Objective To construct recombinant full length genotype B and C hepatitis B virus (HBV)and to examine HBV DNA replication and hepatitis B surface antigen(HBsAg),hepatitis B e antigen(HBeAg)expressions in Huh7 cells. Methods The full length genotype B and C HBV DNA were extracted and amplified from two HBV infected patients. The recombinant plasmids were constructed by inserting the amplified HBV fragments into the eukaryotic expression vector,pHY106,which were then transfected into Huh7 cells. The cells transfected with blank pHY106 vector were used as control. HBV DNA replication at 72 hours of transfection was detected by Southern blot. The HBV DNA levels in Huh7 cells at 24,48,72,96 and 120 hours of transfection were determined by real-time polymerase chain reaction(PCR).Meanwhile, the HBsAg and HBeAg expression levels in the supernatants at 24,48,72,96 and 120 hours were determined by enzyme linked immunosorbent assay(ELISA).Results The recombinant plasmids expressing genotype B or C HBV DNA were successfully constructed.The HBV replicative intermediates in HBV core particles,including rcDNA dsDNA and ssDNA,were detected by Southern blot.HBV DNA level could reach 8 lg copy/mL which was by real-time PCR. HBsAg and HBeAg levels determined by ELISA peaked at 72 hours after transfection and then declined gradually. Conclusions The recombinant plasmids inserted with genotype B or C HBV DNA are constructed successfully, which can express high levels of HBsAg and HBeAg in Huh7 cells. This system provides a platform for studying the pathogenesis of B and C genotype HBV, the interaction between HBV and host, as well as exploiting new drugs against HBV.

Streptomyces sp. A048 was cultured in a complete medium to the last stage of log phase,the hyphae were washed and collected by centrifugation. Then the hyphae were inoculated in liquid medium for chitinase production using two-step fermentation. Activity of chitinase produced by two-step fermentation was 1.1 times higher than that from one-step fermentation,and ferment cycle was for 54 hours,which was 66 hours shorter than that of one-step fermentation. The hyphae and the powder of chitin were co-immobilizated and cultured in liquid medium for 36 hours,activity of chitinase was 1.8 times higher than that from one-step fermentation,and ferment cycle was 54h shorter than that of one-step fermentation. By adding 0.4% cellulose to two-step fermentation,activity of chitinase was 18.52 U/mL that was 4 times higher than that from the control and 10 times higher than that from one-step fermentation. Two step fermentation with chitin and cellulose may be the optimal fermentation process to produce Chitinase from Streptomyces sp. A048.

OBJECTIVETo observe the anti-inflammatory effect of total saponins of Panax japonicus on LPS-induced RAW264. 7 macrophages.

METHODThe effect of total saponins of P. japonicus of different concentrations on RAW264. 7 cell viability was determined with the MTT method. The NO kit assay was adopted to detect the NO release of total saponins of P. japonicus to LPS-induced RAW264. 7 cells. The enzyme linked immunosorbent assay (ELISA) was used to detect the secretion of tumor necrosis factor-alpha (TNF-alpha) and interleukin 1-beta (IL-1beta). The reverse transeriptase-polymerase chain reaction (RT-PCR) was used to determine the expression of inducible nitric oxide synthase (iNOS) ,TNF-alpha,IL-1beta. The protein expression of nuclear transcription factor-kappaB p65 (NF-kappaB p65) was tested by Western blot.

RESULTThe safe medication range of total saponins of P. japonicus was less than 80 mg x L(-1). Compared with the LPS model group, total saponins of P. japonicus high, middle and low dose groups (0.1, 1, 10, 40 mg x L(-1)) could significantly reduce the secretion of NO, TNF-alpha, IL-1beta of LPS-induced RAW264. 7 cells, and inhibit the expressions of iNOS, TNF-alpha and IL-1beta mRNA and the protein expression of NF-kappaB p65.

CONCLUSIONThis study preliminarily proves the protective effect of total saponins of P. japonicus on LPS-induced RAW264.7 macrophages. Its action mechanism may be related to NF-kappaB signal pathway.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Inflammation ; drug therapy ; genetics ; immunology ; Interleukin-1beta ; genetics ; immunology ; Lipopolysaccharides ; adverse effects ; Macrophages ; drug effects ; immunology ; Mice ; NF-kappa B ; genetics ; immunology ; Nitric Oxide ; immunology ; Nitric Oxide Synthase Type II ; genetics ; immunology ; Panax ; chemistry ; Protective Agents ; pharmacology ; Saponins ; pharmacology

Objective:To assess the applicability of the symptom inventory scale of DSM-Ⅳ ADHD in 112 Chinese children.Methods:Samples from out patients were diagnosed with the scale and CCMD-3.Results:The tester-consistency reliability Kappa value of the symptom inventory scale is 0.89,P

Heterokaryon incompatibility is a widespread phenomenon among fungi,controlled by specific loci termed het (for heterokaryon incompatibility).This review focuses on recent developments in our understanding of the molecular mechanisms of nonself recognition and the relationship between the death progresses of heterokaryon incompatibility and associated proteins in fungi.The deep research of heterokaryon incompatibility mechanism will hopefully reveal underlying principles of the evolution of nonself recognition systems and will find some effective method for settling the instability of protoplast fusant of fungi.

Objective To discuss the pathogenesis,diagnosis and treatment of delayed traumatic intracerebral hematoma.Methods The clinic data of traumatic delayed intracranial hematoma patients in this hospital were retro- spectively analyzed.According to clinic observation and CT re-examination,47 cases were diagnosed as delayed trau- matic intracranial hematoma(45 cases by operative treatment,and the other 2 by conservative treatment).Results There were 21 cases of recovery,10 cases of slight disability,8 cases of severe disability,8 cases of death.The total mortality rate was 17 %.Conclusion Brain contusion,subarachnoid hemorrhage and skull base fracture were impor- tant factors of DTICH.Fine-observation and prompt CT re-examination offered excellent results for DTICH.

OBJECTIVE: To explore the treatment and possible prognostic factor of nasal sinuses adenoid cystic carcinoma. METHOD: Retrospectively analysed the records of 18 patients with complete clinical and pathological data,which including 4 patients given up treatment, 5 patients taken surgical treatment and 9 patients taken surgical treatment as well as radiotherapy and chemotherapy. RESULT: Fifty percent of the patients got 2-year survival and 3 cases of death due to intracranial tumor invasion and 2 patients died of the disease distant metastasis. CONCLUSION If patients got Nasal sinuses adenoid cystic carcinoma, they should take comprehensive treatment based on surgery, in order to improve the survival rate. The prognosis depends on the tumor early detection and early treatment, the sooner the treatment, the better.
Adult ; Aged ; Carcinoma, Adenoid Cystic ; therapy ; Female ; Humans ; Male ; Middle Aged ; Nasal Cavity ; pathology ; Paranasal Sinus Neoplasms ; therapy ; Prognosis ; Retrospective Studies

Objective To express nonstructural protein 2 transactivated protein (NS2TP) of hepatitis C virus (HCV) in the prokaryotic expression system and prepare polyclonal antibody,and to study the expressions in different liver tissues.Methods NS2TP gene was amplified by polymerase chain reaction (PCR) technique and cloned into the prokaryotic expression vector pET-32a(+),which was transformed into E.coli BL21.The protein was induced by isopropyl thiogalactose (IPTG) and analyzed with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and confirmed by Western blotting assay.The recombinant protein were expressed and purified in a large amount.The rabbit was immunized with the purified protein to prepare polyelonal antibody.The liver tissues of patients with chronic HCV infection and healthy controls were detected by immunohistochemistry method.Results The recombinant NS2TP protein (relative molecular mass:33×103 ) and polyclonal antibody with high titer and specificity were successfully prepared.NS2TP expressions in the liver of patients with chronic HCV infection were higher than those of healthy controls,and were mainly distributed in the nucleus of hepatocytes.Conclusions The NS2TP expression level and intracellular location in liver tissue of patients with chronic HCV infection are understanded,which could bring new clues for further study of the biological function of NS2TP and the pathogenesis of HCV infection.

To investigate the effects of rennin angiotensin system blockade on the microvessel density in islets of diabetic rats and its relationship with islet function, diabetes model was created by feeding of high-caloric laboratory chow plus intraperitoneal injection of a small dose of streptozotocin (30 mg/kg). After 8 weeks intervention with perindopril (AE, n=10) or valsartan (AR, n=10), the islet function of the animals was evaluated by intravenous insulin release test (IVIRT). The pancreases were immunohistochemically stained to analyze the content of insulin and vascular endothelial growth factor (VEGF) in the islets. The microvessel density (MVD) of islets was detected by counting CD34 positive cells. The hypoxia inducible factor (HIF)-1alpha mRNA expression in the islets was detected by RT-PCR. Compared with normal control group (NC, n=10), the area under the curve for insulin from 0 to 30 min (AUCI(0-30)) of diabetes group (DM, n=8) was decreased by 66.3%; the insulin relative concentration (IRC) of betacell was decreased significantly; the relative content of VEGF was increased obviously [(-4.21+/-0.13) vs (-4.06+/-0.29)]; MVD in islets was decreased by 71.4%; the relative expression of HIF-1alpha mRNA was increased by 1.19 times (all P<0.01). Compared with DM group, the AUCI(0-30) of AE and AR group was increased by 44.6% and 34.9% respectively; IRC was also increased significantly; the relative content of VEGF was decreased by 21.2% and 21.7% respectively; MVD was increased by 62.5% and 75.0% respectively; the relative expression of HIF-1alpha was decreased by 27.2% and 29.0% respectively (all P<0.01 or P<0.05). There were no significant differences in the said indexes between group AE and AR. It is concluded that the blockade of RAS may ameliorate islets function of diabetic rats by increasing the MVD in islets.

Rheumatoid arthritis (RA), as a common systemic inflammatory autoimmune disease, affects approximately 1 in 100 individuals. Effective treatment for RA is not yet available because current research does not have a clear understanding of the etiology and pathogenesis of RA. Xinfeng Capsule, a patent Chinese herbal medicine, has been used in the treatment of RA in recent years. Despite its reported clinical efficacy, there are no large-sample, multicenter, randomized trials that support the use of Xinfeng Capsule for RA. Therefore, we designed a randomized, double-blind, multicenter, placebo-controlled trial to assess the efficacy and safety of Xinfeng Capsule in the treatment of RA.