Effect of muscle origin and phenotype on satellite cell muscle-specific gene expression.

Satellite cells from adult mouse tongue, diaphragm, vastus lateralis, rectus femoris, tibialis anterior, soleus, and extensor digitorum longus muscles were isolated, expanded, and differentiated under identical culture conditions. Proliferating myoblasts and differentiated myofiber cultures were analyzed via SDS-PAGE, immunochemical, and PCR methods for expression of myosin heavy chains (MyHC) and muscle creatine kinase (MCK) as indices of muscle fiber type. Contralateral muscles were harvested for simultaneous, parallel analysis utilizing these assays. The MyHC profile of differentiated primary satellite cells was equivalent across all cultures with MyHC(2A) and MyHC(1/slow) co-expressed in all myotube and myofiber structures. Trace amounts of MyHC(2B) and MyHC(neo) were detected in a few myofibers. MCK was expressed at a uniform, similar level among these cultures. In contrast, contralateral muscles expressed each muscle-specific indicator at levels correlated with the fiber-type distribution within each muscle. MM14 and C2C12 cells, mouse satellite cell lines, were expanded and compared to primary cell cultures. MM14 cells had a high differentiation index (>95%) and co-expressed MyHC(2A) and MyHC(1/slow) along with trace amounts of MyHC(neo) throughout myotube cultures. Myofibers obtained from C2C12 cells exhibited less differentiation (~75%) with MyHC(2A) as the dominant isoform. These data indicate that primary satellite cells from adult muscle form a uniform differentiated cell type regardless of the fiber-type, anatomic location, and embryonic origin of the donor muscles. MM14 cells expressed an adult MyHC isoform profile similar to primary satellite cells. The results suggest that satellite cells provide a uniform cell source for use in autologous transplantation studies and do not acquire a heritable fiber-type-specific phenotype from their host muscle.

p53, retinoblastoma gene product, and cyclin protein expression in human papillomavirus virus DNA-positive cervical intraepithelial neoplasia and invasive cancer.

Human papillomavirus interacts with cyclin protein and tumor suppressor genes, p53, and retinoblastoma gene (Rb). Expression of these gene products was examined in 69 formalin-fixed, paraffin-embedded cervical biopsies by immunohistochemistry utilizing antibodies against p53, Rb, and proliferating cell nuclear antigen (PCNA) and by human papillomavirus DNA in-situ hybridization assays. Samples selected for this study included 27 normal/reactive atypia cases that were all human papillomavirus DNA in-situ hybridization negative, 37 cervical intraepithelial neoplasia (CIN) lesions, and 5 invasive carcinomas. The CIN and invasive carcinoma cases were all human papillomavirus DNA in-situ hybridization positive. p53 protein expression was detected in approximately one-third of the reactive atypia and CIN lesions and in 60% of invasive cancers. Neither the amount or the location of p53 staining was correlated with the histologic diagnosis. Rb staining was more frequently found in the CIN/invasive carcinoma cases compared to the normal/reactive atypia samples (39/42 [93%] versus 21/27 [78%], respectively; P < 0.05 by chi 2. PCNA staining was detected in virtually all samples tested. However, the location of both PCNA and Rb staining differed when the normal/reactive atypia cases were compared to the CIN cases. Only 10% of the former group demonstrated Rb staining throughout the basal two-thirds layer or full thickness of the epithelium compared with 65% of the latter group (P < 0.001 by chi2). Likewise, PCNA staining of the basal two-thirds or full-thickness of the epithelium was found in only 58% of normal/reactive atypia cases, but in 97% of the CIN group (P < 0.001). Our results suggest that the location of Rb and PCNA staining is quite different between normal/reactive atypia cervical biopsies and CIN lesions.