ENDOTHELIAL & KERATINOCYTE STEM CELLS IN WOUND HEALING

在过去的几年里，人们对干细胞作为潜在的治疗剂的兴趣激增，部分原因是人们认识到，干细胞存在于成人的无数组织中，而且这些成人躯体干细胞群体是多能性的。我们一直在研究两种干细胞群体，血源性干细胞和角质形成细胞干细胞。在目标I中，我们将通过确定内源性血源细胞对非糖尿病和糖尿病小鼠受伤和未受伤皮肤的贡献来检查血源细胞参与伤口愈合的程度。这将通过在造血嵌合体小鼠(其内源性造血系统已被可与宿主区分开的细胞取代的小鼠)的皮肤伤口上实现。创面将通过组织学、免疫组织化学和形态计量学进行评估。我们的假设是，糖尿病会减少血源性细胞对皮肤结构的正常贡献，特别是在伤口愈合期间。如果这一假设是正确的，那么针对这一目标的更多实验将开始勾勒出导致这种减少的机制。在这项提案的第二部分，我们将测试血源性干细胞或角质形成细胞干细胞是否可以用于治疗糖尿病小鼠模型的伤口愈合。为此，我们将测试血细胞亚群或表皮干细胞的添加是否会加速非糖尿病和糖尿病小鼠的伤口愈合。我们的假设是，加入外源性血源性或角质形成细胞干细胞可以加速伤口愈合，而且这种影响在糖尿病小鼠比非糖尿病小鼠更明显。我们还假设，血源性干细胞和角质形成干细胞群体都是多能的，并将有助于伤口中的各种细胞类型。DIL标记的白细胞，DIO标记的皮肤角质形成细胞干细胞，或两者都将被直接注射到非糖尿病和糖尿病裸鼠皮肤伤口结痂下的伤口中。在此后的不同时间，将采集创面并进行组织学和免疫组织化学检查，以评估创面愈合率。我们还将调查糖尿病是否影响这些标记细胞结合到不同的皮肤成分中。

In the past few years interest in stem cells as potential therapeutic agents has exploded, due in part to the recognition that stem cells are present in a myriad of tissues in the adult, and that these adult somatic stem cell populations are pluripotent. We have been studying two stem cell populations, blood-derived and keratinocyte stem cells. In Aim I we will examine the extent to which blood-derived cells are involved in wound healing by determining the contribution Of endogenous blood-derived cells to injured and uninjured skin in non-diabetic and diabetic mice. This will be accomplished by making skin wounds in hematopoietic chimeric mice (mice whose endogenous hematopoietic system has been replaced with cells that can be distinguished from those of the host). Wounds will be evaluated using histology, immunohistochemistry, and morphometry. Our hypothesis is that diabetes reduces the normal contribution of blood- derived cells to structures in the skin, especially during wound healing. If this hypothesis is correct, additional experiments in this aim will begin to delineate the mechanisms responsible for the reduction. In the second part of this proposal we will test whether blood-derived or keratinocyte stem cells can be used therapeutically to augment wound healing in a mouse model of diabetes. For this we will test whether addition of subsets of blood cells or epidermal stem cells accelerate wound healing in non-diabetic and diabetic mice. Our hypotheses are that addition of exogenous blood-derived or keratinocyte stem cells can accelerate wound healing, and that this effect will be more pronounced in diabetic than in non-diabetic mice. We also hypothesize that both blood-derived and keratinocyte stem cell populations are multipotent and will contribute to a variety of cell types in the wounds. Dil labeled leukocytes, DiO labeled skin keratinocyte stem cells, or both will be injected directly into the wound beneath the scabs of skin wounds in non-diabetic and diabetic nude mice. At various times thereafter, wounds will be harvested and examined histologically and immunohistochemically to assess the rate of wound healing. We will also investigate whether diabetes affects the incorporation of these labeled cells into different skin components.