Suzan Schwertheim1, Julia Kälsch1,2, Holger Jastrow3, Christoph Matthias Schaefer1, Sarah Theurer1, Saskia Ting1, Ali Canbay4, Heiner Wedemeyer2, Kurt Werner Schmid1,5, Hideo Andreas Baba6. 1. Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Hufelandstr. 55, 45147, Essen, Germany. 2. Department of Gastroenterology and Hepatology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany. 3. Institute of Anatomy and Electron Microscopy Unit of Imaging Center Essen, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany. 4. Department of Medicine, Ruhr University Bochum, University Hospital Knappschaftskrankenhaus Bochum, 44892, Bochum, Germany. 5. West German Cancer Centre Essen (WTZE), Essen, Germany. 6. Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Hufelandstr. 55, 45147, Essen, Germany. hideo.baba@uk-essen.de.
Abstract
Nuclear inclusions (NI) are a common finding in hepatocytes from patients with liver disease especially in diabetes mellitus and non-alcoholic fatty liver disease (NAFLD) but studies examining the shape and content of these inclusions in detail are lacking. In this study we define two distinct types of NI in NAFLD: inclusions bounded by the nuclear membrane, containing degenerative cell organelles and heterolysosomes (type1) and inclusions with deposits of glycogen but without any kind of organelles and delimiting membrane (type2). NI in 77 paraffin-embedded patients of NAFLD including NAFL and non-alcoholic steatohepatitis (NASH) were analyzed. In 4-12% of type1 NI immunopositivity for the autophagy-associated proteins LC3B, ubiquitin, p62/sequestosome1, cathepsin D and cathepsin B were detected with co-localizations of ubiquitin and p62; type2 NI showed no immunoreactivity. Three-dimensional reconstructions of isolated nuclei revealed that NI type1 are completely enclosed within the nucleus, suggesting that NI, although probably derived from cytoplasmic invaginations, are not just simple invaginations. Our study demonstrates two morphologically different types of inclusions in NAFLD, whereby both gained significantly in number in advanced stages. We suggest that the presence of autophagy-associated proteins and degenerated organelles within type1 NI plays a role in disease progression.
Nuclear inclusions (NI) are a common finding in hepatocytes from patients with liver disease especially in diabetes mellitus and non-alcoholic fatty liver disease (NAFLD) but studies examining the shape and content of these inclusions in detail are lacking. In this study we define two distinct types of NI in NAFLD: inclusions bounded by the nuclear membrane, containing degenerative cell organelles and heterolysosomes (type1) and inclusions with deposits of glycogen but without any kind of organelles and delimiting membrane (type2). NI in 77 paraffin-embedded patients of NAFLD including NAFL and non-alcoholic steatohepatitis (NASH) were analyzed. In 4-12% of type1 NI immunopositivity for the autophagy-associated proteins LC3B, ubiquitin, p62/sequestosome1, cathepsin D and cathepsin B were detected with co-localizations of ubiquitin and p62; type2 NI showed no immunoreactivity. Three-dimensional reconstructions of isolated nuclei revealed that NI type1 are completely enclosed within the nucleus, suggesting that NI, although probably derived from cytoplasmic invaginations, are not just simple invaginations. Our study demonstrates two morphologically different types of inclusions in NAFLD, whereby both gained significantly in number in advanced stages. We suggest that the presence of autophagy-associated proteins and degenerated organelles within type1 NI plays a role in disease progression.
The most common chronic liver disease in Western countries is non-alcoholic fatty liver disease (NAFLD), affecting up to 30% of the adult population[1]. NAFLD is already considered as the hepatic manifestation of the metabolic syndrome[2]. Both clinically and histologically, NAFLD shows a broad spectrum of disease variety. Its spectrum embraces simple steatosis in non-alcoholic fatty liver (NAFL) to advanced non-alcoholic steatohepatitis (NASH), which may be associated with fibrosis and progression to cirrhosis or hepatocellular carcinoma (HCC)[3,4]. Morphologic alterations in NAFLD might not only alter the cytoplasm but also the nucleus. Nuclear vacuolation in hepatocytes was first reported over a century ago by Paul Ehrlich[5]. He showed glycogen filled vacuoles in nuclei with eccentrically displaced chromatin in liver tissue obtained from autopsies of diabeticpatients. When stained with haematoxylin and eosin (HE) glycogen was leached out and these formations appeared as an empty nucleus[5]. These so called glycogenated nuclei are found in up to 75% in non-alcoholic steatohepatitis (NASH)[6]. On the ultrastructural level these “vacuoles” are filled with typical glycogen granules with diameters of 3–7 nm and are not delimited by a membrane[7]. In subsequent studies, nuclear vacuolation was described in different liver diseases, such as non-alcoholic fatty liver disease (NAFLD), alcoholic steatohepatitis or Wilson’s disease[8-12].Intranuclear inclusions (NI) have been studied for over 100 years[13]. They have been considered to have a homogenous morphology until Cowdry et al. were the first to divide these NI in patients with viral infections into two groups depending on alterations of cell morphology as a reaction to a virus, the cowdry bodies[14,15]. These early studies were based on mere description of morphological aspects of inclusions in light microscopy. Later, in vivo studies demonstrated the induction of NI by injections of colchicine or thioacetamide in mouse hepatocytes[16,17]. Ultrastructural characterization of NI in mouse liver and HCC revealed inclusions bounded by a double membrane originating from the nuclear envelope[18].In NAFLD, nuclear vacuolation is linked to aging process or senescence but no differentiation is made between NI with or without a limiting membrane[19]. Our recent study of membrane-bounded NI in HCC[20] and in thyroid carcinomas[21] has shown the presence of autophagy-associated proteins and proteases within the inclusions; in HCC the occurrence of inclusions was associated with patients’ survival benefit[20]. Jaskolski et al. described that autophagy is involved in the pathogenesis of intranuclear vacuolation in meningeomas[22]; their electron microscopic studies revealed lysosomal bodies and autophagic vacuoles within these NI, suggesting an active macroautophagy process[22].Thus, the aim of our current study was to characterize NI in NAFLD morphologically in detail and to clarify a possible association with autophagy and disease progression. Therefore, we analyzed the NI found in our NAFLD cohort first on the presence of a limiting membrane and secondly on storage of glycogen. Staining with periodic acid-Schiff (PAS) gave proof of glycogen infiltration. We performed three-dimensional (3D) reconstructions of isolated hepatocyte nuclei to clarify the shape of the NI; transmission electron microscopy (TEM) studies were used to analyze the NI regarding limiting membrane and content in more detail. To investigate if these inclusions might have a biological function we further examined them with immunohistochemistry (IHC) for the presence of the autophagy-associated proteins p62/sequestosome1, ubiquitin, LC3B, cathepsin B and cathepsin D[23] whereby p62 is involved, among others, in NFκB[24,25] and Keap1-Nrf2[26] signaling pathways. Three-dimensional reconstruction of double immunofluorescence (IF) -labelled tissue sections was performed to elucidate the exact spatial location of the inclusions and accumulated proteins.
Results
Occurrence of two different types of NI in NAFLD
Analysis of HE sections in the 58 morbidly obesepatients and 19 organ donor patients (control group) revealed NI in 76 out of the 77 cases. The one case in which we did not detect any NI at all was in the control group. These NI were different in shape and size (Fig. 1). The most obvious difference was the presence of a membrane around some NI. We designated these NI as “type1” (left column of Fig. 1). Whereas all the inclusions’ internal area appeared empty or heterogeneous in HE staining, type1 NI were limited by a basophilic membrane. However, the inclusions we called as “type2” NI were not surrounded by a basophilic membrane; these inclusions are also known as glycogenated nuclei[5,6] (right column of Fig. 1). For both types the staining reaction of the inclusions’ internal area was often similar to that of the cytoplasm but differed from that of the karyoplasm. We detected in both types of NI accumulation of glycogen. Lamin AC immunofluorescence analysis demonstrated clearly the difference between both types of inclusions: type1 inclusion were bounded by a membrane with lamin AC immunopositivity, whereby type2 inclusions appeared to be either surrounded by no membrane at all or at least by no membrane that was positive for lamin AC (Fig. 1). Since lamin AC is a nuclear membrane marker, our discovery of lamin AC at the membrane bordering type1 inclusions, suggests its nuclear membrane origin.
Figure 1
Characterization of type1 and type2 NI in NAFLD. The images show two representative HE-stainings of tissue sections from type1 and type2 NI demonstrating that type1 NI are surrounded by a basophilic membrane, which is lacking in type2 NI. Positive PAS staining is detected both for type1 and type2 NI and the negative staining for PAS diastase depicts glycogen accumulation as the origin for this positivity. Immunofluorescence analysis on lamin AC expression demonstrates that lamin AC is at the membrane bordering type1 NI. The white bar at the bottom right, representative for all images indicates 10 µm. Original magnifications: 1,000 X.
Characterization of type1 and type2 NI in NAFLD. The images show two representative HE-stainings of tissue sections from type1 and type2 NI demonstrating that type1 NI are surrounded by a basophilic membrane, which is lacking in type2 NI. Positive PAS staining is detected both for type1 and type2 NI and the negative staining for PAS diastase depicts glycogen accumulation as the origin for this positivity. Immunofluorescence analysis on lamin AC expression demonstrates that lamin AC is at the membrane bordering type1 NI. The white bar at the bottom right, representative for all images indicates 10 µm. Original magnifications: 1,000 X.
Correlation of the presence of type1/type2 NI and NAFLD disease severity
Additionally, NI were studied quantitatively. The total cell count within the whole tissue section for each case was performed by automatic counting identifying nuclei by positive DAPI staining. Then from the same paraffin block a HE stained tissue section was analyzed on the number of type1 and type2 NI. We counted the number of NI with (type1) or without (type2) delimiting basophilic membrane within the whole section. If a nucleus contained more than one intranuclear inclusion, we counted this as one intranuclear inclusion. To eliminate the bias effect of different tissue size, we divided the number of NI by the total cell count and multiplied it with factor 1,000 for more convenience. Details are depicted in Fig. 2 showing the median values for the number of NI type1 (Fig. 2A) and type2 (Fig. 2B) per 1,000 cells. The median number of type2 NI/1,000 cells in NASH was with 4.03 (range: 0–78.5) 3.5 times higher than that of type1 NI/1,000 cells with 1.15 (range: 0–10). Number of valid cases was 19 Control, 23 NAFL and 35 NASH cases. To investigate whether there was a correlation between the occurrence of NI and NAFLD, we performed the Kruskal–Wallis test. Interestingly, both membrane-bounded NI (type1) and vacuolated nuclei i.e. non-membrane bounded NI (type2) were significantly more in NASH compared to controls (P ≤ 0.001; Fig. 2A,B). Additionally, the number of type1 NI was significantly higher in NASH than in NAFL (P = 0.012; Fig. 2A) and type1 NI were significantly more in NAFL than in controls (P = 0.016; Fig. 2A). Study of Spearman's rho correlations showed significant correlations between type1/type2 NI and disease stages. We detected for both type1 NI (P < 001; rho = 0.659) and type2 NI (P < 001; rho = 0.428) strong associations with NAFLD progression; this correlation was slightly stronger for type1 NI than for type2 NI.
Figure 2
Associations between the number of NI and disease progression. (a) Number of type1 NI/1,000 cells increases with progression of NAFLD; NASH vs. controls: ***P < 0.001; NASH vs. NAFL: *P = 0.012; NAFL vs. controls: *P = 0.016. Number of type1 NI/1,000 cells: Control 0.06 (0–0.21), NAFL 0.26 (0–2.19), NASH 1.15 (0–10). (b) Type2 NI/1,000 cells are more in NASH than in controls: ***P = 0.001. Number of type2 NI/1,000 cells: Control 0.17 (0–14.96), NAFL 0.26 (0–38.25), NASH 4.03 (0–78.49); values are medians with ranges presented in parentheses. (All Kruskal–Wallis test; bold lines inside the box plot represent median levels; the values are shown). Results are significant at *P ≤ 0.05 and ***P ≤ 0.001.
Associations between the number of NI and disease progression. (a) Number of type1 NI/1,000 cells increases with progression of NAFLD; NASH vs. controls: ***P < 0.001; NASH vs. NAFL: *P = 0.012; NAFL vs. controls: *P = 0.016. Number of type1 NI/1,000 cells: Control 0.06 (0–0.21), NAFL 0.26 (0–2.19), NASH 1.15 (0–10). (b) Type2 NI/1,000 cells are more in NASH than in controls: ***P = 0.001. Number of type2 NI/1,000 cells: Control 0.17 (0–14.96), NAFL 0.26 (0–38.25), NASH 4.03 (0–78.49); values are medians with ranges presented in parentheses. (All Kruskal–Wallis test; bold lines inside the box plot represent median levels; the values are shown). Results are significant at *P ≤ 0.05 and ***P ≤ 0.001.
Studies regarding correlations between clinical/laboratory parameters and the number of type1/type2 NI
We have classified fasting blood glucose into three levels: normal range: 70–99 mg/dl, pre-diabetes: 100–125 mg/dl; and diabetes: ≥ 126 mg/dl according to the criteria of the American Diabetes Association (ADA)[27]. Then we split the patients into two groups: we combined the patients with pre-diabetes and diabetes into one group; in the second group were all patients with normal glucose levels. We found a significantly higher number of NI type1 in the pre-diabetes and diabetes group than in the normal group (P = 0.032); details are shown in Table 1. Study of Spearman's rho correlations showed significant correlation between the number of NI type1 and fasting glucose levels (P = 0.043; rho = 0.276). Additionally, we analyzed the data on an association between elevated cholesterol, triglyceride, AST and GGT levels and the number of NI type1 and 2. We detected a significant association between elevated triglyceride levels and the number of NI type1 (P = 0.006) and NI type2 (P = 0.001). Further, patients with elevated GGT levels had significant more NI type2 (P = 0.050). Also the association between the number of NI type1/2 and the steatosis, ballooning and lobular inflammation grades was significant (P < 0.001, Table 1).
Table 1
Correlation of clinical/laboratory parameters with the number of NI type1/type2.
Parameters
n valid cases
Number of NI type1
Number of NI type2
Median valueb
P value*
Median valueb
P value*
Fasting glucose (mg/dl)
Normal (70–99)
19
0.42 (0–1.98)
0.032
0.35 (0–27.23)
0.068
Pre-diabetes and diabetesa
35
1.12 (0–8.46)
4.03 (0–78.49)
Total cholesterol (mg/dl)
Normal (< 200)
20
0.87 (0–8.46)
0.598
4.1 (0–61.39)
0.730
Elevated (> 200)
13
1.12 (0.26–3.79)
5.9 (0–78.49)
Triglyceride (mg/dl)
Normal (< 200)
18
0.66 (0–2.32)
0.006
0.8 (0–31.04)
0.001
Elevated (> 200)
7
2.33 (0.7–8.46)
27.2 (10.4–78)
ALT (U/L)
Normal (< 35/ < 50)
40
0.59 (0–8.46)
0.078
0.73 (0–61.4)
0.444
Elevated (> 35/ > 50)
24
1.05 (0–6.18)
2.73 (0–78.5)
AST (U/L)
Normal (< 35/ < 50)
28
0.59 (0–8.46)
0.617
0.33 (0–61.4)
0.204
Elevated (> 35/ > 50)
16
1.05 (0–6.18)
6.09 (0–78.5)
GGT (U/L)
Normal (< 35/ < 55)
42
0.61 (0–6.18)
0.134
0.37 (0–61.4)
0.050
Elevated (> 35/ > 55)
22
0.92 (0–8.46)
4.2 (0–78.5)
Fibrosis grade
0
10
1.15 (0–2.19)
0.55 (0–38.2)
1
16
0.61 (0–2.32)
0.95 (0–31)
2
28
0.96 (0–10)
0.179
4.8 (0–78.5)
0.159
3
4
0.48 (0–1.3)
2.04 (0–8.67)
Steatosis grade
0
19
0.06 (0–0.21)
0.17 (0–14.96)
1
28
0.6 (0–2.19)
0.4 (0–38.25)
2
18
1.17 (0–8.43)
< 0.001
2.25 (0–61.39)
< 0.001
3
12
1.99 (0.61–10)
12.8 (0.1–78.5)
Ballooning grade
0
37
0.11 (0–1.86)
0.17 (0–27.23)
1
21
1.15 (0–3.53)
< 0.001
1.88 (0–61.39)
< 0.001
2
19
1.67 (0.21–10)
8.67 (0.1–78.5)
Lobular inflammation grade
0
35
0.11 (0–2.19)
0.21 (0–38.25)
1
16
1.05 (0–8.46)
1.51 (0–37.24)
2
24
1.47 (0–10)
< 0.001
4.06 (0–78.49)
0.004
3
2
0.44 (0.28–0.6)
1.8 (0.76–2.84)
aWe defined fasting blood glucose level of 100–125 mg/dl as pre-diabetes and glucose levels of ≥ 126 mg/dl as diabetes.
bValues are median number of type1/2 NI per 1,000 cells with ranges presented in parentheses. ALT and AST threshold for normal values were < 35 U/l for females and < 50 U/l for males; GGT threshold for normal values were < 35 U/l for females and < 55 U/l for males.
*P values correspond to the analysis of correlation between clinical/laboratory parameters with the number of type1/type2 NI in the HCC cohort. Mann–Whitney U-test and Kruskal–Wallis-tests were used for statistical analysis of the difference between two or more groups. Results are significant at P ≤ 0.05.
Correlation of clinical/laboratory parameters with the number of NI type1/type2.aWe defined fasting blood glucose level of 100–125 mg/dl as pre-diabetes and glucose levels of ≥ 126 mg/dl as diabetes.bValues are median number of type1/2 NI per 1,000 cells with ranges presented in parentheses. ALT and AST threshold for normal values were < 35 U/l for females and < 50 U/l for males; GGT threshold for normal values were < 35 U/l for females and < 55 U/l for males.*P values correspond to the analysis of correlation between clinical/laboratory parameters with the number of type1/type2 NI in the HCC cohort. Mann–Whitney U-test and Kruskal–Wallis-tests were used for statistical analysis of the difference between two or more groups. Results are significant at P ≤ 0.05.
Ultrastructural depiction of NI
To investigate NI in NAFLD in more detail, we performed transmission electron microscopy (TEM) and observed that these NI were filled differently (Fig. 3). One type of intranuclear inclusion was coated by a membrane resembling the double-layer nuclear membrane (Fig. 3A,C). Our type1 membrane-bordered NI were filled with degenerated cell organelles and heterolysosomes. The content of the NI appeared to be more compressed compared to the cytoplasm (Fig. 3A,C). We detected membrane-limited NI both with invaginations of surrounding cytoplasm (data not shown) and totally lacking contact to the cytoplasm. In contrast to this our type2 NI had no additional membrane and appeared to be an “empty” nucleus with deposits of glycogen and did not contain any kind of cell organelles (Fig. 3B,D).
Figure 3
Ultrastructural analysis of type1 and type2 NI. (a) TEM image of a type1 intranuclear inclusion (NI) in NAFLD shows a hepatocyte nucleus (N) with NI coated by a membrane resembling the double-layer nuclear membrane (arrows). The NI contains three (hetero-) lysosomes (triangle), a lipid droplet (LD) and an artificial hole (+) in the sample. (b) The image reveals a type2 NI with deposit of glycogen (*). The nuclear membrane (arrow) delimiting the nucleus is shown and a lipid droplet (LD) within the cytoplasm. (c,d) The images depict additional examples of a type1 NI (c) and a type2 NI (d). Ultrathin sections were mounted on copper grids, double-stained with uranyl acetate (1%) and lead citrate (0.4%). Electron microscopy scale bars: Image A: 1 µm and 0.5 µm (in increased magnification); Image B: 5 µm and 1 µm (in increased magnification); Image C: 2 µm; Image D: 5 µm.
Ultrastructural analysis of type1 and type2 NI. (a) TEM image of a type1 intranuclear inclusion (NI) in NAFLD shows a hepatocyte nucleus (N) with NI coated by a membrane resembling the double-layer nuclear membrane (arrows). The NI contains three (hetero-) lysosomes (triangle), a lipid droplet (LD) and an artificial hole (+) in the sample. (b) The image reveals a type2 NI with deposit of glycogen (*). The nuclear membrane (arrow) delimiting the nucleus is shown and a lipid droplet (LD) within the cytoplasm. (c,d) The images depict additional examples of a type1 NI (c) and a type2 NI (d). Ultrathin sections were mounted on copper grids, double-stained with uranyl acetate (1%) and lead citrate (0.4%). Electron microscopy scale bars: Image A: 1 µm and 0.5 µm (in increased magnification); Image B: 5 µm and 1 µm (in increased magnification); Image C: 2 µm; Image D: 5 µm.
3D nuclear imaging: membrane-bounded NI are completely separated from the cytoplasm and closed
We performed 3D reconstruction of isolated hepatocyte nuclei and stained them with immunofluorescence labelled lamin AC to examine the shape and the origin of the NI. We detected that type1 NI were delimited by a lamin AC stained intact membrane and were completely closed in all studied section planes (Fig. 4). There was no connection between the intranuclear inclusion and the cytoplasm. As the membrane bounding the inclusion was lamin AC positive, this indicates an origin from the nuclear membrane. Further, we demonstrate that the type1 intranuclear inclusion was completely enclosed within the nucleus (Fig. 4, Supplementary Movie 1, Supplementary Digital Content 1). In contrast, analysis of the type2 intranuclear inclusion showed no lamin AC stained membrane limitation (Fig. 4, Supplementary Movie 2, Supplementary Digital Content 2).
Figure 4
Three-dimensional (3D) reconstruction of immunofluorescence-labelled (lamin AC) isolated cell nuclei in X-, Y- and Z-axes. The images reveal representative cases with type1 and type2 NI. For the type1 intranuclear inclusion 161 optical sections of 0.1 µm were imaged and 113 of them were used for 3D reconstruction. The type2 intranuclear inclusion was reconstructed from 173 of 213 optical Sects. (0.1 µm). The type1 intranuclear inclusion is limited by a lamin AC (nuclear membrane marker) immunopositive membrane. The inclusion is completely closed in all planes and there is no DAPI (blue) staining within it. Lamin AC-positive immunostaining (light green) is also present in the nuclear membrane. The images of the type2 intranuclear inclusion in the bottom row show only immunopositivity for lamin AC in the nuclear membrane while the nucleoplasm appears “empty” in all planes.
Three-dimensional (3D) reconstruction of immunofluorescence-labelled (lamin AC) isolated cell nuclei in X-, Y- and Z-axes. The images reveal representative cases with type1 and type2 NI. For the type1 intranuclear inclusion 161 optical sections of 0.1 µm were imaged and 113 of them were used for 3D reconstruction. The type2 intranuclear inclusion was reconstructed from 173 of 213 optical Sects. (0.1 µm). The type1 intranuclear inclusion is limited by a lamin AC (nuclear membrane marker) immunopositive membrane. The inclusion is completely closed in all planes and there is no DAPI (blue) staining within it. Lamin AC-positive immunostaining (light green) is also present in the nuclear membrane. The images of the type2 intranuclear inclusion in the bottom row show only immunopositivity for lamin AC in the nuclear membrane while the nucleoplasm appears “empty” in all planes.
Type1 NI contain autophagy-associated proteins
Using immunohistochemistry, we examined whether there is a relationship between the manifestation of NI and autophagy. p62, ubiquitin, LC3B, cathepsin B and cathepsin D immunohistochemistry was performed in all 77 cases (Fig. 5). One case did not have any NI, therefore the total number of cases was 76. NI, which showed positive immunostaining for these autophagy-associated proteins, were counted individually. Most remarkably, none of the NI without a limiting membrane (type2) showed any positive immunostaining within the inclusions; instead these inclusions remained “empty” (right column of Fig. 5). We detected immunoreactivity for the autophagy-associated proteins exclusively in the membrane-bounded NI (type1). For quantitative evaluation of the results, first we focused on the HE stainings of the cases and counted the number of type1 and type2 NI. Thus, for all sections, a total sum of 6,618 NI without a limiting membrane (type2) and 959 membrane-bounded NI (type1) were counted. The total sum of immunopositive NI was for p62: 72 NI (72/959; 7.5%), for LC3B: 35 NI (35/959; 3.6%), for ubiquitin: 52 NI (52/959; 5.4%), for cathepsin B: 120 NI (120/959; 12.5%) and for cathepsin D: 96 NI (96/959; 10%), shown also in the Supplementary Table 1 (Supplementary Digital Content 3). We detected an accumulation of these proteins within NI; regarding p62, LC3B and ubiquitin, the immunostaining intensity was stronger in the NI compared to the surrounding cytoplasm.
Figure 5
Immunohistochemical analysis for autophagy-associated proteins in type1 and type2 NI. Images depict positive immunoreactivity (brown) for autophagy-associated proteins within type1 NI, whereby type2 NI are all immunonegative. Analysis of type1 NI demonstrates strong accumulation of p62, LC3B, ubiquitin, cathepsin B and cathepsin D within the inclusions. Regarding p62, LC3B and ubiquitin this immunopositivity is almost exclusively within the inclusions and nearly lacking in the cytoplasm. Examination of type2 NI shows that these inclusions do not contain any autophagy-associated proteins. The white bar at the bottom left indicates 20 µm and the white bars at the bottom middle and right equals 10 µm. Original magnifications: 400 X (left) and 1,000 X (right).
Immunohistochemical analysis for autophagy-associated proteins in type1 and type2 NI. Images depict positive immunoreactivity (brown) for autophagy-associated proteins within type1 NI, whereby type2 NI are all immunonegative. Analysis of type1 NI demonstrates strong accumulation of p62, LC3B, ubiquitin, cathepsin B and cathepsin D within the inclusions. Regarding p62, LC3B and ubiquitin this immunopositivity is almost exclusively within the inclusions and nearly lacking in the cytoplasm. Examination of type2 NI shows that these inclusions do not contain any autophagy-associated proteins. The white bar at the bottom left indicates 20 µm and the white bars at the bottom middle and right equals 10 µm. Original magnifications: 400 X (left) and 1,000 X (right).Clinical and laboratory data of the study groups.Values are medians with ranges presented in parentheses. *P values correspond to the comparison of NAFL / NASH and n valid cases reports the number of valid NAFL/NASH cases used for the statistical analysis. Statistic tests used were Mann–Whitney-U-tests for continuous factors and two-sided Fisher’s exact test for categorical parameters. P ≤ 0.05 was defined as statistically significant.n, number; NAFL, non-alcoholic fatty liver; NASH, non-alcoholic steatohepatitis; ALT, alanine aminotransferase; BMI, Body Mass Index; AST, aspartate aminotransferase; GGT, gamma-glutamyl transferase; CK18, Cytokeratin18; Lob. Inflam. Grade, lobular inflammation grade.Further, we examined correlations between disease progression and the number of type1 NI with positive immunoreactivity for the autophagy-associated proteins. Since we had shown in Fig. 2 that the number of type1 NI was significantly higher in NASH than in the controls, we related the respective total sum of immunopositive type1 NI in the individual disease groups to the total sum of type1 NI in the individual disease groups. We observed in 2.8–23.8% of the type1 NI immunopositivity for autophagy-associated proteins. However, since the number of type1 NI in the individual groups was different, a comparison of these groups regarding what percentage of these NI are immunopositive remains problematic. Therefore, only a descriptive study was possible. Details are listed in the Supplementary Table 1 (Supplementary Digital Content 3). Interestingly, there was an accumulation of autophagy-associated proteins in NI and these accumulations were only found in type1 NI.
Presence of co-localized autophagy-associated proteins within type1 NI
To investigate the role of autophagy-associated proteins within type1 NI in more detail, we performed 3D reconstruction of immunofluorescence double-labelled (ubiquitin and p62) tissue sections (Fig. 6). The top image in Fig. 6 depicts a representative DAPI-stained nucleus with a membrane-bounded intranuclear inclusion. The images below show positive immunoreactivity for the autophagy-associated proteins p62 and ubiquitin within the same inclusion. On several planes, the co-localization of ubiquitin (red) and p62 (green) is revealed within the intranuclear inclusion shown by the evolution of the merged colour yellow (merged image in Fig. 6).
Figure 6
Three-dimensional imaging of double immunofluorescence-labelled (ubiquitin and p62) tissue sections in X-, Y- and Z-axis. A total of 152 optical sections of 0.1 µm were imaged and 98 of them were used for 3D reconstruction. The image demonstrates a representative case with type1 NI. DAPI staining (blue) depicts the nucleus with an intranuclear inclusion, which contains the autophagy-associated proteins ubiquitin and p62. Additionally, co-localizations of ubiquitin and p62 in the same inclusion are seen, proven by the formation of the strong merged colour yellow (merged image); ubiquitin = red, p62 = green.
Three-dimensional imaging of double immunofluorescence-labelled (ubiquitin and p62) tissue sections in X-, Y- and Z-axis. A total of 152 optical sections of 0.1 µm were imaged and 98 of them were used for 3D reconstruction. The image demonstrates a representative case with type1 NI. DAPI staining (blue) depicts the nucleus with an intranuclear inclusion, which contains the autophagy-associated proteins ubiquitin and p62. Additionally, co-localizations of ubiquitin and p62 in the same inclusion are seen, proven by the formation of the strong merged colour yellow (merged image); ubiquitin = red, p62 = green.Additionally, we prepared a summary figure (Fig. 7), which presents the most important results of our study.
Figure 7
Summary figure representing our main findings. The image depicts a control cell lacking NI and two cells with NI type 1 and 2 pointing out the main characteristics of NI type1 and type2. NI type1 contain autophagy assoziated proteins, proteases and degenerated cell organelles whereby we also detected co-localizations of p62 and ubiquitin in the same NI.
Summary figure representing our main findings. The image depicts a control cell lacking NI and two cells with NI type 1 and 2 pointing out the main characteristics of NI type1 and type2. NI type1 contain autophagy assoziated proteins, proteases and degenerated cell organelles whereby we also detected co-localizations of p62 and ubiquitin in the same NI.
Discussion
As the prevalence of NAFLD is rapidly rising in industrialized countries and because of its association with diabetes and progression to advanced forms such as NASH, it becomes more important to clarify the factors driving its progression. Nuclear vacuolation is often observed in NAFLD[11,28-30] but studies examining the shape and content of these inclusions in detail are lacking. Further, there are no investigations in NAFLD regarding biological function of these inclusions and a possible association with progression. Nuclear inclusions (NI) are generally considered as a morphological feature with no significant biological function.We analyzed these NI in detail and detected two morphologically different NI in NAFLD: first NI delimited by the nuclear membrane (designated as “type1”) and secondly NI without limiting membrane (designated as “type2”) which, appear empty in HE and are known as glycogenated nuclei[5,6]. In the literature also two types of NI are documented; briefly, inclusions without limiting membrane, serving for accumulation of virus particles and glycogen and on the other hand membrane-bounded inclusions[31,32]. This is in line with our results, however, membrane-bounded inclusions are mostly described as pseudoinclusions representing invaginations of cytoplasm into the nucleus[31,32].In contrast, we demonstrate by 3D reconstruction of isolated hepatocyte nuclei (Fig. 4 and Supplementary Movie 1, Supplementary Digital Content 1) that these type1 NI are completely enclosed by the nuclear membrane and thus are located within the nucleus with no contact to the cytoplasm. Our ultrastructural investigations (TEM) of type1 NI verified the nuclear membrane as the origin of the limiting membrane (Fig. 3A,C). We documented accumulations of degenerated cell organelles and heterolysosomes in these NI, which were considerably less in the surrounding cytoplasm (Fig. 3A,C).Similar observations were made by Kleinfeld et al. who described that degenerated organelles such as rough ER and mitochondria are more likely to occur in NI if the opening of the invagination is occluded[33].Thus, we can exclude that these type1 NI are mere invaginations of cytoplasm. Leduc et al. showed by TEM analysis inclusions in HCC bordered by a double membrane originating from the nuclear envelope[18]. They stated these inclusions to be an invagination of the nuclear membrane with a remaining opening to the cytoplasm of varying size. A later closure of the opening would lead to formation of a completely closed intranuclear inclusion without any contact to the cytoplasm. Nonetheless, some invaginations may not close.Taking together the published TEM observations and our data, it is very likely that the inclusions are initially formed by invaginations, later the connection to the cytoplasm closes with the result that the invaginations/inclusions are completely localized in the nucleus.In addition to the ultrastructural differences between type1 and type2 NI both types also differ in their immunohistochemical profile. In type1 NI we detected autophagy-associated proteins and proteases typically located in lysosomes, whereas type2 NI were negative for these proteins. The lysosome proteases cathepsin B and D together with the presence of heterolysosomes in TEM are suggestive for an active proteolytic activity in type1 NI. This proteolytic activity could also explain the nearby degenerated cell organelles, which we and others have observed in NI[20,21,33,34]. This hypothesis is supported by the identification of autophagy related proteins such as LC3B, p62, ubiquitin and cathepsins in NI. The spatial co-localization of p62 and ubiquitin within the same inclusion suggests a proteolytic process similar to autophagy. However, the canonical autophagy takes place in the cytoplasm but not in the nucleus.We observed both NI types in NAFL and NASH but in NASH the number of type2 was about 3 times higher than that of type1. Both types were significantly more frequent in NASH than in the control-group without steatosis (P ≤ 0.001). This indicates that disease progression impacts the presence of NI regardless of the type of inclusion.Further, the number of type1 NI was significantly higher in NAFL than in the controls (P = 0.016) and type1 NI were significantly more frequent in NASH than in NAFL (P = 0.012). Spearman correlations showed significant positive associations between the number of NI for type1 and type2 and the disease progression (Spearman`s rho = 0.659 and rho = 0.428, P ≤ 0.000). However, we have to consider that the number of NI was too small for a possible application in diagnostics. Nevertheless, our study shows that there is a significant correlation between the number of NI and disease progression; this is also consistent with the literature. Though NI can be observed both in normal and neoplastic cells[6,32,33], they are more frequently found in neoplastic cells[20] and are associated with clinical conditions such as diabetes, inflammatory, arteriosclerotic and neoplastic lesions, obesity and NAFLD[8,9,35]. It is documented that NI can be caused by damaging external influences, for example NI induction was observed in mouse livers after colchicine application and in kidney epithelia after chronic lead poisoning and X-ray irradiation[17]. Studies of De Oliveira et al. in a NAFLD/NASH-HCC zebrafish model showed that glycogen accumulation/ballooning degeneration were associated with inflammation in the liver and with cancer progression[36]. Thakur et al. studied in a mouse model of viral hepatitismurine hepatocytes exhibiting nuclear inclusions[37]. They found that the presence of NI correlated with oxidative stress and cellular proliferation.We found that the percentage of type1 NI with immunoposivity for p62 and cathepsin B was higher in NASH than in the controls without steatosis. The association of hepatocellular carcinogenesis with p62 is documented[38-41]. In mouse HCC models it was shown that p62 is required in progression from premalignancy to malignancy; high p62 expression protected HCC-initiating cells from oxidative stress-induced death[41]. In addition, p62/SQSTM1-positive aggregates in liver tissue contributed to progression in NASH and HCC[38] and p62 induced reactive-oxygen-species production and promoted hepatocarcinogenesis by an amplification of inflammation[39]. The involvement of cathepsin B in hepatic injury and fibrogenesis is also documented[42]. Canbay et al. observed that cathepsin B inactivation attenuates tissue damage in the hepatocytes of mice. Since we detected a higher percentage of cathepsin B immunopositive NI in NASH than in the controls we assume that also proteolytic proteases play a role in disease progression.In our study, we were able to clearly define two different types of NI in NAFLD, which can be distinguished in terms of morphological features, shape, and content. Thus, we suggest that these type1 NI are not simple invaginations but probably play a role in autophagy and proteolytic processes. We have established reliable criteria for distinguishing the inclusions in NAFLD that accumulate glycogen and virus particles from the inclusions enclosed by the nuclear membrane. Intriguingly, NI gained in number with disease progression and both types were significantly more frequent in advanced NAFLD than in cases without steatosis. We suggest that the occurrence of intranuclear inclusions play a role in NAFLD progression.
Materials and methods
Patients
This study incorporated 77 patients. Of these, a total of 58 were morbidly obesepatients (40 females, 18 males, median age 40.5, range: 20–67 and median BMI 51.7, range: 27.4–78.2) undergoing surgery in a centre for bariatric surgery; this study group consisted of 23 patients with steatosis (NAFL) and 35 with non-alcoholic steatohepatitis (NASH). According to the guidelines of the National Institutes of Health (NIH), the indication for bariatric surgery was established (BMI ≥ 40 kg/m2 or ≥ 35 kg/m2, plus co-morbidities) as described before[3]. The selection criteria for the patients were based on the same system as described in our earlier study[3]. Briefly, patients with excessive alcohol consumption (> 20 g/day in males or > 10 g/day in females) indicating alcoholic liver disease were excluded. The surgeon’s choice—i. e. adjustable gastric band; Roux-Y; or gastric bypass surgery—was in accordance with current guidelines adapted to the patient's clinical conditions and co-morbidities, and on the basis of clinical experience; during the procedure wedge liver biopsies were taken[3]. A control group of 19 organ donors (9 females, 10 males, median age 45, range: 1–80), without steatosis, had a median BMI of 23 (range: 13–44) kg/m2. Fatty infiltration of the liver was totally absent as all organs were dedicated for transplantation.The diagnostic criteria for NASH have been discussed contradictory[43]. Briefly, the activity of NAFLD can be quantified according to NAFLD-activity score (NAS), as established by Kleiner et al.[44] or according to the FLIP algorithm (fatty liver inhibition and progression) proposed by Bedossa et al.[45]. We used the FLIP algorithm by Bedossa for the classification of liver injury in morbid obesity since it allows a more accurate distinction from NAFL to NASH. Based on the histological characteristics “steatosis”, “ballooning of hepatocytes” and “inflammation”, slides were classified as “no steatosis”, “NAFL” or “NASH”. The control group showed no steatosis. More detailed information on the characteristics of the patients is given in Table 2.
Table 2
Clinical and laboratory data of the study groups.
Characteristics
Control (n = 19)
NAFL(n = 23)
NASH(n = 35)
n valid cases
P value*
Age (years)
45 (1–80)
38 (24–67)
45 (20–67)
23/35
0.242
Gender (male/female)
10/9
4/19
14/21
23/35
0.087
BMI (kg/m2)
23 (13–44)
49.9 (29.4–66.9)
52.9 (27.4–78.2)
21/33
0.231
Adiponectin (µg/mL)
–
3.1 (1.3–8.3)
2.95 (0.83–11.9)
21/25
0.991
CK18 M30 (IU/L)
–
184.9 (61.8–807.7)
367.6 (80.1–1,573.9)
21/25
0.001
CK18 M65 (IU/L)
–
333.4 (87.9–960.1)
636.7 (255.8–5,273.1)
20/25
< 0.001
Fasting glucose (mg/dl)
–
96.00 (73–150)
126 (72–385)
23/31
0.001
Total cholesterol (mg/dl)
–
198 (120–261)
177.5 (116–247)
15/18
0.320
Triglyceride (mg/dl)
–
149 (34–218)
207 (55–421)
13/12
0.041
ALT (U/L)
52 (9–3,272)
21 (13–65)
39 (14–120)
23/33
< 0.001
AST (U/L)
86 (20–3,078)
23 (16–49)
34 (23–90)
15/21
< 0.001
GGT (U/L)
135 (0–743)
20 (1.5–93)
41 (14–1,213)
23/33
< 0.001
Fibrosis grade
0/1
–
4/7
6/9
2/3
–
10/2
18/2
23/35
0.79
Steatosis grade
0/1
19/0
0/20
0/8
2/3
0/0
3/0
15/12
23/35
< 0.001
Ballooning grade
0/1
19/0
18/3
0/18
2
0
2
17
23/35
0.002
Lob. Inflam. grade
0/1
19/0
16/4
0/12
2/3
0/0
3/0
21/2
23/35
< 0.001
Values are medians with ranges presented in parentheses. *P values correspond to the comparison of NAFL / NASH and n valid cases reports the number of valid NAFL/NASH cases used for the statistical analysis. Statistic tests used were Mann–Whitney-U-tests for continuous factors and two-sided Fisher’s exact test for categorical parameters. P ≤ 0.05 was defined as statistically significant.
n, number; NAFL, non-alcoholic fatty liver; NASH, non-alcoholic steatohepatitis; ALT, alanine aminotransferase; BMI, Body Mass Index; AST, aspartate aminotransferase; GGT, gamma-glutamyl transferase; CK18, Cytokeratin18; Lob. Inflam. Grade, lobular inflammation grade.
Liver samples of each patient were retrieved from the files of the Institute of Pathology, University Hospital of Essen, Germany. In all cases standard processed formalin fixed and paraffin embedded (FFPE) material was stained with HE and immunohistochemical staining was performed according to institutional standards as described before[20]. From each patient an informed consent was received. The study was conducted in conformity with the Helsinki Declaration of 1975 and was approved by the Ethics Committee of the University Hospital Essen (reference number: 09–4252).
Sample evaluation and immunohistochemistry
Two observers (HAB and JK) evaluated the HE stained slides and diagnosed NASH according to the FLIP algorithm by Bedossa et al.[45]. The NI were counted in the whole slide while total cell count was determined by DAPI staining. From each paraffin block, additional Sections (3–5 μm) were cut, dewaxed and pretreated. Immunohistochemistry was performed as described previously[20]; immunostainings of p62, ubiquitin, LC3B, cathepsin B and cathepsin D were carried out on an automated staining device (Dako Autostainer, Dako, Glostrup, Denmark). The antibodies used were: anti-p62 (#sc-28359, Santa Cruz Biotechnology, CA, USA; diluted 1:1,000 for 30 min at RT), anti-cathepsin B (#sc-6490-R, Santa Cruz Biotechnology; diluted 1:500 for 30 min at RT), anti-cathepsin D (#sc-6486, Santa Cruz Biotechnology; diluted 1:6,000 for 60 min at RT), anti-ubiquitin (#Z0458, Dako; diluted 1:1,000 for 30 min at RT) and anti-LC3B (#3868, Cell Signaling Danvers, MA, USA; diluted 1:20 for 60 min at RT). Negative controls were included in every run and incubated with non-immune immunoglobulin in the same concentrations but instead of the primary antibody. One observer (JK) evaluated the immunohistochemical stains and examined the immunostainings within the nuclear inclusions. Nuclear inclusions were classified as positively or negatively stained. Additionally, tissue sections were examined for lamin AC (nuclear membrane marker) immunoreactivity in the NI by immunofluorescence studies; briefly the primary antibody anti-lamin AC (#ab193904, Abcam, Cambridge, UK; diluted 1:50 for 60 min at RT) was labelled with the secondary antibody Alexa Fluor 488-conjugated chicken anti-rabbit IgG (#A21441, Thermo Fisher Scientific, Waltham, MA, USA; diluted 1:100 for 60 min at RT). DNA staining was performed with DAPI (Sigma-Aldrich, Steinheim, Germany) and after this cells were mounted in anti-quenching medium (Vectashield; Vector Laboratories, Inc. Burlingame, CA, USA). Image analysis was performed as described before[20]. Accumulation of glycogen in the NI was checked by PAS staining in combination with PAS-Diastase to proof glycogen infiltration.
Transmission electron microscopy (TEM)
Ultrastructural analysis of NI was performed as described previously[20]. Briefly, for TEM, fresh liver tissue was taken as a wedge biopsy during bariatric surgery. It was fixed in 2% glutaraldehyde in 0.1 M cacodylate buffer (cB), pH 7.3 for 4 h at room temperature (RT), then washed in cb, post-fixed in 1% osmium tetroxide in cb, dehydrated in a graded series of alcohol and embedded in epoxy resin. Semi-thin sections were stained with basic fuchsin and methylene blue to specify capable blocks for ultrathin sections to mount on copper grids. The ultrathin sections were contrasted with uranyl acetate (1%) and lead citrate (0.4%) and examined using a Zeiss TEM 902A (Zeiss, Oberkochen, Germany). We used a slow-scan-CCD camera and the ITEM 5.2 software (both Olympus Soft-imaging-Systems, Münster, Germany) for digital image acquisition.
3D reconstruction of immunofluorescence-labelled isolated nuclei
An accurate location of an inclusion can only be retraced by scanning the whole nucleus. Thus, we accomplished 3D imaging on isolated hepatoycte nuclei as previously reported with minor modifications[20]. For immunoflouorescence nuclei of representative samples were isolated. Paraffin Sections (60 μm) were deparaffinized using xylene in a 1.5 ml reaction tube. After discarding the supernatant, the pellet was rehydrated in 99%, 96% and 70% ethanol and the supernatant was discarded. The pellet was washed in Target Retrievel Solution pH9 (#S2367, Dako), centrifuged and the supernatant was removed. After resuspension of the pellet in 400 µl Target Retrievel Solution pH9 and disruption by a homogenizer, Heat-induced epitope retrievel (HIER) was performed at 98 °C for 60 min and the suspension was cooled at RT for 30 min. Cells were equilibrated in a antibody diluent composite with Dako REAL Antibody Diluent (#S2022, Dako) containing additionally 2% BSA and 0.5% saponin for 30 min at RT and afterwards incubated with the primary antibody lamin AC (#Ab108595, Abcam, diluted 1:50 in the antibody diluent composite) overnight at 4°. After centrifugation and washing, cells were incubated with the secondary antibody Alexa Fluor 488-conjugated chicken anti-rabbit IgG (#A21441, Thermo Fisher Scientific) in a dilution of 1:100 in the antibody diluent composite for 60 min at RT. Nuclear staining was performed with DAPI (Sigma-Aldrich) before applying anti-quenching medium to the cells (Vectashield). Images were attained using a Leica TCS SP8 STED confocal microscope (Leica Microsystems, Illinois, USA) and were analyzed with the Application Suite X software (Leica Microsystems). The open source software Fiji (ImageJ; www.fiji.sc) was used for the 3D reconstruction and the Huygens Professional software (Scientific Volume Imaging; www.svi.nl/ContactSVI) was used for generation of the z-stacks.
Double immunofluorescence and 3D reconstruction
Three-dimensional localization of p62 and ubiquitin was analyzed by double immunofluorescence staining and followed by reconstruction as previously described with minor modifications[20]. One-μm FFPE tissue sections were cut, dewaxed, rehydrated and pretreated with Target Retrieval Solution pH9 (#S2367, Dako) for 20 min at 97 °C.For double labelling immunofluorescence the primary antibodies anti-p62 (#BML PW9860, Enzo, Life Sciences, NY, USA, diluted 1:250 for 60 min at RT) and anti-ubiquitin (#NB300-130, Novus, Littleton, CO, diluted 1:100 for 30 min at RT) were used. Secondary antibodies used were Alexa Fluor 488-conjugated donkey anti-rabbit IgG (#A21206, Thermo Fisher Scientific) and Alexa Fluor 594-conjugated goat anti-mouseIgG1 (#A21125, Thermo Fisher Scientific). Anti-p62 antibody was labelled with Alexa Fluor 488 and anti-ubiquitin antibody with Alexa Fluor 594. After 1:100 dilution they were incubated for 60 min at RT. DNA was stained with DAPI and image analysis was performed as described above.
Statistics
Analyses were performed with the Statistical Package for Social Sciences (SPSS 24.0, Chicago, IL, USA). We used Mann–Whitney-U-test and Kruskal–Wallis test for continuous factors and two-sided Fisher’s exact test for categorical parameters. Additionally, Spearman's rho correlations were performed to determine correlations between the variables. All data are shown as medians with ranges presented in parentheses, if not stated otherwise; P ≤ 0.05 was defined as statistically significant.Supplementary Table 1.Supplementary Movie 1.Supplementary Movie 2.
Authors: David E Kleiner; Elizabeth M Brunt; Mark Van Natta; Cynthia Behling; Melissa J Contos; Oscar W Cummings; Linda D Ferrell; Yao-Chang Liu; Michael S Torbenson; Aynur Unalp-Arida; Matthew Yeh; Arthur J McCullough; Arun J Sanyal Journal: Hepatology Date: 2005-06 Impact factor: 17.425
Authors: Daniel J Klionsky; Kotb Abdelmohsen; Akihisa Abe; Md Joynal Abedin; Hagai Abeliovich; Abraham Acevedo Arozena; Hiroaki Adachi; Christopher M Adams; Peter D Adams; Khosrow Adeli; Peter J Adhihetty; Sharon G Adler; Galila Agam; Rajesh Agarwal; Manish K Aghi; Maria Agnello; Patrizia Agostinis; Patricia V Aguilar; Julio Aguirre-Ghiso; Edoardo M Airoldi; Slimane Ait-Si-Ali; Takahiko Akematsu; Emmanuel T Akporiaye; Mohamed Al-Rubeai; Guillermo M Albaiceta; Chris Albanese; Diego Albani; Matthew L Albert; Jesus Aldudo; Hana Algül; Mehrdad Alirezaei; Iraide Alloza; Alexandru Almasan; Maylin Almonte-Beceril; Emad S Alnemri; Covadonga Alonso; Nihal Altan-Bonnet; Dario C Altieri; Silvia Alvarez; Lydia Alvarez-Erviti; Sandro Alves; Giuseppina Amadoro; Atsuo Amano; Consuelo Amantini; Santiago Ambrosio; Ivano Amelio; Amal O Amer; Mohamed Amessou; Angelika Amon; Zhenyi An; Frank A Anania; Stig U Andersen; Usha P Andley; Catherine K Andreadi; Nathalie Andrieu-Abadie; Alberto Anel; David K Ann; Shailendra Anoopkumar-Dukie; Manuela Antonioli; Hiroshi Aoki; Nadezda Apostolova; Saveria Aquila; Katia Aquilano; Koichi Araki; Eli Arama; Agustin Aranda; Jun Araya; Alexandre Arcaro; Esperanza Arias; Hirokazu Arimoto; Aileen R Ariosa; Jane L Armstrong; Thierry Arnould; Ivica Arsov; Katsuhiko Asanuma; Valerie Askanas; Eric Asselin; Ryuichiro Atarashi; Sally S Atherton; Julie D Atkin; Laura D Attardi; Patrick Auberger; Georg Auburger; Laure Aurelian; Riccardo Autelli; Laura Avagliano; Maria Laura Avantaggiati; Limor Avrahami; Suresh Awale; Neelam Azad; Tiziana Bachetti; Jonathan M Backer; Dong-Hun Bae; Jae-Sung Bae; Ok-Nam Bae; Soo Han Bae; Eric H Baehrecke; Seung-Hoon Baek; Stephen Baghdiguian; Agnieszka Bagniewska-Zadworna; Hua Bai; Jie Bai; Xue-Yuan Bai; Yannick Bailly; Kithiganahalli Narayanaswamy Balaji; Walter Balduini; Andrea Ballabio; Rena Balzan; Rajkumar Banerjee; Gábor Bánhegyi; Haijun Bao; Benoit Barbeau; Maria D Barrachina; Esther Barreiro; Bonnie Bartel; Alberto Bartolomé; Diane C Bassham; Maria Teresa Bassi; Robert C Bast; Alakananda Basu; Maria Teresa Batista; Henri Batoko; Maurizio Battino; Kyle Bauckman; Bradley L Baumgarner; K Ulrich Bayer; Rupert Beale; Jean-François Beaulieu; George R Beck; Christoph Becker; J David Beckham; Pierre-André Bédard; Patrick J Bednarski; Thomas J Begley; Christian Behl; Christian Behrends; Georg Mn Behrens; Kevin E Behrns; Eloy Bejarano; Amine Belaid; Francesca Belleudi; Giovanni Bénard; Guy Berchem; Daniele Bergamaschi; Matteo Bergami; Ben Berkhout; Laura Berliocchi; Amélie Bernard; Monique Bernard; Francesca Bernassola; Anne Bertolotti; Amanda S Bess; Sébastien Besteiro; Saverio Bettuzzi; Savita Bhalla; Shalmoli Bhattacharyya; Sujit K Bhutia; Caroline Biagosch; Michele Wolfe Bianchi; Martine Biard-Piechaczyk; Viktor Billes; Claudia Bincoletto; Baris Bingol; Sara W Bird; Marc Bitoun; Ivana Bjedov; Craig Blackstone; Lionel Blanc; Guillermo A Blanco; Heidi Kiil Blomhoff; Emilio Boada-Romero; Stefan Böckler; Marianne Boes; Kathleen Boesze-Battaglia; Lawrence H Boise; Alessandra Bolino; Andrea Boman; Paolo Bonaldo; Matteo Bordi; Jürgen Bosch; Luis M Botana; Joelle Botti; German Bou; Marina Bouché; Marion Bouchecareilh; Marie-Josée Boucher; Michael E Boulton; Sebastien G Bouret; Patricia Boya; Michaël Boyer-Guittaut; Peter V Bozhkov; Nathan Brady; Vania Mm Braga; Claudio Brancolini; Gerhard H Braus; José M Bravo-San Pedro; Lisa A Brennan; Emery H Bresnick; Patrick Brest; Dave Bridges; Marie-Agnès Bringer; Marisa Brini; Glauber C Brito; Bertha Brodin; Paul S Brookes; Eric J Brown; Karen Brown; Hal E Broxmeyer; Alain Bruhat; Patricia Chakur Brum; John H Brumell; Nicola Brunetti-Pierri; Robert J Bryson-Richardson; Shilpa Buch; Alastair M Buchan; Hikmet Budak; Dmitry V Bulavin; Scott J Bultman; Geert Bultynck; Vladimir Bumbasirevic; Yan Burelle; Robert E Burke; Margit Burmeister; Peter Bütikofer; Laura Caberlotto; Ken Cadwell; Monika Cahova; Dongsheng Cai; Jingjing Cai; Qian Cai; Sara Calatayud; Nadine Camougrand; Michelangelo Campanella; Grant R Campbell; Matthew Campbell; Silvia Campello; Robin Candau; Isabella Caniggia; Lavinia Cantoni; Lizhi Cao; Allan B Caplan; Michele Caraglia; Claudio Cardinali; Sandra Morais Cardoso; Jennifer S Carew; Laura A Carleton; Cathleen R Carlin; Silvia Carloni; Sven R Carlsson; Didac Carmona-Gutierrez; Leticia Am Carneiro; Oliana Carnevali; Serena Carra; Alice Carrier; Bernadette Carroll; Caty Casas; Josefina Casas; Giuliana Cassinelli; Perrine Castets; Susana Castro-Obregon; Gabriella Cavallini; Isabella Ceccherini; Francesco Cecconi; Arthur I Cederbaum; Valentín Ceña; Simone Cenci; Claudia Cerella; Davide Cervia; Silvia Cetrullo; Hassan Chaachouay; Han-Jung Chae; Andrei S Chagin; Chee-Yin Chai; Gopal Chakrabarti; Georgios Chamilos; Edmond Yw Chan; Matthew Tv Chan; Dhyan Chandra; Pallavi Chandra; Chih-Peng Chang; Raymond Chuen-Chung Chang; Ta Yuan Chang; John C Chatham; Saurabh Chatterjee; Santosh Chauhan; Yongsheng Che; Michael E Cheetham; Rajkumar Cheluvappa; Chun-Jung Chen; Gang Chen; Guang-Chao Chen; Guoqiang Chen; Hongzhuan Chen; Jeff W Chen; Jian-Kang Chen; Min Chen; Mingzhou Chen; Peiwen Chen; Qi Chen; Quan Chen; Shang-Der Chen; Si Chen; Steve S-L Chen; Wei Chen; Wei-Jung Chen; Wen Qiang Chen; Wenli Chen; Xiangmei Chen; Yau-Hung Chen; Ye-Guang Chen; Yin Chen; Yingyu Chen; Yongshun Chen; Yu-Jen Chen; Yue-Qin Chen; Yujie Chen; Zhen Chen; Zhong Chen; Alan Cheng; Christopher Hk Cheng; Hua Cheng; Heesun Cheong; Sara Cherry; Jason Chesney; Chun Hei Antonio Cheung; Eric Chevet; Hsiang Cheng Chi; Sung-Gil Chi; Fulvio Chiacchiera; Hui-Ling Chiang; Roberto Chiarelli; Mario Chiariello; Marcello Chieppa; Lih-Shen Chin; Mario Chiong; Gigi Nc Chiu; Dong-Hyung Cho; Ssang-Goo Cho; William C Cho; Yong-Yeon Cho; Young-Seok Cho; Augustine Mk Choi; Eui-Ju Choi; Eun-Kyoung Choi; Jayoung Choi; Mary E Choi; Seung-Il Choi; Tsui-Fen Chou; Salem Chouaib; Divaker Choubey; Vinay Choubey; Kuan-Chih Chow; Kamal Chowdhury; Charleen T Chu; Tsung-Hsien Chuang; Taehoon Chun; Hyewon Chung; Taijoon Chung; Yuen-Li Chung; Yong-Joon Chwae; Valentina Cianfanelli; Roberto Ciarcia; Iwona A Ciechomska; Maria Rosa Ciriolo; Mara Cirone; Sofie Claerhout; Michael J Clague; Joan Clària; Peter Gh Clarke; Robert Clarke; Emilio Clementi; Cédric Cleyrat; Miriam Cnop; Eliana M Coccia; Tiziana Cocco; Patrice Codogno; Jörn Coers; Ezra Ew Cohen; David Colecchia; Luisa Coletto; Núria S Coll; Emma Colucci-Guyon; Sergio Comincini; Maria Condello; Katherine L Cook; Graham H Coombs; Cynthia D Cooper; J Mark Cooper; Isabelle Coppens; Maria Tiziana Corasaniti; Marco Corazzari; Ramon Corbalan; Elisabeth Corcelle-Termeau; Mario D Cordero; Cristina Corral-Ramos; Olga Corti; Andrea Cossarizza; Paola Costelli; Safia Costes; Susan L Cotman; Ana Coto-Montes; Sandra Cottet; Eduardo Couve; Lori R Covey; L Ashley Cowart; Jeffery S Cox; Fraser P Coxon; Carolyn B Coyne; Mark S Cragg; Rolf J Craven; Tiziana Crepaldi; Jose L Crespo; Alfredo Criollo; Valeria Crippa; Maria Teresa Cruz; Ana Maria Cuervo; Jose M Cuezva; Taixing Cui; Pedro R Cutillas; Mark J Czaja; Maria F Czyzyk-Krzeska; Ruben K Dagda; Uta Dahmen; Chunsun Dai; Wenjie Dai; Yun Dai; Kevin N Dalby; Luisa Dalla Valle; Guillaume Dalmasso; Marcello D'Amelio; Markus Damme; Arlette Darfeuille-Michaud; Catherine Dargemont; Victor M Darley-Usmar; Srinivasan Dasarathy; Biplab Dasgupta; Srikanta Dash; Crispin R Dass; Hazel Marie Davey; Lester M Davids; David Dávila; Roger J Davis; Ted M Dawson; Valina L Dawson; Paula Daza; Jackie de Belleroche; Paul de Figueiredo; Regina Celia Bressan Queiroz de Figueiredo; José de la Fuente; Luisa De Martino; Antonella De Matteis; Guido Ry De Meyer; Angelo De Milito; Mauro De Santi; Wanderley de Souza; Vincenzo De Tata; Daniela De Zio; Jayanta Debnath; Reinhard Dechant; Jean-Paul Decuypere; Shane Deegan; Benjamin Dehay; Barbara Del Bello; Dominic P Del Re; Régis Delage-Mourroux; Lea Md Delbridge; Louise Deldicque; Elizabeth Delorme-Axford; Yizhen Deng; Joern Dengjel; Melanie Denizot; Paul Dent; Channing J Der; Vojo Deretic; Benoît Derrien; Eric Deutsch; Timothy P Devarenne; Rodney J Devenish; Sabrina Di Bartolomeo; Nicola Di Daniele; Fabio Di Domenico; Alessia Di Nardo; Simone Di Paola; Antonio Di Pietro; Livia Di Renzo; Aaron DiAntonio; Guillermo Díaz-Araya; Ines Díaz-Laviada; Maria T Diaz-Meco; Javier Diaz-Nido; Chad A Dickey; Robert C Dickson; Marc Diederich; Paul Digard; Ivan Dikic; Savithrama P Dinesh-Kumar; Chan Ding; Wen-Xing Ding; Zufeng Ding; Luciana Dini; Jörg Hw Distler; Abhinav Diwan; Mojgan Djavaheri-Mergny; Kostyantyn Dmytruk; Renwick Cj Dobson; Volker Doetsch; Karol Dokladny; Svetlana Dokudovskaya; Massimo Donadelli; X Charlie Dong; Xiaonan Dong; Zheng Dong; Terrence M Donohue; Kelly S Doran; Gabriella D'Orazi; Gerald W Dorn; Victor Dosenko; Sami Dridi; Liat Drucker; Jie Du; Li-Lin Du; Lihuan Du; André du Toit; Priyamvada Dua; Lei Duan; Pu Duann; Vikash Kumar Dubey; Michael R Duchen; Michel A Duchosal; Helene Duez; Isabelle Dugail; Verónica I Dumit; Mara C Duncan; Elaine A Dunlop; William A Dunn; Nicolas Dupont; Luc Dupuis; Raúl V Durán; Thomas M Durcan; Stéphane Duvezin-Caubet; Umamaheswar Duvvuri; Vinay Eapen; Darius Ebrahimi-Fakhari; Arnaud Echard; Leopold Eckhart; Charles L Edelstein; Aimee L Edinger; Ludwig Eichinger; Tobias Eisenberg; Avital Eisenberg-Lerner; N Tony Eissa; Wafik S El-Deiry; Victoria El-Khoury; Zvulun Elazar; Hagit Eldar-Finkelman; Chris Jh Elliott; Enzo Emanuele; Urban Emmenegger; Nikolai Engedal; Anna-Mart Engelbrecht; Simone Engelender; Jorrit M Enserink; Ralf Erdmann; Jekaterina Erenpreisa; Rajaraman Eri; Jason L Eriksen; Andreja Erman; Ricardo Escalante; Eeva-Liisa Eskelinen; Lucile Espert; Lorena Esteban-Martínez; Thomas J Evans; Mario Fabri; Gemma Fabrias; Cinzia Fabrizi; Antonio Facchiano; Nils J Færgeman; Alberto Faggioni; W Douglas Fairlie; Chunhai Fan; Daping Fan; Jie Fan; Shengyun Fang; Manolis Fanto; Alessandro Fanzani; Thomas Farkas; Mathias Faure; Francois B Favier; Howard Fearnhead; Massimo Federici; Erkang Fei; Tania C Felizardo; Hua Feng; Yibin Feng; Yuchen Feng; Thomas A Ferguson; Álvaro F Fernández; Maite G Fernandez-Barrena; Jose C Fernandez-Checa; Arsenio Fernández-López; Martin E Fernandez-Zapico; Olivier Feron; Elisabetta Ferraro; Carmen Veríssima Ferreira-Halder; Laszlo Fesus; Ralph Feuer; Fabienne C Fiesel; Eduardo C Filippi-Chiela; Giuseppe Filomeni; Gian Maria Fimia; John H Fingert; Steven Finkbeiner; Toren Finkel; Filomena Fiorito; Paul B Fisher; Marc Flajolet; Flavio Flamigni; Oliver Florey; Salvatore Florio; R Andres Floto; Marco Folini; Carlo Follo; Edward A Fon; Francesco Fornai; Franco Fortunato; Alessandro Fraldi; Rodrigo Franco; Arnaud Francois; Aurélie François; Lisa B Frankel; Iain Dc Fraser; Norbert Frey; Damien G Freyssenet; Christian Frezza; Scott L Friedman; Daniel E Frigo; Dongxu Fu; José M Fuentes; Juan Fueyo; Yoshio Fujitani; Yuuki Fujiwara; Mikihiro Fujiya; Mitsunori Fukuda; Simone Fulda; Carmela Fusco; Bozena Gabryel; Matthias Gaestel; Philippe Gailly; Malgorzata Gajewska; Sehamuddin Galadari; Gad Galili; Inmaculada Galindo; Maria F Galindo; Giovanna Galliciotti; Lorenzo Galluzzi; Luca Galluzzi; Vincent Galy; Noor Gammoh; Sam Gandy; Anand K Ganesan; Swamynathan Ganesan; Ian G Ganley; Monique Gannagé; Fen-Biao Gao; Feng Gao; Jian-Xin Gao; Lorena García Nannig; Eleonora García Véscovi; Marina Garcia-Macía; Carmen Garcia-Ruiz; Abhishek D Garg; Pramod Kumar Garg; Ricardo Gargini; Nils Christian Gassen; Damián Gatica; Evelina Gatti; Julie Gavard; Evripidis Gavathiotis; Liang Ge; Pengfei Ge; Shengfang Ge; Po-Wu Gean; Vania Gelmetti; Armando A Genazzani; Jiefei Geng; Pascal Genschik; Lisa Gerner; Jason E Gestwicki; David A Gewirtz; Saeid Ghavami; Eric Ghigo; Debabrata Ghosh; Anna Maria Giammarioli; Francesca Giampieri; Claudia Giampietri; Alexandra Giatromanolaki; Derrick J Gibbings; Lara Gibellini; Spencer B Gibson; Vanessa Ginet; Antonio Giordano; Flaviano Giorgini; Elisa Giovannetti; Stephen E Girardin; Suzana Gispert; Sandy Giuliano; Candece L Gladson; Alvaro Glavic; Martin Gleave; Nelly Godefroy; Robert M Gogal; Kuppan Gokulan; Gustavo H Goldman; Delia Goletti; Michael S Goligorsky; Aldrin V Gomes; Ligia C Gomes; Hernando Gomez; Candelaria Gomez-Manzano; Rubén Gómez-Sánchez; Dawit Ap Gonçalves; Ebru Goncu; Qingqiu Gong; Céline Gongora; Carlos B Gonzalez; Pedro Gonzalez-Alegre; Pilar Gonzalez-Cabo; Rosa Ana González-Polo; Ing Swie Goping; Carlos Gorbea; Nikolai V Gorbunov; Daphne R Goring; Adrienne M Gorman; Sharon M Gorski; Sandro Goruppi; Shino Goto-Yamada; Cecilia Gotor; Roberta A Gottlieb; Illana Gozes; Devrim Gozuacik; Yacine Graba; Martin Graef; Giovanna E Granato; Gary Dean Grant; Steven Grant; Giovanni Luca Gravina; Douglas R Green; Alexander Greenhough; Michael T Greenwood; Benedetto Grimaldi; Frédéric Gros; Charles Grose; Jean-Francois Groulx; Florian Gruber; Paolo Grumati; Tilman Grune; Jun-Lin Guan; Kun-Liang Guan; Barbara Guerra; Carlos Guillen; Kailash Gulshan; Jan Gunst; Chuanyong Guo; Lei Guo; Ming Guo; Wenjie Guo; Xu-Guang Guo; Andrea A Gust; Åsa B Gustafsson; Elaine Gutierrez; Maximiliano G Gutierrez; Ho-Shin Gwak; Albert Haas; James E Haber; Shinji Hadano; Monica Hagedorn; David R Hahn; Andrew J Halayko; Anne Hamacher-Brady; Kozo Hamada; Ahmed Hamai; Andrea Hamann; Maho Hamasaki; Isabelle Hamer; Qutayba Hamid; Ester M Hammond; Feng Han; Weidong Han; James T Handa; John A Hanover; Malene Hansen; Masaru Harada; Ljubica Harhaji-Trajkovic; J Wade Harper; Abdel Halim Harrath; Adrian L Harris; James Harris; Udo Hasler; Peter Hasselblatt; Kazuhisa Hasui; Robert G Hawley; Teresa S Hawley; Congcong He; Cynthia Y He; Fengtian He; Gu He; Rong-Rong He; Xian-Hui He; You-Wen He; Yu-Ying He; Joan K Heath; Marie-Josée Hébert; Robert A Heinzen; Gudmundur Vignir Helgason; Michael Hensel; Elizabeth P Henske; Chengtao Her; Paul K Herman; Agustín Hernández; Carlos Hernandez; Sonia Hernández-Tiedra; Claudio Hetz; P Robin Hiesinger; Katsumi Higaki; Sabine Hilfiker; Bradford G Hill; Joseph A Hill; William D Hill; Keisuke Hino; Daniel Hofius; Paul Hofman; Günter U Höglinger; Jörg Höhfeld; Marina K Holz; Yonggeun Hong; David A Hood; Jeroen Jm Hoozemans; Thorsten Hoppe; Chin Hsu; Chin-Yuan Hsu; Li-Chung Hsu; Dong Hu; Guochang Hu; Hong-Ming Hu; Hongbo Hu; Ming Chang Hu; Yu-Chen Hu; Zhuo-Wei Hu; Fang Hua; Ya Hua; Canhua Huang; Huey-Lan Huang; Kuo-How Huang; Kuo-Yang Huang; Shile Huang; Shiqian Huang; Wei-Pang Huang; Yi-Ran Huang; Yong Huang; Yunfei Huang; Tobias B Huber; Patricia Huebbe; Won-Ki Huh; Juha J Hulmi; Gang Min Hur; James H Hurley; Zvenyslava Husak; Sabah Na Hussain; Salik Hussain; Jung Jin Hwang; Seungmin Hwang; Thomas Is Hwang; Atsuhiro Ichihara; Yuzuru Imai; Carol Imbriano; Megumi Inomata; Takeshi Into; Valentina Iovane; Juan L Iovanna; Renato V Iozzo; Nancy Y Ip; Javier E Irazoqui; Pablo Iribarren; Yoshitaka Isaka; Aleksandra J Isakovic; Harry Ischiropoulos; Jeffrey S Isenberg; Mohammad Ishaq; Hiroyuki Ishida; Isao Ishii; Jane E Ishmael; Ciro Isidoro; Ken-Ichi Isobe; Erika Isono; Shohreh Issazadeh-Navikas; Koji Itahana; Eisuke Itakura; Andrei I Ivanov; Anand Krishnan V Iyer; José M Izquierdo; Yotaro Izumi; Valentina Izzo; Marja Jäättelä; Nadia Jaber; Daniel John Jackson; William T Jackson; Tony George Jacob; Thomas S Jacques; Chinnaswamy Jagannath; Ashish Jain; Nihar Ranjan Jana; Byoung Kuk Jang; Alkesh Jani; Bassam Janji; Paulo Roberto Jannig; Patric J Jansson; Steve Jean; Marina Jendrach; Ju-Hong Jeon; Niels Jessen; Eui-Bae Jeung; Kailiang Jia; Lijun Jia; Hong Jiang; Hongchi Jiang; Liwen Jiang; Teng Jiang; Xiaoyan Jiang; Xuejun Jiang; Xuejun Jiang; Ying Jiang; Yongjun Jiang; Alberto Jiménez; Cheng Jin; Hongchuan Jin; Lei Jin; Meiyan Jin; Shengkan Jin; Umesh Kumar Jinwal; Eun-Kyeong Jo; Terje Johansen; Daniel E Johnson; Gail Vw Johnson; James D Johnson; Eric Jonasch; Chris Jones; Leo Ab Joosten; Joaquin Jordan; Anna-Maria Joseph; Bertrand Joseph; Annie M Joubert; Dianwen Ju; Jingfang Ju; Hsueh-Fen Juan; Katrin Juenemann; Gábor Juhász; Hye Seung Jung; Jae U Jung; Yong-Keun Jung; Heinz Jungbluth; Matthew J Justice; Barry Jutten; Nadeem O Kaakoush; Kai Kaarniranta; Allen Kaasik; Tomohiro Kabuta; Bertrand Kaeffer; Katarina Kågedal; Alon Kahana; Shingo Kajimura; Or Kakhlon; Manjula Kalia; Dhan V Kalvakolanu; Yoshiaki Kamada; Konstantinos Kambas; Vitaliy O Kaminskyy; Harm H Kampinga; Mustapha Kandouz; Chanhee Kang; Rui Kang; Tae-Cheon Kang; Tomotake Kanki; Thirumala-Devi Kanneganti; Haruo Kanno; Anumantha G Kanthasamy; Marc Kantorow; Maria Kaparakis-Liaskos; Orsolya Kapuy; Vassiliki Karantza; Md Razaul Karim; Parimal Karmakar; Arthur Kaser; Susmita Kaushik; Thomas Kawula; A Murat Kaynar; Po-Yuan Ke; Zun-Ji Ke; John H Kehrl; Kate E Keller; Jongsook Kim Kemper; Anne K Kenworthy; Oliver Kepp; Andreas Kern; Santosh Kesari; David Kessel; Robin Ketteler; Isis do Carmo Kettelhut; Bilon Khambu; Muzamil Majid Khan; Vinoth Km Khandelwal; Sangeeta Khare; Juliann G Kiang; Amy A Kiger; Akio Kihara; Arianna L Kim; Cheol Hyeon Kim; Deok Ryong Kim; Do-Hyung Kim; Eung Kweon Kim; Hye Young Kim; Hyung-Ryong Kim; Jae-Sung Kim; Jeong Hun Kim; Jin Cheon Kim; Jin Hyoung Kim; Kwang Woon Kim; Michael D Kim; Moon-Moo Kim; Peter K Kim; Seong Who Kim; Soo-Youl Kim; Yong-Sun Kim; Yonghyun Kim; Adi Kimchi; Alec C Kimmelman; Tomonori Kimura; Jason S King; Karla Kirkegaard; Vladimir Kirkin; Lorrie A Kirshenbaum; Shuji Kishi; Yasuo Kitajima; Katsuhiko Kitamoto; Yasushi Kitaoka; Kaio Kitazato; Rudolf A Kley; Walter T Klimecki; Michael Klinkenberg; Jochen Klucken; Helene Knævelsrud; Erwin Knecht; Laura Knuppertz; Jiunn-Liang Ko; Satoru Kobayashi; Jan C Koch; Christelle Koechlin-Ramonatxo; Ulrich Koenig; Young Ho Koh; Katja Köhler; Sepp D Kohlwein; Masato Koike; Masaaki Komatsu; Eiki Kominami; Dexin Kong; Hee Jeong Kong; Eumorphia G Konstantakou; Benjamin T Kopp; Tamas Korcsmaros; Laura Korhonen; Viktor I Korolchuk; Nadya V Koshkina; Yanjun Kou; Michael I Koukourakis; Constantinos Koumenis; Attila L Kovács; Tibor Kovács; Werner J Kovacs; Daisuke Koya; Claudine Kraft; Dimitri Krainc; Helmut Kramer; Tamara Kravic-Stevovic; Wilhelm Krek; Carole Kretz-Remy; Roswitha Krick; Malathi Krishnamurthy; Janos Kriston-Vizi; Guido Kroemer; Michael C Kruer; Rejko Kruger; Nicholas T Ktistakis; Kazuyuki Kuchitsu; Christian Kuhn; Addanki Pratap Kumar; Anuj Kumar; Ashok Kumar; Deepak Kumar; Dhiraj Kumar; Rakesh Kumar; Sharad Kumar; Mondira Kundu; Hsing-Jien Kung; Atsushi Kuno; Sheng-Han Kuo; Jeff Kuret; Tino Kurz; Terry Kwok; Taeg Kyu Kwon; Yong Tae Kwon; Irene Kyrmizi; Albert R La Spada; Frank Lafont; Tim Lahm; Aparna Lakkaraju; Truong Lam; Trond Lamark; Steve Lancel; Terry H Landowski; Darius J R Lane; Jon D Lane; Cinzia Lanzi; Pierre Lapaquette; Louis R Lapierre; Jocelyn Laporte; Johanna Laukkarinen; Gordon W Laurie; Sergio Lavandero; Lena Lavie; Matthew J LaVoie; Betty Yuen Kwan Law; Helen Ka-Wai Law; Kelsey B Law; Robert Layfield; Pedro A Lazo; Laurent Le Cam; Karine G Le Roch; Hervé Le Stunff; Vijittra Leardkamolkarn; Marc Lecuit; Byung-Hoon Lee; Che-Hsin Lee; Erinna F Lee; Gyun Min Lee; He-Jin Lee; Hsinyu Lee; Jae Keun Lee; Jongdae Lee; Ju-Hyun Lee; Jun Hee Lee; Michael Lee; Myung-Shik Lee; Patty J Lee; Sam W Lee; Seung-Jae Lee; Shiow-Ju Lee; Stella Y Lee; Sug Hyung Lee; Sung Sik Lee; Sung-Joon Lee; Sunhee Lee; Ying-Ray Lee; Yong J Lee; Young H Lee; Christiaan Leeuwenburgh; Sylvain Lefort; Renaud Legouis; Jinzhi Lei; Qun-Ying Lei; David A Leib; Gil Leibowitz; Istvan Lekli; Stéphane D Lemaire; John J Lemasters; Marius K Lemberg; Antoinette Lemoine; Shuilong Leng; Guido Lenz; Paola Lenzi; Lilach O Lerman; Daniele Lettieri Barbato; Julia I-Ju Leu; Hing Y Leung; Beth Levine; Patrick A Lewis; Frank Lezoualc'h; Chi Li; Faqiang Li; Feng-Jun Li; Jun Li; Ke Li; Lian Li; Min Li; Min Li; Qiang Li; Rui Li; Sheng Li; Wei Li; Wei Li; Xiaotao Li; Yumin Li; Jiqin Lian; Chengyu Liang; Qiangrong Liang; Yulin Liao; Joana Liberal; Pawel P Liberski; Pearl Lie; Andrew P Lieberman; Hyunjung Jade Lim; Kah-Leong Lim; Kyu Lim; Raquel T Lima; Chang-Shen Lin; Chiou-Feng Lin; Fang Lin; Fangming Lin; Fu-Cheng Lin; Kui Lin; Kwang-Huei Lin; Pei-Hui Lin; Tianwei Lin; Wan-Wan Lin; Yee-Shin Lin; Yong Lin; Rafael Linden; Dan Lindholm; Lisa M Lindqvist; Paul Lingor; Andreas Linkermann; Lance A Liotta; Marta M Lipinski; Vitor A Lira; Michael P Lisanti; Paloma B Liton; Bo Liu; Chong Liu; Chun-Feng Liu; Fei Liu; Hung-Jen Liu; Jianxun Liu; Jing-Jing Liu; Jing-Lan Liu; Ke Liu; Leyuan Liu; Liang Liu; Quentin Liu; Rong-Yu Liu; Shiming Liu; Shuwen Liu; Wei Liu; Xian-De Liu; Xiangguo Liu; Xiao-Hong Liu; Xinfeng Liu; Xu Liu; Xueqin Liu; Yang Liu; Yule Liu; Zexian Liu; Zhe Liu; Juan P Liuzzi; Gérard Lizard; Mila Ljujic; Irfan J Lodhi; Susan E Logue; Bal L Lokeshwar; Yun Chau Long; Sagar Lonial; Benjamin Loos; Carlos López-Otín; Cristina López-Vicario; Mar Lorente; Philip L Lorenzi; Péter Lõrincz; Marek Los; Michael T Lotze; Penny E Lovat; Binfeng Lu; Bo Lu; Jiahong Lu; Qing Lu; She-Min Lu; Shuyan Lu; Yingying Lu; Frédéric Luciano; Shirley Luckhart; John Milton Lucocq; Paula Ludovico; Aurelia Lugea; Nicholas W Lukacs; Julian J Lum; Anders H Lund; Honglin Luo; Jia Luo; Shouqing Luo; Claudio Luparello; Timothy Lyons; Jianjie Ma; Yi Ma; Yong Ma; Zhenyi Ma; Juliano Machado; Glaucia M Machado-Santelli; Fernando Macian; Gustavo C MacIntosh; Jeffrey P MacKeigan; Kay F Macleod; John D MacMicking; Lee Ann MacMillan-Crow; Frank Madeo; Muniswamy Madesh; Julio Madrigal-Matute; Akiko Maeda; Tatsuya Maeda; Gustavo Maegawa; Emilia Maellaro; Hannelore Maes; Marta Magariños; Kenneth Maiese; Tapas K Maiti; Luigi Maiuri; Maria Chiara Maiuri; Carl G Maki; Roland Malli; Walter Malorni; Alina Maloyan; Fathia Mami-Chouaib; Na Man; Joseph D Mancias; Eva-Maria Mandelkow; Michael A Mandell; Angelo A Manfredi; Serge N Manié; Claudia Manzoni; Kai Mao; Zixu Mao; Zong-Wan Mao; Philippe Marambaud; Anna Maria Marconi; Zvonimir Marelja; Gabriella Marfe; Marta Margeta; Eva Margittai; Muriel Mari; Francesca V Mariani; Concepcio Marin; Sara Marinelli; Guillermo Mariño; Ivanka Markovic; Rebecca Marquez; Alberto M Martelli; Sascha Martens; Katie R Martin; Seamus J Martin; Shaun Martin; Miguel A Martin-Acebes; Paloma Martín-Sanz; Camille Martinand-Mari; Wim Martinet; Jennifer Martinez; Nuria Martinez-Lopez; Ubaldo Martinez-Outschoorn; Moisés Martínez-Velázquez; Marta Martinez-Vicente; Waleska Kerllen Martins; Hirosato Mashima; James A Mastrianni; Giuseppe Matarese; Paola Matarrese; Roberto Mateo; Satoaki Matoba; Naomichi Matsumoto; Takehiko Matsushita; Akira Matsuura; Takeshi Matsuzawa; Mark P Mattson; Soledad Matus; Norma Maugeri; Caroline Mauvezin; Andreas Mayer; Dusica Maysinger; Guillermo D Mazzolini; Mary Kate McBrayer; Kimberly McCall; Craig McCormick; Gerald M McInerney; Skye C McIver; Sharon McKenna; John J McMahon; Iain A McNeish; Fatima Mechta-Grigoriou; Jan Paul Medema; Diego L Medina; Klara Megyeri; Maryam Mehrpour; Jawahar L Mehta; Yide Mei; Ute-Christiane Meier; Alfred J Meijer; Alicia Meléndez; Gerry Melino; Sonia Melino; Edesio Jose Tenorio de Melo; Maria A Mena; Marc D Meneghini; Javier A Menendez; Regina Menezes; Liesu Meng; Ling-Hua Meng; Songshu Meng; Rossella Menghini; A Sue Menko; Rubem Fs Menna-Barreto; Manoj B Menon; Marco A Meraz-Ríos; Giuseppe Merla; Luciano Merlini; Angelica M Merlot; Andreas Meryk; Stefania Meschini; Joel N Meyer; Man-Tian Mi; Chao-Yu Miao; Lucia Micale; Simon Michaeli; Carine Michiels; Anna Rita Migliaccio; Anastasia Susie Mihailidou; Dalibor Mijaljica; Katsuhiko Mikoshiba; Enrico Milan; Leonor Miller-Fleming; Gordon B Mills; Ian G Mills; Georgia Minakaki; Berge A Minassian; Xiu-Fen Ming; Farida Minibayeva; Elena A Minina; Justine D Mintern; Saverio Minucci; Antonio Miranda-Vizuete; Claire H Mitchell; Shigeki Miyamoto; Keisuke Miyazawa; Noboru Mizushima; Katarzyna Mnich; Baharia Mograbi; Simin Mohseni; Luis Ferreira Moita; Marco Molinari; Maurizio Molinari; Andreas Buch Møller; Bertrand Mollereau; Faustino Mollinedo; Marco Mongillo; Martha M Monick; Serena Montagnaro; Craig Montell; Darren J Moore; Michael N Moore; Rodrigo Mora-Rodriguez; Paula I Moreira; Etienne Morel; Maria Beatrice Morelli; Sandra Moreno; Michael J Morgan; Arnaud Moris; Yuji Moriyasu; Janna L Morrison; Lynda A Morrison; Eugenia Morselli; Jorge Moscat; Pope L Moseley; Serge Mostowy; Elisa Motori; Denis Mottet; Jeremy C Mottram; Charbel E-H Moussa; Vassiliki E Mpakou; Hasan Mukhtar; Jean M Mulcahy Levy; Sylviane Muller; Raquel Muñoz-Moreno; Cristina Muñoz-Pinedo; Christian Münz; Maureen E Murphy; James T Murray; Aditya Murthy; Indira U Mysorekar; Ivan R Nabi; Massimo Nabissi; Gustavo A Nader; Yukitoshi Nagahara; Yoshitaka Nagai; Kazuhiro Nagata; Anika Nagelkerke; Péter Nagy; Samisubbu R Naidu; Sreejayan Nair; Hiroyasu Nakano; Hitoshi Nakatogawa; Meera Nanjundan; Gennaro Napolitano; Naweed I Naqvi; Roberta Nardacci; Derek P Narendra; Masashi Narita; Anna Chiara Nascimbeni; Ramesh Natarajan; Luiz C Navegantes; Steffan T Nawrocki; Taras Y Nazarko; Volodymyr Y Nazarko; Thomas Neill; Luca M Neri; Mihai G Netea; Romana T Netea-Maier; Bruno M Neves; Paul A Ney; Ioannis P Nezis; Hang Tt Nguyen; Huu Phuc Nguyen; Anne-Sophie Nicot; Hilde Nilsen; Per Nilsson; Mikio Nishimura; Ichizo Nishino; Mireia Niso-Santano; Hua Niu; Ralph A Nixon; Vincent Co Njar; Takeshi Noda; Angelika A Noegel; Elsie Magdalena Nolte; Erik Norberg; Koenraad K Norga; Sakineh Kazemi Noureini; Shoji Notomi; Lucia Notterpek; Karin Nowikovsky; Nobuyuki Nukina; Thorsten Nürnberger; Valerie B O'Donnell; Tracey O'Donovan; Peter J O'Dwyer; Ina Oehme; Clara L Oeste; Michinaga Ogawa; Besim Ogretmen; Yuji Ogura; Young J Oh; Masaki Ohmuraya; Takayuki Ohshima; Rani Ojha; Koji Okamoto; Toshiro Okazaki; F Javier Oliver; Karin Ollinger; Stefan Olsson; Daniel P Orban; Paulina Ordonez; Idil Orhon; Laszlo Orosz; Eyleen J O'Rourke; Helena Orozco; Angel L Ortega; Elena Ortona; Laura D Osellame; Junko Oshima; Shigeru Oshima; Heinz D Osiewacz; Takanobu Otomo; Kinya Otsu; Jing-Hsiung James Ou; Tiago F Outeiro; Dong-Yun Ouyang; Hongjiao Ouyang; Michael Overholtzer; Michelle A Ozbun; P Hande Ozdinler; Bulent Ozpolat; Consiglia Pacelli; Paolo Paganetti; Guylène Page; Gilles Pages; Ugo Pagnini; Beata Pajak; Stephen C Pak; Karolina Pakos-Zebrucka; Nazzy Pakpour; Zdena Palková; Francesca Palladino; Kathrin Pallauf; Nicolas Pallet; Marta Palmieri; Søren R Paludan; Camilla Palumbo; Silvia Palumbo; Olatz Pampliega; Hongming Pan; Wei Pan; Theocharis Panaretakis; Aseem Pandey; Areti Pantazopoulou; Zuzana Papackova; Daniela L Papademetrio; Issidora Papassideri; Alessio Papini; Nirmala Parajuli; Julian Pardo; Vrajesh V Parekh; Giancarlo Parenti; Jong-In Park; Junsoo Park; Ohkmae K Park; Roy Parker; Rosanna Parlato; Jan B Parys; Katherine R Parzych; Jean-Max Pasquet; Benoit Pasquier; Kishore Bs Pasumarthi; Daniel Patschan; Cam Patterson; Sophie Pattingre; Scott Pattison; Arnim Pause; Hermann Pavenstädt; Flaminia Pavone; Zully Pedrozo; Fernando J Peña; Miguel A Peñalva; Mario Pende; Jianxin Peng; Fabio Penna; Josef M Penninger; Anna Pensalfini; Salvatore Pepe; Gustavo Js Pereira; Paulo C Pereira; Verónica Pérez-de la Cruz; María Esther Pérez-Pérez; Diego Pérez-Rodríguez; Dolores Pérez-Sala; Celine Perier; Andras Perl; David H Perlmutter; Ida Perrotta; Shazib Pervaiz; Maija Pesonen; Jeffrey E Pessin; Godefridus J Peters; Morten Petersen; Irina Petrache; Basil J Petrof; Goran Petrovski; James M Phang; Mauro Piacentini; Marina Pierdominici; Philippe Pierre; Valérie Pierrefite-Carle; Federico Pietrocola; Felipe X Pimentel-Muiños; Mario Pinar; Benjamin Pineda; Ronit Pinkas-Kramarski; Marcello Pinti; Paolo Pinton; Bilal Piperdi; James M Piret; Leonidas C Platanias; Harald W Platta; Edward D Plowey; Stefanie Pöggeler; Marc Poirot; Peter Polčic; Angelo Poletti; Audrey H Poon; Hana Popelka; Blagovesta Popova; Izabela Poprawa; Shibu M Poulose; Joanna Poulton; Scott K Powers; Ted Powers; Mercedes Pozuelo-Rubio; Krisna Prak; Reinhild Prange; Mark Prescott; Muriel Priault; Sharon Prince; Richard L Proia; Tassula Proikas-Cezanne; Holger Prokisch; Vasilis J Promponas; Karin Przyklenk; Rosa Puertollano; Subbiah Pugazhenthi; Luigi Puglielli; Aurora Pujol; Julien Puyal; Dohun Pyeon; Xin Qi; Wen-Bin Qian; Zheng-Hong Qin; Yu Qiu; Ziwei Qu; Joe Quadrilatero; Frederick Quinn; Nina Raben; Hannah Rabinowich; Flavia Radogna; Michael J Ragusa; Mohamed Rahmani; Komal Raina; Sasanka Ramanadham; Rajagopal Ramesh; Abdelhaq Rami; Sarron Randall-Demllo; Felix Randow; Hai Rao; V Ashutosh Rao; Blake B Rasmussen; Tobias M Rasse; Edward A Ratovitski; Pierre-Emmanuel Rautou; Swapan K Ray; Babak Razani; Bruce H Reed; Fulvio Reggiori; Markus Rehm; Andreas S Reichert; Theo Rein; David J Reiner; Eric Reits; Jun Ren; Xingcong Ren; Maurizio Renna; Jane Eb Reusch; Jose L Revuelta; Leticia Reyes; Alireza R Rezaie; Robert I Richards; Des R Richardson; Clémence Richetta; Michael A Riehle; Bertrand H Rihn; Yasuko Rikihisa; Brigit E Riley; Gerald Rimbach; Maria Rita Rippo; Konstantinos Ritis; Federica Rizzi; Elizete Rizzo; Peter J Roach; Jeffrey Robbins; Michel Roberge; Gabriela Roca; Maria Carmela Roccheri; Sonia Rocha; Cecilia Mp Rodrigues; Clara I Rodríguez; Santiago Rodriguez de Cordoba; Natalia Rodriguez-Muela; Jeroen Roelofs; Vladimir V Rogov; Troy T Rohn; Bärbel Rohrer; Davide Romanelli; Luigina Romani; Patricia Silvia Romano; M Isabel G Roncero; Jose Luis Rosa; Alicia Rosello; Kirill V Rosen; Philip Rosenstiel; Magdalena Rost-Roszkowska; Kevin A Roth; Gael Roué; Mustapha Rouis; Kasper M Rouschop; Daniel T Ruan; Diego Ruano; David C Rubinsztein; Edmund B Rucker; Assaf Rudich; Emil Rudolf; Ruediger Rudolf; Markus A Ruegg; Carmen Ruiz-Roldan; Avnika Ashok Ruparelia; Paola Rusmini; David W Russ; Gian Luigi Russo; Giuseppe Russo; Rossella Russo; Tor Erik Rusten; Victoria Ryabovol; Kevin M Ryan; Stefan W Ryter; David M Sabatini; Michael Sacher; Carsten Sachse; Michael N Sack; Junichi Sadoshima; Paul Saftig; Ronit Sagi-Eisenberg; Sumit Sahni; Pothana Saikumar; Tsunenori Saito; Tatsuya Saitoh; Koichi Sakakura; Machiko Sakoh-Nakatogawa; Yasuhito Sakuraba; María Salazar-Roa; Paolo Salomoni; Ashok K Saluja; Paul M Salvaterra; Rosa Salvioli; Afshin Samali; Anthony Mj Sanchez; José A Sánchez-Alcázar; Ricardo Sanchez-Prieto; Marco Sandri; Miguel A Sanjuan; Stefano Santaguida; Laura Santambrogio; Giorgio Santoni; Claudia Nunes Dos Santos; Shweta Saran; Marco Sardiello; Graeme Sargent; Pallabi Sarkar; Sovan Sarkar; Maria Rosa Sarrias; Minnie M Sarwal; Chihiro Sasakawa; Motoko Sasaki; Miklos Sass; Ken Sato; Miyuki Sato; Joseph Satriano; Niramol Savaraj; Svetlana Saveljeva; Liliana Schaefer; Ulrich E Schaible; Michael Scharl; Hermann M Schatzl; Randy Schekman; Wiep Scheper; Alfonso Schiavi; Hyman M Schipper; Hana Schmeisser; Jens Schmidt; Ingo Schmitz; Bianca E Schneider; E Marion Schneider; Jaime L Schneider; Eric A Schon; Miriam J Schönenberger; Axel H Schönthal; Daniel F Schorderet; Bernd Schröder; Sebastian Schuck; Ryan J Schulze; Melanie Schwarten; Thomas L Schwarz; Sebastiano Sciarretta; Kathleen Scotto; A Ivana Scovassi; Robert A Screaton; Mark Screen; Hugo Seca; Simon Sedej; Laura Segatori; Nava Segev; Per O Seglen; Jose M Seguí-Simarro; Juan Segura-Aguilar; Ekihiro Seki; Christian Sell; Iban Seiliez; Clay F Semenkovich; Gregg L Semenza; Utpal Sen; Andreas L Serra; Ana Serrano-Puebla; Hiromi Sesaki; Takao Setoguchi; Carmine Settembre; John J Shacka; Ayesha N Shajahan-Haq; Irving M Shapiro; Shweta Sharma; Hua She; C-K James Shen; Chiung-Chyi Shen; Han-Ming Shen; Sanbing Shen; Weili Shen; Rui Sheng; Xianyong Sheng; Zu-Hang Sheng; Trevor G Shepherd; Junyan Shi; Qiang Shi; Qinghua Shi; Yuguang Shi; Shusaku Shibutani; Kenichi Shibuya; Yoshihiro Shidoji; Jeng-Jer Shieh; Chwen-Ming Shih; Yohta Shimada; Shigeomi Shimizu; Dong Wook Shin; Mari L Shinohara; Michiko Shintani; Takahiro Shintani; Tetsuo Shioi; Ken Shirabe; Ronit Shiri-Sverdlov; Orian Shirihai; Gordon C Shore; Chih-Wen Shu; Deepak Shukla; Andriy A Sibirny; Valentina Sica; Christina J Sigurdson; Einar M Sigurdsson; Puran Singh Sijwali; Beata Sikorska; Wilian A Silveira; Sandrine Silvente-Poirot; Gary A Silverman; Jan Simak; Thomas Simmet; Anna Katharina Simon; Hans-Uwe Simon; Cristiano Simone; Matias Simons; Anne Simonsen; Rajat Singh; Shivendra V Singh; Shrawan K Singh; Debasish Sinha; Sangita Sinha; Frank A Sinicrope; Agnieszka Sirko; Kapil Sirohi; Balindiwe Jn Sishi; Annie Sittler; Parco M Siu; Efthimios Sivridis; Anna Skwarska; Ruth Slack; Iva Slaninová; Nikolai Slavov; Soraya S Smaili; Keiran Sm Smalley; Duncan R Smith; Stefaan J Soenen; Scott A Soleimanpour; Anita Solhaug; Kumaravel Somasundaram; Jin H Son; Avinash Sonawane; Chunjuan Song; Fuyong Song; Hyun Kyu Song; Ju-Xian Song; Wei Song; Kai Y Soo; Anil K Sood; Tuck Wah Soong; Virawudh Soontornniyomkij; Maurizio Sorice; Federica Sotgia; David R Soto-Pantoja; Areechun Sotthibundhu; Maria João Sousa; Herman P Spaink; Paul N Span; Anne Spang; Janet D Sparks; Peter G Speck; Stephen A Spector; Claudia D Spies; Wolfdieter Springer; Daret St Clair; Alessandra Stacchiotti; Bart Staels; Michael T Stang; Daniel T Starczynowski; Petro Starokadomskyy; Clemens Steegborn; John W Steele; Leonidas Stefanis; Joan Steffan; Christine M Stellrecht; Harald Stenmark; Tomasz M Stepkowski; Stęphan T Stern; Craig Stevens; Brent R Stockwell; Veronika Stoka; Zuzana Storchova; Björn Stork; Vassilis Stratoulias; Dimitrios J Stravopodis; Pavel Strnad; Anne Marie Strohecker; Anna-Lena Ström; Per Stromhaug; Jiri Stulik; Yu-Xiong Su; Zhaoliang Su; Carlos S Subauste; Srinivasa Subramaniam; Carolyn M Sue; Sang Won Suh; Xinbing Sui; Supawadee Sukseree; David Sulzer; Fang-Lin Sun; Jiaren Sun; Jun Sun; Shi-Yong Sun; Yang Sun; Yi Sun; Yingjie Sun; Vinod Sundaramoorthy; Joseph Sung; Hidekazu Suzuki; Kuninori Suzuki; Naoki Suzuki; Tadashi Suzuki; Yuichiro J Suzuki; Michele S Swanson; Charles Swanton; Karl Swärd; Ghanshyam Swarup; Sean T Sweeney; Paul W Sylvester; Zsuzsanna Szatmari; Eva Szegezdi; Peter W Szlosarek; Heinrich Taegtmeyer; Marco Tafani; Emmanuel Taillebourg; Stephen Wg Tait; Krisztina Takacs-Vellai; Yoshinori Takahashi; Szabolcs Takáts; Genzou Takemura; Nagio Takigawa; Nicholas J Talbot; Elena Tamagno; Jerome Tamburini; Cai-Ping Tan; Lan Tan; Mei Lan Tan; Ming Tan; Yee-Joo Tan; Keiji Tanaka; Masaki Tanaka; Daolin Tang; Dingzhong Tang; Guomei Tang; Isei Tanida; Kunikazu Tanji; Bakhos A Tannous; Jose A Tapia; Inmaculada Tasset-Cuevas; Marc Tatar; Iman Tavassoly; Nektarios Tavernarakis; Allen Taylor; Graham S Taylor; Gregory A Taylor; J Paul Taylor; Mark J Taylor; Elena V Tchetina; Andrew R Tee; Fatima Teixeira-Clerc; Sucheta Telang; Tewin Tencomnao; Ba-Bie Teng; Ru-Jeng Teng; Faraj Terro; Gianluca Tettamanti; Arianne L Theiss; Anne E Theron; Kelly Jean Thomas; Marcos P Thomé; Paul G Thomes; Andrew Thorburn; Jeremy Thorner; Thomas Thum; Michael Thumm; Teresa Lm Thurston; Ling Tian; Andreas Till; Jenny Pan-Yun Ting; Vladimir I Titorenko; Lilach Toker; Stefano Toldo; Sharon A Tooze; Ivan Topisirovic; Maria Lyngaas Torgersen; Liliana Torosantucci; Alicia Torriglia; Maria Rosaria Torrisi; Cathy Tournier; Roberto Towns; Vladimir Trajkovic; Leonardo H Travassos; Gemma Triola; Durga Nand Tripathi; Daniela Trisciuoglio; Rodrigo Troncoso; Ioannis P Trougakos; Anita C Truttmann; Kuen-Jer Tsai; Mario P Tschan; Yi-Hsin Tseng; Takayuki Tsukuba; Allan Tsung; Andrey S Tsvetkov; Shuiping Tu; Hsing-Yu Tuan; Marco Tucci; David A Tumbarello; Boris Turk; Vito Turk; Robin Fb Turner; Anders A Tveita; Suresh C Tyagi; Makoto Ubukata; Yasuo Uchiyama; Andrej Udelnow; Takashi Ueno; Midori Umekawa; Rika Umemiya-Shirafuji; Benjamin R Underwood; Christian Ungermann; Rodrigo P Ureshino; Ryo Ushioda; Vladimir N Uversky; Néstor L Uzcátegui; Thomas Vaccari; Maria I Vaccaro; Libuše Váchová; Helin Vakifahmetoglu-Norberg; Rut Valdor; Enza Maria Valente; Francois Vallette; Angela M Valverde; Greet Van den Berghe; Ludo Van Den Bosch; Gijs R van den Brink; F Gisou van der Goot; Ida J van der Klei; Luc Jw van der Laan; Wouter G van Doorn; Marjolein van Egmond; Kenneth L van Golen; Luc Van Kaer; Menno van Lookeren Campagne; Peter Vandenabeele; Wim Vandenberghe; Ilse Vanhorebeek; Isabel Varela-Nieto; M Helena Vasconcelos; Radovan Vasko; Demetrios G Vavvas; Ignacio Vega-Naredo; Guillermo Velasco; Athanassios D Velentzas; Panagiotis D Velentzas; Tibor Vellai; Edo Vellenga; Mikkel Holm Vendelbo; Kartik Venkatachalam; Natascia Ventura; Salvador Ventura; Patrícia St Veras; Mireille Verdier; Beata G Vertessy; Andrea Viale; Michel Vidal; Helena L A Vieira; Richard D Vierstra; Nadarajah Vigneswaran; Neeraj Vij; Miquel Vila; Margarita Villar; Victor H Villar; Joan Villarroya; Cécile Vindis; Giampietro Viola; Maria Teresa Viscomi; Giovanni Vitale; Dan T Vogl; Olga V Voitsekhovskaja; Clarissa von Haefen; Karin von Schwarzenberg; Daniel E Voth; Valérie Vouret-Craviari; Kristina Vuori; Jatin M Vyas; Christian Waeber; Cheryl Lyn Walker; Mark J Walker; Jochen Walter; Lei Wan; Xiangbo Wan; Bo Wang; Caihong Wang; Chao-Yung Wang; Chengshu Wang; Chenran Wang; Chuangui Wang; Dong Wang; Fen Wang; Fuxin Wang; Guanghui Wang; Hai-Jie Wang; Haichao Wang; Hong-Gang Wang; Hongmin Wang; Horng-Dar Wang; Jing Wang; Junjun Wang; Mei Wang; Mei-Qing Wang; Pei-Yu Wang; Peng Wang; Richard C Wang; Shuo Wang; Ting-Fang Wang; Xian Wang; Xiao-Jia Wang; Xiao-Wei Wang; Xin Wang; Xuejun Wang; Yan Wang; Yanming Wang; Ying Wang; Ying-Jan Wang; Yipeng Wang; Yu Wang; Yu Tian Wang; Yuqing Wang; Zhi-Nong Wang; Pablo Wappner; Carl Ward; Diane McVey Ward; Gary Warnes; Hirotaka Watada; Yoshihisa Watanabe; Kei Watase; Timothy E Weaver; Colin D Weekes; Jiwu Wei; Thomas Weide; Conrad C Weihl; Günther Weindl; Simone Nardin Weis; Longping Wen; Xin Wen; Yunfei Wen; Benedikt Westermann; Cornelia M Weyand; Anthony R White; Eileen White; J Lindsay Whitton; Alexander J Whitworth; Joëlle Wiels; Franziska Wild; Manon E Wildenberg; Tom Wileman; Deepti Srinivas Wilkinson; Simon Wilkinson; Dieter Willbold; Chris Williams; Katherine Williams; Peter R Williamson; Konstanze F Winklhofer; Steven S Witkin; Stephanie E Wohlgemuth; Thomas Wollert; Ernst J Wolvetang; Esther Wong; G William Wong; Richard W Wong; Vincent Kam Wai Wong; Elizabeth A Woodcock; Karen L Wright; Chunlai Wu; Defeng Wu; Gen Sheng Wu; Jian Wu; Junfang Wu; Mian Wu; Min Wu; Shengzhou Wu; William Kk Wu; Yaohua Wu; Zhenlong Wu; Cristina Pr Xavier; Ramnik J Xavier; Gui-Xian Xia; Tian Xia; Weiliang Xia; Yong Xia; Hengyi Xiao; Jian Xiao; Shi Xiao; Wuhan Xiao; Chuan-Ming Xie; Zhiping Xie; Zhonglin Xie; Maria Xilouri; Yuyan Xiong; Chuanshan Xu; Congfeng Xu; Feng Xu; Haoxing Xu; Hongwei Xu; Jian Xu; Jianzhen Xu; Jinxian Xu; Liang Xu; Xiaolei Xu; Yangqing Xu; Ye Xu; Zhi-Xiang Xu; Ziheng Xu; Yu Xue; Takahiro Yamada; Ai Yamamoto; Koji Yamanaka; Shunhei Yamashina; Shigeko Yamashiro; Bing Yan; Bo Yan; Xianghua Yan; Zhen Yan; Yasuo Yanagi; Dun-Sheng Yang; Jin-Ming Yang; Liu Yang; Minghua Yang; Pei-Ming Yang; Peixin Yang; Qian Yang; Wannian Yang; Wei Yuan Yang; Xuesong Yang; Yi Yang; Ying Yang; Zhifen Yang; Zhihong Yang; Meng-Chao Yao; Pamela J Yao; Xiaofeng Yao; Zhenyu Yao; Zhiyuan Yao; Linda S Yasui; Mingxiang Ye; Barry Yedvobnick; Behzad Yeganeh; Elizabeth S Yeh; Patricia L Yeyati; Fan Yi; Long Yi; Xiao-Ming Yin; Calvin K Yip; Yeong-Min Yoo; Young Hyun Yoo; Seung-Yong Yoon; Ken-Ichi Yoshida; Tamotsu Yoshimori; Ken H Young; Huixin Yu; Jane J Yu; Jin-Tai Yu; Jun Yu; Li Yu; W Haung Yu; Xiao-Fang Yu; Zhengping Yu; Junying Yuan; Zhi-Min Yuan; Beatrice Yjt Yue; Jianbo Yue; Zhenyu Yue; David N Zacks; Eldad Zacksenhaus; Nadia Zaffaroni; Tania Zaglia; Zahra Zakeri; Vincent Zecchini; Jinsheng Zeng; Min Zeng; Qi Zeng; Antonis S Zervos; Donna D Zhang; Fan Zhang; Guo Zhang; Guo-Chang Zhang; Hao Zhang; Hong Zhang; Hong Zhang; Hongbing Zhang; Jian Zhang; Jian Zhang; Jiangwei Zhang; Jianhua Zhang; Jing-Pu Zhang; Li Zhang; Lin Zhang; Lin Zhang; Long Zhang; Ming-Yong Zhang; Xiangnan Zhang; Xu Dong Zhang; Yan Zhang; Yang Zhang; Yanjin Zhang; Yingmei Zhang; Yunjiao Zhang; Mei Zhao; Wei-Li Zhao; Xiaonan Zhao; Yan G Zhao; Ying Zhao; Yongchao Zhao; Yu-Xia Zhao; Zhendong Zhao; Zhizhuang J Zhao; Dexian Zheng; Xi-Long Zheng; Xiaoxiang Zheng; Boris Zhivotovsky; Qing Zhong; Guang-Zhou Zhou; Guofei Zhou; Huiping Zhou; Shu-Feng Zhou; Xu-Jie Zhou; Hongxin Zhu; Hua Zhu; Wei-Guo Zhu; Wenhua Zhu; Xiao-Feng Zhu; Yuhua Zhu; Shi-Mei Zhuang; Xiaohong Zhuang; Elio Ziparo; Christos E Zois; Teresa Zoladek; Wei-Xing Zong; Antonio Zorzano; Susu M Zughaier Journal: Autophagy Date: 2016 Impact factor: 16.016
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7