R.N. Sharan

Betel Nut and Susceptibility to Cancer

Betel nut is a widely masticated natural product, which is consumed by over 600 million people across the globe. The ancient habit of betel nut chewing, either as dry or raw/wet nut, in association with betel leaf and a host of region specific additives, including chewing tobacco, is believed to be an important etiological factor for human cancer. Alkaloids and their betel nut specific nitrosamine derivatives produced upon metabolic activation interact with DNA and other cellular targets to produce highly variable mutagenic, genotoxic, cytostatic, immunostatic and teratogenic effects. At molecular level the betel nut or its constituents strongly influence gene expression patterns, especially that of tumor suppressor genes. Structural damage to nucleus and mitochondria, etc. are also induced. The review dwells upon these aspects of betel nut induced carcinogenesis to show that genetic susceptibility to cancer through generations progressively increased due to exposure to betel nut.

Biological effects induced by swift heavy ions of lithium on aqueous solution of plasmid pMTa4

Biological samples exposed to swift heavy ions sustain damage on different components. Damage to DNA, a critical component of a living system, has considerable biological implications. In this study aqueous solution of plasmid pMTa4 was exposed to varying fluence of swift 7Li ions and its different topological forms were analysed by agarose gel electrophoresis to study the induced damage. To monitor radiation labile nucleotide sequence the 7Li ions exposed plasmid was degraded by three different restriction endonucleases and also analysed by agarose gel electrophoresis. The results show that 7Li ions predominantly induced double strand breaks in the plasmid DNA in a dose-dependent manner and affected preferentially the GC-rich motifs of the DNA. The results suggest that 7Li ions induce premutagenic lesions at an enhanced frequency in segments of the DNA which are rich in CG content as compared to GC-poor segments.

Evaluation of endogenous control gene(s) for gene expression studies in human blood exposed to 60Co γ-rays ex vivo.

In gene expression studies, it is critical to normalize data using a stably expressed endogenous control gene in order to obtain accurate and reliable results. However, we currently do not have a universally applied endogenous control gene for normalization of data for gene expression studies, particularly those involving 60Co γ-ray-exposed human blood samples. In this study, a comparative assessment of the gene expression of six widely used housekeeping endogenous control genes, namely 18S, ACTB, B2M, GAPDH, MT-ATP6 and CDKN1A, was undertaken for a range of 60Co γ-ray doses (0.5, 1.0, 2.0 and 4.0 Gy) at 8.4 Gy min−1 at 0 and 24 h post-irradiation time intervals. Using the NormFinder algorithm, real-time PCR data obtained from six individuals (three males and three females) were analyzed with respect to the threshold cycle (Ct) value and abundance, ΔCt pair-wise comparison, intra- and inter-group variability assessments, etc. GAPDH, either alone or in combination with 18S, was found to be the most suitable endogenous control gene and should be used in gene expression studies, especially those involving qPCR of γ-ray-exposed human blood samples.

Consensus reference gene(s) for gene expression studies in human cancers: end of the tunnel visible?

BackgroundGene expression studies are increasingly used to provide valuable information on the diagnosis and prognosis of human cancers. Also, for in vitro and in vivo experimental cancer models gene expression studies are widely used. The complex algorithms of differential gene expression analyses require normalization of data against a reference or normalizer gene, or a set of such genes. For this purpose, mostly invariant housekeeping genes are used. Unfortunately, however, there are no consensus (housekeeping) genes that serve as reference or normalizer for different human cancers. In fact, scientists have employed a wide range of reference genes across different types of cancer for normalization of gene expression data. As a consequence, comparisons of these data and/or data harmonizations are difficult to perform and challenging. In addition, an inadequate choice for a reference gene may obscure genuine changes and/or result in erroneous gene expression data comparisons.MethodsIn our effort to highlight the importance of selecting the most appropriate reference gene(s), we have screened the literature for gene expression studies published since the turn of the century on thirteen of the most prevalent human cancers worldwide.ConclusionsBased on the analysis of the data at hand, we firstly recommend that in each study the suitability of candidate reference gene(s) should carefully be evaluated in order to yield reliable differential gene expression data. Secondly, we recommend that a combination of PPIA and either GAPDH, ACTB, HPRT and TBP, or appropriate combinations of two or three of these genes, should be employed in future studies, to ensure that results from different studies on different human cancers can be harmonized. This approach will ultimately increase the depth of our understanding of gene expression signatures across human cancers.

Gp78 involvement in cellular proliferation: Can act as a promising modulator for cell cycle regulatory proteins?

In cells, protein synthesis and degradation are normal processes, which are tightly regulated by various cellular metabolic pathways. Cellular protein quality control (PQC) mechanisms always present a continuous and rigorous check over all intracellular proteins before they can participate in various cellular physiological processes with the help of PQC pathways like autophagy and ubiquitin proteasome system (UPS). The UPS employs few selective E3 ubiquitin ligases for the intracellular degradation of cyclin‐dependent kinase inhibitor 1B (p27Kip1) that tightly controls cell cycle progression. But, the complex mechanistic interactions and the interplay between E3 ubiquitin ligases involved in the functional regulation as well as expression of p27 are not well known. Here, we demonstrate that cell surface glycoprotein Gp78, a putative E3 ubiquitin ligase, is involved in the stabilization of intracellular steady‐state levels of p27. Transient overexpression of Gp78 increases the accumulation of p27 in cells in the form of massive inclusions like structures, which could be due to its cumulative increased stability in cells. We have also monitored how under stress condition, E3 ubiquitin ligase Gp78 regulates endogenous levels of p27 in cells. ER stress treatment generates a marginal increase in Gp78 endogenous levels, and this elevation effect was prominent for intracellular accumulation of p27 in cells. Taken together, our current findings suggest a valuable multifactorial regulatory mechanism and linkage of p27 with UPS pathway.

Identification and preliminary validation of radiation response protein(s) in human blood for a high-throughput molecular biodosimetry technology for the future

The absence of a rapid and high-throughput technology for radiation biodosimetry has been a great obstacle in our full preparedness to cope with large-scale radiological incidents. The existing cytogenetic technologies have limitations, primarily due to their time-consuming methodologies, which include a tissue culture step, and the time required for scoring. This has seriously undermined its application in a mass casualty scenario under radiological emergencies for timely triage and medical interventions. Recent advances in genomics and proteomics in the postgenomic era have opened up new platforms and avenues to discover molecular biomarkers for biodosimetry in the future. Using a genomic-to-proteomic approach, we have identified a basket of twenty “candidate” radiation response genes.(RRGs) using DNA microarray and tools of bioinformatics immediately after ex vivo irradiation of freshly drawn whole blood of consenting and healthy human volunteers. The candidate RRGs have partially been validated using real-time quantitative polymerase chain reaction.(RT-qPCR or qPCR) to identify potential “candidate” RRGs at mRNA level. Two potential RRGs, CDNK1A and ZNF440, have so far been identified as genes with potentials to form radiation response proteins in liquid biopsy of blood, which shall eventually form the basis of fluorescence- or ELISA-based quantitative immunoprobe assay for a high-throughput technology of molecular biodosimetry in the future. More work is continuing.

Metal ion-catalyzed mineralization and biodetoxification studies of Calconcarboxylic acid in aqueous solution: effect of –COOH group

AbstractMineralization and biodetoxification studies of Calconcarboxylic acid (CCA), an azo dye, were undertaken by metal ion-catalyzed oxidation processes, viz. Fenton and Fenton-like processes and the effect of presence of –COOH group thereon were established. The metal ions used were Fe2+ and Ag+ and the oxidants used were hydrogen peroxide (HP) and ammonium persulfate (APS). For a given oxidant, Fe2+ is found to be the most suitable catalyst and among the oxidants, APS proves to be most suitable one. As all the systems yield maximum mineralization at pH 1, Fe2+/APS at pH 1 is the most suitable system. It is observed that mineralization is favored by Fe2+/HP process due to the presence of –COOH group in CCA and by Fe2+/APS process in its absence. Removal of –COOH group causes a shift in optimum pH, for Fenton process, to 3 accompanied by a corresponding drop in mineralization from 70.6 to 35.6% and an increase from 73.2 to 91.1% in Fe2+/APS system. Bioassay experiment reveals that the treated solutions...