1.Inhibitory effect of melatonin on pyroptosis of lens epithelium cells and its mechanism
Xin YANG ; Xuhui LIU ; Jia MENG ; Mengyuan FANG ; Fengyan ZHANG
Chinese Journal of Experimental Ophthalmology 2022;40(5):414-421
Objective:To investigate the inhibitory effect of melatonin on pyroptosis of lens epithelium cells (HLECs) induced by hydrogen peroxide (H 2O 2) and its mechanism. Methods:The cultured HLECs were divided into normal control group, model control group, melatonin group, vitamin E group, and vitamin E solvent group.Cells in melatonin group, vitamin E group and vitamin E solvent group were pre-cultured with 1×10 -6 mol/L melatonin, 100 μmol/L vitamin E or equal volume of vitamin E solvent, then cultured with 100 μmol/L H 2O 2, respectively, and the cells in the normal control group and model control group were cultured with normal condition or 100 μmol/L H 2O 2, respectively.The HLECs transfected with nuclear factor erythroid 2-related factor 2 short hairpin RNA (shNrf2) or shNrf2 negtive control lentivirus and following with 100 μmol/L H 2O 2 treatment were served as shNrf2 group and shNrf2 negative control group, respectively; and the transfected cells treated with 1×10 -6 mol/L melatonin and subsequent 100 μmol/L H 2O 2 treatment were served as melatonin+ shNrf2 group and melatonin+ shNrf2 negative control group.The activity of lactic dehydrogenase (LDH) and the expression levels of interleukin (IL)-1β and IL-18 in the culture supernatant were detected by the enzyme linked immunosorbent assay (ELISA) kit.The concentration of reactive oxygen species (ROS) was assessed by flow cytometry.The expression level of Nrf2, NLRP3, apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC), caspase-1 p20 and GSDMD-N proteins were evaluated by Western blot. Results:Compared with model control group, the activity of LDH and the concentrations of IL-1β and IL-18 were significantly decreased in melatonin group and vitamin E group, showing statistically significant differences (all at P<0.05). The ROS fluorescence intensities were 13 040.67±1 550.66 and 12 593.67±1 677.06 in melatonin group and vitamin E group, respectively, which were significantly lower than 18 310.33±1 248.01 in model control group (both at P<0.05). The relative expression levels of Nrf2 protein were 4.24±0.44 and 3.73±0.38 in melatonin group and vitamin E group, respectively, which were significantly higher than 2.28±0.34 in model control group, and the relative expression levels of NLRP3, ASC, caspase-1 p20 and GSDMD-N in melatonin group and vitamin E group were significantly decreased than model control group (all at P<0.05). The relative expression level of Nrf2 protein in shNrf2 group and melatonin+ shNrf2 group was significantly reduced, and the expression levels of LDH, IL-1β, IL-18, ROS content, NLRP3, ASC, caspase-1 p20 and GSDMD-N were significantly increased in comparison with shNrf2 negative control group and melatonin+ shNrf2 negative control group, respectively (all at P<0.05). Conclusions:Melatonin can inhibit the release of NLRP3 inflammasome by activating Nrf2, and has an inhibitory effect on the pyroptosis of HLECs.
2.Progress of reversible parkinsonism
Ting PU ; Tao CHEN ; Mengyuan XU ; Xu MENG ; Jia TANG ; Xiaoguang LEI
Chinese Journal of Neurology 2022;55(7):775-782
Parkinsonism is a clinical syndrome caused by many reasons, mainly manifested as bradykinesia, stiffness, static tremor and postural instability. Common disease development patterns include occult onset, gradual development, and little natural remission. However, clinically there are some Parkinsonism that will improve, naturally alleviate or "cure", called reversible parkinsonism (RP). By searching the relevant literature, RP was classified into 12 different types: drugs induced, poisoning induced, infection induced, intracranial vascular induced, structural encephalopathy related, changes in intracranial pressure related, imbalance of internal environment induced, visceral diseases related, alcohol withdrawal related, surgery related, immunization and radiotherapy induced RP. This article aims to provide clinicians with more ideas for the clinical diagnosis and treatment of parkinsonism, so as to promote clinicians to make reasonable identification and diagnosis and treatment of parkinsonism as soon as possible.
3. Effects of microRNA-34a on regulating silent information regulator 1 and influence of the factor on myocardial damage of rats with severe burns at early stage
Xiaozhi BAI ; Ting HE ; Julei ZHANG ; Yang LIU ; Mengyuan CAO ; Jianing ZHANG ; Weixia CAI ; Yanhui JIA ; Jihong SHI ; Linlin SU ; Dahai HU
Chinese Journal of Burns 2018;34(1):21-28
Objective:
To explore the effects of microRNA-34a on regulating silent information regulator 1 (SIRT1) and influence of SIRT1 on myocardial damage of rats with severe burns at early stage.
Methods:
(1) Twenty-four Sprague-Dawley (SD) rats were divided into sham injury (SI) group, simple burns (SB) group and SIRT1 agonist (SA) group according to the random number table (the same grouping method below), with 8 rats in each group. Rats in groups SB and SA were inflicted with 30% total body surface area full-thickness scald (hereinafter referred to as burns) on the back, and rats in group SI were sham injuried on the back. Immediately after injury, rats in groups SI and SB were intraperitoneally injected with normal saline of 50 mL/kg, and rats in group SA were intraperitoneally injected with normal saline of 50 mL/kg and 1 mg/mL resveratrol of 50 mg/kg. At 6 h post injury, abdominal aortic blood was collected to make serum and myocardial tissue of rats was collected. (2) Myocardial cells of twelve neonatal SD rats were collected and divided into microRNA-34a mimic control (MMC) group, microRNA-34a mimic (MM) group, microRNA-34a inhibitor control (MIC) group, and microRNA-34a inhibitor (MI) group, which were respectively transfected with gene sequences of mimic control, mimic, inhibitor control, and inhibitor of microRNA-34a. The microRNA-34a expression level and protein expression level of SIRT1 in myocardial cells were respectively detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blotting. Another batch of myocardial cells were divided into microRNA-34a inhibitor control+ burn serum (MCB) group, microRNA-34a inhibitor+ burn serum (MB) group, and microRNA-34a inhibitor+ burn serum + EX527 (MBE) group. Myocardial cells in group MCB were transfected with gene sequence of inhibitor control, and myocardial cells in the later groups were transfected with gene sequence of inhibitor of microRNA-34a. After transfection of 48 h, myocardial cells in group MBE were cultured in Dulbecco′s modified Eagle′s medium (DMEM) solution for 6 hours, with serum in group SB of volume fraction of 10% and final amount-of-substance concentration of 1 mol/L, and myocardial cells in the other 2 groups were cultured in DMEM solution with serum from rats of group SB of volume fraction of 10%. The protein expression levels of myocardial cells of SIRT1, cleaved-caspase-3, and Bax were detected by Western blotting. (3) Myocardial tissue from (1) was collected to detect expression levels of microRNA-34a and mRNA of SIRT1 in groups SI and SB by real-time fluorescence quantitative RT-PCR. Morphology of myocardial tissue of rats in groups SI, SB, and SA was observed with biological image navigator. The mRNA expression levels of interleukin 1β (IL-1β) and tumor necrosis factor (TNF-α) of rats in groups SI, SB, and SA were detected by real-time fluorescence quantitative RT-PCR. The expression levels of cleaved-caspase-3, and Bax of myocardial tissue of rats in groups SI, SB, and SA were detected by Western blotting. Data were processed with one-way analysis of variance and least-significant difference test.
Results:
(1) After transfection of 48 h, the expression level of microRNA-34a of myocardial cells in group MM was 4.67±0.92, significantly higher than 1.03±0.04 in group MMC (
4.Effect of Pangshi Antai Zhixue Decoction on Th1/Th2 Balance in Decidua and Membrane of Spontaneous Abortion Rats with Heat Syndrome by Regulating p38 MAPK Signal Pathway
Liya MA ; Yuanyuan CAI ; Xuelin ZHANG ; Yitong YANG ; Mengyuan JIA ; Danya WANG ; Jingyi WANG ; Dawei ZHANG
Chinese Journal of Experimental Traditional Medical Formulae 2022;28(18):62-69
ObjectiveTo explore the anti-abortional effect of Pangshi Antai Zhixue decoction and its mechanism in helper T lymphocyte 1 (Th1)/Th2 balance in the decidual tissues of spontaneous abortion rats with heat syndrome, based on the p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway. MethodAconiti Lateralis Radix Praeparata, Zingiberis Rhizoma, and Cinnamomi Cortex decoction was used to replicate the rat model of spontaneous abortion with heat syndrome. The spontaneous abortion rats with heat syndrome were randomly divided into model group, aspirin group (5.25 mg·kg-1), dydrogesterone group (3.02 mg·kg-1), Pangshi Antai Zhixue decoction high-dose (44 g·kg-1), medium-dose (22 g·kg-1), and low-dose (11 g·kg-1) groups, with ten rats in each group. Ten normal rats were divided into a normal group. Rats in each group were given corresponding drugs, Once a day for 12 d. After 24 h of the last administration, blood was collected from the abdominal aorta. The enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of β-human chorionic gonadotropin (β-HCG), progesterone (P), estradiol (E2), γ interferon (IFN-γ), and interleukin-4 (IL-4) in rat serum. The uterus and meconium tissues of rats were collected to determine the number and rate of miscarriages. Western blot was used to detect GATA3, T-bet, p38 MAPK, and its phosphorylation in the decidual tissue. ResultAs compared with the normal group, the number of live births, the β-HCG, P, E2, and IL-4 in the serum, and the GATA3 protein expression in the decidual tissue in the model group were reduced (P<0.01), whereas the number and rate of miscarriages, IFN-γ in the serum, and the expression of p-p38 MAPK and T-bet protein levels in the demolded tissues increased (P<0.01). As compared with the model group, the number of live births, the β-HCG, P, E2, and IL-4 in the serum, and the GATA3 protein expression in the decidual tissue in the Pangshi Antai Zhixue decoction medium-dose group increased (P<0.01), whereas the number and rate of miscarriages, IFN-γ in the serum, and the expression of p-p38 and T-bet protein levels in the demolded tissues reduced (P<0.01). As compared with the aspirin group, the P, E2, and IL-4 in the serum of rats in the dydrogesterone group and the Pangshi Antai Zhixue decoction high-dose and medium-dose groups increased (P<0.01), the number of live births in the Pangshi Antai Zhixue decoction medium-dose group increased (P<0.01), and the β-HCG and IFN-γ in the serum of rats in the dydrogesterone group decreased (P<0.01). The number and rate of miscarriages, IFN-γ in the serum, and T-bet and GATA3 levels in the decidual tissues of rats in the Pangshi Antai Zhixue decoction medium-dose group decreased (P<0.05). Compared with the Pangshi Antai Zhixue decoction medium-dose group, the low-dose group, high-dose group, and dydrogesterone group showed increased number and rate of miscarriages (P<0.05), and the high-dose group and dydrogesterone group decreased the number of live birth (P<0.01). The IFN-γ in the serum and p-p38 MAPK and T-bet protein in the decidual tissue in the low-dose group, and the p-p38 MAPK and T-bet protein in the decidual tissue in the high-dose group all increased (P<0.05). The β-HCG, P, and E2 in the serum of rats in the Pangshi Antai Zhixue decoction low-dose group, dydrogesterone group, and aspirin group decreased (P<0.01), and the IL-4 in the serum and GATA3 in the decidual tissue of rats in the Pangshi Antai Zhixue decoction low-dose and high-dose group and the dydrogesterone group decreased (P<0.01). ConclusionPangshi Antai Zhixue decoction realizes the effect of fetal protection by regulating the activation of p38 MAPK signal pathways and Th1/Th2 balance.