Modulation of vocal fold scar fibroblast by adipose derived stem cell

从两周前电烙的雪貂声带中分离瘢痕成纤维细胞（SF）（N=2）。从雪貂脂肪抽吸的皮下腹部脂肪（N=2）分离脂肪来源的干细胞（ASC）。对于共培养实验，将两种细胞类型在Transwell板中组合6天，然后在移除上室后进行1天或3天的单培养。然后对来自底室的细胞和培养基进行测定。我们通过酶联免疫吸附试验（ELISA）测定（1）透明质酸（HA）、胶原和肝细胞生长因子（HGF）的产生，（2）α-平滑肌肌动蛋白（α-SMA）的表达，（3）SF的细胞增殖和（4）SF的凋亡（通过流式细胞术测定2、3和4）。其他实验检查了HGF对SF的影响以及共培养系统中HGF中和的影响。我们证明，共培养导致SF胶原蛋白产生（p<0.05）、增殖（p<0.05）和α-SMA表达（p<0.05）显著降低，而HA产生增加（p<0.05）。共培养也增加了ASC的HGF分泌（p<0.05）。HGF的中和消除了ASC对SF胶原合成的抑制作用（p<0.05）。我们的结论是，ASCs影响SFs采取较少的纤维化配置文件。看来，HGF是至少一个负责这种效果的可溶性因子。植入的ASCs可能通过作为HGF的长期内在来源而改善声带瘢痕

Scar fibroblasts (SFs) were isolated from ferret vocal folds electrocauterized two weeks previously (N=2). Adipose-derived stem cells (ASCs) were isolated from ferret lipoaspirated subcutaneous abdominal fat (N=2). For co-culture experiments, the two cell types were combined in Transwell plates for 6 days, followed by 1 or 3 days of monoculture after removing the upper chamber. Assays were then performed on cells and media from the bottom chamber. We measured (1) the production of hyaluronic acid (HA), collagen and hepatocite growth factor (HGF) via enzyme-linked immunosorbent assays (ELISA) (2) the expression of α-smooth muscle actin (α-SMA), (3) cell proliferation and (4) apoptosis of SFs (2, 3 & 4 via flow cytometry). Other experiments examined the effects of HGF on SFs and the effects of HGF neutralization in the co-culture system. We demonstrated that co-culture led to significant decreases in SF collagen production (p<.05), proliferation (p<.05), and α-SMA expression (p<.05), while HA production increased (p<.05). Co-culture also increased HGF secretion from ASCs (p<.05). Neutralization of HGF abolished the inhibitory effects of ASCs on SF collagen synthesis (p<.05). We concluded that ASCs influence SFs to adopt a less fibrotic profile. It appears that HGF is at least one of the soluble factors responsible for this effect. Implanted ASCs could potentially ameliorate vocal fold scar by acting as a long-term, intrinsic source of HGF