1.Protective effect of p38 mitogen-activated protein kinase inhibitors on zebrafish larvae brain after hypoxia/ reoxygenation injury
Jun CHEN ; Yusha HUANG ; Cheng ZENG ; Bin WANG
Chinese Journal of Applied Clinical Pediatrics 2017;32(2):135-140
Objective To examine the effects of p38 mitogen-activated protein kinase (MAPK) inhibitor on the behavioral response to zebrafish larvae after hypoxia/reoxygenation brain injury and to identify whether the protective effect is mediated by inhibiting apoptosis and protein,and mRNA related to apoptosis.Methods The 5-day post-fertilization zebrafish larvae were randomly assigned to 3 groups:control group,model group and intervention group.Fishes in the intervention group were separated into 3 subgroups according to p38 MAPK inhibitor concentration (5,10,20 μmol/L).The activity levels of the larvae were analyzed by using quantization mode of ZebraLab software,swimming distance and moving speed were recorded.Terminal transferase dUTP nick end labeling (TUNEL) assays of brain assays were performed.The protein levels of phosphorylation of p38 MAPK,apoptosis related proteins of B-cell lymphoma-2 (Bcl-2),Bcl-2 associated X protein(Bax) and Caspase-3 were determined by Western blot.The mRNA expressions of Bcl-2,Bax,and caspase-3 were also analyzed by reverse transcription-quantitative PCR (RT-qPCR).Results The activity movement analysis of the intervention group 2 and 3 demonstrated a significantly increase in the swimming distance compared with the model group(P < 0.05).After irradiation under strong light,all groups showed dramatically increasing in the moving speed.After removal of strong light,a significant decrease in moving speed was found in the control group and intervention group 2 and intervention group 3.The TUNEL assay showed that apoptosis index decreased in the intervention group (21.7 ±2.0,12.8 ± 1.9,17.7 ±2.6) compared with model group (46.8 ±5.3) (all P <0.01).Western blot assays demonstrated a significant increase protein level of phosphorylation of p38 MAPK after hypoxia and reoxygenation,and the inhibitor reduced the p-p38 MAPK expression.Compared with the model group,p38 MAPK inhibitor increased the protein and mRNA expression level of Bcl-2,whereas reduced the Bax and caspase-3 expression in the brain.Conclusions Under the influences of p38 MAPK inhibitor,zebrafish larvae improved the behavioral changes after hypoxia-induced brain injury.The inhibitor (10 μmol/L) optimally reduces hypoxia-induced apoptosis in brain by up-regulating Bcl-2,down-regulating Bax/caspase-3 protein and their mRNA level.
2.DAPT regulates Treg/Th17 cells immune balance to inhibit atherosclerosis
Jingning YANG ; Su ZHANG ; Yusha LUO ; Jun CHEN
Chinese Journal of Immunology 2017;33(3):343-346
Objective:To observe the effect of Notch signal inhibitor DAPT (γ-secretase inhibitor ) on the pathological changes of atherosclerosis mice and the immune balance of Treg/Th17.Methods:24 ApoE knockout C57BL mice were randomly divided into blank group,model group and DAPT group.The blank group were fed with normal diet ,the model group and the experimental group were fed with high fat diet.After 5 weeks of feeding,the mice in the experimental group were injected with DAPT [100 mg/(kg· d),re-suspended in DMSO],and the other two groups were injected with the equivalent amount of DMSO .After another 5 weeks,pathological changes of the mice in each group were analyzed by HE staining .ELISA was used to detect the level of IL-17 in plasma,and the propor-tion of splenic Treg/Th17 cells in each group was detected by flow cytometry .Results:HE staining results showed that the model group had obvious plaque formation and foam cell formation ,which showed that the AS model was successfully prepared .The degree of arterial disease in the DAPT group was significantly less than that in the model group .The plasma levels of IL-17 in the blank group , model group and DAPT group were(293.94±28.59),(454.05±172.68) and (335.40±89.57) pg/ml,respectively .The percentages of Treg cells in the blank group,model group and DAPT group were(3.80±0.56)%,(2.54±0.38)%and(4.73±0.64)%,respective-ly.The Th17 cell subsets of mice in the blank group , model group and DAPT group were ( 3.46 ±0.23 )%, ( 4.52 ±0.85 )% and (1.38±0.37)%,respectively .Conclusion:DAPT decreased the plasma level of IL-17 in AS mice,inhibited the differentiation of Th17 cell subsets,and promoted the differentiation of Treg ,and reduced the atherosclerosis by changing the Treg/Th17 cells immune balance.
3.Effect of Taurine on the expression of glucose regulated protein 78 and growth arrest and DNA damage - in-ducible protein 34 in zebrafish larvae brain after hypoxia/reoxygenation brain injury
Yusha HUANG ; Yan CHENG ; Xiaohui CHEN ; Jixuan LUO ; Cheng ZENG ; Bin WANG
Chinese Journal of Applied Clinical Pediatrics 2016;31(6):457-461
Objective To investigate the expression of endoplasmic reticulum stress - related factors, glucose - regulated protein 78(GRP78)and growth arrest and DNA damage - inducible protein 34( GADD34)and neuronal apoptosis in the brain of zebrafish larvae after hypoxia/ reoxygenation brain injury,and the neuroprotective role of Taurine. Methods The 5 day post - fertilization zebrafish larvae were randomly assigned to 3 groups:control group, hypoxia/ reoxygenation model group(model group)and Taurine treatment group(Taurine group). According to the dif-ferent time points for observation,each group was subdivided into 5 subgroups(1 h,3 h,6 h,24 h,48 h)with 100 ze-brafish larvae each. The pathological changes in the brain tissues and cell apoptosis were detected by fluorescence ter-minal deoxynucleotidyltransferase - mediated dUTP nick end - labeling( TUNEL). The expression of GRP78 and GADD34 mRNA in the brain of zebrafish larvae were detected by real - time quantitative reverse transcription PCR (qRT - PCR). The changes in GRP78 and GADD34 protein were detected by Western blot. Results (1)TUNEL:apoptosis index(AI)was increased after hypoxia/ reoxygenation,and reached the peak at 3 h in model group,the AI in Taurine group was decreased compared with that in the model group at the same time point(1 h:22. 83 ± 1. 80 vs 30. 18 ± 1. 81,3 h:23. 22 ± 2. 46 vs 42. 97 ± 4. 01,6 h:16. 80 ± 1. 69 vs 22. 97 ± 1. 91,all P ﹤ 0. 05).(2)qRT -PCR:the expression of GRP78 and GADD34 mRNA was increased at 1 h after hypoxia/ reoxygenation,and reached the peak at 3 h in the model group,the expression in Taurine group was decreased compared with that in the model group at the same time point(GRP78 mRNA:1 h:2. 35 ± 0. 13 vs 5. 36 ± 0. 35,3 h:3. 08 ± 0. 33 vs 4. 27 ± 0. 52,6 h:1. 57 ± 0. 12 vs 3. 00 ± 0. 13,all P ﹤ 0. 05;GADD34 mRNA:1 h:5. 14 ± 0. 55 vs 7. 45 ± 0. 67,3 h:2. 79 ± 0. 58 vs 5. 83 ± 0. 51,6 h:1. 79 ± 0. 22 vs 3. 67 ± 0. 30,all P ﹤ 0. 05).(3)Western blot:the expression of GRP78 and GADD34 pro-tein was increased at 1 h,reached the peak at 6 h,but it was decreased in Taurine group(GRP78 protein:1 h:1. 12 ± 0. 11 vs 1. 37 ± 0. 13,3 h:0. 79 ± 0. 11 vs 1. 25 ± 0. 10,6 h:0. 55 ± 0. 10 vs 1. 52 ± 0. 14,all P ﹤ 0. 05;GADD34 pro-tein:1 h:0. 92 ± 0. 11 vs 1. 11 ± 0. 13,3 h:0. 96 ± 0. 11 vs 1. 52 ± 0. 09,6 h:0. 76 ± 0. 05 vs 1. 89 ± 0. 06,all P ﹤0. 05).(4)The expression of GRP78 and GADD34 protein was positively correlated with AI in the model group(r =0. 53,0. 56 respectively,all P ﹤ 0. 05). Conclusion One of neuroprotective mechanisms of Taurine against hypoxia/reoxygenation brain injury may down - regulate GRP78 and GADD34 expression.
4.Aviation flight and neurological diseases
Yusha TANG ; Tingting SONG ; Lei CHEN
Journal of Chinese Physician 2022;24(4):630-633
In the air flight, individuals rise rapidly to the environment of low-pressure hypoxia, low humidity and high noise. If we can't adapt to the change of the environment, we may suffer headache, stroke, epilepsy and other common nervous system diseases. With the increasing popularity of aviation, more and more in-flight medical emergencies are reported. Aviation medical is gradually concerned and studied by social people, medical internists and scientific research workers. This article will summarize the clinical characteristics and pathogenesis of common paroxysmal nervous system diseases in flight, and discuss the prospect of early warning and prevention of in-flight medical emergencies.
5.Functional Magnetic Resonance Imaging Study of Oxcarbazepine Effect on Auditory Cortex
Rui LI ; Lei CHEN ; Yusha TANG
Journal of Medical Research 2023;52(12):32-36
Objective Functional magnetic resonance imaging was used to explore the effects of oxcarbazepine on auditory cortex function in patients with epilepsy.Methods 21 patients with epilepsy treated with oxcarbazepine and twenty-one patients with epilepsy who matched age and gender but did not treat with oxcarbazepine were collected.Amplitude of low frequency fluctuation(ALFF)and re-gional homogeneity(ReHo)were used to compare the differences in the functional activity of the auditory cortex between the two groups.Results Compared with the patients without oxcarbazepine,the patients with oxcarbazepine had higher ALFF values in the right superior temporal gyrus,bilateral middle temporal gyrus and visual union cortex,and lower ALFF values in the bilateral orbitofrontal area and infe-rior temporal gyrus,the patients with oxcarbazepine had higher ReHo values in right superior temporal gyrus,bilateral middle temporal gy-rus,orbitofrontal area,and left visual union cortex,and lower ReHo values in bilateral superior temporal gyrus,inferior temporal gyrus and right middle temporal gyrus.In the same cross-sectional area,ALFF and ReHo values were enhanced in the right superior temporal gyrus,right cingulate gyrus,right somatosensory cortex and left visual union cortex,while ALFF and ReHo values were weakened in the bilateral inferior temporal gyrus.However,there was no significant difference in ALFF and ReHo values of bilateral transverse temporal gyrus and transverse temporal sulcus between the two groups.Conclusion Oxcarbazepine may affect auditory cortex function in patients with epilepsy,and it mainly acts on the secondary auditory cortex or regulates the excitatory balance of the auditory network.
6.Effect of docosahexaenoic acid on long-term learning and memory disorders after hypoxic ischemic brain damage in rats
Cheng ZENG ; Siyun SHU ; Yusha HUANG ; Yan CHENG ; Jun CHEN ; Bin WANG
Chinese Journal of Neuromedicine 2016;15(7):678-684
Objective To explore the effect of docosahexaenoic acid (DHA) on long-term learning and memory disorders and potential mechanism in rats after hypoxic ischemic brain damage. Methods Sixty neonatal 7-day-old SD rats were ramdonly divided into three groups: group S (sham operation+vehicle treatment), group C (hypoxic-ischemic brain damage [HIBD]+vehicle treatment) and group D (HIBD+DHA treatment). After left common carotid artery was isolated and ligated for 2.5 h, rats of group C and group D were put into a condition which oxygen concentration was about 8%for 2 h;rats in the group S were only isolated the left carotid artery, without ligation or hypoxia treatment;rats in the group D were intraperitoneally injected DHA of 15 mg/kg after modeling, and rats in the group S and group C were intraperitoneally injected equivalent volume of vehcle, once a day for 10 consecutive days. The pathomorphology changes of the hypocampal CA1 area, and marginal division of striatum were observed by Nissl staining 48 h after modling; the apoptosis cells were measured by TUNEL;immunohistochemical method was used to detect the expressions of Bax and Caspase-3 positive cells in the two brain areas. Morris water maza test was used to evaluate the long-term lerning and momory functions of 2-month-old rats, and the expressions of N-methyl-D-aspartate receptor 1 (NMDAR1) positive cells were detected by immunohistochemical method. Results The pathomorphology damage was significantly improved, the expressions of Bax and Caspase-3 positive cells and the neuron apoptosis in hypocampal CA1 areas and marginal division of striatum in group D were all signficantly decreased as compared with those in the group C (P<0.05). Rats in group D had significantly decreased escape latency as compared with those in group C in Morris water maze test (P<0.05), and the expression of NMDAR1 positive cells in the two brain areas of group D was significantly increased as compared with that in the group C (P<0.05). Conclusion DHA has the ameliorative effect on long-term learning and memory disorders after hypoxic ischemic brain damage in rats, which may be associated with inhibitory action of cell apoptosis at early phase and up-regulation of expression of NMDA1 at the late phase.
7.A case of classical Vohwinkel syndrome caused by the mutation p.N54H in the GJB2 gene
Deyu SONG ; Jiayue WANG ; Jia GENG ; Meirong ZOU ; Zhongtao LI ; Yusha CHEN ; Sheng WANG
Chinese Journal of Dermatology 2023;56(7):669-672
Objective:To detect gene mutations in 1 patient with Vohwinkel syndrome who presented with palmoplantar keratoderma, pseudo-ainhum and deafness.Methods:Clinical data were collected from the proband, and a genetic test was performed to identify mutation sites.Results:Clinical manifestations of the proband were consistent with classical Vohwinkel syndrome. The genetic test revealed a heterozygous mutation c.160A>C (p.N54H) in the GJB2 gene, which was not detected in her parents or healthy controls.Conclusion:The heterozygous mutation c.160A>C (p.N54H) in the GJB2 gene was first identified in a patient with classical Vohwinkel syndrome, and there were overlaps in mutation sites between classical Vohwinkel syndrome and palmoplantar keratoderma with deafness.
8.Isoflavones' effects on pharmacokinetic profiles of main iridoids from Gardeniae Fructus in rats
Ruirui CHANG ; Jialin LIU ; Yusha LUO ; Taohong HUANG ; Qiang LI ; Jun WEN ; Weidong CHEN ; Tingting ZHOU
Journal of Pharmaceutical Analysis 2020;10(6):571-580
Gardeniae Fructus (GF) and Semen Sojae Praeparatum (SSP) are both medicine food homologies and widely used in Chinese clinical prescriptions together. The research investigated the pharmacokinetics of four iridoids in normal rats and isolfavones-fed rats, which were administered with isolfavones from SSP for 7, 14, 21 and 28 consecutive days. A validated LC-MS/MS method was developed for determining shanzhiside, genipin-1-gentiobioside, geniposide and their metabolite genipin in rat plasma. Plasma samples were pretreated by solid-phase extraction using paeoniflorin as the internal standard. The chromatographic separation was performed on a Waters Atlantis T3 (4.6 mm × 150 mm, 3μm) column using a gradient mobile phase consisting of acetonitril and water (containing 0.06%acetic acid). The mass detection was under the multiple reaction monitoring (MRM) mode via polarity switching between negative and positive ionization modes. The calibration curves exhibited good linearity (r>0.997) for all components. The lower limit of quantitation was in the range of 1-10 ng/mL. The intra-day and inter-day precisions (RSD) at three different levels were both less than 12.2% and the accuracies (RE) ranged from -10.1% to 16.4%. The extraction recovery of them ranged from 53.8% to 99.7%. Pharmacokinetic results indicated the bioavailability of three iridoid glycosides and the metabolite, genipin in normal rats was higher than that in rats exposed to isoflavones. With the longer time of administration of iso-flavones, plasma concentrations of iridoids decreased, while genipin sulfate, the phase II metabolite of genposide and genipin-1-gentiobioside, appeared the rising exposure. The pharmacokinetic profiles of main iridoids from GF were altered by isoflavones.