Lacour J P, Gordon P R, Eller M, Bhawan J, Gilchrest B A
USDA Human Nutrition Research Center on Aging, Tufts University, Boston, Massachusetts.
J Cell Physiol. 1992 May;151(2):287-99. doi: 10.1002/jcp.1041510210.
In contrast to neurite outgrowth, pigment cell dendrite formation is relatively unstudied. Keratinocyte-conditioned medium (KCM) induces a striking dendricity in human melanocytes and B16 melanoma cells that is detectable within 30 min, maximal in 24-48 hr, and quantifiable by computerized image analysis. Cytochalasin B (CB), known to disrupt actin microfilaments, completely blocks dendrite formation if added to cultures before or with KCM. This effect is rapidly reversible, and dendrites appear within 1 hr after refeeding with KCM alone. In contrast, CB treatment fails to disrupt existing dendrites previously induced by KCM. Agents known to cause microtubule disassembly (colchicine, nocodazole, or vinblastine) do not inhibit dendrite formation if added before or with KCM. In contrast, these agents disrupt established dendrites. Inhibition of protein synthesis with cycloheximide or actinomycin D completely blocks dendrite formation, but if cultures are provided fresh KCM lacking protein synthesis inhibitors, dendrites reappear within 24 hr. Actin microfilaments visualized with a monoclonal antibody or rhodamine-phalloidin are poorly organized in untreated cells, but form numerous fibers localized along dendrites in KCM-treated cells. Microtubules visualized with a monoclonal anti-tubulin antibody are localized in the center of dendrites. These cytoskeletal changes occur without altering beta actin or beta tubulin mRNA levels. Taken together, these data implicate actin microfilaments in dendrite outgrowth, but not in maintenance, and conversely microtubules in dendrite maintenance but not in formation. These keratinocyte-induced changes involving beta actin and beta tubulin polymerization appear to require both new protein synthesis and post-translational regulation. The observed similarities between melanocytes and other neural crest-derived cells suggest that cutaneous pigment cells might serve as an alternative model for studies of neurite outgrowth.
与神经突生长不同，色素细胞树突形成的研究相对较少。角质形成细胞条件培养基（KCM）可诱导人黑素细胞和B16黑色素瘤细胞形成显著的树突，30分钟内即可检测到，24 - 48小时达到最大值，并可通过计算机图像分析进行量化。已知细胞松弛素B（CB）会破坏肌动蛋白微丝，如果在培养物中与KCM同时添加或在添加KCM之前添加，会完全阻断树突形成。这种作用可迅速逆转，仅用KCM重新培养1小时后树突就会出现。相比之下，CB处理不会破坏先前由KCM诱导形成的现有树突。已知会导致微管解聚的药物（秋水仙碱、诺考达唑或长春碱）如果在添加KCM之前或同时添加，不会抑制树突形成。相反，这些药物会破坏已形成的树突。用环己酰亚胺或放线菌素D抑制蛋白质合成会完全阻断树突形成，但如果为培养物提供不含蛋白质合成抑制剂的新鲜KCM，树突会在24小时内重新出现。用单克隆抗体或罗丹明 - 鬼笔环肽可视化的肌动蛋白微丝在未处理的细胞中组织不良，但在KCM处理的细胞中会形成许多沿着树突定位的纤维。用单克隆抗微管蛋白抗体可视化的微管位于树突的中心。这些细胞骨架变化在不改变β - 肌动蛋白或β - 微管蛋白mRNA水平的情况下发生。综上所述，这些数据表明肌动蛋白微丝参与树突生长，但不参与维持，相反，微管参与树突维持，但不参与形成。这些由角质形成细胞诱导的涉及β - 肌动蛋白和β - 微管蛋白聚合的变化似乎既需要新的蛋白质合成，也需要翻译后调控。黑素细胞与其他神经嵴衍生细胞之间观察到的相似性表明，皮肤色素细胞可能作为神经突生长研究的替代模型。
