REGULATION OF OUTFLOW FACILITY BY GENE TRANSFER

描述：判断蛋白质贡献的一种有效方法 对细胞功能的调节是通过将其引入到 细胞核酸，相同的(正义的)或互补的序列 (反义)到编码目标蛋白的mRNAs序列。 PI假说是细胞骨架相关基因表达 房水流出通道的细胞动力学可影响房水流出 设施；对编码的蛋白质浓度的操纵 这些基因将改变流出设施；复制缺陷 腺病毒载体将成功地将基因输送到流出途径 细胞；以及对这种药物的有效性和毒性的严格评估 递送系统将促进眼基因的潜在开发 青光眼的治疗。 由于在体内和体内观察到的流出设施的增加 体外用微管解聚药物治疗后，我们希望 初步研究B-微管蛋白、微管相关蛋白和 转录因子NFkB。PI将表征哪些异构体来自 这些基因家族在小梁网中表达。她会的 使用腺病毒转移技术过度表达和低表达其 器官培养中的特定区域灌流了猪和人的眼睛。她会的 测量流出设施。 初步结果表明，复制缺陷型腺病毒 可能是理想的载体来研究我们的假设。他们进入的细胞 小梁网状结构非常有效地工作，但本身并不 干扰细胞骨架网络，也不影响细胞外流速度。 即使这些最初的操作不会改变房水流出，这 调查仍应有助于对蜂窝电话的宝贵见解 房水流出的机制。此外，完整的 分子生物学与生理学相结合的系统研制 测量，将允许测试未来的多个假设，包括 蛋白质/酶(受体、生长因子、离子通道、激动酶...)那 已经被概念化为房水生理学中的重要部分。

DESCRIPTION: One effective means of judging the contribution of a protein to cellular function is to modulate its expression by introducing into the cell nucleic acid, sequences that are identical (sense) or complementary (antisense) to the mRNA sequence that encodes the target protein. The PI hypothesis is that expression of genes involved in cytoskeleton dynamics in cells of the outflow pathway can influence aqueous humor outflow facility; that manipulation of the concentration of the proteins coded by these genes will alter outflow facility; that replication deficient adenovirus vectors will successfully deliver genes to the outflow pathway cells; and that the rigorous evaluation of the efficacy and toxicity of this delivery system will advance the potential development of ocular gene therapy for glaucoma. Because of the increase in outflow facility observed both in vivo and in vitro after treatment with microtubule depolymerization drugs, we wish to initially investigate B-tubulins, microtubule-associated proteins and the transcription factor NFkB. The PI will characterize which isoforms from these gene families are expressed in the trabecular meshwork. She will employ adenovirus transfer technology to overexpress and underexpress their specific regions in organ culture perfused porcine and human eyes. She will measure outflow facility. Preliminary results indicate that replication-deficient adenoviruses are likely ideal vectors to investigate our hypotheses. They enter the cells of the trabecular meshwork very efficiently and, by themselves, do not interfere with the cytoskeleton network nor with the rate of outflow. Even if these initial manipulations do not alter aqueous outflow, this investigation should still contribute valuable insight about cellular mechanisms involved in aqueous humor outflow. Furthermore, the complete development of this system, coupling molecular biology with physiological measurements, will allow testing of future multiple hypotheses involving proteins/enzymes (receptors, growth factors, ion channels, kinases...) that have been conceptualized as being important in aqueous humor physiology.