Biallelic variants in CHST3 cause Spondyloepiphyseal dysplasia with joint dislocations in three Pakistani kindreds.

BACKGROUND: Skeletal dysplasia is a heterogeneous group of disorders. Spondyloepiphyseal dysplasias comprise one subgroup. Deficiency of carbohydrate sulfotransferase 3 has been reported in a small number of patients with recessively inherited spondyloepiphyseal dysplasia with joint dislocation, short stature and scoliosis. We report here molecular and clinical findings of affected individuals in three consanguineous Pakistani families. Affected individuals in all three families had a uniform phenotype including severe short stature, multiple dislocated joints, progressive scoliosis and facial dysmorphism. METHODS: Clinical evaluation was done for three unrelated families. Radiological survey of bones was completed for patients from two of the families. Whole exome sequencing index patients from each family was performed followed by Sanger sequencing for validation of segregation of identified variants in respective families. In-silico analysis for determining pathogenicity of identified variants and conservation was done. RESULTS: Whole-exome sequencing revealed biallelic variants c.590 T > C;p.(Leu197Pro), c.603C > A;p.(Tyr201Ter) and c.661C > T;p.(Arg221Cys) in CHST3 (NM_004273.5) in the three families with eight, five and two affected individuals, respectively. Contrary to previous reports, affected individuals in none of the families exhibited a hearing loss. CONCLUSION: We describe genotypic and phenotypic findings of three unrelated families with spondyloepiphyseal dysplasia. Our study confirms phenotypic variability and adds to the genotypic spectrum of spondyloepiphyseal dysplasia.

Correction to: Novel mutation G324C in WNT1 mapped in a large Pakistani family with severe recessively inherited Osteogenesis Imperfecta.

In the original publication of this article [1], there are two errors in the article which the cDNA position of the pathogenic variant WNT1 p.Gly324Cys should be c.970G>T instead of c.1168G>T.

Novel mutation G324C in WNT1 mapped in a large Pakistani family with severe recessively inherited Osteogenesis Imperfecta.

INTRODUCTION: Osteogenesis imperfecta (OI) is a clinically and genetically heterogeneous disease with skeletal fragility and variable extra-skeletal manifestations. To date several point mutations in 18 different genes causing different types of OI have been identified. Mutations in WNT1 compromise activity of the osteoblasts leading to disturbed bone mass accrual, fragility fractures and progressive skeletal abnormalities. The present study was conducted to determine the underlying genetic cause of an autosomal recessive skeletal dysplasia in a large consanguineous family from Chinute, Pakistan. MATERIALS AND METHODS: Blood was collected from 24 individuals of affected family along with clinical data. Homozygosity mapping was performed to confirm consanguinity. SNPs were identified, followed by whole exome and Sanger sequencing. In silico characterization of WNT1 mutation was performed using multiple platforms. RESULTS: Nine affected family members exhibited severe bone deformities, recurrent fractures, short stature and low bone mineral density. SNP array data revealed homozygous segments > 1 Mb in length accounting for 2.1-12.7% of the genome in affected individuals and their siblings and a single 6,344,821 bp homozygous region in all affected individuals on chromosome 12q12-q13. This region includes two potential OI candidate genes WNT1 and VDR. We did whole-exome sequencing for both genes in two patients and identified a novel damaging missense mutation in exon 4 of WNT1: c.1168G > T (NM_005430) resulting in p.G324C. Sanger sequencing confirmed segregation of mutation with the disease in family. CONCLUSION: We report a novel mutation responsible for OI and our investigation expands the spectrum of disease-causing WNT1 mutations and the resulting OI phenotypes.

Biallelic variants in four genes underlying recessive osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is an inherited heterogeneous rare skeletal disorder characterized by increased bone fragility and low bone mass. The disorder mostly segregates in an autosomal dominant manner. However, several rare autosomal recessive and X-linked forms, caused by mutations in 18 different genes, have also been described in the literature. Here, we present five consanguineous families segregating OI in an autosomal recessive pattern. Affected individuals in the five families presented severe forms of skeletal deformities. It included frequent bone fractures with abnormal healing, short stature, facial dysmorphism, osteopenia, joint laxity, and severe scoliosis. In order to search for the causative variants, DNA of at least one affected individual in three families (A-C) were subjected to whole exome sequencing (WES). In two other families (D-E), linkage analysis using highly polymorphic microsatellite markers was followed by Sanger sequencing. Sequence analysis revealed two novels and three previously reported disease-causing variants. The two novel homozygous variants including [c.824G > A; p.(Cys275Tyr)] in the SP7 gene and [c.397C > T, p.(Gln133*)] in the SERPINF1 gene were identified in families A and B, respectively. The three previously reported homozygous variants including [c.497G > A; p.(Arg166His)] in the SPARC gene, (c.359-3C > G; intron 2) and [c.677C > T; p.(Ser226Leu)] in the WNT1 gene were identified in family C, D, and E. In conclusion, our findings provided additional evidence of involvement of homozygous sequence variants in the SP7, SERPINF1, SPARC and WNT1 genes causing severe OI. It also highlights the importance of extensive genetic investigations to search for the culprit gene in each case of skeletal deformity.

Recessive multiple epiphyseal dysplasia - Clinical characteristics caused by rare compound heterozygous SLC26A2 genotypes.

Pathogenic sequence variants in the solute carrier family 26 member 2 (SLC26A2) gene result in lethal (achondrogenesis Ib and atelosteogenesis II) and non-lethal (diastrophic dysplasia and recessive multiple epiphyseal dysplasia, rMED) chondrodysplasias. We report on two new patients with rMED and very rare compound heterozygous mutation combinations in non-consanguineous families. Patient I presented in childhood with waddling gait and joint stiffness. Radiographs showed epiphyseal changes, bilateral coxa plana-deformity and knee valgus deformity, for which he underwent surgeries. At present 33 years his height is 165 cm. Patient II presented with cleft palate, small jaw, short limbs, underdeveloped thumbs and on radiographs, cervical kyphosis with an underdeveloped C4. He also developed severe scoliosis but has grown at -2.9 SD curve. Molecular analysis revealed that patient I is heterozygous for two known pathogenic variants in SLC26A2, a splice site variant c.-26+2T > C and a missense variant c.1957T > A (p.Cys653Ser), while patient II is compound heterozygous for missense variants c.835C > T (p.Arg279Trp) and c.1535C > A (p.Thr512Lys). These patients further elucidate the variability of the phenotypic and genetic presentations of rMED.

A Novel Homozygous Frameshift Variant in XYLT2 Causes Spondyloocular Syndrome in a Consanguineous Pakistani Family.

We report on three new patients with spondyloocular syndrome (SOS) in a consanguineous Pakistani family. All three patients present progressive generalized osteoporosis, short stature, recurrent fractures, hearing loss and visual impairments. WES revealed a novel homozygous frameshift variant in exon 11 of XYLT2 (NG 012175.1, NP_071450.2) resulting in loss of evolutionary conserved amino acid sequences (840 - 865/865) at C-terminus p.R840fs∗115. Sanger Sequencing confirmed the presence of the novel homozygous mutation in all three patients while the parents were heterozygous carriers of the mutation, in accordance with an autosomal recessive inheritance pattern. Only nine variants worldwide have previously been reported in XYLT2 in patients with SOS phenotype. These three patients with novel homozygous variant extend the genotypic and phenotypic spectrum of SOS.

Osteoporosis and skeletal dysplasia caused by pathogenic variants in SGMS2.

Mechanisms leading to osteoporosis are incompletely understood. Genetic disorders with skeletal fragility provide insight into metabolic pathways contributing to bone strength. We evaluated 6 families with rare skeletal phenotypes and osteoporosis by next-generation sequencing. In all the families, we identified a heterozygous variant in SGMS2, a gene prominently expressed in cortical bone and encoding the plasma membrane-resident sphingomyelin synthase SMS2. Four unrelated families shared the same nonsense variant, c.148C>T (p.Arg50*), whereas the other families had a missense variant, c.185T>G (p.Ile62Ser) or c.191T>G (p.Met64Arg). Subjects with p.Arg50* presented with childhood-onset osteoporosis with or without cranial sclerosis. Patients with p.Ile62Ser or p.Met64Arg had a more severe presentation, with neonatal fractures, severe short stature, and spondylometaphyseal dysplasia. Several subjects had experienced peripheral facial nerve palsy or other neurological manifestations. Bone biopsies showed markedly altered bone material characteristics, including defective bone mineralization. Osteoclast formation and function in vitro was normal. While the p.Arg50* mutation yielded a catalytically inactive enzyme, p.Ile62Ser and p.Met64Arg each enhanced the rate of de novo sphingomyelin production by blocking export of a functional enzyme from the endoplasmic reticulum. SGMS2 pathogenic variants underlie a spectrum of skeletal conditions, ranging from isolated osteoporosis to complex skeletal dysplasia, suggesting a critical role for plasma membrane-bound sphingomyelin metabolism in skeletal homeostasis.

SGCD Homozygous Nonsense Mutation (p.Arg97∗) Causing Limb-Girdle Muscular Dystrophy Type 2F (LGMD2F) in a Consanguineous Family, a Case Report.

Background: Limb-girdle muscular dystrophy (LGMD) is an increasingly heterogeneous category of inherited muscle diseases, mainly affecting the muscles of shoulder areas and the hip, segregating in both autosomal recessive and dominant manner. To-date, thirty-one loci have been identified for LGMD including seven autosomal dominant (LGMD type 1) and twenty four autosomal recessive (LGMD type 2) inherited loci. Methodology/Laboratory Examination: The present report describes a consanguineous family segregating LGMD2F in an autosomal recessive pattern. The affected individual is an 11-year-old boy having two brothers and a sister. Direct targeted next generation sequencing was performed for the single affected individual (VI-1) followed by Sanger sequencing. Results: Targeted next generation sequencing revealed a novel homozygous nonsense mutation (c.289C>T; p.Arg97∗) in the exon 3 of the delta-sarcoglycan (SGCD) gene, that introduces a premature stop codon (TCA), resulting in a nonsense mediated decay or a truncated protein product. Discussion and Conclusion: This is the first report of LGMD2F caused by an SGCD variant in a Pakistani population. The mutation identified in the present investigation extends the body of evidence implicating the gene SGCD in causing LGMD2F and might help in genetic counseling, which is more important to deliver the risk of carrier or affected in the future pregnancies.

Pharmacological evaluation of Rhazya stricta root extract / Evaluación farmacológica del extracto de raíz de Rhazya stricta

The present study aimed to screen the Rhazya stricta Decne root for its antihyperglycemic and antioxidants potential through invitro assays along with phytochemical and elemental analyses. The crude extract was prepared through maceration and fractionated using solvent-solvent extraction technique. The spectroscopic studies indicated the presence of various phytochemical classes in the extract and its fractions. The antioxidant assays showed notable results along with a good concentration of phenolic and flavonoid contents. Enzyme inhibition assays demonstrated glucose-lowering effects by inhibiting the enzyme activity which could reduce post-prandial blood glucose level. The Dipeptidyl peptidase-IV (DPP-IV) inhibition assay results showed the novel DPP-IV inhibition activity of the plant extract and all fractions showed noteworthy enzyme inhibition and antihyperglycemic activity. Conclusively, the Rhazya stricta root extract displayed its antioxidant and antihyperglycemic potential due to the presence of various classes of phytochemicals and micro-nutrients.

El presente estudio tuvo como objetivo examinar la raíz de Rhazya stricta Decne por su potencial antihiperglicémico y antioxidante a través de ensayos in vitro junto con análisis fitoquímicos y elementales. El extracto crudo se preparó por maceración y se fraccionó usando una técnica de extracción solvente-solvente. Los estudios espectroscópicos indicaron la presencia de varias clases fitoquímicas en el extracto y sus fracciones. Los ensayos antioxidantes mostraron resultados notables junto con una importante concentración de contenido fenólico y flavonoide. Los ensayos de inhibición enzimática demostraron efectos reductores de la glucosa al inhibir la actividad enzimática que podría reducir el nivel de glucosa posprandial en sangre. Los resultados del ensayo de inhibición de Dipeptidyl peptidase-IV (DPP-IV) mostraron la nueva actividad de inhibición de DPP-IV del extracto de la planta y todas las fracciones mostraron una notable inhibición enzimática y actividad antihiperglicémica. En conclusión, el extracto de raíz de Rhazya stricta Decne mostró su potencial antioxidante y antihiperglicémico debido a la presencia de varias clases de fitoquímicos y micronutrientes.