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Objective:To investigate the role of mitofusion 2 (Mfn2) in high glucose (HG)-induced endoplasmic reticulum stress (ERS) and apoptosis of podocytes.Methods:(1) Streptozocin was used to induce a diabetes mellitus (DM) rat model. Renal histopathological changes in rats were observed by HE staining. Expression of Mfn2 and CCAAT/enhancer-binding protein homologous protein (CHOP) in glomeruli was observed by immunohistochemistry. Protein levels of Mfn2, protein kinase RNA-like ER kinase (PERK), phospho(p)-PERK, and CHOP in glomeruli were analyzed by Western blotting. (2) Conditionally immortalized human podocytes (HPC) cultured in vitro were divided into control, mannitol (MA) and HG groups. Expression of Mfn2 was observed by immunofluorescence. Protein levels of Mfn2, p-PERK, PERK and CHOP in HPC were analyzed by Western blotting. Podocyte apoptosis in each group was evaluated by flow cytometry with AnnexinⅤ-PE/7AAD double staining method. (3) HPC were divided into control, HG, HG+Mfn2-Myc plasmid-transfected and HG+control plasmid-transfected groups. Protein levels of Mfn2, p-PERK, PERK and CHOP in HPC were analyzed by Western blotting. Expression of CHOP was observed by immunofluorescence. Mitochondrial membrane potential in each group was observed by mitochondrial membrane potential assay kit with JC-1. Podocyte apoptosis in each group was evaluated by flow cytometry with AnnexinⅤ-PE/7AAD double staining method. Results:(1) Compared with the control group, the glomerular mesangial matrix of the DM group rats was significantly proliferated, and the expression of Mfn2 was down-regulated with the expression of ERS-related proteins p-PERK/PERK and CHOP up-regulated (all P<0.05). (2) Compared with the control group, Mfn2 was down-regulated and p-PERK/PERK and CHOP were up-regulated in HPC of HG group (all P<0.05). Apoptosis of HPC was also increased in HG group. There was no significant difference in the above indicators between the control group and the mannitol group (all P>0.05). (3) Compared with the HG group, mitochondrial membrane potential of HPC was alleviated and apoptosis of HPC was decreased in HG+Mfn2-Myc plasmid-transfected group ( P<0.05). P-PERK/PERK and CHOP were down-regulated in HG+Mfn2-Myc plasmid-transfected group (both P<0.05). There was no significant difference in the above indicators between the HG group and the HG+control plasmid-transfected group (all P>0.05). Conclusions:Mfn2 down-regulation in HG-stimulated podocytes may induce ERS to increase apoptosis of podocytes. Up-regulation of Mfn2 can alleviate the HG-induced ERS and apoptosis in podocytes.
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AimsStudies have indicated that chloroquine (CQ) shows antagonism against COVID-19 in vitro. However, evidence regarding its effects in patients is limited. This study aims to evaluate the efficacy of hydroxychloroquine (HCQ) in the treatment of patients with COVID-19. Main methodsFrom February 4 to February 28, 2020, 62 patients suffering from COVID-19 were diagnosed and admitted to Renmin Hospital of Wuhan University. All participants were randomized in a parallel-group trial, 31 patients were assigned to receive an additional 5-day HCQ (400 mg/d) treatment, Time to clinical recovery (TTCR), clinical characteristics, and radiological results were assessed at baseline and 5 days after treatment to evaluate the effect of HCQ. Key findingsFor the 62 COVID-19 patients, 46.8% (29 of 62) were male and 53.2% (33 of 62) were female, the mean age was 44.7 (15.3) years. No difference in the age and sex distribution between the control group and the HCQ group. But for TTCR, the body temperature recovery time and the cough remission time were significantly shortened in the HCQ treatment group. Besides, a larger proportion of patients with improved pneumonia in the HCQ treatment group (80.6%, 25 of 31) compared with the control group (54.8%, 17 of 31). Notably, all 4 patients progressed to severe illness that occurred in the control group. However, there were 2 patients with mild adverse reactions in the HCQ treatment group. Significance: Among patients with COVID-19, the use of HCQ could significantly shorten TTCR and promote the absorption of pneumonia. SignificanceAmong patients with COVID-19, the use of HCQ could significantly shorten TTCR and promote the absorption of pneumonia. Trial registrationURL: https://www.clinicaltrials.gov/. The unique identifier: ChiCTR2000029559.
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BackgroundWith the emergence of 4rd generation transmission, the prevention and treatment of the novel coronavirus disease 2019 (COVID-19) has entered a new period. We aimed to report several changes in the clinical characteristics at admission of patients with COVID-19. MethodsClinical records and laboratory results of patients suffering from COVID-19 were retrospectively reviewed and matched with the admission dates to analyze the changes in characteristics at the onset of illness. ResultsOf the 89 affected patients, 31 [34.8%] patients were admitted from January 16 to 22, and 58 [65.2%] were admitted from January 23 to 29. Patients were admitted with more systemic symptoms, such as fever (21 [67.7%] of 31), fatigue (13 [41.9%] of 31), and myalgia (7 [22.6%] of 31), before January 23. More patients (10 [32.3%] of 31) admitted before January 23 had a small amount of sputum production compared with a smaller proportion (4 [6.9%] of 58) of the patients admitted after January 23. Other symptoms, such as cough, nausea, diarrhea, and chest tightness, were not significantly different between the two groups. In addition, the group admitted before January 23 had a larger proportion of patients with reduced lymphocyte (13 [54.2%] of 24), CD3 (11 [54.4%] of 21), and CD8 (9 [42.9%] of 21) counts and elevated serum amyloid A (SAA, 18 [75%] of 24). ConclusionsThe initial symptoms of recently infected patients seem more insidious, indicating that the new coronavirus may gradually evolve into a virus similar to influenza and latent in asymptomatic carriers for a long time. SummaryCompared with the cases admitted earlier, more hidden initial symptoms and improved immune system disorders appeared in COVID-19 patients infected recently.
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Objective To investigate the role of autophagy in high glucose-induced podocyte lipid droplet metabolism.Methods (1) Cultured,conditionally immortalized human podocytes (HPC)were divided into normal control group,high glucose group and mannitol group.Oil red O staining and oil red O staining extraction assay was used to observe the degree of lipid accumulation;Protein level of SREBP-1 was analyzed by Western blotting.(2) HPC were cultured and divided into normal control group,high glucose group,high glucose+3-methyladenine (3-MA) group,and mannitol group.Acridine orange staining was used to observe the formation of autophagosomes.Western blotting was used to detect the protein levels of beclin-1 and LC3-Ⅱ/LC3-Ⅰ.Oil red O staining and oil red O staining extraction assay was used to observe the degree of lipid accumulation;Western blotting was used to analyze the expression of SREBP-1.Results (1) Compared with the normal control group,the lipid accumulation in the high glucose group was increased and the lipid metabolism related molecule SREBP-1 was up-regulated (P ＜ 0.05);There was no significant difference between the normal control group and the mannitol group in lipid accumulation (P ＞ 0.05).(2) Compared with the normal group,the number of autophagosomes was increased and autophagy-related proteins beclin-1 and LC3-Ⅱ/LC3-Ⅰ were up-regulated in high glucose group (all P ＜ 0.05).After intervened with 3-methyladenine,a significant decrease in autophagosomes was observed;Protein levels of autophagy-related proteinsbeclin-1 and LC3-Ⅱ/LC3-Ⅰ were decreased (all P ＜ 0.05);The lipid droplets in the high glucose+3-MA group was decreased and lipid metabolism related molecule SREBP-1 was down-regulated (all P ＜0.05).Conclusion Autophagy may be involved in the process of high-glucose-induced podocyte lipid accumulation by affecting SREBP-1 expression,and inhibition of autophagy can alleviate the high-glucose-induced podocyte lipid accumulation.
