ABSTRACT
Objective To observe the changes of mechanical pain thresholds and autophagy related proteins microtubule-associated protein 1 light chain 3 (LC3) and sequestosome 1 (SQSTM1 also known as p62) expression levels in the C57BL/6 mouse models of chronic prostatitis/ chronic pelvic pain syndrome (CP/CPPS),and provide animal experimental evidence for CP/CPPS pain and autophagy study.Methods 36 male C57BL/6 mice were randomly divided into three groups: the model group,control group and na(i)ve group.The CP/CPPS model was established by subcutaneous injection in the lower abdomen region with suspension liquid,containing protein extract of male SD rat prostate gland and complete Freund adjuvant.At 1month and 6 months after modeling,the mice were sacrificed and prostate tissues were harvested for histological examination using HE staining.Mechanical tactile hyperalgesia was measured with von Frey filaments.The autophagy-related proteins LC3 and p62 expression levels were detected by immunohistochemistry,respectively.The average IOD was measured by Image Pro Plus 6.0,and the statistical analysis was performed with GraphPad Prism 5 software.Results The histopathology showed the appearance of chronic prostatitis in the model group,representing hyperplasia and lymphocytic infiltration to a different degree and lasted for 6 months after modeling.Moreover,prostate intraepithelial neoplasia (PIN) appeared in the model group at 6 months after modeling,characterized by the disappearence of basement membrane and obvious nuclear abnormality,while the control and na(i)ve groups showed normal histology during the 1-6 months.Compared with the control and na(i)ve groups,the mechanical pain threshold in the model group was significantly decreased along with the time from (0.353±0.154) g at 0 week to (0.008±0.00) g at 22 weeks (P<0.05).The average IOD of LC3 and p62 expression in the model group was significantly increased with timing from [(2.767±0.464)%,(2.872±1.642)%] at 1month to [(13.501±1.900)%,(9.07±0.49)%] at 6 month,P<0.05.Conclusions A CP/CPPS model is successfully established in C57BL/6 mice.For the model group,the mechanical pain threshold is decreased and autophagy levels are increased gradually with time.These phenomena show that chronic inflammation microenvironment may promote pain and autophagy activity in the prostate,which is closely related with the occurrence and development of prostatic intraepithelial neoplasia.
ABSTRACT
OBJECTIVE:To study the anti-coagulation effect and mechanism of fibrinolytic enzyme SNFE in sipuculus nudus, and provide reference for further development of SNFE. METHODS:40 mice were randomly divided into blank control group(nor-mal saline),Xueshuantong group(positive control,15 mg/kg)and SNFE low-dose,high-dose group(15,30 mg/kg),10 in each group. After intravenous injection in tail,tail bleeding time (BT) and clotting time (CT) were respectively determined to investi-gate the anti-coagulation effect of SNFE. After taking blood in abdominal aorta of rats,test was divided into blank control group, positive control group and SNFE low-mass concentration,medium-mass concentration,high-mass concentration groups (0.25, 0.50,1.00 mg/mL). Prothrombin time(PT),re-calcium time(PRT)(using orokinase as positive drug,100000 U/mL),and max-mum platelet aggregation rate (PAG) in 5 min under adenosine diphosphate (ADP) inducer (using asprin as positive drug,0.50 mg/mL) were respectively determined,and anti-coagulation effect mechanism of SNFE was analyzed. RESULTS:Compared with blank control group,BT,CT of mice in each group were prolonged,with statistical significance in Xueshuantong group and SNFE high-dose group (P<0.05 or P<0.01). Plasma PT of rats in positive control group,SNFE medium-dose,high-dose groups and PRT in each administration group were significantly prolonged(P<0.05 or P<0.01);and PAG in administration group was signifi-cantly reduced(P<0.01). CONCLUSIONS:The fibrinolytic enzyme SNFE in sipuculus nudus can play its anti-coagulant effect by inhibiting the activity of coagulation factors in internal and external sources and ADP-induced platelet aggregation.
ABSTRACT
OBJECTIVE:To study the anti-coagulation effect and mechanism of fibrinolytic enzyme SNFE in sipuculus nudus, and provide reference for further development of SNFE. METHODS:40 mice were randomly divided into blank control group(nor-mal saline),Xueshuantong group(positive control,15 mg/kg)and SNFE low-dose,high-dose group(15,30 mg/kg),10 in each group. After intravenous injection in tail,tail bleeding time (BT) and clotting time (CT) were respectively determined to investi-gate the anti-coagulation effect of SNFE. After taking blood in abdominal aorta of rats,test was divided into blank control group, positive control group and SNFE low-mass concentration,medium-mass concentration,high-mass concentration groups (0.25, 0.50,1.00 mg/mL). Prothrombin time(PT),re-calcium time(PRT)(using orokinase as positive drug,100000 U/mL),and max-mum platelet aggregation rate (PAG) in 5 min under adenosine diphosphate (ADP) inducer (using asprin as positive drug,0.50 mg/mL) were respectively determined,and anti-coagulation effect mechanism of SNFE was analyzed. RESULTS:Compared with blank control group,BT,CT of mice in each group were prolonged,with statistical significance in Xueshuantong group and SNFE high-dose group (P<0.05 or P<0.01). Plasma PT of rats in positive control group,SNFE medium-dose,high-dose groups and PRT in each administration group were significantly prolonged(P<0.05 or P<0.01);and PAG in administration group was signifi-cantly reduced(P<0.01). CONCLUSIONS:The fibrinolytic enzyme SNFE in sipuculus nudus can play its anti-coagulant effect by inhibiting the activity of coagulation factors in internal and external sources and ADP-induced platelet aggregation.
ABSTRACT
The protein production system using a baculovirus Antheraea pernyi nucleopolyhedrovirus (AnpeNPV) as a gene expression vector and its host insect as a natural bioreactor was successful established and its excellent performance in the protein production has been demonstrated. In this paper, the system is used to produce recombinant human epidermal growth factor (rhEGF), which have been widely used in medical and cosmetic treatment. A recombinant AnpehEGF virus has been constructed by replacing the viral polyhedrin gene with the rhEGF gene, and then injected it to Samia cynthia ricini pupae. Amplification and expression of rhEGF gene in the pupae was clearly detected by PCR, Western blot and ELISA analyses. These analyses have also revealed that rhEGF in the pupae was significantly increased at 6 days post-infection, and reached maximum level at the 12th day. The concentrations of rhEGF were 19.77, 24.90, 618.59 and 1 952.46 ng/g pupae at 3, 6, 9 and 12 days post-infection, respectively. However, the rhEGF concentration reduced at later stage (days 15). The rhEGF in the pupae could be purified using ammonium sulfate precipitation and Ni-NTA agrose affinity chromatography. Results demonstrate that Samia cynthia ricini pupae can be used as a bioreactor to produce rhEGF and, if successfully improved, will be a novel method of rhEGF production with lower cost and more efficient.
Subject(s)
Animals , Humans , Amino Acid Sequence , Bombyx , Genetics , Metabolism , Epidermal Growth Factor , Genetics , Genetic Vectors , Genetics , Molecular Sequence Data , Nucleopolyhedroviruses , Genetics , Recombinant Proteins , GeneticsABSTRACT
AIM To study the thrombolytic effect of non hemorrhagic fibrinolytic enzyme from Agkistrodon acutus venom on animal thrombosis. METHODS By means of pulmonary embolism,arterial thrombosis model in rabbits and venous thrombosis model in rats. The non hemorrhagic fibrinolytic enzyme from Agkistrodon acutus was given by intravenous injection at 0 14, 0 7, 1.4 mg?kg -1 three different doses respectively. Normal saline was used as negative control and thrombolytic effects was observed. RESULTS It was showed that moist weights of arterial thrombosis, venous thrombosis and pulmonary embolism were markedly lightened and has statistical significance compared with normal saline ( P
ABSTRACT
Aim To investigate the inhibitory effect of peptide from Agkistrodon Acutus snake venom(fraction K)on the growth of human ovarian carcimona A2780 cell line and its mechanism.Methods The A2780 cells were treated with fraction K at different concentrations.The proliferation of A2780 cells was assayed with MTT colorimetric method.The histologi-cal and ultras-tructural changes of cell were scored using light and electron microseopy.The apoptosis of A2780 cells induced by fraction K was studied by fluorescent staining.Crystal violet staining and MTT assay were employed to determine the effect of fraction K on A2780 cells that they had been adhered to fibronectin.Result Proliferation of A2780 cells was inhibited significantly by fraction K in a dose-and time-dependant manner.Under electron-microseop and fluorescence microscope,some A2780 cells underwent a typical apoptosis and morphology changes after 24 hours incubation with fraction K.Fraction K could inhibit the adhesion of A2780 cells to fibronectin in a dose-dependent manner.Conclusion fraction K can inhibit proliferation of A2780 cells,which may be related to the mechanism of inducing cellular apoptosis and cellular adhesion.
ABSTRACT
Aim To purify a small platide inhibiting platelet aggregation from Agkistrodon Acutus snake venom,and to identify its physical and chemical properties and its effects on the platelet aggregation stimulated by ADP, collagen, and thrombin. Methods Extract snake venom through Superdex 75 gel filtration, ultrafiltration and DEAE-Sepharose CL-6B ionexchange. The purified product was identified by HPLC C_ 18 . The molecular weight was determined by SDS-polyacrylamid gel electrphoresis. Platelet aggragation was measured by nephelometry. Results The molecular weight of the peptide was 7 862 u and its purified from Agkistrodon Acutus snake venom is oelectric point was pH 4.29. This peptide dose-dependently inhibited the platelet aggregation induced by ADP,collagen,and thrombin. Conclusion The method has been proved to be successful for the purification of low molecular weight peptide fraction that inhibits platelet aggregation.
ABSTRACT
AIM To study the inhibitory effect of the cytotoxin (CTX) from guangxi cobra venom on human nasopharyngeal cancer cell (CNE) and other tumons cells. METHODS The cytotoxin was isolated and purified from Guangxi cobra venom by successive chromatography on Sephadex G-50 and CM Sepharose CL-6B columns. Its cytotoxic effects and does-effect relationship on human tumors cell lines were examined by MTT assay. RESULTS The inhibitory effects of CTX CM-5 on CNE, human ovarian carcinoma cell (Ho8990), uterus cervical carcinoma cell (HELA) and lymphoma (YAC) cell lines showed a definite does-effect relationship. The IC 50 (48 h in cubation) was 1.84, 2.59, 1.84 and 0.75 mg?L -1 respctively. The inhibitory effects of CTX CM-5 on CNE increased with time. The IC 50 (3 h and 24 h cubation) was 4.78 and 1.04 mg?L -1 respctively. CONCLUSION CTX from Guangxi cobra venom exhibites strong suppressive effect on cultured tumor cells line in vitro.