Department of Animal Science, Michigan State University, East Lansing, MI 48824-1225, USA.
J Dairy Sci. 2011 Dec;94(12):6036-46. doi: 10.3168/jds.2011-4590.
Mammary remodeling in dairy cows involves coordinated changes in stromal and epithelial tissue. Tissue remodeling is characterized by changes in cell proliferation, activation of fibroblasts into myofibroblasts, and changes in extracellular matrix content. Transforming growth factor β-1 (TGF-β1) increases differentiation of fibroblasts to myofibroblasts, regulates expression of extracellular matrix proteins and proteases, and has cell-type dependent effects on proliferation. The objective of this study was to determine whether TGF-β1 treatment of mammary tissue from cows in late lactation and the dry period affects cell proliferation, expression of matrix metalloproteinase-3 (MMP-3) and fibronectin (FN), and the differentiation of fibroblasts into myofibroblasts that express smooth muscle α actin (SMA). Tissue was biopsied from 7 Holstein cows at 4 time points: late lactation, 1 wk after dry-off, 3 wk before expected calving, and 1 wk before expected calving. Explants of biopsied tissue were incubated for 2h in Waymouth's medium containing insulin, hydrocortisone, and 0 or 5 ng of TGF-β1/mL; a subset of cultures was also incubated with bromodeoxyuridine to measure epithelial and stromal cell proliferation. Tissues were fixed, embedded in paraffin, sectioned, and stained by immunohistochemistry. Stage at biopsy had an overall effect on rate of epithelial and stromal cell proliferation, and TGF-β1 treatment increased rate of bromodeoxyuridine incorporation more than 2-fold in both cell types at 1 wk after dry-off. The number of fibroblasts expressing SMA was 19% higher in the intralobular stroma at 1 wk after dry-off compared with that at 1 wk before expected calving, and the percentage of activated fibroblasts tended to be higher in tissue incubated with TGF-β1. Biopsy stage had an overall effect on percentage area of epithelium expressing FN and MMP-3. Incubation with TGF-β1 had no effect on percentage intralobular stroma area expressing FN or MMP-3. Effects of TGF-β1 treatment were most apparent at 1 wk after dry-off, indicating that the first week of dry period may be an ideal target for testing effects in vivo.
奶牛的乳腺重塑涉及间质和上皮组织的协调变化。组织重塑的特征是细胞增殖、成纤维细胞向肌成纤维细胞的激活以及细胞外基质含量的变化。转化生长因子β-1（TGF-β1）增加成纤维细胞向肌成纤维细胞的分化，调节细胞外基质蛋白和蛋白酶的表达，并对增殖具有细胞类型依赖性效应。本研究的目的是确定在泌乳后期和干奶期对奶牛乳腺组织进行 TGF-β1 处理是否会影响细胞增殖、基质金属蛋白酶-3（MMP-3）和纤维连接蛋白（FN）的表达，以及分化成表达平滑肌α肌动蛋白（SMA）的成肌纤维细胞。从 7 头荷斯坦奶牛在 4 个时间点采集乳腺组织活检：泌乳后期、干奶后 1 周、预计产犊前 3 周和预计产犊前 1 周。将活检组织的外植体在含有胰岛素、氢化可的松和 0 或 5ng/ml TGF-β1 的 Waymouth 培养基中孵育 2 小时；部分培养物还与溴脱氧尿苷一起孵育，以测量上皮和基质细胞的增殖。组织固定、石蜡包埋、切片，并用免疫组织化学染色。活检时相总体上影响上皮和基质细胞的增殖速度，并且在干奶后 1 周，TGF-β1 处理使两种细胞类型的溴脱氧尿苷掺入率增加了两倍以上。与预计产犊前 1 周相比，干奶后 1 周小叶内间质中表达 SMA 的成纤维细胞数量增加了 19%，并且用 TGF-β1 孵育的组织中激活的成纤维细胞百分比也有增加的趋势。活检时相总体上影响上皮组织中 FN 和 MMP-3 的表达百分比区域。TGF-β1 孵育对小叶内间质中 FN 或 MMP-3 的表达百分比区域无影响。TGF-β1 处理的影响在干奶后 1 周最为明显，这表明干奶期的第一周可能是体内试验的理想目标。
