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R-spondin-1 (Rspol) is an intestinal growth factor known to exert its effects through activation of the canonical Wnt (cWnt) signaling pathway and subsequent expression of cWnt target genes. We have detected Rspol mRNA in murine islets and the murine MIN6 and βTC 18-cell lines, and Rspol protein in MIN6 β-cells. Rspol activated cWnt signaling in MIN6 β-cells by increasing nuclear β-catenin and c-myc, a cWnt target gene. Rspol also induced insulin mRNA expression in MIN6 cells. Analysis of MIN6 and mouse β-cell proliferation by [³H]thymidine and BrdU incorporation, respectively, revealed that Rspol stimulated cell growth. Incubation of MIN6 and mouse B-cells with cytokines (IL1β/TNFα/interferon-γ) significantly increased cellular apoptosis; this increase was abolished by pretreatment with Rspol. Rspol also stimulated insulin secretion in a glucose-independent fashion. We further demonstrated that the glucagon-like peptide-1 receptor agonist, exendin4 (EX4), stimulated Rspol mRNA transcript levels in MIN6 cells in a glucose-, time-, dose-, and PI3-kinase-dependent fashion. This effect was not limited to this B-cell line, as similar time-dependent increases in Rspol were also observed in the βTC β-cell line and mouse islets in response to EX4 treatment. Together, these studies demonstrate that Rspol is a novel β-cell growth factor and insulin secretagogue that is regulated by EX4. These findings suggest that Rspol and the cWnt signaling pathway may serve as a novel target to enhance 18-cell growth and function in patients with type 2 diabetes. [ABSTRACT FROM AUTHOR]

Inhibition of histone deacetylase 6 (HDAC6) was shown to support axon growth on the nonpermissive substrates myelinassociated glycoprotein (MAG) and chondroitin sulfate proteoglycans (CSPGs). Though HDAC6 deacetylates a-tubulin, we find that another HDAC6 substrate contributes to this axon growth failure. HDAC6 is known to impact transport of mitochondria, and we show that mitochondria accumulate in distal axons after HDAC6 inhibition. Miro and Milton proteins link mitochondria to motor proteins for axon transport. Exposing neurons to MAG and CSPGs decreases acetylation of Miro1 on Lysine 105 (K105) and decreases axonal mitochondrial transport. HDAC6 inhibition increases acetylated Miro1 in axons, and acetyl-mimetic Miro1 K105Q prevents CSPG-dependent decreases in mitochondrial transport and axon growth. MAG- and CSPG-dependent deacetylation of Miro1 requires RhoA/ROCK activation and downstream intracellular Ca2+ increase, and Miro1 K105Q prevents the decrease in axonal mitochondria seen with activated RhoA and elevated Ca2+. These data point to HDAC6-dependent deacetylation of Miro1 as a mediator of axon growth inhibition through decreased mitochondrial transport. [ABSTRACT FROM AUTHOR]

Background: Neuropathic pain impairs quality of life, is widely prevalent, and incurs significant costs. Current pharmacological therapies have poor/no efficacy and significant adverse effects; safe and effective alternatives are needed. Hyperpolarisation-activated cyclic nucleotide-regulated (HCN) channels are causally implicated in some forms of peripherally mediated neuropathic pain. Whilst 2,6-substituted phenols, such as 2,6-di-tert-butylphenol (26DTB-P), selectively inhibit HCN1 gating and are antihyperalgesic, the development of therapeutically tolerable, HCN-selective antihyperalgesics based on their inverse agonist activity requires that such drugs spare the cardiac isoforms and do not cross the blood-brain barrier., Methods: In silico molecular dynamics simulation, in vitro electrophysiology, and in vivo rat spared nerve injury methods were used to test whether 'hindered' variants of 26DTB-P (wherein a hydrophilic 'anchor' is attached in the para-position of 26DTB-P via an acyl chain 'tether') had the desired properties., Results: Molecular dynamics simulation showed that membrane penetration of hindered 26DTB-Ps is controlled by a tethered diol anchor without elimination of head group rotational freedom. In vitro and in vivo analysis showed that BP4L-18:1:1, a variant wherein a diol anchor is attached to 26DTB-P via an 18-carbon tether, is an HCN1 inverse agonist and an orally available antihyperalgesic. With a CNS multiparameter optimisation score of 2.25, a >100-fold lower drug load in the brain vs blood, and an absence of adverse cardiovascular or CNS effects, BP4L-18:1:1 was shown to be poorly CNS penetrant and cardiac sparing., Conclusions: These findings provide a proof-of-concept demonstration that anchor-tethered drugs are a new chemotype for treatment of disorders involving membrane targets., (Copyright © 2023 British Journal of Anaesthesia. Published by Elsevier Ltd. All rights reserved.)

Charcot-Marie-Tooth type 2A (CMT2A) peripheral neuropathy, the most common axonal form of CMT, is caused by dominantly inherited point mutations in the Mitofusin 2 (Mfn2) gene. It is characterized by progressive length-dependent degeneration of motor and sensory nerves with corresponding clinical features of motor and sensory impairment. There is no cure for CMT, and therapeutic approaches are limited to physical therapy, orthopedic devices, surgery, and analgesics. In this study we focus on histone deacetylase 6 (HDAC6) as a therapeutic target in a mouse model of mutant MFN2 (MFN2 R94Q )-induced CMT2A. We report that these mice display progressive motor and sensory dysfunction as well as a significant decrease in α-tubulin acetylation in distal segments of long peripheral nerves. Treatment with a new, highly selective HDAC6 inhibitor, SW-100, was able to restore α-tubulin acetylation and ameliorate motor and sensory dysfunction when given either prior to or after the onset of symptoms. To confirm HDAC6 is the target for ameliorating the CMT2A phenotype, we show that genetic deletion of Hdac6 in CMT2A mice prevents the development of motor and sensory dysfunction. Our findings suggest α-tubulin acetylation defects in distal parts of nerves as a pathogenic mechanism and HDAC6 as a therapeutic target for CMT2A., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Objective: The cWnt activator, R-spondin1 (Rspo1), regulates β-cell growth, function, and neogenesis, although its role in conditions such as streptozotocin (STZ)-induced diabetes is unknown. We hypothesized that Rspo1 deficiency enhances β-cell neogenesis in STZ-induced diabetes., Methods: Wild-type (Rspo1) and knockout (Rspo1) mice were injected with STZ (40 mg/kg) for 5 days, followed by analysis of oral glucose and insulin tolerance, and were killed on day 6 (acute; 9-11 mice) or 32 (chronic; 11-16 mice). Immunohistochemistry was performed for β-cell apoptosis, proliferation, neogenesis, and markers of β-cell maturity., Results: There was no difference in oral glucose handling between STZ-induced Rspo1 and Rspo1 mice, although Rspo1 mice demonstrated increased insulin sensitivity. β-cell mass, islet number, and islet size distribution did not differ between STZ-induced Rspo1 and Rspo1 mice, but Rspo1 animals had reduced β-cell apoptosis and increased numbers of insulin-positive ductal cells, indicating β-cell neogenesis. Furthermore, the increased β-cell regeneration observed in the Rspo1 animals was associated with a more differentiated/mature β-cell phenotype as assessed by increased immunopositivity for Nkx6.1, MafA, and GLUT2., Conclusions: These findings indicate that Rspo1 is a negative regulator of β-cell neogenesis, development, and survival in the face of STZ-induced diabetes, providing a therapeutic target for the enhancement of β-cell mass.