Deciphering the mechanism of Tinospora cordifolia extract on Th17 cells through in-depth transcriptomic profiling and in silico analysis.

Naive CD4+ T cells differentiate into effector (Th1, Th2, Th17) cells and immunosuppressive (Treg) cells upon antigenic stimulation in the presence of a specific cytokine milieu. The T cell in vitro culture system provides a very efficient model to study compounds' therapeutic activity and mechanism of action. Tinospora cordifolia (Willd.) Hook.f. & Thomson (Family. Menispermaceae) is one of the widely used drugs in Ayurveda (ancient Indian system of medicine) for various ailments such as inflammatory conditions, autoimmune disorders, and cancer as well as for promoting general health. In vitro and in vivo studies on immune cells comprising dendritic cells, macrophages, and B cells suggest its immune-modulating abilities. However, to date, the effect of T. cordifolia on individual purified and polarized T cell subsets has not been studied. Studying drug effects on T cell subsets is needed to understand their immunomodulatory mechanism and to develop treatments for diseases linked with T cell abnormalities. In this study, we examined the immunomodulatory activity of T. cordifolia on primary CD4+ T cells, i.e., Th1, Th17, and iTreg cells. An aqueous extract of T. cordifolia was non-cytotoxic at concentrations below 1500 µg/ml and moderately inhibited the proliferation of naive CD4+ T cells stimulated with anti-CD3ε and anti-CD28 for 96 h. T. cordifolia treatment of naive CD4+ T cells differentiated under Th17-polarizing conditions exhibited reduced frequency of IL-17 producing cells with inhibition of differentiation and proliferation. For the first time, in-depth genome-wide expression profiling of T. cordifolia treated naive CD4+ T cells, polarized to Th17 cells, suggests the broad-spectrum activity of T. cordifolia. It shows inhibition of the cytokine-receptor signaling pathway, majorly via the JAK-STAT signaling pathway, subsequently causing inhibition of Th17 cell differentiation, proliferation, and effector function. Additionally, the molecular docking studies of the 69 metabolites of T. cordifolia further substantiate the inhibitory activity of T. cordifolia via the cytokine-receptor signaling pathway. Furthermore, in vitro polarized Th1 and iTreg cells treated with T. cordifolia extract also showed reduced IFN-γ production and FoxP3 expression, respectively. This study provides insight into the plausible mechanism/s of anti-inflammatory activity of T. cordifolia involving T cells, mainly effective in Th17-associated autoimmune and inflammatory diseases.

Events of alternative splicing in head and neck cancer via RNA sequencing - an update.

BACKGROUND: Alternative splicing (AS) is a regulatory mechanism used to create many forms of mature messengers RNAs (mRNAs) from the same gene. Sequencing of RNA (RNA-Seq) is an advanced technology, which has been utilized by different studies to find AS mechanisms in head and neck cancer (HNC). Hitherto, there is no available review that could inform us of the major findings from these studies. Hence, we aim to perform a systematic literature search following PRISMA guidelines to study AS events in HNC identified through RNA-Seq studies. RESULTS: A total of five records were identified that utilized RNA-Seq data for identifying AS events in HNC. Five software was used in these records to identify AS events. Two genes influenced by AS i.e. MLL3 and RPS9 were found to be common in 4 out of 5 records. Likewise, 38 genes were identified to be similar in at least 3 records. CONCLUSIONS: Alternative splicing in HNC is a multifaceted regulatory mechanism of gene expression. It can be studied via RNA-Seq using different bioinformatics tools. Genes MLL3, as well as RPS9, were repeatedly found to be associated with HNC, however needs further functional validation.

Dohra- a mixture of potent carcinogens.

BACKGROUND & OBJECTIVES: Dohra is a areca nut preparation used with or without tobacco in a few of the areas of Uttar Pradesh (UP), India. There is evidence that it causes potentially malignant disorders and oral cancer. This study was undertaken to provide information on dohra by searching through literature and also through a survey in three areas of Uttar Pradesh (UP), India. METHODS: The information on dohra was collected through literature search, study tour to different areas of UP, where group discussions with dohra vendors and with community members of different age group were done to obtain information. RESULTS: Dohra was prepared by the users for their personal use or prepared by small-scale industry for sale. It was available mostly in betel shops or any other store/kiosks and was also available in special dohra shops. Dohra was available in both dry and wet form. Its common constituents were areca nut, catechu (Acacia catechu), edible lime, peppermint (Mentha piperita), cardamom (Elettaria cardamomum) and some flavoring agents. Dohra was consumed as such or with tobacco. INTERPRETATION & CONCLUSIONS: Different varieties of Dohra were available such as sukha dohra, sukha mulethi dohra and geela dohra. Different processing methods for producing dohra existed. As dohra increases the risk of cancer, it needs to be banned or it should be sold in packets with the details of its constituents and also statutory warning about its adverse health effects.

Reconstructing the demographic history of the Himalayan and adjoining populations.

The rugged topography of the Himalayan region has hindered large-scale human migrations, population admixture and assimilation. Such complexity in geographical structure might have facilitated the existence of several small isolated communities in this region. We have genotyped about 850,000 autosomal markers among 35 individuals belonging to the four major populations inhabiting the Himalaya and adjoining regions. In addition, we have genotyped 794 individuals belonging to 16 ethnic groups from the same region, for uniparental (mitochondrial and Y chromosomal DNA) markers. Our results in the light of various statistical analyses suggest a closer link of the Himalayan and adjoining populations to East Asia than their immediate geographical neighbours in South Asia. Allele frequency-based analyses likely support the existence of a specific ancestry component in the Himalayan and adjoining populations. The admixture time estimate suggests a recent westward migration of populations living to the East of the Himalaya. Furthermore, the uniparental marker analysis among the Himalayan and adjoining populations reveal the presence of East, Southeast and South Asian genetic signatures. Interestingly, we observed an antagonistic association of Y chromosomal haplogroups O3 and D clines with the longitudinal distance. Thus, we summarise that studying the Himalayan and adjoining populations is essential for a comprehensive reconstruction of the human evolutionary and ethnolinguistic history of eastern Eurasia.

Signature of genetic associations in oral cancer.

Oral cancer etiology is complex and controlled by multi-factorial events including genetic events. Candidate gene studies, genome-wide association studies, and next-generation sequencing identified various chromosomal loci to be associated with oral cancer. There is no available review that could give us the comprehensive picture of genetic loci identified to be associated with oral cancer by candidate gene studies-based, genome-wide association studies-based, and next-generation sequencing-based approaches. A systematic literature search was performed in the PubMed database to identify the loci associated with oral cancer by exclusive candidate gene studies-based, genome-wide association studies-based, and next-generation sequencing-based study approaches. The information of loci associated with oral cancer is made online through the resource "ORNATE." Next, screening of the loci validated by candidate gene studies and next-generation sequencing approach or by two independent studies within candidate gene studies or next-generation sequencing approaches were performed. A total of 264 loci were identified to be associated with oral cancer by candidate gene studies, genome-wide association studies, and next-generation sequencing approaches. In total, 28 loci, that is, 14q32.33 (AKT1), 5q22.2 (APC), 11q22.3 (ATM), 2q33.1 (CASP8), 11q13.3 (CCND1), 16q22.1 (CDH1), 9p21.3 (CDKN2A), 1q31.1 (COX-2), 7p11.2 (EGFR), 22q13.2 (EP300), 4q35.2 (FAT1), 4q31.3 (FBXW7), 4p16.3 (FGFR3), 1p13.3 (GSTM1-GSTT1), 11q13.2 (GSTP1), 11p15.5 (H-RAS), 3p25.3 (hOGG1), 1q32.1 (IL-10), 4q13.3 (IL-8), 12p12.1 (KRAS), 12q15 (MDM2), 12q13.12 (MLL2), 9q34.3 (NOTCH1), 17p13.1 (p53), 3q26.32 (PIK3CA), 10q23.31 (PTEN), 13q14.2 (RB1), and 5q14.2 (XRCC4), were validated to be associated with oral cancer. "ORNATE" gives a snapshot of genetic loci associated with oral cancer. All 28 loci were validated to be linked to oral cancer for which further fine-mapping followed by gene-by-gene and gene-environment interaction studies is needed to confirm their involvement in modifying oral cancer.

De novo fatty acid synthesis controls the fate between regulatory T and T helper 17 cells.

Interleukin-17 (IL-17)-secreting T cells of the T helper 17 (TH17) lineage play a pathogenic role in multiple inflammatory and autoimmune conditions and thus represent a highly attractive target for therapeutic intervention. We report that inhibition of acetyl-CoA carboxylase 1 (ACC1) restrains the formation of human and mouse TH17 cells and promotes the development of anti-inflammatory Foxp3(+) regulatory T (Treg) cells. We show that TH17 cells, but not Treg cells, depend on ACC1-mediated de novo fatty acid synthesis and the underlying glycolytic-lipogenic metabolic pathway for their development. Although TH17 cells use this pathway to produce phospholipids for cellular membranes, Treg cells readily take up exogenous fatty acids for this purpose. Notably, pharmacologic inhibition or T cell-specific deletion of ACC1 not only blocks de novo fatty acid synthesis but also interferes with the metabolic flux of glucose-derived carbon via glycolysis and the tricarboxylic acid cycle. In vivo, treatment with the ACC-specific inhibitor soraphen A or T cell-specific deletion of ACC1 in mice attenuates TH17 cell-mediated autoimmune disease. Our results indicate fundamental differences between TH17 cells and Treg cells regarding their dependency on ACC1-mediated de novo fatty acid synthesis, which might be exploited as a new strategy for metabolic immune modulation of TH17 cell-mediated inflammatory diseases.

Selective and efficient generation of functional Batf3-dependent CD103+ dendritic cells from mouse bone marrow.

Multiple subsets of FMS-like tyrosine kinase 3 ligand (FLT3L)-dependent dendritic cells (DCs) control T-cell tolerance and immunity. In mice, Batf3-dependent CD103(+) DCs efficiently enter lymph nodes and cross-present antigens, rendering this conserved DC subset a promising target for tolerance induction or vaccination. However, only limited numbers of CD103(+) DCs can be isolated with current methods. Established bone marrow culture protocols efficiently generate monocyte-derived DCs or produce a mixture of FLT3L-dependent DC subsets. We show that CD103(+) DC development requires prolonged culture time and continuous action of both FLT3L and granulocyte macrophage colony-stimulating factor (GM-CSF), explained by a dual effect of GM-CSF on DC precursors and differentiating CD103(+) DCs. Accordingly, we established a novel method to generate large numbers of CD103(+) DCs (iCD103-DCs) with limited presence of other DC subsets. iCD103-DCs develop in a Batf3- and Irf8-dependent fashion, express a CD8α/CD103 DC gene signature, cross-present cell-associated antigens, and respond to TLR3 stimulation. Thus, iCD103-DCs reflect key features of tissue CD103(+) DCs. Importantly, iCD103-DCs express high levels of CCR7 upon maturation and migrate to lymph nodes more efficiently than classical monocyte-derived DCs. Finally, iCD103-DCs induce T cell-mediated protective immunity in vivo. Our study provides insights into CD103(+) DC development and function.

Unravelling the distinct strains of Tharu ancestry.

The northern region of the Indian subcontinent is a vast landscape interlaced by diverse ecologies, for example, the Gangetic Plain and the Himalayas. A great number of ethnic groups are found there, displaying a multitude of languages and cultures. The Tharu is one of the largest and most linguistically diverse of such groups, scattered across the Tarai region of Nepal and bordering Indian states. Their origins are uncertain. Hypotheses have been advanced postulating shared ancestry with Austroasiatic, or Tibeto-Burman-speaking populations as well as aboriginal roots in the Tarai. Several Tharu groups speak a variety of Indo-Aryan languages, but have traditionally been described by ethnographers as representing East Asian phenotype. Their ancestry and intra-population diversity has previously been tested only for haploid (mitochondrial DNA and Y-chromosome) markers in a small portion of the population. This study presents the first systematic genetic survey of the Tharu from both Nepal and two Indian states of Uttarakhand and Uttar Pradesh, using genome-wide SNPs and haploid markers. We show that the Tharu have dual genetic ancestry as up to one-half of their gene pool is of East Asian origin. Within the South Asian proportion of the Tharu genetic ancestry, we see vestiges of their common origin in the north of the South Asian Subcontinent manifested by mitochondrial DNA haplogroup M43.

Phylogenetic analysis, based on EPIYA repeats in the cagA gene of Indian Helicobacter pylori, and the implications of sequence variation in tyrosine phosphorylation motifs on determining the clinical outcome.

The population of India harbors one of the world's most highly diverse gene pools, owing to the influx of successive waves of immigrants over regular periods in time. Several phylogenetic studies involving mitochondrial DNA and Y chromosomal variation have demonstrated Europeans to have been the first settlers in India. Nevertheless, certain controversy exists, due to the support given to the thesis that colonization was by the Austro-Asiatic group, prior to the Europeans. Thus, the aim was to investigate pre-historic colonization of India by anatomically modern humans, using conserved stretches of five amino acid (EPIYA) sequences in the cagA gene of Helicobacter pylori. Simultaneously, the existence of a pathogenic relationship of tyrosine phosphorylation motifs (TPMs), in 32 H. pylori strains isolated from subjects with several forms of gastric diseases, was also explored. High resolution sequence analysis of the above described genes was performed. The nucleotide sequences obtained were translated into amino acids using MEGA (version 4.0) software for EPIYA. An MJ-Network was constructed for obtaining TPM haplotypes by using NETWORK (version 4.5) software. The findings of the study suggest that Indian H. pylori strains share a common ancestry with Europeans. No specific association of haplotypes with the outcome of disease was revealed through additional network analysis of TPMs.

Phylogenetic analysis, based on EPIYA repeats in the cagA gene of Indian Helicobacter pylori, and the implications of sequence variation in tyrosine phosphorylation motifs on determining the clinical outcome

The population of India harbors one of the world's most highly diverse gene pools, owing to the influx of successive waves of immigrants over regular periods in time. Several phylogenetic studies involving mitochondrial DNA and Y chromosomal variation have demonstrated Europeans to have been the first settlers in India. Nevertheless, certain controversy exists, due to the support given to the thesis that colonization was by the Austro-Asiatic group, prior to the Europeans. Thus, the aim was to investigate pre-historic colonization of India by anatomically modern humans, using conserved stretches of five amino acid (EPIYA) sequences in the cagA gene of Helicobacter pylori. Simultaneously, the existence of a pathogenic relationship of tyrosine phosphorylation motifs (TPMs), in 32 H. pylori strains isolated from subjects with several forms of gastric diseases, was also explored. High resolution sequence analysis of the above described genes was performed. The nucleotide sequences obtained were translated into amino acids using MEGA (version 4.0) software for EPIYA. An MJ-Network was constructed for obtaining TPM haplotypes by using NETWORK (version 4.5) software. The findings of the study suggest that Indian H. pylori strains share a common ancestry with Europeans. No specific association of haplotypes with the outcome of disease was revealed through additional network analysis of TPMs.

Relevance of Helicobacter pylori genotypes in gastric pathology and its association with plasma malondialdehyde and nitric oxide levels.

Persistent infection with Helicobacter pylori confers an increased risk of peptic ulceration and gastric adenocarcinoma. Reactive oxygen and nitrogen species play a crucial role in the progression from normal gastric mucosa to cancer. The aim of the present study was to investigate the plasma malondialdehyde and nitric oxide levels in H. pylori related gastroduodenal diseases and associate their levels with gastric pathology and genotypes of H. pylori. Malondialdehyde and nitric oxide levels in plasma samples of 250 subjects were spectrophotometrically determined. Subsequently, genotypic and histopathological assessment was performed in gastric biopsies obtained during endoscopy. The levels of MDA and NO exceeded in subjects infected with genotype-1 of Hp than those with other genotypes suggesting more precise interaction of highly virulent strains of Hp in eliciting severe tissue damage. In conclusion, the study demonstrates close relationship between the plasma malondialdehyde and nitric oxide levels, gastric histopathology and genotypes of H. pylori.

Dopamine transporter (DAT1) VNTR polymorphism in 12 Indian populations.

The dopamine transporter (DAT1) is a membrane spanning protein that binds the neurotransmitter dopamine and performs re-uptake of dopamine from the synapse into a neuron. The gene encoding DAT1 consists of 15 exons spanning 60 kb on chromosome 5p15.32. Several studies have investigated the possible associations between variants in DAT1 gene and psychiatric disorders. The present study aimed to determine the distribution of the variable number of tandem repeat (VNTR) polymorphism in the 3' untranslated region of DAT1 in 12 Indian populations. A total of 471 healthy unrelated individuals in 12 Indian populations from 3 linguistic groups were included in the present study. The analysis was carried out using PCR and electrophoresis. Overall, 4 alleles of the DAT1 40-bp VNTR, ranging from 7 to 11 repeats were detected. Heterozygosity indices were low and varied from 0.114 to 0.406. The results demonstrate the variability of the DAT1 40-bp VNTR polymorphism in Indian populations and revealed a high similarity with East Asian populations.

Deep rooting in-situ expansion of mtDNA Haplogroup R8 in South Asia.

BACKGROUND: The phylogeny of the indigenous Indian-specific mitochondrial DNA (mtDNA) haplogroups have been determined and refined in previous reports. Similar to mtDNA superhaplogroups M and N, a profusion of reports are also available for superhaplogroup R. However, there is a dearth of information on South Asian subhaplogroups in particular, including R8. Therefore, we ought to access the genealogy and pre-historic expansion of haplogroup R8 which is considered one of the autochthonous lineages of South Asia. METHODOLOGY/PRINCIPAL FINDINGS: Upon screening the mtDNA of 5,836 individuals belonging to 104 distinct ethnic populations of the Indian subcontinent, we found 54 individuals with the HVS-I motif that defines the R8 haplogroup. Complete mtDNA sequencing of these 54 individuals revealed two deep-rooted subclades: R8a and R8b. Furthermore, these subclades split into several fine subclades. An isofrequency contour map detected the highest frequency of R8 in the state of Orissa. Spearman's rank correlation analysis suggests significant correlation of R8 occurrence with geography. CONCLUSIONS/SIGNIFICANCE: The coalescent age of newly-characterized subclades of R8, R8a (15.4+/-7.2 Kya) and R8b (25.7+/-10.2 Kya) indicates that the initial maternal colonization of this haplogroup occurred during the middle and upper Paleolithic period, roughly around 40 to 45 Kya. These results signify that the southern part of Orissa currently inhabited by Munda speakers is likely the origin of these autochthonous maternal deep-rooted haplogroups. Our high-resolution study on the genesis of R8 haplogroup provides ample evidence of its deep-rooted ancestry among the Orissa (Austro-Asiatic) tribes.