Klotho基因突變鼠腎臟病理特性之分析

腎臟是Klotho表現量最豐富的地方，老化會造成腎臟結構性改變，而Klotho基因缺陷小鼠有加速老化的現象，而且會因為無法抑制生長因子和增強內質網壓力造成腎臟纖維化增加。Klotho可以增加EpoR （erythropoietin receptor, EpoR）表現，並活化MAPK（mitogen activated protein kinase）/ERK（extracellular signaling related kinase）路徑，增加對細胞的保護作用。VEGF（vascular endothelial growth factor）會刺激內皮細胞增殖和分化，並藉由內皮細胞讓血管擴張，以增加血管通透性，當缺乏Klotho基因，會造成VEGFR2（vascular endothelial growth factor receptor 2）/TRPC-1（transient-receptor potential canonical Ca2+ channel 1）複合體作用受損，造成血管內皮細胞受損。為了更深入探討Klotho基因缺陷小鼠腎臟結構性改變，以及Klotho基因缺陷對EpoR、ERK、VEGF和VEGFR2蛋白表現之影響，本研究藉由動物實驗，觀察Klotho基因缺陷小鼠腎臟組織切片，藉由一般染色法、Trichrome染色和免疫組織化學IHC染色（immunohistochemical stain），以探討Klotho基因缺陷對於腎臟病理變化及生理機制之影響。 實驗結果發現，Klotho基因缺陷會造成腎臟腎小管細胞核縮小和濃染，腎絲球繫膜細胞增生，腎小管和血管纖維化，並使氧化壓力顯著增加，也會顯著增加血管、腎絲球和腎小管中VEGFR2以及腎小管VEGF的表現。 Klotho基因缺陷對於EpoR表現之影響並不明顯，不過仍然會影響EpoR之下游蛋白P-Erk1/2，使得亨利氏環粗支及腎絲球之P-Erk1/2表現顯著增加，推測可能Klotho和P-Erk1/2之間可能還有其他路徑介入調控，有待後續研究證實。本研究結果支持Klotho在調控血管新生作用，以及保護腎臟細胞作用上扮演重要角色。

Klotho is expressed most abundantly in the kidney. While aging causes structural changes in the kidney, Klotho gene-deficient mice are known to exhibit accelerated aging and are expected to have pre-mature renal aging phenotypes. Since Klotho gene-deficient mice can’t inhibit growth factors, concomitant with elevated endoplasmic reticulum pressure, kidney fibrosis is expected to accumulate in the mutant mice. Klotho can increase EpoR expression and activate MAPK/ ERK pathway to protect kidney. VEGF can stimulate the proliferation and differentiation of endothelial cells as well as enhance vascular permeability. Klotho gene knockout leads to impairment of VEGFR2/TRPC-1 complex, with subsequent damages to vascular endothelial cells. This study investigated the structural changes of kidneys and the protein expression of EpoR, ERK, VEGF and VEGFR2 under mouse Klotho deficiency status. H-E staining, Trichrome staining and IHC staining were applied to investigated the traits of renal pathological changes and physiological mechanisms in the Klotho mutant mice. The results showed that Klotho gene deficiency caused the nucleus of renal tubule cells smaller and hyperchromatic with excessive mesangial cell proliferation in the glomeruli. The Klotho gene deficiency also leads to renal tubular and vascular fibrosis and significant increase of oxidative stress. VEGFR2 expression was significantly increased in blood vessels, glomeruli, and renal tubules, and the VEGF expression of renal tubular also increased significantly under the influence of Klotho deficiency. Although the effect of Klotho gene deficiency on EpoR expression was not obvious, EpoR downstream protein P-Erk1/2 was evidently increased in the thick segment of the loop of Henle and glomeruli, suggesting that other pathways may be involved in the regulation between Klotho and P-Erk1/2. More studies should confirm this potential linkage. The results of this study support that Klotho plays an important role in the regulation of angiogenesis and protection of renal cells.