CAPACITATIVE CALCIUM ENTRY AND STORES IN KERATINOCYTES

角质细胞中钙的电容性进入和储存

• 批准号: 6375243
• 负责人: Kenneth T Izutsu
• 金额: $ 25.8万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-08-01 至 2003-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6375243
• 关键词: Bacteroides gingivalis; bacteria infection mechanism; biological signal transduction; calcium channel; calcium flux; calcium indicator; cell differentiation; cell growth regulation; cytoplasm; electrophysiology; evoked potentials; gastrointestinal epithelium; gene expression; gingiva; human tissue; intracellular; keratin; keratinocyte; tissue /cell culture; voltage /patch clamp

DESCRIPTION (Adapted from the Investigator's Abstract): Intracellular calcium concentration ([Ca2+]i) is an important regulator of keratinocyte proliferation and differentiation. Hence, changes in [Ca2+]i play an important role in keratinocyte functions involved with tissue differentiation, wound healing, cancer development and bacterial invasion. The Principal Investigator found recently that nuclear [Ca2+] ([Ca2+]n) changes occur during bacterial invasion of keratinocytes. These [Ca2+]n changes were also dependent on Ca2+ influx. Moreover, the magnitude of the store depletion induced capacitative current was found to depend on the differentiation status of the keratinocyte. Since [Ca2+]n changes are important regulators of gene expression in all cells, such changes are likely to play an important role in the host response to bacterial invasion. However, before this invasion process can be understood, the mechanisms by which [Ca2+]n is regulated must be elucidated. In addition, it is necessary to elucidate why the capacitative current varies with keratinocyte differentiation state. The present proposal initiates efforts to understand these [Ca2+] regulatory mechanisms. Hence, this proposal has the following Specific Aims: Specific Aim 1 proposes to determine how intracellular Ca2+ stores affect nuclear [Ca2+]. Specific Aim 2 proposes to characterize the Ca2+ permeable channels activated by Ca2+ store depletion, and to determine whether the resulting Ca2+ influx alters nuclear [Ca2+]. Specific Aim 3 will demonstrate that the nuclear Ca2+ signaling system is altered in Ca2+-activated, differentiated keratinocytes as compared to no differentiated keratinocytes. [Ca2+]i will be measured with wide-field, deconvolution microscopy using fura-2; Ca2+ channel properties with whole-cell and single channel, patch clamp techniques. Keratinocyte differentiation will be induced by exposure to Ca2+ in culture, and monitored by induction of involucrin and cytokeratin 13. Completion of the proposed Specific Aims will establish the characteristics of the capacitatively activated Ca current and the Ca stores that affect [Ca2+]n in differentiated and undifferentiated keratinocytes, and their relationships to the [Ca2+]n changes that can affect gene expression.

描述（改编自研究者摘要）：细胞内钙 [Ca 2 +]i是角质形成细胞增殖的重要调节因子 和差异化。因此，[Ca 2 +]i的变化在其中发挥着重要作用 涉及组织分化，伤口愈合， 癌症发展和细菌入侵。首席研究员发现 最近发现，在细菌入侵过程中，细胞核[Ca 2 +]（[Ca 2 +]n）发生变化， 角质形成细胞。这些[Ca ~（2+）]n变化也依赖于Ca ~（2+）内流。 此外，存储耗尽引起的电容电流的大小为 发现取决于角质形成细胞的分化状态。以来 [Ca2+]n变化是所有细胞中基因表达的重要调节因子，例如 变化可能在宿主对细菌的反应中起重要作用。 入侵然而，在了解这一入侵过程之前， 必须阐明[Ca 2 +]n调节的机制。此外还 有必要阐明为什么电容电流随角质形成细胞 分化状态本提案旨在努力了解 这些[Ca 2 +]调节机制。因此，这项建议有以下几点 具体目标：具体目标1提出确定细胞内Ca 2 + 钙库影响细胞核[Ca 2 +]。具体目标2建议表征Ca 2 + Ca 2+储存耗尽激活的渗透性通道，并确定是否 由此产生的Ca 2+内流改变核[Ca 2 +]。第3章将 表明细胞核Ca 2+信号系统被改变， Ca 2+激活的分化角质形成细胞与未分化角质形成细胞相比 角质形成细胞[Ca2+]i将用宽场反卷积测量 使用fura-2的显微镜;全细胞和单个细胞的Ca 2+通道特性 通道，膜片钳技术。诱导角质形成细胞分化 通过暴露于培养物中的Ca 2+，并通过外皮蛋白的诱导和 细胞角蛋白13。完成拟议的具体目标将建立 电容激活钙电流和钙库特性 影响分化和未分化角质形成细胞中的[Ca 2 +]n， 它们与[Ca 2 +]n变化的关系可以影响基因表达。