Computational validation of ABCB1 gene polymorphism and its effect on tacrolimus dose concentration/levels in renal transplant individuals of South India.

Patients with end-stage renal failure require hemodialysis and peritoneal dialysis; however, kidney transplantation is considered a better treatment option for renal failure patients, improving their quality of life and longevity. Among several potent immunosuppressive agents, tacrolimus (TAC) has shown progressive improvement in the graft survival rates after renal transplantation. Fifty kidney transplant patients undergoing TAC immunosuppressive treatment were included. The human genomic DNA was isolated using the phenol-chloroform extraction procedure. CYP3A5*6, CYP3A5*2, and ABCB1 exon 21 G2677 T/A polymorphisms were genotyped using the polymerase chain reaction-restriction fragment length polymorphism method. Fisher's exact test and Chi-square analysis were performed to analyze the data, where p < 0.05 was considered statistically significant. In addition, we implemented bioinformatics studies on ABCB1 protein to determine the mutation's effect sequentially and structurally. Among the genotyped single nucleotide polymorphisms (SNPs), SNPs of CYP3A5*2 and CYP3A5*6 did not vary in the studied population. The concentration/dose (C/D) ratio of TT genotype of the ABCB1 gene was higher (95% CI: 177.38-269.46) when compared to TA and AA. However, there were no substantial differences between the ABCB1 genotypes and TAC C/D ratio (p = 0.953). The TAC dose mg/kg/day (p = 0.002) and C/D ratio (p = 0.004) exhibited a statistically significant difference. However, no significant difference was found with respect to the ABCB1 gene between the non-toxicity and toxicity groups. Mutation and residue interaction analysis results showed that the S893T mutation destabilizes the ABCB1 protein, thus reducing the protein's flexibility. The present study demonstrated a substantial relationship between the TAC dose and C/D ratio, including the non-toxicity and toxicity groups. However, no possible correlation was observed between the ABCB1 gene polymorphism and renal transplant.

Potential application of γ-H2AX as a biodosimetry tool for radiation triage.

Radiation triage and biological dosimetry are two initial steps in the medical management of exposed individuals following radiological accidents. Well established biodosimetry methods such as the dicentric (DC) assay, micronucleus (MN) assay, and fluorescence in-situ hybridization (FISH) translocation assay (for residual damage) have been used for this purpose for several decades. Recent advances in scoring methodology and networking among established laboratories have increased triage capacity; however, these methods still have limitations in analysing large sample numbers, particularly because of the ∼ 48 h minimum culture time required prior to analysis. Hence, there is a need for simple, and high throughput markers to identify exposed individuals in case of radiological/nuclear emergencies. In recent years, a few markers were identified, one being phosphorylated histone 2AX (γ-H2AX), which measured a nuclear foci or nuclear staining intensity that was found to be suitable for triage. Measurement of γ-H2AX foci formed at and around the sites of DNA double-strand breaks is a rapid and sensitive biodosimetry method which does not require culturing and is thus promising for the analysis of a large number of samples. In this review, we have summarized the recent developments of γ-H2AX assay in radiation triage and biodosimetry, focusing chiefly on: i) the importance of baseline frequency and reported values among different laboratories, ii) the influence of known and unknown variables on dose estimation, iii) quality assurance such as inter-laboratory comparison between scorers and scoring methods, and iv) current limitations and potential for future development.