Bioluminescent Monitoring of Graft Survival in an Adoptive Transfer Model of Autoimmune Diabetes in Mice

Type 1 diabetes is characterized by the autoimmune destruction of the insulin-producing beta cells of the pancreas. A promising treatment for this disease ...

Harvard Medical School

Perspectives on the activities of ANGPTL8/betatrophin.

The β-cell mitogenic effects of ANGPTL8 have been subjected to substantial debate. The original findings suggested that ANGPTL8 overexpression in mice induced a 17-fold increase in β-cell proliferation. Subsequent studies in mice contested this claim, but a more recent report in rats supported the original observations. These conflicting results might be explained by variable ANGPTL8 expression and differing methods of β-cell quantification. To resolve the controversy, three independent labs collaborated on a blinded study to test the effects of ANGPTL8 upon β-cell proliferation. Recombinant human betatrophin (hBT) fused to maltose binding protein (MBP) was delivered to mice by intravenous injection. The results demonstrate that ANGPTL8 does not stimulate significant β-cell proliferation. Each lab employed different methods for β-cell identification, resulting in variable quantification of β-cell proliferation and suggests a need for standardizing practices for β-cell quantification. We also observed a new action of ANGPTL8 in stimulating CD45+ hematopoietic-derived cell proliferation which may explain, in part, published discrepancies. Overall, the hypothesis that ANGPTL8 induces dramatic and specific β-cell proliferation can no longer be supported. However, while ANGPTL8 does not stimulate robust β-cell proliferation, the original experimental model using drug-induced (S961) insulin resistance was validated in subsequent studies, and thus still represents a robust system for studying signals that are either necessary or sufficient for β-cell expansion. As an added note, we would like to commend collaborative group efforts, with repetition of results and procedures in multiple laboratories, as an effective method to resolve discrepancies in the literature. 

Resolving Discrepant Findings on ANGPTL8 in β-Cell Proliferation: A Collaborative Approach to Resolving the Betatrophin Controversy.

Investigation of cell-cycle kinetics in mammalian pancreatic β cells has mostly focused on transition from the quiescent (G0) to G1 phase. Here, we report that centromere protein A (CENP-A), which is required for chromosome segregation during the M-phase, is necessary for adaptive β cell proliferation. Receptor-mediated insulin signaling promotes DNA-binding activity of FoxM1 to regulate expression of CENP-A and polo-like kinase-1 (PLK1) by modulating cyclin-dependent kinase-1/2. CENP-A deposition at the centromere is augmented by PLK1 to promote mitosis, while knocking down CENP-A limits β cell proliferation and survival. CENP-A deficiency in β cells leads to impaired adaptive proliferation in response to pregnancy, acute and chronic insulin resistance, and aging in mice. Insulin-stimulated CENP-A/PLK1 protein expression is blunted in islets from patients with type 2 diabetes. These data implicate the insulin-FoxM1/PLK1/CENP-A pathway-regulated mitotic cell-cycle progression as an essential component in the β cell adaptation to delay and/or prevent progression to diabetes.

Insulin Signaling Regulates the FoxM1/PLK1/CENP-A Pathway to Promote Adaptive Pancreatic β Cell Proliferation.

Type 1 diabetes (T1D) is caused by the autoimmune destruction of pancreatic beta cells. Pluripotent stem cells can now be differentiated into beta cells, thus raising the prospect of a cell replacement therapy for T1D. However, autoimmunity would rapidly destroy newly transplanted beta cells. Using a genome-scale CRISPR screen in a mouse model for T1D, we show that deleting RNLS, a genome-wide association study candidate gene for T1D, made beta cells resistant to autoimmune killing. Structure-based modelling identified the U.S. Food and Drug Administration-approved drug pargyline as a potential RNLS inhibitor. Oral pargyline treatment protected transplanted beta cells in diabetic mice, thus leading to disease reversal. Furthermore, pargyline prevented or delayed diabetes onset in several mouse models for T1D. Our results identify RNLS as a modifier of beta cell vulnerability and as a potential therapeutic target to avert beta cell loss in T1D.

Genome-scale in vivo CRISPR screen identifies RNLS as a target for beta cell protection in type 1 diabetes.

The CRISPR/Cas9 genome editing system has been one of the greatest scientific discoveries in the last decade. The highly efficient and precise editing ability of this technology is of great therapeutic value and benefits the basic sciences as an advantageous research tool. In recent years, forward genetic screens using CRISPR technology have been widely adopted, with genome-wide or pathway-focused screens leading to important and novel discoveries. CRISPR screens have been used primarily in cancer biology, virology, and basic cell biology, but they have rarely been applied to diabetes research. A potential reason for this is that diabetes-related research can be more complicated, often involving cross talk between multiple organs or cell types. Nevertheless, many questions can still be reduced to the study of a single cell type if assays are carefully designed. Here we review the application of CRISPR screen technology and provide perspective on how it can be used in diabetes research.

Applying CRISPR Screen in Diabetes Research.

Prevention or amelioration of declining β cell mass is a potential strategy to cure diabetes. Here, we report the pathways utilized by β cells to robustly replicate in response to acute insulin resistance induced by S961, a pharmacological insulin receptor antagonist. Interestingly, pathways that include CENP-A and the transcription factor E2F1 that are independent of insulin signaling and its substrates appeared to mediate S961-induced β cell multiplication. Consistently, pharmacological inhibition of E2F1 blocks β-cell proliferation in S961-injected mice. Serum from S961-treated mice recapitulates replication of β cells in mouse and human islets in an E2F1-dependent manner. Co-culture of islets with adipocytes isolated from S961-treated mice enables β cells to duplicate, while E2F1 inhibition limits their growth even in the presence of adipocytes. These data suggest insulin resistance-induced proliferative signals from adipocytes activate E2F1, a potential therapeutic target, to promote β cell compensation.

Peng Yi

Section for Islet Cell and Regenerative Biology,
Joslin Diabetes Center,
Section for Islet Cell and Regenerative Biology, Joslin Diabetes Center

Harvard Medical School

Bioluminescent Monitoring of Graft Survival in an Adoptive Transfer Model of Autoimmune Diabetes in Mice

Peng Yi

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Section for Islet Cell and Regenerative Biology,Joslin Diabetes Center,Section for Islet Cell and Regenerative Biology, Joslin Diabetes Center

Harvard Medical School

Bioluminescent Monitoring of Graft Survival in an Adoptive Transfer Model of Autoimmune Diabetes in Mice

Section for Islet Cell and Regenerative Biology,
Joslin Diabetes Center,
Section for Islet Cell and Regenerative Biology, Joslin Diabetes Center