Aims/hypothesisThe Coxsackie and adenovirus receptor (CAR) is a transmembrane cell-adhesion protein that serves as an entry receptor for enteroviruses and may be essential for their ability to infect cells. Since enteroviral infection of beta cells has been implicated as a factor that could contribute to the development of type 1 diabetes, it is often assumed that CAR is displayed on the surface of human beta cells. However, CAR exists as multiple isoforms and it is not known whether all isoforms subserve similar physiological functions. In the present study, we have determined the profile of CAR isoforms present in human beta cells and monitored the subcellular localisation of the principal isoform within the cells.MethodsFormalin-fixed, paraffin-embedded pancreatic sections from non-diabetic individuals and those with type 1 diabetes were studied. Immunohistochemistry, confocal immunofluorescence, electron microscopy and western blotting with isoform-specific antisera were employed to examine the expression and cellular localisation of the five known CAR isoforms. Isoform-specific qRT-PCR and RNA sequencing (RNAseq) were performed on RNA extracted from isolated human islets.ResultsAn isoform of CAR with a terminal SIV motif and a unique PDZ-binding domain was expressed at high levels in human beta cells at the protein level. A second isoform, CAR-TVV, was also present. Both forms were readily detected by qRT-PCR and RNAseq analysis in isolated human islets. Immunocytochemical studies indicated that CAR-SIV was the principal isoform in islets and was localised mainly within the cytoplasm of beta cells, rather than at the plasma membrane. Within the cells it displayed a punctate pattern of immunolabelling, consistent with its retention within a specific membrane-bound compartment. Co-immunofluorescence analysis revealed significant co-localisation of CAR-SIV with zinc transporter protein 8 (ZnT8), prohormone convertase 1/3 (PC1/3) and insulin, but not proinsulin. This suggests that CAR-SIV may be resident mainly in the membranes of insulin secretory granules. Immunogold labelling and electron microscopic analysis confirmed that CAR-SIV was localised to dense-core (insulin) secretory granules in human islets, whereas no immunolabelling of the protein was detected on the secretory granules of adjacent exocrine cells. Importantly, CAR-SIV was also found to co-localise with protein interacting with C-kinase 1 (PICK1), a protein recently demonstrated to play a role in insulin granule maturation and trafficking.Conclusions/interpretationThe SIV isoform of CAR is abundant in human beta cells and is localised mainly to insulin secretory granules, implying that it may be involved in granule trafficking and maturation. We propose that this subcellular localisation of CAR-SIV contributes to the unique sensitivity of human beta cells to enteroviral infection.Electronic supplementary materialThe online version of this article (10.1007/s00125-018-4704-1) contains peer-reviewed but unedited supplementary material, which is avai...

Summary Iron (ferric carboxymaltose) infusion therapy is used to treat severe iron deficiency which is not responding to the first-line oral iron therapy. However, it can also cause severe renal wasting of phosphate resulting in severe hypophosphataemia in some patients. Despite the growing number of case reports, this side effect is not well known to healthcare professionals. The product labelling information sheet does mention that hypophosphataemia can be a side effect, but also says that this side effect is usually transient and asymptomatic. We report a challenging case of a patient who developed severe, symptomatic and prolonged hypophosphataemia after an intravenous iron infusion for severe iron deficiency. Learning points: Clinicians prescribing ferric carboxymaltose (Ferinject®) should be aware of the common side effect of hypophosphataemia, which could be mild, moderate or severe. Patients receiving iron infusion should be educated concerning this potential side effect. Pre-existing vitamin D deficiency, low calcium levels, low phosphate levels or raised parathyroid hormone levels may be risk factors, and these should be evaluated and corrected before administering intravenous iron. Patients may require phosphate and vitamin D replacement along with monitoring for a long period after iron infusion-induced hypophosphataemia. Every incident should be reported to the designated body so that the true prevalence and management thereof can be ascertained.

Coxsackie B viruses (CV-B) are important pathogens associated with several central nervous system (CNS) disorders. CV-B are mainly transmitted by the faecal-oral route, but there is also evidence for vertical transmission. The outcome of in utero CV-B infections on offspring's CNS is poorly explored. The aim of this study was to investigate vertical transmission of CV-B to the CNS. For this purpose, pregnant Swiss albino mice were intraperitoneally inoculated with CV-B4 E2 at gestational days 10G or 17G. Different CNS compartments were collected and analyzed for virus infection and histopathological changes. Using plaque assays, we demonstrated CV-B4 E2 vertical transmission to offspring's CNS. Viral RNA persisted in the CNS up to 60 days after birth, as evidenced by a sensitive semi-nested(sn) reverse transcripton(RT)-PCR method. This was despite infectious particles becoming undetectable at later time points. Persistence was associated with inflammatory lesions, lymphocyte infiltration and viral dsRNA detected by immunohistochemistry. Offspring born to dams mock- or virus-infected at day 17G were challenged by the same virus at day 21 after birth (-+ and ++ groups, respectively). Sn-RT-PCR and histology results compared between both ++ and -+ groups, show that in utero infection did not enhance CNS infection during challenge of the offspring with the same virus.

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