Worst pattern of invasion in oral squamous cell carcinoma is an independent prognostic factor.

Introduction: Biologic aggressiveness of OSCC (Oral Cavity Squamous Cell Carcinoma), has intrigued research in various prognosticating histopathological markers over past few decades. DOI (Depth of Invasion) is one such histopathological factor which affects outcomes and was included in the AJCC 8th edition TNM staging. Pattern of Invasion (POI) has been widely reported as an adverse prognostic factor associated with higher locoregional failure and poor prognosis. However, these factors are not utilized for treatment decision making and for outcome assessment. Materials and methods: This is a retrospective analysis of 320 patients with OSCC who underwent treatment, from October 2018-February 2020. Clinic demographic details were extracted from electronic medical records. Univariate and multivariate analysis was done for the parameters. WPOI (Worst Pattern of Invasion) was correlated with all histopathological prognostic factors. Survival analysis was done using Kaplan Meier for WPOI type's I-V. DFS (Disease free Survival) was evaluated for different grades of WPOI. Results: We analyzed the results comparing, early and advanced T (Tumor) stages, cohesive WPOI I-III, non-cohesive WPOI IV-V. Univariate analysis showed a significant association of T-stage (p = 0.001), N (Nodal) -stage (p = 0.002), DOI (p = 0.008), PNI (Peri-neural invasion) (0.001) and Tumor differentiation Grade (p = 0.001). On multivariate analysis, non-cohesive WPOI (IV & V) showed significant association with grade, PNI, DOI (0.002, 0.033 & 0.033 respectively). Non-cohesive WPOI had significantly higher locoregional failures and short DFS. Conclusion: Presence of invasive WPOI is associated with advanced T stage, poor differentiation, PNI, greater depth of invasion, and higher chances of nodal metastasis. WPOI is associated with poor DFS, treatment intensification in early stage disease with WPOI type IV & V may improve survival.

Extramedullary Solitary Plasmacytoma of Larynx: A Rare Entity and Therapeutic Challenge.

Solitary extramedullary plasmacytoma is a rare entity. The diagnosis is established by clinical evaluation, detailed radiological workup and histopathology with immunohistochemistry markers. There is no clear standard of care for a solitary laryngeal plasmacytoma. A case of solitary laryngeal plasmacytoma is being reported here with the rationale for the chosen treatment modality.

Molecular characterization of a series of solitary fibrous tumors, including immunohistochemical expression of STAT6 and NATB2-STAT6 fusion transcripts, using Reverse Transcriptase(RT)-Polymerase chain reaction(PCR) technique: An Indian experience.

A solitary fibrous tumor (SFT) is characterized by a diverse clinicopathologic spectrum. Recent studies have unraveled STAT6 as a useful diagnostic immunohistochemical (IHC) marker for a SFT and NAB2-STAT6 as its specific gene fusion transcript. Thirty-three SFTs were tested for STAT6 immunostaining by polymer detection technique. STAT6 immunoexpression was further graded, based on intensity (mild, moderate and strong) and percentage of immunopositive tumor cells, ranging from 1 to 25%(1+); 26-50%(2+); 51-75%(3+) and in more than 75%(4+) tumor nuclei. These cases along with 17 other tumors were tested for 8 variants of NAB2-STAT6, using qualitative endpoint reverse-transcriptase (RT)-PCR technique. RNA extraction was performed using Recover All Total nucleic acid extraction kit. The selected cases were screened for all the 8 fusion variants, using 8 primer pairs for NAB2 and STAT6 genes. Thirty-three SFTs occurred in 18 men and 15 women (M: F=1.2:1), with age varying from 13 to 74 years(average=49.6); across various body sites. Immunohistochemically, most SFTs (30/33) (90.9%) displayed moderate to strong immunostaining for STAT6, including 3+ and 4+ immunostaining patterns in 27/33 (81.8%) tumors. By RT-PCR, 30/33(90.9%) cases of SFT were positive for NAB2-STAT6 fusions, including NAB2ex4/STAT6ex2 (7cases), NAB2ex7/STAT6ex2 (7cases), NAB2ex6/STAT6ex3 (6cases), NAB2ex6/SAT6ex16 (4cases), NAB2ex3/STAT6ex19 (4cases), NAB2ex6/STAT6ex17 (single case), NAB2ex4/STAT6ex4 (single case) and NAB2ex6/STAT6ex18 (none). NAB2-STAT6 fusions were not observed in 9 cases of synovial sarcoma, 4 of Ewing sarcoma, 2 of MPNST and 2 cases of dedifferentiated liposarcomas (100% specificity). On comparing with clinical outcomes, more cases (7/11)(63.6%) of classic SFT were associated with favorable outcomes, while more cases(5/8)(62%) of atypical and malignant SFTs were associated with aggressive outcomes. This study reinforces high sensitivity and specificity of NAB2-STAT6 fusion and its correlation with strong and diffuse IHC expression of STAT6 in a SFT, irrespective of its occurrence in various body sites and its histopathologic types. NAB2ex4-STAT6ex2 and NAB2ex7-STAT6ex2 fusions were relatively more frequently observed in our patients. Atypical and malignant SFTs, together, were more frequently associated with relatively aggressive clinical outcomes.

A Comprehensive Inter-Tissue Crosstalk Analysis Underlying Progression and Control of Obesity and Diabetes.

Obesity is a metabolic state associated with excess of positive energy balance. While adipose tissues are considered the major contributor for complications associated with obesity, they influence a variety of tissues and inflict significant metabolic and inflammatory alterations. Unfortunately, the communication network between different cell-types responsible for such systemic alterations has been largely unexplored. Here we study the inter-tissue crosstalk during progression and cure of obesity using multi-tissue gene expression data generated through microarray analysis. We used gene expression data sets from 10 different tissues from mice fed on high-fat-high-sugar diet (HFHSD) at various stages of disease development and applied a novel analysis algorithm to deduce the tissue crosstalk. We unravel a comprehensive network of inter-tissue crosstalk that emerges during progression of obesity leading to inflammation and insulin resistance. Many of the crosstalk involved interactions between well-known modulators of obesity and associated pathology like inflammation. We then used similar datasets from mice that in addition to HFHSD were also administered with a herbal concoction known to circumvent the effects of HFHSD in the diet induced model of obesity in mice. We propose, the analysis presented here could be applied to understand systemic details of several chronic diseases.

Molecular signatures for obesity and associated disorders identified through partial least square regression models.

BACKGROUND: Obesity is now a worldwide epidemic disease and poses a major risk for diet related diseases like type 2 diabetes, cardiovascular disease, stroke and fatty liver among others. In the present study we employed the murine model of diet-induced obesity to determine the early, tissue-specific, gene expression signatures that characterized progression to obesity and type 2 diabetes. RESULTS: We used the C57BL/6 J mouse which is known as a counterpart for diet-induced human diabetes and obesity model. Our initial experiments involved two groups of mice, one on normal diet (ND) and the other on high-fat and high-sucrose (HFHSD). The later were then further separated into subgroups that either received no additional treatment, or were treated with different doses of the Ayurvedic formulation KAL-1. At different time points (week3, week6, week9, week12, week15 and week18) eight different tissues were isolated from mice being fed on different diet compositions. These tissues were used to extract gene-expression data through microarray experiment. Simultaneously, we also measured different body parameters like body weight, blood Glucose level and cytokines profile (anti-inflammatory & pro-inflammatory) at each time point for all the groups. Using partial least square discriminant analysis (PLS-DA) method we identified gene-expression signatures that predict physiological parameters like blood glucose levels, body weight and the balance of pro- versus anti-inflammatory cytokines. The resulting models successfully predicted diet-induced changes in body weight and blood glucose levels, although the predictive power for cytokines profiles was relatively poor. In the former two instances, however, we could exploit the models to further extract the early gene-expression signatures that accurately predict the onset of diabetes and obesity. These extracted genes allowed definition of the regulatory network involved in progression of disease. CONCLUSION: We identified the early gene-expression signature for the onset of obesity and type 2 diabetes. Further analysis of this data suggests that some of these genes could be used as potential biomarkers for these two disease-states.

Immunomodulatory Role of an Ayurvedic Formulation on Imbalanced Immunometabolics during Inflammatory Responses of Obesity and Prediabetic Disease.

Kal-1 is a polyherbal decoction of seven different natural ingredients, traditionally used in controlling sugar levels, inflammatory conditions particularly regulating metabolic and immunoinflammatory balance which are the major factors involved in obesity and related diseases. In the present study, we aimed to investigate the effect of Kal-1 (an abbreviation derived from the procuring source) on diet-induced obesity and type II diabetes using C57BL/6J mice as a model. The present study was performed with two experimental groups involving obese and prediabetic mice as study animals. In one, the mice were fed on high-fat with increased sucrose diet, and different amounts (5, 20, and 75 µ L) of Kal-1 were administered with monitoring of disease progression over a period of 21 weeks whereas in the second group the mice were first put on the same diet for 21 weeks and then treated with the same amounts of Kal-1. A significant reduction in body weight, fat pads, fasting blood glucose levels, insulin levels, biochemical parameters, immunological parameters, and an array of pro- and anticytokines was observed in obese and diabetic mice plus Kal-1 than control (lean) mice fed on normal diet. In conclusion, Kal-1 has immunomodulatory potential for diet-induced obesity and associated metabolic disorders.

Gene set enrichment analysis identifies LIF as a negative regulator of human Th2 cell differentiation.

In this study we show that IL-4 is crucial during reinforcement window of human Th2 differentiation for optimal Th2 development. We have also shown here that during this stage, IL-4 helps in cellular decision-making process of differentiation versus proliferation. We have combined computational and experimental methods to analyze Th2 transcription network to name novel players of the process of Th2 differentiation. Here we report that LIF through STAT3 negatively regulates Th2 differentiation. This approach can be generalized to analyze "omics" data to identify key regulatory modules.

A novel mechanism for ERK-dependent regulation of IL4 transcription during human Th2-cell differentiation.

We demonstrate that the mitogen-activated protein kinases extracellular signal-regulated kinase (ERK)-1 and ERK-2 have a central role in mediating T-cell receptor-dependent induction of IL4 expression in human CD4(+) T cells. Significantly, this involved a novel mechanism wherein receptor cross-linking induced activated ERK to physically associate with a promoter element on the IL4 gene. The proximally localized ERK then facilitated recruitment of the key transcription factors necessary for initiating IL4 gene transcription. Although both ERK-1 and ERK-2 bound to the promoter, recruitment of either one alone was found to be sufficient. We thus identify a novel mode of function for ERK wherein its physical association with the promoter serves as a prerequisite for enhanceosome assembly. This unusual pathway is also indispensable for human Th2-cell differentiation.

Innate and specific gut-associated immunity and microbial interference.

Certain bacterial species isolated from the gastrointestinal microbial communities release low-molecular-weight peptides into milk products using bacteria-derived proteases that degrade milk casein, and thereby generate peptides, triggering immune responses. The intestinal microbial communities contributes to the processing of food antigens in the gut. The present study was designed to investigate the immunomodulatory effects of microbial interference to determine whether casein degraded by probiotic bacteria-derived enzymes could modulate the cytokine production and peripheral blood mononuclear cells in atopic infants with cow or other synthetic milk allergy. Without hydrolyzation, casein reduced the production of interleukin-4, which indicates that probiotics modify the structure of potentially harmful antigens and thereby alter the mode of their immunogenicity. Intraluminal bacterial antigens have been reported to elicit specific responses in the gut-associated lymphoid tissue (GALT) through the binding capacity of intraluminal bacterial antigens to epithelial cells, which allows antigen entry via enterocytes and aids in evading the tolerance function in Peyer's patches. Such tonic immune responses in the GALT may allow control of the metabolic activity and balance of the gut microbial communities.

The role of nitric oxide in inflammatory reactions.

Nitric oxide (NO) was initially described as a physiological mediator of endothelial cell relaxation, an important role in hypotension. NO is an intercellular messenger that has been recognized as one of the most versatile players in the immune system. Cells of the innate immune system--macrophages, neutrophils and natural killer cells--use pattern recognition receptors to recognize the molecular patterns associated with pathogens. Activated macrophages then inhibit pathogen replication by releasing a variety of effector molecules, including NO. In addition to macrophages, a large number of other immune-system cells produce and respond to NO. Thus, NO is important as a toxic defense molecule against infectious organisms. It also regulates the functional activity, growth and death of many immune and inflammatory cell types including macrophages, T lymphocytes, antigen-presenting cells, mast cells, neutrophils and natural killer cells. However, the role of NO in nonspecific and specific immunity in vivo and in immunologically mediated diseases and inflammation is poorly understood. This Minireview will discuss the role of NO in immune response and inflammation, and its mechanisms of action in these processes.

Immune response to leishmania: paradox rather than paradigm.

The leishmaniases are a group of diseases caused by protozoan parasites of the genus Leishmania. Various Leishmania species can cause human infection, producing a spectrum of clinical manifestations. It is estimated that 350 million people are at risk, with a global yearly incidence of 1-1.5 million for cutaneous and 500,000 for visceral leishmaniasis (VL). VL is a major cause of morbidity and mortality in East Africa and the Indian subcontinent. Coinfection with HIV enhances the risk of the disease. The only control measure currently available in India is case detection and treatment with antimonial drugs, which are expensive, not always available and cannot be self-administered. Newer drugs like oral miltefosine have not become widely available. Vector and reservoir control is difficult due to the elusive nature of the vector and the diversity of the animal reservoir. A detailed knowledge of immune response to the parasite would help in designing prophylactic and therapeutic strategies against this infection.

Nitric oxide and immune response.

Nitric oxide (NO), initially described as a physiological mediator of endothelial cell relaxation plays an important role in hypotension. It is an intercellular messenger and has been recognized as one of the most versatile players in the immune system. Cells of the innate immune system--macrophages, neutrophils and natural killer (NK) cells use pattern recognition receptors to recognize molecular patterns associated with pathogens. Activated macrophages then inhibit pathogen replication by releasing a variety of effector molecules, including NO. In addition to macrophages, a large number of other immune system cells produce and respond to NO. Thus, NO is important as a toxic defense molecule against infectious organisms. It also regulates the functional activity, growth and death of many immune and inflammatory cell types including macrophages, T lymphocytes, antigen-presenting cells, mast cells, neutrophils and NK cells. However, the role of NO in non-specific and specific immunity in vivo and in immunologically mediated diseases and inflammation is poorly understood. This review discusses the role of NO in immune response and inflammation and its mechanisms of action in these processes.

Leishmania donovani: identification of stimulatory soluble antigenic proteins using cured human and hamster lymphocytes for their prophylactic potential against visceral leishmaniasis.

Most of the studies for the identification of prophylactic antigens that elicit T cell responses were concentrated on membrane proteins of Leishmania donovani. This study was taken up to assess L. donovani soluble promastigote antigens for their ability to stimulate proliferation of peripheral blood mononuclear cells (PBMCs) from cured visceral leishmaniasis (VL) patients, endemic and non-endemic controls and lymphocytes/peritoneal macrophages of cured hamsters. The soluble protein was subjected to sequential precipitation with saturated ammonium sulphate (20%, 40%, 60% and 80%), of which largely 80% fractioned protein showed significant cellular responses in cured patients and hamsters. This fraction was further fractionated into five sub fractions by preparative SDS-PAGE and subjected to re-evaluation for their ability to induce cellular responses. Out of these, only F2 sub fraction belonging to the MW of 97.4-68 kDa stimulated remarkable lymphoproliferative and IFN-gamma responses in cured VL patients and in endemic controls. Similarly, significant lymphoproliferative responses and nitric oxide production were also noticed in cured Leishmania infected animals indicating an element of uniformity in responses between hamster and human. F2 sub fraction, when evaluated for its prophylactic efficacy with BCG against L. donovani challenge in hamster exhibited significant parasite inhibition in spleen (71.1%; p<0.001) and liver (68.2%; p<0.001) as compared to their unvaccinated counterpart. The vaccinated animals showed significant lymphoproliferative response and nitric oxide production but leishmania specific IgG level were suppressed. The results indicate the presence of immunostimulatory and protective molecules in F2 sub fraction which may further be exploited for the development of a vaccine against VL, hitherto an unrealized goal.

Human S-antigen: peptide determinant recognition in uveitis patients.

Uveitis is an inflammation of the uveal tract and is one of the major causes of visual impairment. Several lines of evidence suggest an important role for activated T lymphocytes in the perpetuation of posterior uveitis. In sequel to our preliminary observations with human S-antigen, we have further investigated the proliferative response of peripheral blood lymphocytes of posterior uveitis patients against 20 linear and 9 overlapping peptides of retinal S-antigen. The expression of surface markers CD4, CD8, CD29, CD45RA in peripheral blood was detected by flow cytometry. We have also assessed the pattern of cytokines present in peripheral blood mononuclear cells (PBMCs) using ribonuclease protection assay (RPA). Nineteen out of 32 patients' lymphocytes showed proliferative response to S-antigen, one or more of its 20 linear and nine overlapping synthetic peptides. Six patients showed significant lymphoproliferative response against various peptides. The maximum response was found to peptides from the 231-270 amino acid region of human S-antigen sequence. The percentage of CD29(+) (memory cells) and CD45RA(+) (naive cells) T-lymphocytes was higher in patients compared to healthy volunteers. There was a demonstrable difference in the percentage of CD4(+) and CD8(+) lymphocytes in the patients (P <== 0.05) as compared to controls. Higher message for interleukin (IL)-5, IL-10, IL-15, IL-9, IL-2, IL-13, and interferon (IFN)-gamma was observed in uveitis patients than in healthy individuals. In brief, our study suggests that a particular region of S-antigen plays an important role in idiopathic uveitis.

Gene therapy in ocular diseases.

Gene therapy is a novel form of drug delivery that enlists the synthetic machinery of the patient's cells to produce a therapeutic agent. Genes may be delivered into cells in vitro or in vivo utilising viral or non-viral vectors. Recent technical advances have led to the demonstration of the molecular basis of various ocular diseases. Ocular disorders with the greatest potential for benefit of gene therapy include hereditary diseases such as retinitis pigmentosa, tumours such as retinoblastoma or melanoma, and acquired proliferative and neovascular retinal disorders. Gene transfer into ocular tissues has been demonstrated with growing functional success and may develop into a new therapeutic tool for clinical ophthalmology in future.