Exome sequencing revealed variants in SGCA and SIL1 genes underlying limb girdle muscular dystrophy and Marinesco-Sjögren syndrome patients.

BACKGROUND: Inherited neuromuscular (NMD) and neurodegenerative diseases (NDD) belong to two distinct categories that disturb different components of the nervous system, leading to a variety of different symptoms and clinical manifestations. Both NMD and NDD are a heterogeneous group of genetic conditions. Genetic variations in the SGCA and SIL1 genes have been implicated in causing Limb Girdle Muscular Dystrophy (LGMD), a type of neuromuscular disorder, and Marinesco-Sjögren Syndrome (MSS) which is a neurodegenerative disorder. METHODS: In the present study, we have investigated four patients presenting LGMD and five patients with MSS features. After collecting detailed clinical and family history, necessary laboratory investigations, including estimation of a skeletal muscle marker enzyme serum creatine kinase (CK), nerve conduction study (NCS), electromyography (EMG), echocardiography (Echo), Magnetic resonance imaging (MRI -brain), CT-brain and X-rays were performed. Whole exome followed by Sanger sequencing was employed to search for the disease-causing variants. RESULTS: Physical examination in LGMD patients revealed poor muscle tone and facing difficulty in straightening up from the floor. Clinical history revealed frequent falls and strenuousness in climbing stairs. They started toe-walking in early childhood. Laboratory investigations confirmed elevated CK levels and abnormal NCS and EMG. The MSS patients showed abnormalities in gate and jerking movement, abnormal speech, and strabismus with cataract. MRI-brain showed cerebral atrophy in some MSS patients with elevated CK levels. Whole exome sequencing revealed a nonsense variant [c.C574T, p.(Arg192*)] in the SGCA gene and a frameshift [c.936dupG, p.(Leu313AlaFs*39)] in the SIL1 gene in LGMD and MSS patients, respectively. CONCLUSION: Our study emphasizes the significance of integrating clinical and genetic analyses for precise diagnosis and tailored management strategies in inherited NMD and NDD disorders. To the best of our knowledge, this is the first study documenting SGCA and SIL1 recurrent variants in subcontinent populations with few rare clinical features. The recurrent mutations expanding the global understanding of the mutation's geographic and ethnic distribution and contributing valuable epidemiological data. The study will facilitate genetic counseling for families experiencing similar clinical features, both within Pakistani populations and in other regions.

Relative Permittivity Measurement of Microliter Volume Liquid Samples through Microwave Filters.

This paper proposes a concept of dielectric characterization of low-volume liquid samples using the coupling coefficient of filters. The concept is validated through a two-pole substrate integrated waveguide filter in which the liquid under test is mounted on the coupling section between the two resonators. Unlike the conventional resonator perturbation method reported many times in the literature, this technique uses the coupling coefficient for sensing. The liquid sample is collected in a capillary tube and carefully positioned on the coupling section of the filter; the coupling coefficient of the two resonators varies compared to the relative permittivity of the sample; thus, an empirical model is established. The proposed sensor has been tested to compute the permittivity of different alcohols. Binary solutions of ethanol and water have also been characterized to calculate the volume ratio and relative permittivity as a proof-of-concept. The obtained results show that the proposed sensing technique is capable of characterizing a low quantity of liquids (≈44 µL) with good accuracy, and a worst case measured error of only 6.8% is noted. The ease of integration with other circuitry, low cost, reusability with no deterioration, and adaptability of the proposed sensor makes it a suitable choice for the chemical as well as for the pharmaceutical industry.

Exome sequencing reveals the first intragenic deletion in ABCA5 underlying autosomal recessive hypertrichosis.

BACKGROUND: Hereditary hypertrichosis (HH) is characterized by excessive hair growth on various body areas, which is independent of the individual's age. This rare hair disorder has been classified by its origin (genetic or acquired), age of onset, breadth of hair distribution (universal or localized) and the affected body areas. HH is often linked to several additional congenital abnormalities involving teeth, heart and bones. Human HH is associated with heterozygous genomic duplications and deletions in the chromosomal region 17q24.2-q24.3, containing genes such as ABCA5, ABCA6, ABCA10 and MAP2K6. Recently, a homozygous splice-site variant in ABCA5 has been reported to cause autosomal recessive congenital generalized hypertrichosis terminalis (CGHT; OMIM 135400). AIM: To investigate the clinical and genetic basis of autosomal recessive hypertrichosis in a large consanguineous Pakistani family. METHODS: In the present study, we characterized a family of Pakistani origin segregating CGHT in an autosomal recessive pattern, using whole exome sequencing followed by Sanger sequencing. RESULTS: We identified a novel 2-bp intragenic deletion [NM_172232.4(ABCA5);c.977_978delAT] causing a frameshift variant (p.His326ArgfsTer5) in ABCA5. CONCLUSIONS: To our knowledge, this is the first intragenic deletion in ABCA5 underlying CGHT. The findings further validate the involvement of ABCA5 in hair development. The study will facilitate genetic counselling of families carrying CGHT-related features in Pakistani and other populations.

Is age just a number or does it really affect the surgical and oncological outcomes after pancreatoduodenectomy?

OBJECTIVE: To review the impact of age on perioperative and postoperative outcomes in patients following pancreaticoduodenectomy. METHODS: The retrospective study was conducted at the Shaukat Khanum Memorial Cancer Hospital and Research Centre, Lahore, Pakistan, and comprised data from January 2014 to December 2018 of all patients who underwent pancreatoduodenectomy. Postoperative morbidity and oncological outcomes were compared between patients aged ≤60 years in group A and those aged >60 years in group B. Data was analysed using SPSS 20. RESULTS: Of the 161 patients, 103(64%) were males and 58(36%) were females. There were 117(73%) patients in group A; 72(61.5%) males and 45(38.5%) females with an overall mean age of 46±11 years. The remaining 44(27%) were in group B; 31(70.5%) males and 13(29.5%) females with an overall mean age of 67±05 years. The most common pathology was adenocarcinoma 130(81%), commonest site was periampullary 85(53%) and the most common pancreatic reconstruction technique employed was pancreaticogastrostomy 110(68%). Patients in group B had significantly higher comorbidities compared to those in group A (p<0.05). Estimated blood loss during surgery was significantly higher in group B (p=0.004). There was no significant difference in overall morbidity (p=0.856), reoperation (p=1.000), 30-day readmission rate (p=0.097), 90-day mortality rate (p=0.324)) and overall survival (p=0.551) between the groups. CONCLUSIONS: Pancreatoduodenectomy could be performed in the elderly with comparable morbidity and oncological outcomes as younger patients. Comorbid conditions remained higher in elderly patients and preoperative optimisation may help improve postoperative outcomes.

Wolfram-like syndrome with bicuspid aortic valve due to a homozygous missense variant in CDK13.

BACKGROUND: Wolfram syndrome (WFS) is characterized by deafness, diabetes mellitus, and diabetes insipidus along with optic atrophy. WFS has an autosomal recessive mode of inheritance and is due to variants in WFS1 and CISD2. METHODS: We evaluated the underlying molecular etiology of three affected members of a consanguineous family with hearing impairment, bicuspid aortic valve, diabetes mellitus and insipidus, clinodactyly, and gastrointestinal tract abnormalities via exome sequencing approach. We correlated clinical and imaging data with the genetic findings and their associated phenotypes. RESULTS: We identified a homozygous missense variant p.(Asn1097Lys) in CDK13, a gene previously associated with autosomal dominant congenital heart defects, dysmorphic facial features, clinodactyly, gastrointestinal tract abnormalities, intellectual developmental disorder, and seizures with variable phenotypic features. CONCLUSION: We report a homozygous variant in CDK13 and suggest that this gene causes an autosomal recessive disorder with hearing impairment, bicuspid aortic valve, diabetes mellitus and insipidus, clinodactyly, and gastrointestinal tract abnormalities.

Pancreaticoduodenectomy: Outcomes of a complex surgical procedure from a developing country.

BACKGROUND: Pancreaticoduodenectomy (PD) plays an integral part in the management of pancreatic, periampullary and duodenal cancers, along with a few other pathologies of this region. Despite advances in surgery PD continues to have significant morbidity and noteworthy mortality. The aim of this study is to provide an in-depth report on the patient characteristics, indications and the outcomes of PD) in a tertiary cancer hospital in Pakistan. MATERIALS AND METHODS: The study population included patients who underwent PD between January 1, 2014 and march 31, 2019, at Shaukat Khanum Memorial Cancer Hospital and Research Center (SKMCH&RC) in Pakistan. The data was retrospectively analyzed from the Hospital Information System (HIS), which is a prospectively maintained patient electronic database of SKMCH&RC. Patient characteristics, procedural details and post-operative outcomes according to internationally accepted definitions were reported. RESULTS: A total of 161 patients underwent PD at our hospital in the study period at a median age of 53 years, ranging from 19 to 78 years. 62% of the patients were males while 37% were females. Jaundice was the most common presenting symptom (64.6%), followed by abdominal pain (26.7%). PD with pancreaticogastrostomy was performed in 110 patients (68.3%), while pancreaticojejunostomy was performed in the rest of the cohort. Surgical site infection (SSI) was observed in 64 patients (40%). The incidence of Pancreatic Fistula grade C based on the International Study Group on Pancreatic Fistula (ISGPF) definition was 7.45% (n = 12). The 30 days mortality rate was 3.1%. Median survival of the cohort was 21 ±1.13 months and disease-free survival was 16±2.62 months. CONCLUSION: PD can be performed with acceptable morbidity and mortality in a resource constrained country, as long as it is undertaken in a high-volume center. This is in keeping with data published from other well-reputed international centers.

Co-prescription of ciprofloxacin and statins; a dangerous combination: Case Report.

Drug interaction is a common clinical problem which is often underestimated by physicians. Statins are one of the commonly prescribed medicines worldwide that are generally well tolerated. Muscle-related symptoms have a varied clinical presentation which usually increases if a new medicine is co-prescribed. We report the case of a 65- year-old woman who presented with a 10-day history of extreme fatigue, slowly progressing muscle weakness and insomnia. Drug-induced myopathy was diagnosed with concomitant use of atorvastatin and ciprofloxacin. Muscle weakness improved after the medicines were withdrawn. Co-prescription of Ciprofloxacin and statin therapy appeared to have contributed to muscle weakness in this patient. Drug interaction should also be kept in mind, when managing patients on statins as it may be underappreciated as a cause of muscle weakness and its consequences can have potentially serious outcomes.

Autosomal-Recessive Hearing Impairment Due to Rare Missense Variants within S1PR2.

The sphingosine-1-phosphate receptors (S1PRs) are a well-studied class of transmembrane G protein-coupled sphingolipid receptors that mediate multiple cellular processes. However, S1PRs have not been previously reported to be involved in the genetic etiology of human traits. S1PR2 lies within the autosomal-recessive nonsyndromic hearing impairment (ARNSHI) locus DFNB68 on 19p13.2. From exome sequence data we identified two pathogenic S1PR2 variants, c.323G>C (p.Arg108Pro) and c.419A>G (p.Tyr140Cys). Each of these variants co-segregates with congenital profound hearing impairment in consanguineous Pakistani families with maximum LOD scores of 6.4 for family DEM4154 and 3.3 for family PKDF1400. Neither S1PR2 missense variant was reported among ∼120,000 chromosomes in the Exome Aggregation Consortium database, in 76 unrelated Pakistani exomes, or in 720 Pakistani control chromosomes. Both DNA variants affect highly conserved residues of S1PR2 and are predicted to be damaging by multiple bioinformatics tools. Molecular modeling predicts that these variants affect binding of sphingosine-1-phosphate (p.Arg108Pro) and G protein docking (p.Tyr140Cys). In the previously reported S1pr2(-/-) mice, stria vascularis abnormalities, organ of Corti degeneration, and profound hearing loss were observed. Additionally, hair cell defects were seen in both knockout mice and morphant zebrafish. Family PKDF1400 presents with ARNSHI, which is consistent with the lack of gross malformations in S1pr2(-/-) mice, whereas family DEM4154 has lower limb malformations in addition to hearing loss. Our findings suggest the possibility of developing therapies against hair cell damage (e.g., from ototoxic drugs) through targeted stimulation of S1PR2.

Identification of CACNA1D variants associated with sinoatrial node dysfunction and deafness in additional Pakistani families reveals a clinical significance.

Sinoatrial node dysfunction and deafness (SANDD) syndrome is rare and characterized by a low heart beat and severe-to-profound deafness. Additional features include fatigue, dizziness, and episodic syncope. The sinoatrial node (SAN) drives heart automaticity and continuously regulates heart rate. The CACNA1D gene encoding the Cav1.3 protein expressed in inner hair cells, atria and SAN, induces loss-of-function in channel activity and underlies SANDD. To date, only one variant c.1208_1209insGGG:p.(G403_V404insG) has been reported for SANDD syndrome. We studied five Pakistani families with SANDD and characterized a new missense variant p.(A376V) in CACNA1D in one family, and further characterized the founder variant p.(G403_V404insG) in four additional pedigrees. We show that affected individuals in the four families which segregate p.(G403_V404insG) share a 1.03 MB haplotype on 3p21.1 suggesting they share a common distant ancestor. In conclusion, we identified new and known variants in CACNA1D in five Pakistani families with SANDD. This study is of clinical importance as the CACNA1D founder variant is only observed in families from the Khyber Pakhtunkhwa (KPK) province, in Pakistan. Therefore, screening patients with congenital deafness for SAN dysfunction in this province could ensure adequate follow-up and prevent cardiac failure associated with SAN.

The learning environment of four undergraduate health professional schools: Lessons learned.

BACKGROUND AND OBJECTIVES: Learning is an interplay between cognition and environmental factors. Any learning environment, that fulfills the intrinsic and extrinsic needs of the students will probably lead to better and more promising learning outcomes. This study aimed to investigate the student perceptions of Learning Environment (LE) in four health schools of a large university and compare between schools, years of study, and gender. METHODS: Dundee Ready Education Environment Measure (DREEM) questionnaire and a socio-demographic questionnaire were completed by 1185 undergraduate students enrolled in the school of Medicine, Dentistry, Nursing and Applied Medical Sciences (AMS) of a large university during the academic year 2012-2013. Chi-square test was used to compare categorical variables. Independent student t-test or ANOVA (with Tukey post-hoc test) was used for continuous variables at a significance level of p≤0.05. RESULTS: The mean total DREEM score was 89.23±33.3. The total DREEM mean scores for Dentistry (120.54±23.45) and Medicine (110.72±19.33) were higher compared with AMS (63.48±21.36) and Nursing (57.48±22.80) (p=0.000) (Post hoc Tukey p=0.000). First year students gave significantly higher positive perceptions ratings than the rest of the years (p=0.000). Total scores were significantly higher for male (92.78±33.86) than female students (84.70±32.25) p=0.000. CONCLUSION: The LE significantly differed by year and gender. The students from non-integrated curricula (nursing and AMS) perceived the LE less positively than their integrated curriculum counterparts (medicine and dentistry). A qualitative study is needed to investigate the variation in the perception of LE among these groups.

Relationship of S/R warfarin ratio with CYP2C9 genotypes in Pakistani population.

Warfarin is administered as a racemic preparation of R- and S-enantiomers. S-warfarin is more potent than R-warfarin, so changes in blood levels of S-warfarin affect the anticoagulant response. This study was carried out to determine the effect of CYP2C9*2 and CYP2C9*3 polymorphisms on S/R warfarin ratio. A single blood sample was collected 12-16 hours after drug administration from 170 stable patients fulfilling the criteria. Genotyping of the CYP2C9 polymorphisms was done by polymerase chain reaction-restriction fragment length polymorphism assay. S- and R-warfarin enantiomers extraction from plasma was accomplished by a validated HPLC method. The concentration of S-warfarin was significantly different among CYP2C9 genotypes (p =0.018) whereas there was no effect on R-warfarin (p =0.134). There was statistically significant effect of different CYP2C9 genotypes on S/R warfarin ratio (p=0.000). It is concluded that CYP2C9 polymorphisms influence CYP2C9 enzymatic activity in turn affecting S-warfari levels but not R-warfarin, thus leading to different S/R warfarin enantiomers ratio among different CYP2C9 genotypes.

Role of gut microbiota against calcium oxalate.

Nephrolithiasis is a condition marked by the presence or formation of stones in kidneys. Several factors contribute to kidney stones development such as environmental conditions, type of dietary intake, gender and gastrointestinal flora. Most of the kidney stones are composed of calcium phosphate and calcium oxalate, which enter in to the body through diet. Both sources of oxalates become dangerous when normal flora of gastrointestinal tract is disturbed. Oxalobacter and Lactobacillus species exist symbiotically in the human gut and prevent stone formation by altering some biochemical pathways through production of specific enzymes which help in the degradation of oxalate salts. Both Oxalobacter and Lactobacillus have potential probiotic characteristics for the prevention of kidney stone formation and this avenue should be further explored.

Frontal Sinus Mucocele Formation as a Late Complication of External Dacrocystorhinostomy.

Two cases of frontal sinus mucocele post external approach dacrocystorhinostomy (DCR) surgery are reported. The possible anatomical causes of this condition are discussed and in particular, attention is drawn to the consideration of frontal sinus mucocele in patients presenting with frontal sinus symptoms post-DCR surgery.

A homozygous missense variant in type I keratin KRT25 causes autosomal recessive woolly hair.

BACKGROUND: Woolly hair (WH) is a hair abnormality that is primarily characterised by tightly curled hair with abnormal growth. METHODS: In two unrelated consanguineous Pakistani families with non-syndromic autosomal recessive (AR) WH, homozygosity mapping and linkage analysis identified a locus within 17q21.1-q22, which contains the type I keratin gene cluster. A DNA sample from an affected individual from each family underwent exome sequencing. RESULTS: A homozygous missense variant c.950T>C (p.(Leu317Pro)) within KRT25 segregated with ARWH in both families, and has a combined maximum two-point LOD score of 7.9 at Ï´=0. The KRT25 variant is predicted to result in disruption of the second α-helical rod domain and the entire protein structure, thus possibly interfering with heterodimerisation of K25 with type II keratins within the inner root sheath (IRS) of the hair follicle and the medulla of the hair shaft. CONCLUSIONS: Our findings implicate a novel gene involved in human hair abnormality, and are consistent with the curled, fragile hair found in mice with Krt25 mutations, and further support the role of IRS-specific type I keratins in hair follicle development and maintenance of hair texture.

Diagnosis, Management And Prevention Of Hepatitis C In Pakistan 2017.

AIMS AND OBJECTIVES: Since the advent of direct acting antiviral agents, there is a revolutionary change in the management of HCV infection. Newer drugs with different mechanism of action are being introduced and are expected to be available in coming few months in Pakistan as well. The main purpose of the guideline is to review and induct the latest research in field of HCV infection in Pakistani perspective so that our healthcare professionals can apply the new recommendations in timely and judicial manner. Target groups of guidelines are general physicians treating hepatitis C, hepatologists and gastroenterologists. Other beneficiaries of these guidelines are public health institutions of Pakistan, which provide free treatment to deserving patients under National Hepatitis Prevention and Control Program and Pakistan Bait-ul- Mal Program. METHODOLOGY: These guidelines are based on the review of National consensus practice guidelines: Diagnosis, Management and Prevention of Hepatitis C Pakistan 2009. Published data in National and International Journals searched with the help of Google search and pub med, and 2015-16 guidelines of HCV by AASLD, EASL, APASL and WHO. Local studies are preferably added with references to enhance the Pakistani perspective. Evidence was also taken from published studies. Recommendations have been based upon evidence from national publications on the subject and scientific presentations at national liver meeting as well from experts' personal experience and opinion.

In silico analysis of SIGMAR1 variant (rs4879809) segregating in a consanguineous Pakistani family showing amyotrophic lateral sclerosis without frontotemporal lobar dementia.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder affecting upper motor neurons in the brain and lower motor neurons in the brain stem and spinal cord, resulting in fatal paralysis. It has been found to be associated with frontotemporal lobar degeneration (FTLD). In the present study, we have described homozygosity mapping and gene sequencing in a consanguineous autosomal recessive Pakistani family showing non-juvenile ALS without signs of FTLD. Gene mapping was carried out in all recruited family members using microsatellite markers, and linkage was established with sigma non-opioid intracellular receptor 1 (SIGMAR1) gene at chromosome 9p13.2. Gene sequencing of SIGMAR1 revealed a novel 3'-UTR nucleotide variation c.672*31A>G (rs4879809) segregating with disease in this family. The C9ORF72 repeat region in intron 1, previously implicated in a related phenotype, was excluded through linkage, and further confirmation of exclusion was obtained by amplifying intron 1 of C9ORF72 with multiple primers in affected individuals and controls. In silico analysis was carried out to explore the possible role of 3'-UTR variant of SIGMAR1 in ALS. The Regulatory RNA motif and Element Finder program revealed disturbance in miRNA (hsa-miR-1205) binding site due to this variation. ESEFinder analysis showed new SRSF1 and SRSF1-IgM-BRCA1 binding sites with significant scores due to this variation. Our results indicate that the 3'-UTR SIGMAR1 variant c.672*31A>G may have a role in the pathogenesis of ALS in this family.

A novel homozygous nonsense mutation in the PVRL4 gene and expansion of clinical spectrum of EDSS1.

Ectodermal dysplasias (EDs) belong to a group of genetic diseases which result from alterations in ectoderm-derived appendages including hair, nail, teeth and sweat glands. Ectodermal dysplasia syndactyly syndrome (EDSS1) is one of the rare forms of ED caused by mutations in nectin-4, encoded by the PVRL4 gene. The present study described clinical investigation of the EDSS1 identified in a large consanguineous family. Furthermore, DNA sequence analysis revealed a novel homozygous nonsense mutation (181C>T, p.Asp61*) in the PVRL4 gene.

Emergency contraception: Awareness, attitudes and barriers of Saudi Arabian Women.

OBJECTIVES: To assess knowledge, attitude, and barriers about emergency contraception (EC) among married women of child bearing age. METHODS: A quantitative cross-sectional study was conducted over a 6-month period, commencing in March 2013 at Family Practice Clinics of King Khalid University Hospital (KKUH), Riyadh, Saudi Arabia. Data was collected using a structured pretested questionnaire and analyzed using SPSS version 21.0 statistical software. RESULT: A total of 242 women were enrolled in the study. Only 6.2% (15/242) had some knowledge of EC and of these only two had ever used it. Health care professionals were the least reported source of EC information (6.6%, n=1). Majority (73.3%) had negative attitude toward EC being available over-the-counter without a prescription. The most common barriers to using EC were concerns about possible health effects. Only two women (13.3%) considered religious belief as a major hindrance to its use. CONCLUSION: Awareness of emergency contraception is very low among women of Saudi Arabia. Health care professionals were the least reported source of information, which is a cause for concern. Our findings reveal an urgent need to educate women about EC, keeping in view the social norms and the Islamic values.