Studies on insulin secretory granule formation capacity by the control of proprotein-processing endoprotease furin.

我们以前报道过，具有较高弗林蛋白酶表达的胰岛细胞诱导生长因子的产生增加，这导致使用自分泌/旁分泌机制增加细胞生长。在胰腺β细胞中寻找弗林蛋白酶的底物时，我们发现甲状旁腺相关蛋白（PTHrP）和转化生长因子β（TGF β）具有弗林蛋白酶切割基序Arg-X-（Lys/Arg）-Arg。PTHrP常在胰腺内分泌肿瘤中产生。PTHrP作为前体pro-PTHrP合成，并且前肽从其前体的切割对于其生物活化是至关重要的。我们证明，弗林蛋白酶是高表达的大鼠胰岛生长在围产期。由此，我们怀疑弗林蛋白酶可能与PTHrP在胰岛素瘤中共表达。我们检查了21例人类内分泌肿瘤，弗林蛋白酶在所有10例胰岛素瘤中均呈阳性染色。同样，在相同的胰岛素瘤中检测到PTHrP。但其他非胰岛素瘤内分泌肿瘤 关于我们 MORS不显示弗林蛋白酶和PTHrP阳性。因此，弗林蛋白酶和pro-PTHrP在胰岛素瘤中特异性共表达。据报道，PTHrP的产生在更分化的β细胞培养系中更高，尽管胰岛素瘤比正常胰岛β细胞分化更少。PTHrP诱导某些细胞类型的分化和其他细胞类型的去分化或生长。我们检查了PTHrP的产生是在生长的β细胞中还是在分化良好的β细胞中更大，以及PTHrP是否诱导β细胞的分化或生长。我们使用了分化良好的小鼠β细胞系MIN6的四个亚系（17、25、31和41代）和小鼠胰岛。随着MIN6传代次数的增加，胰岛β细胞特异性分化特征如胰岛素含量和TGF β表达逐渐减少，而PTHrP、furin表达和细胞生长逐渐增加。PTHrP（1 - 34）和PTHrP（1 - 86）均能增加MIN6 - 17细胞胰岛素含量和mRNA水平，且作用强度大于MIN6 - 41细胞。PTHrP诱导的胰岛素含量的增加也注意到在原代培养的胰岛。相比之下，PTHrP在MIN6 - 41细胞中比在MIN6 - 17细胞中更广泛地增加DNA合成。PTHrP可增加MIN6 - 17细胞TGF β的表达，降低MIN6 - 41细胞TGF β的表达。MIN 6细胞中PTH/PTHrP受体和TGF β Ⅱ型受体的表达相似。双丁酰cAMP在MIN6亚系中再现了PTHrP对胰岛素含量和DNA合成的影响。PTHrP似乎通过cAMP途径诱导胰岛素表达。我们得出结论，PTHrPb通过cAMP途径增加分化良好的β细胞中的分化功能，如胰岛素和TGF β的表达，并刺激生长中的β细胞的生长。少

We previously reported that the islet cells with higher furin expression induces increased production of growth factors, which result in an increase in cell growth using an autocrine/paracrine mechanism. When searching for substrates for furin in pancreatic β cells, we noticed that parathyroid hormone-related protein (PTHrP) and transforming growth factor β (TGFb) possess a furin cleavage motif Arg-X-(Lys/Arg)-Arg. PTHrP is frequently produced in pancreatic endocrine tumors. PTHrP is synthesized as the precursor pro-PTHrP and the cleavage of the propeptide from its precursor is crucial for its biological activation. We demonstrated that furin is highly expressed in rat pancreatic islets during the perinatal stage growth. From this, we suspected that furin may be co-expressed with PTHrP in insulinomas. We examined 21 human endocrine tumors, and furin was positively stained in all 10 insulinomas. Likewise, PTHrP was detected in the same insulinomas. But other non-insulinomas endocrine tu … More mors did not display furin- and PTHrP- positivity. Thus, furin and pro-PTHrP are co-expressed specifically in insulinomas.Production of PTHrP is reportedly higher in more differentiated β cell culture lines, although insulinomas are less differentiated than normal islet β cells. PTHrP induces differentiation in some cell-types and de-differentiation or growth in others. We examined whether PTHrP production is greater in growing β cells or in well-differentiated β cells, and whether PTHrP induces differentiation or growth in β cells. We used four sublines of the well-differentiated mouse β cell line, MIN6, with 17, 25, 31, and 41 passages, and mouse pancreatic islets. With insreasing MIN6 passage number, β cell-specific differentiated features such as insulin content diminished together with TGFβ expression, whereas the expression of PTHrP, furin, and cell growth gradually increased. Both PTHrP(1-34) and PTHrP(1-86) increased insulin content and mRNA levels more in MIN6-17 cells than in MIN6-41 cells. A PTHrP-induced increase in insulin content was also noted in primary-cultured islets. In contrast, PTHrP increased DNA synthesis more extensively in MIN6-41 cells than in MIN6-17 cells. PTHrP increased TGFβ expression in MIN6-17 cells and decreased its expression in MIN6-41 cells. The expression of PTH/PTHrP receptor and TGFβ type II receptor were similar in all MIN6 sublines. Dibutyryl cAMP reproduced PTHrP's effect on insulin content and DNA synthesis in the MIN6 sublines. PTHrP appears to induce insulin expression by a cAMP pathway. We conclude that PTHrPb increases differentiated functions such as insulin and TGFβ expressions in well-differentiated β cells through the cAMP pathway, and stimulates growth in growing β cells. Less