Alteration of the risk of pre-oral cancer and cancer in North Indian population by NAT1 and NAT2 polymorphisms genotypes and haplotypes.

PURPOSE: The risk of oral cancer is strongly related to consumption of tobacco, smoking and drinking alcohol. N-acetyl transferases 1,2 are phase II metabolic enzymes, metabolize aryl and heterocyclic amines which are present in tobacco. NAT2 slows acetylator phenotype and the genotype is related to reduced ability to detoxify these xenobiotic that are carcinogenic to tissues. The aim of our study to determine the risk of oral cancer as well as oral precancerous lesions in North Indian population with polymorphisms in these two N-acetyl transferases 1,2 genes. MATERIALS AND METHODS: A total of 250 patients with pre oral cancer, oral cancer and 250 healthy volunteers were genotypes for the NAT1 and NAT2 gene polymorphisms. Genotypes were identified by PCR and RFLP. Genotype frequencies were evaluated by Chi-square test and risk of disease was estimated by Odds ratio (OR) with 95% confidence interval. RESULT: Our results showed that individuals with CT and TT genotypes of NAT1 C > T polymorphism were significantly lower risk of oral diseases (p value = 0.02, OR = 0.60 and p value = 0.04, OR = 0.58, respectively). For NAT2 C > T polymorphism, the TT genotype significantly increased the risk of OSMF (Oral Sub mucous Fibrosis) and Leukoplakia (p value = 0.001, OR = 4.16; p value = 0.002, OR = 4.38, respectively). In contrary, the CC genotype for NAT2 T > C polymorphism increased the risk of OSMF (p value = 0.01, OR = 3.00, 95% CI = 1.31-6.86). CONCLUSION: Our study concludes that the NAT1 polymorphism shows protective association with oral diseases and NAT2 polymorphism and haplotypes also influence the susceptibility to oral diseases in North Indian population subjects.

Coronavirus disease: Dental review.

COVID-19 was declared a pandemic outbreak by the World Health Organization, creating a significant impact on health care system. Realizing the high risk associated with this disease and its high rate of transmission, dentists were instructed by health authorities, to stop providing treatment which includes aerosols and droplets and only except emergency complaints. This was mainly for protection of dental healthcare personnel, their families, contacts, and their patients from the transmission of virus. Hence, this review focuses on the life cycle of COVID-19, its clinical symptoms and several issues concerned directly to dental practice in terms of prevention, treatment, and orofacial clinical manifestations.

Lichen planus drugs re-purposing as potential anti COVID-19 therapeutics through molecular docking and molecular dynamics simulation approach.

Background and Aim: The present study intends to investigate COVID-19 by targeting their main proteins with 17 selected drugs used for treating Oral Lichen Planus (OLP) which is a chronic muco-cutaneous disorder. Here, an attempt is made to gain better insight into the structure of various drugs targeting specific proteins which will be helpful in developing drugs useful for therapeutic and preventive measures. Method: In silico studies, molecular docking and molecular dynamic simulations were performed to repurpose the therapeutic drugs (n = 17) which were used to treat OLP against COVID-19. In addition, the maximum binding affinities of the key protein spike glycoprotein, main-protease (Mpro) of coronavirus, and Angiotensin-Converting Enzyme-2 (ACE-2) in the human body were evaluated with the selected drugs. Results: Epigallocatechin-3-gallate (EGCG) showed the highest docking values among the drugs selected for repurposing. Among the target proteins, EGCG has shown maximum binding affinity with ACE-2 receptor. Further, according to the molecular dynamic simulation studies, EGCG has shown the least conformational fluctuations with Mpro. Conclusion: EGCG can be a potential inhibitor drug which can bind with ACE-2 receptor thus inhibiting the interaction of mainly Mpro protein and spike glycoprotein of SARS-CoV-2. Relevance for Patients: EGCG, a natural compound shows antiviral potential having considerably high affinity and stability with SARS-CoV-2. It might be further employed as a lead drug against selective inhibitors of SARS-CoV-2 for the therapeutic management of COVID-19 patients after necessary clinical trials.

Lichen planus drugs re-purposing as potential anti COVID-19 therapeutics through molecular docking and molecular dynamics simulation approach.

Background and Aim: The present study intends to investigate COVID-19 by targeting their main proteins with 17 selected drugs used for treating Oral Lichen Planus (OLP) which is a chronic muco-cutaneous disorder. Here, an attempt is made to gain better insight into the structure of various drugs targeting specific proteins which will be helpful in developing drugs useful for therapeutic and preventive measures. Method: In silico studies, molecular docking and molecular dynamic simulations were performed to repurpose the therapeutic drugs (n = 17) which were used to treat OLP against COVID-19. In addition, the maximum binding affinities of the key protein spike glycoprotein, main-protease (Mpro) of coronavirus, and Angiotensin-Converting Enzyme-2 (ACE-2) in the human body were evaluated with the selected drugs. Results: Epigallocatechin-3-gallate (EGCG) showed the highest docking values among the drugs selected for repurposing. Among the target proteins, EGCG has shown maximum binding affinity with ACE-2 receptor. Further, according to the molecular dynamic simulation studies, EGCG has shown the least conformational fluctuations with Mpro. Conclusion: EGCG can be a potential inhibitor drug which can bind with ACE-2 receptor thus inhibiting the interaction of mainly Mpro protein and spike glycoprotein of SARS-CoV-2. Relevance for Patients: EGCG, a natural compound shows antiviral potential having considerably high affinity and stability with SARS-CoV-2. It might be further employed as a lead drug against selective inhibitors of SARS-CoV-2 for the therapeutic management of COVID-19 patients after necessary clinical trials.

Role of molecular techniques in PMI estimation: An update.

The time frames between death and reporting of the cadaver, known as post Mortem interval (PMI), is essential in investigation of homicide deaths, suspicious deaths, or other untimely deaths as well as natural deaths. Such information helps to connect the missing links in homicide or other relevant cases. Over the time several methods are developed which depends upon factors as several methods physiological, biochemical, entomological, and archaeological for the estimation of degradation of body with time. These methods lack precision, require expertise to achieve worthy results or authentic estimate. Although these methods are currently in use but, these evaluations are still unreliable and imprecise. Hence, we still need new methods for better estimation of PMI. Initially, the predictable morphological and chemical changes in cadaver are used as PMI indicators but, as the time since death increases, the above methods become less useful for as they can't pin point the time of death rather give a ballpark idea. With the advent of the field of molecular biology, the estimation of PMI is proposed to be executed by evaluating the degradation pattern of the biological markers (DNA, RNA, and Proteins). It is now proved that the DNA is fairly unwavering over long post-mortem phases, RNA is much more labile in nature, and sensitive to degradation in a tissue-specific manner. Thus, the main purpose (aim, agenda) of this document is to provide review that mainly focuses on potential use of RNA markers in estimation of PMI. For this Critical Review, the systematic evaluation of 47 studies is executed according to the chosen inclusion and exclusion criteria.

Immunotherapy: Newer Therapeutic Armamentarium against Cancer Stem Cells.

Mounting evidence from the literature suggests the existence of a subpopulation of cancer stem cells (CSCs) in almost all types of human cancers. These CSCs possessing a self-renewal capacity inhabit primary tumors and are more defiant to standard antimitotic and molecularly targeted therapies which are used for eliminating actively proliferating and differentiated cancer cells. Clinical relevance of CSCs emerges from the fact that they are the root cause of therapy resistance, relapse, and metastasis. Earlier, surgery, chemotherapy, and radiotherapy were established as cancer treatment modalities, but recently, immunotherapy is also gaining importance in the management of various cancer patients, mostly those of the advanced stage. This review abridges potential off-target effects of inhibiting CSC self-renewal pathways on immune cells and some recent immunological studies specifically targeting CSCs on the basis of their antigen expression profile, even though molecular markers or antigens that have been described till date as expressed by cancer stem cells are not specifically expressed by these cells which is a major limitation to target CSCs. We propose that owing to CSC stemness property to mediate immunotherapy response, we can apply a combination therapy approach by targeting CSCs and tumor microenvironment (TME) along with conventional treatment strategies as an effective means to eradicate cancer cells.

Isolation and characterization of mesenchymal stem cells from human fetus heart.

BACKGROUND: Mesenchymal stem cells (MSCs) are promising cells for cardiovascular regenerative medicine. However, their potential may be limited, because of their restricted cardiovascular differentiation potential and decline in their number and functional characteristics with increasing donor age. We have previously shown that rat fetus heart harbors primitive MSCs and administration of these cells improved left ventricular (LV) function after ischemia/reperfusion injury in rats. To evaluate their potential as a new cell type for clinical cardiovascular cell therapy, we have undertaken this study on the isolation and characterization of human fetal cardiac MSCs (hfC-MSCs). METHODS: MSCs were isolated from the heart of five 14-16-week-old aborted human fetuses and studied for their growth characteristics, karyotypic stability and senescence over successive passages, expression of mesenchymal and embryonal markers by flow cytometry and immunocytochemistry, constitutive expression of cardiovascular genes by RT-PCR, differentiation into cells of the cardiovascular lineage and their immunomodulatory properties. RESULTS: The hfC-MSCs grew as adherent monolayer with spindle shaped morphology and exhibited rapid proliferation with an average population doubling time of 34 hours and expansion to up to more than 80 population doublings with maintenance of a normal karyotype and without senescence. Immunophenotyping showed that they had similar phenotype as human bone marrow mesenchymal stem cells (hBM-MSCs) expressing CD73, CD90, CD105 and lacking expression of CD31, CD34, CD45, HLA-DR. However, hfC-MSCs expressed significantly higher levels of CD117 and SSEA-4 compared to hBM-MSCs. In addition, hfC-MSCs expressed the embryonal markers Oct-4, Nanog and Sox-2 as compared to hBM-MSCs. Further, hfC-MSCs had significantly higher expression of the cardiovascular genes viz. ISL-1, flk-1, GATA-4, NKX2.5 and MDR-1 as compared to hBM-MSCs, and could be differentiated into major cardiovascular cells (cardiomyocytes, endothelial cells, smooth muscle cells). Interestingly, hfC-MSCs markedly reduced T-lymphocyte proliferation with an increased secretion of TGF-ß and IL-10. CONCLUSIONS: Our results show that human fetus heart is a novel source of primitive MSCs with cardiovascular commitment which may have a potential therapeutic application in cardiovascular regenerative medicine.

Cardiomyogenic Heterogeneity of Clonal Subpopulations of Human Bone Marrow Mesenchymal Stem Cells.

We have evaluated the cardiomyogenic potential of clonal populations of human bone marrow mesenchymal stem cells (BM-MSC). Four rapidly proliferating clones of BM-MSC were obtained from the BM of a healthy donor which were then treated with 5-azacytidine and evaluated for the expression of GATA-4, NKx-2.5, FOG-2, TDGF-1, ß-MHC, MEF2D and NPPA genes and cTnT, Desmin and ß-MHC proteins. Of the four clones (i) Clone-1 had high expression of GATA-4 (1.89 fold (p<0.05), Nkx2.5 (2.29 fold; p<0.05), FOG2 (2.76 fold; p<0.05), TDGF1 (6.97 fold, p<0.005), ßMHC (10.22 fold; p<0.005), MEF-2D (1.91 fold; p<0.005) and NPPA (1.65 fold; p<0.005); (ii) clone-2 had up-regulation of Nkx2.5 (1.98 fold; p<0.05) but down-regulation of rest of the genes; (iii) clone-3 had up-regulation of Nkx2.5 (2.11 fold; p<0.05), TDGF1 (1.88 fold; p<0.05), MEF-2D (1.30 fold; p<0.05) and NPPA (1.21 fold; p<0.05), down regulation of GATA-4 and Fog-2 but no change in ßMHC gene; and (iv) clone-4 had up-regulation of MEF-2D (1.17 fold; p<0.05) and down regulation of GATA-4, Nkx2.5 but no change in other genes compared to untreated cells of the clones. At the protein level, clone-1 expressed cTnT, Desmin, and ßMHC; clone-2 Desmin only while clones-3 and 4 each expressed cTnT, Desmin, and ßMHC. Our data shows that BM-MSC are a heterogenous population of stem cells with sub-populations exhibiting a marked difference in the expression of cardiac markers both at gene and protein levels. This highlights that administering selected sub-populations of BM-MSC with a cardiomyogenic potential may be more efficacious than whole population of cells for cardiac regeneration.

Therapeutic efficacy of multipotent adult progenitor cells versus mesenchymal stem cells in experimental autoimmune encephalomyelitis.

AIM: In this study, we have evaluated the therapeutic efficacy of mouse multipotent adult progenitor cells (mMAPCs) in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, and compared it with mouse mesenchymal stem cells (mMSCs). MATERIALS & METHODS: We administered PKH26-labeled mMAPC and mMSC into EAE mice and evaluated their therapeutic efficacy. RESULTS: The mMAPC-treated mice in comparison with the mMSC group exhibited a higher suppression of EAE (p < 0.05), and a higher fold expression of neuronal genes GAP43, NG2, PDGFR, Nestin, SMI 32, BDNF and NT 3 in spinal cord (p < 0.05), suggesting a better neuroprotective and regenerative potential of mMAPC than mMSC. CONCLUSION: MAPC may be a potential cell type, which is superior to mesenchymal stem cell for the treatment of EAE/multiple sclerosis.

Expression Levels of Tetraspanin KAI1/CD82 in Breast Cancers in North Indian Females.

BACKGROUND: Carcinogenesis is a multifaceted intricate cellular mechanism of transformation of the normal functions of a cell into neoplastic alterations. Metastasis may result in failure of conventional treatment and death Hence, research on metastatic suppressors in cancer is a high priority. The metastatic suppressor gene CD82, also known as KAI1, is a member of the transmembrane 4 superfamily which was first identified in carcinoma of prostate. Little work has been done on this gene in breast cancer. Herein, we aimed to determine the gene and protein level expression of CD82/KAI1 in breast cancer and its role as a prognosticator. MATERIALS AND METHODS: In this study, 83 histologically proven cases of breast cancer and a similar number of controls were included. Patient age ranged from 1870 years. Quantitative Real Time Polymerase Chain Reaction (qRT PCR) and immunohistochemistry (IHC) were used to investigate KAI1 expression at gene and protein levels, respectively. Statistical analysis was done to correlate expression of KAI1 and clinicopathological parameters. RESULTS: It was revealed that: (i) KAI1 was remarkably diminished in metastatic vs non metastatic breast cancer both at the gene and the protein levels (P<.05); (ii) KAI1 expression levels were strongly correlated with TNM staging, histological grade and advanced stage (p<0.001) and no association was found with any other studied parameter; (iii) Lastly, a significant correlation was observed between expression of KAI1 and overall median survival of BC patients (P = 0.04). CONCLUSIONS: Our results suggest that lack of expression of the KAI1 might indicate a more aggressive form of breast cancer. Loss of KAI1 may be considered a significant prognostic marker in predicting metastatic manifestation. When evaluated along with the clinical and pathological factors, KAI1 expression may be beneficial to tailor aggressive therapeutic strategies for such patients.

Evaluation of KiSS1 as a Prognostic Biomarker in North Indian Breast Cancer Cases.

BACKGROUND: Breast cancer is the commonest female cancer worldwide and its propensity to metastasize negatively impacts on therapeutic outcome. Several clinicopathological parameters with prognostic/predictive significance have been associated with metastatic suppressor expression levels. The role of metastatic suppressor gene (MSG) KiSS1 in breast cancer remains unclear. Our goal was to investigate the possible clinical significance of KiSS1 breast cancer. MATERIALS AND METHODS: The study was conducted on 87 histologically proven cases of breast cancer and background normal tiisue. Quantitative reverse transcriptase polymerase chain reaction (qRT PCR) and immunohistochemistry (IHC) were used to investigate KiSS1 at gene and protein levels, respectively, for correlation with several patient characteristics including age, family history, hormonal receptor status, stage, tumor size, nodal involvement and metastatic manifestation and finally with median overall survival (OS). RESULTS: Our study revealed (i) KiSS1 levels were generally elevated in breast cancer vs normal tissue (< 0.05). (ii) however, a statistically significant lower expression of KiSS1 was observed in metastatic vs non metastatic cases (P = 0.04). (iii) KiSS1 levels strongly correlated with T,N,M category, histological grade and advanced stage (<0.001) but not other studied parameters. (iv) Lastly, a significant correlation between expression of KiSS1 and median OS was found (P = 0.04). CONCLUSIONS: Conclusively, less elevated KiSS1 expression is a negative prognostic factor for OS, advancing tumor stage, axillary lymph node status, metastatic propensity and advancing grade of the breast cancer patient. Patients with negative KiSS1 expression may require a more intensive therapeutic strategy.

Enhanced adipogenicity of bone marrow mesenchymal stem cells in aplastic anemia.

Fatty bone marrow (BM) and defective hematopoiesis are a pathologic hallmark of aplastic anemia (AA). We have investigated adipogenic and osteogenic potential of BM mesenchymal stem cells (BM-MSC) in 10 AA patients (08 males and 02 females) with median age of 37 years (range: 06 to 79 years) and in the same number of age and sex matched controls. It was observed that BM-MSC of AA patients had a morphology, phenotype, and osteogenic differentiation potential similar to control subjects but adipocytes differentiated from AA BM-MSC had a higher density and larger size of lipid droplets and they expressed significantly higher levels of adiponectin and FABP4 genes and proteins as compared to control BM-MSC (P < 0.01 for both). Thus our data shows that AA BM-MSC have enhanced adipogenicity, which may have an important implication in the pathogenesis of the disease.

Characterization of a Novel Putative S-Adenosylmethionine Decarboxylase-Like Protein from Leishmania donovani.

In addition to the S-adenosylmethionine decarboxylase (AD) present in all organisms, trypanosomatids including Leishmania spp. possess an additional copy, annotated as the putative S-adenosylmethionine decarboxylase-like proenzyme (ADL). Phylogenetic analysis confirms that ADL is unique to trypanosomatids and has several unique features such as lack of autocatalytic cleavage and a distinct evolutionary lineage, even from trypanosomatid ADs. In Trypanosoma ADL was found to be enzymaticaly dead but plays an essential regulatory role by forming a heterodimer complex with AD. However, no structural or functional information is available about ADL from Leishmania spp. Here, in this study, we report the cloning, expression, purification, structural and functional characterization of Leishmania donovani (L. donovani) ADL using biophysical, biochemical and computational techniques. Biophysical studies show that, L. donovani ADL binds S-adenosylmethionine (SAM) and putrescine which are natural substrates of AD. Computational modeling and docking studies showed that in comparison to the ADs of other organisms including human, residues involved in putrescine binding are partially conserved while the SAM binding residues are significantly different. In silico protein-protein interaction study reveals that L. donovani ADL can interact with AD. These results indicate that L. donovani ADL posses a novel substrate binding property and may play an essential role in polyamine biosynthesis with a different mode of function from known proteins of the S-adenosylmethionine decarboxylase super family.

Comparison of phenotypic markers and neural differentiation potential of multipotent adult progenitor cells and mesenchymal stem cells.

AIM: To compare the phenotypic and neural differentiation potential of human bone marrow derived multipotent adult progenitor cells (MAPC) and mesenchymal stem cells (MSC). METHODS: Cultures of MAPC and MSC were established in parallel from same samples of human bone marrow (n = 5). Both stem cell types were evaluated for expression of pluripotency markers including Oct-4 and Nanog by immunocytochemistry and reverse-transcription polymerase chain reaction (RT-PCR) and expression of standard mesenchymal markers including CD14, CD34, CD44, CD45, CD73, CD90, CD105 and human leukocyte antigen (HLA)-ABC by flow cytometry. After treatment with neural induction medium both MAPC and MSC were evaluated for expression of neural proteins [neuronal filament-200 (NF-200) and glial fibrillar acidic protein (GFAP)] by immunocytochemistry and Western blotting and neural genes [NF-200, GFAP, Tau, microtubule-associated protein (MAP)-1B, MAP-2, neuron-specific enolase (NSE) and oligodendrocyte-1 (Olig-1)] by quantitative real-time-PCR. RESULTS: MAPC had small trigonal shaped while MSC had elongated spindle-shaped morphology. The MAPC expressed Oct-4 and Nanog both at gene and protein levels, whereas MSC were negative for these pluripotent markers. MAPC were negative for HLA-ABC while MSC had high expression of HLA-ABC. In addition, MAPC as compared to MSC had significantly lower expression of CD44 (36.56% ± 1.92% vs 98.23% ± 0.51%), CD73 (15.11% ± 2.24% vs 98.53% ± 2.22%) and CD105 (13.81% ± 3.82% vs 95.12% ± 5.65%) (P < 0.001, for all) MAPC cultures compared to MSC cultures treated with neural induction medium had significantly higher fold change expression of NF-200 (0.64), GFAP (0.52), Tau (0.59), MAP-2 (0.72), Olig-1 (0.18) and NSE (0.29) proteins (P < 0.01 for Olig-1 and P < 0.001 for rest) as well as higher fold change expression of genes of NF-200 (1.34), GFAP (1.12), Tau (1.08), MAP-1B (0.92), MAP-2 (1.14) and NSE (0.4) (P < 0.001 for all). CONCLUSION: MAPC can be differentially characterized from MSC as Oct-4 and Nanog positive stem cells with no expression of HLA-ABC and low expression of mesenchymal markers CD44, CD73 and CD105 and when compared to MSC they possess greater predilection for differentiation into neuro-ectodermal lineage.