AIM: To observe the effect of mouse Sim2 (mSim2) eukaryotic expression vector transfection on the cell cycle in PC12 cells in vitro and to explore the role of Sim2 in the pathogenesis of Down syndrome. METHODS: The full open reading frame of mSim2 was amplified by reverse transcription-polymerase chain reaction (RT-PCR) and cloned into the vector pcDNA3. Then the constructed pcDNA3-Sim2 vectors were transiently transfected into PC12 with Lipofectamine~ TM . The expression of mSim2 was detected by RT-PCR. The effect of mSim2 on the cell cycle was observed by flow cytometry. RESULTS: The eukaryotic expression vector mSim2 was successfully constructed. There was notable expression of mSim2 in the cells transfected with pcDNA3-Sim2. There were more cells in G_0/G_1 phase in the pcDNA3-Sim2 transfected cells than that in the control (P

To investigate the effects of ischemia-reperfusion on the levels of nitric oxide and nitric oxide synthase isoforms (nNOS and iNOS), rat organotypic hippocampus slice were cultured in vitro and subjected to ischemia by oxygen-glucose deprivation (OGD) for 30 min and then placed in the normal culture condition. The ischemia-reperfusion produced a time-dependent increase in nitrite levels in the culture medium. Reverse transcriptional-polymerase chain reaction showed augmented levels of mRNA for both nNOS and iNOS when compared with control at 12 h and remained increase at 36 h after OGD (P < 0.05). The protein levels of both nitric oxide synthase isoforms increased significantly as determined by Western Blot. OGD also caused neurotoxicity in this model as revealed by the elevated lactate dehydrogenase (LDH) efflux into the incubation solution. The results suggest that organotypic hippocampus slice is a useful model in studying ischemia-reperfusion brain injury. NO and NOS may play a critical role in the ischemia-reperfusion brain damage in vitro.
Animals, Newborn ; Cell Hypoxia ; Hippocampus/cytology ; Hippocampus/*metabolism ; Nitric Oxide/*metabolism ; Nitric Oxide Synthase Type I/*metabolism ; Nitric Oxide Synthase Type II/*metabolism ; RNA, Messenger/metabolism ; Rats, Sprague-Dawley ; Reperfusion Injury/*metabolism ; Tissue Culture Techniques

The aim of this study was to express and purify human histydyl-tRNA synthetase related gene and to prepare its polyantibody. The open reading frame was amplified by PCR, and then recombined into prokaryotic expression vector pQE30 and transformed into E. coli M15 for expression. The expressed products were induced by IPTG after the reconstructed pQE30 was transferred into M15. After purified by Ni affinity chromatography, the product was identified to be a single band by SDS-PAGE. The rabbits were inoculated with purified products. High-titer polyantibody was successfully prepared. Highly-purified expression product and prepared polyantibody may provide a good basis for further study.
Antibodies/*genetics ; Antibodies/immunology ; Escherichia coli/genetics ; Escherichia coli/metabolism ; Histidine-tRNA Ligase/biosynthesis ; Histidine-tRNA Ligase/*genetics ; Histidine-tRNA Ligase/immunology ; Open Reading Frames/genetics ; Prokaryotic Cells/metabolism

Objective To explore the three criteria of Ranson,CT severity index (CTSI) and the bedside index of severity in acute pancreatitis (BISAP) in assessment of severity and prognosis of acute pancreatitis.Methods A total of 503 diagnosed acute pancreatitis patients from January 2008 to April 2011 were studied retrospectively,including 356 mild acute pancreatitis (MAP) cases and 147 severe acute pancreatitis (SAP) cases.The differences of the three criteria of Rason,CTSI and BISAP in assessment of severity and prognosis of acute pancreatitis were compared with receiver operating characteristic (ROC) curve.The SAP group was divided into no organ failure group and organ failure group,and the predictive value of the three criteria in acute pancreatitis with organ failure was compared.Results There was significant differences between MAP group and SAP group with Rason,CTSI and BISAP score (x2 =236.88、126.24 and 101.27,P＜0.01).The sensitivity (97.3％) and AUC value (0.93) of Ranson criterion was highest.In no organ failure group and organ failure group of 147 SAP patients,there was significant difference between Ranson criterion and BISAP criterion (x2 =17.67 and 26.12,P＜0.01),both sensitivity were 100％,the specificity were 96％ and 85％ respectively,and the area under ROC curve (AUC) value of BISAP criterion (0.8) was the highest.In illness improved group and deteriorated group,the difference of score between Ranson criterion and BISAP criterion was significant (x2 ＝ 9.53 and 10.19,P＜0.05),and the AUC value of BISAP criterion (0.74) was the highest.Conclusions All three criteria can be used to determine the severity of acute pancreatitis.In predicting the risk of SAP with organ failure and prognosis,BISAP criterion is better than Ranson criterion.BISAP criterion is simple and easy to practice,which provides an important rule for evaluating the clinical condition of acute pancreatitis.

Objective: To study the effects of Rhizoma Polygoni Cuspidati on expression of transforming growth factor beta 1(TGF-?1) in rats with pulmonary fibrosis induced by bleomycin.Method: 120 male SD rats were randomly divided into five groups: the control group(n=30),the bleomycin(BLM) model group(n=30),the Rhizoma Polygoni Cuspidati prophylactic group(n=30),the 7 th day treatment group(n=20) and the 28 th day treatment group(n=10).The rat models of pulmonary fibrosis were established by intratracheal injection of bleomycin, while the control group was given with normal saline instead.The Rhizoma Polygoni Cuspidati prophylactic group received intragastric administration with 4ml?kg-1?d-1 Rhizoma Polygoni Cuspidati 2 days before setting up models.The 7 th day treatment group and the 28 th day treatment group received the treatment on the seventh day and the twenty-eighth days after establishing models respectively.The other two groups received saline instead.On the 3rd,7th,14th,28th,42th and 56th day after instillation of bleomycin,lung samples were obtained and the content of hydroxyproline in lung tissue homogenate was detected to judge the degree of fibrosis and the therapeutic efficacy.At the same time the express of TGF-?1mRNA and TGF-?1 protein was examined by RT-PCR and immunohistochemical method.Results: Compared with the model group,the content of hydroxyproline in the Rhizoma Polygoni Cuspidati prophylactic group and the 7th day treatment group decreased significantly(P

The effect of electroacupuncture (EA) on TRPM7 mRNA expression of focal cerebral ischemia in rats and further the role of EA in the relationship between TRPM7 and trkA pathway was investigated. Thirty SD rats were randomly divided into 5 groups : normal group, ischemia/reperfusion group, EA treated group (ischemic rats with EA treatment), TE infusion group (ischemic rats with EA treatment and TE buffer infusion), AS-ODN group (ischemic rats with EA treatment and antisense trkA oligonucleotide infusion). The stroke animal model was established by the modified method of middle cerebral artery occlusion. Antisense trkA oligonucleotide that blocked NGFs effects was injected into cerebroventricle before EA. The TRPM7 mRNA was detected by RT-PCR method. The results showed that there were low TRPM7 mRNA levels in cortex and hippocampus in normal group. Compared with normal group, TRPM7 mRNA expression was increased significantly in ischemia/reperfusion group (P<0.05). A significant reduction in the expression of TR-PM7 mRNA was found in EA treated group in contrast to ischemia/reperfusion group (P<0.05). The expression of TRPM7 mRNA in AS-ODN group was remarkably increased compared with EA treated group and TE infusion group (P<0.05). The results indicated that TRPM7 channels in the ischemic cortex and hippocampus in rats might play a key role in ischemic brain injury. EA could reverse the overexpression of TRPM7 in cerebral ischemia/reperfusion rats. And the inhibitory effect of EA on TRPM7 channels might be through trkA pathway.

The aim of this study was to express and purify human histydyl-tRNA synthetase related gene and to prepare its polyantibody. The open reading frame was amplified by PCR, and then recombined into prokaryotic expression vector pQE30 and transformed into E. coli M15 for expression. The expressed products were induced by IPTG after the reconstructed pQE30 was transferred into M15. After purified by Ni affinity chromatography, the product was identified to be a single band by SDS-PAGE. The rabbits were inoculated with purified products. High-titer polyantibody was successfully prepared. Highly-purified expression product and prepared polyantibody may provide a good basis for further study.
Antibodies ; genetics ; immunology ; Escherichia coli ; genetics ; metabolism ; Histidine-tRNA Ligase ; biosynthesis ; genetics ; immunology ; Humans ; Open Reading Frames ; genetics ; Prokaryotic Cells ; metabolism

Summary: The effect of electroacupuncture (EA) on TRPM7 mRNA expression of focal cerebral ischemia in rats and further the role of EA in the relationship between TRPM7 and trkA pathway was investigated. Thirty SD rats were randomly divided into 5 groups : normal group, ischemia/reperfusion group, EA treated group (ischemic rats with EA treatment), TE infusion group (ischemic rats with EA treatment and TE buffer infusion),AS-ODN group (ischemic rats with EA treatment and antisense trkA oligonucleotide infusion). The stroke animal model was established by the modified method of middle cerebral artery occlusion. Antisense trkA oligonucleotide that blocked NGF's effects was injected into cerebroventricle before EA. The TRPM7 mRNA was detected by RT-PCR method. The results showed that there were low TRPM7 mRNA levels in cortex and hippocampus in normal group. Compared with normal group, TRPM7 mRNA expression was increased significantly in ischemia/reperfusion group (P<0.05). A significant reduction in the expression of TRPM7 mRNA was found in EA treated group in contrast to ischemia/reperfusion group (P<0.05). The expression of TRPM7 mRNA in AS-ODN group was remarkably increased compared with EA treated group and TE infusion group (P<0.05). The results indicated that TRPM7 channels in the ischemic cortex and hippocampus in rats might play a key role in ischemic brain injury. EA could reverse the overexpression of TRPM7 in cerebral ischemia/reperfusion rats. And the inhibitory effect of EA on TRPM7 channels might be through trkA pathway.

To investigate the effects of ischemia-reperfusion on the levels of nitric oxide and nitric oxide synthase isoforms (nNOS and iNOS), rat organotypic hippocampus slice were cultured in vitro and subjected to ischemia by oxygen glucose deprivation (OGD) for 30 min and then placed in the normal culture condition. The ischemia-reperfusion produced a time-dependent increase in nitrite levels in the culture medium. Reverse transcriptional-polymerase chain reaction showed augmented levels of mRNA for both nNOS and iNOS when compared with control at 12 h and remained increase at 36 h after OGD (P＜0.05). The protein levels of both nitric oxide synthase isoforms increased significantly as determined by Western Blot. OGD also caused neurotoxicity in this model as revealed by the elevated lactate dehydrogenase (LDH) efflux into the incubation solution. The results suggest that organotypic hippocampus slice is a useful model in studying ischemia-reperfusion brain injury. NO and NOS may play a critical role in the ischemia-reperfusion brain damage in vitro.

Objective To explore the effect of laparoscopic salpingectomy on ovarian reserve function and serum neuropeptide Y(NPY)and cortisol(Cor)in patients with ectopic pregnancy.Methods 82 patients with ectopic pregnancy who had visited Chaohu Hospital Affiliated to Anhui Medical University from June 2018 to June 2021 were randomly divided into two groups,with 41 in each group.The control group underwent laparoscopic salpingostomy and suturing for embryo retrieval,while the study group underwent laparoscopic salpingectomy.Surgical duration,intraoperative blood loss,time to postsurgical mobilization,length of hospital stay,serum NPY and Cor levels,antral follicle count(AFC),indexes for peak flow velocity and resistance of ovarian artery were compared between the two groups.Follow-up was conducted for two years after surgery,and the success rate of pregnancy and the ectopic pregnancy rate were recorded.Results The surgical duration in the study group was shorter than that in the control group(P<0.05).The differences in serum NPY and Cor values between the two groups immediately after surgery and at postoperative week one were not statistically significant(P>0.05).At postoperative months1 and 6,the peak flow velocity of the affected ovary was lower in the study group than in the control group,whereas the resistance index was higher(P<0.05).At postoperative month 6,both groups showed an increase in peak flow velocity and a decrease in resistance index(P<0.05).There were no statistically signifi-cant differences in AFC and the success rate of pregnancy between the two groups postoperatively(P>0.05).The ectopic pregnancy rate in the study group was lower than that in the control group(P<0.05).Conclusion As compared with laparoscopic salpingostomy,laparoscopic salpingectomy for ectopic pregnancy can also preserve postoperative ovarian antral follicle count(AFC),and it has greater advantage in reducing the risk of recurrent ectopic pregnancy after surgery.