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Objective:To investigate the neuroprotective effect of cerebroprotein hydrolysate (CH) -Ⅰ on cerebral ischemia-reperfusion injury in rats and its mechanism.Methods:Eighty adult healthy male SD rats were randomly divided into sham operation group, model group, CH-Ⅰ intervention group and cerebrolysin (CBL) positive control group. The model of ischemia-reperfusion injury was induced by temporarily occluding the left middle cerebral artery with suture-occluded method. The CH-Ⅰ and CBL groups intraperitoneally injected with CH-Ⅰ and CBL at 0, 3, 6 and 12 h after reperfusion at the dose of 20 mg/kg. The sham operation group and the model group were injected with the same volume of normal saline. At 24 h after reperfusion, the behavior changes of the rats were detected by the modified neurological severity score (mNSS). The volume of cerebral infarction was detected by TTC staining. The morphology and structure of neurons in ischemic cortex were observed by Nissl staining. The apoptosis of neurons in ischemic cortex was detected by TUNEL staining. The expression changes of phosphorylated extracellular signal-regulated kinase (pERK) 1/2, phosphorylated mitogen-activated protein kinase/extracellular signal-regulated kinase (pMEK) 1/2, phosphorylated cAMP response element binding protein (pCREB) and brain-derived neurotrophic factor (BDNF) in the ischemic cortex were detected by Western blot.Results:At 24 h after reperfusion, the mNSS score and cerebral infarct volume in the model group were significantly higher and larger than those in the sham group (all P<0.001). The mNSS scores and cerebral infarct volumes in the CH-Ⅰ and CBL groups were significantly reduced compared with those in the model group (all P<0.05), but there was no significant difference between the CH-Ⅰ group and the CBL group. Nissl and TUNEL staining showed that the degenerative cell index and apoptotic cell index in the CH-Ⅰ group were significantly lower than those in the model group (all P<0.01), but there were no significant difference between the CH-Ⅰ group and the CBL group. Western blot analysis showed that compared with the sham operation group, the pMEK1/2, pERK1/2 and pCREB expressions in ischemic cortex were significantly enhanced and the BDNF expression was significantly attenuated in the model group ( P<0.05). Compared with the model group, pMEK1/2, pERK1/2, and pCREB expressions in the CH-Ⅰ group were significantly decreased (all P<0.05), and the BDNF expression was significantly increased ( P<0.05). Conclution:CH-Ⅰ can reduce cerebral infarct volume and improve neurological function, and its mechanism may be associated with the inhibition of the MEK-ERK-CREB pathway as well as the enhancement of BDNF expression.
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Objective:To study the effects of regenerated tissue extracts after liver injury on the proliferation, differentiation, migration and invasion of SK-HEP1 cells.Methods:Regenerated tissue extracts after liver injury were used to induce SK-HEP1 cells after enrichment, their effects on the proliferation, differentiation, migration and invasion of SK-HEP1 cells were observed through in vitro cell culture, MTT, flow cytometry and transwell assays.Results:In response to the action of regenerated tissue extracts after liver injury, SK-HEP1 cells were blocked in GConclusions:To a certain extent, regenerated tissue extracts after liver injury can inhibit the proliferation, differentiation, migration and invasion of hepatoma cells, showing an important potential of being a differentiating agent for the treatment of liver cancer.
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Objective: To study the effects of regenerated tissue extracts after liver injury on the proliferation, differentiation, migration and invasion of SK-HEP1 cells. Methods: Regenerated tissue extracts after liver injury were used to induce SK-HEP1 cells after enrichment, their effects on the proliferation, differentiation, migration and invasion of SK-HEP1 cells were observed through in vitro cell culture, MTT, flow cytometry and transwell assays. Results: In response to the action of regenerated tissue extracts after liver injury, SK-HEP1 cells were blocked in G
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Objective To simulate the chemical microenvironment of traumatic brain injury (TBI) under mild hypothermia, and investigate the effect of such microenvironment on umbilical cord mesenchymal stem cells (UCMSCs) in vitro.Methods Eighteen SD rats were allocated to shamoperated group, TBI group and mild hypothermia group according to the random number table, with 6 rats per group.Rat models of TBI were made by electric cortical contusion impactor.After systemic mild hypothermia (33℃) for 4 h, brain tissue homogenate extracts were harvested.Polyacrylamide gels mimicking the elastic modulus of brain were manufactured.Human UCMSCs were isolated and cultured on the gels, added with brain tissue extracts from each group.After 24 h, the apoptosis level of UCMSCs was checked, and the medium was changed with normal one.Cell growth and morphological changes in each group were given dynamic observation.Seven days later, cell immunofluorescence was implemented, with the differentiation level of each group estimated.Results Apoptotic rate in TBI group was 73.47％,significantly higher than 10.42％ in sham-operated group (P ＜0.01).While the apoptotic rate was 28.57％ in mild hypothermia group, indicating mild hypothermia significantly reversed the apoptosis of cells in TBI group (P ＜ 0.01).Cell immunofluorescence demonstrated rate of neuronal differentiation of UCMSCs in sham-operated group, TBI group and mild hypothermia group was 16.48％, 2.59％ and 11.83％ respectively.Mild hypothermia resulted in significantly improved neuronal differentiation of UCMSCs after TBI (P ＜ 0.05).Conclusions More apoptosis and lower neuronal differentiation ability are observed in UCMSCs in the chemical microenvironment after TBI.However, mild hypothermia significantly reverses the elevation of apoptosis and restores the neuronal differentiation capacity of UCMSCs after TBI.
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Objective To investigate the effect of early using periplaneta americana extract on gastrointesti-nal function in critically patients with infection,and to evaluate its possible mechanism.Methods 66 patients with infection in Intensive Care Unit(ICU)were divided into control group and treatment group(33 cases in each group) according to the random number table.All patients in the two groups were given conventional cluster treatment,the patients in the treatment group additionally received periplaneta americana extract.At the beginning of treatment and 3,7 and 10 days after treatment,the indicators of bowel sounds,gastrointestinal function,APACHEⅡ and blood lactic acid level were respectively tested or recorded.Results At the beginning of treatment,there were no statistically sig-nificant differences between the two groups in terms of gastrointestinal function,bowel sounds,APACHE II score and blood lactic acid level(t =0.132,0.201,0.329,0.190,all P >0.05).3,7 days after treatment,the gastrointestinal function[(1.56 ±0.49)points vs.(1.32 ±0.45)points,(1.27 ±0.44)points vs.(1.02 ±0.42)points],bowel sounds[(1.58 ±0.76)times/min vs.(2.27 ±0.75)times/min,(2.15 ±0.91)times/min vs.(3.11 ±0.97)times/min],APACHE II score[(19.61 ±4.02)points vs.(17.50 ±4.37)points,(17.69 ±3.94)points vs.(15.24 ± 3.82)points]and blood lactic acid level[(2.94 ±0.88)mmol/L vs.(2.45 ±0.75)mmol/L,(2.51 ±0.59)mmol/L vs.(2.20 ±0.55)mmol/L],the terms in the treatment were better than those in the control group,the differences were statistically significant(t3 =2.072,t7 =2.361;t3 =3.712,t7 =4.146;t3 =2.041,t7 =2.565;t3 =2.434,t7 =2.208,all P <0.05).Conclusion Early using periplaneta americana extract has protective effect on gastrointestinal function in critically patients with infection,and improve the condition in a certain degree.
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Objective To study the effectiveness of vaccinia vaccination rabbit inflammatory skin extracts on blood homocysteine(Hcy) and insulin-like growth factor-1(IGF-1) in patients with diabetic peripheral neuropathy. Methods 100 patients with diabetic peripheral neuropathy were randomly divided into the two groups,the observa-tion group(n=50 cases) and the control group(n=50 cases).The observation group was treated through vaccinia vaccination rabbit inflammatory skin extracts, while the control group was treated through mecobalamin.They were treated for 15 days.Median nerve,peroneal motor nerve conduction velocity (MCV) and sensory nerve conduction velocity ( SCV) were determined before and after treatment.Blood IGF-1 and Hcy were detected.Results TSS were significantly lowered after treatment (t=9.772,13.624,all P<0.01).Compared with the control group,the observa-tion group was significantly decreased (t=3.925,P<0.05).After treatment,the median nerve,peroneal nerve MCV and SCV were significantly increased (t=5.103,3.019,4.998,2.928,5.128,3.112,5.286,3.118,all P<0.05). Compared with the control group,theobservation group was observed to increase more significantly(t=2.826,2.743, 3.268,3.096,all P<0.05).After treatment,serum IGF-1 were significantly elevated,and compared with the control group after treatment,post-treatment observation group increased more significantly(t=4.179,P<0.05).After treat-ment plasma Hcy were significantly lower than that of the control group after treatment,the observation group after treatment significantly decreased(t=3.165,P<0.05).Conclusion Vaccinia vaccination rabbit inflammatory skin extracts can improve blood Hcy and IGF-1 in patients with diabetic peripheral neuropathy.
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Objective To compare three methods for the extraction of mycobacterial DNA.Methods Two commercial DNA extraction kits and an ordinary freeze-thawing method were used to extract DNA from the pure suspensions of three species of Mycobacteria (M.tuberculosis,M.leprae and M.smegmatis) at different densities (1 × 10 to 1 × 105 cells/ml),simulated clinical specimens containing different concentrations of mycobacterial cells (1 × 10 to 1 × 104 cells/ml).The purity and concentration of the extracted DNA were evaluated.Then,PCR was performed to amplify the 16S rRNA region of Mycobacteria.The performance of the three methods was compared by the purity and concentration of extracted DNA as well as the results of PCR.Further more,76 clinical skin specimens suspected to be infected with Mycobacteria were used to further validate the performance of these methods.Results All the extracted DNA samples could be detected by PCR.The highest purity of DNA was obtained by the kit A,followed sequentially by the freeze-thawing method and the kit B.When pure suspensions were used,the detection limit was consistently 1 × 102 cells/ml for all the three methods.With simulated specimens,the detection rate was consistently 100％ for all the three methods at the concentration of 1 × 103 cells/ml,60％ (12/20),55％ (11/20) and 55％ (11/20) for the kit A,kit B and freeze-thawing method respectively at the concentration of 1 × 102 cells/ml.The analysis of clinical specimens showed that the kit B could be used to extract DNA from paraffin-embedded specimens,with the detection rate similar to that of kit A and freeze-thawing method.Conclusions The kit A could rapidly yield high-quality genomic DNA of Mycobacteria by repeated cleaning of columns,and may serve as the optimal method for scientific and clinical studies,and the kit B is suitable for extracting mycobacterial DNA from fresh tissue specimens besides paraffin-embedded specimens.
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Objective To investigate the protective effects of extract of Periplaneta americana (APA) against carbon tetrachloride (CCl4)-induced hepatic fibrosis in rats. Methods Seventy SD rats were divided into five groups:normal con-trol group (C), model control group (M), extract of APA 1 group (APA1), extract of APA 2 group (APA2) and reduced glutathi-one group (R). The liver fibrosis model was induced by injecting 40%CCl4-olive solution subcutaneously for seven weeks in M, APA1, APA 2 and R groups. Drugs were given at the same time. Rats were killed at 7-week (C, M, APA1 and R groups), and were killed at 9-week (APA2 group). The serum values of transaminase (ALT, AST), albumin (ALB), hyaluronic acid (HA) and laminin (LN), and hepatic inflammation and fibrosis were detected respectively. The expressions of apoptotic relat-ed gene Bcl-2 and Bax protein in central veins and portal areas of hepatic lobules were detected by immunohistochemical method. Results Rats showed poor nutritional status, and significantly increased transaminase, HA and LN in M group, but the serum level of ALB was significantly lower than that in C group. There was extensive necrosis of liver cells, obviously fi-brosis or cirrhosis in model rats’liver tissues. The serum contents of ALT, AST, HA and LN were significantly decreased, the serum level of ALB were significantly increased in APA1,APA2 and R groups (P<0.05). There were complete hepatic lobule and no obvious fibrous tissue hyperplasia in liver biopsy of APA1 and R groups. The values of Bcl-2/Bax proteins in central vein of liver tissues were significantly higher in APA1,APA2 and R groups than those of M group (P<0.05), but the ratio of Bcl-2/Bax was lower in periportal area of liver tissues (P<0.05). There were significantly higher body weight and ALB levels in APA1 group than those in R group (P<0.05). The level of transaminase was slightly higher in APA1 group than that of R group. The degeneration of liver cells was found mostly in R group. Conclusion The extract of Periplaneta americana has a role in protecting liver cells, inhibiting the progression of hepatic fibrosis, and improving the nutritional sta-tus of trial rats.
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Objective To investigate the cell cycle and mechanisms of HCC Bel-7404 cell line by treatment of extract of centipede ( ECP). Method Bel-7404 cell line was cultured in vitro. ECP was applied to the interference of the growth of Bel-7404 with different drug concentration. Cell cycles and apoptosis ratios of ECP group were detected by flow cytometry (FCM). To investigate the expression of apop-tosis related genes XIAP and Bax, we also detect HCC Bel-7404 cell line after 48 hours treated with ECP by immunohistochemical method. Result The results of FCM displayed that ECP in treatment group mainly retard cell cycle phase in G0/G1 after 48 hours. The ratios of G0/ G1 .proliferation index (PI) and apoptosis ratios were significantly different between treatment group and control group when the concentrations were different. It was also significantly different between treatment group and control group when the same concentration of ECP was used. It showed that XIAP gene expression decreased gradually while Bax gene expression increased with the increase of ECP concentration, and these were statistical significance in contrast to control group ( P <0.05). Conclusion ECP mainly retarded cell cycle phase of Bel-7404 in G0/G1, suppressed the proliferation, and could induce it to apoptosis. The inhibitory effect was time - concentration dependent. The mechanisms were due to promote Bax gene and inhibit XIAP gene expression in HCC Bel-7404 cell line.
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Effect of cell-free preparation of human fetal liver (cytosol and its dialysa-te) , human fetal muscle cytosol(FMC) and human adult liver cytosol (ALC) on survival rate of rats with acute hepatic failure induced by D-galactosamine was studied. The components of the 4 preparations were analysed preliminarily. Results: 1 ,fetal liver cytosol(FLC) could increase the survival rate(SR) of rats with hepatic failure(HF) comparing with normal saline(P0.05) . The SR of rats with HF was reduced by ALC markedly) 2,IgG,IgA,IgM and C3 could not be detected in FLC, but a few kinds of hormones, trace elements, glucose and lipids, and biological large molecule substances (BLMS) such as nucleic acids and proteins were found. As to free amino acid,content of branch chain amino acids in FLC was significantly higher than that in ALC, but the content of aromatic amino acids and ammonia was markedly lower than that in ALC.The components in FLD were almost the same as in FLC except BLMS.These data suggest that the soluble BLMS in FLC may play a key role in treating HF. ALC could reduce SR and contained a large amount of poisonous substances such as aromatic amino acids and ammonia.