Objective To investigate the localization and the morphological changes of manchette during mouse spermiogenesis.Methods Immunofluorescence staining with FITC and costaining with DAPI were used to demonstrate the cellular localization of the manchette at different stages during mouse spermiogenesis.The structural changes of the manchette were observed during the maturing of the spermatid.Results Immunofluorescence staining showed that manchette existed exactly around the nuclei of the spermatids.Manchette began to form,when the shape of the nucleus changed from spherical to slightly elongated.While the nucleus of the spermatids condensed and elongated at later stages,manchette moved gradually to the caudal position of the spermatids.At last,the manchette diminished as the spermatids became mature.During mouse spermiogenesis,manchette underwent a transition from a cap-like to a tubular configuration.ConclusionThe formation and diminishment of the manchette is in step with the condensation and elongation of the nucleus of the spermatid.Both the structural and positional changes of the manchette coincide with the changes of the nucleus.These results imply that manchette might play an important role in mouse spermiogenesis.

Objective To investigate the expression and distribution of Diacylglycerol Kinase α (DGKα) mRNA in human hepatocellular carcinoma (HCC),and to explore the function of DGKα in the metastasis of hepatocellular carcinoma.Methods Tissues from 30 cases of HCC and 5 normal liver tissues were collected immediately after surgical resection.Semi-quantitative RT-PCR and in situ hybridization were used to detect the expression levels and distribution of DGKα mRNA,respectively.Results Semi-quantitative RT-PCR showed that the expression level of DGKα mRNA in HCC (0.798±0.317) and normal tissues (0.908±0.425) was significantly higher than those in carcinoma adjacent tissue with cirrhosis (0.205±0.102,P<0.05).In situ hybridization demonstrated that the number of DGKα mRNA positive hepatocytes in HCC [(57.6±6.3)/mm2] and normal tissues [69.8±8.7)/mm2] was significantly higher than those in carcinoma adjacent tissue with cirrhosis [(26.3±4.9)/mm2,P<0.05]; DGKα mRNA was expressed in the cytoplasm of hepatocytes in HCC and carcinoma adjacent tissue with cirrhosis,and in the nuclear of hepatocytes in normal tissues.Conclusion The present study suggests that DGKa may play important roles in carcinogenesis and progressing of HCC.

Objective To study the inhibition effect of non custodial terpenes-3β-alcohol to experimentally in-duced autoimmune encephalomyelitis in guinea pigs. Methods Different doses (25 mg/kg, 50 mg/kg and 100 mg/kg) of non custodial terpenes-3β-alcohol were given to the experimentally induced autoimmune encephalomyelitis model of guinea pigs by gavage for 8 weeks. Plasma levels of CD4+/CD8+, IL-1, IL-2, IL-6, IL-10, neuropeptide Y (NPY), beta endorphin (β-EP) , transforming growth factor-β(TGF-β), matrix metalloproteinase (MMP-2), nitric oxide synthase (NOS) and leuko-cyte differentiation antigen CD3 were assessed. The brain neuron morphology changes was observed under light microscopy while its ultrastructure changes was observed under electron microscope. NOS expression in neurons was observed through immunofluoresce technology. Results Non custodialterpenes-3β-alcohol inhibited the increase of plasma CD4+/CD8+, IL-1, IL-2, IL-6, IL-10, MMP-2, CD3 and NPY while decrease of plasmaβ-EP, brain TGF-β. It also increase NOS expres-sion in neuronal cytoplasm and maintained neuron morphology. Conclusion Non custodial terpenes-3β-alcohol inhibit-ed the experimental autoimmune encephalomyelitis in guinea pig.

Objective:To ascertain whether the immune complexes (ICs) formed by Dengue virus 1 non-structure protein 1 (DENV1 NS1)and its IgG antibodies could mediate passive systemic anaphylaxis (PSA) and to explain the pathogenesis of Dengue hemorrhagic fever or Dengue shock syndrome (DHF/DSS).Methods:The monoclonal antibodies (mAbs) or mAb cocktails from 20 IgG mAbs of DENV1 NS1 prepared in this lab were screened to initiate PSA and passive cutaneous anaphylaxis (PCA) in mice.Meanwhile, the effects of GdCl3 and platelet activating factor ( PAF) antagonist CV-3988 on PSA induced by the NS1-IgG ICs were observed.Results:Two groups of monoclonal antibody cocktails with purified NS 1 were proved to be capable of provoking PCA and PSA in mice,whereas the other mAbs or mAb cocktails could be not .The murine PSA initiated by NS1-IgG(5D25+3B1) ICs could be sig-nificantly inhibited by in vivo treatment with GdCl3 or PAF antagonist CV-3988.Conclusion: The NS1-IgG ICs formed with DENV1 NS1 and IgG mAb cocktails can mediate PSA and PCA ,but not all of ICs formed by DENV 1 NS1 mAbs or mAb cocktails with DENV 1 NS1 can induce PSA ,indicating that it may be related to the special epitopes of DENV 1 NS1.The monocyte/macrophages and PAF may be as major effector cells and the major mediator for PSA induced by NS 1-IgG ICs,respectively.

Objective To compare the detection efficiency between multiplex RT-PCR method and liquichip technology for screening the viral etiological agents of diarrhea.Methods The development of the multiplex RT-PCR method.A total of 107 feces samples from patients who suffered from diarrhea and attended to Zhujiang Hospital of Southern University from September 2013 to February 2014 were collected and tested in parallel by both multiplex RT-PCR and xTAG Gastrointestinal Pathogen Panel ( xTAG GPP) for Adenovirus, Norovirus genogroupⅠandⅡ, as well as by both multiplex RT-PCR and monoplex RT-PCR for Astrovirus and Sapovirus.To evaluate the sensitivity and specificity of multiplex RT-PCR, xTAG GPP and monoplex RT-PCR were used as reference.Kappa coefficient test was used to evaluate the consistency among the methods.The detection limit and accuracy of multiplex RT-PCR were evaluated by detection of serial dilution of positive plasmids and products sequencing for the five viral agents.Results The multiplex RT-PCR showed high consistency with xTAG GPP and monoplex RT-PCR, in which Kappa value was 0.885 and 1.000 respectively( P=0.000 ).Compared to xTAG GPP, the sensitivity and specificity of the multiplex RT-PCR were at average of 80.8%( 21/26 ) and 100%( 295/295 ) respectively.The detection limit and accuracy of multiplex RT-PCR were 104 copies /μl-106 copies/μl.Conclusion The high consistency indicated that both the multiplex RT-PCR and xTAG GPP are useful as a special,sensitive, high throughput and rapid diagnostic tools for the detection of the major viral pathogens related to diarrhea in clinical laboratory.

Objective To construct a recombinant expression vector for expression of the function-al domains of dengue virus serotype 1 ( DENV1 ) envelope ( E ) protein in native soluble form. Methods The genes encoding the functional domains of DENV1-E protein (1-394 aa) were amplified with PCR and then cloned into the Psectag2B-Fc eukaryotic expression vector.The 293T cells were transfected with the recombinant vector by cationic lipid-based delivery.The cell clones expressing the fusion DENV1-E-Fc protein were screened out with 2 mg/ml of Zeocin.Immunofluorescence assay ( IFA) was performed to analyze the antigenicity and integrity of the fusion protein.The fusion proteins were purified from cell lysate with Protein-G and further identified by Western blot assay.Results The soluble form of fusion protein with a molecular weight of about 90×103 was obtained at a yield of about 25 μg per 1×107 cells.The results of IFA indicated that the fusion protein kept its integrity with right conformational epitopes.The fusion protein was successfully expressed with the advantage of good specificity as indicated by IFA and Western blot assay. Conclusion The recombinant fusion protein in soluble form was successfully expressed in eukaryotic ex-pression system, which paved the way for further investigation on the function of DENV1 E protein and its protective epitopes.