In This Issue of Diabetes

A new approach to generating pancreatic β-cell–specific gene knockout mice is reported by Ueki et al. (p. 1609). The approach, which is based on CRISPR-Cas9 technology, avoids the expensive and timeconsuming steps usually involved in generating mice with the Cre-loxP system, knocking months off the time needed and potentially leading to a faster discovery process for treatments for β-cell failure and thus insulin needs in patients with diabetes. Called βCas9, the new approach uses mice expressing Cas9 in pancreatic β-cells and adeno-associated virus 8 to deliver genomic DNA via intraperitoneal injection. The approach is widely applicable, but the authors focus on β-cell function in the context of diabetes and demonstrate a process that should help with research in the area. According to the authors, they crossbred LSL-Cas9-P2A-EGFP mice with Ins-Cre mice to generate the βCas9 mice. Glucose tolerance and insulin secretion were comparable with those of control mice at 11 and 15 weeks, and experiments with isolated islets largely pointed in the same direction. They also found that gene knockout of EGFP with adeno-associated virus 8 delivery of genomic RNA was accurate in terms of targeting β-cells and was highly efficient, at 80%. Notably, the authors also managed to dose the virus and RNA with a simple intraperitoneal injection. As a proof of concept, the authors used the system to target Pdx1, which is a gene linked to maturity-onset diabetes of the young 4. They found that mice with a knockout for that gene had glucose intolerance that was due to β-cells changing to α-cells that expressed glucagon. They note that the system still has several limitations, including issues with efficiency, and the cost-effectiveness may not be superior to that of the original Cre-loxP approach due to issues with purifying the adeno-associated virus. Commenting further, author Yuya Nishida said, “We believe our βCas9 system, which allows us to save time in generating β-cell–specific knockout mice, will significantly advance the research field of diabetes.”

Inhibition of the activity of an enzyme called lysophosphatidylcholine acyltransferase 3, or LPCAT3, might be the basis for a new treatment for insulin resistance and hyperlipidemia, according to He et al. (p. 1547). Specifically, they found that it was possible to target LPCAT3 in adipose tissue to manipulate the level of saturation in membrane phospholipids, particularly polyunsaturated phosphatidylcholines, ultimately resulting in changes in insulin sensitivity. The findings center on the idea that polyunsaturated versions of phosphatidylcholine increase cell membrane fluidity and reduce the size of lipid rafts, where many receptors, such as insulin receptors, are located, thus influencing receptor-mediated signal transduction. After initially identifying LPCAT3 as the major type of lysophosphatidylcholine acyltransferase in adipose tissue, the authors generated adipose-specific Lpcat3 knockout mice to investigate the role of the enzyme in insulin signaling. They found that the deficiency reduced the level of polyunsaturated phosphatidylcholines in adipocyte membranes and resulted in increased insulin sensitivity. Moreover, there was higher LPCAT3 activity in adipose tissue in obese and diabetic mouse models, and this activity made a critical contribution to insulin resistance. They also found that treatment of human or mouse adipocytes with polyunsaturated phosphatidylcholines could significantly reduce insulin signaling. The authors suggest that LPCAT3 deficiency–mediated enlargement of lipid rafts can recruit more insulin receptors, thereby promoting insulin signaling. Despite some limitations and the need for further research, the authors say the findings, along with those of previous studies, suggest that inhibition of LPCAT3 activity can be used to treat insulin resistance. They note that various aspects of the underlying mechanisms that might be involved and other issues deserve further investigation. Commenting further, author Xian-Cheng Jiang said, “Given the combined results of LPCAT3 studies in the small intestine, liver, muscle, and adipose tissue, we believe that inhibition of LPCAT3 activity could be a novel approach for treatment of metabolic diseases, including hyperlipidemia and insulin resistance.”

The loss of size selectivity in the glomerular barrier is an early marker of glomerular injury in diabetes, according to Saulnier et al. (p. 1682). There was also a significant association between the severity of such an injury and progression to kidney failure, independent of albuminuria and glomerular filtration rate. According to the authors, this suggests that uncovering the molecular mechanisms underlying such early changes in barrier function might eventually reveal novel therapeutic targets for diabetic kidney disease in type 2 diabetes. The findings come from a longitudinal study of 185 American Indians with type 2 diabetes who underwent detailed study of kidney function over many years. Specifically, measurements included glomerular filtration rate and effective renal plasma flow as well as assessment of dextran sieving characteristics of the glomerular capillary. The authors used modeling to examine the hazards for developing kidney failure according to alterations in the size-selective properties of glomerular capillary and after adjusting for a series of confounding factors. They found that of the 185 participants, 67 developed kidney failure over ~18 years of follow-up. After adjusting for confounding, they found that increments in a specific measure of kidney function and filtration capabilities (termed ω₀) were associated with higher risk of kidney failure in later life, with a hazard ratio of 1.55 (95% CI 1.17, 2.05). The risk was also independent of albuminuria and glomerular filtration rate, suggesting that impaired selectivity is an independent determinant of diabetic kidney disease progression. The authors note that there could be concerns around generalizability based on their study population. However, they point out that many of the diabetes characteristics and the clinical course seen in the population consistently replicate in other populations, meaning their findings could have general implications in wider type 2 diabetes populations.

Meteorin-like hormone, or Metrnl, appears to accelerate wound healing in both healthy and diabetic mice, according to Song et al. (p. 1692). Specifically, it appears that Metrnl is produced in keratinocytes and macrophages upon wounding (although less so when diabetes is present) and that topical application of a recombinant version of the cytokine appears to aid wound healing irrespective of diabetes status. According to the authors, this finding suggests that a hydrogel with Metrnl can be used in minor or nonsevere diabetic wound cases. However, they caution that it is too early to know its effectiveness in more serious wounds, such as diabetic foot ulcers with exposed bone or osteomyelitis, and more research is needed before it has clinical application. The findings come from experiments with mice, mouse cells, and human tissue samples that broadly investigated the role of Metrnl in wounding and wound healing and specifically whether a recombinant version in a hydrogel could improve healing. They found that Metrnl expression did increase during physiological wound healing, but the increase was notably less in the presence of diabetes. Topical application of recombinant Metrnl also accelerated wound closure in normal models and various models of diabetes in mice compared with controls. They note that treated and untreated wounds both showed a trend toward healing that does not necessarily replicate diabetic wounds, which tend not to close in clinical cases. Mechanistic studies revealed that keratinocytes secrete Metrnl, which has multiple roles in promoting angiogenesis and epithelialization via cell proliferation, migration, tube formation, and macrophage M2 polarization. In the latter case, they found such macrophages also secrete Metrnl to further stimulate angiogenesis and that Metrnl appears to drive angiogenesis and reepithelialization (i.e., wound healing) via a specific AKT phosphorylation that is dependent on KIT receptor tyrosine kinase.