Kisspeptin藉由細胞自噬調控 β 細胞在葡萄糖刺激下的胰島素分泌

Abstract

已知kisspeptins及其受體Kiss1r在生殖系統中扮演了重要的角色。然而，對於他們在其他周邊組織中的功能及角色了解的仍然不多。文獻中指出升糖素 (Glucagon) 會刺激肝臟分泌kisspeptin而進一步影響葡萄糖刺激的胰島素分泌功能 (Glucose-stimulated insulin secretion, GSIS)。而值得注意的是，在二型糖尿病的患者血液中，會有較高的kisspeptin濃度，且葡萄糖刺激的胰島素分泌功能也會隨著病程的發展而有受損的情形。再加上細胞自噬 (Autophagy) 被認為參與了胰島素的分解作用而能調控β細胞的胰島素分泌功能。因此，我們提出假設，kisspeptin可能會藉由調控β細胞中的細胞自噬而直接影響胰島素的分泌，而這樣的現象可能在二型糖尿的的病程中扮演了重要的角色。 首先，在經由專一性的測試後，利用自製的抗體組織免疫染色法中證實Kiss1r表現於β細胞中，因而直接傳遞kisspeptin的下游訊息。接著，利用小鼠β細胞的細胞株，NIT-1，建立體外試驗模式以了解長期給予kisspeptin對β細胞胰島素分泌的影響及其可能的機制。從結果可以看到，在利用過度表達kisspeptin於細胞中而模擬長期影響的模式下，NIT-1細胞中葡萄糖刺激的胰島素分泌會受到抑制，且同時會促進細胞自噬的發生進而影響胰島素在細胞中的含量。除此之外，阻檔細胞自噬能夠有效回復過度表達kisspeptin對胰島素含量的影響。 綜合以上結果，說明了受到kisspeptin影響而受損的胰島素分泌功能可能是藉由調控細胞自噬而改變胰島素在細胞中的含量所致。由於長期處於高濃度kisspeptin對胰島素分泌的影響可能在二型糖尿病的病程中扮演了重要的角色，透過我們的實驗更加了解其機制則有助於提供未來可能的治療方向。

Kisspeptin and its receptor Kiss1r are known to play important roles in regulating animal reproduction. However, their functions in other peripheral tissues are poorly understood. Couple with the proved fact that hepatic kisspeptin stimulated by glucagon would inhibit GSIS, the report also showed that type 2 diabetes patients and obese mice model exhibited not only an increase in liver kisspeptin mRNA and protein level but also impaired GSIS function (Song et al., 2014). It gave us a hint that kisspeptin/Kiss1r system might plays a role in the development of type 2 diabetes. In addition, previous studies have shown that the capacity of GSIS in β cell is negatively regulated by autophagic degradation of insulin. Based on these studies, we hypothesized that kisspeptin may activate autophagy to suppress GSIS in β cells. To validate our hypothesis, we first conducted IHC staining experiments using our previously developed anti-Kiss1r polyclonal IgY antibody to confirm the expression of Kiss1r in mouse β cell. Our IHC staining results showed that Kiss1r is positively stained in insulin expressing β cells. Then, we chose mouse pancreatic β cell line, NIT-1, that positively respond to kisspeptin as in vitro model to study the effects of kisspeptin in β cells. In our study, we established Kiss1 over-expression model that we used to mimic long-term kisspeptin exposure to study the posible roles of kisspeptin in type 2 diabetes. It was shown that long-term treatment of kisspeptin resulted in potently suppressed GSIS and decreased insulin protein level in NIT-1 cells. Importantly, our Western Blot results proved the fact that long-term treatment of kisspeptin impaired GSIS through induced autophagy and thus decreased insulin content in NIT-1 cells. Taken together, our data suggested that long-term kisspeptin exposure could suppress GSIS in β cell by inducing autophagic degradation of insulin, which might play an important role in type 2 diabetes development.