•Generation of LGMDR1 patient specific iPSC lines as cellular models.•LGMDR1 patient iPSCs differentiate to muscle in vitro and in vivo.•CAPN3 isoforms can switch during in vitro myogenic ...differentiation of iPSCs.•Reduced DMD expression in LGMDR1 patient-derived cultures.
Limb-girdle muscular dystrophy recessive 1 (LGMDR1) represents one of the most common types of LGMD in the population, where patients develop a progressive muscle degeneration. The disease is caused by mutations in calpain 3 gene, with over 500 mutations reported to date. However, the molecular events that lead to muscle wasting are not clear, nor the reasons for the great clinical variability among patients, and this has so far hindered the development of effective therapies. Here we generate human induced pluripotent stem cells (iPSCs) from skin fibroblasts of 2 healthy controls and 4 LGMDR1 patients with different mutations. The generated lines were able to differentiate into myogenic progenitors and myotubes in vitro and in vivo, upon a transient PAX7 overexpressing protocol. Thus, we have generated myogenic cellular models of LGMDR1 that harbor different CAPN3 mutations within a human genetic background, and which do not derive from muscular biopsies. These models will allow us to investigate disease mechanisms and test therapies. Despite the variability found among iPSC lines that was unrelated to CAPN3 mutations, we found that patient-derived myogenic progenitors and myotubes express lower levels of DMD, which codes a key protein in satellite cell regulation and myotube maturation.
Purpose
Focused parathyroidectomy has been proven to be a safe technique for the treatment of single-gland primary hyperparathyroidism (PHPT). The CaPTHUS scoring model has been reported to be an ...accurate preoperative diagnostic tool for distinguishing single-gland (SGD) from multiglandular disease (MGD), including preoperative serum calcium and PTH values plus ultrasound and Sestamibi scanning. The purpose of the present study was to validate the CaPTHUS model for the population in southern Europe, since the North American and the European populations show different clinicopathological profiles in PHPT.
Methods
This is a retrospective review of a prospectively maintained database of patients diagnosed with PHPT who underwent surgical treatment in a single referral center. Differences between SGD and MGD groups were analyzed using chi-square and Fisher’s exact tests for categorical variables and Student’s
t
test for continuous variables. Overall diagnostic accuracy of the scoring model was assessed by the area under the receiver operating characteristic (ROC) curve (AUC). A
p
< 0.05 level was accepted as significant.
Results
From January 2001 to November 2014, 241 patients were included in the study, of whom 92.1 % had SGD and 71.8 % had a CaPTHUS score ≥3. SGD was distinguished from MGD (
p
< 0.001) using the dichotomous scoring model based on an AUC value of 0.762. Scores ≥3 had a sensitivity of 76.5 % and a positive predictive value of 96 % for SGD.
Conclusions
Despite good test performance, a CaPTHUS score ≥3 does not discard MGD definitely. Intraoperative adjuncts are still needed to further reduce the risk of missing MGD during selective parathyroidectomy.
Congenital Disorders of Glycosylation (CDG) are scarcely reported from Latin America. We here report on a Mexican mestizo with a multi-systemic syndrome including neurological involvement and a type ...I transferrin (Tf) isoelectric focusing (IEF) pattern. Clinical exome sequencing (CES) showed known compound missense variants in PMM2 c.422G > A (p.R141H) and c.395 T > C (p.I132T), coding for the phosphomanomutase 2 (PMM2). PMM2 catalyzes the conversion of mannose-6-P to mannose-1-P required for the synthesis of GDP-Man and Dol-P-Man, donor substrates for glycosylation reactions. This is the third reported Mexican CDG patient and the first with PMM2-CDG. PMM2 has been recently identified as one of the top 10 genes carrying pathogenic variants in a Mexican population cohort.
Induced pluripotent stem cells (iPSC) offer an unprecedented opportunity to model human disease in relevant cell types, but it is unclear whether they could successfully model age‐related diseases ...such as Parkinson's disease (PD). Here, we generated iPSC lines from seven patients with idiopathic PD (ID‐PD), four patients with familial PD associated to the G2019S mutation in the Leucine‐Rich Repeat Kinase 2 (LRRK2) gene (LRRK2‐PD) and four age‐ and sex‐matched healthy individuals (Ctrl). Over long‐time culture, dopaminergic neurons (DAn) differentiated from either ID‐PD‐ or LRRK2‐PD‐iPSC showed morphological alterations, including reduced numbers of neurites and neurite arborization, as well as accumulation of autophagic vacuoles, which were not evident in DAn differentiated from Ctrl‐iPSC. Further induction of autophagy and/or inhibition of lysosomal proteolysis greatly exacerbated the DAn morphological alterations, indicating autophagic compromise in DAn from ID‐PD‐ and LRRK2‐PD‐iPSC, which we demonstrate occurs at the level of autophagosome clearance. Our study provides an iPSC‐based in vitro model that captures the patients' genetic complexity and allows investigation of the pathogenesis of both sporadic and familial PD cases in a disease‐relevant cell type.
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