Isolation of Nanjing local strains of HHV-7 and their biological and immunological characteristics.

To investigate the biological and immunological characteristics of the Nanjing local strains of HHV-7, four strains of herpesvirus were isolated from saliva specimens of one healthy individual and three children suffering from a kidney disease in Nanjing. The viruses were identified by transmission electron microscopy (EM), indirect immunofluorescence assay (IFA) with a specific monoclonal antibody; nested polymerase chain reaction, restriction mapping and DNA sequencing. The virus-infected cells showed the typical cytophathic effect (CPE) under microscopy and could be detected by IFA with the human herpesvirus-7 (HHV-7) specific antibody. Under EM, herpesvirus-like and virions capsids could be found in their cytoplasm or nucleoplasm. HHV-7 DNA fragments amplified from infected cells by nested PCR were confirmed by restriction mapping and DNA sequencing. Similarly to DC strain, an known HHV-7 strain used in the present study as the positive control, the virus could be inactivated by ultraviolet irradiation for 10 min, heated at 45 centigrade degree for 30 min, pH<5 or>9 at 4 centigrade degree for 2 h and ether or chloroform for 10 h. The virus induced the production of TNF-alpha, IL-10 and IL-12 p 70 while inhibited IFN-alpha secretion, increased the percentage of CD2(+) cells while decreased that of CD4(+) or CD 45 RA(+) cells. The results indicate that the viruses isolated in Nanjing are HHV-7, which has similar biological characteristic to the known HHV-7 strain, DC. Infection with HHV-7 in vitro could affect immune function of lymphocytes by disturbing cytokine production and CD antigen expression.

Rotating microgravity-bioreactor cultivation enhances the hepatic differentiation of mouse embryonic stem cells on biodegradable polymer scaffolds.

Embryonic stem (ES) cells are pluripotent cells that are capable of differentiating all the somatic cell lineages, including those in the liver tissue. We describe the generation of functional hepatic-like cells from mouse ES (mES) cells using a biodegradable polymer scaffold and a rotating bioreactor that allows simulated microgravity. Cells derived from ES cells cultured in the three-dimensional (3D) culture system with exogenous growth factors and hormones can differentiate into hepatic-like cells with morphologic characteristics of typical mature hepatocytes. Reverse-transcription polymerase chain-reaction testing, Western blot testing, immunostaining, and flow cytometric analysis show that these cells express hepatic-specific genes and proteins during differentiation. Differentiated cells on scaffolds further exhibit morphologic traits and biomarkers characteristic of liver cells, including albumin production, cytochrome P450 activity, and low-density lipoprotein uptake. When these stem cell-bearing scaffolds are transplanted into severe combined immunodeficient mice, the 3D constructs remained viable, undergoing further differentiation and maturation of hepatic-like cells in vivo. In conclusion, the growth and differentiation of ES cells in a biodegradable polymer scaffold and a rotating microgravity bioreactor can yield functional and organizational hepatocytes useful for research involving bioartificial liver and engineered liver tissue.