Autosomal recessive non-syndromic hearing loss genes in Pakistan during the previous three decades.

Hearing loss is a clinically and genetically heterogeneous disorder, with over 148 genes and 170 loci associated with its pathogenesis. The spectrum and frequency of causal variants vary across different genetic ancestries and are more prevalent in populations that practice consanguineous marriages. Pakistan has a rich history of autosomal recessive gene discovery related to non-syndromic hearing loss. Since the first linkage analysis with a Pakistani family that led to the mapping of the DFNB1 locus on chromosome 13, 51 genes associated with this disorder have been identified in this population. Among these, 13 of the most prevalent genes, namely CDH23, CIB2, CLDN14, GJB2, HGF, MARVELD2, MYO7A, MYO15A, MSRB3, OTOF, SLC26A4, TMC1 and TMPRSS3, account for more than half of all cases of profound hearing loss, while the prevalence of other genes is less than 2% individually. In this review, we discuss the most common autosomal recessive non-syndromic hearing loss genes in Pakistani individuals as well as the genetic mapping and sequencing approaches used to discover them. Furthermore, we identified enriched gene ontology terms and common pathways involved in these 51 autosomal recessive non-syndromic hearing loss genes to gain a better understanding of the underlying mechanisms. Establishing a molecular understanding of the disorder may aid in reducing its future prevalence by enabling timely diagnostics and genetic counselling, leading to more effective clinical management and treatments of hearing loss.

Novel and known minor alleles of CNTNAP2 gene variants are associated with comorbidity of intellectual disability and epilepsy phenotypes: a case-control association study reveals potential biomarkers.

BACKGROUND: Neurodevelopmental disorders are heterogeneous due to underlying multiple shared genetic pathways and risk factors. Intellectual disability, epilepsy and autism spectrum disorder phenotypes overlap which indicates the diverse effects of common genes. Recent studies suggested the probable contribution of CNTNAP2 gene polymorphisms to the comorbidity of these neurological conditions. METHODS AND RESULTS: This study was conducted to investigate the role of CNTNAP2 polymorphisms rs147815978 (G>T) and rs2710102 (A>G) as a risk factor for comorbidity of intellectual disability and epilepsy in a group of 345 individuals including 170 patients and 175 healthy controls recruited from various ethnic groups of Pakistani population. Our case-control study group was genotyped by tetra primer ARMS-PCR technique and results were analysed to know the effects of CNTNAP2 rs147815978 (G>T) and rs2710102 (A>G) polymorphisms in the group. The frequency of risk allele T (rs147815978) and risk allele G (rs2710102) for homozygous recessive genotypes (TT/GG) in our study group was 36.47% while odds ratios for risk allele T (rs147815978) was 5.45 (3.90-7.61: 95% CI, P = 0.000) and that for risk allele G (rs2710102) was 2.39 (1.76-3.24: 95% CI, P = 0.0001). Homozygous recessive genotypes (TT/GG) appeared only in cases and not in control group which indicated these as suspected risk genotypes and the significant association (p < 0.05%) of CNTNAP2 gene polymorphisms rs147815978 (G>T) and rs2710102 (A>G) with co-occurrence of intellectual disability and epilepsy phenotypes in our study group which is in HWE (χ2 = 174, P < 0.0001). Logistic regression analysis shows additive (p < 0.0001) and multiplicative (p < 0.001) models which confirms significant association of both the polymorphisms in our data, which are closely located on same haplotype (D' = - 0.168). CONCLUSIONS: We propose that CNTNAP2 rs147815978 (G>T) and rs2710102 (A>G) polymorphisms are possible risk loci for overlapping neurodevelopmental disorders in Pakistani population. We propose the role of a previously reported common SNP rs2710102 (A>G) with a rarely reported novel SNP rs147815978 (G>T) for CNTNAP2 gene association with neurodevelopmental disorders in our data. Our study has expanded the knowledge of CNTNAP2 gene polymorphisms as probable biomarkers for susceptibility of co-occurrence of intellectual disability and epilepsy phenotypes in Pakistani population. We hope that our study will open new horizons of CNTNAP2 gene variants research to cure the neurological conditions in Pakistani population where consanguinity is a tradition and prevalence of neurodevelopmental disorders has increased from 1 to 2% during last 5 years.

Association of APO B gene polymorphisms with the development of myocardial infarction in Pakistani population.

Myocardial infarction (MI) is when a blood clot in the coronary artery obstructs blood flow to a specific part of the heart, leading to the death of myocardium in that area. The development of MI is influenced by various environmental factors, genetic components, and their interactions, even though the exact cause has not been fully established. This is the first case-control study examining the possible association between the human Apo B gene and MI in the Punjab region of Pakistan. The study included 100 patients and 50 healthy individuals. Genomic DNA was isolated from blood samples using manual extraction methods. Subsequently, primers were optimized, and genotyping was performed using PCR, followed by DNA sequencing and RFLP analysis. The research focused on two specific APO B gene SNPs, codon 4154 G/A (rs1801701) and codon 2488 G/A (rs1042031). Both SNPs involved the substitution of guanine with adenine. It was found that individuals carrying the minor allele A of SNP rs1801701 (p < 0.001) and the minor allele A of rs1042031 (p < 0.001) had a significantly higher risk of developing MI. Additionally, haplotype analysis revealed that the AA haplotype (comprising both rs1801701 and rs1042031 SNPs) was associated with a substantially increased risk of MI (OR = 3.845). In conclusion, the study provides evidence supporting the association between specific mutations in the APOB gene and the risk of myocardial infarction in the Pakistani population.

The miRNA variants MIR196A2 (rs11614913) and MIR423 (rs6505162) contribute to an increase in the risk of myocardial infarction.

INTRODUCTION: MicroRNAs (miRNAs) are small, single-stranded RNA molecules that negatively regulate gene expression and play a key role in the pathogenesis of human diseases. Recent studies have suggested that miRNAs contribute to cardiovascular diseases (CVDs). However, the association between single-nucleotide polymorphisms (SNPs) in miRNAs and myocardial infarction (MI) remains in infancy. AIM: The current study was designed to find out the association of SNPs in MIR196A2 and MIR423 (rs11614913 and rs6505162, respectively). METHODS: Using Tetra-Primer Amplification Refractory Mutation System-Polymerase Chain Reaction (T-ARMS PCR) in 400 cases (MI patients) and 336 healthy controls. Using different inheritance models (co-dominant, homozygous dominant, homozygous recessive, and additive models), the association of these SNPs was genotyped with MI risk. RESULTS: For variant rs11614913, significant distribution of the genotypes among the cases and controls was determined by co-dominant [χ2 = 29.19, 2; p value < 0.0001], dominant (C/C vs. C/T + T/T) [OR = 0.45 (0.34 to 0.61); p < 0.0001], recessive (T/T vs. C/T + C/C) [OR = 1.009 (0.63 to 1.63); p-value p value > 0.999], and additive models [OR = 0.65 (0.52 to 0.80); p value = 0.0001]. Similarly, a significant association of rs6505162 was determined by co-dominant [χ2 = 24.29, 2; p value < 0.0001], dominant (C/C vs. A/C+ A/A) [OR = 0.44 (0.32 to 0.61); p value < 0.0001], recessive (A/A vs. A/C + C/C) [OR = 1.29 (0.85 to 1.98); p value = 0.28], and additive models [OR = 0.65 (0.52 to 0.81); p value = 0.0001]. CONCLUSION: Therefore, the current study showed that both variants rs11614913 and rs6505162 are significantly associated with MI in the Pakistani population.

Myths and Misconceptions of Breast Cancer in the Pakistani Population.

Background Breast cancer is one of the most prevalent types of cancer in the female population. The cultural diversity, religious beliefs, myths, and misinformation regarding the disease contribute to diagnostic delays and enhanced burden on the healthcare system. This study aimed to ascertain the extent of knowledge and prevalence of erroneous beliefs and misconceptions regarding breast cancer among Pakistani women belonging to diverse socioeconomic and educational backgrounds. Methodology This cross-sectional study was performed in a tertiary care hospital in Karachi, Pakistan. A total of 350 women were enrolled in the study as a representative female population, and 300 participants were included who met the inclusion criteria. Participants were conveniently interviewed using a pre-piloted questionnaire designed to assess the prevalent myths and misconceptions about breast cancer. The data were analyzed by SPSS version 23 (IBM Corp., Armonk, NY, USA) using descriptive statistics. Results The study findings point to a significant prevalence of erroneous beliefs and a lack of accurate information on breast cancer. The mean age of the participants was 20.8 ± 10.4 years. The majority of the participants belonged to a middle socioeconomic status (70%) and were undergraduates (61.4%). The participants' friends and family members were the most frequent sources of information regarding breast cancer. The most common myth was "breast-feeding offers immunity to breast cancer completely" (76.6%), followed by "breast cancer spreads after biopsy" (63.8%). Participants also believed that breast tissue biopsy can lead to the spread of cancer (63.4%) and that faith healers and alternative medicine can cure breast cancer (47.5%). One-third (33.3%) of the participants considered all lumps to be breast cancer; however, approximately half (41.6%) of the participants thought that only painful lumps were associated with breast cancer. A significant number of participants believed breast cancer to be a result of God's curse (31.4%) or evil eye (38.7%). Conclusions The findings suggest a critical need for community-based breast health education initiatives that take into account Pakistani women's distinctive cultural and societal attitudes and work to dispel common misconceptions about the condition.

A homozygous founder variant in PDE2A causes paroxysmal dyskinesia with intellectual disability.

Intellectual developmental disorder with paroxysmal dyskinesia or seizures (IDDPADS, OMIM#619150) is an ultra-rare childhood-onset autosomal recessive movement disorder manifesting paroxysmal dyskinesia, global developmental delay, impaired cognition, progressive psychomotor deterioration and/or drug-refractory seizures. We investigated three consanguineous Pakistani families with six affected individuals presenting overlapping phenotypes partially consistent with the reported characteristics of IDDPADS. Whole exome sequencing identified a novel missense variant in Phosphodiesterase 2A (PDE2A): NM_002599.4: c.1514T > C p.(Phe505Ser) that segregated with the disease status of individuals in these families. Retrospectively, we performed haplotype analysis that revealed a 3.16 Mb shared haplotype at 11q13.4 among three families suggesting a founder effect in this region. Moreover, we also observed abnormal mitochondrial morphology in patient fibroblasts compared to controls. Belonging to diverse age groups (13 years-60 years), patients presented paroxysmal dyskinesia, developmental delay, cognitive abnormalities, speech impairment, and drug-refractory seizures with variable onset of disease (as early as 3 months of age to 7 years). Together with the previous reports, we observed that intellectual disability, progressive psychomotor deterioration, and drug-refractory seizures are consistent outcomes of the disease. However, permanent choreodystonia showed variability. We also noticed that the later onset of paroxysmal dyskinesia manifests severe attacks in terms of duration. Being the first report from Pakistan, we add to the clinical and mutation spectrum of PDE2A-related recessive disease raising the total number of patients from six to 12 and variants from five to six. Together, with our findings, the role of PDE2A is strengthened in critical physio-neurological processes.

Effects of body mass index on propofol-induced cardiovascular depression in the Pakistani population.

Objective: To determine the relationship between the patient's Body Mass Index (BMI) and the cardiovascular effects produced by propofol at a dose of 1.5 mg/kg in the Pakistani population. Methods: This descriptive cross-sectional study was conducted in the Holy Family Hospital Rawalpindi from August 2021 to January 2022. According to their BMI, one hundred twenty Pakistani individuals 18 to 60 years of age were equally divided into three groups. Group N (n = 40) with a BMI of 18 to 24.9, group OW (n=40) with a BMI of 25 to 29.5, and group O (n=40) with a BMI of 30 to 34.9 were randomized to receive propofol injections at a 1.5 mg/kg dose for induction of anesthesia. We measured mean blood pressure before the propofol and then at one, three, and ten minutes after the injection. Data were analyzed by using SPSS 22. Results: Mean blood pressure decreases significantly in all groups, as shown by p-values of <0.001 for the first two readings. In group N, blood pressure returned to near normal within ten minutes (p-value 0.061), but in groups, OW and O, mean blood pressure was significantly lower even after ten minutes (p-values 0.005 and 0.001, respectively). Individual variations in propofol response were also observed. Conclusion: In the Pakistani population, propofol at an induction dose of 1.5 mg/kg to patients with different body weights produces cardiovascular effects with marked standard deviations in each group, which indicate different individual responses. Clinical Trial Number: NCT05383534 https://register.clinicaltrials.gov/.

Genetic studies in the Pakistani population reveal novel associations with ventricular septal defects (VSDs).

BACKGROUND: With prevalence up to 4%, Ventricular Septal Defect (VSD) is one of the leading causes of neonatal deaths. VSD is a common complex genetic disorder that has been associated with many genetic determinants. Variants from genes for the transcription factors including T-Box TBX5 and NFATc1 (nuclear factor of activated T cells, cytoplasmic 1), Vascular endothelial growth factor (VEGF), ISLET1 (encoded by the ISL1 gene) and enzyme MTHFR, a methylene tetrahydrofolate reductase were selected. Genetic risk score (GRS) is a widely accepted approach used to convert the genetic data into prediction and assessment tool for disease susceptibility. METHODS: A total of 200 participants were recruited for the current study, 100 VSD patients and 100 controls. Genotyping of the ISL1: rs1017, NFATc1: rs7240256, VEGF: rs36208048, TBX5: rs11067075, and MTHFR: rs1801133 variants was performed using tetra primer ARMS PCR and PCR-RFLP. For the statistical analysis, the software SPSS version 23 was used. Genotypic frequencies of cases and controls were calculated using chi-square (χ²) whereas allelic frequencies were calculated by using the SNPStats tool. The association of GRS quartiles with VSD was examined using binary logistic regression. Adjusted p-value 0.01 was used as significance threshold for all analyses. RESULTS: The ISL1 (OD: 0.242, CI: 0.158-0.37, p-value: 2.15 × 10- 4 :), NFATc1 (OD: 2.53, CI: 1.64-3.89, p-value: 2.11 × 10- 5), TBX5 (OD: 2.24, CI: 1.47-3.41, p-value:1.6 × 10- 4) and MTHFR (OD: 10.46, CI: 5.68-19.26, p-value: 2.09 × 10- 9:) variants were found to be in association with VSD. In contrast, the VEGF (OD: 0.952, CI: 0.56-1.62, p-value: 0.8921) variant did not show significance association with the VSD. For cases, the mean GRS score was 3.78 ± 1.285 while in controls it was 2.95 ± 1.290 (p-value: 0.479, CI: 0.474-1.190). Comparison of GRS between cases and control showed that mean GRS of cases was 1.90 ± 0.480 while in controls it was 1.68 ± 0.490 (p-value: 0.001, CI: 0.086-0.354). Higher quartiles were more prevalent in cases whereas lower quartiles were more prevalent in controls. CONCLUSION: GRS of these five loci was strongly associated with VSD. Moreover, genetic risk score can provide better information for the association between variants and disease as compared to a single SNP. We also illustrated that the cumulative power of GRS is greater over the single SNP effect. This is a pilot scale study with a relatively small sample size whose findings should be replicated in a larger sample size for the unique local Pakistani population.

Genome-wide analysis of runs of homozygosity in Pakistani controls with no history of speech or language-related developmental phenotypes.

BACKGROUND: Runs of homozygosity (ROHs) analysis of controls provide a convenient resource to minimize the association of false positive results of disease-associated ROHs and genetic variants for simple and complex disorders in individuals from the same population. Evidence for the value of ROHs to speech or language-related traits is restricted due to the absence of population-matched behaviourally defined controls and limited family-based studies. AIM: This study aims to identify common ROHs in the Pakistani population, focussing on the total length and frequency of ROHs of variable sizes, shared ROHs, and their genomic distribution. SUBJECTS AND METHODS: We performed homozygosity analysis (in PLINK) of 86 individuals (39 males, 47 females) with no history of speech or language-related phenotypes (controls) who had been genotyped with the Illumina Infinium QC Array-24. RESULTS: ROHs of 1-<4 megabases (Mb) were frequent in unrelated individuals. We observed ROHs over 20 Mb among six individuals. Over 30 percent of the identified ROHs were shared among several individuals, indicating consanguinity's effect on the Pakistani population. CONCLUSION: Our findings serve as a foundation for family-based genetic studies of consanguineous families with speech or language-related disorders to ultimately narrow the homozygosity regions of interest to identify pathogenic variants.

Evaluation of the Nasopalatine Canal (NPC) in a subset of Pakistani population, using Cone Beam Computed Tomography (CBCT).

OBJECTIVE: To evaluate variations in the shape, diameter, length and width of the nasopalatine canal along with the width of the buccal cortical bone anterior to it, using cone beam computed tomography imaging. METHODS: The retrospective, cross-sectional study was conducted at the Aga Khan University Hospital, Karachi, from September to October 2020, and comprised pre-existing cone beam computed tomography scans taken between 2015 and 2020 of patients of either gender aged 18-60 years who had maxillary central incisors present. The shapes and dimensions of the nasopalatine canal were observed along with the buccal bone anterior to the nasopalatine canal. Data was compared with respect to age and gender. Data was analysed using SPSS 23. RESULTS: Of the 90 scans evaluated, 46(51.1%) belonged to females with a mean age of 37.85±18.19 years, and 44(48.9%) belonged to males with a mean age of 38.07±13.58 years. The mean length and width of the nasopalatine canal was 11.28±1.90mm and 2.62±0.91mm, respectively. The nasopalatine canal was significantly longer (p<0.01) and wider (p=0.02) in males than females. The mean diameter of foramen of Stenson was 2.99±1.17mm and incisive foramen was 6.09±1.80mm. The mean width of the buccal cortical bone at the most coronal, middle and most incisal levels was 7.20±1.70mm, 6.12±1.31mm and 6.12±1.31mm, respectively. Buccal bone width was wider in males than females, but the difference was significant only at the midpoint (p<0.05). CONCLUSIONS: There was a significant difference in the dimensions of the width and length of the NPC with respect to gender. No significant differences were observed with respect to age.

Association of the BB genotype of the ABO gene with the risk of acute myocardial infarction in hospital-based study.

Objectives: The ABO gene locus has been identified to be associated with myocardial infarction in patients with coronary heart disease. The primary focus of this hospital-based study was to explore the relationship of ABO blood groups and ABO genotypes with acute myocardial infarction (AMI) in Karachi, Pakistan. Methods: In a comparative cross-sectional study, an equal number of adult AMI patients and healthy controls (n=275 in each group; age range 30-70 years, both males and females) were recruited from the Aga Khan University and NICVD, Karachi, with informed consent. The blood samples were analyzed for ABO blood groups and other biomarkers. PCR followed by RFLP techniques were employed for determining the ABO genotypes. Multinomial regression was used to evaluate the association of genotypes with the risk of AMI. Results: Thirteen different combinations of ABO genotypes were observed while the O2O2 and A2A2 genotypes were not detected. No significant association based on the distribution of blood groups A, B, O and AB among AMI patients and healthy individuals was observed. The odds of AMI were 3.32 times in subjects with BB genotype as compared to subjects with OO genotypes after adjustment of age, gender, body mass index, heart rate, total cholesterol, and waist circumference [AOR (95% CI) =3.32 (1.36-8.08), p-value =0.008]. Conclusion: Our hospital-based study indicates that ABO genotype BB was significantly associated with the risk of AMI. This harmful effect of the BB genotype could have a possible relationship with AMI's development in the Pakistani population.

COVID-19 Fear Among Pakistanis: Psychometric Evaluation of the Fear of COVID-19 Scale Using Item Response Theory and Confirmatory Factor Analysis.

The Fear of COVID-19 Scale (FCV-19S) assesses the fear of the novel coronavirus disease 2019 (COVID-19) and has been translated and validated into over 20 languages. The present study conducted confirmatory factor analysis (CFA) and item response theory (IRT) analyses on the FCV-19S among a sample of 937 Pakistani adults (mean [SD] age of 25.83 [11.80] years; 537 [57.3%] females). The CFA and IRT confirmed the unidimensionality of the FCV-19S. The Likert-type scale used in the FCV-19S was supported by the proper threshold orderings. Additionally, no DIF contrast had an absolute value larger than 0.5 regarding the participants' characteristics of gender, age, living status, and education in the IRT findings. The FCV-19S was found to be valid and reliable with strong psychometric properties among the Pakistani adult population.

Gender Determination in Pakhtun Pakistani Population Using Dental Arch Dimensions: A Digital Model Study

@#Introduction: In circumstances where the ante mortem list is unknown, gender determination would exclude onehalf of the population, aid in a more precise search of the ante mortem records. This study aims to formulate gender prediction models in the Pakhtun Pakistani population using digital dental arch dimensions. Methods: Data collection and analysis of the dental casts were conducted on 128 subjects, 64 males and 64 females from the Pakistani population. The mean age of the subjects was 19.2 years old. Several linear dental arch dimensions were measured and recorded for both upper and lower arches. Results: It was found that gender differences in linear arch dimensions were statistically significant for both males and females (p<0.05); in which the arch dimensions for the males were larger than the arch dimensions for the females. Stepwise discriminant function analysis found that the highest discriminant power of the variables was present within the inter-second premolar width for the upper arch and inter-molar width for the lower arch. These variables significantly contributed to gender variance. Moreover, the prediction of 67.2% of original grouped cases for the upper arch and 66.4% of cross-validated group cases was correct. Similarly, the correct prediction was made on 64.8% of original grouped cases for the lower arch and 64.1% of cross-validated group cases. Conclusion: The dental arch dimensions were larger among the males compared to the females. Prediction models obtained in this study were moderately strong predictors which may be used as an adjunct to predict gender.

Association of Fc Gamma Receptor 3B Gene Copy Number Variation with Rheumatoid Arthritis Susceptibility.

Structural variations such as copy number variants (CNVs) have been associated with multiple autoimmune diseases. In this study, we explored the association of the Fc gamma receptor 3B gene (FCGR3B) copy number variation (CNV) with rheumatoid arthritis (RA) susceptibility and related serological traits in the Pakistani population. We also performed a meta-analysis of four published FCGR3B CNV studies along with the current study. A total of 927 subjects (597 RA cases, 330 healthy controls) were recruited from three rheumatology centers in Pakistan. Anti-cyclic citrullinated peptide (anti-CCP) antibodies and rheumatoid factor (RF) were measured in RA patients. FCGR3B copy number was assayed using the TaqMan® CN assay (Hs04211858_cn, Applied Biosystems, Foster City, CA, USA) and the copy number was estimated by using CopyCaller® software (version 2.1; Applied Biosystems, USA). Logistic regression was applied to calculate the odds ratio (OR) of RA risk associated with FCGR3B CNV using sex and age as covariates in R. Meta-analysis on four previously published studies and the current study was performed using the random-effect model. We observed a significant association between FCGR3B copy number < 2 and RA susceptibility (OR = 1.53; 95% CI: 1.05 to 2.22; p = 0.0259) and anti-CCP seropositivity (OR 2.56; 95% CI: 1.34 to 4.89; p = 0.0045). A non-significant association of FCGR3B copy number < 2 was also observed between increased rheumatoid factor (RF) seropositivity (OR = 1.74; 95% CI:0.93 to 3.26; p = 0.0816). Meta-analysis on 13,915 subjects (7005 RA cases and 6907 controls) also showed significant association of copy number < 2 with the increased risk of RA (OR = 1.30; 95% CI: 1.07 to 1.56; p = 0.00671). FCGR3B copy number < 2 is associated with increased RA risk and anti-CCP seropositivity.

Primary thyroid lymphoma: Case series of patients from a developing country.

Primary Thyroid Lymphoma (PTL) is a rare disease, mostly affecting middle-to-old aged females. It has a strong association with pre-existing thyroiditis, which increases the risk of its development. The disease has been studied in detail in the Western populations. However, there is lack of data for south Asians, especially in Pakistani population. This study was carried out to learn more about the disease characteristics in our area. This was a retrospective study where we reviewed four diagnosed and/or treated cases of primary thyroid Lymphoma at tertiary care hospital of Pakistan (Aga Khan University Hospital), Karachi, Pakistan. Each patient's medical record was reviewed and studied. The participants' average age was 62.8±10.12 years,, with two males and two females. The histology of all of the patients was compatible with the Diffuse B cell Lymphoma subtype. One patient died as a result of complications, one patient was lost to follow up, and the remaining two patients are doing well with no active complaints and are being followed up on a regular basis.

Study of variants associated with ventricular septal defects (VSDs) highlights the unique genetic structure of the Pakistani population.

BACKGROUND: Ventricular septal defects (VSDs) are one of the leading causes of death due to cardiac anomalies during the first months of life. The prevalence of VSD in neonates is reported up to 4%. Despite the remarkable progress in medication, treatment and surgical procedure for VSDs, the genetic etiology of VSDs is still in infancy because of the complex genetic and environmental interactions. METHODS: Three hundred fifty subjects (200 VSD children and 150 healthy controls) were recruited from different pediatric cardiac units. Pediatric clinical and demographic data were collected. A total of six variants, rs1017 (ISL1), rs7240256 (NFATc1), rs36208048 (VEGF), variant of HEY2, rs11067075 (TBX5) and rs1801133 (MTHFR) genes were genotyped by tetra-ARMS PCR and PCR-RFLP methods. RESULTS: The results showed that in cases, the rs1017 (g.16138A > T) variant in the ISL1 gene has an allele frequency of 0.42 and 0.58 respectively for the T and A alleles, and 0.75 and 0.25 respectively in the controls. The frequencies of the AA, TA and TT genotypes were, 52%, 11% and 37% in cases versus 21%, 8% and 71% respectively in the controls. For the NFATc1 variant rs7240256, minor allele frequency (MAF) was 0.43 in cases while 0.23 in controls. For the variant in the VEGF gene, genotype frequencies were 0% (A), 32% (CA) and 68% (CC) in cases and 0.0%, 33% and 67% respectively in controls. The allele frequency of C and A were 0.84 and 0.16 in cases and 0.83 and 0.17 respectively in controls. The TBX5 polymorphism rs11067075 (g.51682G > T) had an allelic frequency of 0.44 and 0.56 respectively for T and G alleles in cases, versus 0.26 and 0.74 in the controls. We did not detect the presence of the HEY2 gene variant (g.126117350A > C) in our pediatric cohort. For the rs1801133 (g.14783C > T) variant in the MTHFR gene, the genotype frequencies were 25% (CC), 62% (CT) and 13% (TT) in cases, versus 88%, 10% and 2% in controls. The ISL1, NFATc1, TBX5 and MTHFR variants were found to be in association with VSD in the Pakistani pediatric cohort whilst the VEGF and HEY2 variants were completely absent in our cohort. CONCLUSION: We propose that a wider programme of genetic screening of the Pakistani population for genetic markers in heart development genes would be helpful in reducing the risk of VSDs.

Assessment of Contributing Factors and Treatment Practices for Therapeutic Efficacy and Drug-Related Problems in Suicidal Psychotic Patients.

Suicide, a deliberate act of self-harm with the intention to die, is an emerging health concern but, unfortunately, the most under-researched subject in Pakistan, especially in Khyber Pukhtunkhwa (KPK). In this study, we aimed to identify risk factors that can be associated with suicidal behavior (SB) and to evaluate the prevailing treatment practices for therapeutic efficacy and drug-related problems (DRPs) in psychotic patients among the local population of KPK. A prospective, multicenter study was conducted for suicidal cases admitted to the study centers by randomized sampling. Socio-demographics and data on suicidal behavior were assessed using the Columbia-Suicide Severity Rating Scale (C-SSRS), socioeconomic condition by Kuppuswamy socioeconomic scale (KSES) and treatment adherence by Morisky Medication-Taking Adherence Scale (MMAS-4). Drug-related problems and the therapeutic efficacy of prevailing treatment practices were assessed at baseline and follow-up after 3 months of treatment provided. Regarding suicidality (N = 128), females reported more ideations (63.1%), while males witnessed more suicidal behavior (66.6%, p < 0.001). Suicide attempters were mostly married (55.6%, p < 0.002); highly educated (53.9%, p = 0.004); dissatisfied with their life and had a previous history (p < 0.5) of suicide attempt (SA) (20.6%), self-injurious behavior (SIB) (39.7%) and interrupted (IA) or aborted attempts (AA) (22.2%). A greater improvement was observed in patients receiving combination therapy (p = 0.001) than pharmacotherapy (p = 0.006) or psychotherapy (p = 0.183), alone. DRPs were also detected, including drug-selection problems (17.88%), dose-related problems (20.64%), potential drug−drug interactions (24.31%), adverse drug reactions (11.46%) and other problems like inadequate education and counseling (21.55%). Furthermore, it was also found that psychotic patients with suicidal ideations (SI) were significantly (p = 0.01) more adherent to the treatment as compared to those with suicidal attempts. We concluded that suicide attempters differed significantly from patients with suicidal ideations in psychotic patients and presented with peculiar characteristics regarding socio-demographic factors. A combination of therapies and adherence to the treatment provided better outcomes, and targeted interventions are warranted to address drug-related problems.

Clinical Response of Nuberol Forte® for Pain Management With Musculoskeletal Conditions in Routine Pakistani Practice (NFORTE-EFFECT).

Background Musculoskeletal pain is the most common complaint presented to the health practitioner. It is well-known that untreated or under-treated pain can have a significant negative impact on an individual's quality of life (QoL). Objectives The current study aimed to assess the clinical response of Nuberol Forte® (paracetamol 650 mg + orphenadrine 50 mg) to musculoskeletal pain in routine Pakistani practice and its impact on improving the patient's QoL. Methods A prospective, observational multicenter study (NFORT-EFFECT: Safety & Efficacy of Nuberol Forte in Pain Management). Three hundred ninety-nine patients with known prescreened musculoskeletal pain were recruited from 10 major healthcare facilities across six (6) major cities of Pakistan, as per the inclusion/exclusion criteria. After the baseline visit (Visit 1), the patients were followed up one to two weeks (Visit 2) after the treatment as per the physician's discretion. Data were collected using the Case Report Form (CRF) designed for the study, and adverse events (AEs) were also monitored to assess drug safety. Pain intensity was assessed through a visual analog scale (VAS), and QoL was assessed using the Muscle and Joint Measure (MJM) scale. Results Out of 399 enrolled patients, 49.4% were males and 50.6% were females with a mean age of 47.24 ± 14.20 years. Most patients were presented with knee osteoarthritis (OA), i.e., 148 (38%), followed by backache 70 (18.2%). A significant reduction in the mean pain score was observed after treatment with the combination of paracetamol and orphenadrine (p<0.05). Furthermore, an overall improvement in the patient's QoL was also observed. During the study, only 10 patients reported mild adverse events (AEs), namely, dryness of the mouth, dizziness, gastric irritation, tachycardia, restlessness, etc. Conclusion The combination of paracetamol and orphenadrine (Nuberol Forte) exhibited effective pain management among patients with musculoskeletal conditions and improved their QoL.

Correlation of lip profile preferences in Pakistani population; and its comparison with orthodontic norms.

OBJECTIVE: To determine the most desired lip profile and compare the subjective sense of aesthetics among orthodontists, general dentists and the general population. METHODS: The cross-sectional study was conducted at the Orthodontic Department of the Armed Forces Institute of Dentistry, Rawalpindi, Pakistan, from January 1 to February 25, 2020, and comprised different silhouettes for each gender with increasing lip procumbence from -6mm to +6mm with respect to Rickett's E-line which were created using Photoshop CS 8.0 after cephalometric analysis of 20 cephalograms. The sample comprised an equal number of orthodontists in group A, general dentists in group B and orthodontic treatment-seekers in group C with equal representation of the two genders. Data was analysed using SPSS 24. RESULTS: Of the 180 subjects, there were 60(33.3%) in each of the three groups, with 30(50%) males and as many females in all the groups. All the three groups preferred the average lip profile for males (p=0.018) and 2mm procumbent lips for females (p=0.008). There was significant difference of opinion between groups A and C (p=0.034) and between groups B and C (p=0.022). CONCLUSIONS: There was found to be a marked difference of opinion among the orthodontists, the general dentists and the orthodontic treatment-seekers regarding the desired lip profile.

Germline Mutation Analysis in Sporadic Breast Cancer Cases With Clinical Correlations.

Demographics for breast cancers vary widely among nations. The frequency of germline mutations in breast cancers, which reflects the hereditary cases, has not been investigated adequately and accurately in highly-consanguineous Pakistani population. In the present discovery case series, germ-line mutations in twenty-seven breast cancer candidate genes were investigated in eighty-four sporadic breast cancer patients along with the clinical correlations. The germ-line variants were also assessed in two healthy gender-matched controls. The clinico-pathological features were evaluated by descriptive analysis and Pearson χ2 test (with significant p-value <0.05). The most frequent parameters associated with hereditary cancer cases are age and ethnicity. Therefore, the analyses were stratified on the basis of age (≤40 years vs. >40 years) and ethnicity. The breast cancer gene panel assay was carried out by BROCA, which is a genomic capture, massively parallel next generation sequencing assay on Illumina Hiseq2000 with 100bp read lengths. Copy number variations were determined by partially-mapped read algorithm. Once the mutation was identified, it was validated by Sanger sequencing. The ethnic analysis stratified on the basis of age showed that the frequency of breast cancer at young age (≤40 years) was higher in Sindhis (n = 12/19; 64%) in contrast to patients in other ethnic groups. Majority of the patients had stage III (38.1%), grade III (50%), tumor size 2-5 cm (54.8%), and invasive ductal carcinoma (81%). Overall, the analysis revealed germ-line mutations in 11.9% of the patients, which was not significantly associated with younger age or any particular ethnicity. The mutational spectrum was restricted to three genes: BRCA1, BRCA2, and TP53. The identified mutations consist of seven novel germ-line mutations, while three mutations have been reported previously. All the mutations are predicted to result in protein truncation. No mutations were identified in the remaining twenty-four candidate breast cancer genes. The present study provides the framework for the development of hereditary-based preventive and treatment strategies against breast cancers in Pakistani population.