Skeletal muscle and the nervous system depend on efficient protein quality control, and they express chaperones and cochaperones at high levels to maintain protein homeostasis. Mutations in many of these proteins cause neuromuscular diseases, myopathies, and hereditary motor and sensorimotor neuropathies. In this review, we cover mutations in DNAJB6, DNAJB2, αB-crystallin (CRYAB, HSPB5), HSPB1, HSPB3, HSPB8, and BAG3, and discuss the molecular mechanisms by which they cause neuromuscular disease. In addition, previously unpublished results are presented, showing downstream effects of BAG3 p.P209L on DNAJB6 turnover and localization.

To apply in vivo corneal confocal microscopy (IVCM) to study the pathogenesis of keratitis (keratoendotheliitis) fugax hereditaria, an autosomal dominant cryopyrin-associated periodic keratitis, associated with the c.61G>C pathogenic variant in the NLRP3 gene, in its acute and chronic phase, and to report histopathologic findings after penetrating keratoplasty.DESIGN: This was an observational case series. METHODS: The study population included 6 patients during an acute attack, 18 patients in the chronic phase, and 1 patient who underwent penetrating keratoplasty. Interventions included Sanger sequencing for the NLRP3 variant c.61C>G, a clinical examination, corneal photography, IVCM, light microscopy, and immunohistochemistry. Our primary outcome measures included IVCM and histopathologic findings.RESULTS: During the acute attack, hyperreflective cellular structures consistent with inflammatory cells transiently occupied the anterior to middle layers of the corneal stroma. Other corneal layers were unremarkable.With recurring attacks, central oval stromal opacities accumulated. IVCM revealed that they contained long, hyperreflective, needle-shaped structures in the extracellular matrix. Using light microscopy, the anterior half of the stroma displayed thin and finely vacuolated lamellae, and keratocytes throughout the stroma were immunopositive for syndecan.CONCLUSIONS: The acute attacks and chronic stromal deposits mainly involve the anterior to middle layers of the corneal stroma, and the disease is primarily a keratitis rather than a keratoendotheliitis. IVCM shows that inflammatory cells invade only the stroma during an acute attack. IVCM and light microscopic findings suggest that the central corneal opacities represent gradual depo-Supplemental Material available at AJO.com.

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PurposeTo describe four Finnish families with epithelial recurrent erosion dystrophy (ERED) caused by the pathogenic variant c.3156C>T in collagen type XVII alpha 1 chain gene (COL17A1).MethodsEleven affected and two unaffected individuals underwent clinical ophthalmological examination, anterior segment photography, and corneal topography. Two of them underwent phototherapeutic keratectomy (PTK). Genetic analysis included both next‐generation and Sanger sequencing. Specimens from the manual keratectomy of one patient were available for ophthalmic pathologic examination, including immunohistochemistry.ResultsThe common splice‐site altering synonymous variant c.3156C > T, p.(Gly1052=) in COL17A1 was confirmed in 15 individuals with ERED from the four families. Subepithelial corneal scarring grades varied and increased with age, leading to decreased best‐corrected visual acuity. PTK improved vision in 58‐ and 67‐year‐old individuals without reactivating the disease. The keratectomy specimens showed an uneven epithelium and a spectrum of basement membrane abnormalities, including breaks, fragmentation, multiplication and entrapment within the subepithelial scar, reflecting recurrent erosions. The stromal cells consisted of varying proportions of bland and activated fibroblasts and myofibroblasts, reflecting different ages of scars. The family with the largest number of known affected generations originated from Southern Sweden.ConclusionThe phenotype in the Finnish ERED families is consistent with earlier reports of the c.3156C > T variant, although the severity has varied between reports. The phenotype may be modulated by other genes. This study suggests a likely founder effect of the variant in both Finnish and Swedish populations due to their shared population histories. If vision is compromised, PTK can be considered especially in older patients.

AimsTo elucidate the effect ofNLRP3variant c.61G>C on interleukin-1β (IL-1β) secretion in keratitis fugax hereditaria (KFH), a cryopyrin-associated periodic syndrome limited to the eye, and to probe the potential modifying role of prednisolone.MethodsPeripheral blood mononuclear cells (PBMCs) isolated from whole blood of patients with KFH and healthy controls were grown under steady-state conditions or primed with lipopolysaccharide (LPS) with or without prednisolone, and subsequently activated with ATP. Cell lysates and proteins precipitated from the cell culture media were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. NLRP3, procaspase-1, and IL-1β were visualised by western blotting. The concentration of secreted IL-1β in the culture media was quantified by ELISA.ResultsFollowing priming of the NLRP3 inflammasome with LPS, a lower threshold for IL-1β secretion was observed in patient-derived PBMCs, compared with healthy controls (median, 124 vs 10 pg/mL, respectively). Interestingly, in PBMCs derived from patients with frequent KFH symptoms, LPS priming alone was able to trigger substantial IL-1β secretion (median, 522 pg/mL), whereas those of patients experiencing occasional KFH attacks showed a subtler release of IL-1β (median, 85 pg/mL). NLRP3 expression was significantly enhanced with LPS stimulation (p=0.03) whereas procaspase-1 expression was not affected. LPS and ATP treated PBMCs from patients with KFH showed significantly diminished IL-1β secretion with prednisolone treatment (p=0.04).ConclusionsPBMCs from patients with KFH are more prone to secrete IL-1β, confirming the presumption that the c.61G>C is a gain-of-function variant. Furthermore, prednisolone is confirmed as a potent drug to reduce IL-1β secretion in KFH.

Purpose To elucidate by using in vivo corneal confocal microscopy (IVCM) the pathogenesis of keratoendotheliitis fugax hereditaria, an autosomal dominant cryopyrin‐associated periodic keratitis associated with a pathogenic variant in NLRP3, in its acute and chronic phase. Methods Cross‐sectional, hospital‐based study of 7 patients during an acute attack and 18 patients in the chronic phase. Corneal photography, IVCM and Sanger sequencing to confirm NLRP3 variant c.61C>G were performed. Results During the acute attack, the stroma was transiently infiltrated with hyperreflective small roundish bodies consistent with inflammatory cells. Other layers of the cornea were essentially normal. With multiple recurring attacks, a central corneal stromal opacity gradually develops. IVCM revealed that the opacities correspond to hyperreflective needle‐shaped structures in the extracellular matrix of the stroma. Conclusions Keratoendotheliitis fugax hereditaria mainly involves the middle layers of the corneal stroma, and is stromal keratitis rather than a true keratoendotheliitis. IVCM shows that inflammatory cells invade only the stroma during an acute attack. With repeated attacks, needle‐shaped hyperreflective extracellular matrix elements emerge and cause a persistent opacity. The disease will be excellent in vivo human model to study the activation of the NLRP3 inflammasome and the tissue specificity in cryopyrin‐associated periodic syndromes.

PurposeKeratitis fugax hereditaria (KFH) is a rare periodic autoinflammatory corneal disease caused by a heterozygous pathogenic variant c.61G>C in the NLRP3 gene. Other NLRP3 variants cause cryopyrin‐associated periodic syndromes (CAPS), characterized by enhanced production of IL‐1β. While more than 50% of KFH patients use prednisolone to relieve pain, there are no data on how the c.61G>C NLRP3 variant or prednisolone treatment affects IL‐1β production in KFH patients. We carried out a functional study to elucidate these questions.MethodsPeripheral blood mononuclear cells (PBMCs) isolated from whole blood of patients with KFH (n = 8) and healthy controls (n = 4) were grown under steady‐state conditions or exposed to NLRP3 inflammasome priming with LPS (500 ng/ml) together with or without prednisolone (100 µM) and subsequently activating inflammasome with ATP(1 mM). Proteins precipitated from the cell culture media were separated by SDS‐PAGE and transferred to a membrane by western blotting. IL‐1β was visualized from the membranes by immunodetection. Alternatively, the concentration of secreted IL‐1β in the media was quantified by ELISA.ResultsFollowing NLRP3 inflammasome activating stimuli, a reduced threshold for IL‐1β secretion was observed in severely affected KFH patients compared to mildly affected patients or healthy controls. In these patients, LPS priming alone was able to trigger IL‐1β secretion while in the mildly affected patients and healthy controls IL‐1β secretion required a canonical two‐hit activation of the NLRP3 inflammasome by both LPS and ATP. In both KFH and healthy control‐derived PBMCs, IL‐1β secretion declined after prednisolone treatment.ConclusionsOur study demonstrated that patients with severe KFH are more prone to IL‐1β secretion, thus indicating that the c.61G>C is a gain‐of‐function variant. Furthermore, prednisolone is a potent treatment for reducing IL‐1β secretion in KFH.

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