Members of the CUGBP Elav-like family of RNA-binding proteins are expressed in distinct populations of primary sensory neurons.

Primary sensory dorsal root ganglia (DRG) neurons are diverse, with distinct populations that respond to specific stimuli. Previously, we observed that functionally distinct populations of DRG neurons express mRNA transcript variants with different 3' untranslated regions (3'UTRs). 3'UTRs harbor binding sites for interaction with RNA-binding proteins (RBPs) for transporting mRNAs to subcellular domains, modulating transcript stability, and regulating the rate of translation. In the current study, analysis of publicly available single-cell RNA-sequencing data generated from adult mice revealed that 17 3'UTR-binding RBPs were enriched in specific populations of DRG neurons. This included four members of the CUG triplet repeat (CUGBP) Elav-like family (CELF): CELF2 and CELF4 were enriched in peptidergic, CELF6 in both peptidergic and nonpeptidergic, and CELF3 in tyrosine hydroxylase-expressing neurons. Immunofluorescence studies confirmed that 60% of CELF4+ neurons are small-diameter C fibers and 33% medium-diameter myelinated (likely Aδ) fibers and showed that CELF4 is distributed to peripheral termini. Coexpression analyses using transcriptomic data and immunofluorescence revealed that CELF4 is enriched in nociceptive neurons that express GFRA3, CGRP, and the capsaicin receptor TRPV1. Reanalysis of published transcriptomic data from macaque DRG revealed a highly similar distribution of CELF members, and reanalysis of single-nucleus RNA-sequencing data derived from mouse and rat DRG after sciatic injury revealed differential expression of CELFs in specific populations of sensory neurons. We propose that CELF RBPs may regulate the fate of mRNAs in populations of nociceptors, and may play a role in pain and/or neuronal regeneration following nerve injury.

Conditioning electrical stimulation fails to enhance sympathetic axon regeneration.

Peripheral nerve injuries are common, and there is a critical need for the development of novel therapeutics to complement surgical repair. Conditioning electrical stimulation (CES) is a novel variation to the well-studied perioperative electrical stimulation, both of which have displayed success in enhancing the regeneration of motor and sensory axons in an injured peripheral nerve. CES is a clinically attractive alternative not only because of its ability to be performed at the bedside prior to a scheduled nerve repair surgery, but it has also been shown to be superior to perioperative electrical stimulation in the enhancement of motor and sensory regeneration. However, the effects of CES on sympathetic regeneration are unknown. Therefore, we tested the effects of two clinically relevant CES paradigms on sympathetic axon regeneration and distal target reinnervation. Because of the long history of evidence for the enhancement of motor and sensory axons in response to electrical stimulation, we hypothesize that CES will also enhance sympathetic axon regeneration. Our results indicate that the growth of sympathetic axons is acutely inhibited by CES; however, at a longer survival time point post-injury, there is no difference between sham CES and the CES groups. There has been evidence to suggest that the growth of sympathetic axons is inhibited by a conditioning lesion, and that sympathetic axons may respond to electrical stimulation by sprouting rather than elongation. Our data indicate that sympathetic axons may retain some regenerative ability after CES, but no enhancement is exhibited, which may be accounted for by the inability of the current clinically relevant electrical stimulation paradigm to recruit the small-caliber sympathetic axons into activity. Further studies will be needed to optimize electrical stimulation parameters in order to enhance the regeneration of all neuron types.

Sensitive detection of multiple islet autoantibodies in type 1 diabetes using small sample volumes by agglutination-PCR.

Islet autoantibodies are predominantly measured by radioassay to facilitate risk assessment and diagnosis of type 1 diabetes. However, the reliance on radioactive components, large sample volumes and limited throughput renders radioassay testing costly and challenging. We developed a multiplex analysis platform based on antibody detection by agglutination-PCR (ADAP) for the sample-sparing measurement of GAD, IA-2 and insulin autoantibodies/antibodies in 1 µL serum. The assay was developed and validated in 7 distinct cohorts (n = 858) with the majority of the cohorts blinded prior to analysis. Measurements from the ADAP assay were compared to radioassay to determine correlation, concordance, agreement, clinical sensitivity and specificity. The average overall agreement between ADAP and radioassay was above 91%. The average clinical sensitivity and specificity were 96% and 97%. In the IASP 2018 workshop, ADAP achieved the highest sensitivity of all assays tested at 95% specificity (AS95) rating for GAD and IA-2 autoantibodies and top-tier performance for insulin autoantibodies. Furthermore, ADAP correctly identified 95% high-risk individuals with two or more autoantibodies by radioassay amongst 39 relatives of T1D patients tested. In conclusion, the new ADAP assay can reliably detect the three cardinal islet autoantibodies/antibodies in 1µL serum with high sensitivity. This novel assay may improve pediatric testing compliance and facilitate easier community-wide screening for islet autoantibodies.