Garrett K L, Anderson J E
Department of Anatomy, University of Manitoba, Winnipeg, Canada.
Dev Biol. 1995 Jun;169(2):596-608. doi: 10.1006/dbio.1995.1172.
Tissue culture studies using muscle cell lines suggest that in addition to mitogenic effects, fibroblast growth factors (FGF) inhibit skeletal muscle differentiation and the expression of members of a family of muscle-specific regulatory genes including MyoD and myogenin. We examined the possible coexpression of bFGF and myogenin by tandem in situ hybridization (detecting mRNA) and immunocytochemistry studies (detecting protein) to determine whether myogenic cells in vivo endogenously produce bFGF. Mdx mouse muscle, which shows characteristic dystrophic damage and regeneration, demonstrated mononuclear cells containing myogenin and bFGF transcripts in similar regions of adjacent sections of focal degeneration and repair, particularly near recent segmental fiber damage. Using immunocytochemistry and in situ hybridization concurrently on the same sections, bFGF protein and myogenin mRNA were colocalized in both muscle precursors and new myotubes. The in vivo results were confirmed in vitro using primary explant cultures of mdx muscle. Approximately one-half of mononuclear cells in vivo were myogenic by the criterion of myogenin mRNA expression. Both myogenin and bFGF mRNAs were also colocalized with bFGF protein, indicating endogenous expression of bFGF in a subpopulation of myogenic cells. Small numbers of myogenic mononuclear cells were differentiated, as determined by the presence of developmental myosin heavy chain protein (DevMHC). These cells and new myotubes also colocalized myogenin, DevMHC, and bFGF. Since bFGF and myogenin are colocalized in mpec and myotubes in vivo and in vitro, endogenous expression of bFGF is not mutually exclusive of myogenic regulatory gene expression, either before or after differentiation of the skeletal muscle phenotype. Such features of coexpression suggest an important and complex role for bFGF in muscle regeneration in vivo.
利用肌肉细胞系进行的组织培养研究表明，成纤维细胞生长因子（FGF）除了具有促有丝分裂作用外，还能抑制骨骼肌分化以及包括MyoD和肌细胞生成素在内的肌肉特异性调节基因家族成员的表达。我们通过串联原位杂交（检测mRNA）和免疫细胞化学研究（检测蛋白质）来检测碱性成纤维细胞生长因子（bFGF）和肌细胞生成素的可能共表达，以确定体内的生肌细胞是否内源性产生bFGF。mdx小鼠肌肉表现出典型的营养不良性损伤和再生，在局灶性变性和修复的相邻切片的相似区域，尤其是在近期节段性纤维损伤附近，显示出含有肌细胞生成素和bFGF转录本的单核细胞。在同一切片上同时使用免疫细胞化学和原位杂交，bFGF蛋白和肌细胞生成素mRNA在肌肉前体细胞和新的肌管中均共定位。使用mdx肌肉的原代外植体培养在体外证实了体内结果。根据肌细胞生成素mRNA表达标准，体内约一半的单核细胞具有生肌能力。肌细胞生成素和bFGF mRNA也与bFGF蛋白共定位，表明bFGF在生肌细胞亚群中内源性表达。通过发育性肌球蛋白重链蛋白（DevMHC）的存在确定有少量分化的生肌单核细胞。这些细胞和新的肌管也共定位了肌细胞生成素、DevMHC和bFGF。由于bFGF和肌细胞生成素在体内和体外的肌卫星细胞（mpec）和肌管中共定位，bFGF的内源性表达在骨骼肌表型分化之前或之后都与生肌调节基因表达并非相互排斥。这种共表达特征表明bFGF在体内肌肉再生中具有重要而复杂的作用。
