- 學位論文
遠端腎小管鈣及鎂離子運送之分子機制
Molecular Mechanism of Calcium and Magnesium Transport by Renal Distal Tubule
摘要
腎臟是維持身體鈣離子及鎂離子恆定重要的器官之一，藉由腎絲球的過濾及腎小管的重吸收以達到調控離子排出，調節整體鈣離子與鎂離子的恆定。在腎小管的運送會直接影響這些離子從尿液排出量的多寡，而運送的過程則依不同節段腎小管有各自的方式與運送分子。遠端腎小管除了具受特殊的荷爾蒙調節及穿透細胞的方式之特色外，藉由近年被找出的第一個腎小管鈣離子通道TRPV5及鎂鈣離子通道TRPM6已可完整地將此段腎小管的分子運送機制呈現。 我們的研究從以calbindin-D28k基因剔除鼠的動物實驗瞭解到遠端腎小管對鈣離子重吸收的重要為開始，此一老鼠有明顯尿鈣增加的現象。此外，在副甲狀腺切除的實驗中，我們也證實副甲狀腺荷爾蒙對TRPV5、calbindin-D28k的正向調節作用，而鎂離子通道TRPM6也有同樣的變化，而亨利氏上行支的paracellin-1則不受影響。而生理狀況如鹽分及體液的改變，也直接影響到鈣、鎂離子在遠端腎小管的運送。為進一步瞭解位於遠端腎小管上的這些運送分子在其他方面可能扮演的角色，我們主要的三個研究分別探討：（一）calcineurin inhibitors引起高尿鈣及低血鎂的分子機制 ：在此一研究中，環孢靈影響維他命D3的代謝途徑，抑制維他命D受器的表現，因而抑制了包括TRPV5及calbindins這些分子，而FK506則並不影響維他命D於腎臟的代謝；（二）不同利尿劑對鈣離子運送的影響 ：研究結果發現thiazide類的利尿劑會直接刺激遠端腎小管增加鈣離子重吸收，而furosemide導致的抑制重吸收則使得遠端腎小管鈣離子的運送增加；（三）糖尿病合併的尿鈣及尿鎂增加的機制：研究結果推測鈣及鎂運送的障礙並不在遠端腎小管，但鈣及鎂離子在遠端腎小管的重吸收有代償性的增加。 綜述我們的研究，除了瞭解這些運送分子的生理功能及荷爾蒙調控外，利用實驗動物模式分別探討這些分子參與的程度及實際的生理病理及藥理分子機制，對此一領域的研究，增添新的發現，將有助於更多研究的進展，俾利於臨床應用。
並列摘要
The kidney plays a crucial role in maintaining the homeostasis of calcium and magnesium. There are many factors modulate renal calcium and magnesium handling, and they act either directly on transport mechanism with specific effects on transport molecules or indirectly via various signal transduction pathways. Renal distal tubule has distinguishable role in regulating the calcium and magnesium transport. Recent identification of novel apical calcium and magnesium channels, TRPV5 and TRPM6, provide the direct entry as the initial step of the transcellular transport. Our studies aim to investigate the role of these transport molecules in health and disease status. Based on our previous work of calbindin-D28k knockout mice, we continue to assess the effect of calcineurin inhibitors, including cyclosporine A and tacrolimus on distal tubule calcium and magnesium handling. Both drugs induce calciuria and magnesiuria and significant downregulation of calbindin-D28k and calbindin-D9k associated with decreased expression of TRPV5, TRPM6 were noted. However, cyclosporine A inhibited vitamin D receptor expression but tacrolimus did not affect vitamin D metabolism. Treatment with thiazide can reduce urinary calcium excretion. Results of our study show that the hypocalciuric effect was associated with upregulation of calcium transporters in the absence of diuretics-induced volume depletion. While increased expression of these downstream transporters of distal tubule after furosemide administration represents a load-dependent physiological response. This result was demonstrated in our study irrespective of volume status. Diabetes mellitus is associated with renal calcium and magnesium wasting. Impaired tubular reabsorption is considered to be the underlying defect. Our animal study demonstrated that in untreated diabetic rats, both calcium and magnesium transporters were increased in expression and insulin therapy corrected hyperglycemia and also normalized the expression of transport molecules. This result suggests distal tubule might not be involved in diabetes-associated calciuria and magnesiuria. In summary, using animal models, our series of physiological and pharmacological investigations demonstrate that the novel epithelial calcium and magnesium channels play an important role in maintaining calcium and magnesium homeostasis and that modulation of their expression results in abnormalities in calcium and/or magnesium homeostasis.