OBJECTIVE To clone the MSI-78 gene for the purpose of providing evidence for further studies in prokaryotic expression and activities of antimicrobial peptides. METHODS According to the amino acid sequences of MSI-78,the MSI-78 gene was designed favorable for the Escherichia coli codons. After EcoRⅠand PstⅠ disgestion,cohesive ends were added to both ends respectively and the MSI-78 gene was synthesized by chemical methods. Then,the MSI-78 gene was ligated with pUC-18,transformed into the E. coli DH5?. Through filtration of ? complementary screening,the positive recombinant was finally identified by enzyme digestion of ECORⅠand ECORⅠ/PstⅠ and by PCR. RESULTS The MSI-78 gene was ligated with pUC-18 and transformed into the E. coli DH5?. As a result,MSI-78 gene was cloned in E. coli DH5? successfully. CONCLUSIONS The cloning of the MSI-78 gene provides evidence for further studies of its prokaryotic expression and activities of antimicrobial peptides.

Objective To investigate the effects of breviscapine on the renal structure, function and PKC-?mRNA and its protein expression in brain-dead BA-Ma mini pigs. Methods Fifteen BA-Ma mini pigs were randomly divided into 3 groups: brain-dead group (group A), breviscapine pretreatment group (group B), and control group (group C), 5 pigs in each group. The brain-dead models were established by increasing intracranial pressure in a modified, slow and intermittent way. At 3, 6, 12, 18 and 24 h after the initial brain death, serum BUN, Cr, TNF-?, IL-1?, and IL-6 were determined. At 3, 6, 12, and 24 h, the changes of renal tissues were observed by HE staining, and the expression of PKC-?mRNA and protein was detected by RT-PCR and immnohistochemistry respectively. The ultrastructural changes of hepatic cells were observed under electron microscopy. Results (1) At 3rd h after the initial brain death, IL-1?, IL-6, and TNF-?in group A and group B began to increase. Serum BUN and Cr in group A and group B began to increase at 12 th after brain death and were higher at each time point (P

BACKGROUND: The cel density is one of the factors involved in the state of cel differentiation, and the effect of cel density on transforming growth factor-β1-induced epithelial-mesenchymal transition of HaCaT cells is stil unclear. OBJECTIVE: To observe the effect of cel density on transforming growth factor-β1-induced epithelial-mesenchymal transition of HaCaT cells. METHODS: HaCaT cells was seeded in 6-wel plates at low density of 103/cm2 and high density of 105/cm2 then treated by 2 μg/L transforming growth factor-β1 for 48 hours, thereafter observed the changes in cel morphology. The transcription levels of epithelial cadherin, tight junction protein-1, vimentin, neuronal-cadherin were detected by real-time PCR, and expression levels of epithelial cadherin and vimentin were detected by Western blot. RESULTS AND CONCLUSION: Cel gap of HaCaT cells grew larger after treated with transforming growth factor-β1 for 48 hours, and cel morphology was long spindle rather than polygonal in low-density group, while in high-density group without obvious morphological changes. The real-time PCR showed that the transcriptions of epithelial cel marker epithelial cadherin and tight junction protein-1 were suppressed when compared with the control group (P < 0.05), and the decreasing deree in the high-density group was higher than that in the low-density group (P <0.05), mesenchymal cel marker neuronal-cadherin and vimentin were upregulated in the high-density group and the low-density group when compared with the control group (P < 0.05), and there were no significant differences between high-density group and low-density group. Western blot results verified the changes of neuronal-cadherin and vimentin expression level. These results suggest that the high seeding density can inhibit transforming growth factor-β1-induced epithelial-mesenchymal transition of HaCaT cells.

Objective To study the effect and mechanism of increase in radio sensitivity of kidney cancer cells(GRC-1) induced by ?-elemenen in vitro. Methods GRC-1 cells were divided into 3 groups, blank group (added with 2 mL culture medium), emulsion group (added with 2 mL blank emulsion culture medium) and drug group (added with 2 mL 50 mg?L -1 ?-elemenen culture medium). After been cultivated for 24 hours, the cells were irradiated using 6MeV X-linear accelerator in different doses at the rate of 400cGy per minute. Number of cell clones was counted, and radiation-survival curve of GRC-1 cells was drawn. Flow cytometry (FCM) was used to measure cell cycle and apoptosis. Cells of climbing flake were dyed by immunocytochemical method, the gene expression of bcl-2 and PCNA was measured by imaging system. Results The cell cycle showed that the G 2M blocking caused by 50 mg?L -1 ?-elemenen was enhanced with time increase. It reached peak at 24 hours. FCM showed that the level of apoptosis increased with increase in drug dose and action time. The gene expression of bcl-2 was decreased by 20% in drug group than that in blank group, but there was no expression of PCNA in the two groups. Conclusion The radiosensitivity of GRC-1 cells can be enhanced by ?-elemenen. The mechanism of effect may be associated with the cell cycle blocking, inducing cell apoptosis and down-regulating expression of bcl-2 gene.

Objective To investigate the metabolic changes in free fatty acids in the livers of obese rats with type 2 diabetes.Methods Thirty male SD rats were randomly divided into control group(Con group) and diabetes mellitus group(DM group),with 15 in each group.Rats in Con group and DM group were fed with normal diet and high-fat diet,respectively.Eight weeks later,OGTT and ISI test were performed to identify insulin resistance.Then the insulin-resistant rats received intraperitoneal injection of low-dose streptozocin(STZ) to induce type 2 diabetes.After giving high-fat diet further for six more weeks,8 rats of each group were sacrificed and artery blood and liver sample were obtained for further analysis.The mRNA levels of diacylglycerol O-acyltransferase 1(DGAT1),carnitine palmitoyltransferaseⅠ(CPTⅠ),palmitoyl-CoA oxidase(ACOX1),D-bifunctional protein(DBP) and L-bifunctional protein(LBP),which were involved in fatty acids metabolism,were evaluated by RT-PCR.The protein level of DBP was evaluated by Western blotting Peroxisome fatty acids ?-oxidation was measured by spectrophotometry.Free fatty acids in blood and liver were determined by gas chromatography.Oil red staining was used to determine the fat accumulation in liver.Results The mRNA expressions of DGAT1,CPTⅠ,ACOX1 and LBP increased(P

Animals ; Heart ; drug effects ; Hippophae ; Liver ; drug effects ; metabolism ; Male ; Myocardium ; metabolism ; Oxidative Stress ; Plant Oils ; pharmacology ; Rats ; Rats, Sprague-Dawley

Objective To evaluate the effect of 36-neuro-motorial examination on early diagnosis of children with cerebral palsy within 1 year-old.Methods36-neuro-motorial examination was analysed in 210 children with cerebral palsy from 2 to 12 months. Results4-29 items reflecting the abnormity of reflex, muscle tone, posture and motion were observeed in patients with cerebral palsy.If these abnormities were laid according as frequency among 2-6 months,reflex abnormities are knee tendon reflex, palmar grasp reflex,Babinski sign,asymmetrical tonic neck reflex,ankle clonus,sitting equilibrium reflex,stepping reflex,crossed extension reflex,tonic labyrinthine reflex,trunk incurvation reflex,Moro reflex; posture abnormities are head to back≥15°,from supine to the side position, axillar suspension reflex,posterior neck fovea≥1cm in the supine position,landau reflex,Vojta reflex,traction reflex,tonic torticollis,abnormal spontaneous posture;muscle tone abnormities are foot dorsiflexion angle,scarf sign,tonic palmar grasp,to up the foot heel≥30° in the stand position,thumb decussation to the palm,poples angle,adductor angle,heel-ear angle; others are head cycle≤-2s, squint, active movement attenuation or abnormity, auditory abnormity, visual abnormity, ophthalmodonesis, epilepsy.Conclusions 36-neuro-motorial examination is effective for early diagnosis of cerebral palsy within 1 year.

Objective To evaluate the effect of NGX6 combined with cisplatin on the inhibition rate of A549 cells and NCI-H1975 cells and antitumor effects in vivo.Methods The NGX6 was loaded in the LPD, and prepare Liposome protamine DNA complexes.A549 cells and NCI-H1975 cells were seperately divided into NGX6 group ( 30 μg/mL NGX6 concentration ) , cisplatin group, NGX6 +cisplatin group, PBS as negative control group.The effect of cytotoxicity of four group on A549 cells and NCI-H1975 cell were evaluated by MTT assay.the clone forming rate and the inhibition rate were determined by Cell colony count.lung transplantation tumor model were successfully established, then nude mice were divided into four groups as abeve, each group of 10, tumor size and survival period were determined tumor cell apoptosis were observed.Results The cell viability of A549 cells and NCI-H1975 cells of (NGX6 +cisplatin) were lower than that of NGX6 group, cisplatin group and saline group, respectively(P<0.01).The cloning efficiency of A549 cells and NCI-H1975 cells of ( NGX6 +cisplatin) were lower than that of NGX6 group, cisplatin group and saline group, respectively(P<0.01).The tumor inhibitory rate was in vivo for (NGX6 +cisplatin) was higher than other group(P<0.01).The median survival of nude mice in (NGX6 +cisplatin), NGX6, cisplatin and saline group were 43,31,29 and 15 days.Conclusion NGX6 combination with cisplatin can inhibit the cell proliferate of lung cancer cells and inhibit the tumor growth and the combination of NGX6 and cisplatin may be a potentially effective treatment for lung cancer.

Objective This study was designed to determine the in vitro sensitivity of LMWH caused by different reagents,and to explore whether the ACT can be used to monitor LMWH.Methods This study was performed in vitro.ACT was measured with different reagents(glass beads,celite,and kaolin)on volunteer(n =30)blood samples spiked with increasing concentrations of LMWH(datleparin,0.2-1.8IU/ml).Linear regression analysis was performed to establish a regression equation from different concentration of datleparin and corresponding ACT values.Results Analysis of dose-response curves obtained in vitro,an excellent linear relationship was observed between the ACT and dalteparin concentrations for all three reagents(p less than 0.01).Differences in slope of the regression curves of ACT were observed with all the reagents tested(glass beads 249.7s/IU,celite 77.7s/IU,and kaolin 59.3s/IU,p less than 0.01).Reagents vary widely in their in-vitro sensitivity related to dalteparin.In the concentration range of 0.2-1.8 IU/ml,the gaolin reagent was insensitive to dalteparin,and glass beads was the most suitable reagent for monitoring the anticoagulant effect of dalteparin.Conclusions Glass beads,celite,and kaolin.Glass beads were the most suitable reagent for monitoring the anticoagulant effect of dalteparin.Vary widely in their in-vitro sensitivity related to datleparin.

Objective To observe the effects of an enriched environment (EE) on cognitive functioning and the synaptic plasticity of mice modeling post-stroke cognitive impairment (PSCI) and explore the possible mechanisms involved.Methods Mice modeling PSCI and sham-operated mice were randomly divided into 3 groups:sham-operated mice in a standard environment (the Sham+SE group),PSCI mice in a standard environment (the PSCI+SE group) and PSCI mice in an enriched environment (the PSCI+EE group).The cognitive functioning of all of the mice was quantified using a Morris water maze and their hippocampal long-term potentiation (LTP) was recorded using an electrophysiological method.The level of synaptophysin was detected using Western blotting.Synaptic ultrastructure in the hippocampus was imaged using electron microscopy.Results Compared with the Sham +SE group,the PSCI+SE group showed significantly poorer water maze performance and failed induction of contralateral LTP.Their average level of synaptophysin was significantly lower,and significant adverse changes in the synaptic ultrastructure of the hippocampus were observed,including a decreased number of synapses.The average width of the synaptic cleft,postsynaptic density and the interface curvature of the synapses were all less desirable.All of the measurements of the PSCI+EE group improved significantly compared to those of the PSCI+SE group,but were still significantly poorer than those of the Sham+SE group.Conclusions An enhanced environment can improve the cognitive functioning of mice modelling PSCI.It may be that an EE can improve synaptic plasticity in the hippocampus contralateral to the stroke.