N-acetyltransferase polymorphism among northern Sudanese

Interindividual and interethnic differences in allele frequencies of N-acetyltransferase (NAT2) single nucleotide polymorphisms (SNPs) are responsible for phenotypic variability of adverse drug reactions and susceptibility to cancer. We genotyped the seven NAT2 common SNPs in 127 randomly selected unrelated northern Sudanese subjects using allele-specific and RFLP polymerase chain reaction (PCR) based methods. Molecular genotyping was enough to designate alleles for 41 individuals unambiguously, whereas 63 individuals' alleles were inferred from haplotypes previously described. In the remaining 23 individuals, however, the phase of the SNPs could not be decided because of multiple SNP heterozygotes. Using computational methods in the HAP and Phase programs, we confirmed the inferred alleles of the 62 individuals and predicted the remaining 23 ambiguous alleles. Twelve NAT2 alleles were identified. Four alleles coded for rapid acetylators (18%), and eight alleles coded for slow acetylators (82%). Two genotypes coded for rapid acetylation (3.9%), 10 for intermediate acetylation (27.6%), and 13 for slow acetylation (68.5%). The G191A African SNP and the G857A predominantly Asian SNP were each detected at a low frequency of 3.1%. The combination of molecular and computational analysis was useful in resolving ambiguous genotypes of NAT2 in multiple SNP heterozygotes. Among the northern Sudanese the SNPs associated with slow acetylation are more prevalent than in Caucasians and Asians. This and other African studies are suggestive of an African origin for NAT2-associated polymorphism.     

Identifying people at high risk for developing sleep apnea syndrome (SAS): a cross-sectional study in a Pakistani population

Background  

Obstructive Sleep Apnea (OSA) is associated with many cardiovascular and psychiatric diseases. Day-time sleepiness is a common consequence of sleep apnea and correlates with road-traffic accidents (RTA). Pakistan has a high prevalence of factors which predispose an individual to OSA and death from RTAs are a huge burden. However there is a dearth of prevalence studies in this regard. We aim to understand local relevance of the disease and estimate the prevalence of individuals high-risk for OSA.     

Role of urinary NMP-22 combined with urine cytology in follow-up surveillance of recurring superficial bladder urothelial carcinoma

OBJECTIVE: To determine efficacy and utility of NMP-22 in follow-up of bladder urothelial carcinoma (UC) and compare NMP-22 as a single evaluating test vs combination with cytology. STUDY DESIGN: Ninety-four consecutive urine cytology samples of bladder UC were identified. Patients received follow-up urine cytology, NMP-22 testing and cystoscopy with surgical biopsy. RESULTS: NMP-22 specificity was 100%, sensitivity 45%, positive predictive value (PPV) 100% and negative predictive value (NPV) 87%. NMP-22 showed lower sensitivity for high-grade lesions and higher for low-grade lesions. Cytologic diagnosis had a high inconclusive rate; when regarded as positive, it resulted in 75% sensitivity, 58% specificity, 33% PPV and 89% NPV. NMP-22 correctly classified 60% of false negative cases diagnosed by cytology with low-grade UC and clarified 27 inconclusive cytologic diagnoses. NMP-22 misclassified 9 cases as false negative, all with high-grade UC; all were correctly identified on cytology as true positive. Combined interpretation showed 90% sensitivity, 92% specificity, 75% PPV and 98% NPV. CONCLUSION: NMP-22 complements cytology by its higher sensitivity for low-grade lesions; its values are not affected by bacillus Calmette Guérin therapy changes, which are limiting in cytology. Combined interpretation of NMP-22 and cytology shows promise as an effective, noninvasive method for surveillance of UC.     

Generation of human embryonic stem cell-derived mesoderm and cardiac cells using size-specified aggregates in an oxygen-controlled bioreactor

The ability to generate human pluripotent stem cell-derived cell types at sufficiently high numbers and in a reproducible manner is fundamental for clinical and biopharmaceutical applications. Current experimental methods for the differentiation of pluripotent cells such as human embryonic stem cells (hESC) rely on the generation of heterogeneous aggregates of cells, also called "embryoid bodies" (EBs), in small scale static culture. These protocols are typically (1) not scalable, (2) result in a wide range of EB sizes and (3) expose cells to fluctuations in physicochemical parameters. With the goal of establishing a robust bioprocess we first screened different scalable suspension systems for their ability to support the growth and differentiation of hESCs. Next homogeneity of initial cell aggregates was improved by employing a micro-printing strategy to generate large numbers of size-specified hESC aggregates. Finally, these technologies were integrated into a fully controlled bioreactor system and the impact of oxygen concentration was investigated. Our results demonstrate the beneficial effects of stirred bioreactor culture, aggregate size-control and hypoxia (4% oxygen tension) on both cell growth and cell differentiation towards cardiomyocytes. QRT-PCR data for markers such as Brachyury, LIM domain homeobox gene Isl-1, Troponin T and Myosin Light Chain 2v, as well as immunohistochemistry and functional analysis by response to chronotropic agents, documented the impact of these parameters on cardiac differentiation. This study provides an important foundation towards the robust generation of clinically relevant numbers of hESC derived cells.     

Evaluation of ARG protein expression in mature B cell lymphomas compared to non-neoplastic reactive lymph node

The participation of Abl-Related Gene (ARG) is demonstrated in pathogenesis of different human malignancies. However there is no conclusive evidence on ARG expression level in mature B cell lymphomas. In this study we evaluated ARG protein expression in Follicular Lymphoma (FL), Burkitt’s Lymphoma (BL) and Diffused Large B Cell Lymphoma (DLBCL) in comparison with non-neoplastic lymph nodes. Semi-quantitative fluorescent ImmunoHistoChemistry was applied on 14, 7 and 4 patients with DLBCL, FL and BL respectively, adding to 4 normal and 4 reactive lymph nodes. The mean ratio of ARG/GAPDH expression was significantly different (p < 0.00) between lymphomas and control samples, with DLBCL having the highest ARG expression amongst all. Over expression of ARG was seen in FL and BL, with FL expressing statistically more ARG than BL. Moreover, the ARG/GAPDH expression ratio increased from DLBCL stage I towards stage VI, all showing significantly more ARG expression than FL and BL (in all cases p < 0.00).     

Offset configurations for single- and double-strand DNA inside single-walled carbon nanotubes

Nanotechnology is a rapidly expanding research area, and it is believed that the unique properties of molecules at the nano-scale will prove to be of substantial benefit to mankind, especially so in medicine and electronics. Here we use applied mathematical modelling exploiting the basic principles of mechanics and the 6–12 Lennard-Jones potential function together with the continuum approximation, which assumes that intermolecular interactions can be approximated by average atomic surface densities. We consider the equilibrium offset positions for both single-strand and double-strand DNA molecules inside a single-walled carbon nanotube, and we predict offset positions with reference to the cross-section of the carbon nanotube. For the double-strand DNA, the potential energy is determined for the general case for any helical phase angle ?, but we also consider a special case when ? = π, which leads to a substantial simplification in the analytical expression for the energy. As might be expected, our results confirm that the global minimum energy positions for a single-strand DNA molecule and a double-strand DNA molecule will lie off axis and they become closer to the tube wall as the radius of the tube increases.     

The Development of the Pediatric NAFLD Fibrosis Score (PNFS) to Predict the Presence of Advanced Fibrosis in Children with Nonalcoholic Fatty Liver Disease

Background

Noninvasive hepatic fibrosis scores that predict the presence of advanced fibrosis have been developed and validated in adult patients with NAFLD. The aims of our study were to assess the utility of commonly used adult fibrosis scores in pediatric NAFLD and to develop a pediatric specific fibrosis score that can predict advanced fibrosis.

Methods

Consecutive children with biopsy-proven NAFLD were included. Fibrosis was determined by an experienced pathologist (F0–4). Advanced fibrosis was defined as fibrosis stage ≥3. The following adult fibrosis scores were calculated for each child: AST/ALT ratio, AST/platelet ratio index (APRI), NAFLD fibrosis score (NFS), and FIB-4 Index. Multivariable logistic regression analysis was performed to build a new pediatric model for predicting advanced fibrosis.

Results

Our cohort consisted of 242 children with a mean age of 12.4±3.1 years and 63% were female. 36 (15%) subjects had advanced fibrosis. APRI and FIB-4 were higher in patients with advanced fibrosis compared to those with fibrosis stage 0–2; however, AST/ALT ratio and NFS were not different between the two groups. We used our data to develop a new model to predict advanced fibrosis which included: ALT, alkaline phosphatase, platelet counts and GGT. The multivariable logistic regression model (z) was defined as follows: z = 1.1+(0.34*sqrt(ALT))+(0.002*alkaline phosphatase) – (1.1*log(platelets) – (0.02*GGT). This value was then converted into a probability distribution (p) with a value between 0 to 100 by the following formula: p = 100×exp(z)/[1+exp(z)]. The AUCROC for this model was 0.74 (95% CI: 0.66, 0.82). This was found to be significantly better than APRI, NAFLD Fibrosis Score and FIB-4 Index.

Conclusion

Noninvasive hepatic fibrosis scores developed in adults had poor performance in diagnosing advanced fibrosis in children with NAFLD. We developed a new pediatric NAFLD fibrosis score with improved performance characteristics.     

Therapeutic Efficacy of an ω-3-Fatty Acid-Containing 17-β Estradiol Nano-Delivery System against Experimental Atherosclerosis

Atherosclerosis and its consequences remain prevalent clinical challenges throughout the world. Initiation and progression of atherosclerosis involves a complex, dynamic interplay among inflammation, hyperlipidemia, and endothelial dysfunction. A multicomponent treatment approach targeted for delivery within diseased vessels could prove beneficial in treating atherosclerosis. This study was undertaken to evaluate the multimodal effects of a novel ω-3-fatty acid-rich, 17-β-estradiol (17-βE)-loaded, CREKA-peptide-modified nanoemulsion system on experimental atherosclerosis. In vitro treatment of cultured human aortic endothelial cells (ECs) with the 17-βE-loaded, CREKA-peptide-modified nanoemulsion system increased cellular nitrate/nitrite, indicating improved nitric oxide formation. In vivo, systemic administration of this nanoemulsion system to apolipoprotein-E knock out (ApoE^-/-) mice fed a high-fat diet significantly improved multiple parameters related to the etiology and development of occlusive atherosclerotic vasculopathy: lesion area, circulating plasma lipid levels, and expression of aortic-wall inflammatory markers. These salutary effects were attributed selectively to the 17-βE and/or ω-3 polyunsaturated fatty acid components of the nano-delivery system. At therapeutic doses, the 17-βE-loaded, CREKA-peptide modified nanoemulsion system appeared to be biocompatible in that it elicited no apparent adverse/toxic effects, as indexed by body weight, plasma alanine aminotransferase/aspartate aminotransferase levels, and liver and kidney histopathology. The study demonstrates the therapeutic potential of a novel, 17-βE-loaded, CREKA-peptide-modified nanoemulsion system against atherosclerosis in a multimodal fashion by reducing lesion size, lowering the levels of circulating plasma lipids and decreasing the gene expression of inflammatory markers associated with the disease.     

Comparative genomic hybridization arrays in clinical pathology: progress and challenges

Array-based comparative genomic hybridization (array CGH) genome scanning is a powerful method for the global detection of gains and losses of genetic material in both congenital and neoplastic disorders. When used as a clinical diagnostic test, array CGH combines the whole genome perspective of traditional G-banded cytogenetics with the targeted identification of cryptic chromosomal abnormalities characteristic of fluorescence in situ hybridization (FISH). However, the presence of structural variants in the human genome can complicate analysis of patient samples, and array CGH does not provide morphologic information about chromosome structure, balanced translocations, or the actual chromosomal location of segmental duplications. Identification of such anomalies has significant diagnostic and prognostic implications for the patient. We therefore propose that array CGH should be used as a guide to the presence of genomic structural rearrangements in germline and tumor genomes that can then be further characterized by FISH or G-banding, depending on the clinical scenario. In this article, we share some of our experiences with diagnostic array CGH and discuss recent progress and challenges involved with the integration of array CGH into clinical laboratory medicine.