Bowness J M
Can J Biochem. 1982 Aug;60(8):777-81. doi: 10.1139/o82-096.
Punch wounds (3 mm) were made in the skin of rats and the animals were killed after 1 or 3 days. Plugs (4 mm) of wounded and unwounded skin were incubated in vitro with [3H]fucose. The labelled plugs were homogenized and subjected to sequential extraction with buffered salt solutions, ethanol-ether, and 8 M urea - 50 mM dithiothreitol (DTT). Nondialysable counts in the extracts and insoluble residue were determined and the incorporation of label by wounded and unwounded skin plugs was compared. Wound plugs showed a greater total incorporation of [3H]fucose. In addition, a greater proportion of [3H]fucose was found in the urea-DTT extracts. The highest specific activity (disintegrations per minute [3H]fucose per milligram dry weight) was found in a finely dispersed precipitate, sedimenting at 10000 x g but not at 1000 x g. The transglutaminase inhibitors aminoacetonitrile and dansyl cadaverine were found to increase the extractability of a portion of the material which incorporated [3H]fucose without affecting the total incorporation. These results show that healing wounds have an increased biosynthetic capacity for an insoluble fucosylated glycoprotein fraction and they suggest that transglutaminase is necessary to make this fraction fully insoluble.
在大鼠皮肤上制造3毫米的冲孔伤口，1天或3天后处死动物。将受伤和未受伤皮肤的4毫米组织块在体外与[3H]岩藻糖一起孵育。将标记的组织块匀浆，并用缓冲盐溶液、乙醇-乙醚和8M尿素-50mM二硫苏糖醇（DTT）进行顺序提取。测定提取物和不溶性残渣中的非透析性计数，并比较受伤和未受伤皮肤组织块对标记物的掺入情况。伤口组织块显示出[3H]岩藻糖的总掺入量更高。此外，在尿素-DTT提取物中发现了更大比例的[3H]岩藻糖。在以10000×g离心沉淀但不以1000×g离心沉淀的细分散沉淀物中发现了最高的比活性（每毫克干重每分钟[3H]岩藻糖的衰变数）。发现转谷氨酰胺酶抑制剂氨基乙腈和丹磺酰尸胺可增加掺入[3H]岩藻糖的一部分物质的可提取性，而不影响总掺入量。这些结果表明，愈合伤口对不溶性岩藻糖基化糖蛋白部分具有增强的生物合成能力，并且表明转谷氨酰胺酶是使该部分完全不溶所必需的。
