Transcriptional modulation of calcium-permeable AMPA receptor subunits in glioblastoma by MEK-ERK1/2 inhibitors and their role in invasion.

Glioblastoma is the most common primary brain tumor. Glioblastoma cells secrete a significant amount of glutamate, which serve as a potential growth factor in glioma pathobiology through their specific receptor subtypes including α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR). Glioblastoma express AMPAR subunits; however, its regulation and activation with downstream intracellular signaling are not well-understood. Phosphorylated-extracellular signaling-regulated kinase (ERK)1/2 is known to regulate the ionotropic glutamate receptors in cortical neurons. The mitogen-activated protein kinase cascade is frequently activated in several tumors, including glioma. Nonetheless, the association of ERK signaling with AMPAR subunits in glioblastoma is undetermined. Here, we demonstrated potential role of AMPAR in invasion, and the modulation of AMPAR subunits at transcript level by ERK signaling in glioblastoma cells. Inhibition of ERK signaling specifically downregulated the expression of calcium-permeable AMPAR subunits, GluA1 and GluA4, and upregulated calcium-impermeable AMPAR subunit GluA2 implying differential regulation of the expression of calcium-permeable AMPAR subunits of glioblastoma. Concomitantly, it significantly decreased the invasion of U87MG cells. Taken together, these findings suggest that the AMPAR enhances invasion of glioblastoma, and ERK signaling modulates the differential expression of calcium-permeable AMPAR phenotype that might play a crucial role in the invasive propensity of glioblastoma cells.

TNF-α mediated MEK-ERK signaling in invasion with putative network involving NF-κB and STAT-6: a new perspective in glioma.

Glioblastoma is the most common malignant primary brain tumor with poor prognosis. Invasion involves pro-inflammatory cytokines and major signaling hubs. Tumor necrosis factor-α (TNF-α) acts as a master switch in establishing an intricate link between inflammation and cancer. The present study attempted to explore the possible implication of MAPK extracellular signaling-regulated kinase kinase (MEK)-extracellular signaling-regulated kinase (ERK) signaling pathway and expression of nuclear factor-κB (NF-κB), signal transducers and activators of transcription-6 (STAT-6), ERK, and phosphorylated-ERK (p-ERK) signaling proteins in TNF-α microenvironment. U0126 and PD98059 were used to inhibit the MEK-ERK1/2 pathway. TNF-α stimulation enhanced invasion in U87MG, U251MG and patient-derived primary glioma cells, whereas cell viability was not altered. Matrix metalloproteinase-2 (MMP-2) activity was increased only in U251MG glioma cells. These data suggest that TNF-α microenvironment plays an important role in the invasion of U251MG, U87MG, and patient-derived primary glioma cells, without any cytotoxic effect. The MMP-2 activity is differentially regulated by TNF-α stimulation in these cells. TNF-α stimulation upregulated the protein expression of ERK-1, ERK-2 and also increased the level of p-ERK1/2. TNF-α stimulation further upregulated the expression of NF-κB1, STAT-6 in tandem with Ras-MEK signaling system in U87MG cells, which emphasized the possible involvement of these signaling hubs in the glioma microenvironment. MEK-ERK inhibitors significantly attenuated the invasion of U87MG cells mediated by the TNF-α stimulation, probably through their inhibitory impact on p-ERK1/2 and ERK-2. This study provides the possible rationale of invasion by glioma cells in a TNF-α-induced pro-inflammatory milieu, which involves direct role of MEK-ERK signaling, with possible implication of NF-κB and STAT-6.

Survival of glioblastoma cells in response to endogenous and exogenous oxidative challenges: possible implication of NMDA receptor-mediated regulation of redox homeostasis.

Cancer cells are highly metabolically active and produce high levels of reactive oxygen species (ROS). Drug resistance in cancer cells is closely related to their redox status. The role of ROS and its impact on cancer cell survival seems far from elucidation. The mechanisms through which glioblastoma cells overcome aberrant ROS and oxidative stress in a milieu of hypermetabolic state is still elusive. We hypothesize that the formidable growth potential of glioma cells is through manipulation of tumor microenvironment for its survival and growth, which can be attributed to an astute redox regulation through a nexus between activation of N-methyl-d-aspartate receptor (NMDAR) and glutathione (GSH)-based antioxidant prowess. Hence, we examined the NMDAR activation on intracellular ROS level, and cell viability on exposure to hydrogen peroxide (H2 O2 ), and antioxidants in glutamate-rich microenvironment of glioblastoma. The activation of NMDAR attenuated the intracellular ROS production in LN18 and U251MG glioma cells. MK-801 significantly reversed this effect. On evaluation of GSH redox cycle in these cells, the level of reduced GSH and glutathione reductase (GR) activity were significantly increased. NMDAR significantly enhanced the cell viability in LN18 and U251MG glioblastoma cells, by attenuating exogenous H2 O2 -induced oxidative stress, and significantly increased catalase activity, the key antioxidant that detoxifies H2 O2 . We hereby report an unexplored role of NMDAR activation induced protection of the rapidly multiplying glioblastoma cells against both endogenous ROS as well as exogenous oxidative challenges. We propose potentiation of reduced GSH, GR, and catalase in glioblastoma cells through NMDAR as a novel rationale of chemoresistance in glioblastoma.

IL-1ß microenvironment promotes proliferation, migration, and invasion of human glioma cells.

Among the primary brain tumors, glioblastoma is the most common and severe. Glioblastoma have poor prognosis because of their highly diffusive growth pattern and invasion into surrounding brain tissue. Human glioblastoma cells overexpress interleukin-1ß (IL-1ß) and also the levels of IL-1ß in U87MG glioma cells are significantly higher than in U373, T98G, A172 glioma cell lines. Malignant tumors are characterized by unlimited proliferation, migration and invasion. This study examines the effect of IL-1ß microenvironment on proliferation, migration and invasion in human glioma cell line U87MG that expresses wild-type p53 protein, and U251MG which expresses mutant p53. Proliferation was investigated by MTT assay, migration by wound-healing migration assay and invasion by in vitro transwell Matrigel invasion assay. An IL-1ß microenvironment significantly increased migration and invasion of both wild-type and mutant p53 expressing glioma cells, but significantly increased proliferation only in U87MG glioma cells. These effects were inhibited by IL-1 receptor antagonist (IL-1Ra), thus giving evidence that an IL-1ß milieu promotes glioma cell migration, invasion, and proliferation.

Activation of NMDA receptor of glutamate influences MMP-2 activity and proliferation of glioma cells.

Glioblastoma multiforme (GBM) is the most common malignant glioma, which has high proliferative rate and an extremely invasive phenotype. Major limitations in the effective treatment of malignant gliomas are the proliferation and infiltration into the surrounding brain tissue. Although studies have shown that various stimuli promote glioma cell proliferation and invasion, the underlying mechanisms remain largely unknown. Glioma cells secrete significant amount of glutamate into surrounding tissue and intracellular signaling is thought to be initiated upon glutamate-induced modulation of the ion channels in GBM cells. The objective of the study was to investigate the effect of activation of NMDA (N-methyl-D-aspartate) receptors of glutamate on gelatinase subfamily MMPs and on proliferation of glioma cells. U251MG and U87MG cell lines were maintained in Dulbecco's Modified Eagle's Medium. Proliferation assay was investigated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, a yellow tetrazole (MTT) assay. Matrix metalloproteinase (MMP)-2 and MMP-9 activity was investigated by gelatin zymography assay. We demonstrate that activated NMDA receptors (NMDAR) increased the activity of MMP-2 only in U251MG glioma cells at concentrations of 100 and 200 µM and increased the proliferation of both U87MG and U251MG glioma cells at concentrations of 50, 100, 150 and 200 µM. Inhibition of NMDAR using MK-801, a non-competitive antagonist of the NMDAR, significantly inhibited the effect of activation of NMDAR on MMP-2 activity and on proliferation. We conclude that NMDA receptor activation has role in activity of MMP-2 and proliferation of glioma cells.

Circulating microRNA profiling identifies microRNAs linked to prediabetes associated with alcohol dependence syndrome.

BACKGROUND: MicroRNAs are abundant in serum and have emerged as important regulators of gene expression, implicating them in a wide range of diseases. The purpose of this study was to discover and validate serum miRNAs in prediabetes associated with alcohol dependence syndrome (ADS). METHOD: Serum samples from ADS patients with or without prediabetes and normoglycemic controls were subjected to microarray. Validation of identified candidate miRNAs was performed by RT-qPCR. Additionally, GO and KEGG pathway analyses were carried out to uncover target genes anticipated to be controlled by the candidate miRNAs. RESULTS: Notably, 198, and 172 miRNAs were differentially expressed in ADS-patients with or without prediabetes compared to healthy controls, and 7 miRNAs in ADS-patients with prediabetes compared to ADS-normoglycemic patients, respectively. Furthermore, hsa-miR-320b and hsa-miR-3135b were differentially expressed exclusively in ADS-patients with prediabetes, and this was further validated. Interestingly, GO and KEGG pathway analysis revealed that genes predicted to be modulated by the candidates were considerably enriched in numerous diabetes-related biological processes and pathways. CONCLUSION: Our findings revealed that ADS-patients with or without prediabetes have different sets of miRNAs compared to normoglycemic healthy subjects. We propose serum hsa-miR-320b and hsa-miR-3135b as potential biomarkers for the diagnosis of prediabetes in ADS-patients.

Plasma microRNAs as a Potential Biomarker for Identification of Progressive Supranuclear Palsy.

Progressive supranuclear palsy (PSP) is the second most common Parkinsonian disorder with complex etiology. The underlying molecular mechanism of PSP pathogenesis remains unclear. The present study aims to find the feasibility of using plasma miRNAs as novel biomarkers. Plasma-focused qPCR panels were used for microRNA profiling and identified differentially expressed microRNAs in PSP compared to controls. The DIANA-miRPath v3.0 was used to perform KEGG pathway analysis. We then confirmed the expression of selected candidates by RT-qPCR and their clinical utility was assessed by ROC analysis. Profiling data revealed 28 differentially expressed microRNAs in PSP. Five overexpressed miRNAs were selected for further analysis. The KEGG pathway analysis revealed 48 high-risk pathways. The study revealed that as a single marker-miR-19b-3p, miR-33a-5p, miR-130b-3p, miR-136-3p, and miR-210-3p had a specificity of 64.71%, 82.35%, 68.75%, 82.35%, and 70.59% at sensitivity 77.78%, 77.78%, 66.67%, 73.33%, and 66.67%, respectively. The result suggests that circulating plasma miRNAs were altered in PSP compared to control. The findings of this study may provide potential biomarkers and pathways associated with PSP. Further large-scale validation studies are required to confirm the same.

Inflammatory bowel diseases in Tamil Nadu: A survey of demographics, clinical profile, and practices.

BACKGROUND: Inflammatory bowel disease (IBD) is increasingly diagnosed in South Asia. This survey by the Tamil Nadu Chapter of the Indian Society of Gastroenterology (TNISG) documents the demography, clinical profile, and therapeutic practices related to IBD in Tamil Nadu. METHODS: TNISG members from 32 institutions completed an online cross-sectional questionnaire on IBD patients from March 2020 to January 2021. RESULTS: Of 1295 adult IBD patients, 654 had Crohn's disease (CD), 499 ulcerative colitis (UC), and 42 IBD-unclassified (IBD-U). CD and UC showed a unimodal age distribution. A total of 55% were graduates or postgraduates. A positive family history was noted in 30, other risk factors were uncommon. In CD, the pattern of involvement was ileocolonic (42.8%), ileal (34.7%), colonic (18.9%), and upper gastrointestinal (3.5%); while in UC, disease was characterized as extensive (44.9%), left-sided (41.7%), or proctitis (13.4%). Perineal disease, perianal fistulae, and bowel obstruction were noted in 4.3, 14.0, and 23.5%, respectively, of CD. The most widely used drugs were mesalamine, azathioprine, and corticosteroids. Surgery was undertaken in 141 patients with CD and 23 patients with UC. Of the 138 patients with pediatric IBD (≤16 years), 23 were characterized as very early onset IBD (VEO-IBD), 27 as early-onset, and 88 as adolescent IBD. VEO-IBD were more likely to have a positive family history of IBD and were more likely to have perineal disease and to have the IBD-U phenotype. Among pediatric IBD patients, corticosteroids, mesalamine, and azathioprine were the most commonly used medications, while 25 pediatric patients received biologics. CONCLUSION: This study provides important information on demography, clinical profile, and treatment practices of IBD in India.

Clinical Application of Circulating MicroRNAs in Parkinson's Disease: The Challenges and Opportunities as Diagnostic Biomarker.

Discovery of evolutionarily conserved, nonprotein-coding, endogenous microRNAs has induced a paradigm shift in the overall understanding of gene regulation. Now, microRNAs are considered and classified as master regulators of gene expression as they regulate a wide range of processes - gene regulation, splicing, translation and posttranscriptional modifications. Besides, dysregulated microRNAs have been related to many diseases, including Parkinson's and related disorders. Several studies proposed that differentially expressed microRNAs as a potential biomarker. So far, there is no accepted clinical diagnostic test for Parkinson's disease based on biochemical analysis of biological fluids. However, circulating microRNAs possess many vital features typical of reliable biomarkers and discriminates Parkinson's patients from healthy control with much higher sensitivity and specificity. Though they show tremendous promise as a putative biomarker, translating these research findings to clinical application is often met with many obstacles. Most of the candidate microRNAs reported as a diagnostic biomarker is not organ-specific, and their overlap is low between studies. Therefore this review aimed to highlight the challenges in the application of microRNA in guiding disease discrimination decisions and its future prospects as a diagnostic biomarker in Parkinson's Disease.

Role of MEK-ERK signaling mediated adhesion of glioma cells to extra-cellular matrix: Possible implication on migration and proliferation.

BACKGROUND: Glioblastoma represents the most common primary brain tumor with a worst prognosis despite developments in neurosurgery and chemoradiotherapy. Detachment of the cells from the primary tumor tissue is a prerequisite for their dispersion and spreading. Initial and incessant dispersal of tumor cells from the primary tumor tissue renders GBM refractory to comprehensive surgical removal and increases the chance of recurrence and poorer prognosis. PURPOSES: The current study was designed to investigate the effect of inhibition of MEK-ERK1/2 signaling by PD98059 and U0126 on the growth and migration of glioma cells as well as their adhesion to extracellular matrix. METHODS: MEK-ERK1/2 signaling in U87-MG cells was inhibited by PD98059 and U0126. Migration, proliferation and adhesion were analyzed by scratch-wound assay, MTT assay, cell adhesion assay respectively. RESULTS: PD98059 and U0126 significantly not only reduced the proliferation of glioma cells and attenuated their migration but also increased their adhesion to gelatin of extracellular matrix. CONCLUSION: This study provides the evidence that inhibition of MEK-ERK1/2 signaling enhances the adhesion of glioma cells to gelatin/collagen component of ECM, and decreases the proliferation and migration of the glioma cells. We propose the possible rationale of association between ERK signaling and cell-cell adhesion molecules in glioma microenvironment which regulates the glioma initiation, growth and progression.

MicroRNAs to differentiate Parkinsonian disorders: Advances in biomarkers and therapeutics.

Parkinsonian disorders are a set of progressive neurodegenerative movement disorders characterized by rigidity, tremor, bradykinesia, postural instability and their distinction has significant implications in terms of management and prognosis. Parkinson's disease (PD) is the most common among them. Its clinical diagnosis is challenging and, it can be misdiagnosed in the early stages. Multiple system atrophy and progressive supranuclear palsy are the close mimickers in early stages, due to overlapping clinical features. MicroRNAs are a class of stable non-coding small RNA molecules implicated in post-transcriptional gene regulation. Current studies propose that miRNAs play an essential role in the pathobiology of multiple neurodegenerative disorders including Parkinsonism, and they seem to be one of the reasonably available methods to aid in the differential diagnosis between PD and related disorders. MicroRNA-based diagnostic biomarkers and therapeutics are a powerful tool to understand and explore the function of the pathogenic gene/s, their mechanism in the disease pathobiology, and to validate drug targets. In this review, we emphasize on the recent developments in the usage of miRNAs as diagnostic biomarkers to identify PD and to differentiate it from atypical parkinsonian conditions, their role in disease pathogenesis, and their possible utility in the therapy of these disorders.

Duodenal Lipomatosis as a Curious Cause of Upper Gastrointestinal Bleed: A Report with Review of Literature.

Lipomas of the gastrointestinal tract are rare. Duodenal lipomas are incidental and mostly asymptomatic. Tumours may produce symptoms of abdominal pain and discomfort or cause bleeding due to ulceration or intestinal obstruction due to intussusception. We describe a 45-year-old man presenting in emergency with 3 days history of melena with normal gastroduodenoscopy and contrast enhanced computed tomography revealing multiple polypoid lesion in duodenum and proximal jejunum suggestive of lipoma. Due to ongoing bleed, he underwent laparotomy with duodenectomy and uneventful postoperative recovery. Our review of cases published in last 67 years indicate that duodenal lipomas are rare to occur but commonly found in second part, they may be seen in third and fourth part of duodenum which may be missed on endoscopy. They can be multiple and may present as severe UGI bleeding which could be managed surgically. Though CT is diagnostic, histopathology confirms the diagnosis which shows lipomatous lesion composed of mature adipose arranged in lobules.

Sevoflurane and thiopental preconditioning attenuates the migration and activity of MMP-2 in U87MG glioma cells.

BACKGROUND: Tumor cell migration and diffuse infiltration into brain parenchyma are known causes of recurrence after treatment in glioblastoma (GBM), mediated in part by the interaction of glioma cells with the extracellular matrix, followed by degradation of matrix by tumor cell derived proteases, particularly the matrix metalloproteinases (MMP). Sevoflurane and thiopental are anesthetics commonly used in cancer surgery. However, their effect on the progression of glioma cells remains unclear. The aim of this study was to explore the role of these anesthetics on the migration and activity of MMP-2 in glioma cells. METHODOLOGY: Cultured U87MG cells were pretreated with sevoflurane or thiopental and in vitro wound healing scratch assay was carried out to analyze their effect on migration of these cells. Gelatin zymography was carried out to examine the effect of these anesthetics on tumor cell MMP-2 activity using the conditioned media 24 h after pretreatment. Cell viability was analyzed using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay. RESULTS: U87MG cells exposed to 2.5% sevoflurane or different concentrations of thiopental significantly decreased migration and activity of MMP-2 compared to control. No effect was seen on the viability of these cells after pretreatment with sevoflurane or thiopental. CONCLUSION/SIGNIFICANCE: These results suggest that both sevoflurane and thiopental have inhibitory effect on the migration and MMP-2 activity in glioma cells. Thus, it is important that the choice of anesthetics to be used during glioma surgery takes into account their inhibitory properties against the tumor cells.