scholarly journals The protective effects of melatonin in high glucose environment by alleviating autophagy and apoptosis on primary cortical neurons

2021 ◽  
Author(s):  
Xiong Lijiao ◽  
Song Liu ◽  
Chaoming Liu ◽  
Tianting Guo ◽  
Zhihua Huang ◽  
...  
Keyword(s):  
Protein Expression ◽  
Protective Effect ◽  
Cell Viability ◽  
High Glucose ◽  
Cortical Neurons ◽  
Neuroprotective Effect ◽  
Control Group ◽  
Primary Cortical Neurons ◽  
Apoptosis Rate ◽  
Glucose Group

Abstract Cognitive dysfunction has been regarded as a complication of diabetes. Melatonin shows a neuroprotective effect on various neurological diseases. However, it’s protective effect on cortical neurons in high glucose environment has not been reported. Our present study aims to observe the protective effect of melatonin on rat cortical neurons and its relationship with autophagy in high glucose environment. The rat primary cortical neurons damaged model was induced by high glucose. The CCK-8, flow cytometry, Western Blot and immunofluorescence methods were used to examine the cell viability, apoptosis rate and proteins expression. Our results showed that there were no differences in cell viability, apoptosis rate, and protein expression among the control MLT and mannitol group. The cell viability of the glucose group was significantly lower than that of the control group, and the apoptosis rate of the glucose group was significantly higher than that of the control group. Compared with the glucose group, the glucose + melatonin group showed a significant increase in cell viability and a notable decrease in apoptosis rate. Melatonin concentration of 0.1-1 mmol/L can significantly reduce the injury of cortical neurons by high glucose. Compared with the control group, the glucose group showed a significant reduction of Bcl-2 protein expression, while remarkable elevations of Bax, caspase-3, Beclin-1 and LC3B levels. The neurons pre-administered with melatonin obtained significantly reversed these changes induced by high glucose. The phosphorylation levels of Akt and mTOR in the glucose group were significantly lower than those in the control group, were significantly increased in the glucose + MLT group compared with the glucose group. These data indicated that melatonin has a neuroprotective effect on cortical neurons under high glucose environment, which may work by activating Akt/mTOR pathway and following the down-regulation of autophagy.

scholarly journals Protective Effects of Oxymatrine on Vascular Endothelial Cells from High-Glucose-Induced Cytotoxicity by Inhibiting the Expression of A2B Receptor

2018 ◽  
Vol 45 (2) ◽  
pp. 558-571 ◽  
Author(s):  
Yun Yi ◽  
Yulin Shen ◽  
Qin Wu ◽  
Jingan Rao ◽  
Shu Guan ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cell Viability ◽  
Western Blotting ◽  
High Glucose ◽  
Vascular Endothelial Cells ◽  
Control Group ◽  
Vascular Endothelial ◽  
A2b Receptor ◽  
High Glucose Group ◽  
Glucose Group

Background/Aims: Diabetes mellitus (DM) has become an increasingly epidemic metabolic disease. Vascular endothelial cells play a key role in developing the cardiovascular complications of DM. The A2B receptor is expressed in vascular endothelial cells, and may help regulate the function of endothelial cells. The aim of this study was to investigate the protective effects of oxymatrine (OMT) on human umbilical vein endothelial cells (HUVECs) from high glucose-induced cytotoxicity. Methods: Homology modeling and molecular docking analysis were used to detect the binding sites between the adenosine A2B receptor and OMT. HUVECs were cultured with control (5.5 mM) or elevated glucose (22.2 mM) in the presence or absence of 3 µM OMT or A2B siRNA for 3 days. The MTS cell viability assay was used to measure the toxicity of high glucose on HUVECs and the protective effect of OMT or A2B siRNA. The expression of the adenosine A2B receptor and CCL5 in HUVECs was detected with real-time quantitative PCR (qPCR) and Western blotting methods in each group. Levels of IL-1β and TNF-α were measured using an enzyme-linked immunosorbent assay (ELISA) kit, and the concentration of NO was detected with the nitrate reductase method. Monocyte chemotactic activity in each group was detected using Transwell chambers. Furthermore, the phosphorylation of p38 and ERK1/2 in each group was observed through the Western blotting method. Results: Homology modeling and molecular docking analysis showed that OMT contains well-fitted binding sites to the A2B receptor. After chronic culture at high glucose, the rate of cell viability was significantly lower than that of the control group. After co-treatment with OMT or A2B siRNA, cell viability was significantly increased compared with the high-glucose group. The results from real-time quantitative RT-PCR (qRT-PCR) and Western blotting indicated that high glucose could increase the expression of A2B receptors in HUVECs, an effect that was inhibited by OMT. In addition, the results revealed that the expression of CCL5, IL-1β and TNF-α was increased in the high-glucose group, and that the NO produced by HUVECs decreased due to hyperglycemia; however, co-culture with OMT or A2B siRNA abolished these effects. Meanwhile, the chemotaxis activity of monocytes to HUVECs cultured in high-glucose medium was enhanced 2.59-fold compared to the control cells. However, the inflammatory reactions in HUVECs were completely relieved by co-treatment with OMT or A2B siRNA. Moreover, the phosphorylation of p38 and ERK1/2 in HUVECs in the high-glucose group was significantly higher than that of the control group; these effects were reversed after co-treatment with OMT or A2B siRNA. Conclusion: OMT may protect the HUVECs from high glucose-induced cytotoxicity through inhibitting the expression of A2B receptor and inflammatory factors as well as decreasing the phosphorylation of p38 and ERK1/2.

scholarly journals Vascular Endothelial Growth Factor Ameliorated Palmitate Induced Cardiomyocyte Injury via JNK Pathway

2020 ◽  
Author(s):  
Shiya Wang ◽  
Cao Zou ◽  
Xiaofeng Liu ◽  
Yonjin Yan ◽  
Shunzhon Gu ◽  
...  
Keyword(s):  
Protein Expression ◽  
Western Blot ◽  
Cell Viability ◽  
Mtt Assay ◽  
Caspase 3 ◽  
Time Dependent ◽  
Control Group ◽  
Apoptosis Rate ◽  
Dependent Relation ◽  
Quantitative Fluorescence

Abstract Objective To investigate the effect of palmitate (PAL) on apoptosis and the timing and activity of VEGF expression in HHHM2 myocardial cells (a human embryonic cardiomyocyte cell- line). Methods 1. Cardiomyocytes were divided into the following five groups: the control group and the 0.2 mM, 0.5 mM, 0.8 mM, and 1.2 mM PAL groups. We examined the changes in cell viability by MTT assay after PAL incubation for 24 h and the cardiomyocyte apoptosis rate by FACS examination, and thus determined the effective concentration of PAL. The transcription levels of CASP3, Bcl-2, Bax, and VEGF were detected by quantitative fluorescence PCR and the protein expression of caspase 3 and VEGF by western blot. 2. To observe the time-dependent effects on cell injury induced by 0.5 mM PAL, cardiomyocytes were divided into 0, 4, 8, 16, 24, and 48 h groups. The variation in cell viability was examined by MTT assay. The transcription levels of CASP3, Bcl-2, Bax, and VEGF were detected by quantitative fluorescence PCR and the protein expression of caspase 3 and VEGF by western blot. 3. To observe the effects of VEGF on the PAL induced apoptosis of cardiomyocytes, the cells were divided into the control group and the VEGF overexpression group. At 24 h after transfection, cells were incubated with 0.5 mM PAL for 6, 12, 24, and 48 h. Cell viability was examined by MTT assay. The apoptosis rate was measured by FACS using the Annexin V-FITC kit. The transcription levels of CASP3, Bcl-2, Bax, NF-kB p65, and VEGF were measured by quantitative fluorescence PCR, the protein expression of VEGF, caspase 3, Bcl-2, Bax, NF-κB p65, p-JNK/JNK, and p-ERK/ERK were measured by western blot, as well as caspase 3 activity. Results 1. A dose-dependent relation between the concentration of PAL and H9c2 cardiomyocyte injury was observed. In the 0.5 mM group, the apoptosis rate was increased significantly, while cell viability was decreased, indicating that 0.5 mM PAL was the ideal concentration to induce cardiomyocyte apoptosis. The expression of caspase 3 and Bax was significantly increased, and the expression of VEGF was enhanced, while the levels of Bcl-2 remained unchanged during the process. 2. A significant time-dependent relation between PAL and cardiomyocyte injury was observed. The apoptosis rate was increased greatly after 16 h treatment with 0.5 mM PAL. 3. Cell viability was restored by VEGF overexpression during treatment with 0.5 mM PAL. The apoptosis rate was also reduced by VEGF overexpression, as detected by FACS. The expression of caspase 3, Bax, and NF-κB p65 was significantly decreased, Bcl-2 and VEGF expression was dramatically increased, p-JNK/JNK expression was significantly enhanced, p-ERK/ERK levels did not exhibit a significant change, and the activity of caspase 3 was significantly decreased. Conclusions 1. PAL can induce injury and apoptosis in HHHM2 myocardial cells, and these effects are time-dependent. A PAL concentration of 0.5 mM was ideal to establish the cardiac cell injury model. 2. PAL at a concentration of 0.5 mM can effectively induce cardiomyocyte injury and enhance the expression of caspase 3, Bax, and VEGF, especially after 24 h and 48 h of PAL treatment. 3. VEGF overexpression can reverse the effects of PAL on apoptosis and cell viability. In addition, VEGF overexpression inhibited the expression of proapoptotic and inflammatory factors, caspase 3 activity, and transduction of the MAPK signaling pathway.

scholarly journals Effects of Qijin granules on high glucose-induced proliferation, apoptosis and expression of nuclear factor- κB and MCP-1 in rat glomerular mesangial cells

2021 ◽  
Vol 20 (9) ◽  
pp. 1819-1826
Author(s):  
Yuanfeng Yang ◽  
Gaocai Xiong ◽  
Renhui Yang ◽  
Yuchuan Li ◽  
Yuling Luo ◽  
...  
Keyword(s):  
Cell Cycle ◽  
High Glucose ◽  
Mesangial Cells ◽  
Control Group ◽  
Glomerular Mesangial Cells ◽  
Smad Signaling ◽  
Apoptosis Rate ◽  
Proliferation And Apoptosis ◽  
Protein Expressions ◽  
Tgf Β1

Purpose: To investigate the effects of Qijin granules on high glucose-induced proliferation and apoptosis in rat glomerular mesangial cells (MC).Methods: MC cells from rats were passaged and cultured, and randomly divided into control group (CNG), high glucose group (HGG), Western medicine group (WMG, high glucose + Benazepril + Gliquidone), and Qijin granules 1/2/3 group (high glucose + different doses of Qijin granules). Mesangial cells proliferation was measured using MTT assay. The NF-κB, MCP-1 and inflammatory factors in supernatant were determined by ELISA. Apoptosis rate and cell cycle were assessed by flow cytometry. The apoptosis-related TGF-β1/Smad signaling pathway-related protein expressions were measured by Western blot.Results: The A-value and early apoptosis rate, apoptosis rate and S-phase percentage, and protein expressions of NF-κB, MCP-1, IL-6, IL-2, TNF-ɑ, Bax, Cyt-C, caspase-3, TGF-β1, and p-Smad3 of MC cells in the HGG at 12 h, 24 h and 48 h were higher than those in the CNG. The above indices were lower in the WMG, and Qijin granules 1/2/3 groups than in the HGG. The Bcl-2, Smad7 protein expression level and the percentage of G1 and G2/M phase were lower in the HGG than in the CNG, and the above indeices were higher in the WMG and Qijin granules 1/2/3 group than in HGG.Conclusion: Qijin granules can dose-dependently inhibit high glucose-induced proliferation and apoptosis in rat MC cells, block the cell cycle and reduce inflammatory responses. This may be related to the regulation of NF-κB, MCP-1 and TGF-β1/Smad signaling pathways. These findings provide theoretical and experimental basis for the clinical treatment of early diabetic nephropathy.

scholarly journals Effect and Mechanism of 808 nm Light Pretreatment of Hypoxic Primary Neurons

2014 ◽  
Vol 2014 ◽  
pp. 1-5
Author(s):  
Xin Chen ◽  
Qiang-Li Wang ◽  
Shuang Li ◽  
Xian-qiang Mi
Keyword(s):  
Cell Viability ◽  
Cortical Neurons ◽  
Colorimetric Assay ◽  
Neuronal Morphology ◽  
Light Irradiation ◽  
Control Group ◽  
Primary Neurons ◽  
Atp Content ◽  
Low Intensity ◽  
Primary Mouse

This study investigated the effect of low intensity 808 nm light pretreatment of hypoxic primary neurons. Cobalt chloride (CoCl2) has been used to induce hypoxic injury in primary mouse cortical neurons. Low intensity 808 nm light was from light-emitting diode (LED). Cells were randomly divided into 4 groups: normal control group, CoCl2-induced group, CoCl2-induced group with 808 nm light irradiation pretreatment, and normal group with 808 nm light irradiation pretreatment. Effect of low intensity 808 nm light on neuronal morphology has been observed by microscope. MTT colorimetric assay has been used to detect the effect of low intensity 808 nm light on neuronal activity. Adenosine triphosphate (ATP) concentration and cytochrome C oxidase (COX) activity has been detected to study the effect of low intensity 808 nm light on neuronal mitochondria function. The results indicated that low intensity 808 nm light pretreatment alone did not affect cell viability, COX activity, and ATP content of neurons and low intensity 808 nm light pretreatment promoted the cell viability, COX activity, and ATP content of neurons with CoCl2exposure; however, low intensity 808 nm light pretreatment did not completely recover COX activity and cellular ATP content of primary neurons with CoCl2exposure to the level of the normal neurons.

scholarly journals Protective Effect of Green Perilla-Derived Chalcone Derivative DDC on Amyloid β Protein-Induced Neurotoxicity in Primary Cortical Neurons

2019 ◽  
Vol 42 (11) ◽  
pp. 1942-1946 ◽  
Author(s):  
Mami Iwasaki ◽  
Naotaka Izuo ◽  
Yasuhiko Izumi ◽  
Yuki Takada-Takatori ◽  
Akinori Akaike ◽  
...  
Keyword(s):  
Protective Effect ◽  
Cortical Neurons ◽  
Amyloid Β ◽  
Amyloid Β Protein ◽  
Primary Cortical Neurons ◽  
Chalcone Derivative ◽  
Β Protein

Neuroprotective effect of TAT PTD-Ngb fusion protein on primary cortical neurons against hypoxia-induced apoptosis

2013 ◽  
Vol 34 (10) ◽  
pp. 1771-1778 ◽  
Author(s):  
Guoyu Zhou ◽  
Peiyan Shan ◽  
Xueqiang Hu ◽  
Xueping Zheng ◽  
Shengnian Zhou
Keyword(s):  
Fusion Protein ◽  
Cortical Neurons ◽  
Neuroprotective Effect ◽  
Primary Cortical Neurons ◽  
Induced Apoptosis

scholarly journals Protective effect of estrogen on apoptosis in a cell culture model of Parkinson's disease

2008 ◽  
Vol 31 (5) ◽  
pp. 258 ◽  
Author(s):  
Xue-Li Li ◽  
Wei-Dong Cheng ◽  
Juan Li ◽  
Xian-Ling Guo ◽  
Cun-Ju Guo ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Cell Culture ◽  
Parkinson's Disease ◽  
Protective Effect ◽  
Cell Viability ◽  
Cell Model ◽  
Control Group ◽  
Beneficial Effects ◽  
Apoptotic Protein ◽  
Cell Groups

Objectives: The protective effect of estrogen on the neurons in Parkinson’s disease (PD) is unclear. The present study aimed to investigate the effect of estrogen on the apoptosis and dopaminergic function on a cultured cell model of PD. Methods: The PD model was established by addition of 1-methyl-4-phenylpyridinium (MPP+) to PC12 cell culture. Estrogen was added to cell groups with MPP+ (Estrogen+MPP+), and without MPP+ (Estrogen only group). Cell viability, content of tyrosine hydroxylase (TH), apoptosis ratio, expression of apoptosis-suppression protein Bcl-x and apoptosis-acceleration protein IL-1 beta converting enzyme (ICE) were measured. Results: Cell viability in the Estrogen+MPP+ group was similar to the control group but was higher than in the MPP+ group (P < 0.05). The apoptosis ratios in the Estrogen+MPP+ group (33.6%), and the control group (31.3%), were also similar, but it was lower than in the MPP+ group (63.5%, P < 0.05). Concentrations of Bcl-x were higher in the Estrogen+MPP+ group, whereas ICE concentrations were lower than in the MPP+ group (P < 0.05). Conclusions: Estrogen suppresses apoptosis and improves cell viability in MPP+ induced injuries in the PC12 cells. The beneficial effects of estrogen on the PD model are due to the suppression of pro-apoptotic protein ICE, and stimulation of anti-apoptotic protein Bcl-x.

scholarly journals Hypaconitine confers protection on ketamine-induced neuronal injury in neonatal rat brain via a mechanism involving PI3K/Akt/Bcl-2 pathway

2021 ◽  
Vol 18 (9) ◽  
pp. 1839-1844
Author(s):  
Junbao Liu ◽  
Yubo Hu ◽  
Linlin Li ◽  
Longyun Li
Keyword(s):  
Cell Viability ◽  
Caspase 3 ◽  
Neuroprotective Effect ◽  
Neuronal Cells ◽  
Specific Inhibitor ◽  
Neuronal Cell ◽  
Neuronal Injury ◽  
Neonatal Rat ◽  
Control Group ◽  
Neonatal Rats

Purpose: To investigate the neuroprotective effect of hypaconitine against ketamine-induced neuronal injury in the brains of neonatal rats, and the underlying mechanism of action. Methods: Seven day-old Sprague-Dawley pups weighing 15.0 to 20.0 g (mean weight = 17.5 ± 2.5 g), and aged 7 days were used for this study. The pups were sacrificed, and their forebrains isolated and used to prepare cell suspensions. The isolated cells were treated with ketamine (100 µM) or varied concentrations of hypaconitine (0.1 – 2 µM) or LY294002 (10 µM). The cells were trypsinized and cultured at 37 °C in 10 % fetal bovine serum (FBS) supplemented Dulbecco's modified Eagle's medium (DMEM) in a humidified incubator containing 5 % CO2. Cell viability was determined using MTT assay, while TUNEL assay was used to determine the extent of apoptosis in the cells. The expressions of pAkt, Bcl-2 and caspase-3 were determined using Western blotting. Results: There were only few viable cells in the ketamine-treated group, and cell viability was significantly and dose-dependently increased in hypaconitine-treated groups (p < 0.05). The extent of apoptosis was significantly higher in ketamine-treated cells than in control cells, but treatment with hypaconitine significantly reduced the number of apoptotic cells (p < 0.05). However, in the presence of LY294002 (a PI3K-specific inhibitor), the effect of hypaconitine on neuronal cell apoptosis was significantly reversed (p < 0.05). The expressions of p-Akt and Bcl-2 were significantly down-regulated while the expression of caspase-3 was significantly upregulated in ketamine-treated neuronal cells, when compared with control group (p < 0.05). However, in cells treated with hypaconitine, the expressions of p-Akt and Bcl-2 were significantly upregulated, while the expression of caspase-3 was significantly down-regulated (p < 0.05). Treatment of neuronal cells with hypaconitine in the presence of LY294002 significantly reversed the effect of hypaconitine on the expressions of p-Akt, Bcl-2 and caspase-3 (p < 0.05). Conclusion: These results suggest that hypaconitine ameliorates ketamine-induced neuronal injury in neonatal rats via a mechanism involving the PI3K/Akt/Bcl-2 pathway.

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