Ahn Miwon, Dhawan Sangeeta, McCown Erika M, Garcia Pablo A, Bhattacharya Supriyo, Stein Roland, Thurmond Debbie C
Department of Molecular and Cellular Endocrinology, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of Hope, Duarte, CA, USA.
Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, Beckman Research Institute, City of Hope, Duarte, CA, USA.
Diabetologia. 2025 Jan;68(1):152-165. doi: 10.1007/s00125-024-06286-2. Epub 2024 Oct 15.
AIMS/HYPOTHESIS: p21 (CDC42/RAC1) activated kinase 1 (PAK1) is depleted in type 2 diabetic human islets compared with non-diabetic human islets, and acute PAK1 restoration in the islets can restore insulin secretory function ex vivo. We hypothesised that beta cell-specific PAK1 enrichment in vivo can mitigate high-fat-diet (HFD)-induced glucose intolerance by increasing the functional beta cell mass.
Human islets expressing exogenous PAK1 specifically in beta cells were used for bulk RNA-seq. Human EndoC-βH1 cells overexpressing myc-tagged PAK1 were used for chromatin immunoprecipitation (ChIP) and ChIP-sequencing (ChIP-seq). Novel doxycycline-inducible beta cell-specific PAK1-expressing (iβPAK1-Tg) mice were fed a 45% HFD pre-induction for 3 weeks and for a further 3 weeks with or without doxycycline induction. These HFD-fed mice were evaluated for GTT, ITT, 6 h fasting plasma insulin and blood glucose, body composition, islet insulin content and apoptosis.
Beta cell-specific PAK1 enrichment in type 2 diabetes human islets resulted in decreased beta cell apoptosis and increased insulin content. RNA-seq showed an upregulation of INS gene transcription by PAK1. Using clonal human beta cells, we found that PAK1 protein was localised in the cytoplasm and the nucleus. ChIP studies revealed that nuclear PAK1 enhanced pancreatic and duodenal homeobox1 (PDX1) and neuronal differentiation 1 (NEUROD1) binding to the INS promoter in a glucose-responsive manner. Importantly, the iβPAK1-Tg mice, when challenged with HFD and doxycycline induction displayed enhanced glucose tolerance, increased islet insulin content and reduced beta cell apoptosis when compared with iβPAK1-Tg mice without doxycycline induction.
CONCLUSIONS/INTERPRETATION: PAK1 plays an unforeseen and beneficial role in beta cells by promoting insulin biogenesis via enhancing the expression of PDX1, NEUROD1 and INS, along with anti-apoptotic effects, that culminate in increased insulin content and beta cell mass in vivo and ameliorate diet-induced glucose intolerance.
The raw and processed RNA-seq data and ChIP-seq data, which has been made publicly available at Gene Expression Omnibus (GEO) at https://www.ncbi.nlm.nih.gov/geo/ , can be accessed in GSE239382.
目的/假设：与非糖尿病患者的胰岛相比，2型糖尿病患者的胰岛中p21（CDC42/RAC1）激活激酶1（PAK1）含量减少，胰岛中PAK1的急性恢复可在体外恢复胰岛素分泌功能。我们假设，体内β细胞特异性富集PAK1可通过增加功能性β细胞量来减轻高脂饮食（HFD）诱导的葡萄糖不耐受。
使用在β细胞中特异性表达外源性PAK1的人胰岛进行批量RNA测序。使用过表达myc标签PAK1的人EndoC-βH1细胞进行染色质免疫沉淀（ChIP）和ChIP测序（ChIP-seq）。新型强力霉素诱导的β细胞特异性表达PAK1（iβPAK1-Tg）小鼠在诱导前用45%的HFD喂养3周，然后在有或没有强力霉素诱导的情况下再喂养3周。对这些HFD喂养的小鼠进行葡萄糖耐量试验（GTT）、胰岛素耐量试验（ITT）、6小时空腹血浆胰岛素和血糖、身体组成、胰岛胰岛素含量和细胞凋亡评估。
2型糖尿病人胰岛中β细胞特异性富集PAK1可导致β细胞凋亡减少和胰岛素含量增加。RNA测序显示PAK1上调了胰岛素（INS）基因转录。使用克隆人β细胞，我们发现PAK1蛋白定位于细胞质和细胞核。ChIP研究表明，细胞核中的PAK1以葡萄糖反应性方式增强胰腺和十二指肠同源盒1（PDX1）和神经分化1（NEUROD1）与INS启动子的结合。重要的是，与未用强力霉素诱导的iβPAK1-Tg小鼠相比，用HFD和强力霉素诱导的iβPAK1-Tg小鼠表现出增强的葡萄糖耐量、增加的胰岛胰岛素含量和减少的β细胞凋亡。
结论/解读：PAK1通过增强PDX1、NEUROD1和INS的表达促进胰岛素生物合成，同时具有抗凋亡作用，最终导致体内胰岛素含量增加和β细胞量增加，并改善饮食诱导的葡萄糖不耐受，从而在β细胞中发挥了意想不到的有益作用。
已在基因表达综合数据库（GEO）（https://www.ncbi.nlm.nih.gov/geo/）上公开提供的原始和处理后的RNA测序数据及ChIP测序数据可在GSE239382中获取。
