Clinical and genetic investigation of 14 families with various forms of short stature syndromes.

Skeletal dysplasias are a heterogeneous group of disorders presenting mild to lethal defects. Several factors, such as genetic, prenatal, and postnatal environmental may contribute to reduced growth. Fourteen families of Pakistani origin, presenting the syndromic form of short stature either in the autosomal recessive or autosomal dominant manner were clinically and genetically investigated to uncover the underlying genetic etiology. Homozygosity mapping, whole exome sequencing, and Sanger sequencing were used to search for the disease-causing gene variants. In total, we have identified 13 sequence variants in 10 different genes. The variants in the HSPG2 and XRCC4 genes were not reported previously in the Pakistani population. This study will expand the mutation spectrum of the identified genes and will help in improved diagnosis of the syndromic form of short stature in the local population.

Identification of a Novel Nonsense Variant in the DLL3 Gene Underlying Spondylocostal Dysostosis in a Consanguineous Pakistani Family.

Introduction: Spondylocostal dysostosis (SCD) is characterized by multiple vertebral abnormalities associated with abnormalities of the ribs. Five genes causative for the disease have been identified. These include DLL3 (OMIM *602768), MESP2 (OMIM #608681), LFNG (OMIM #609813), TBX6 (OMIM *602427), and HES7 (OMIM *608059). Methods: In the current study, we investigated a Pakistani consanguineous family segregating spondylocostal dysotosis. Whole-exome sequencing (WES) followed by Sanger sequencing was performed using DNA of affected and unaffected individuals to identify pathogenic variant(s). The identified variant was interpreted using ACMG classification. Literature review was performed to summarize currently known mutated alleles of DLL3 and the underlying clinical phenotypes. Results: Clinical examination using anthropometric measurements and radiographs diagnosed the patients to be afflicted with SCD. Pedigree analysis of the affected family showed an autosomal recessive inheritance pattern of the disease. WES followed by Sanger sequencing identified a novel homozygous nonsense variant (DLL3(NM_016941.4): c.535G>T; p.Glu179Ter) in the DLL3 gene located on chromosome 19q13.2. Conclusion: The study will be helpful in carrier testing and genetic counseling to prevent segregation of the disease to the next generations within this family. It also provides knowledge for clinicians and researchers in search of a better understanding of SCD anomalies.

Variants in the PNPLA1 Gene in Families with Autosomal Recessive Congenital Ichthyosis Reveal Clinical Significance.

The term autosomal recessive congenital ichthyosis (ARCI) is the subgroup of ichthyosis, which describes a highly heterogeneous group of genetic disorders of the skin characterized by cornification and defective keratinocytes differentiation associated with mutations in at least 14 genes including PNPLA1. To study the molecular basis of the Pakistani kindreds (A and B) affected by ARCI, whole-exome sequencing (WES) in the DNA samples of affected members was performed followed by Sanger sequencing of the candidate gene to hunt down the disease-causing sequence variant/s. WES data analysis led to the identification of a novel nonsense sequence variant (c.892C>T; p.Arg298*, family A) and a recurrent missense variant (c.102C>A; p.Asp34Glu, family B) in PNPLA1 mapped to the ARCI locus in chromosome 6p21.31. Validation and cosegregation analysis of the variants in the remaining family members of the respective families were confirmed by Sanger sequencing. The current investigation expands the spectrum of PNPLA1 mutations and helps establish the proper clinico-genetic diagnosis and correct genotype-phenotype correlation.

Chloroplast genome evolution in the Dracunculus clade (Aroideae, Araceae).

Chloroplast (cp) genomes are considered important for the study of lineage-specific molecular evolution, population genetics, and phylogenetics. Our aim here was to elucidate the molecular evolution in cp genomes of species in the Dracunculus clade (Aroideae, Araceae). We report de novo assembled cp genomes for eight species from eight genera and also retrieved cp genomes of four species from the National Center for Biotechnology Information (NCBI). The cp genomes varied in size from 162,424 bp to 176,835 bp. Large Single Copy (LSC) region ranged in size from 87,141 bp to 95,475 bp; Small Single Copy (SSC) from 14,338 bp to 23,981 bp; and Inverted Repeats (IRa and IRb) from 25,131 bp to 32,708 bp. The expansion in inverted repeats led to duplication of ycf1 genes in four species. The genera showed high similarity in gene content and yielded 113 unique genes (79 protein-coding, 4 rRNA, and 30 tRNA genes). Codon usage, amino acid frequency, RNA editing sites, microsatellites repeats, transition and transversion substitutions, and synonymous and non-synonymous substitutions were also similar across the clade. A previous study reported deletion of ycf1, accD, psbE, trnL-CAA, and trnG-GCC genes in four Amorphophallus species. Our study supports conservative structure of cp genomes in the Dracunculus clade including Amorphophallus species and does not support gene deletion mentioned above. We also report suitable polymorphic loci based on comparative analyses of Dracunculus clade species, which could be useful for phylogenetic inference. Overall, the current study broad our knowledge about the molecular evolution of chloroplast genome in aroids.

Sequence Variants in the WNT10B and TP63 Genes Underlying Isolated Split-Hand/Split-Foot Malformation.

Aims: Split-hand/split-foot malformation (SHFM) is a developmental and congenital limb malformation characterized by variable degrees of medial clefting or absence of one or more digits in hands and/or feet. The aim of this study was to identify the underlying cause of three consanguineous Pakistani families showing various types of SHFM-related features. Materials and Methods: Standard molecular methods, including whole-genome sequencing (WGS), whole-exome sequencing (WES), microsatellite markers-based genotyping, and Sanger sequencing were performed to search for the likely causative variants. Results: In family A, WES revealed a novel homozygous missense variant [c.338G>A, p.(Gly113Asp)] in the WNT10B gene. In family B, microsatellite-based genotyping followed by Sanger sequencing revealed a novel homozygous 13 base pairs deletion [c.884-896delTCCAGCCCCGTCT, p.(Phe295Cysfs*87)] in the same gene. In family C, WGS divulged a previously reported heterozygous missense variant [c.956G>A, p.(Arg319His)] in the TP63 gene. Conclusions: Mapping and sequencing genes and variants for severe skeletal disorders, such as SHRM, will facilitate establishing specific genotype-phenotype correlations and providing genetic counseling for the families suffering from such conditions.

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.

Identification of a novel biallelic missense variant in the KIAA0825 underlies postaxial polydactyly type A.

Postaxial polydactyly (PAP) is characterized by development of extra digits, which mostly segregates in autosomal recessive pattern. The underlying genetic cause of recessive non-syndromic PAP type A has been associated with sequence variants in five different genes (ZNF141, IQCE, GLI1, FAM92A, KIAA0825). The present study was aimed to investigate clinical and genetic causes of PAPA in a consanguineous family of Pakistani origin. Microsatellite-based linkage analysis was used to search for the disease-causing gene. Linkage in the family was established at chromosome 5q15 harbouring a candidate gene KIAA0825. Subsequently, Sanger sequencing revealed a novel homozygous missense variant [c.50T>C; p. (Leu17Ser)] in the gene, which co-segregated with the disease within the family. Protein structural analysis predicted a substantial change in the secondary structure of the mutant protein affecting its function. This is the third disease causing variant identified in the KIAA0825. This has not only expanded spectrum of the mutations in the gene but also further substantiated its role in the limb development in human.

Nonsyndromic Split-Hand/Foot Malformation: Recent Classification.

Split-hand/foot malformation (SHFM) is a genetic limb anomaly disturbing the central rays of the autopod. SHFM is a genetically heterogeneous disorder with variable expressivity inherited as syndromic and nonsyndromic forms. We provide an update of the clinical and molecular aspects of nonsyndromic SHFM. This rare condition is highly complex due to the clinical variability and irregular genetic inheritance observed in the affected individuals. Nonsyndromic SHFM types have been reviewed in terms of major molecular genetic alterations reported to date. This updated overview will assist researchers, scientists, and clinicians in making an appropriate molecular diagnosis, providing an accurate recurrence risk assessment, and developing a management plan.

A novel missense variant in the BBS7 gene underlying Bardet-Biedl syndrome in a consanguineous Pakistani family.

Bardet-Biedl syndrome (BBS) is characterized by six major features: postaxial polydactyly, obesity, learning disabilities, renal anomalies, retinitis pigmentosa and hypogonadism and is inherited in an autosomal recessive manner. BBS is caused by disease causing sequence variants in the 22 BBS genes identified to date. In the present study, a single consanguineous Pakistani Family with BBS was clinically and genetically characterized. After establishing linkage to a BBS gene on chromosome 4q27, Sanger sequencing was performed in all available affected and unaffected members. Sequence analysis of the BBS7 gene revealed novel substitution mutation (c.719G>T; p. Gly240Val). Our findings further extend the body of evidence implicating BBS7 in causing BBS and expand the mutation spectrum.

Biallelic Missense Mutation in the ECEL1 Underlies Distal Arthrogryposis Type 5 (DA5D).

Distal arthrogryposis (DA) is a heterogeneous sub-group of arthrogryposis multiplex congenita (AMC), mostly characterized by having congenital contractures affecting hands, wrists, feet, and ankles. Distal arthrogryposis is mostly autosomal dominantly inherited, while only one sub-type DA type 5D is inherited in an autosomal recessive manner. Clinically, DA5D is described having knee extension contractures, micrognathia, distal joint contractures, clubfoot, ptosis, contractures (shoulders, elbows, and wrists), and scoliosis. Using whole exome sequencing (WES) followed by Sanger sequencing, we report on a first familial case of DA5D from Pakistani population having a novel biallelic missense mutation (c.158C>A, p.Pro53Leu) in the ECEL1 gene. Our result support that homozygous mutations in ECEL1 causes DA5D and expands the clinical and allelic spectrum of ECEL1 related contracture syndromes.

A Fatigue Measuring Protocol for Wireless Body Area Sensor Networks.

As players and soldiers preform strenuous exercises and do difficult and tiring duties, they are usually the common victims of muscular fatigue. Keeping this in mind, we propose FAtigue MEasurement (FAME) protocol for soccer players and soldiers using in-vivo sensors for Wireless Body Area Sensor Networks (WBASNs). In FAME, we introduce a composite parameter for fatigue measurement by setting a threshold level for each sensor. Whenever, any sensed data exceeds its threshold level, the players or soldiers are declared to be in a state of fatigue. Moreover, we use a vibration pad for the relaxation of fatigued muscles, and then utilize the vibrational energy by means of vibration detection circuit to recharge the in-vivo sensors. The induction circuit achieves about 68 % link efficiency. Simulation results show better performance of the proposed FAME protocol, in the chosen scenarios, as compared to an existing Wireless Soccer Team Monitoring (WSTM) protocol in terms of the selected metrics.

DEADS: Depth and Energy Aware Dominating Set Based Algorithm for Cooperative Routing along with Sink Mobility in Underwater WSNs.

Performance enhancement of Underwater Wireless Sensor Networks (UWSNs) in terms of throughput maximization, energy conservation and Bit Error Rate (BER) minimization is a potential research area. However, limited available bandwidth, high propagation delay, highly dynamic network topology, and high error probability leads to performance degradation in these networks. In this regard, many cooperative communication protocols have been developed that either investigate the physical layer or the Medium Access Control (MAC) layer, however, the network layer is still unexplored. More specifically, cooperative routing has not yet been jointly considered with sink mobility. Therefore, this paper aims to enhance the network reliability and efficiency via dominating set based cooperative routing and sink mobility. The proposed work is validated via simulations which show relatively improved performance of our proposed work in terms the selected performance metrics.

Spectrofluorimetric analysis of famotidine in pharmaceutical preparations and biological fluids by derivatization with benzoin.

A sensitive and simple spectrofluorimetric method has been developed for the analysis of famotidine, from pharmaceutical preparations and biological fluids after derivatization with benzoin. The reaction was carried out in alkaline medium with measurement of fluorescence intensity at 446 nm with excitation wavelength at 286 nm. Linear calibration was obtained with 0.5-15 µg/ml with coefficient of determination (r(2)) 0.997. The factors affecting the fluorescence intensity were optimized. The pharmaceutical additives and amino acid did not interfere in the determination. The mean percentage recovery (n=4) calculated by standard addition from pharmaceutical preparation was 94.8-98.2% with relative standard deviation (RSD) 1.56-3.34% and recovery from deproteinized spiked serum and urine of healthy volunteers was 98.6-98.9% and 98.0-98.4% with RSD 0.34-0.84% and 0.29-0.87% respectively.

Listening to patients' voices: linguistic indicators related to diabetes self-management.

A great deal of research in health care has examined a wide range of variables to better understand the degree to which patients follow the advice of medical professionals in managing their health, known as adherence. This paper explains the development of the linguistic systems to describe and evaluate two psychosocial constructs (i.e. control orientation and agency) that have been found to be related to adherence in previous research for subjects with diabetes (Trento et al. 2007; Wangberg 2007; O'Hea et al. 2009). The present data came from 43 semi-structured in-depth interviews of subjects with Type 2 diabetes. One-on-one interviews with open-ended questions elicited subjects' 'stories' about living with diabetes, and the transcribed interviews were analyzed to develop the linguistic systems of control orientation and agency. The resultant systems were applied to the 43 interviews by raters with high inter-rater reliability. The results showed demarcations of clearly identified codings of patient types. The paper presents the linguistic coding systems developed in the study, the results of their application to the patient interview data, and recommendations for improved communication with patients.

Long-term kidney transplant outcome in obese patients in a predominantly African American population.

The impact of obesity on long-term kidney transplant outcome has largely been studied in non-African American patients. This study seeks to determine differences in outcome between obese and non-obese patients after kidney transplantation, in a predominantly African American population. We reviewed 642 adult renal transplant recipients who received their transplants at SUNY Downstate Medical Center between 1998 and 2007. Sixty-six percent of the patients studied were African American. The patients were divided into five groups according to their BMI status: underweight <20, normal 20-24.9, overweight 25-29.9, obese 30-34.9, and morbidly obese ≥35. There were no differences in race, gender, cytomegalovirus infection, type of transplant, panel-reactive antibody, retransplant status, flow cytometry cross-match results, mycophenolate mofetil therapy, and total HLA mismatch status. The mean discharge serum creatinine in the morbidly obese group was significantly higher than in other groups (p < 0.001). The difference in creatinine level disappeared at six wk and six months (p > 0.5), respectively. Acute rejection rates, delayed graft function, graft survival, and patient survival were not different between the groups. The findings from this large single-center study suggest that obese and morbidly obese patients had similar outcomes compared to other weight groups. Obese and morbidly obese African American patients should not be excluded from kidney transplantation on the basis of weight alone.

New-onset diabetes after hemodialysis initiation: impact on survival.

BACKGROUND: The incidence of new-onset diabetes after initiation of hemodialysis (NODAD) and its impact on survival is not known. METHODS: We used data from the United States Renal Data System (USRDS) from January 2000 to December 2001, with at least 3 years of follow-up for this study. Patients aged 18-80 years were included. NODAD was defined as two Medicare institutional claims for diabetes in patients with no history of diabetes prior to starting hemodialysis (HD). Incidence (per 1,000 patient-years), prevalence (%) and hazard ratios for mortality in patients with NODAD were calculated. RESULTS: There were 59,340 incident patients with no history of diabetes prior to starting HD, of which 3,853 met criteria for NODAD. The overall incidence and prevalence of NODAD were 20 per 1,000 patient-years and 7.6%, respectively. In a cohort of 444 patients without diabetes and documented glycosylated hemoglobin A1c, <6% prior to starting HD (from January 2005 and March 2006), at a mean follow-up of 4.7 +/- 2.6 months, 6.8% developed NODAD defined by two Medicare claims for diabetes after initiation of HD. NODAD was associated with a significantly increased risk of death as compared to non-diabetes patients (hazard ratio 1.20, 95% confidence interval 1.14-1.25). CONCLUSION: The USRDS showed a high incidence of NODAD, associated with significantly higher mortality compared to those who did not develop NODAD. The mechanism of NODAD needs to be explored further in experimental and clinical studies.

Effect of ethnicity on the progression of diabetic kidney disease independent of glycemic control.

BACKGROUND/AIMS: The prevalence of diabetic kidney disease (DKD) and risk of progression to end-stage renal disease is higher in African-Americans as compared to Caucasians. Whether the higher rate of estimated glomerular filtration rate (eGFR) decline in African-Americans is mediated by poor glycemic control is unclear. METHODS: We conducted a prospective study of 183 (African-American, n = 95; Caucasian, n = 88, mean age 66 +/- 10 vs. 70 +/- 11 years) patients with a diagnosis of DKD followed over a period of 12 months. eGFR (ml/min/1.73 m(2)) was calculated by MDRD formula and grouped into stage 1-2 (> or =60 ml/min), stage 3 (30-60 ml/min) and stage 4 (<30 ml/min). In addition, glycosylated hemoglobin A1C (HbA1c) was categorized into tertiles (<7, 7-8 and >8%) at each time point. RESULTS: There were no significant differences in eGFR at any time point between African-American and Caucasian in any stage of CKD during this period. There were also no significant differences in eGFR at any time point in each category of HbA1c. CONCLUSIONS: Our data indicate that there were no ethnic differences in the rate of progression of DKD under equivalent glycemic control. Further research is needed to explore the mechanisms associated with higher prevalence and rapid progression of CKD in African-Americans compared to Caucasians.

Long-term outcomes of dual kidney transplantation-a single center experience.

The shortage of kidney donors has led to broadening of the acceptance criteria for deceased donor organs beyond the traditional use of young donors. We determined long-term post-transplant outcomes in recipients of dual expanded criteria donor kidneys (dECD, n = 44) and compared them to recipients of standard criteria donor kidneys (SCD, n = 194) and single expanded criteria donor kidneys (sECD, n = 62). We retrospectively reviewed these 300 deceased donor kidney transplants without primary non-function (PNF) or death in the first two wk, at our center from 1996 to 2003. The three groups were similar in baseline characteristics. Kidney allograft survival and patient survival (nine yr) were similar in the three respective donor groups, SCD, sECD and dECD (60% vs. 59% vs. 64% and 82% vs. 73% vs. 73%). Acute rejection in the first three months was 23.2%, 16.1%, and 22.7% in SCD, sECD and dECD, respectively (p = 0.49) and delayed graft function was 25.2%, 31.9% and 17.1% in the three groups, respectively (p = 0.28). When PNF and death within the first two wk was included, there was no significant difference in graft survival between the three groups. In our population, recipients of dECD transplants have acceptable patient and graft survival with kidneys that would have usually been discarded.

Posttransplant diabetes and hypertension: pathophysiologic insights and therapeutic rationale.

New-onset diabetes after transplantation and hypertension are very common after renal transplantation and are associated with adverse graft and cardiovascular outcomes. A thorough understanding of the unique factors that operate in renal transplant recipients is essential for the proper evaluation and management of these important disorders. This review outlines the pathogenesis, diagnostic workup, and therapeutic rationale for diabetes and hypertension after transplantation.

Erythropoietin stimulating agents in the management of anemia of chronic kidney disease.

Anemia is a very common clinical problem in patients with chronic kidney disease (CKD) and is associated with increased morbidity and mortality in these patients. Erythropoietin is a hormone synthesized in the kidney responsible for red blood cell maturation in the bone marrow. It is deficient in the majority of patients with advanced kidney disease thereby predisposing to anemia. Since the approval of recombinant human erythropoietin (epoetin alfa) by the US FDA in 1989, epoetin alfa and similar agents now collectively known as erythropoietin stimulating agents (ESA) have become the standard of care for the treatment of the erythropoietin-deficient anemia that occurs in most patients with CKD. In this review, we have outlined the considerations that need to be taken into account when prescribing ESA for the treatment of anemia in CKD.