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Objective To observe the effects of total saponins of Trillium tschonoskii Maxim(TST)on vascular cognitive impairment(VCI),neurovascular units(NVUs),and neural circuit integrity in rats.Methods Forty-eight male Sprague-Dawley(SD)rats were randomly divided into sham-operated group,model group,TST group(intragastric administration,100 mg/kg),and donepezil group(intragastric administration,0.45 mg/kg),and then subjected to ischemic stroke by 2-VO method(bilateral common carotid artery ligation)or sham surgery.After 28 days of intragastric administration,Mirros water maze test was performed to evaluate the spatial learning and memory abilities of rats in each group.HE and Nissl staining were used to observe the pathological changes of brain tissue in rats.The expression of synuclein(SYN)in rat hippocampus was observed by immunohistochemical staining.Changes in dendritic spines in rat's hippocampal neurons were observed by Golgi staining.Western blotting was used to detect the expression levels of IL-1β,IL-10,vascular endothelial growth factor A(VEGFA),postsynaptic density protein 95(PSD95),and growth associated protein 43(GAP43)in rat's hippocampus in each group.Results In Mirros water maze test,rats in model group showed significant prolonged escape latency(P<0.05),and a significant reduction in the number of crossing platforms and the percentage of activity time in the target quadrant(P<0.05)than those in sham-operated group;while rats in TST group and donepezil group showed significant shortened escape latency(P<0.01),and significant increase of the number of times of crossing platforms and the percentage of activity time in the target quadrant(P<0.05)than those in model group.Compared of sham-operated group,model group showed a decrease in the expression of SYN and the number of neurons,Nissl bodies,and dendritic spines in the CA1 region of the hippocampus(P<0.05).Compared with model group,TST group and donepezil group showed an increase in the expression of SYN and the number of neurons,Nissl bodies,and dendritic spines in the CA1 region of the hippocampus(P<0.05).Western blotting showed a significant increase in the expression of IL-1β and VEGF(P<0.05),and a decrease in the expression of IL-10,PSD95,and GAP43(P<0.01)in rat's hippocampus of model group than those in sham-operated group.Compared with model group,TST group and donepezil group showed a significant decrease in the expression of IL-1β(P<0.05),and an increase in the expression of VEGFA,IL-10,and GAP43(P<0.05).Conclusions TST could alleviate cognitive impairment through promoting synaptic plasticity and neurovascular unit remodeling in 2-VO model rats,suggesting its significance as a potential drug for apoplexy.
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Aim To investigate the effects of total saponins from Trillium tschonoskii maxim(TST)on cognitive impairment and mitochondrial autophagy in aging rats induced by D-galactose(D-gal). Methods Male SD rats were randomly divided into normal control group,model group(D-gal,subcutaneous injection),intervention group(TST,low,medium and high dose groups by intragastric administration),with 10 rats in each group,and administered for 6 weeks. Morris water maze was used to evaluate the cognitive function. HE and Nissl staining were used to test the hippocampal and brain cortex morphology. Immunohistochemistry staining was applied to detect the localization expression of Pink1 and Parkin. Western blot was employed to detect the expressions of Pink1,Parkin,LC3-Ⅱ,p62 and Beclin1. Results Compared with the normal control group,the escape latency time was prolonged and the number of crossing platform decreased in D-gal model group(P<0.05). The number of neurons in hippocampus significantly decreased. The positive cells labeled by Pink1 and Parkin staining in hippocampus significantly decreased. The expressions of Pink1,Parkin,LC3-Ⅱ and Beclin1 were markedly reduced,while the expression of p62 was significantly raised(P<0.05). Compared with D-gal model group,the escape latency time of TST dose groups was shortened,the Times of crossing the platform was more,and the time of staying in the platform quadrant increased(P<0.05). The number of neurons in hippocampus significantly increased. The positive cells labeled by Pink1 and Parkin staining in hippocampus significantly increased. The expressions of Pink1,Parkin,LC3-Ⅱ and Beclin1 in hippocampus were apparently up-regulated,while the protein expression of p62 was evidently down-regulated(P<0.05). Conclusions TST has neuroprotective effects on the learning and memory capacities in aging rats induced by D-gal,which may be related to the increasing levels of Pink1,Parkin,LC3-Ⅱ and Beclin1 proteins and the activation of mitochondrial autophagy.
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Aim To study the therapeutic effect of Balanophora polysaccharide(BPS)on gastric ulcer(GU)induced by acetic acid in rats and to investigateits mechanisms. Methods Sixty male SD rats were randomly divided into sham-operated group, GU model group, omeprazole positive group(3.6 mg·kg-1), and low, medium and high dose of BPS treatment groups(100, 200 and 400 mg·kg-1). The GU model group was prepared by acetic acid cautery method, and the morphology and pathological changes of ulcers were observed by visual observation combined with HE staining, and the ulcer area and inhibition rate were measured and calculated; superoxide dismutase(SOD)activity, malondialdehyde(MDA)content and glutathione peroxidase(GSH-PX)activity were measured by enzymatic assay; tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)content were detected by ELISA. The expression levels of epidermal growth factor(EGF)and epidermal growth factor receptor(EGFR)were measured by immunohistochemistry staining and Western blot. Results Compared with the sham-operated group, obvious ulcer damage was seen in the model group. Compared with the model group, the BPS-treated group showed a significant reduction in ulcer area, an increase in SOD and GSH-PX activity and EGF and EGFR expression levels, and a significant decrease in MDA, TNF-α and IL-6 content. Conclusions BPS has a therapeutic effect on GU in rats, and its mechanism may be related to the inhibition of oxidative stress, suppression of inflammatory stimuli and promotion of regenerative repair of gastric mucosa.
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MicroRNAs (miRNAs) are mediators of the aging process. The purpose of this work was to analyze the miRNA expression profiles of spermatozoa from men of different ages with normal fertility. Twenty-seven donors were divided into three groups by age (Group A, n = 8, age: 20-30 years; Group B, n = 10, age: 31-40 years; and Group C, n = 9, age: 41-55 years) for high-throughput sequencing analysis. Samples from 65 individuals (22, 22, and 21 in Groups A, B, and C, respectively) were used for validation by quantitative real-time polymerase chain reaction (qRT-PCR). A total of 2160 miRNAs were detected: 1223 were known, 937 were newly discovered and unnamed, of which 191 were expressed in all donors. A total of 7, 5, and 17 differentially expressed microRNAs (DEMs) were found in Group A vs B, Group B vs C, and Group A vs C comparisons, respectively. Twenty-two miRNAs were statistically correlated with age. Twelve miRNAs were identified as age-associated miRNAs, including hsa-miR-127-3p, mmu-miR-5100_L+2R-1, efu-miR-9226_L-2_1ss22GA, cgr-miR-1260_L+1, hsa-miR-652-3p_R+1, pal-miR-9993a-3p_L+2R-1, hsa-miR-7977_1ss6AG, hsa-miR-106b-3p_R-1, hsa-miR-186-5p, PC-3p-59611_111, hsa-miR-93-3p_R+1, and aeca-mir-8986a-p5_1ss1GA. There were 9165 target genes of age-associated miRNAs. Gene Ontology (GO) analysis of the target genes identified revealed enrichment of protein binding, membrane, cell cycle, and so on. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of age-related miRNAs for target genes revealed 139 enriched pathways, such as signaling pathways regulating stem cell pluripotency, metabolic pathways, and the Hippo signaling pathway. This suggests that miRNAs play a key role in male fertility changes with increasing age and provides new evidence for the study of the mechanism of age-related male fertility decline.
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Humains , Mâle , Jeune adulte , Adulte , Adulte d'âge moyen , microARN/génétique , Transduction du signal/génétique , Spermatozoïdes/métabolisme , Analyse de profil d'expression de gènesRésumé
OBJECTIVES@#To study the effect of ligustrazine injection on mitophagy in neonatal rats with hypoxic-ischemic encephalopathy (HIE) and its molecular mechanism.@*METHODS@#Neonatal Sprague-Dawley rats, aged 7 days, were randomly divided into a sham-operation group with 8 rats, a model group with 12 rats, and a ligustrazine group with 12 rats. The rats in the model group and the ligustrazine group were used to establish a neonatal rat model of HIE by ligation of the left common carotid artery followed by hypoxia treatment, and blood vessels were exposed without any other treatment for the rats in the sham-operation group. The rats in the ligustrazine group were intraperitoneally injected with ligustrazine (20 mg/kg) daily after hypoxia-ischemia, and those in the sham-operation group and the model group were intraperitoneally injected with an equal volume of normal saline daily. Samples were collected after 7 days of treatment. Hematoxylin and eosin staining and Nissl staining were used to observe the pathological changes of neurons in brain tissue; immunohistochemical staining was used to observe the positive expression of PINK1 and Parkin in the hippocampus and cortex; TUNEL staining was used to measure neuronal apoptosis; Western blotting was used to measure the expression levels of the mitophagy pathway proteins PINK1 and Parkin and the autophagy-related proteins Beclin-1, microtubule-associated protein 1 light chain 3 (LC3), and ubiquitin-binding protein (P62).@*RESULTS@#Compared with the sham-operation group, the model group had a significant reduction in the number of neurons, an increase in intercellular space, loose arrangement, lipid vacuolization, and a reduction in Nissl bodies. The increased positive expression of PINK1 and Parkin, apoptosis rate of neurons, and protein expression levels of PINK1, Parkin, Beclin1 and LC3 (P<0.05) and the decreased protein expression level of P62 in the hippocampus were also observed in the model group (P<0.05). Compared with the model group, the ligustrazine group had a significant increase in the number of neurons with ordered arrangement and an increase in Nissl bodies, significant reductions in the positive expression of PINK1 and Parkin, the apoptosis rate of neurons, and the protein expression levels of PINK1, Parkin, Beclin1, and LC3 (P<0.05), and a significant increase in the protein expression level of P62 (P<0.05).@*CONCLUSIONS@#Ligustrazine can alleviate hypoxic-ischemic brain damage and inhibit neuronal apoptosis in neonatal rats to a certain extent, possibly by inhibiting PINK1/Parkin-mediated autophagy.
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Rats , Animaux , Hypoxie-ischémie du cerveau/métabolisme , Animaux nouveau-nés , Rat Sprague-Dawley , Bécline-1 , Autophagie , Ubiquitin-protein ligases/métabolisme , Protein kinases/métabolismeRésumé
AIM:Observation of neuroprotective effects of Toddalia asiatica(TA)on cerebral ischemia-reper-fusion injury(CIRI)in rats by investigating the effects and mechanisms of drugs on the polarization of microglia M1/M2 subtype and the Toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)/nuclear factor-κB(NF-κB)path-way.METHODS:The modified thread occlusion method was used to establish a rat model of CIRI,and the rats were ran-domly divided into the model group,Toddalia asiatica(1.08 g/kg)group,donepezil hydrochloride(0.45 mg/kg)group,and sham group,with 16 rats in each group.Based on the assessment of neurofunctional changes in each group of rats,HE and Nissl staining were used to observe the pathological changes in brain tissue.TUNEL staining was performed to de-tect neuronal apoptosis,immunohistochemistry was used to detect the expression of M1 microglia marker ionized calcium-binding adapter molecule 1(Iba1),M2 microglia marker arginase 1(Arg1),and TLR4 localization.Western blot was used to detect the expression of microglia polarization proteins and proteins related to TLR4/MyD88/NF-κB pathway in the hippocampus region.RESULTS:Compared with sham group,the model group rats had higher neurological function scores(P<0.01),and neuronal arrangement in the hippocampus and cortex was loose and disordered,Nissl bodies de-creased,and neuronal apoptosis increased.The numbers of M1 microglia marker Iba1-,M2 microglia marker Arg1-,and TLR4-positive cells were significantly increased.In addition,the protein levels of TLR4,MyD88,p-NF-κB p65,NF-κB p65,p-NF-κB inhibitory factor(p-IκB),Iba1,interleukin-6(IL-6),and Arg1 in the hippocampus were elevated(P<0.05),while IL-4 and IL-10 expression were decreased(P<0.01).Compared with model group,the Toddalia asiatica group and Donepezil hydrochloride group showed increased protein expression of Arg1,IL-4 and IL-10(P<0.05),while the other indicators were decreased(P<0.05,P<0.01).CONCLUSION:Toddalia asiatica possesses neuroprotective effects on CIRI rats,which may be attributed to its ability to regulate M1/M2 polarization and inhibit the TLR4/MyD88/NF-κB-mediated inflammatory pathway.
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This study was designed to investigate the effect of dihydromyricetin (DHM) on inducing apoptosis of ovarian cancer cells A2780 through endoplasmic reticulum stress (ERS) pathway and the mechanisms involved in vitro and in vivo. A2780 cells were treated with different concentrations of DHM, and the protein expression levels of glucose-regulated protein 78 (GRP78) which is related to ERS increased, apoptotic proteins C/EBP-homologous protein (CHOP), and cysteinyl aspartate specific proteinase-12 (caspase-12) elevated. After pretreatment with ERS inhibitor, 4-phenyl butyric acid (4-PBA), following the intervention with DHM, the A2780 cell viability decreased and apoptotic rate increased. All animal welfare and experimental procedures were approved by the Animal Ethics Committee of Chongqing Medical University. Intraperitoneal injection of DHM suspension into nude mice with ovarian cancer could significantly inhibit the growth of transplanted tumor in vivo, increase the protein expression levels of GRP78, CHOP, and caspase-3. Moreover, swollen and broken endoplasmic reticulum could be observed in tumor tissues, suggesting that DHM intervention induces apoptosis mediated by ERS. The results indicated that DHM could induce apoptosis of ovarian cancer cells and inhibit the growth of transplanted tumors in nude mice, which might be related to the activation of ERS pathway.
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Objective:To explore the effect of trillin on oxidative stress response and nuclear factor E2-related factor 2/antioxidant response element (Nrf2/ARE) pathway in rats after spinal cord injury (SCI). Methods:A total of 108 male Sprague-Dawley rats were randomly divided into sham group (n = 36), model group (n = 36) and trillin group (n = 36), each group was divided into one day, three days and seven days subgroups, with twelve rats in each subgroup. The SCI model was established by modified Allen's heavy strike method in the model group and the trillin group, but no obvious injury in the sham group. The trillin group was given trillin 200 mg/kg every day, and the same amount of normal saline was given in the sham group and model group, twice a day. BBB score was performed one day, three days and seven days after modeling. Morphological changes were tested by Nissl's staining, and the changes of malonaldehyde (MDA) content and superoxide dismutase (SOD) activity were detected by ELISA seven days after modeling. The expression of Nrf2, Kelch like ECH associated protein 1 (Keap1), NAD(P)H quinone oxidoreductase (NQO1) and haemoxygenase 1 (HO-1) were detected by Western blotting one day, three days and seven days after modeling. Results:Compared with the model group, BBB scores increased (P < 0.05); the structure of spinal cord was more complete and the number of Nissl bodies increased; SOD activity increased (P < 0.05) and MDA content decreased (P < 0.05); the expression of Nrf2, Keap1, NQO1 and HO-1 increased (P < 0.05) in the trillin group. Conclusion:Trillin may play a protective role in spinal cord injury by inhibiting oxidative stress response and improving the motor function.
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Aim To explore the effect of Trillium tschonoskii Maxim (TTM) on the expression of miR-NA-155-3p in rats with brain aging induced by D-gal. Methods Fifty SD rats were divided into five groups randomly. The rats were administered with 0. 9% normal saline (NS) subeutaneously every day in control group, 200 mg • kg • d-1 of D-galactose (D-gal) daily inD-galmodelgroup,and50,100and200mg • kg • d-1 of TTM by gavage 2 hours before D-gal injection everyday in TTM treatment groups for 6 weeks. After 5 weeks, Morris water maze was used to test the ability of spatial learning and memory every day. At the 6th week, rats were sacrificed arid hippocampi were tested by Nissl staining and 8-OHdG immunohistochemical (8-OHdC) staining. The expression of miR-155-3p was determined by Real-time PCR, and the levels of Rheb. mTOR, p-inTOR and p70S6K were detected by Western blot. Results The length of escape latency time and path distance in five groups showed a trend of shortening gradually in the orientation navigation experiments. The average escape latency and the distance in D-gal group were longer than those in control group and TTM group (P <0. 01 and 0. 05) , and the number of crossing platform limes less too (P < 0. 05). The arrangement of neurons was irregular and the intercellular space widened in D-gal group compared with those in TTM group by HE staining. There were more Nissl particles in neurons of the hippocampal CA1 area in con-trol group than that in D-gal group, and TTM treatment could increase the number of Nissl bodies-induced by D-gal. Compared with control group, the fluorescence density of 8-OHdG in D-gal group significantly in-creased (P <0. 01) , while that in TTM group was lower than that in D-gal group (P < 0. 05). The expression of miR-155-3p in the hippocampi in D-gal group was significantly higher than that in normal group (P < 0. 05) , while TTM treatment could alleviate D-gal-in-duced increase of miR-155-3p (P < 0. 05) , followed by an increase of the levels of Rheb and p70S6K, and decrease of mTOR. Conclusions The expression of miR-155-3p increased in the hippocampi of aging rats induced by D-gal. TTM could execute the anti-aging process of brain and down-regulate the level of iniR-155-3p through Rheb/mTOR/p70S6K signaling pathway.
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Objective To investigate the effect of Trillium tschonoskii Maxim extract on the expression of ciliary neurotrophic factor (CNTF) and its receptor (CNTFRα) after spinal cord injury in rats. Methods Forty-five rats were equally and randomly divided into control group (group A), model group (group B) and Trillium tschonoskii Maxim treated group (group C). Allen's weight drop method was used to reproduce acute spinal cord injury (SCI) model in rats of the group B and C. In group C, the rats were gavaged with Trillium tschonoskii Maxim extract 2 weeks before the injury, while rats in group A and B were fed a same quantity of distilled water.1, 7 or 14 days after injury, the rats were sacrificed to observe the structure of nerve cells after HE and Nissl staining, and the expression of CNTF and CNTFRα with immunohistochemical method, RT-PCR and Western blotting. Results HE staining showed that the structure of spinal cord in the the rats group C was more discernible, with milder edema and necrosis of nerve cells, as compared with that of group B. Nissl staining showed that Nissl bodies were decreased or disappeared in anterior horn motor neurons in both group B and C, but it was significantly less marked in group C than that in group B. Immunohistochemical staining, Western blotting and RT-PCR revealed that the protein and mRNA of CNTF and CNTFRα were positively expressed in rats of every group. The mRNA levels of CNTF and CNTFRα in group C were higher than those in group B. Conclusions Extract of Trillium tschonoskii Maxim can up-regulate the expression of CNTF and CNTFRα, and plays a protective role against injury to spinal cord.
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<p><b>OBJECTIVE</b>To explore the effect of Se-riched soybean peptide (SSP) on antioxidant function in rats of fatty liver caused by high-fat diet.</p><p><b>METHODS</b>Forty Wistar rats were divided into 4 groups randomly and fed with standard diet and water (NC), high-fat diet and water (HC), high-fat diet and SSP (0.1 g/d) (SeH), standard diet and SSP (0.1 g/d) (SeN) respectively. After 10 weeks, the rats were killed to investigate the pimelosis level in liver tissues by Sudan III staining and the expression of hepatic GRP78 by immunohistochemical analysis. We also analyzed the changes of liver function, blood lipid, the glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in livers and serum.</p><p><b>RESULTS</b>The pimelosis level, total cholesterol (TC), triglyceride (TG), MDA contents and the expression of GRP78 in HC group were significantly higher than those in NC, SeN, SeH groups. The activities of GSH-Px and SOD in liver and serum were markedly up-regulated in SeH (P < 0.01). There was no significant difference between NC and SeN groups.</p><p><b>CONCLUSION</b>SSP can improve liver cell injury and the antioxidant functions in rats with fatty liver effectively and decrease the expression of GRP78 in liver.</p>