Reactive oxygen species (ROS) and peroxiredoxin signaling appear to have a role in oleate-induced β-cell proliferation, according to Vivoli et al. (p. 45). Confirmed by blocking specific parts of the suspected signaling pathway, the findings further decipher the underlying mechanisms of β-cell proliferation. Type 1 and type 2 diabetes are both characterized by a loss of functional β-cells; thus, any approach to increasing β-cell mass could also be a lead for therapies. The study, which focuses of isolated rat islets, centers around the idea that glucose and fatty acids are central to how β-cells react to nutrients. The authors had previously shown that in the presence of elevated glucose, oleate (an unsaturated fatty acid) promotes β-cell proliferation, while palmitate (a saturated fatty acid) does not have the same effect. The underlying mechanisms, however, were not known, which motivated the study. To unravel the mechanisms, the authors exposed isolated rat islets to high levels of glucose plus either oleate or palmitate (or vehicle) for 48 h. They then used single-cell RNA sequencing to investigate effects on islet transcriptome and flow cytometry to investigate β-cell proliferation. They then used unsupervised clustering to look for subclusters with different functions. One cluster (β2) had proliferating β-cells (the others had different activities). As expected, β-cell proliferation increased in the presence of oleate but not palmitate. Investigating gene expression, they found that both fatty acids enhanced energy metabolism and mitochondrial activity. By comparing proliferating β-cells and nonproliferating β-cells, they then found that ROS and peroxiredoxin signaling are potentially important in β-cell proliferation. Unspecific and specific blockers effectively confirmed a key role for ROS signaling via peroxiredoxin activation in oleate-induced β-cell proliferation. “Our findings suggest that ROS and peroxiredoxin signaling . . . are required for oleate-induced β-cell proliferation in rat islets, providing important information on the molecular mechanisms of β-cell adaptation to nutrient excess,” they write.

Plot of pooled β-cells showing five different β-cell clusters

Plot of pooled β-cells showing five different β-cell clusters

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Vivoli et al. Single-cell RNA sequencing reveals a role for reactive oxygen species and peroxiredoxins in fatty acid–induced rat β-cell proliferation. Diabetes 2023;72:45–58

A series of “unconventional” CD4+ T cell epitopes may exist that additionally contribute to the loss of tolerance in autoimmune type 1 diabetes, according to Guyer et al. (p. 85). While the development of type 1 diabetes is usually attributed to the appearance of islet autoantibodies, there is additional loss of T-cell tolerance to insulin and other islet antigens. For several reasons, there has been suspicion that T-cell epitopes exist beyond those for the “classical” antigens, prompting the investigation. The authors, who initially used a screening algorithm, identified 145 candidate peptides for subsequent binding assays. Thirty-five candidate peptides had detectable binding. Of those, the authors then identified three peptides with the required immunogenicity (i.e., toward CD4+ T cells). One peptide, termed CCNI, was novel (i.e., a neoepitope), while the other two, PCSK2 and UCN3, were conventional epitopes but were also considered unconventional in that they usually show immunogenicity for CD8+ T cells. In T-cell expansions from 15 individuals with type 1 diabetes, 8 showed positive staining for UCN3, 4 had positive staining for PCSK2, and 5 had positive staining for CCNI. They could also be detected in nearly all peripheral blood samples of 10 individuals with type 1 diabetes. T cells targeting the different epitopes had various phenotypes, with those targeting CCNI and PCSK2 having a Th1-like surface phenotype and those targeting UCN3 a Th2-like surface phenotype. The combined frequency of epitope-specific T cells was significantly higher in individuals with type 1 diabetes than in matched control participants. The authors note that frequencies of T cells for the individual epitopes mostly trended toward being higher in type 1 diabetes, but there was no significant difference compared with controls. T cells that recognize PCSK2 could also be detected in islets from a Network for Pancreatic Organ Donors with Diabetes (nPOD) pancreas donor, which the authors say supports the relevance of the three epitopes in disease. “Collectively, our findings introduce UCN3 as a potentially relevant CD4+ T-cell antigen and strongly suggest that the CCNI splice variant and PCSK2 are important T-cell targets in type 1 diabetes,” they write.

Heat map showing variable frequencies of T cells specific for novel epitopes in 10 subjects with type 1 diabetes.

Heat map showing variable frequencies of T cells specific for novel epitopes in 10 subjects with type 1 diabetes.

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Guyer et al. Recognition of mRNA splice variant and secretory granule epitopes by CD4+ T cells in type 1 diabetes. Diabetes 2023;72:85–96

Sustained upregulation of NADPH oxidase 4, or Nox4, in retinal endothelial cells appears likely to result in long-term retinal blood vessel damage in diabetic retinopathy, according to Tang et al. (p. 112). The effect appears to be mediated, at least in part, by reactive oxygen species (ROS) and damage to mitochondria. The findings potentially set up Nox4 as a target for therapy to reduce or reverse the progression of damage after diabetic retinopathy onset. The authors caution, however, that it remains to be seen if this is the case, not least because Nox4 likely interacts with other cell types. The findings come from a series of experiments with a humanized transgenic mouse model with endothelium-specific Nox4 overexpression and an equivalent mouse model with a Nox4 knockout. According to the authors, the mice with Nox4 overexpression did not experience any abnormal development in retinal vasculature. However, after 10–12 months, Nox4 overexpression led to significant retinal vascular damage compared with wild-type controls. By exploring mechanisms (in vitro), they found Nox4 overexpression led to significant mitochondrial ROS generation, increased lipid peroxidation, and decreased mitochondrial membrane potential and respiratory function. Knockout of Nox4 meanwhile reduced ROS generation and damage in mitochondria, reduced endothelial cell apoptosis, and, in the context of streptozotocininduced diabetes in mice, protected the retina from acellular capillary formation and vascular permeability. Given the various effects of overexpression and knockout, the authors point to Nox4 being implicated in how mitochondria handle cellular energy and potentially mitochondrial injury when it is overexpressed, as seems to be the case in diabetes. “Collectively, our study identified targeting endothelial Nox4 as a potential approach for maintaining mitochondrial function, restoring redox balance, and reducing oxidative damage of retinal blood vessels to prevent [diabetic retinopathy] progression,” they write. While the authors do discuss potential underlying mechanisms, they do not explore any actual treatment routes for reducing Nox4 activity and indeed caution that it remains to be seen if targeting Nox4 has any beneficial effects.

Leakage from blood vessels. Shown are wild-type (WT) mice (top) and transgenic (TG) mice (bottom).

Leakage from blood vessels. Shown are wild-type (WT) mice (top) and transgenic (TG) mice (bottom).

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Tang et al. Sustained upregulation of endothelial Nox4 mediates retinal vascular pathology in type 1 diabetes. Diabetes 2023;72:112–125

Early preclinical data suggest that the diphenylbutylpiperidine (DPBP) drug class has potent glucose-lowering actions in the context of obesity and type 2 diabetes. According to Tabatabaei Dakhili et al. (p. 126), there may be a case for drug class repurposing, although careful monitoring will be needed to avoid ketoacidosis. The DPBP drug class was originally approved as a treatment for schizophrenia and other mental health disorders in the 1960s. Crucially, it was via their action as dopamine 2 receptor antagonists that they originally received approval as treatments in mental disorders. However, in this case the authors have noted they also have a completely different potential mechanism of action: inhibition of succinyl-CoA:3-ketoacid CoA transferase, or SCOT, which happens to be the rate-limiting enzyme of ketone oxidation. They previously noted that elevated levels of SCOT activity contribute to hyperglycemia and that the DPBP drug pimozide can alleviate hyperglycemia in obesity. Using in silico molecular dynamics modeling, the authors show that SCOT is likely a noncanonical (i.e., not the original) target of at least three members (pimozide, penfluridol, and fluspirilene) of the DPBP drug class. All three drugs improved glycemia in obese mice. However, obese mice with SCOT knocked out failed to improve glucose tolerance following administration of each of the drugs. Further experiments back up the authors’ case for repurposing the drug class toward diabetes, but crucially they also demonstrate that the effects likely have nothing to do with the original purpose of the drug class as dopamine 2 receptor antagonists. Both acute and chronic treatment with lurasidone, a structurally unrelated dopamine 2 receptor antagonist, failed to have glucose-lowering actions in obese mice. “The DPBPs appear to be relatively safe in humans, where they have been previously approved for the treatment of schizophrenia/psychosis,” they write, concluding that “this drug class may have utility in being repurposed for the treatment of [type 2 diabetes], though careful monitoring of circulating ketones will be necessary. ”

Structural comparison of pimozide (top) and lurasidone (bottom).

Structural comparison of pimozide (top) and lurasidone (bottom).

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Tabatabaei Dakhili et al. The antipsychotic dopamine 2 receptor antagonist diphenylbutylpiperidines improve glycemia in experimental obesity by inhibiting succinyl-CoA:3-ketoacid CoA transferase. Diabetes 2023;72:126–134

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