Weihong Li1, Pengtao Li2, Ziwang Liu3, Qinghong Du4, Andre Steinmetz5, Ning Wang5, Huan Du6, Jinghong Hu4. 1. School of Preclinical Medicine, Beijing University of Chinese Medicine, 11 Bei San Huan Dong Lu, Chao Yang District, Beijing 100029, China. Electronic address: liweihong.403@163.com. 2. Dongzhimen Hospital Affiliated with Beijing University of Chinese Medicine, 5 Hai Yun Cang, Dongcheng District, Beijing 100700, China. 3. The Third Affiliated Hospital of Beijing University of Chinese Medicine, 51 An Wai Xiao Guan, Chao Yang District, Beijing 100029, China. 4. School of Preclinical Medicine, Beijing University of Chinese Medicine, 11 Bei San Huan Dong Lu, Chao Yang District, Beijing 100029, China. 5. Public Research Center for Health (CRP-Santé), 84 Val Fleuri, Luxembourg L-1526, Luxembourg. 6. Department of Pathology, Tianjin University of Traditional Chinese Medicine, 88 Yu Quan Lu, Nan Kai District, Tianjin 300193, China.
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE: Tong Luo Jiu Nao injection (TLJN), a Chinese medicine preparation, was extracted from the Chinese herbs Panax notoginseng and Gardenia jasminoides. Its pharmacological effect on cerebral ischemia was observed in the study. We previously reported that paracrine signaling of brain microvascular endothelial cells (BMECs) had a direct impact on the survival of neurons in mimicked cerebral ischemia in vitro. The current study was designed to investigate whether paracrine signaling of BMECs could be regulated by drug to achieve neuroprotection. MATERIALS AND METHODS: First, an in vitro model of cerebral ischemia in BMECs or neurons was established by oxygen-glucose-deprivation (OGD). TLJN was used as a medicine of intervention. Injured neurons were cultured in the conditioned media from normal and injured BMECs treated with TLJN. The changes in neurons, including the expression of N-methyl-D-aspartate receptor 1(NMDAR1), Ca(2+) concentration, cytochrome c release, the mitochondrial membrane potential, were determined by the immunofluorescence staining and molecular Probes. The content of Vascular endothelial growth factor (VEGF) and platelet activating factor (PAF) in various BMECs were also examined by Western blotting and Elisa. RESULTS: The results showed the activity of injured neurons was significantly increased when they were grown in conditioned media of normal or injured BMECs treated with TLJN, compared with that of normal or injured BMECs. These changes include a decrease of Ca(2+) concentration, of NMDAR1 expression, and of cytochrome c release, also an increase of the mitochondrial membrane potential. Moreover, the VEGF expression was up-regulated and the PAF expression was down-regulated by TLJN in BMECs. CONCLUSION: The results suggest that a neuroprotective effect of TLJN could be achieved by regulating paracrine signaling of BMECs, which could in part be explained by a TLJN-induced up-regulation of VEGF and a down-regulation of PAF in BMECs. Therefore, regulating the paracrine of BMECs could be the important target of the drug action on injured-neurons, which may be a novel path for therapeutic intervention in ischemic injury.
ETHNOPHARMACOLOGICAL RELEVANCE: Tong Luo Jiu Nao injection (TLJN), a Chinese medicine preparation, was extracted from the Chinese herbs Panax notoginseng and Gardenia jasminoides. Its pharmacological effect on cerebral ischemia was observed in the study. We previously reported that paracrine signaling of brain microvascular endothelial cells (BMECs) had a direct impact on the survival of neurons in mimicked cerebral ischemia in vitro. The current study was designed to investigate whether paracrine signaling of BMECs could be regulated by drug to achieve neuroprotection. MATERIALS AND METHODS: First, an in vitro model of cerebral ischemia in BMECs or neurons was established by oxygen-glucose-deprivation (OGD). TLJN was used as a medicine of intervention. Injured neurons were cultured in the conditioned media from normal and injured BMECs treated with TLJN. The changes in neurons, including the expression of N-methyl-D-aspartate receptor 1(NMDAR1), Ca(2+) concentration, cytochrome c release, the mitochondrial membrane potential, were determined by the immunofluorescence staining and molecular Probes. The content of Vascular endothelial growth factor (VEGF) and platelet activating factor (PAF) in various BMECs were also examined by Western blotting and Elisa. RESULTS: The results showed the activity of injured neurons was significantly increased when they were grown in conditioned media of normal or injured BMECs treated with TLJN, compared with that of normal or injured BMECs. These changes include a decrease of Ca(2+) concentration, of NMDAR1 expression, and of cytochrome c release, also an increase of the mitochondrial membrane potential. Moreover, the VEGF expression was up-regulated and the PAF expression was down-regulated by TLJN in BMECs. CONCLUSION: The results suggest that a neuroprotective effect of TLJN could be achieved by regulating paracrine signaling of BMECs, which could in part be explained by a TLJN-induced up-regulation of VEGF and a down-regulation of PAF in BMECs. Therefore, regulating the paracrine of BMECs could be the important target of the drug action on injured-neurons, which may be a novel path for therapeutic intervention in ischemic injury.
Keywords:
BMECs; Brain microvascular endothelial cells; I-CM; IT-CM; Mitochondria; N-CM; NT-CM; Neurons; OGD; Paracrine signaling; TLJN; Tong Luo Jiu Nao Injection; brain microvascular endothelial cells; conditioned medium from OGD-injured BMECs; conditioned medium from OGD-injured BMECs treated with Tong Luo Jiu Nao Injection; conditioned medium from normal BMECs; conditioned medium from normal BMECs treated with Tong Luo Jiu Nao Injection; oxygen–glucose-deprived