Expression of lactate dehydrogenase, myosin heavy chain and myogenic regulatory factor genes in rabbit embryonic muscle cell cultures.

International audienceThe expression of myogenic regulatory factors (MRFs), lactate dehydrogenase (LDH) and myosin heavy chains (MyHC), as markers of myogenesis, metabolism and contractility respectively, were investigated during differentiation of rabbit embryonic muscle cells in primary culture. Myf5, MyoD and myogenin mRNAs were abundantly expressed at day 1 of culture. The expression of Myf5 and MyoD mRNA transcripts decreased sharply as myoblasts fused and differentiated into myotubes, whilst myogenin mRNA was maintained throughout the duration of the culture. In contrast, MRF4 mRNA was weakly expressed on day 1 of culture, its expression increased slightly as myoblasts fused and reached a maximum level in 7-day-old cultures containing striated myofibres. The specific activity of LDH increased linearly during myoblast proliferation and fusion. In 7-day-old cultures, LDH-M mRNA (dominant in glycolytic muscles) and LDH-H mRNA (predominant in perinatal and oxidative muscles) represented 38% and 62% of total LDH mRNA respectively. At this stage, immunocytochemical staining with perinatal and adult-type MyHC antibodies showed that embryonic and perinatal MyHC isoforms were expressed in all myotubes, while few of them were stained by type I MyHC antibody. However, none of them expressed adult type II MyHC. The latter results were further supported by RT-PCR analysis of adult-type MyHC mRNA which showed that only the type I MyHC mRNA transcript was expressed. These data were in agreement with those reported in vivo on perinatal rabbit muscles. They differed from those obtained on cultured satellite cells isolated from adult rabbit fast-twitch or slow-twitch muscles which did not express embryonic MyHC, and instead expressed fast- or slow-type MyHC according to their muscle origin. Taken together, these results further suggest that myogenic mononucleated cells express different properties in vitro according to their developmental origin as well as properties related to those of the muscles from which they were isolated

Real-Time PCR Quantification of Human Cytomegalovirus DNA in Amniotic Fluid Samples from Mothers with Primary Infection

A real-time PCR assay was developed to quantify human cytomegalovirus (HCMV) DNA in amniotic fluid (AF) samples collected from 30 pregnant women with primary HCMV infection as detected either from HCMV-immunoglobulin G (IgG) seroconversion or by the presence of HCMV-specific IgG and IgM associated with a low IgG avidity. Clinical information available for each case included ultrasonographic examination and fetal or newborn outcome. HCMV infection of fetuses or newborns was confirmed for the 30 studied cases. AF samples were subdivided into three groups. In group A (n = 13), fetuses presented major ultrasound abnormalities, and pregnancy was terminated. In group B (n = 13), fetuses had normal ultrasound findings, the pregnancy went to term, and the newborns were asymptomatic at birth. In group C (n = 4), fetuses had no or minor ultrasonographic signs, and pregnancy was terminated. The HCMV DNA load values in AF samples were significantly higher in group A (median, 2.8 × 10(5) genome equivalents [GE]/ml) than in group B (median, 8 × 10(3) GE/ml) (P = 0.014). Our findings suggest that HCMV load level in AF samples correlates with fetal clinical outcome but might also be dependent on other factors, such as the gestational age at the time of AF sampling and the time elapsed since maternal infection

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