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Hypohidrotic ectodermal dysplasia (HED) is a congenital anomaly characterized by hypohydrosis, hypotrichosis and hypodontia. Mutations in at least four genes (EDAR, EDARADD, WNT10A, TRAF6) have been reported to cause both autosomal recessive and autosomal dominant forms of HED. Mutations in two other genes (EDA and IKBKG) have been reported to cause X-linked HED. To clinically characterize three consanguineous families (A-C) segregating with autosomal recessive HED and identify possible disease-causing variants of EDAR and EDARADD genes. The genes, EDAR and EDARADD, were sequenced in Family A and C, and exome sequencing was performed in Family B. Additionally, in Family A and C, the effect of the identified variants was examined by analysis of EDAR mRNA, extracted from hair follicles from both affected and unaffected members. Sequence analysis revealed three possible disease-causing EDAR variants including a novel splice acceptor site variant (IVS3-1G > A) in Family A and two previously reported mutations (p.[Ala26Val], p.[Arg25*]) in the two other families. Previously, the nonsense variant p.(Arg25*) was reported only in the heterozygous state. Analysis of the RNA, extracted from hair follicles, revealed skipping of a downstream exon in EDAR and complete degradation of EDAR mRNA in affected members in family A and C, respectively. Computational modelling validated the pathogenic effect of the two variants identified in Family B and C. The three variants reported here expand the spectrum of EDAR mutations associated with HED which may further facilitate genetic counselling of families segregating with similar disorders in the Pakistani population.

Green nanotechnology-based approaches have been acquired as environmentally friendly and cost effective with many biomedical applications. The present study reports the synthesis of silver nanoparticles (AgNPs) from the leaves of

The aim of the current study was to evaluate the effect of dietary supplementation of Pinus ponderosa leaves (pine leaves) and α-tocopherol acetate (vitamin E) powder on male reproductive system, serum metabolites and carcass characteristics of Japanese quails. A total of 360-day-old male quails were purchased from the open market and kept at poultry shed for ninety-four days. After ten days of adaptation, all quails were randomly assigned into 4 groups, control (IC); supplemented with α-tocopherol acetate (IE) at the rate of 150 mg/L; Pinus ponderosa leaves (IP) at the rate of 150 mg/L; and 70 mg α-tocopherol acetate and 70 mg Pinus ponderosa leaves (IEP). Pinus ponderosa leaves and α-tocopherol acetate supplementation had not significantly (p .05) effected on final body weight gain, feed intake and feed conversion ratio of quails. The high-density lipoprotein cholesterol (HDLC) and total cholesterol (TC) were significantly (p .05) affected by IE and IP groups as compared to IC and IEP groups. Triglyceride (TG), glutathione peroxidase (GPx) and superoxide dismutase (SOD) significantly (p .05) increased in all treatment groups except for the IC group. Aspartate transaminase (AST) significantly (p .05) decreased in treatment groups as compared to control group. Overall, the mineral levels significantly (p .05) increased in treatment groups as compared to control. Cloacal gland index values, the quantity of foam production and testis weight were significantly (p .05) increased in treatment groups. It was concluded that the supplementation of Pinus ponderosa leaves and α-tocopherol acetate improved the testis weight, foam production, serum antioxidant enzymes and mineral level especially zinc in Japanese quail considered an indicative characteristic of higher sperm production rate and improved sexual activity. Further, higher gametogenesis rate, sperm production or reproductive behaviour including different hormonal level will be analysed in future study.

Frontonasal dysplasia (FND) is a heterogeneous group of disorders characterized by hypertelorism, telecanthus, broad nasal root, wide prominent nasal bridge, short and wide nasal ridge, broad columella and smooth philtrum. To date one X-linked and three autosomal recessive forms of FND have been reported in different ethnic groups. We sought to identify the gene responsible for FND in a consanguineous Pakistani family segregating the disorder in autosomal recessive pattern. Genome-wide homozygosity mapping using 250KNsp array revealed five homozygous regions in the selected affected individuals. Exome sequencing found a novel splice acceptor site variant (c.661-1G>C: NM_006982.2) in ALX1. Sanger sequencing confirmed the correct segregation of the pathogenic variant in the whole family. Our study concludes that the splice site variant identified in the ALX1 gene causes mild form of FND.

Frontorhiny is one of the two forms of mid-facial malformations characterized by ocular hypertelorism, wide and short nasal ridge, bifid nasal tip, broad columella, widely separated nares, long and wide philtrum and V-shaped hairline. Sometimes these phenotypes are associated with ptosis and midline dermoid cysts. Frontorhiny inherits in an autosomal recessive pattern. Sequence variants in the Aristaless-like homeobox 3 (ALX3) gene underlying frontorhiny have been reported previously. Here, in the present study, we have investigated four patients in a consanguineous family of Pakistani origin segregating frontorhiny in autosomal recessive manner. Genome scan using 250k Nsp1 array followed by exome and Sanger sequence analysis revealed a novel homozygous nonsense variant (c.604C>T, p.Gln202*) in the ALX3 gene resulting in frontorhiny in the family. This is the first mutation in the ALX3 gene, underlying frontorhiny, in Pakistani population.

BACKGROUND: Postaxial polydactyly (PAP) type A is characterised by well-formed functionally developed 5th digit duplication in hands and/or feet. It is genetically heterogeneous condition, inherited both in autosomal recessive and dominant manners. To date one autosomal recessive and four autosomal dominant loci have been mapped on human chromosomes. In the present study we have investigated a consanguineous Pakistani family segregating autosomal recessive PAP type A to identify the gene responsible for this phenotype. METHODS: Whole exome sequencing combined with homozygosity mapping and array comparative genomic hybridisation (aCGH) analysis was used to search for a genetic cause of PAP type A in the present study. RESULTS: Exome sequencing identified a missense mutation (c.1420C>T; p.Thr474Ile) in all the affected individuals of the family, in the gene ZNF141, mapped to the telomeric region on chromosome 4p16.3. CONCLUSION: This study revealed involvement of a zinc finger gene ZNF141 in causing autosomal recessive PAP type A, which may open up interesting perspectives into the function of this protein in limb development.

Background: Ellis-van Creveld (EVC) syndrome is a rare autosomal recessive disorder characterized by skeletal, ectodermal and cardiac defects. This syndrome is caused by mutations in EVC and EVC2 genes, which are separated by 2.6 kb of genomic sequence on chromosome 4p16. Methods: In the present study we ascertained a four-generation pedigree of Pakistani origin with features of EVC. Linkage was searched by genotyping microsatellite markers linked to chromosome 4p16. Affected individuals showed homozygosity to the microsatellite markers tightly linked to EVC and EVC2 genes on chromosome 4p16. It was then subjected to direct sequencing of the EVC and EVC2 genes. Results: Mutation analysis of the EVC and EVC2 genes identified a novel missense change (c.617G>A; p.S206N) in the EVC gene. Conclusions: We herein report on the first family from Pakistan with a large number of individuals affected by EVC. DNA sequence analysis led to the identification of the fifth missense mutation in the EVC gene.

To evaluate diabetes awareness among general population of Bahawalpur, Pakistan.A questionnaire based descriptive cross sectional study was performed during 2014. Descriptive and inferential statistics were applied using the IBM Statistical Package for Social Sciences version 20.Total 383 respondents participated in this study giving a response rate of about 88%. Mean age of the participants was 33±13.3 years and the majority of the participants were females (50.4%). Mean scores of the participants were 20.77±6.65 out of 34 and only one participant scored 34/34 marks. Kruskal-Wallis and Mann-Whitney U test revealed that females scored significantly higher diabetes awareness score than males (p0.05). Moreover, urban residential status, better education and higher socioeconomic status were significantly associated with higher diabetes awareness.Majority of our study population had adequate awareness regarding diabetes. However, lack of awareness was observed among illiterate, poor and rural participants that indicated the immediate need of diabetes awareness programs for these participants.

To evaluate the role of Fluorine-18-fluorodexoyglucose Positron Emission Tomography (FDG-PET) scan in staging and its implications on the treatment of lymphoma, and to study the concordance between visual assessment and Deauville criteria for the interpretation of interim scans.The prospective single-arm experimental study was conducted at the Shaukat Khanum Memorial Cancer Hospital, Lahore, from May 2011 to October 2011. It comprised 53 newly diagnosed lymphoma patients who agreed to participate in the study. All patients underwent scans with contrast-enhanced computerised tomography at baseline. Treatment plan was formulated based on the final stage. Interim scans were acquired after 2 cycles of chemotherapy and were reported using visual criteria and compared with the 5-point Deauville criteria. Score of 1-3 was taken as disease-negative, while 4-5 was taken as disease-positive. SPSS 19 was used for statistical analysis.Of the 53 patients, 35 (66%) had Hodgkin's Lymphoma, while 18 (34%) had Non-Hodgkin's Lymphoma. Scans resulted in disease upstaging in 4 (7.5%) patients, and detecting increased disease burden in 12 (23%). On interim scans, complete remission was achieved in 38 (71%) patients (Deauville score 1-3); 12(23%) showed partial response (Deauville score 4-5); and 3 (6%) had progression. Kappa test was statistically significant (kappa 0.856; p0.001).The positron emission tomography helped to upstage lymphoma and reflected increased disease burden. The Deauville criteria correlated very well with visual assessment criteria and can be applied in the patient population.

Postaxial Polydactyly (PAP) is characterized by fifth digit duplication in hands and/or feet. Two types of PAP including PAP-A, representing the development of well-formed extra digit, and PAP-B, representing the presence of rudimentary fifth digit, have been described. Both isolated and syndromic forms of PAP have been reported. Isolated forms of PAP usually segregate as an autosomal dominant trait and to date four loci have been identified. In the present study, we have described mapping of the first locus of autosomal recessive PAP type A on chromosome 13q13.3–13q21.2 in a consanguineous Pakistani family. Using polymorphic microsatellite markers, the disease locus was mapped to a 17.87-cM (21.13 Mb) region flanked by markers D13S1288 and D13S632, on chromosome 13q13.3–13q21.2. A maximum multipoint LOD score of 3.84 was obtained with several markers along the disease interval. DNA sequence analysis of exons and splice-junction sites of ten candidate genes (CHM-I, TSC22D1, FOXO1, DIAPH3, CCDC122, CKAP2, SUGT1, RANKL, LPAR6, C13ORF31) did not reveal potentially causal variants.

Autosomal recessive hypotrichosis/woolly hair is a rare genetic hair loss disorder characterized by sparse scalp hair/woolly hair, sparse to absent eyebrows and eyelashes, sparse axillary and body hair in affected individuals. This form of hair loss results from mutations in either LPAR6 or LIPH gene.To identify mutations in LPAR6 and LIPH genes in 17 consanguineous Pakistani families showing features of hypotrichosis/woolly hair.Genotyping in 17 families was carried out using polymorphic microsatellite markers linked to genes causing autosomal recessive hypotrichosis/woolly hair phenotype. To screen for mutations in LPAR6 and LIPH genes, all of their exons and splice junction sites were amplified by PCR and sequenced using an automated DNA sequencer.Genotyping with polymorphic microsatellite markers showed linkage in eight families to LPAR6 and in nine families to LIPH gene. Sequence analysis revealed four recurrent mutations (p.Phe24HisfsX28; p.Asp63Val; p.Gly146Arg; p.Ile188Phe) in LPAR6 and two recurrent mutations (p.Trp108Arg; p.Ile220ArgfsX29) in LIPH gene. Comparison of the haplotypes generated by typing LPAR6 and LIPH genes linked microsatellite markers in different families suggested common founder natures of the two mutations (c.66_69insCATG and c.659_660delTA).Mutations identified in the present study extend the body of evidence implicating LPAR6 and LIPH genes in pathogenesis of human hereditary hair loss.

DFNB89 is a novel autosomal recessive non-syndromic hearing impairment (ARNSHI) locus that was mapped to 16q21-q23.2. Linkage to the region was established by carrying out genome-wide linkage scans in two unrelated, consanguineous Pakistani families segregating ARNSHI. The maximum multipoint LOD score is 9.7 for both families and for each family, a significant maximum LOD score of 6.0 and 3.7 were obtained. The 3-unit support interval and the region of homozygosity for the two families extend from rs717293 (chr16: 62.1 Mb) to rs728929 (chr16: 78.2 Mb) and contain 16.1 Mb of sequence. A total of 146 genes are within the DFNB89 interval. Eight candidate genes, CALB2, CDH1, CDH3, CDH11, HAS3, NOB1, PLEKHG4 and SMPD3, were sequenced, but no potentially causal variants were discovered. DFNB89 is the second ARNSHI locus mapped to chromosome 16.