Mutational spectrum of autosomal recessive limb-girdle muscular dystrophies in a cohort of 112 Iranian patients and reporting of a possible founder effect

Background: Limb-girdle muscular dystrophies are a group of genetically heterogeneous diseases that are inherited in both autosomal dominant (LGMDD) and autosomal recessive forms (LGMDR), the latter is more common especially in populations with high consanguineous marriages like Iran. In the present study, we aimed to investigate the genetic basis of patients who are suspicious of being affected by LGMDR. DNA samples of 60 families suspected of LGMD were extracted from their whole blood. Four short tandem repeat (STR) markers for each candidate genes related to LGMD R1 (calpain3 related)- R6 (δ-sarcoglycan-related) were selected, and all these 24 STRs were applied in two sets of multiplex PCR. After autozygosity mapping, Sanger sequencing and variant analysis were done. Predicting identified variants' effect was performed using in-silico tools, and they were interpreted according to the American College of Medical Genomics and Genetics (ACMG) guideline. MLPA was used for those patients who had large deletions. Fresh muscle specimens were taken from subjects and were evaluated using the conventional panel of histochemical stains. Results: forty out of sixty families showed homozygote haplotypes in CAPN3, DYSF, SGCA, and SGCB genes. The exons and intron-exon boundaries of the relevant genes were sequenced and totally 38 mutations including CAPN3 (n = 15), DYSF (n = 9), SGCB (n = 11), and SGCA (n = 3) were identified. Five out of them were novel. The most prevalent form of LGMDs in our study was calpainopathy followed by sarcoglycanopathy in which beta-sarcoglycanopathy was the most common form amongst them. Exon 2 deletion in the SGCB gene was the most frequent mutation in this study. We also reported evidence of a possible founder effect in families with mutations in DYSF and SGCB genes. We also detected a large consanguineous family suffered from calpainopathy who showed allelic heterogeneity. Conclusions: This study can expand our knowledge about the genetic spectrum of LGMD in Iran, and also suggest the probable founder effects in some Iranian subpopulations which confirming it with more sample size can facilitate our genetic diagnosis and genetic counseling. © 2020 The Author(s)

Hints From the Cellular Functions to the Practical Outlook of Circular RNAs

Although it has been about 30 years since the discovery of circular RNAs (circRNAs) in mammalian cells, these subtypes of RNAs� capabilities have come into focus in recent years. The unique structure and various functional roles of circRNAs in many cellular processes have aroused researchers� interest and raised many questions about whether circRNAs can facilitate the diagnosis and treatment of diseases. To answer these questions, we will illustrate the main known functions and regulatory roles of circRNAs in the cell after presenting a brief history of the discovery of circRNAs and the main proposed theories of the biogenesis of circRNAs. Afterward, the practical application of circRNAs as biomarkers of different pathophysiological conditions will be discussed, mentioning some examples and challenges in this area. We also consider one of the main questions that human beings have always been faced, �the origin of life,� and its possible connection to circRNAs. Finally, focusing on the various capabilities of circRNAs, we discuss their potential therapeutic applications considering the immunity response toward exogenous circRNAs. However, there are still disputes about the exact immune system reaction, which we will discuss in detail. © Copyright © 2021 Yesharim, Mojbafan and Abiri

Reporting a rare form of myopathy, myopathy with extrapyramidal signs, in an Iranian family using next generation sequencing: A case report

Background: Myopathy with extrapyramidal signs (MPXPS) is an autosomal recessive mitochondrial disorder which is caused by mutation in mitochondrial calcium uptake 1 (MICU1) gene located on chromosome 10q22.1. Next Generation Sequencing (NGS) technology is the most effective method for identification of pathogenic variants with the ability to overcome some limitations which Sanger sequencing may encountered. There are few reports on this rare disease around the world and here in this study we first revealed genetic identification of two affected individuals in an Iranian family with a novel mutation. Case presentation: The proband was a 5-year-old girl from consanguenous parents. She was first clinically suspicious of affected with limb-girdle muscular dystrophy (LGMD). Muscle biopsy studies and autozygosity mapping, using four short tandem repeat (STR) markers linked to 6 genes of the most prevalent forms of LGMD, ruled out calpainopathy, dysferlinopathy, and sarcoglycanopathis. DNA sample of the proband was sent for NGS. Whole exome sequencing (WES) revealed a novel mutation c.1295delA in exon 13 of MICU1 gene. This homozygous deletion creates a frameshift and a premature stop codon downstream of canonical EF4 calcium binding motif of MICU1. According to the American College of Medical Genetics and Genomics (ACMG) guidline for sequence interpretation, this variant was a pathogenic one. Sanger sequencing in all family members confirmed the results of the WES. Conclusions: This study was the first report of MPXPS in Iranian population which also revealed a novel mutation in the MICU1 gene. © 2020 The Author(s)

Isolated hypercholesterolemia leads to steatosis in the liver without affecting the pancreas

Abstract Background Lipid accumulation in the liver and pancreas is primarily caused by combined hyperlipidemia. However, the effect of isolated hypercholesterolemia without hypertriglyceridemia is not fully described. Therefore, our aim was to investigate whether hypercholesterolemia alone leads to alterations both in hepatic and pancreatic lipid panel and histology in rats. Methods Male Wistar rats were fed with 2% cholesterol +0.25% cholate-supplemented diet or standard chow for 12 weeks. Blood was collected at weeks 0, 4, 8 and 12 to measure serum cholesterol and triglyceride levels. At week 12, both the pancreas and the liver were isolated for further histological and biochemical analysis. Hepatic and plasma fatty acid composition was assessed by gas chromatography. Expression of mRNA of major enzymes involved in saturated/unsaturated fatty acid synthesis was analyzed by qPCR. In separate experiments serum enzyme activities and insulin levels were measured at week 9. Results At week 12, rats fed with 2% cholesterol +0.25% cholate-supplemented diet were characterized by elevated serum cholesterol (4.09 ± 0.20 vs. 2.89 ± 0.22 mmol/L, *p < 0.05) while triglyceride (2.27 ± 0.05 vs. 2.03 ± 0.03 mmol/L) and glucose levels (5.32 ± 0.14 vs. 5.23 ± 0.10 mmol/L) remained unchanged. Isolated hypercholesterolemia increased hepatic lipid accumulation, hepatic cholesterol (5.86 ± 0.22 vs. 1.60 ± 0.15 ng/g tissue, *p < 0.05) and triglyceride contents (19.28 ± 1.42 vs. 6.78 ± 0.71 ng/g tissue, *p < 0.05), and hepatic nitrotyrosine level (4.07 ± 0.52 vs. 2.59 ± 0.31 ng/mg protein, *p < 0.05). The histology and tissue lipid content of the pancreas was not affected. Serum total protein level, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities remained unchanged in response to isolated hypercholesterolemia while serum alkaline phosphatase activity (ALP) significantly increased. Plasma insulin levels did not change in response to isolated hypercholesterolemia suggesting an intact endocrine function of the pancreas. Isolated hypercholesterolemia caused a significantly increased hepatic and serum fatty acid level associated with a marked alteration of fatty acid composition. Hepatic expression of Δ9-desaturase (SCD1) was increased 4.92×, while expression of Δ5-desaturase and Δ6-desaturase were decreased (0.447× and 0.577×, respectively) due to isolated hypercholesterolemia. Conclusions Isolated hypercholesterolemia leads to hepatic steatosis and marked alterations in the hepatic lipid profile without affecting the pancreas. Altered fatty acid profile might mediate harmful effects of cholesterol in the liver

Molecular genetic study of Calpainopathy in Iran

Introduction: Calpainopathy is an autosomal recessive form of limb girdle muscular dystrophies (LGMDs) caused by mutations in the CAPN3 gene. CAPN3 is a Ca2+-dependent cystein protease consisting of 821 amino acids. LGMD is a highly heterogeneous disorder and mutation identification of this disease by Sanger sequencing of all genes is expensive and time consuming. Using autozygosity mapping is an effective approach to address this issue. Methods: We used two sets of multiplex STR (Short tandem repeat) markers linked to CAPN3, DYSF, SGCA, SGCB, SGCG, SGCD genes following sequencing of the CAPN3 gene. In silico analysis and mutation detection in one hundred ethnically matched healthy individuals were carried out to determine the pathogenicity of novel mutations. Sequence variant interpretation was performed using the American College of Medical Genetics and Genomics (ACMG) guideline. Results: Sixteen out of 50 families linked to the CAPN3 gene. In this study, mutations were found in 14 out of 16 families including 4 novel (c.1894A &gt; T, c.567delG, c.2254-2256delAAC, and c.2373C &gt; T) and 9 previously reported mutations consisting of 5 missense (c.2105C &gt; T, c.2243G &gt; A, c.1714C &gt; T, c.291C &gt; A, c.956C &gt; T), 3 splice site (c.2380 + 2 T &gt; G, c.946-2A &gt; G, c.380G &gt; A), and one indel (c.2257delinsAA) mutations. Discussion: The c.2105C &gt; T was found to be the most frequent mutation in this study. The results of this study revealed that most cases with splicing, frame shift and nonsense mutations experienced more severe clinical manifestations. Nonetheless, this should be confirmed by further studies on larger sample size. © 2018 Elsevier B.V