Arif Tasleem Jan

Heavy Metals and Human Health: Mechanistic Insight into Toxicity and Counter Defense System of Antioxidants

Heavy metals, which have widespread environmental distribution and originate from natural and anthropogenic sources, are common environmental pollutants. In recent decades, their contamination has increased dramatically because of continuous discharge in sewage and untreated industrial effluents. Because they are non-degradable, they persist in the environment; accordingly, they have received a great deal of attention owing to their potential health and environmental risks. Although the toxic effects of metals depend on the forms and routes of exposure, interruptions of intracellular homeostasis include damage to lipids, proteins, enzymes and DNA via the production of free radicals. Following exposure to heavy metals, their metabolism and subsequent excretion from the body depends on the presence of antioxidants (glutathione, α-tocopherol, ascorbate, etc.) associated with the quenching of free radicals by suspending the activity of enzymes (catalase, peroxidase, and superoxide dismutase). Therefore, this review was written to provide a deep understanding of the mechanisms involved in eliciting their toxicity in order to highlight the necessity for development of strategies to decrease exposure to these metals, as well as to identify substances that contribute significantly to overcome their hazardous effects within the body of living organisms.

Outer Membrane Vesicles (OMVs) of Gram-negative Bacteria: A Perspective Update

Outer Membrane Vesicles (OMVs) of Gram-negative bacteria are spherical membrane-enclosed entities of endocytic origin. Reported in the consortia of different bacterial species, production of OMVs into extracellular milieu seems essential for their survival. Enriched with bioactive proteins, toxins, and virulence factors, OMVs play a critical role in the bacteria-bacteria and bacteria-host interactions. Emergence of OMVs as distinct cellular entities helps bacteria in adaptating to diverse niches, in competing with other bacteria to protect members of producer species and more importantly play a crucial role in host-pathogen interaction. Composition of OMV, their ability to modulate host immune response, along with coordinated secretion of bacterial effector proteins, endows them with the armory, which can withstand hostile environments. Study of the OMV production under natural and diverse stress conditions has broadened the horizons, and also opened new frontiers in delineating the molecular machinery involved in disease pathogenesis. Playing diverse biological and pathophysiological functions, OMVs hold a great promise in enabling resurgence of bacterial diseases, in concomitance with the steep decline in the efficiency of antibiotics. Having multifaceted role, their emergence as a causative agent for a series of infectious diseases increases the probability for their exploitation in the development of effective diagnostic tools and as vaccines against diverse pathogenic species of Gram-negative origin.

3-Deoxyglucosone: A Potential Glycating Agent Accountable for Structural Alteration in H3 Histone Protein through Generation of Different AGEs

Advanced glycation end-products (AGEs) are heterogeneous group of compounds, known to be implicated in diabetic complications. One of the consequences of the Maillard reaction is attributed to the production of reactive intermediate products such as α-oxoaldehydes. 3-deoxyglucosone (3-DG), an α-oxoaldehyde has been found to be involved in accelerating vascular damage during diabetes. In the present study, calf thymus histone H3 was treated with 3-deoxyglucosone to investigate the generation of AGEs (Nε-carboxymethyllysine, pentosidine), by examining the degree of side chain modifications and formation of different intermediates and employing various physicochemical techniques. The results clearly indicate the formation of AGEs and structural changes upon glycation of H3 by 3-deoxyglucosone, which may hamper the normal functioning of H3 histone, that may compromise the veracity of chromatin structures and function in secondary complications of diabetes.

Physicochemical Analysis of Structural Alteration and Advanced Glycation End Products Generation During Glycation of H2A Histone by 3-Deoxyglucosone

Advanced glycation end‐products comprise a complex and heterogeneous group of compounds that have been implicated in diabetes‐related complications. The importance of the Maillard reaction is depicted by the formation of reactive intermediate products known as α‐oxoaldehydes, such as 3‐deoxyglucosone (3‐DG). This product has been found to be involved in accelerated vascular damage in diabetes. In the present study, calf thymus histone H2A was reacted with 3‐DG, and the generation of advanced glycation end products was investigated by determining the degree of side chain modifications (lysine and arginine residues), Amadori products, carbonyl content, Nε‐carboxymethyl lysine, and pentosidine using various physicochemical techniques. Moreover, fluorescence, absorbance as well as structural characteristics of glycated‐H2A were comprehensively investigated. Overall, this study demonstrates structural perturbation, formation of different intermediates, and AGEs that are believed to hamper the normal functioning of H2A histone, compromising the integrity of chromatin structures and function in secondary complications of diabetes.

Binding of erucic acid with human serum albumin using a spectroscopic and molecular docking study

Erucic acid (EA) is one of the key fatty acids usually found in canola oil, mustard oil and rapeseed oil. Consumption of EA in primates was found to cause myocardial lipidosis and cardiac steatosis. To have an insight of the effect of EA in humans, we performed in vitro interaction studies of EA with the primary plasma protein, human serum albumin (HSA). Spectroscopic (UV-vis and fluorescence) analysis of the HSA-EA interaction revealed a static mode of quenching with binding constant Kb ∼104 reflecting high affinity of EA for HSA. The negative value of ΔG° for binding of EA to HSA in the fluorescence studies indicates the process to be spontaneous. Thermodynamic signatures of the HSA-EA interaction in the complex reflect dominance of hydrogen bonds. Despite predominance of hydrogen bonds, hydrophobic interactions in the HSA-EA complex were found acting as a contributing factor in the binding of EA to HSA, observed as structural change in the far-UV CD spectra. Försters resonance energy transfer of the EA-HSA complex revealed a distance of 3.2nm between acceptor molecules (EA) and the donor Trp residue of HSA. To have a deeper insight of the structural dependence of the HSA-EA interaction in the complex, thermodynamic study was supplemented with molecular docking. The molecular docking analysis further highlighted the EA binding in the subdomain IIIA (Sudlow site II) of HSA. The information generated in the study reflects greater pharmacological significance of EA and highlights its importance in the clinical medicine.

Network Analysis for the Identification of Differentially Expressed Hub Genes Using Myogenin Knock-down Muscle Satellite Cells

Muscle, a multinucleate syncytium formed by the fusion of mononuclear myoblasts, arises from quiescent progenitors (satellite cells) via activation of muscle-specific transcription factors (MyoD, Myf5, myogenin: MYOG, and MRF4). Subsequent to a decline in Pax7, induction in the expression of MYOG is a hallmark of myoblasts that have entered the differentiation phase following cell cycle withdrawal. It is evident that MYOG function cannot be compensated by any other myogenic regulatory factors (MRFs). Despite a plethora of information available regarding MYOG, the mechanism by which MYOG regulates muscle cell differentiation has not yet been identified. Using an RNA-Seq approach, analysis of MYOG knock-down muscle satellite cells (MSCs) have shown that genes associated with cell cycle and division, DNA replication, and phosphate metabolism are differentially expressed. By constructing an interaction network of differentially expressed genes (DEGs) using GeneMANIA, cadherin-associated protein (CTNNA2) was identified as the main hub gene in the network with highest node degree. Four functional clusters (modules or communities) were identified in the network and the functional enrichment analysis revealed that genes included in these clusters significantly contribute to skeletal muscle development. To confirm this finding, in vitro studies revealed increased expression of CTNNA2 in MSCs on day 12 compared to day 10. Expression of CTNNA2 was decreased in MYOG knock-down cells. However, knocking down CTNNA2, which leads to increased expression of extracellular matrix (ECM) genes (type I collagen α1 and type I collagen α2) along with myostatin (MSTN), was not found significantly affecting the expression of MYOG in C2C12 cells. We therefore propose that MYOG exerts its regulatory effects by acting upstream of CTNNA2, which in turn regulates the differentiation of C2C12 cells via interaction with ECM genes. Taken together, these findings highlight a new mechanism by which MYOG interacts with CTNNA2 in order to promote myoblast differentiation.

Towards Identifying Protective B-Cell Epitopes: The PspA Story

Pneumococcal surface protein A (PspA) is one of the most abundant cell surface protein of Streptococcus pneumoniae (S. pneumoniae). PspA variants are structurally and serologically diverse and help evade complement-mediated phagocytosis of S. pneumoniae, which is essential for its survival in the host. PspA is currently been screened for employment in the generation of more effective (serotype independent) vaccine to overcome the limitations of polysaccharide based vaccines, providing serotype specific immune responses. The cross-protection eliciting regions of PspA localize to the α-helical and proline rich regions. Recent data indicate significant variation in the ability of antibodies induced against the recombinant PspA variants to recognize distinct S. pneumoniae strains. Hence, screening for the identification of the topographical repertoire of B-cell epitopes that elicit cross-protective immune response seems essential in the engineering of a superior PspA-based vaccine. Herein, we revisit epitope identification in PspA and the utility of hybridoma technology in directing the identification of protective epitope regions of PspA that can be used in vaccine research.

Fibromodulin: A regulatory molecule maintaining cellular architecture for normal cellular function

Fibromodulin (FMOD) is a small leucine-rich proteoglycan that plays roles in a series of biological and pathophysiological processes. The interaction between FMOD and lysyl oxidase (LOX; collagen cross-linking enzyme) helps regulate extracellular matrix composition, and thereby, provides a permissive environment for regulating cellular turnover. FMOD has been mostly studied in the context of matrix component assembly, but during recent years its association with muscle development, cell reprogramming, and the angiogenic program have demonstrated its activities well beyond extracellular matrix maintenance. In fact, the involvement of FMOD in these cellular processes places it the centrum of cellular behaviour and ultimately of tissue properties. Thus, a clear view of the impact FMOD has on tissue integrity would aid its exploitation for tissue modelling and in the treatment of different disorders.

blaCTX-M-152, a Novel Variant of CTX-M-group-25, Identified in a Study Performed on the Prevalence of Multidrug Resistance among Natural Inhabitants of River Yamuna, India

Natural environment influenced by anthropogenic activities creates selective pressure for acquisition and spread of resistance genes. In this study, we determined the prevalence of Extended Spectrum β-Lactamases producing gram negative bacteria from the River Yamuna, India, and report the identification and characterization of a novel CTX-M gene variant blaCTX-M-152. Of the total 230 non-duplicate isolates obtained from collected water samples, 40 isolates were found positive for ESBL production through Inhibitor-Potentiation Disc Diffusion test. Based on their resistance profile, 3% were found exhibiting pandrug resistance (PDR), 47% extensively drug resistance (XDR), and remaining 50% showing multidrug resistant (MDR). Following screening and antimicrobial profiling, characterization of ESBLs (blaTEM and blaCTX-M), and mercury tolerance determinants (merP, merT, and merB) were performed. In addition to abundance of blaTEM-116 (57.5%) and blaCTX-M-15 (37.5%), bacteria were also found to harbor other variants of ESBLs like blaCTX-M-71 (5%), blaCTX-M-3 (7.5%), blaCTX-M-32 (2.5%), blaCTX-M-152 (7.5%), blaCTX-M-55 (2.5%), along with some non-ESBLs; blaTEM-1 (25%) and blaOXY (5%). Additionally, co-occurrence of mercury tolerance genes were observed among 40% of isolates. In silico studies of the new variant, blaCTX-M-152were conducted through modeling for the generation of structure followed by docking to determine its catalytic profile. CTX-M-152 was found to be an out-member of CTX-M-group-25 due to Q26H, T154A, G89D, P99S, and D146G substitutions. Five residues Ser70, Asn132, Ser237, Gly238, and Arg273 were found responsible for positioning of cefotaxime into the active site through seven H-bonds with binding energy of -7.6 Kcal/mol. Despite small active site, co-operative interactions of Ser237 and Arg276 were found actively contributing to its high catalytic efficiency. To the best of our knowledge, this is the first report of blaCTX-M-152 of CTX-M-group-25 from Indian subcontinent. Taking a note of bacteria harboring such high proportion of multidrug and mercury resistance determinants, their presence in natural water resources employed for human consumption increases the chances of potential risk to human health. Hence, deeper insights into mechanisms pertaining to resistance development are required to frame out strategies to tackle the situation and prevent acquisition and dissemination of resistance determinants so as to combat the escalating burden of infectious diseases.

Fibromodulin: a master regulator of myostatin controlling progression of satellite cells through a myogenic program

Differentiation of muscle satellite cells (MSCs) involves interaction of the proteins present in the extracellular matrix (ECM) with MSCs to regulate their activity, and therefore phenotype. Herein, we report fibromodulin (FMOD), a member of the proteoglycan family participating in the assembly of ECM, as a novel regulator of myostatin (MSTN) during myoblast differentiation. In addition to having a pronounced effect on the expression of myogenic marker genes [myogenin (MYOG) and myosin light chain 2 (MYL2)], FMOD was found to maintain the transcriptional activity of MSTN. Moreover, coimmunoprecipitation and in silico studies performed to investigate the interaction of FMOD helped confirm that it antagonizes MSTN function by distorting its folding and preventing its binding to activin receptor type IIB. Furthermore, in vivo studies revealed that FMOD plays an active role in healing by increasing satellite cell recruitment to sites of injury. Together, these findings disclose a hitherto unrecognized regulatory role for FMOD in MSCs and highlight new mechanisms whereby FMOD circumvents the inhibitory effects of MSTN and triggers myoblast differentiation. These findings offer a basis for the design of novel MSTN inhibitors that promote muscle regeneration after injury or for the development of pharmaceutical agents for the treatment of different muscle atrophies.—Lee, E. J., Jan, A. T., Baig M. H., Ashraf, J. M., Nahm, S.‐S., Kim, Y.‐W., Park, S.‐Y., Choi, I. Fibromodulin: a master regulator of myostatin controlling progression of satellite cells through a myogenic program. FASEB J. 30, 2708‐2719 (2016). www.fasebj.org