Caloric restriction is an anti-aging intervention known to extend lifespan in several experimental models, at least in part, by stimulating autophagy. Caloric restriction increases neuropeptide Y (NPY) in the hypothalamus and plasma ghrelin, a peripheral gut hormone that acts in hypothalamus to modulate energy homeostasis. NPY and ghrelin have been shown to be neuroprotective in different brain areas and to induce several physiological modifications similar to those induced by caloric restriction. However, the effect of NPY and ghrelin in autophagy in cortical neurons is currently not known. Using a cell culture of rat cortical neurons we investigate the involvement of NPY and ghrelin in caloric restriction-induced autophagy. We observed that a caloric restriction mimetic cell culture medium stimulates autophagy in rat cortical neurons and NPY or ghrelin receptor antagonists blocked this effect. On the other hand, exogenous NPY or ghrelin stimulate autophagy in rat cortical neurons. Moreover, NPY mediates the stimulatory effect of ghrelin on autophagy in rat cortical neurons. Since autophagy impairment occurs in aging and age-related neurodegenerative diseases, NPY and ghrelin synergistic effect on autophagy stimulation may suggest a new strategy to delay aging process.
Caloric restriction is an anti-aging intervention known to extend lifespan in several experimental models, at least in part, by stimulating autophagy. Caloric restriction increases neuropeptide Y (NPY) in the hypothalamus and plasma ghrelin, a peripheral gut hormone that acts in hypothalamus to modulate energy homeostasis. NPY and ghrelin have been shown to be neuroprotective in different brain areas and to induce several physiological modifications similar to those induced by caloric restriction. However, the effect of NPY and ghrelin in autophagy in cortical neurons is currently not known. Using a cell culture of rat cortical neurons we investigate the involvement of NPY and ghrelin in caloric restriction-induced autophagy. We observed that a caloric restriction mimetic cell culture medium stimulates autophagy in rat cortical neurons and NPY or ghrelin receptor antagonists blocked this effect. On the other hand, exogenous NPY or ghrelin stimulate autophagy in rat cortical neurons. Moreover, NPY mediates the stimulatory effect of ghrelin on autophagy in rat cortical neurons. Since autophagy impairment occurs in aging and age-related neurodegenerative diseases, NPY and ghrelin synergistic effect on autophagy stimulation may suggest a new strategy to delay aging process.
Entities:
Keywords:
autophagy; caloric restriction; cortical neurons; ghrelin; neuropeptide Y (NPY)
Aging is an age-dependent or age-progressive decline in intrinsic physiological functions. Average human life expectancy has increased, and consequently, the prevalence of cognitive decline dementia, and neurodegenerative diseases. Aging research is now focused in finding strategies that increase both lifespan and healthspan [1].Caloric restriction, reduction of food intake to 30-40 % below ad libitum intake levels without malnutrition and retaining the essential nutrients, is one of the most robust non-pharmacological interventions shown to extend median and maximum lifespan and delay the onset of age-related diseases in several species, including fruit flies, rodents and rhesus monkeys [2-11]. Caloric restriction-induced beneficial effects are mediated, at least in part, by autophagy activation [9, 12-14]. Autophagy is a degradation process of long-lived proteins and organelles and is important for cellular homeostasis maintenance [14, 15]. It is well established that the basal autophagic activity of living cells decreases with age, contributing to the different aspects of the aging phenotype and to the aggravation of detrimental age-related diseases [16, 17]. In fact, several evidences indicate that autophagy impairment is a hallmark of aging and neurodegenerative diseases [16, 18]. The beneficial roles of autophagy in nervous system are mainly associated with maintaining the normal balance between the formation and degradation of cellular proteins as defects in autophagy pathway have been linked to neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease, Huntington's disease, transmissible spongiform encephalopathy or prion disease and Machado-Joseph disease [19-28].Caloric restriction induces a neuroendocrine response such as increasing neuropeptide Y (NPY) levels, in the arcuate nucleus of the hypothalamus [29-32]. NPY is abundantly expressed in numerous brain regions including hypothalamus, hippocampus and cerebral cortex [33]. NPY acts through G-coupled protein NPY receptors, named NPY Y1, Y2, Y4 or Y5 receptors [34]. NPY receptors activation regulates several physiological functions, such as regulation of food intake, blood pressure, body temperature, hormone and neuro-transmitters release, and modulation of pain, sexual behavior, circadian rhythms, memory processing and cognition [35]. In addition, NPY receptors activation has neuroprotective effects in different brain areas and delays neurodegenerative diseases, such as Alzheimer's, Parkinson's and Machado-Joseph disease rodent models [34, 36-38]. Recently, data obtained by our group show that caloric restriction increases NPY levels in hypothalamic neurons and NPY, per se, not only induces autophagy in hypothalamic neurons, but also mediates caloric restriction-induced autophagy, suggesting that NPY may mediate caloric restriction effects on auto-phagy [39, 40]. This effect on other brain regions, such as the cerebral cortex, was never investigated before.Caloric restriction also increases the circulating levels of ghrelin, a peripheral orexigenic hormone synthesized predominantly in the stomach in response to fasting [41-43]. Ghrelin has a ubiquitous expression throughout the body namely in the central nervous system, in particularly in the hypothalamus and cerebral cortex [44, 45]. The actions of ghrelin are mediated through the activation of the G-coupled protein growth hormone secretagogue type 1a receptor (GHS-R1a), which also has a wide tissue distribution [43, 46]. Ghrelin is involved in the regulation of cardiovascular functions, bone metabolism and inflammation [47, 48]. Ghrelin is also involved in memory and learning and has a neuroprotective effect in neurodegenerative diseases and ischemic brain injury models [46, 48-52].Since caloric restriction increases autophagy and both NPY and ghrelin, the aim of this study was to investigate whether NPY and ghrelin stimulates autophagy and if these peptides mediate caloric restriction-induced autophagy in rat cortical neurons. Understanding how NPY and ghrelin may act as caloric restriction mimetics by increasing autophagic clearance in cortical neurons, provides a new anti-aging mechanisms of caloric restriction that could be further explored.
RESULTS
Caloric restriction induces autophagy in rat cortical neurons
To investigate whether caloric restriction regulates autophagy in rat cortical cortical neurons, we monitored autophagy in rat cortical neurons exposed to a caloric restriction mimetic medium (referred as caloric restriction hereafter) by measuring the protein levels of the transient autophagosomal membrane-bound form of LC3B (LC3B-II) and sequestosome 1 (SQSTM1, also known as p62), widely used as markers of the autophagic process [53, 54].As shown in Figure 1A and B, caloric restriction increases LC3B puncta immunoreactivity in rat cortical neurons. While untreated cells (control cells) have a diffuse LC3B cellular distribution, with few small LC3B puncta, in caloric restriction-treated cells an increase in LC3B puncta immunoreactivity was observed, suggesting an increase in autophagosome formation and autophagy induction. The levels of LC3B-II and SQSTM1 were also measured by Western blotting (Figure 1C). The results show that caloric restriction increased LC3B-II protein levels (159.9±9.1% of control) in rat cortical neurons, supporting an increase in the number of auto-phagosomes. However, an increase in LC3B-II levels and the consequent autophagosome formation does not guarantee an increase of the autophagic activity [53]. To rule out the possibility that the increase of LC3B puncta immunoreactivity was due to an inhibited auto-phagosome degradation rather than autophagosome formation, we evaluated endogenous autophagic system in the presence or absence of an inhibitor of lysosomal degradation, chloroquine [53-55]. Since LC3B-II and other autophagic substrates, as is the case of SQSTM1, are degraded at the final stages of autophagy, chloroquine treatment will impair their degradation, leading to the accumulation of both LC3B-II and SQSTM1. In the presence of chloroquine, we observed that the increase of LC3B-II induced by caloric restriction was significantly higher than in cells under caloric restriction without chloroquine (Figure 1C). Concomitant with the increase in LC3B-II steady state levels, caloric restriction decreased SQSTM1 protein content in rat cortical neurons (Figure 1D). The SQSTM1 levels were higher in cells under caloric restriction in the presence of chloroquine than in cells under caloric restriction without chloroquine (132.9±10.9% of control; Figure 1D), indicating that lysosomal degradation was inhibited. Altogether, these results show that caloric restriction increases autophagic clearance in rat cortical neurons. One of the key regulators of autophagy is the mechanistic target of rapamycin (MTOR), a conserved serine/threonine kinase that suppresses the initiation of the autophagic process when nutrients, growth factors, and energy are available [57, 58]. Inhibition of MTOR, therefore, results in activation of autophagy [57, 58]. MTOR activity can be assessed by the analysis of MTOR phosphorylation at its active site Ser2448. As shown in Figure 1E, caloric restriction decreased phospho-MTOR levels (60.2±6.3% of control), indicating that caloric restriction induces autophagy in rat cortical neurons through MTOR inhibition.
Figure 1
Caloric restriction increases autophagy in rat cortical neurons
Primary rat cortical neurons were exposed to caloric restriction mimic medium (CR), DMEM low glucose, for 6 h. Untreated cells were used as control (Ctrl). (A) LC3B puncta immunoreactivity was assessed by immunocytochemistry, as described in Materials and Methods. Cells were immunolabeled for LC3B (green) and MAP2 (red). Nuclei were stained with Hoechst 33342 (blue). Figures are representative of three independents experiments. Scale bar, 20 μM. (B) Quantification of LC3B puncta immunoreactivity (green) per cell in each condition (>20 cells per group). ***p<0.001, significantly different compared to control, as determined by Student's t test. (C, D and E) Cells were incubated with chloroquine (ChQ, 100 μM), a lysosomal degradation inhibitor, 30 min before caloric restriction medium for 6 h. Whole cell extracts were assayed for LC3B-II (C), SQSTM1 (D), phospho-MTOR (p-MTOR) (E) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. **p<0.01 and ***p<0.001, significantly different compared to control; ###p<0.001, significantly different from caloric restriction; $$$p<0.001, significantly different from chloroquine-treated cells, as determined by ANOVA, followed by Bonferroni's post test.
Caloric restriction increases autophagy in rat cortical neurons
Primary rat cortical neurons were exposed to caloric restriction mimic medium (CR), DMEM low glucose, for 6 h. Untreated cells were used as control (Ctrl). (A) LC3B puncta immunoreactivity was assessed by immunocytochemistry, as described in Materials and Methods. Cells were immunolabeled for LC3B (green) and MAP2 (red). Nuclei were stained with Hoechst 33342 (blue). Figures are representative of three independents experiments. Scale bar, 20 μM. (B) Quantification of LC3B puncta immunoreactivity (green) per cell in each condition (>20 cells per group). ***p<0.001, significantly different compared to control, as determined by Student's t test. (C, D and E) Cells were incubated with chloroquine (ChQ, 100 μM), a lysosomal degradation inhibitor, 30 min before caloric restriction medium for 6 h. Whole cell extracts were assayed for LC3B-II (C), SQSTM1 (D), phospho-MTOR (p-MTOR) (E) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. **p<0.01 and ***p<0.001, significantly different compared to control; ###p<0.001, significantly different from caloric restriction; $$$p<0.001, significantly different from chloroquine-treated cells, as determined by ANOVA, followed by Bonferroni's post test.
Caloric restriction increases NPY and ghrelin mRNA and protein levels in rat cortical neurons
Since caloric restriction was shown to increase the levels of hypothalamic NPY [28-31] and circulating ghrelin [39, 41, 42], we next investigated whether caloric restriction could also regulate the levels of both peptides in rat cortical neurons. As shown in Figure 2A and B, caloric restriction mimetic medium increased both NPY and ghrelin mRNA levels in primary rat cortical neurons (1.3±0.1 and 2.1±0.3 fold increase over control, respectively). Concomitantly, caloric restriction mimetic medium increased both NPY and ghrelin protein content in primary rat cortical neurons (342.3±49.0 and 237.5±32.3 percentage of control, respectively; Figure 2C and D).
Figure 2
Caloric restriction increases NPY and ghrelin levels in rat cortical neurons
Primary rat cortical neurons were exposed to caloric restriction medium (CR), DMEM low glucose, for 6 h. Cells in neurobasal medium with 2 % B27 supplement were used as control (Ctrl). (A and B) Total RNA was isolated and the transcript levels of NPY and ghrelin were analyzed by qRT-PCR, as described in Materials and Methods. The results represent the mean ± SEM of five independents experiments and are expressed as the relative amount compared to control. **p<0.01 and ***p<0.001, significantly different compared to control, as determined by Student's t test. (C and D) Caloric restriction increased NPY and ghrelin protein content, determined by Enzyme-Linked Immunosorbent Assays, as described in Material and Methods. The results represent the mean ± SEM of 3-4 independents experiments and are expressed as the relative amount compared to control. **p<0.01 significantly different compared to control, as determined by Student's t test.
Caloric restriction increases NPY and ghrelin levels in rat cortical neurons
Primary rat cortical neurons were exposed to caloric restriction medium (CR), DMEM low glucose, for 6 h. Cells in neurobasal medium with 2 % B27 supplement were used as control (Ctrl). (A and B) Total RNA was isolated and the transcript levels of NPY and ghrelin were analyzed by qRT-PCR, as described in Materials and Methods. The results represent the mean ± SEM of five independents experiments and are expressed as the relative amount compared to control. **p<0.01 and ***p<0.001, significantly different compared to control, as determined by Student's t test. (C and D) Caloric restriction increased NPY and ghrelin protein content, determined by Enzyme-Linked Immunosorbent Assays, as described in Material and Methods. The results represent the mean ± SEM of 3-4 independents experiments and are expressed as the relative amount compared to control. **p<0.01 significantly different compared to control, as determined by Student's t test.
Caloric restriction stimulates autophagy through NPY receptors activation
We observed that caloric restriction induces autophagy in rat cortical neurons and this is accompanied by an increase in NPY levels. Given that NPY and NPY receptors are expressed in rat cortical neurons [56], we hypothesized that NPY receptors could play a role on caloric restriction-induced autophagy in rat cortical neurons. We observed that NPY Y1, Y2 or Y5 receptor antagonists inhibited the stimulatory effect of caloric restriction on autophagy markers: the increase in LC3B-II (Figure 3A) and the decrease in SQSTM1 (Figure 3B) levels. These results suggest that caloric restriction-induced autophagy in rat cortical neurons is mediated by NPY Y1, Y2 or Y5 receptor activation.
Figure 3
NPY receptor antagonists inhibit the stimulatory effect of caloric restriction on autophagy in rat cortical neurons
Primary rat cortical neurons were incubated with NPY Y1 receptor antagonist BIBP3226 (Y1ant, 1 μM), NPY Y2 receptor antagonist BIIE0246 (Y2ant, 1 μM) or NPY Y5 receptor antagonist L152,800 (Y5ant, 1 μM), 30 min before caloric restriction medium (CR) for 6 h. Whole cell extracts were assayed for LC3B-II (A), SQSTM1 (B) and β-tubulin (loading control) immunoreactivity by Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05 and ***p<0.001, significantly different compared to control; #p<0.05, ##p<0.01 and ###p<0.001, significantly different from caloric restriction, as determined by ANOVA, followed by Bonferroni's post test.
NPY receptor antagonists inhibit the stimulatory effect of caloric restriction on autophagy in rat cortical neurons
Primary rat cortical neurons were incubated with NPY Y1 receptor antagonist BIBP3226 (Y1ant, 1 μM), NPY Y2 receptor antagonist BIIE0246 (Y2ant, 1 μM) or NPY Y5 receptor antagonist L152,800 (Y5ant, 1 μM), 30 min before caloric restriction medium (CR) for 6 h. Whole cell extracts were assayed for LC3B-II (A), SQSTM1 (B) and β-tubulin (loading control) immunoreactivity by Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05 and ***p<0.001, significantly different compared to control; #p<0.05, ##p<0.01 and ###p<0.001, significantly different from caloric restriction, as determined by ANOVA, followed by Bonferroni's post test.
NPY induces autophagy in rat cortical neurons
As the activation of NPY receptors is involved in caloric restriction-induced autophagy, we then investigated the effect of NPY per se on rat cortical neurons autophagy. We observed that NPY, similarly to caloric restriction, increased LC3B puncta immunoreactivity (Figure 4A and B) and LC3B-II steady state levels (129.8±4.6% of control; Figure 4C) in rat cortical neurons. Moreover, in the presence of chloroquine, the increase in LC3B-II levels was higher (277.7±28.2% of control) than in cells treated with NPY without chloroquine (Figure 4C). NPY also decreased SQSTM1 content in rat cortical neurons (Figure 4D) and this effect was inhibited in the presence of chloroquine. Moreover, we observed that NPY Y1, Y2 or Y5 receptor antagonists inhibited LC3B-II increase (Figure 4E) and SQSTM1 decrease (Figure 4F) induced by NPY. Overall, these results show that NPY enhances autophagic activity in rat cortical neurons, through NPY Y1, Y2 or Y5 receptors activation. As shown in Figure 4G, NPY decreases phospho-MTOR (Ser2448) levels (78.4±3.3% of control) in rat cortical neurons, which suggest that NPY, similarly to caloric restriction, induces autophagy through the inhibition of MTOR.
Figure 4
NPY increases autophagy in rat cortical neurons
Primary rat cortical neurons were exposed to NPY (100 nM) for 6 h. Untreated cells were used as control (Ctrl). (A) LC3B distribution was assessed by immunocytochemistry assay, as described in Materials and Methods. Cells were immunolabeled for LC3B (green) and MAP2 (red, neurons). Nuclei were stained with Hoechst 33342 (blue). Figures are representative of three independents experiments. Scale bar, 20 μM. (B) Quantification of the number of LC3B puncta immunoreactivity (green) per cell in each condition (>20 cells per group). ***p<0.001, significantly different compared to control, as determined by Student's t test. (C-G) Cells were incubated with chloroquine (ChQ, 100 μM) (C and D), or with Y1 receptor antagonist BIBP3226 (Y1ant, 1 μM), Y2 receptor antagonist BIIE0246 (Y2ant, 1 μM) or Y5 receptor antagonist L152,800 (Y5ant, 1 μM) (E and F), 30 min before NPY (100 nM). Whole cell extracts were assayed for LC3B-II (C and E), SQSTM1 (D and F), phospho-MTOR (p-MTOR) (G) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05, **p<0.01 and ***p<0.001, significantly different compared to control; #p<0.05, ##p<0.01 and ###p<0.001, significantly different from NPY treatment; $$$p<0.001, significantly different from chloroquine-treated cells, as determined by ANOVA, followed by Bonferroni's post test.
NPY increases autophagy in rat cortical neurons
Primary rat cortical neurons were exposed to NPY (100 nM) for 6 h. Untreated cells were used as control (Ctrl). (A) LC3B distribution was assessed by immunocytochemistry assay, as described in Materials and Methods. Cells were immunolabeled for LC3B (green) and MAP2 (red, neurons). Nuclei were stained with Hoechst 33342 (blue). Figures are representative of three independents experiments. Scale bar, 20 μM. (B) Quantification of the number of LC3B puncta immunoreactivity (green) per cell in each condition (>20 cells per group). ***p<0.001, significantly different compared to control, as determined by Student's t test. (C-G) Cells were incubated with chloroquine (ChQ, 100 μM) (C and D), or with Y1 receptor antagonist BIBP3226 (Y1ant, 1 μM), Y2 receptor antagonist BIIE0246 (Y2ant, 1 μM) or Y5 receptor antagonist L152,800 (Y5ant, 1 μM) (E and F), 30 min before NPY (100 nM). Whole cell extracts were assayed for LC3B-II (C and E), SQSTM1 (D and F), phospho-MTOR (p-MTOR) (G) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05, **p<0.01 and ***p<0.001, significantly different compared to control; #p<0.05, ##p<0.01 and ###p<0.001, significantly different from NPY treatment; $$$p<0.001, significantly different from chloroquine-treated cells, as determined by ANOVA, followed by Bonferroni's post test.
Caloric restriction stimulates autophagy through ghrelin receptor activation
Since caloric restriction increases ghrelin mRNA and protein levels in rat cortical neurons (Figure 2B), we hypothesized that ghrelin, similarly to NPY, could be involved in caloric restriction-induced autophagy in rat cortical neurons. As shown in Figure 5A and B, ghrelin receptor (GHS-R1a) antagonist ([D-Lys3]-GHRP-6) inhibited the increase of LC3B-II and the decrease of SQSTM1, induced by caloric restriction in rat cortical neurons. These results suggest that ghrelin receptor GHS-R1a mediates, in part, caloric restriction-induced autophagy in rat cortical neurons.
Figure 5
Ghrelin mediates caloric restriction-induced autophagy in rat cortical neurons
Primary rat cortical neurons were treated with GHS-R1a receptor antagonist [D-Lys3]-GHRP-6 (GHS-R1a ant, 100 μM) 30 min before caloric restriction (CR) for 6 h. Whole cell extracts were assayed for LC3B-II (A), SQSTM1 (B) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05 and ***p<0.001, significantly different compared to control; #p<0.05, ##p<0.01 and ###p<0.001, significantly different from caloric restriction, as determined by ANOVA, followed Bonferroni's post test.
Ghrelin mediates caloric restriction-induced autophagy in rat cortical neurons
Primary rat cortical neurons were treated with GHS-R1a receptor antagonist [D-Lys3]-GHRP-6 (GHS-R1a ant, 100 μM) 30 min before caloric restriction (CR) for 6 h. Whole cell extracts were assayed for LC3B-II (A), SQSTM1 (B) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05 and ***p<0.001, significantly different compared to control; #p<0.05, ##p<0.01 and ###p<0.001, significantly different from caloric restriction, as determined by ANOVA, followed Bonferroni's post test.
Ghrelin induces autophagy in rat cortical neurons
We next evaluated the effect of ghrelin per se on autophagy in rat cortical neurons. Similarly to caloric restriction and NPY, in rat cortical neurons, ghrelin induced autophagy and autophagosome formation, as shown by an increase in LC3B puncta immunoreactivity (Figure 6A), LC3B-II steady state levels (Figure 6B), and autophagic degradation, as shown by SQSTM1 protein decrease (Figure 6C and D). As expected, the GHS-R1a receptor blockage with the ghrelin receptor antagonist ([D-Lys3]-GHRP-6) abolished ghrelin stimulatory effects on both autophagic substrates (Figure 6E and F). Next, we observed that ghrelin decreased phospho-MTOR levels in rat cortical neurons, suggesting that ghrelin, similarly to caloric restriction and NPY, induced autophagy through the canonical inhibition of MTOR activity (Figure 6G).
Figure 6
Ghrelin induces autophagy in rat cortical neurons
Primary rat cortical neurons were exposed to ghrelin (GHRL, 10 nM) for 6 h. Untreated cells were used as control (Ctrl). (A) LC3B cellular distribution was assessed by immunocytochemistry assay, as described in Materials and Methods. Cells were immunolabeled for LC3B (green) and MAP2 (red, neurons). Nuclei were stained with Hoechst 33342 (blue). Figures are representative of three independents experiments. Scale bar, 20 μM. (B) Quantification of the number of LC3B puncta immunoreactivity (green) per cell in each condition (>20 cells per group). ***p<0.001, significantly different compared to control, as determined by Student's t test. (C-G) Cells were incubated with chloroquine (ChQ, 100 μM), a lysosomal degradation inhibitor (C and D) or GHS-R1a receptor antagonist [D-Lys3]-GHRP-6 (GHS-R1a ant, 100 μM) (E and F), 30 min before ghrelin (GHRL, 10 nM) treatment for 6 h. Whole cell extracts were assayed for LC3B-II (C and E), SQSTM1 (D and F), phospho-MTOR (p-MTOR) (G) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05, **p<0.01 and ***p<0.001, significantly different compared to control; #p<0.05 and ###p<0.001, significantly different from ghrelin treatment, as determined by ANOVA, followed Bonferroni's post test.
Ghrelin induces autophagy in rat cortical neurons
Primary rat cortical neurons were exposed to ghrelin (GHRL, 10 nM) for 6 h. Untreated cells were used as control (Ctrl). (A) LC3B cellular distribution was assessed by immunocytochemistry assay, as described in Materials and Methods. Cells were immunolabeled for LC3B (green) and MAP2 (red, neurons). Nuclei were stained with Hoechst 33342 (blue). Figures are representative of three independents experiments. Scale bar, 20 μM. (B) Quantification of the number of LC3B puncta immunoreactivity (green) per cell in each condition (>20 cells per group). ***p<0.001, significantly different compared to control, as determined by Student's t test. (C-G) Cells were incubated with chloroquine (ChQ, 100 μM), a lysosomal degradation inhibitor (C and D) or GHS-R1a receptor antagonist [D-Lys3]-GHRP-6 (GHS-R1a ant, 100 μM) (E and F), 30 min before ghrelin (GHRL, 10 nM) treatment for 6 h. Whole cell extracts were assayed for LC3B-II (C and E), SQSTM1 (D and F), phospho-MTOR (p-MTOR) (G) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05, **p<0.01 and ***p<0.001, significantly different compared to control; #p<0.05 and ###p<0.001, significantly different from ghrelin treatment, as determined by ANOVA, followed Bonferroni's post test.
Neuropeptide Y regulates, in part, ghrelin-induced autophagy in rat cortical neurons
As ghrelin regulates NPY expression in hypothalamic neurons [57], we hypothesized that ghrelin could also regulate NPY levels in rat cortical neurons. As shown in Figure 7A and B, ghrelin increased NPY mRNA (1.9±0.3 fold increase over control) and NPY protein (165.7±23.5 percentage of control) levels in rat cortical neurons. This interesting observation led us to hypothesize whether NPY receptors activation through endogenous NPY could play a role on ghrelin-induced autophagy in rat cortical neurons. We observed that NPY Y1, Y2 or Y5 receptor antagonists significantly decreased autophagy (LC3B-II increase and SQSTM1 decrease) induced by ghrelin (Figure 7C and D). These results suggest that ghrelin enhances autophagy in rat cortical neurons, at least partially, by increasing NPY levels and consequently NPY receptors activation.
Figure 7
Ghrelin increases NPY content and NPY receptor antagonists block the stimulatory role of ghrelin on autophagy in rat cortical neurons
Primary rat cortical neuronal cultures were exposed to ghrelin (GHRL, 10 nM) for 6 h. Untreated cells were used as control (Ctrl). (A) Total RNA was isolated and the transcript levels of NPY were analyzed by qPCR, as described in Materials and Methods. The results represent the mean ± SEM of five independents experiments and are expressed as the relative amount compared to control. *p<0.05, significantly different compared to control, as determined by Student's t test. (B) Ghrelin leads to increased NPY protein content, using an Enzyme-Linked Immunosorbent Assay, as described in Material and Methods. The results represent the mean ± SEM of three independents experiments and are expressed as the relative amount compared to control. (C and D) Cells were incubated with NPY Y1 receptor antagonist BIBP3226 (Y1ant, 1 μM), NPY Y2 receptor antagonist BIIE0246 (Y2ant, 1 μM) or NPY Y5 receptor antagonist L152,800 (Y5ant, 1 μM), 30 min before ghrelin (GHRL, 10 nM) treatment for 6 h. Whole cell extracts were assayed for LC3B-II (C), SQSTM1 (D) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05 significantly different compared to control; #p<0.05 and ##p<0.01, significantly different from ghrelin treatment, as determined by ANOVA, followed by Bonferroni's post test.
Ghrelin increases NPY content and NPY receptor antagonists block the stimulatory role of ghrelin on autophagy in rat cortical neurons
Primary rat cortical neuronal cultures were exposed to ghrelin (GHRL, 10 nM) for 6 h. Untreated cells were used as control (Ctrl). (A) Total RNA was isolated and the transcript levels of NPY were analyzed by qPCR, as described in Materials and Methods. The results represent the mean ± SEM of five independents experiments and are expressed as the relative amount compared to control. *p<0.05, significantly different compared to control, as determined by Student's t test. (B) Ghrelin leads to increased NPY protein content, using an Enzyme-Linked Immunosorbent Assay, as described in Material and Methods. The results represent the mean ± SEM of three independents experiments and are expressed as the relative amount compared to control. (C and D) Cells were incubated with NPY Y1 receptor antagonist BIBP3226 (Y1ant, 1 μM), NPY Y2 receptor antagonist BIIE0246 (Y2ant, 1 μM) or NPY Y5 receptor antagonist L152,800 (Y5ant, 1 μM), 30 min before ghrelin (GHRL, 10 nM) treatment for 6 h. Whole cell extracts were assayed for LC3B-II (C), SQSTM1 (D) and β-tubulin (loading control) immunoreactivity through Western blotting analysis, as described in Materials and Methods. Representative Western blots for each protein are presented above each respective graph. The results represent the mean ± SEM of, at least, five independents experiments, and are expressed as percentage of control. *p<0.05 significantly different compared to control; #p<0.05 and ##p<0.01, significantly different from ghrelin treatment, as determined by ANOVA, followed by Bonferroni's post test.
DISCUSSION
In the present study, we show, for the first time, that NPY and ghrelin mediate autophagy stimulation induced by caloric restriction in rat cortical neurons.These results are in agreement to recent studies that show autophagy induction in primary cortical neurons upon caloric restriction and in rodent cortical neurons upon short-term food restriction [58, 59].Caloric restriction increases NPY in the hypothalamic neurons and herein we show that caloric restriction also increases NPY levels also in rat cortical neurons [29, 39]. In addition, we observed that NPY Y1, Y2 and Y5 receptor antagonists decreased the stimulatory effect of caloric restriction on autophagy, suggesting that caloric restriction-induced autophagy is dependent on NPY Y1, Y2 or Y5 receptor activation in rat cortical neurons. Accordingly, we recently showed that caloric restriction stimulates autophagy in rodent hypothalamic neurons and the NPY Y1, Y2 or Y5 receptors antagonists inhibits this stimulatory effect of caloric restriction in autophagy [39].In the present study we showed that exogenous NPY enhances autophagy in rat cortical neurons through NPY Y1, Y2 or Y5 receptor activation. The similarity between the effects of caloric restriction and NPY on autophagy in cortical neuronal suggests that NPY mediates caloric restriction-induced autophagy and may be considered as a caloric restriction mimetic, as suggested by others [29, 60]. NPY and caloric restriction induce similar physiological effects, such as: hyperphagia, decreased blood glucose levels, reduced core body temperature and reduced fertility [29]. In addition, it has been shown that NPY mediates the antitumorigenic effect of caloric restriction and that caloric restriction does not increase lifespan of NPY KO mice, enlightening NPY role as a lifespan and aging regulator [61, 62]. In fact, in humans, increased NPY levels may also be correlated with lifespan benefits, since long-lived female centenarians have higher NPY plasma levels compared to younger women [63].Aging is associated with attenuated ghrelin signaling [64, 65]. During aging, caloric restriction produces health benefits accompanied by enhanced ghrelin and ghrelin receptor (GHS-R1a) levels [41-43, 66-68]. For the first time, we show that ghrelin levels rise in rat cortical neurons upon caloric restriction, and blocking ghrelin receptor (GHS-R1a), the stimulatory effect of caloric restriction on autophagy was partially inhibited. These results suggest that ghrelin signaling may represent one of the mechanisms activated by caloric restriction. Moreover, similarly to caloric restriction, exogenous ghrelin stimulates autophagy in rat cortical neurons, by GHS-R1a receptor activation. These results suggest the potential role of ghrelin as a caloric restriction mimetic. In fact, like caloric restriction, ghrelin has several beneficial effects of age-related diseases. Ghrelin is involved in the regulation of cardiovascular functions (increase of cardiac output, decrease blood flow, protection against cardiac damage, anti-apoptotic effects), bone metabolism (increase osteoblast differentiation and bone mineral density) and inflammation (suppressing the production of cytokines) [48]. Ghrelin is also involved in memory and learning and has a neuroprotective effect in neurodegenerative diseases and ischemic brain injury models [46, 48, 69]. Indeed, ghrelin is effective in improving cell survival, reducing infarct size and rescuing memory in these animal models. Although it has been proposed that dysfunction of ghrelin signaling, through GHS-R1a ablation, may be beneficial to age-related obesity and insulin resistance [70], it has been reported that ghrelin administration in rodents and humans can possibly reverse certain characteristics of aging [71-78]. In fact, a recent study show that increasing ghrelin signalling ameliorated several age-related disorders and prolonged survival in several animal models of human aging, supporting endogenous ghrelin signalling as an important role in preventing aging-related diseases and premature death [78]. Ghrelin and GHS-R1a functions are diverse and the inter-action between their central and peripheral effects are complex, raising some controversy regarding ghrelin physiological versus pharmacological action [79]. The effectiveness of ghrelin in these roles may be impaired as ghrelin levels decrease with age, perhaps contributing to other age-related conditions like insulin resistance and diabetes, reduced fertility, and decreased performance on cognitive and memory tasks with advancing age [80, 81]. In addition, ghrelin is already being used in several clinical trials as a therapeutic strategy for the treatment of cachexia in chronic heart failure, cancer, end stage- renal disease or cystic fibrosis, frailty in elderly, anorexia nervosa, growth hormone deficientpatients and sleep-wake regulation (e.g. majordepression) [82, 83].The significant overlap between caloric restriction- and ghrelin-induced physiological processes suggest that ghrelin may play a role in the beneficial effects of caloric restriction on health and lifespan. In fact, we observed that ghrelin increases NPY expression in rat cortical neurons and NPY Y1, Y2 or Y5 subtypes receptors antagonists inhibited the stimulatory effect of ghrelin on autophagy. These observations suggest that, similarly to caloric restriction, NPY also mediates ghrelin-induced autophagy in rat cortical neurons. The contribution of NPY in ghrelin effects has been shown by other on feeding behavior, energy balance, growth hormone secretion and gastrointestinal motility [45, 57, 84-87].Cellular metabolic stress underpins the development of pathological conditions, of which the prevalence increases dramatically with age. In fact, a decline of NPY and ghrelin plasma levels in humans correlates with the increase of age [88-90] and the modulation of these peptides has been shown to provide neuroprotection in several neurodegenerative diseases [46, 91-94].Overall, the present study shows that NPY and ghrelin stimulate autophagy and mediate autophagy stimulation induced by caloric restriction in rat cortical neurons. Furthermore, NPY mediates, in part, ghrelin-induced autophagy, which suggests that both peptides have a synergist effect on autophagy in rat cortical neurons. Since caloric restriction increases both NPY and ghrelin levels in cortical neurons, modulation of both peptides may be considered as a protective mechanism against impaired cortical neuronal dysfunction. Moreover, given the difficulty to implement a caloric restriction regimen, and the fact that autophagy impairment occurs in aging and age-related neurodegenerative diseases, NPY and ghrelin synergistic effect on autophagy stimulation may suggest a new approach for the improvement of both healthspan and lifespan in aging societies.
MATERIALS AND METHODS
Animals
Female Wistar rats were purchased from Charles River Laboratories (L'Arbresle, France). All experimental procedures were performed in accordance to the guidelines of the European Community directive for the use of animals in laboratory (2010/63/EU) translated to the Portuguese law in 2013 (Decreto-lei 113/2013). The researchers received adequate training (Felasa-certified course) and certification to perform the experiments from the Portuguese authorities (Direcção Geral de Veterinária). The present study is included in a project approved and financed by the Portuguese Science Foundation that approved the animal experimentation described. CNC – Center for Neuroscience and Cell Biology – University of Coimbra animal experimentation board approved the utilization of animals for this project (reference PTDC/SAU-FCF/099082/2008).
Primary rat cortical neuronal cultures
Primary rat cortical neuronal cultures were obtained from cortical tissue was dissected from rats at embryonic days 18-19 (E18-19) from female rats of 16-24 weeks of age and 250-300 g of body weight. The pregnant females were sacrificed by cervical dislocation and subjected to caesarean section in order to remove the uterine horns containing the embryos. The brains were removed from the skull and cortices were dissected and meninges thoroughly removed. The cells were chemically dissociated in the presence of 0.25 % trypsin (Invitrogen) and 50 μg.mL−1 DNAse I (Sigma-Aldrich). After the cells were isolated, they were resuspended in high glucose (4.5 g.L−1) Neurobasal medium with 500 μM L-Glutamine, 2 % B27 supplement, 100 U.mL−1 penicillin and 100 μg.mL−1 streptomycin (all from Gibco), with no growth factors, and plated at a density of 1.32×105 cells.cm−2 on poly-D-Lysine coated cell culture plates. Cortical neurons were maintained at 37°C in a humidified incubator with 5 % CO2/air for 8 days and the medium was replaced every fourth day by aspirating half of the medium from each well and replacing it with fresh medium.
Experimental conditions
Cells were exposed to nutrient deprivation to mimic a caloric restriction condition, Dulbecco's Modified Eagle Medium (DMEM; Sigma-Aldrich, St. Louis, MO, USA) low glucose medium (1 g.L−1 glucose, 100 U.mL−1 penicillin and 100 μg.mL−1 streptomycin, without B27 supplementation), NPY (100 nM; Phoenix Europe GmbH, Karlsruhe, Germany) or acylated ghrelin (10 nM; Bachem, Bubendorf, Switzerland). Cells were also exposed to the lysosomal protein degradation inhibitor, chloroquine (100 μM; Sigma-Aldrich), NPY receptors selective antagonists (all at 1 μM; all from Bachem, Bubendorf, Switzerland; NPY Y1 antagonist (BIBP3226), NPY Y2 antagonist (BIIE0246) and NPY Y5 antagonist (L-152,804)), and/or ghrelin receptor antagonist ([D-Lys3]-GHRP-6) (100 μM; Tocris Bioscience, Bristol, UK), added to the cell culture medium 30 min prior to caloric restriction, NPY or ghrelin treatment for 6 h.
Isolation of total RNA and cDNA synthesis
Total RNA was isolated using the RNeasy Mini Kit (Qiagen) according to the manufacturer's instructions. Briefly, cells were lysed, the total RNA was adsorbed to a silica matrix, washed with the recommended buffers and eluted with 30 μL of RNase-free water by centrifugation. Total RNA amount was quantified by optical density (OD) measurements using a ND-1000 Nanodrop Spectrophotometer (Thermo Scientific), and the purity was evaluated by measuring the ratio of OD at 260 and 280 nm. RNA samples were treated with RNase-free DNAse (Qiagen) to eliminate any contamination with genomic DNA. Reverse transcription into cDNA was carried out using the iScript Select cDNA Synthesis Kit (Bio-Rad) following the manufacturer's instructions. Briefly, 1 μg of total RNA from each sample was reverse transcribed into cDNA in a 20 μL reaction containing 1x reaction buffer, 1x random primers, and 50 units of reverse transcriptase. Reverse transcription reactions were performed in a thermocycler at 25°C for 5 min, 42°C for 30 min, 85°C for 5 min, and 4°C for 5 min. cDNA samples were then stored at −20°C until use.
Quantitative real-time PCR was performed in an iQ5 thermocycler (Bio-Rad) using 96-well microtiter plates and the QuantiTect SYBR Green PCR Master Mix (Qiagen). The primers for the target rat genes (NPY, NM-012614), (Ghrelin, NM-021669) and the reference gene (ratHPRT, NM-012583) were pre-designed and validated by QIAGEN (QuantiTect Primers, Qiagen). A master mix was prepared for each primer set, containing the appropriate volume of 2× QuantiTect SYBR Green PCR Master Mix and 10× QuantiTectPrimer. For each reaction, 18 μL of master mix were added to 2 μL of template cDNA. All reactions were performed in duplicate (two cDNA reactions per RNA sample) at a final volume of 20 μL per well. Negative controls were performed without RNA sample, which was substituted by water. The reactions were performed according to the manufacturer's recommendations: 95°C for 15 min, followed by 40 cycles at 94°C for 15 sec, 55°C for 30 sec and 72°C for 30 sec. The melting curve protocol started immediately after amplification. The amplification efficiency for each gene and the threshold values for threshold cycle determination (Ct) were determined automatically by the iQ5 Optical System Software (Bio-Rad). Relative mRNA quantification was performed using the ΔCt method for genes with the same amplification efficiency. The results are expressed as the relative amount compared to control.
Determination of NPY and ghrelin protein content
Samples were assayed for NPY and ghrelin concentration using an NPY and ghrelin EIA kit (RayBiotech, Norcross, GA, USA), respectively, according to manufacturer instructions. Cells were lysed with Krebs buffer (132 mM NaCl; 4 mM KCl; 1.4 mM MgCl2; 1 mM CaCl2; 10 mM glucose; 10 mM Hepes, supplemented with 0.001 % Tween 20, pH 7.4). Each lysate was sonicated 6x 5 sec pulses and centrifuged at 16000×g, for 8 min, at 4°C. Each supernatant was collected to a PolySorb™ tube, containing 5 % Tween 20 and 0.03 M EDTA and was stored at −80°C, until use. The protein concentration of each sample was determined by the bicinchoninic acid (BCA) protein assay (Pierce Biotechnology).
Western blotting
Cells were lysed on ice in RIPA buffer (50 mM Tris-HCl, pH 7.4; 150 mM NaCl; 5 mM EDTA; 1 % Triton X-100; 0.5 % deoxycholate; 0.1 % sodium dodecyl sulphate (SDS); 200 μM phenyl-methylsulphonylfluoride (PMSF); 1 mM dithiothreitol (DTT), 1 mM Na3VO4; 10 mM NaF), supplemented with complete mini protease inhibitor cocktail tablet (Roche). Lysates were incubated for 15 min at 4°C, and the insoluble material was pelleted by centrifugation for 10 min at 16,000xg and 4°C. The protein concentration of each sample was determined by the bicinchoninic acid (BCA) protein assay (Pierce Biotechnology). The samples were denaturated by adding 6x concentrated sample buffer (0.5 M Tris, 30 % glycerol, 10 % SDS, 0.6 M DTT, 0.012 % bromophenol blue) and heating for 5 min at 95°C. Samples were stored at −20°C until use. Equal amounts of total protein were loaded per lane and separated by electrophoresis in SDS-PAGE, using 8-12 % gels. The protein samples were then transferred electrophoretically in CAPS buffer (0.1 M CAPS, pH 11.0; 10 % methanol) to PVDF membranes (Millipore). After blotting, the membranes were blocked in 5 % non-fat milk in Tris-buffered saline (137 mM NaCl, 20 mM Tris–HCl, pH 7.6) containing 0.1 % Tween 20 (TBS-T) for one hour at room temperature and then incubated overnight with the primary antibodies at 4°C. The primary antibodies used (all at a dilution of 1:1000; Cell Signaling) were: rabbit polyclonal anti-LC3B, anti-SQSTM1 and anti-phospho-MTOR (Ser2448). There-after, the membranes were incubated with an alkaline phosphatase-linked secondary antibody, specific to rabbit IgG or mouse IgG in a 1:10000 dilution (GE Healthcare) Protein immunoreactive bands were visualized by chemifluorescence using the ECF substrate (GE Healthcare) in a VersaDoc Imaging System (Bio-Rad) and the optical density of the bands was quantified with the Quantity One Software (Bio-Rad). The membranes were reprobed for β-tubulin immuno-reactivity (1:10000; Sigma) for equal protein loading control.
Immunocytochemistry
After treatments, cells were washed with PBS and then fixed with ice-cold 4 % paraformaldehyde/PBS for 15 min. Cells were permeabilized with 0.25 % TX-100/PBS for 10 min, washed in PBS and blocked for one hour blocking in 10 % goat serum/PBS. Cells were incubated with primary antibodies overnight at 4°C. After incubation, cells were washed in PBS and incubated for one hour at room temperature with the respective secondary antibodies. The nuclei were stained with Hoechst 33342 (2 μg.mL−1; Sigma-Aldrich) during secondary antibody incubation. The coverslips were washed in PBS and mounted on glass slides with Dako Fluorescence Mounting Medium (Dako). The primary antibodies used were: rabbit anti-LC3B (1:400; Cell Signaling) and mouse anti-MAP2 (1:500; Sigma-Aldrich). The secondary antibodies (all at a dilution 1:200; Invitrogen) used were: Alexa-Fluor 488-conjugated goat anti-rabbit IgG and Alexa-Fluor 594-conjugated goat anti-mouse IgG. Cells were analyzed on a Zeiss Axiovert fluorescence microscope (Carl Zeiss). The procedure was performed for three independent cell culture preparations. Quantification of LC3B puncta immuno-reactivity in rat cortical neurons and imaging procedures using the Fiji (Fiji is Just ImageJ) software (National Health Institute) were performed. The results are expressed as the relative amount compared with control.
Statistical analysis
Results are expressed as mean ± standard error of the mean (SEM). Data were analyzed using one-way analysis of variance (ANOVA) followed by Bonferroni's post test, or Student's unpaired t test with two-tailed p value, as indicated in figure legends. A value of p<0.05 was considered significant. Prism 5.0 (GraphPad Software) was used for all statistical analysis.
Authors: C Stadelmann; T L Deckwerth; A Srinivasan; C Bancher; W Brück; K Jellinger; H Lassmann Journal: Am J Pathol Date: 1999-11 Impact factor: 4.307
Authors: Michael Lutter; Ichiro Sakata; Sherri Osborne-Lawrence; Sherry A Rovinsky; Jason G Anderson; Saendy Jung; Shari Birnbaum; Masashi Yanagisawa; Joel K Elmquist; Eric J Nestler; Jeffrey M Zigman Journal: Nat Neurosci Date: 2008-06-15 Impact factor: 24.884
Authors: Ligen Lin; Pradip K Saha; Xiaojun Ma; Iyabo O Henshaw; Longjiang Shao; Benny H J Chang; Eric D Buras; Qiang Tong; Lawrence Chan; Owen P McGuinness; Yuxiang Sun Journal: Aging Cell Date: 2011-10-12 Impact factor: 9.304
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Susanna Ambrosio; Amal O Amer; Veena Ammanathan; Zhenyi An; Stig U Andersen; Shaida A Andrabi; Magaiver Andrade-Silva; Allen M Andres; Sabrina Angelini; David Ann; Uche C Anozie; Mohammad Y Ansari; Pedro Antas; Adam Antebi; Zuriñe Antón; Tahira Anwar; Lionel Apetoh; Nadezda Apostolova; Toshiyuki Araki; Yasuhiro Araki; Kohei Arasaki; Wagner L Araújo; Jun Araya; Catherine Arden; Maria-Angeles Arévalo; Sandro Arguelles; Esperanza Arias; Jyothi Arikkath; Hirokazu Arimoto; Aileen R Ariosa; Darius Armstrong-James; Laetitia Arnauné-Pelloquin; Angeles Aroca; Daniela S Arroyo; Ivica Arsov; Rubén Artero; Dalia Maria Lucia Asaro; Michael Aschner; Milad Ashrafizadeh; Osnat Ashur-Fabian; Atanas G Atanasov; Alicia K Au; Patrick Auberger; Holger W Auner; Laure Aurelian; Riccardo Autelli; Laura Avagliano; Yenniffer Ávalos; Sanja Aveic; Célia Alexandra Aveleira; Tamar Avin-Wittenberg; Yucel Aydin; Scott Ayton; Srinivas Ayyadevara; Maria Azzopardi; Misuzu Baba; Jonathan M Backer; Steven K Backues; Dong-Hun Bae; 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Naïma Belgareh-Touzé; Cristina Bellarosa; Francesca Belleudi; Melissa Belló Pérez; Raquel Bello-Morales; Jackeline Soares de Oliveira Beltran; Sebastián Beltran; Doris Mangiaracina Benbrook; Mykolas Bendorius; Bruno A Benitez; Irene Benito-Cuesta; Julien Bensalem; Martin W Berchtold; Sabina Berezowska; Daniele Bergamaschi; Matteo Bergami; Andreas Bergmann; Laura Berliocchi; Clarisse Berlioz-Torrent; Amélie Bernard; Lionel Berthoux; Cagri G Besirli; Sebastien Besteiro; Virginie M Betin; Rudi Beyaert; Jelena S Bezbradica; Kiran Bhaskar; Ingrid Bhatia-Kissova; Resham Bhattacharya; Sujoy Bhattacharya; Shalmoli Bhattacharyya; Md Shenuarin Bhuiyan; Sujit Kumar Bhutia; Lanrong Bi; Xiaolin Bi; Trevor J Biden; Krikor Bijian; Viktor A Billes; Nadine Binart; Claudia Bincoletto; Asa B Birgisdottir; Geir Bjorkoy; Gonzalo Blanco; Ana Blas-Garcia; Janusz Blasiak; Robert Blomgran; Klas Blomgren; Janice S Blum; Emilio Boada-Romero; Mirta Boban; Kathleen Boesze-Battaglia; Philippe Boeuf; Barry Boland; Pascale Bomont; Paolo Bonaldo; Srinivasa Reddy Bonam; Laura Bonfili; Juan S Bonifacino; Brian A Boone; Martin D Bootman; Matteo Bordi; Christoph Borner; Beat C Bornhauser; Gautam Borthakur; Jürgen Bosch; Santanu Bose; Luis M Botana; Juan Botas; Chantal M Boulanger; Michael E Boulton; Mathieu Bourdenx; Benjamin Bourgeois; Nollaig M Bourke; Guilhem Bousquet; Patricia Boya; Peter V Bozhkov; Luiz H M Bozi; Tolga O Bozkurt; Doug E Brackney; Christian H Brandts; Ralf J Braun; Gerhard H Braus; Roberto Bravo-Sagua; José M Bravo-San Pedro; Patrick Brest; Marie-Agnès Bringer; Alfredo Briones-Herrera; V Courtney Broaddus; Peter Brodersen; Jeffrey L Brodsky; Steven L Brody; Paola G Bronson; Jeff M Bronstein; Carolyn N Brown; Rhoderick E Brown; Patricia C Brum; John H Brumell; Nicola Brunetti-Pierri; Daniele Bruno; Robert J Bryson-Richardson; Cecilia Bucci; Carmen Buchrieser; Marta Bueno; Laura Elisa Buitrago-Molina; Simone Buraschi; Shilpa Buch; J Ross Buchan; Erin M Buckingham; Hikmet Budak; Mauricio Budini; Geert Bultynck; Florin Burada; Joseph R Burgoyne; M Isabel Burón; Victor Bustos; Sabrina Büttner; Elena Butturini; Aaron Byrd; Isabel Cabas; Sandra Cabrera-Benitez; Ken Cadwell; Jingjing Cai; Lu Cai; Qian Cai; Montserrat Cairó; Jose A Calbet; Guy A Caldwell; Kim A Caldwell; Jarrod A Call; Riccardo Calvani; Ana C Calvo; Miguel Calvo-Rubio Barrera; Niels Os Camara; Jacques H Camonis; Nadine Camougrand; Michelangelo Campanella; Edward M Campbell; François-Xavier Campbell-Valois; Silvia Campello; Ilaria Campesi; Juliane C Campos; Olivier Camuzard; Jorge Cancino; Danilo Candido de Almeida; Laura Canesi; Isabella Caniggia; Barbara Canonico; Carles Cantí; Bin Cao; Michele Caraglia; Beatriz Caramés; Evie H Carchman; Elena Cardenal-Muñoz; Cesar Cardenas; Luis Cardenas; Sandra M Cardoso; Jennifer S Carew; Georges F Carle; Gillian Carleton; Silvia Carloni; Didac Carmona-Gutierrez; Leticia A Carneiro; Oliana Carnevali; Julian M Carosi; Serena Carra; Alice Carrier; Lucie Carrier; Bernadette Carroll; A Brent Carter; Andreia Neves Carvalho; Magali Casanova; Caty Casas; Josefina Casas; Chiara Cassioli; Eliseo F Castillo; Karen Castillo; Sonia Castillo-Lluva; Francesca Castoldi; Marco Castori; Ariel F Castro; Margarida Castro-Caldas; Javier Castro-Hernandez; Susana Castro-Obregon; Sergio D Catz; Claudia Cavadas; Federica Cavaliere; Gabriella Cavallini; Maria Cavinato; Maria L Cayuela; Paula Cebollada Rica; Valentina Cecarini; Francesco Cecconi; Marzanna Cechowska-Pasko; Simone Cenci; Victòria Ceperuelo-Mallafré; João J Cerqueira; Janete M Cerutti; Davide Cervia; Vildan Bozok Cetintas; Silvia Cetrullo; Han-Jung Chae; Andrei S Chagin; Chee-Yin Chai; Gopal Chakrabarti; Oishee Chakrabarti; Tapas Chakraborty; Trinad Chakraborty; Mounia Chami; Georgios Chamilos; David W Chan; Edmond Y W Chan; Edward D Chan; H Y Edwin Chan; Helen H Chan; Hung Chan; Matthew T V Chan; Yau Sang Chan; Partha K Chandra; Chih-Peng Chang; Chunmei Chang; Hao-Chun Chang; Kai Chang; Jie Chao; Tracey Chapman; Nicolas Charlet-Berguerand; Samrat Chatterjee; Shail K Chaube; Anu Chaudhary; Santosh Chauhan; Edward Chaum; Frédéric Checler; Michael E Cheetham; Chang-Shi Chen; Guang-Chao Chen; Jian-Fu Chen; Liam L Chen; Leilei Chen; Lin Chen; Mingliang Chen; Mu-Kuan Chen; Ning Chen; Quan Chen; Ruey-Hwa Chen; Shi Chen; Wei Chen; Weiqiang Chen; Xin-Ming Chen; Xiong-Wen Chen; Xu Chen; Yan Chen; Ye-Guang Chen; Yingyu Chen; Yongqiang Chen; Yu-Jen Chen; Yue-Qin Chen; Zhefan Stephen Chen; Zhi Chen; Zhi-Hua Chen; Zhijian J Chen; Zhixiang Chen; Hanhua Cheng; Jun Cheng; Shi-Yuan Cheng; Wei Cheng; Xiaodong Cheng; Xiu-Tang Cheng; Yiyun Cheng; Zhiyong Cheng; Zhong Chen; Heesun Cheong; Jit Kong Cheong; Boris V Chernyak; Sara Cherry; Chi Fai Randy Cheung; Chun Hei Antonio Cheung; King-Ho Cheung; Eric Chevet; Richard J Chi; Alan Kwok Shing Chiang; Ferdinando Chiaradonna; Roberto Chiarelli; Mario Chiariello; Nathalia Chica; Susanna Chiocca; Mario Chiong; Shih-Hwa Chiou; Abhilash I Chiramel; Valerio Chiurchiù; Dong-Hyung Cho; Seong-Kyu Choe; Augustine M K Choi; Mary E Choi; Kamalika Roy Choudhury; Norman S Chow; Charleen T Chu; Jason P Chua; John Jia En Chua; Hyewon Chung; Kin Pan Chung; Seockhoon Chung; So-Hyang Chung; Yuen-Li Chung; Valentina Cianfanelli; Iwona A Ciechomska; Mariana Cifuentes; Laura Cinque; Sebahattin Cirak; Mara Cirone; Michael J Clague; Robert Clarke; Emilio Clementi; Eliana M Coccia; Patrice Codogno; Ehud Cohen; Mickael M Cohen; Tania Colasanti; Fiorella Colasuonno; Robert A Colbert; Anna Colell; Miodrag Čolić; Nuria S Coll; Mark O Collins; María I Colombo; Daniel A Colón-Ramos; Lydie Combaret; Sergio Comincini; Márcia R Cominetti; Antonella Consiglio; Andrea Conte; Fabrizio Conti; Viorica Raluca Contu; Mark R Cookson; Kevin M Coombs; Isabelle Coppens; Maria Tiziana Corasaniti; Dale P Corkery; Nils Cordes; Katia Cortese; Maria do Carmo Costa; Sarah Costantino; Paola Costelli; Ana Coto-Montes; Peter J Crack; Jose L Crespo; Alfredo Criollo; Valeria Crippa; Riccardo Cristofani; Tamas Csizmadia; Antonio Cuadrado; Bing Cui; Jun Cui; Yixian Cui; Yong Cui; Emmanuel Culetto; Andrea C Cumino; Andrey V Cybulsky; Mark J Czaja; Stanislaw J Czuczwar; Stefania D'Adamo; Marcello D'Amelio; Daniela D'Arcangelo; Andrew C D'Lugos; Gabriella D'Orazi; James A da Silva; Hormos Salimi Dafsari; Ruben K Dagda; Yasin Dagdas; Maria Daglia; Xiaoxia Dai; Yun Dai; Yuyuan Dai; Jessica Dal Col; Paul Dalhaimer; Luisa Dalla Valle; Tobias Dallenga; Guillaume Dalmasso; Markus Damme; Ilaria Dando; Nico P Dantuma; April L Darling; Hiranmoy Das; Srinivasan Dasarathy; Santosh K Dasari; Srikanta Dash; Oliver Daumke; Adrian N Dauphinee; Jeffrey S Davies; Valeria A Dávila; Roger J Davis; Tanja Davis; Sharadha Dayalan Naidu; Francesca De Amicis; Karolien De Bosscher; Francesca De Felice; Lucia De Franceschi; Chiara De Leonibus; Mayara G de Mattos Barbosa; Guido R Y De Meyer; Angelo De Milito; Cosimo De Nunzio; Clara De Palma; Mauro De Santi; Claudio De Virgilio; Daniela De Zio; Jayanta Debnath; Brian J DeBosch; Jean-Paul Decuypere; Mark A Deehan; Gianluca Deflorian; James DeGregori; Benjamin Dehay; Gabriel Del Rio; Joe R Delaney; Lea M D Delbridge; Elizabeth Delorme-Axford; M Victoria Delpino; Francesca Demarchi; Vilma Dembitz; Nicholas D Demers; Hongbin Deng; Zhiqiang Deng; Joern Dengjel; Paul Dent; Donna Denton; Melvin L DePamphilis; Channing J Der; Vojo Deretic; Albert Descoteaux; Laura Devis; Sushil Devkota; Olivier Devuyst; Grant Dewson; Mahendiran Dharmasivam; Rohan Dhiman; Diego di Bernardo; Manlio Di Cristina; Fabio Di Domenico; Pietro Di Fazio; Alessio Di Fonzo; Giovanni Di Guardo; Gianni M Di Guglielmo; Luca Di Leo; Chiara Di Malta; Alessia Di Nardo; Martina Di Rienzo; Federica Di Sano; George Diallinas; Jiajie Diao; Guillermo Diaz-Araya; Inés Díaz-Laviada; Jared M Dickinson; Marc Diederich; Mélanie Dieudé; Ivan Dikic; Shiping Ding; Wen-Xing Ding; Luciana Dini; Jelena Dinić; Miroslav Dinic; Albena T Dinkova-Kostova; Marc S Dionne; Jörg H W Distler; Abhinav Diwan; Ian M C Dixon; Mojgan Djavaheri-Mergny; Ina Dobrinski; Oxana Dobrovinskaya; Radek Dobrowolski; Renwick C J Dobson; Jelena Đokić; Serap Dokmeci Emre; Massimo Donadelli; Bo Dong; Xiaonan Dong; Zhiwu Dong; Gerald W Dorn Ii; Volker Dotsch; Huan Dou; Juan Dou; Moataz Dowaidar; Sami Dridi; Liat Drucker; Ailian Du; Caigan Du; Guangwei Du; Hai-Ning Du; Li-Lin Du; André du Toit; Shao-Bin Duan; Xiaoqiong Duan; Sónia P Duarte; Anna Dubrovska; Elaine A Dunlop; Nicolas Dupont; Raúl V Durán; Bilikere S Dwarakanath; Sergey A Dyshlovoy; Darius Ebrahimi-Fakhari; Leopold Eckhart; Charles L Edelstein; Thomas Efferth; Eftekhar Eftekharpour; Ludwig Eichinger; Nabil Eid; Tobias Eisenberg; N Tony Eissa; Sanaa Eissa; Miriam Ejarque; Abdeljabar El Andaloussi; Nazira El-Hage; Shahenda El-Naggar; Anna Maria Eleuteri; Eman S El-Shafey; Mohamed Elgendy; Aristides G Eliopoulos; María M Elizalde; Philip M Elks; Hans-Peter Elsasser; Eslam S Elsherbiny; Brooke M Emerling; N C Tolga Emre; Christina H Eng; Nikolai Engedal; Anna-Mart Engelbrecht; Agnete S T Engelsen; Jorrit M Enserink; Ricardo Escalante; Audrey Esclatine; Mafalda Escobar-Henriques; Eeva-Liisa Eskelinen; Lucile Espert; Makandjou-Ola Eusebio; Gemma Fabrias; Cinzia Fabrizi; Antonio Facchiano; Francesco Facchiano; Bengt Fadeel; Claudio Fader; Alex C Faesen; W Douglas Fairlie; Alberto Falcó; Bjorn H Falkenburger; Daping Fan; Jie Fan; Yanbo Fan; Evandro F Fang; Yanshan Fang; Yognqi Fang; Manolis Fanto; Tamar Farfel-Becker; Mathias Faure; Gholamreza Fazeli; Anthony O Fedele; Arthur M Feldman; Du Feng; Jiachun Feng; Lifeng Feng; Yibin Feng; Yuchen Feng; Wei Feng; Thais Fenz Araujo; Thomas A Ferguson; Álvaro F Fernández; Jose C Fernandez-Checa; Sonia Fernández-Veledo; Alisdair R Fernie; Anthony W Ferrante; Alessandra Ferraresi; Merari F Ferrari; Julio C B Ferreira; Susan Ferro-Novick; Antonio Figueras; Riccardo Filadi; Nicoletta Filigheddu; Eduardo Filippi-Chiela; Giuseppe Filomeni; Gian Maria Fimia; Vittorio Fineschi; Francesca Finetti; Steven Finkbeiner; Edward A Fisher; Paul B Fisher; Flavio Flamigni; Steven J Fliesler; Trude H Flo; Ida Florance; Oliver Florey; Tullio Florio; Erika Fodor; Carlo Follo; Edward A Fon; Antonella Forlino; Francesco Fornai; Paola Fortini; Anna Fracassi; Alessandro Fraldi; Brunella Franco; Rodrigo Franco; Flavia Franconi; Lisa B Frankel; Scott L Friedman; Leopold F Fröhlich; Gema Frühbeck; Jose M Fuentes; Yukio Fujiki; Naonobu Fujita; Yuuki Fujiwara; Mitsunori Fukuda; Simone Fulda; Luc Furic; Norihiko Furuya; Carmela Fusco; Michaela U Gack; Lidia Gaffke; Sehamuddin Galadari; Alessia Galasso; Maria F Galindo; Sachith Gallolu Kankanamalage; Lorenzo Galluzzi; Vincent Galy; Noor Gammoh; Boyi Gan; Ian G Ganley; Feng Gao; Hui Gao; Minghui Gao; Ping Gao; Shou-Jiang Gao; Wentao Gao; Xiaobo Gao; Ana Garcera; Maria Noé Garcia; Verónica E Garcia; Francisco García-Del Portillo; Vega Garcia-Escudero; Aracely Garcia-Garcia; Marina Garcia-Macia; Diana García-Moreno; Carmen Garcia-Ruiz; Patricia García-Sanz; Abhishek D Garg; Ricardo Gargini; Tina Garofalo; Robert F Garry; Nils C Gassen; Damian Gatica; Liang Ge; Wanzhong Ge; Ruth Geiss-Friedlander; Cecilia Gelfi; Pascal Genschik; Ian E Gentle; Valeria Gerbino; Christoph Gerhardt; Kyla Germain; Marc Germain; David A Gewirtz; Elham Ghasemipour Afshar; Saeid Ghavami; Alessandra Ghigo; Manosij Ghosh; Georgios Giamas; Claudia Giampietri; Alexandra Giatromanolaki; Gary E Gibson; Spencer B Gibson; Vanessa Ginet; Edward Giniger; Carlotta Giorgi; Henrique Girao; Stephen E Girardin; Mridhula Giridharan; Sandy Giuliano; Cecilia Giulivi; Sylvie Giuriato; Julien Giustiniani; Alexander Gluschko; Veit Goder; Alexander Goginashvili; Jakub Golab; David C Goldstone; Anna Golebiewska; Luciana R Gomes; Rodrigo Gomez; Rubén Gómez-Sánchez; Maria Catalina Gomez-Puerto; Raquel Gomez-Sintes; Qingqiu Gong; Felix M Goni; Javier González-Gallego; Tomas Gonzalez-Hernandez; Rosa A Gonzalez-Polo; Jose A Gonzalez-Reyes; Patricia González-Rodríguez; Ing Swie Goping; Marina S Gorbatyuk; Nikolai V Gorbunov; Kıvanç Görgülü; Roxana M Gorojod; Sharon M Gorski; Sandro Goruppi; Cecilia Gotor; Roberta A Gottlieb; Illana Gozes; Devrim Gozuacik; Martin Graef; Markus H Gräler; Veronica Granatiero; Daniel Grasso; Joshua P Gray; Douglas R Green; Alexander Greenhough; Stephen L Gregory; Edward F Griffin; Mark W Grinstaff; Frederic Gros; Charles Grose; Angelina S Gross; Florian Gruber; Paolo Grumati; Tilman Grune; Xueyan Gu; Jun-Lin Guan; Carlos M Guardia; Kishore Guda; Flora Guerra; Consuelo Guerri; Prasun Guha; Carlos Guillén; Shashi Gujar; Anna Gukovskaya; Ilya Gukovsky; Jan Gunst; Andreas Günther; Anyonya R Guntur; Chuanyong Guo; Chun Guo; Hongqing Guo; Lian-Wang Guo; Ming Guo; Pawan Gupta; Shashi Kumar Gupta; Swapnil Gupta; Veer Bala Gupta; Vivek Gupta; Asa B Gustafsson; David D Gutterman; Ranjitha H B; Annakaisa Haapasalo; James E Haber; Aleksandra Hać; Shinji Hadano; Anders J Hafrén; Mansour Haidar; Belinda S Hall; Gunnel Halldén; Anne Hamacher-Brady; Andrea Hamann; Maho Hamasaki; Weidong Han; Malene Hansen; Phyllis I Hanson; Zijian Hao; Masaru Harada; Ljubica Harhaji-Trajkovic; Nirmala Hariharan; Nigil Haroon; James Harris; Takafumi Hasegawa; Noor Hasima Nagoor; Jeffrey A Haspel; Volker Haucke; Wayne D Hawkins; Bruce A Hay; Cole M Haynes; Soren B Hayrabedyan; Thomas S Hays; Congcong He; Qin He; Rong-Rong He; You-Wen He; Yu-Ying He; Yasser Heakal; Alexander M Heberle; J Fielding Hejtmancik; Gudmundur Vignir Helgason; Vanessa Henkel; Marc Herb; Alexander Hergovich; Anna Herman-Antosiewicz; Agustín Hernández; Carlos Hernandez; Sergio Hernandez-Diaz; Virginia Hernandez-Gea; Amaury Herpin; Judit Herreros; Javier H Hervás; Daniel Hesselson; Claudio Hetz; Volker T Heussler; Yujiro Higuchi; Sabine Hilfiker; Joseph A Hill; William S Hlavacek; Emmanuel A Ho; Idy H T Ho; Philip Wing-Lok Ho; Shu-Leong Ho; Wan Yun Ho; G Aaron Hobbs; Mark Hochstrasser; Peter H M Hoet; Daniel Hofius; Paul Hofman; Annika Höhn; Carina I Holmberg; Jose R Hombrebueno; Chang-Won Hong Yi-Ren Hong; Lora V Hooper; Thorsten Hoppe; Rastislav Horos; Yujin Hoshida; I-Lun Hsin; Hsin-Yun Hsu; Bing Hu; Dong Hu; Li-Fang Hu; Ming Chang Hu; Ronggui Hu; Wei Hu; Yu-Chen Hu; Zhuo-Wei Hu; Fang Hua; Jinlian Hua; Yingqi Hua; Chongmin Huan; Canhua Huang; Chuanshu Huang; Chuanxin Huang; Chunling Huang; Haishan Huang; Kun Huang; Michael L H Huang; Rui Huang; Shan Huang; Tianzhi Huang; Xing Huang; Yuxiang Jack Huang; Tobias B Huber; Virginie Hubert; Christian A Hubner; Stephanie M Hughes; William E Hughes; Magali Humbert; Gerhard Hummer; James H Hurley; Sabah Hussain; Salik Hussain; Patrick J Hussey; Martina Hutabarat; Hui-Yun Hwang; Seungmin Hwang; Antonio Ieni; Fumiyo Ikeda; Yusuke Imagawa; Yuzuru Imai; Carol Imbriano; Masaya Imoto; Denise M Inman; Ken Inoki; Juan Iovanna; Renato V Iozzo; Giuseppe Ippolito; Javier E Irazoqui; Pablo Iribarren; Mohd Ishaq; Makoto Ishikawa; Nestor Ishimwe; Ciro Isidoro; Nahed Ismail; Shohreh Issazadeh-Navikas; Eisuke Itakura; Daisuke Ito; Davor Ivankovic; Saška Ivanova; Anand Krishnan V Iyer; José M Izquierdo; Masanori Izumi; Marja Jäättelä; Majid Sakhi Jabir; William T Jackson; Nadia Jacobo-Herrera; Anne-Claire Jacomin; Elise Jacquin; Pooja Jadiya; Hartmut Jaeschke; Chinnaswamy Jagannath; Arjen J Jakobi; Johan Jakobsson; Bassam Janji; Pidder Jansen-Dürr; Patric J Jansson; Jonathan Jantsch; Sławomir Januszewski; Alagie Jassey; Steve Jean; Hélène Jeltsch-David; Pavla Jendelova; Andreas Jenny; Thomas E Jensen; Niels Jessen; Jenna L Jewell; Jing Ji; Lijun Jia; Rui Jia; Liwen Jiang; Qing Jiang; Richeng Jiang; Teng Jiang; Xuejun Jiang; Yu Jiang; Maria Jimenez-Sanchez; Eun-Jung Jin; Fengyan Jin; Hongchuan Jin; Li Jin; Luqi Jin; Meiyan Jin; Si Jin; Eun-Kyeong Jo; Carine Joffre; Terje Johansen; Gail V W Johnson; Simon A Johnston; Eija Jokitalo; Mohit Kumar Jolly; Leo A B Joosten; Joaquin Jordan; Bertrand Joseph; Dianwen Ju; Jeong-Sun Ju; Jingfang Ju; Esmeralda Juárez; Delphine Judith; Gábor Juhász; Youngsoo Jun; Chang Hwa Jung; Sung-Chul Jung; Yong Keun Jung; Heinz Jungbluth; Johannes Jungverdorben; Steffen Just; Kai Kaarniranta; Allen Kaasik; Tomohiro Kabuta; Daniel Kaganovich; Alon Kahana; Renate Kain; Shinjo Kajimura; Maria Kalamvoki; Manjula Kalia; Danuta S Kalinowski; Nina Kaludercic; Ioanna Kalvari; Joanna Kaminska; Vitaliy O Kaminskyy; Hiromitsu Kanamori; Keizo Kanasaki; Chanhee Kang; Rui Kang; Sang Sun Kang; Senthilvelrajan Kaniyappan; Tomotake Kanki; Thirumala-Devi Kanneganti; Anumantha G Kanthasamy; Arthi Kanthasamy; Marc Kantorow; Orsolya Kapuy; Michalis V Karamouzis; Md Razaul Karim; Parimal Karmakar; Rajesh G Katare; Masaru Kato; Stefan H E Kaufmann; Anu Kauppinen; Gur P Kaushal; Susmita Kaushik; Kiyoshi Kawasaki; Kemal Kazan; Po-Yuan Ke; Damien J Keating; Ursula Keber; John H Kehrl; Kate E Keller; Christian W Keller; Jongsook Kim Kemper; Candia M Kenific; Oliver Kepp; Stephanie Kermorgant; Andreas Kern; Robin Ketteler; Tom G Keulers; Boris Khalfin; Hany Khalil; Bilon Khambu; Shahid Y Khan; Vinoth Kumar Megraj Khandelwal; Rekha Khandia; Widuri Kho; Noopur V Khobrekar; Sataree Khuansuwan; Mukhran Khundadze; Samuel A Killackey; Dasol Kim; Deok Ryong Kim; Do-Hyung Kim; Dong-Eun Kim; Eun Young Kim; Eun-Kyoung Kim; Hak-Rim Kim; Hee-Sik Kim; Jeong Hun Kim; Jin Kyung Kim; Jin-Hoi Kim; Joungmok Kim; Ju Hwan Kim; Keun Il Kim; Peter K Kim; Seong-Jun Kim; Scot R Kimball; Adi Kimchi; Alec C Kimmelman; Tomonori Kimura; Matthew A King; Kerri J Kinghorn; Conan G Kinsey; Vladimir Kirkin; Lorrie A Kirshenbaum; Sergey L Kiselev; Shuji Kishi; Katsuhiko Kitamoto; Yasushi Kitaoka; Kaio Kitazato; Richard N Kitsis; Josef T Kittler; Ole Kjaerulff; Peter S Klein; Thomas Klopstock; Jochen Klucken; Helene Knævelsrud; Roland L Knorr; Ben C B Ko; Fred Ko; Jiunn-Liang Ko; Hotaka Kobayashi; Satoru Kobayashi; Ina Koch; Jan C Koch; Ulrich Koenig; Donat Kögel; Young Ho Koh; Masato Koike; Sepp D Kohlwein; Nur M Kocaturk; Masaaki Komatsu; Jeannette König; Toru Kono; Benjamin T Kopp; Tamas Korcsmaros; Gözde Korkmaz; Viktor I Korolchuk; Mónica Suárez Korsnes; Ali Koskela; Janaiah Kota; Yaichiro Kotake; Monica L Kotler; Yanjun Kou; Michael I Koukourakis; Evangelos Koustas; Attila L Kovacs; Tibor Kovács; Daisuke Koya; Tomohiro Kozako; Claudine Kraft; Dimitri Krainc; Helmut Krämer; Anna D Krasnodembskaya; Carole Kretz-Remy; Guido Kroemer; Nicholas T Ktistakis; Kazuyuki Kuchitsu; Sabine Kuenen; Lars Kuerschner; Thomas Kukar; Ajay Kumar; Ashok Kumar; Deepak Kumar; Dhiraj Kumar; Sharad Kumar; Shinji Kume; Caroline Kumsta; Chanakya N Kundu; Mondira Kundu; Ajaikumar B Kunnumakkara; Lukasz Kurgan; Tatiana G Kutateladze; Ozlem Kutlu; SeongAe Kwak; Ho Jeong Kwon; Taeg Kyu Kwon; Yong Tae Kwon; Irene Kyrmizi; Albert La Spada; Patrick Labonté; Sylvain Ladoire; Ilaria Laface; Frank Lafont; Diane C Lagace; Vikramjit Lahiri; Zhibing Lai; Angela S Laird; Aparna Lakkaraju; Trond Lamark; Sheng-Hui Lan; Ane Landajuela; Darius J R Lane; Jon D Lane; Charles H Lang; Carsten Lange; Ülo Langel; Rupert Langer; Pierre Lapaquette; Jocelyn Laporte; Nicholas F LaRusso; Isabel Lastres-Becker; Wilson Chun Yu Lau; Gordon W Laurie; Sergio Lavandero; Betty Yuen Kwan Law; Helen Ka-Wai Law; Rob Layfield; Weidong Le; Herve Le Stunff; Alexandre Y Leary; Jean-Jacques Lebrun; Lionel Y W Leck; Jean-Philippe Leduc-Gaudet; Changwook Lee; Chung-Pei Lee; Da-Hye Lee; Edward B Lee; Erinna F Lee; Gyun Min Lee; He-Jin Lee; Heung Kyu Lee; Jae Man Lee; Jason S Lee; Jin-A Lee; Joo-Yong Lee; Jun Hee Lee; Michael Lee; Min Goo Lee; Min Jae Lee; Myung-Shik Lee; Sang Yoon Lee; Seung-Jae Lee; Stella Y Lee; Sung Bae Lee; Won Hee Lee; Ying-Ray Lee; Yong-Ho Lee; Youngil Lee; Christophe Lefebvre; Renaud Legouis; Yu L Lei; Yuchen Lei; Sergey Leikin; Gerd Leitinger; Leticia Lemus; Shuilong Leng; Olivia Lenoir; Guido Lenz; Heinz Josef Lenz; Paola Lenzi; Yolanda León; Andréia M Leopoldino; Christoph Leschczyk; Stina Leskelä; Elisabeth Letellier; Chi-Ting Leung; Po Sing Leung; Jeremy S Leventhal; Beth Levine; Patrick A Lewis; Klaus Ley; Bin Li; Da-Qiang Li; Jianming Li; Jing Li; Jiong Li; Ke Li; Liwu Li; Mei Li; Min Li; Min Li; Ming Li; Mingchuan Li; Pin-Lan Li; Ming-Qing Li; Qing Li; Sheng Li; Tiangang Li; Wei Li; Wenming Li; Xue Li; Yi-Ping Li; Yuan Li; Zhiqiang Li; Zhiyong Li; Zhiyuan Li; Jiqin Lian; Chengyu Liang; Qiangrong Liang; Weicheng Liang; Yongheng Liang; YongTian Liang; Guanghong Liao; Lujian Liao; Mingzhi Liao; Yung-Feng Liao; Mariangela Librizzi; Pearl P Y Lie; Mary A Lilly; Hyunjung J Lim; Thania R R Lima; Federica Limana; Chao Lin; Chih-Wen Lin; Dar-Shong Lin; Fu-Cheng Lin; Jiandie D Lin; Kurt M Lin; Kwang-Huei Lin; Liang-Tzung Lin; Pei-Hui Lin; Qiong Lin; Shaofeng Lin; Su-Ju Lin; Wenyu Lin; Xueying Lin; Yao-Xin Lin; Yee-Shin Lin; Rafael Linden; Paula Lindner; Shuo-Chien Ling; Paul Lingor; Amelia K Linnemann; Yih-Cherng Liou; Marta M Lipinski; Saška Lipovšek; Vitor A Lira; Natalia Lisiak; Paloma B Liton; Chao Liu; Ching-Hsuan Liu; Chun-Feng Liu; Cui Hua Liu; Fang Liu; Hao Liu; Hsiao-Sheng Liu; Hua-Feng Liu; Huifang Liu; Jia Liu; Jing Liu; Julia Liu; Leyuan Liu; Longhua Liu; Meilian Liu; Qin Liu; Wei Liu; Wende Liu; Xiao-Hong Liu; Xiaodong Liu; Xingguo Liu; Xu Liu; Xuedong Liu; Yanfen Liu; Yang Liu; Yang Liu; Yueyang Liu; Yule Liu; J Andrew Livingston; Gerard Lizard; Jose M Lizcano; Senka Ljubojevic-Holzer; Matilde E LLeonart; David Llobet-Navàs; Alicia Llorente; Chih Hung Lo; Damián Lobato-Márquez; Qi Long; Yun Chau Long; Ben Loos; Julia A Loos; Manuela G López; Guillermo López-Doménech; José Antonio López-Guerrero; Ana T López-Jiménez; Óscar López-Pérez; Israel López-Valero; Magdalena J Lorenowicz; Mar Lorente; Peter Lorincz; Laura Lossi; Sophie Lotersztajn; Penny E Lovat; Jonathan F Lovell; Alenka Lovy; Péter Lőw; Guang Lu; Haocheng Lu; Jia-Hong Lu; Jin-Jian Lu; Mengji Lu; Shuyan Lu; Alessandro Luciani; John M Lucocq; Paula Ludovico; Micah A Luftig; Morten Luhr; Diego Luis-Ravelo; Julian J Lum; Liany Luna-Dulcey; Anders H Lund; Viktor K Lund; Jan D Lünemann; Patrick Lüningschrör; Honglin Luo; Rongcan Luo; Shouqing Luo; Zhi Luo; Claudio Luparello; Bernhard Lüscher; Luan Luu; Alex Lyakhovich; Konstantin G Lyamzaev; Alf Håkon Lystad; Lyubomyr Lytvynchuk; Alvin C Ma; Changle Ma; Mengxiao Ma; Ning-Fang Ma; Quan-Hong Ma; Xinliang Ma; Yueyun Ma; Zhenyi Ma; Ormond A MacDougald; Fernando Macian; Gustavo C MacIntosh; Jeffrey P MacKeigan; Kay F Macleod; Sandra Maday; Frank Madeo; Muniswamy Madesh; Tobias Madl; Julio Madrigal-Matute; Akiko Maeda; Yasuhiro Maejima; Marta Magarinos; Poornima Mahavadi; Emiliano Maiani; Kenneth Maiese; Panchanan Maiti; Maria Chiara Maiuri; Barbara Majello; Michael B Major; Elena Makareeva; Fayaz Malik; Karthik Mallilankaraman; Walter Malorni; Alina Maloyan; Najiba Mammadova; Gene Chi Wai Man; Federico Manai; Joseph D Mancias; Eva-Maria Mandelkow; Michael A Mandell; Angelo A Manfredi; Masoud H Manjili; Ravi Manjithaya; Patricio Manque; Bella B Manshian; Raquel Manzano; Claudia Manzoni; Kai Mao; Cinzia Marchese; Sandrine Marchetti; Anna Maria Marconi; Fabrizio Marcucci; Stefania Mardente; Olga A Mareninova; Marta Margeta; Muriel Mari; Sara Marinelli; Oliviero Marinelli; Guillermo Mariño; Sofia Mariotto; Richard S Marshall; Mark R Marten; Sascha Martens; Alexandre P J Martin; Katie R Martin; Sara Martin; Shaun Martin; Adrián Martín-Segura; Miguel A Martín-Acebes; Inmaculada Martin-Burriel; Marcos Martin-Rincon; Paloma Martin-Sanz; José A Martina; Wim Martinet; Aitor Martinez; Ana Martinez; Jennifer Martinez; Moises Martinez Velazquez; Nuria Martinez-Lopez; Marta Martinez-Vicente; Daniel O Martins; Joilson O Martins; Waleska K Martins; Tania Martins-Marques; Emanuele Marzetti; Shashank Masaldan; Celine Masclaux-Daubresse; Douglas G Mashek; Valentina Massa; Lourdes Massieu; Glenn R Masson; Laura Masuelli; Anatoliy I Masyuk; Tetyana V Masyuk; Paola Matarrese; Ander Matheu; Satoaki Matoba; Sachiko Matsuzaki; Pamela Mattar; Alessandro Matte; Domenico Mattoscio; José L Mauriz; Mario Mauthe; Caroline Mauvezin; Emanual Maverakis; Paola Maycotte; Johanna Mayer; Gianluigi Mazzoccoli; Cristina Mazzoni; Joseph R Mazzulli; Nami McCarty; Christine McDonald; Mitchell R McGill; Sharon L McKenna; BethAnn McLaughlin; Fionn McLoughlin; Mark A McNiven; Thomas G McWilliams; Fatima Mechta-Grigoriou; Tania Catarina Medeiros; Diego L Medina; Lynn A Megeney; Klara Megyeri; Maryam Mehrpour; Jawahar L Mehta; Alfred J Meijer; Annemarie H Meijer; Jakob Mejlvang; Alicia Meléndez; Annette Melk; Gonen Memisoglu; Alexandrina F Mendes; Delong Meng; Fei Meng; Tian Meng; Rubem Menna-Barreto; Manoj B Menon; Carol Mercer; Anne E Mercier; Jean-Louis Mergny; Adalberto Merighi; Seth D Merkley; Giuseppe Merla; Volker Meske; Ana Cecilia Mestre; Shree Padma Metur; Christian Meyer; Hemmo Meyer; Wenyi Mi; Jeanne Mialet-Perez; Junying Miao; Lucia Micale; Yasuo Miki; Enrico Milan; Małgorzata Milczarek; Dana L Miller; Samuel I Miller; Silke Miller; Steven W Millward; Ira Milosevic; Elena A Minina; Hamed Mirzaei; Hamid Reza Mirzaei; Mehdi Mirzaei; Amit Mishra; Nandita Mishra; Paras Kumar Mishra; Maja Misirkic Marjanovic; Roberta Misasi; Amit Misra; Gabriella Misso; Claire Mitchell; Geraldine Mitou; Tetsuji Miura; Shigeki Miyamoto; Makoto Miyazaki; Mitsunori Miyazaki; Taiga Miyazaki; Keisuke Miyazawa; Noboru Mizushima; Trine H Mogensen; Baharia Mograbi; Reza Mohammadinejad; Yasir Mohamud; Abhishek Mohanty; Sipra Mohapatra; Torsten Möhlmann; Asif Mohmmed; Anna Moles; Kelle H Moley; Maurizio Molinari; Vincenzo Mollace; Andreas Buch Møller; Bertrand Mollereau; Faustino Mollinedo; Costanza Montagna; Mervyn J Monteiro; Andrea Montella; L Ruth Montes; Barbara Montico; Vinod K Mony; Giacomo Monzio Compagnoni; Michael N Moore; Mohammad A Moosavi; Ana L Mora; Marina Mora; David Morales-Alamo; Rosario Moratalla; Paula I Moreira; Elena Morelli; Sandra Moreno; Daniel Moreno-Blas; Viviana Moresi; Benjamin Morga; Alwena H Morgan; Fabrice Morin; Hideaki Morishita; Orson L Moritz; Mariko Moriyama; Yuji Moriyasu; Manuela Morleo; Eugenia Morselli; Jose F Moruno-Manchon; Jorge Moscat; Serge Mostowy; Elisa Motori; Andrea Felinto Moura; Naima Moustaid-Moussa; Maria Mrakovcic; Gabriel Muciño-Hernández; Anupam Mukherjee; Subhadip Mukhopadhyay; Jean M Mulcahy Levy; Victoriano Mulero; Sylviane Muller; Christian Münch; Ashok Munjal; Pura Munoz-Canoves; Teresa Muñoz-Galdeano; Christian Münz; Tomokazu Murakawa; Claudia Muratori; Brona M Murphy; J Patrick Murphy; Aditya Murthy; Timo T Myöhänen; Indira U Mysorekar; Jennifer Mytych; Seyed Mohammad Nabavi; Massimo Nabissi; Péter Nagy; Jihoon Nah; Aimable Nahimana; Ichiro Nakagawa; Ken Nakamura; Hitoshi Nakatogawa; Shyam S Nandi; Meera Nanjundan; Monica Nanni; Gennaro Napolitano; Roberta Nardacci; Masashi Narita; Melissa Nassif; Ilana Nathan; Manabu Natsumeda; Ryno J Naude; Christin Naumann; Olaia Naveiras; Fatemeh Navid; Steffan T Nawrocki; Taras Y Nazarko; Francesca Nazio; Florentina Negoita; Thomas Neill; Amanda L Neisch; Luca M Neri; Mihai G Netea; Patrick Neubert; Thomas P Neufeld; Dietbert Neumann; Albert Neutzner; Phillip T Newton; Paul A Ney; Ioannis P Nezis; Charlene C W Ng; Tzi Bun Ng; Hang T T Nguyen; Long T Nguyen; Hong-Min Ni; Clíona Ní Cheallaigh; Zhenhong Ni; M Celeste Nicolao; Francesco Nicoli; Manuel Nieto-Diaz; Per Nilsson; Shunbin Ning; Rituraj Niranjan; Hiroshi Nishimune; Mireia Niso-Santano; Ralph A Nixon; Annalisa Nobili; Clevio Nobrega; Takeshi Noda; Uxía Nogueira-Recalde; Trevor M Nolan; Ivan Nombela; Ivana Novak; Beatriz Novoa; Takashi Nozawa; Nobuyuki Nukina; Carmen Nussbaum-Krammer; Jesper Nylandsted; Tracey R O'Donovan; Seónadh M O'Leary; Eyleen J O'Rourke; Mary P O'Sullivan; Timothy E O'Sullivan; Salvatore Oddo; Ina Oehme; Michinaga Ogawa; Eric Ogier-Denis; Margret H Ogmundsdottir; Besim Ogretmen; Goo Taeg Oh; Seon-Hee Oh; Young J Oh; Takashi Ohama; Yohei Ohashi; Masaki Ohmuraya; Vasileios Oikonomou; Rani Ojha; Koji Okamoto; Hitoshi Okazawa; Masahide Oku; Sara Oliván; Jorge M A Oliveira; Michael Ollmann; James A Olzmann; Shakib Omari; M Bishr Omary; Gizem Önal; Martin Ondrej; Sang-Bing Ong; Sang-Ging Ong; Anna Onnis; Juan A Orellana; Sara Orellana-Muñoz; Maria Del Mar Ortega-Villaizan; Xilma R Ortiz-Gonzalez; Elena Ortona; Heinz D Osiewacz; Abdel-Hamid K Osman; Rosario Osta; Marisa S Otegui; Kinya Otsu; Christiane Ott; Luisa Ottobrini; Jing-Hsiung James Ou; Tiago F Outeiro; Inger Oynebraten; Melek Ozturk; Gilles Pagès; Susanta Pahari; Marta Pajares; Utpal B Pajvani; Rituraj Pal; Simona Paladino; Nicolas Pallet; Michela Palmieri; Giuseppe Palmisano; Camilla Palumbo; Francesco Pampaloni; Lifeng Pan; Qingjun Pan; Wenliang Pan; Xin Pan; Ganna Panasyuk; Rahul Pandey; Udai B Pandey; Vrajesh Pandya; Francesco Paneni; Shirley Y Pang; Elisa Panzarini; Daniela L Papademetrio; Elena Papaleo; Daniel Papinski; Diana Papp; Eun Chan Park; Hwan Tae Park; Ji-Man Park; Jong-In Park; Joon Tae Park; Junsoo Park; Sang Chul Park; Sang-Youel Park; Abraham H Parola; Jan B Parys; Adrien Pasquier; Benoit Pasquier; João F Passos; Nunzia Pastore; Hemal H Patel; Daniel Patschan; Sophie Pattingre; Gustavo Pedraza-Alva; Jose Pedraza-Chaverri; Zully Pedrozo; Gang Pei; Jianming Pei; Hadas Peled-Zehavi; Joaquín M Pellegrini; Joffrey Pelletier; Miguel A Peñalva; Di Peng; Ying Peng; Fabio Penna; Maria Pennuto; Francesca Pentimalli; Cláudia Mf Pereira; Gustavo J S Pereira; Lilian C Pereira; Luis Pereira de Almeida; Nirma D Perera; Ángel Pérez-Lara; Ana B Perez-Oliva; María Esther Pérez-Pérez; Palsamy Periyasamy; Andras Perl; Cristiana Perrotta; Ida Perrotta; Richard G Pestell; Morten Petersen; Irina Petrache; Goran Petrovski; Thorsten Pfirrmann; Astrid S Pfister; Jennifer A Philips; Huifeng Pi; Anna Picca; Alicia M Pickrell; Sandy Picot; Giovanna M Pierantoni; Marina Pierdominici; Philippe Pierre; Valérie Pierrefite-Carle; Karolina Pierzynowska; Federico Pietrocola; Miroslawa Pietruczuk; Claudio Pignata; Felipe X Pimentel-Muiños; Mario Pinar; Roberta O Pinheiro; Ronit Pinkas-Kramarski; Paolo Pinton; Karolina Pircs; Sujan Piya; Paola Pizzo; Theo S Plantinga; Harald W Platta; Ainhoa Plaza-Zabala; Markus Plomann; Egor Y Plotnikov; Helene Plun-Favreau; Ryszard Pluta; Roger Pocock; Stefanie Pöggeler; Christian Pohl; Marc Poirot; Angelo Poletti; Marisa Ponpuak; Hana Popelka; Blagovesta Popova; Helena Porta; Soledad Porte Alcon; Eliana Portilla-Fernandez; Martin Post; Malia B Potts; Joanna Poulton; Ted Powers; Veena Prahlad; Tomasz K Prajsnar; Domenico Praticò; Rosaria Prencipe; Muriel Priault; Tassula Proikas-Cezanne; Vasilis J Promponas; Christopher G Proud; Rosa Puertollano; Luigi Puglielli; Thomas Pulinilkunnil; Deepika Puri; Rajat Puri; Julien Puyal; Xiaopeng Qi; Yongmei Qi; Wenbin Qian; Lei Qiang; Yu Qiu; Joe Quadrilatero; Jorge Quarleri; Nina Raben; Hannah Rabinowich; Debora Ragona; Michael J Ragusa; Nader Rahimi; Marveh Rahmati; Valeria Raia; Nuno Raimundo; Namakkal-Soorappan Rajasekaran; Sriganesh Ramachandra Rao; Abdelhaq Rami; Ignacio Ramírez-Pardo; David B Ramsden; Felix Randow; Pundi N Rangarajan; Danilo Ranieri; Hai Rao; Lang Rao; Rekha Rao; Sumit Rathore; J Arjuna Ratnayaka; Edward A Ratovitski; Palaniyandi Ravanan; Gloria Ravegnini; Swapan K Ray; Babak Razani; Vito Rebecca; Fulvio Reggiori; Anne Régnier-Vigouroux; Andreas S Reichert; David Reigada; Jan H Reiling; Theo Rein; Siegfried Reipert; Rokeya Sultana Rekha; Hongmei Ren; Jun Ren; Weichao Ren; Tristan Renault; Giorgia Renga; Karen Reue; Kim Rewitz; Bruna Ribeiro de Andrade Ramos; S Amer Riazuddin; Teresa M Ribeiro-Rodrigues; Jean-Ehrland Ricci; Romeo Ricci; Victoria Riccio; Des R Richardson; Yasuko Rikihisa; Makarand V Risbud; Ruth M Risueño; Konstantinos Ritis; Salvatore Rizza; Rosario Rizzuto; Helen C Roberts; Luke D Roberts; Katherine J Robinson; Maria Carmela Roccheri; Stephane Rocchi; George G Rodney; Tiago Rodrigues; Vagner Ramon Rodrigues Silva; Amaia Rodriguez; Ruth Rodriguez-Barrueco; Nieves Rodriguez-Henche; Humberto Rodriguez-Rocha; Jeroen Roelofs; Robert S Rogers; Vladimir V Rogov; Ana I Rojo; Krzysztof Rolka; Vanina Romanello; Luigina Romani; Alessandra Romano; Patricia S Romano; David Romeo-Guitart; Luis C Romero; Montserrat Romero; Joseph C Roney; Christopher Rongo; Sante Roperto; Mathias T Rosenfeldt; Philip Rosenstiel; Anne G Rosenwald; Kevin A Roth; Lynn Roth; Steven Roth; Kasper M A Rouschop; Benoit D Roussel; Sophie Roux; Patrizia Rovere-Querini; Ajit Roy; Aurore Rozieres; Diego Ruano; David C Rubinsztein; Maria P Rubtsova; Klaus Ruckdeschel; Christoph Ruckenstuhl; Emil Rudolf; Rüdiger Rudolf; Alessandra Ruggieri; Avnika Ashok Ruparelia; Paola Rusmini; Ryan R Russell; Gian Luigi Russo; Maria Russo; Rossella Russo; Oxana O Ryabaya; Kevin M Ryan; Kwon-Yul Ryu; Maria Sabater-Arcis; Ulka Sachdev; Michael Sacher; Carsten Sachse; Abhishek Sadhu; Junichi Sadoshima; Nathaniel Safren; Paul Saftig; Antonia P Sagona; Gaurav Sahay; Amirhossein Sahebkar; Mustafa Sahin; Ozgur Sahin; Sumit Sahni; Nayuta Saito; Shigeru Saito; Tsunenori Saito; Ryohei Sakai; Yasuyoshi Sakai; Jun-Ichi Sakamaki; Kalle Saksela; Gloria Salazar; Anna Salazar-Degracia; Ghasem H Salekdeh; Ashok K Saluja; Belém Sampaio-Marques; Maria Cecilia Sanchez; Jose A Sanchez-Alcazar; Victoria Sanchez-Vera; Vanessa Sancho-Shimizu; J Thomas Sanderson; Marco Sandri; Stefano Santaguida; Laura Santambrogio; Magda M Santana; Giorgio Santoni; Alberto Sanz; Pascual Sanz; Shweta Saran; Marco Sardiello; Timothy J Sargeant; Apurva Sarin; Chinmoy Sarkar; Sovan Sarkar; Maria-Rosa Sarrias; Surajit Sarkar; Dipanka Tanu Sarmah; Jaakko Sarparanta; Aishwarya Sathyanarayan; Ranganayaki Sathyanarayanan; K Matthew Scaglione; Francesca Scatozza; Liliana Schaefer; Zachary T Schafer; Ulrich E Schaible; Anthony H V Schapira; Michael Scharl; Hermann M Schatzl; Catherine H Schein; Wiep Scheper; David Scheuring; Maria Vittoria Schiaffino; Monica Schiappacassi; Rainer Schindl; Uwe Schlattner; Oliver Schmidt; Roland Schmitt; Stephen D Schmidt; Ingo Schmitz; Eran Schmukler; Anja Schneider; Bianca E Schneider; Romana Schober; Alejandra C Schoijet; Micah B Schott; Michael Schramm; Bernd Schröder; Kai Schuh; Christoph Schüller; Ryan J Schulze; Lea Schürmanns; Jens C Schwamborn; Melanie Schwarten; Filippo Scialo; Sebastiano Sciarretta; Melanie J Scott; Kathleen W Scotto; A Ivana Scovassi; Andrea Scrima; Aurora Scrivo; David Sebastian; Salwa Sebti; Simon Sedej; Laura Segatori; Nava Segev; Per O Seglen; Iban Seiliez; Ekihiro Seki; Scott B Selleck; Frank W Sellke; Joshua T Selsby; Michael Sendtner; Serif Senturk; Elena Seranova; Consolato Sergi; Ruth Serra-Moreno; Hiromi Sesaki; Carmine Settembre; Subba Rao Gangi Setty; Gianluca Sgarbi; Ou Sha; John J Shacka; Javeed A Shah; Dantong Shang; Changshun Shao; Feng Shao; Soroush Sharbati; Lisa M Sharkey; Dipali Sharma; Gaurav Sharma; Kulbhushan Sharma; Pawan Sharma; Surendra Sharma; Han-Ming Shen; Hongtao Shen; Jiangang Shen; Ming Shen; Weili Shen; Zheni Shen; Rui Sheng; Zhi Sheng; Zu-Hang Sheng; Jianjian Shi; Xiaobing Shi; Ying-Hong Shi; Kahori Shiba-Fukushima; Jeng-Jer Shieh; Yohta Shimada; Shigeomi Shimizu; Makoto Shimozawa; Takahiro Shintani; Christopher J Shoemaker; Shahla Shojaei; Ikuo Shoji; Bhupendra V Shravage; Viji Shridhar; Chih-Wen Shu; Hong-Bing Shu; Ke Shui; Arvind K Shukla; Timothy E Shutt; Valentina Sica; Aleem Siddiqui; Amanda Sierra; Virginia Sierra-Torre; Santiago Signorelli; Payel Sil; Bruno J de Andrade Silva; Johnatas D Silva; Eduardo Silva-Pavez; Sandrine Silvente-Poirot; Rachel E Simmonds; Anna Katharina Simon; Hans-Uwe Simon; Matias Simons; Anurag Singh; Lalit P Singh; Rajat Singh; Shivendra V Singh; Shrawan K Singh; Sudha B Singh; Sunaina Singh; Surinder Pal Singh; Debasish Sinha; Rohit Anthony Sinha; Sangita Sinha; Agnieszka Sirko; Kapil Sirohi; Efthimios L Sivridis; Panagiotis Skendros; Aleksandra Skirycz; Iva Slaninová; Soraya S Smaili; Andrei Smertenko; Matthew D Smith; Stefaan J Soenen; Eun Jung Sohn; Sophia P M Sok; Giancarlo Solaini; Thierry Soldati; Scott A Soleimanpour; Rosa M Soler; Alexei Solovchenko; Jason A Somarelli; Avinash Sonawane; Fuyong Song; Hyun Kyu Song; Ju-Xian Song; Kunhua Song; Zhiyin Song; Leandro R Soria; Maurizio Sorice; Alexander A Soukas; Sandra-Fausia Soukup; Diana Sousa; Nadia Sousa; Paul A Spagnuolo; Stephen A Spector; M M Srinivas Bharath; Daret St Clair; Venturina Stagni; Leopoldo Staiano; Clint A Stalnecker; Metodi V Stankov; Peter B Stathopulos; Katja Stefan; Sven Marcel Stefan; Leonidas Stefanis; Joan S Steffan; Alexander Steinkasserer; Harald Stenmark; Jared Sterneckert; Craig Stevens; Veronika Stoka; Stephan Storch; Björn Stork; Flavie Strappazzon; Anne Marie Strohecker; Dwayne G Stupack; Huanxing Su; Ling-Yan Su; Longxiang Su; Ana M Suarez-Fontes; Carlos S Subauste; Selvakumar Subbian; Paula V Subirada; Ganapasam Sudhandiran; Carolyn M Sue; Xinbing Sui; Corey Summers; Guangchao Sun; Jun Sun; Kang Sun; Meng-Xiang Sun; Qiming Sun; Yi Sun; Zhongjie Sun; Karen K S Sunahara; Eva Sundberg; Katalin Susztak; Peter Sutovsky; Hidekazu Suzuki; Gary Sweeney; J David Symons; Stephen Cho Wing Sze; Nathaniel J Szewczyk; Anna Tabęcka-Łonczynska; Claudio Tabolacci; Frank Tacke; Heinrich Taegtmeyer; Marco Tafani; Mitsuo Tagaya; Haoran Tai; Stephen W G Tait; Yoshinori Takahashi; Szabolcs Takats; Priti Talwar; Chit Tam; Shing Yau Tam; Davide Tampellini; Atsushi Tamura; Chong Teik Tan; Eng-King Tan; Ya-Qin Tan; Masaki Tanaka; Motomasa Tanaka; Daolin Tang; Jingfeng Tang; Tie-Shan Tang; Isei Tanida; Zhipeng Tao; Mohammed Taouis; Lars Tatenhorst; Nektarios Tavernarakis; Allen Taylor; Gregory A Taylor; Joan M Taylor; Elena Tchetina; Andrew R Tee; Irmgard Tegeder; David Teis; Natercia Teixeira; Fatima Teixeira-Clerc; Kumsal A Tekirdag; Tewin Tencomnao; Sandra Tenreiro; Alexei V Tepikin; Pilar S Testillano; Gianluca Tettamanti; Pierre-Louis Tharaux; Kathrin Thedieck; Arvind A Thekkinghat; Stefano Thellung; Josephine W Thinwa; V P Thirumalaikumar; Sufi Mary Thomas; Paul G Thomes; Andrew Thorburn; Lipi Thukral; Thomas Thum; Michael Thumm; Ling Tian; Ales Tichy; Andreas Till; Vincent Timmerman; Vladimir I Titorenko; Sokol V Todi; Krassimira Todorova; Janne M Toivonen; Luana Tomaipitinca; Dhanendra Tomar; Cristina Tomas-Zapico; Sergej Tomić; Benjamin Chun-Kit Tong; Chao Tong; Xin Tong; Sharon A Tooze; Maria L Torgersen; Satoru Torii; Liliana Torres-López; Alicia Torriglia; Christina G Towers; Roberto Towns; Shinya Toyokuni; Vladimir Trajkovic; Donatella Tramontano; Quynh-Giao Tran; Leonardo H Travassos; Charles B Trelford; Shirley Tremel; Ioannis P Trougakos; Betty P Tsao; Mario P Tschan; Hung-Fat Tse; Tak Fu Tse; Hitoshi Tsugawa; Andrey S Tsvetkov; David A Tumbarello; Yasin Tumtas; María J Tuñón; Sandra Turcotte; Boris Turk; Vito Turk; Bradley J Turner; Richard I Tuxworth; Jessica K Tyler; Elena V Tyutereva; Yasuo Uchiyama; Aslihan Ugun-Klusek; Holm H Uhlig; Marzena Ułamek-Kozioł; Ilya V Ulasov; Midori Umekawa; Christian Ungermann; Rei Unno; Sylvie Urbe; Elisabet Uribe-Carretero; Suayib Üstün; Vladimir N Uversky; Thomas Vaccari; Maria I Vaccaro; Björn F Vahsen; Helin Vakifahmetoglu-Norberg; Rut Valdor; Maria J Valente; Ayelén Valko; Richard B Vallee; Angela M Valverde; Greet Van den Berghe; Stijn van der Veen; Luc Van Kaer; Jorg van Loosdregt; Sjoerd J L van Wijk; Wim Vandenberghe; Ilse Vanhorebeek; Marcos A Vannier-Santos; Nicola Vannini; M Cristina Vanrell; Chiara Vantaggiato; Gabriele Varano; Isabel Varela-Nieto; Máté Varga; M Helena Vasconcelos; Somya Vats; Demetrios G Vavvas; Ignacio Vega-Naredo; Silvia Vega-Rubin-de-Celis; Guillermo Velasco; Ariadna P Velázquez; Tibor Vellai; Edo Vellenga; Francesca Velotti; Mireille Verdier; Panayotis Verginis; Isabelle Vergne; Paul Verkade; Manish Verma; Patrik Verstreken; Tim Vervliet; Jörg Vervoorts; Alexandre T Vessoni; Victor M Victor; Michel Vidal; Chiara Vidoni; Otilia V Vieira; Richard D Vierstra; Sonia Viganó; Helena Vihinen; Vinoy Vijayan; Miquel Vila; Marçal Vilar; José M Villalba; Antonio Villalobo; Beatriz Villarejo-Zori; Francesc Villarroya; Joan Villarroya; Olivier Vincent; Cecile Vindis; Christophe Viret; Maria Teresa Viscomi; Dora Visnjic; Ilio Vitale; David J Vocadlo; Olga V Voitsekhovskaja; Cinzia Volonté; Mattia Volta; Marta Vomero; Clarissa Von Haefen; Marc A Vooijs; Wolfgang Voos; Ljubica Vucicevic; Richard Wade-Martins; Satoshi Waguri; Kenrick A Waite; Shuji Wakatsuki; David W Walker; Mark J Walker; Simon A Walker; Jochen Walter; Francisco G Wandosell; Bo Wang; Chao-Yung Wang; Chen Wang; Chenran Wang; Chenwei Wang; Cun-Yu Wang; Dong Wang; Fangyang Wang; Feng Wang; Fengming Wang; Guansong Wang; Han Wang; Hao Wang; Hexiang Wang; Hong-Gang Wang; Jianrong Wang; Jigang Wang; Jiou Wang; Jundong Wang; Kui Wang; Lianrong Wang; Liming Wang; Maggie Haitian Wang; Meiqing Wang; Nanbu Wang; Pengwei Wang; Peipei Wang; Ping Wang; Ping Wang; Qing Jun Wang; Qing Wang; Qing Kenneth Wang; Qiong A Wang; Wen-Tao Wang; Wuyang Wang; Xinnan Wang; Xuejun Wang; Yan Wang; Yanchang Wang; Yanzhuang Wang; Yen-Yun Wang; Yihua Wang; Yipeng Wang; Yu Wang; Yuqi Wang; Zhe Wang; Zhenyu Wang; Zhouguang Wang; Gary Warnes; Verena Warnsmann; Hirotaka Watada; Eizo Watanabe; Maxinne Watchon; Anna Wawrzyńska; Timothy E Weaver; Grzegorz Wegrzyn; Ann M Wehman; Huafeng Wei; Lei Wei; Taotao Wei; Yongjie Wei; Oliver H Weiergräber; Conrad C Weihl; Günther Weindl; Ralf Weiskirchen; Alan Wells; Runxia H Wen; Xin Wen; Antonia Werner; Beatrice Weykopf; Sally P Wheatley; J Lindsay Whitton; Alexander J Whitworth; Katarzyna Wiktorska; Manon E Wildenberg; Tom Wileman; Simon Wilkinson; Dieter Willbold; Brett Williams; Robin S B Williams; Roger L Williams; Peter R Williamson; Richard A Wilson; Beate Winner; Nathaniel J Winsor; Steven S Witkin; Harald Wodrich; Ute Woehlbier; Thomas Wollert; Esther Wong; Jack Ho Wong; Richard W Wong; Vincent Kam Wai Wong; W Wei-Lynn Wong; An-Guo Wu; Chengbiao Wu; Jian Wu; Junfang Wu; Kenneth K Wu; Min Wu; Shan-Ying Wu; Shengzhou Wu; Shu-Yan Wu; Shufang Wu; William K K Wu; Xiaohong Wu; Xiaoqing Wu; Yao-Wen Wu; Yihua Wu; Ramnik J Xavier; Hongguang Xia; Lixin Xia; Zhengyuan Xia; Ge Xiang; Jin Xiang; Mingliang Xiang; Wei Xiang; Bin Xiao; Guozhi Xiao; Hengyi Xiao; Hong-Tao Xiao; Jian Xiao; Lan Xiao; Shi Xiao; Yin Xiao; Baoming Xie; Chuan-Ming Xie; Min Xie; Yuxiang Xie; Zhiping Xie; Zhonglin Xie; Maria Xilouri; Congfeng Xu; En Xu; Haoxing Xu; Jing Xu; JinRong Xu; Liang Xu; Wen Wen Xu; Xiulong Xu; Yu Xue; Sokhna M S Yakhine-Diop; Masamitsu Yamaguchi; Osamu Yamaguchi; Ai Yamamoto; Shunhei Yamashina; Shengmin Yan; Shian-Jang Yan; Zhen Yan; Yasuo Yanagi; Chuanbin Yang; Dun-Sheng Yang; Huan Yang; Huang-Tian Yang; Hui Yang; Jin-Ming Yang; Jing Yang; Jingyu Yang; Ling Yang; Liu Yang; Ming Yang; Pei-Ming Yang; Qian Yang; Seungwon Yang; Shu Yang; Shun-Fa Yang; Wannian Yang; Wei Yuan Yang; Xiaoyong Yang; Xuesong Yang; Yi Yang; Ying Yang; Honghong Yao; Shenggen Yao; Xiaoqiang Yao; Yong-Gang Yao; Yong-Ming Yao; Takahiro Yasui; Meysam Yazdankhah; Paul M Yen; Cong Yi; Xiao-Ming Yin; Yanhai Yin; Zhangyuan Yin; Ziyi Yin; Meidan Ying; Zheng Ying; Calvin K Yip; Stephanie Pei Tung Yiu; Young H Yoo; Kiyotsugu Yoshida; Saori R Yoshii; Tamotsu Yoshimori; Bahman Yousefi; Boxuan Yu; Haiyang Yu; Jun Yu; Jun Yu; Li Yu; Ming-Lung Yu; Seong-Woon Yu; Victor C Yu; W Haung Yu; Zhengping Yu; Zhou Yu; Junying Yuan; Ling-Qing Yuan; Shilin Yuan; Shyng-Shiou F Yuan; Yanggang Yuan; Zengqiang Yuan; Jianbo Yue; Zhenyu Yue; Jeanho Yun; Raymond L Yung; David N Zacks; Gabriele Zaffagnini; Vanessa O Zambelli; Isabella Zanella; Qun S Zang; Sara Zanivan; Silvia Zappavigna; Pilar Zaragoza; Konstantinos S Zarbalis; Amir Zarebkohan; Amira Zarrouk; Scott O Zeitlin; Jialiu Zeng; Ju-Deng Zeng; Eva Žerovnik; Lixuan Zhan; Bin Zhang; Donna D Zhang; Hanlin Zhang; Hong Zhang; Hong Zhang; Honghe Zhang; Huafeng Zhang; Huaye Zhang; Hui Zhang; Hui-Ling Zhang; Jianbin Zhang; Jianhua Zhang; Jing-Pu Zhang; Kalin Y B Zhang; Leshuai W Zhang; Lin Zhang; Lisheng Zhang; Lu Zhang; Luoying Zhang; Menghuan Zhang; Peng Zhang; Sheng Zhang; Wei Zhang; Xiangnan Zhang; Xiao-Wei Zhang; Xiaolei Zhang; Xiaoyan Zhang; Xin Zhang; Xinxin Zhang; Xu Dong Zhang; Yang Zhang; Yanjin Zhang; Yi Zhang; Ying-Dong Zhang; Yingmei Zhang; Yuan-Yuan Zhang; Yuchen Zhang; Zhe Zhang; Zhengguang Zhang; Zhibing Zhang; Zhihai Zhang; Zhiyong Zhang; Zili Zhang; Haobin Zhao; Lei Zhao; Shuang Zhao; Tongbiao Zhao; Xiao-Fan Zhao; Ying Zhao; Yongchao Zhao; Yongliang Zhao; Yuting Zhao; Guoping Zheng; Kai Zheng; Ling Zheng; Shizhong Zheng; Xi-Long Zheng; Yi Zheng; Zu-Guo Zheng; Boris Zhivotovsky; Qing Zhong; Ao Zhou; Ben Zhou; Cefan Zhou; Gang Zhou; Hao Zhou; Hong Zhou; Hongbo Zhou; Jie Zhou; Jing Zhou; Jing Zhou; Jiyong Zhou; Kailiang Zhou; Rongjia Zhou; Xu-Jie Zhou; Yanshuang Zhou; Yinghong Zhou; Yubin Zhou; Zheng-Yu Zhou; Zhou Zhou; Binglin Zhu; Changlian Zhu; Guo-Qing Zhu; Haining Zhu; Hongxin Zhu; Hua Zhu; Wei-Guo Zhu; Yanping Zhu; Yushan Zhu; Haixia Zhuang; Xiaohong Zhuang; Katarzyna Zientara-Rytter; Christine M Zimmermann; Elena Ziviani; Teresa Zoladek; Wei-Xing Zong; Dmitry B Zorov; Antonio Zorzano; Weiping Zou; Zhen Zou; Zhengzhi Zou; Steven Zuryn; Werner Zwerschke; Beate Brand-Saberi; X Charlie Dong; Chandra Shekar Kenchappa; Zuguo Li; Yong Lin; Shigeru Oshima; Yueguang Rong; Judith C Sluimer; Christina L Stallings; Chun-Kit Tong Journal: Autophagy Date: 2021-02-08 Impact factor: 13.391
Authors: Amir Ajoolabady; Shuyi Wang; Guido Kroemer; Josef M Penninger; Vladimir N Uversky; Domenico Pratico; Nils Henninger; Russel J Reiter; Askiel Bruno; Kaumudi Joshipura; Hamid Aslkhodapasandhokmabad; Daniel J Klionsky; Jun Ren Journal: Pharmacol Ther Date: 2021-04-03 Impact factor: 13.400
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