Circulating miRNA-150-5p is associated with immune-mediated early ß-cell loss in a humanized mouse model.

BACKGROUND: Aberrant microRNA (miRNA) expression levels are associated with various graft rejections. We used our humanized mouse model with transplanted human islets to identify miRNAs in islet grafts related to xenograft rejection and circulating miRNAs associated with xenograft rejection-mediated ß-cell loss. METHODS: Diabetic immunodeficient NOD.scid mice were transplanted with human islets and subsequently achieved stable normoglycemia. Lymphocytes from NOD mice were then adoptively transferred to the humanized mice to induce human ß-cell destruction. Islet graft and plasma were collected immediately once blood glucose reached >200 mg/dL. miRNAs in the islet grafts and in the plasma with or without adoptive lymphocyte transfer (ALT) were measured using NanoString nCounter® miRNA Expression Assay and qPCR. RESULTS: A set of immune-related miRNAs was significantly increased in human islet grafts of ALT-treated mice compared to control mice. Of these miRNAs, miR-150-5p was significantly increased in the circulation of ALT-treated mice at tissue collection and the increase was a result of immune activation rather than simply the presence of lymphocytes in circulation. Furthermore, miR-150-5p was significantly increased in human islet graft and circulation prior to the development of hyperglycemia in the ALT-treated mice. CONCLUSIONS: Our data demonstrated that immune-related miRNAs are associated with human islet xenograft rejection in mice. miR-150-5p is increased in human islet graft and in the circulation during islet xenograft rejection and ß-cell destruction prior to hyperglycemia and may be an early biomarker for islet xenograft rejection.

Exploring lncRNAs associated with human pancreatic islet cell death induced by transfer of adoptive lymphocytes in a humanized mouse model.

Background: Long noncoding RNA (lncRNA)-mediated posttranscriptional and epigenetic landscapes of gene regulation are associated with numerous human diseases. However, the regulatory mechanisms governing human ß-cell function and survival remain unknown. Owing to technical and ethical constraints, studying the direct role of lncRNAs in ß-cell function and survival in humans in vivo is difficult. Therefore, we utilized humanized mice with human islets to investigate lncRNA expression using whole transcriptome shotgun sequencing. Our study aimed to characterize lncRNAs that may be crucial for human islet cell function and survival. Methods: Human ß-cell death was induced in humanized mice engrafted with functional human islets. Using these humanized mice harboring human islets with induced ß-cell death, we investigated lncRNA expression through whole transcriptome shotgun sequencing. Additionally, we systematically identified, characterized, and explored the regulatory functions of lncRNAs that are potentially important for human pancreatic islet cell function and survival. Results: Human islet cell death was induced in humanized mice engrafted with functional human islets. RNA sequencing analysis of isolated human islets, islet grafts from humanized mice with and without induced cell death, revealed aberrant expression of a distinct set of lncRNAs that are associated with the deregulated mRNAs important for cellular processes and molecular pathways related to ß-cell function and survival. A total of 10 lncRNA isoforms (SCYL1-1:22, POLG2-1:1, CTRB1-1:1, SRPK1-1:1, GTF3C5-1:1, PPY-1:1, CTRB1-1:5, CPA5-1:1, BCAR1-2:1, and CTRB1-1:4) were identified as highly enriched and specific to human islets. These lncRNAs were deregulated in human islets from donors with different BMIs and with type 2 diabetes (T2D), as well as in cultured human islets with glucose stimulation and induced cell death induced by cytokines. Aberrant expression of these lncRNAs was detected in the exosomes from the medium used to culture islets with cytokines. Conclusion: Islet-enriched and specific human lncRNAs are deregulated in human islet grafts and cultured human islets with induced cell death. These lncRNAs may be crucial for human ß-cell function and survival and could have an impact on identifying biomarkers for ß-cell loss and discovering novel therapeutic targets to enhance ß-cell function and survival.

Activation of GPR119 Stimulates Human ß-Cell Replication and Neogenesis in Humanized Mice with Functional Human Islets.

Using humanized mice with functional human islets, we investigated whether activating GPR119 by PSN632408, a small molecular agonist, can stimulate human ß-cell regeneration in vivo. Human islets were transplanted under the left kidney capsule of immunodeficient mice with streptozotocin- (STZ-) induced diabetes. The recipient mice were treated with PSN632408 or vehicle and BrdU daily. Human islet graft function in the mice was evaluated by nonfasting glucose levels, oral glucose tolerance, and removal of the grafts. Immunostaining for insulin, glucagon, and BrdU or Ki67 was performed in islet grafts to evaluate α- and ß-cell replication. Insulin and CK19 immunostaining was performed to evaluate ß-cell neogenesis. Four weeks after human islet transplantation, 71% of PSN632408-treated mice achieved normoglycaemia compared with 24% of vehicle-treated mice. Also, oral glucose tolerance was significantly improved in the PSN632408-treated mice. PSN632408 treatment significantly increased both human α- and ß-cell areas in islet grafts and stimulated α- and ß-cell replication. In addition, ß-cell neogenesis was induced from pancreatic duct cells in the islet grafts. Our results demonstrated that activation of GPR119 increases ß-cell mass by stimulating human ß-cell replication and neogenesis. Therefore, GPR119 activators may qualify as therapeutic agents to increase human ß-cell mass in patients with diabetes.