2021-01-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/644283摘要： 本計劃擬研究體積感應陰離子通道(VSOAC)於代謝及循環系統中之生理功能。先前研究指出，人類LRRC8A 為構成VSOAC 之必要蛋白質，而另一必要蛋白質LRRC8D 則可調控VSOAC 運輸牛磺酸之活性進而調節細胞體積。在2014 年發現LRRC8A 為VSOAC 之必要元件後，近年來本實驗室與其他研究皆致力於釐清VSOAC 之生理功能。第五型丙球蛋白缺乏貧血症為LRRC8A 基因平衡易位缺損，導致病患循環系統缺少B 細胞。在Lrrc8a 基因剔除小鼠研究中則發現T 細胞發育異常。然而，若小鼠Lrrc8a 只有C 端缺損時，B 細胞及T 細胞卻不受影響。因此，VSOAC 在淋巴細胞發育和功能中的作用仍需進一步研究。此外，其他研究發現血流之剪力會影響血球分化，而我們先前研究也發現牛磺酸缺乏會使心臟發育異常。由於牛磺酸是調控血液動力持衡的重要因子，且VSOAC 為目前已知唯一將牛磺酸運至細胞外之通道。本計畫首先將研究斑馬魚lrrc8a 突變對於VSOAC 生化特性及細胞生理之影響。接著，我們假設牛磺酸透過VSOAC 調節滲透壓進而影響循環及淋巴系統的發育，將藉由添加牛磺酸於lrrc8a 基因突變斑馬魚胚及調降lrrc8d，研究lrrc8a 與牛磺酸、lrrc8d 對心臟血管生成及造血作用之影響，並進一步評估其淋巴器官及淋巴細胞之分佈型態。最後，我們假設牛磺酸透過VSOAC 影響代謝持衡，將以本實驗室先前建立之斑馬魚運動模式及過度餵飼模式，比較添加牛磺酸或抑制lrrc8d對lrrc8a基因突變斑馬魚之運動耐力、體重成長及血糖持衡之影響。綜上所述，本計劃之目標為：一、研究Lrrc8a N 端在VSOAC 通道中的功能。二、VSOAC 對循環及免疫系統的影響。三、研究VSOAC 對代謝持衡的影響。 Abstract our own study began to shed light on the physiological role of VRAC/VSOAC. Human patient with balanced translocation disrupting LRRC8A lead to agammaglobulinemia, but Lrrc8a knockout mouse showed abnormality in T cell development with normal B cell function. Furthermore, hypomorphic mouse mutation with truncated C-terminal of Lrrc8a gene showed normal B and T cells. These controversial evidence indicate that the roles of VRAC/VSOAC in lymphocyte development and function remains elusive. On the other hand, previous studies done by other groups indicated that hematopoiesis is influenced by shear stress triggered by hemodynamics, while our previous study suggested that taurine deficiency led to cardiac abnormalities. Since taurine has been considered an important player in hemodynamic homeostasis and VSOAC is the only known mechanism to efflux taurine from within a cell to extracellular environment, in this project, we will first characterize zebrafish lrrc8a mutant genes for their biochemical, cellular and physiological role as an essential component of VSOAC. Secondly, we hypothesize that taurine, via VSOAC, play a role in the development of circulatory system and lymphatic system. To this end, we have created and acquired zebrafish lrrc8a mutant lines, and will characterize their phenotypes in cargiogenesis, vasculogenesis, angiogenesis and hematopoiesis with or without taurine supplementation and lrrc8d paralogs. Furthermore, the lymphatic organs and lymphocyte profiles in the adult zebrafish will also be evaluated. Thirdly, we hypothesize the taurine, via VSOAC, play a role in the metabolic homeostasis. To this end, we will compare the exercise tolerance, body weight growth and blood sugar homeostasis in our intensive exercise model and overfeeding obese model in lrrc8a mutant zebrafish with or without taurine supplementation and lrrc8d paralogs. Aim 1: To investigate the role of Lrrc8a N-terminus in modulating channel function; Aim 2: To characterize the role of VSOAC in the circulatory and immune system; Aim 3: To determine the role of VSOAC in metabolic homeostasis.心血管系統滲透壓調節牛磺酸斑馬魚血流動力學Cardiovascular systemHemodynamicsOsmoregulationTaurineZebrafish