NF-kB Activation Research Articles

In Vivo Effects of Bay 11-7082 on Fibroid Growth and Gene Expression: A Preclinical Study.

Current medical therapies for fibroids have major limitations due to their hypoestrogenic side effects. Based on our previous work showing the activation of NF-kB in fibroids, we hypothesized that inhibiting NF-kB in vivo would result in the shrinkage of tumors and reduced inflammation. Fibroid xenografts were implanted in SCID mice and treated daily with Bay 11-7082 (Bay) or vehicle for two months. Bay treatment led to a 50% reduction in tumor weight. RNAseq revealed decreased expression of genes related to cell proliferation, inflammation, extracellular matrix (ECM) composition, and growth factor expression. Validation through qRT-PCR, Western blotting, ELISA, and immunohistochemistry (IHC) confirmed these findings. Bay treatment reduced mRNA expression of cell cycle regulators (CCND1, E2F1, and CKS2), inflammatory markers (SPARC, TDO2, MYD88, TLR3, TLR6, IL6, TNFα, TNFRSF11A, and IL1β), ECM remodelers (COL3A1, FN1, LOX, and TGFβ3), growth factors (PRL, PDGFA, and VEGFC), progesterone receptor, and miR-29c and miR-200c. Collagen levels were reduced in Bay-treated xenografts. Western blotting and IHC showed decreased protein abundance in certain ECM components and inflammatory markers, but not cleaved caspase three. Ki67, CCND1, and E2F1 expression decreased with Bay treatment. This preclinical study suggests NF-kB inhibition as an effective fibroid treatment, suppressing genes involved in proliferation, inflammation, and ECM remodeling.

Increased mRNA expression of ADAMTS1 and ADAMTS4 in peripheral blood mononuclear cells of psoriasis patients developed psoriatic arthritis

Background and purpose: Psoriatic arthritis (PsA) is a chronic inflammatory disease associated with psoriasis (PsO) affecting both skin and joint. ADAMTS (A disintegrin-like and metalloproteinase domain with thrombospondin-1 repeats) is a large family of proteoglycanase enzymes and the expression levels of ADAMTS proteases are upregulated in arthritis. In this study, we aimed to determine mRNA expression levels of ADAMTS1, ADAMTS4 and ADAMTS5 and identifying the key signaling pathways involved in the regulation of these proteases in peripheral blood mononuclear cells (PBMCs) of patients with PsO who later developed PsA. Materials and methods: 25 PsA patients, 25 PsO patients and 25 healthy individuals were included in this study. PBMCs were isolated from venous blood and mRNA expression levels of ADAMTS1, ADAMTS4 and ADAMTS5 were measured through Real-time quantitative PCR (RTqPCR). Results: mRNA expression levels of ADAMTS1 and ADAMTS4 were found to be increased in PsA patients compared with control and PsO patients. In response to TNF-a stimulation, the expression of ADAMTS1 in PsA patients was determined to be reduced in a Erk1/2 activity dependent manner, whereas p38 and JNK activities were shown to induce the expression of ADAMTS4 in PsA patients. The reduced ADAMTS1 expression in PsA patients induced by IL-1b stimulation was revealed to be dependent on NFkB activity. Conclusions: mRNA expressions of ADAMTS1 and ADAMTS4 regulated by MAPKs and NFkB were increased in PBMCs of PsA patients. This study supports the hypothesis that ADAMTS1 and ADAMTS4 mRNA level might be diagnostic markers for identifying psoriatic patients who are more likely to develop PsA and a future drug target for PsA treatment.

Complement Membrane Attack Complexes Disrupt Proteostasis to Function as Intracellular Alarmins.

Internalized pools of membrane attack complexes (MACs) promote NF-kB and dysregulated tissue inflammation. Here, we show that C9, a MAC-associated protein, promotes loss of proteostasis to become intrinsically immunogenic. Surface-bound C9 is internalized into Rab5 + endosomes whose intraluminal acidification promotes C9 aggregates. A region within the MACPF/CDC domain of C9 stimulates aggrephagy to induce NF-kB, inflammatory genes, and EC activation. This process requires ZFYVE21, a Rab5 effector, which links LC3A/B on aggresome membranes to RNF34-P62 complexes to mediate C9 aggrephagy. C9 aggregates form in human tissues, C9-associated signaling responses occur in three mouse models, and ZFYVE21 stabilizes RNF34 to promote C9 aggrephagy in vivo. Gene-deficient mice lacking ZFYVE21 in ECs showed reduced MAC-induced tissue injury in a skin model of chronic rejection. While classically defined as cytotoxic effectors, MACs may impair proteostasis, forming aggregates that behave as intracellular alarmins.

Identification of a novel GSK3β inhibitor involved in abrogating KRas dependent pancreatic tumors in Wnt/beta-catenin and NF-kB dependent manner

Pancreatic cancer is an aggressive malignancy with a poor survival rate because it is difficult to diagnose the disease during its early stages. The currently available treatments, which include surgery, chemotherapy and radiation therapy, offer only limited survival benefit. Pharmacological interventions to inhibit Glycogen Synthase Kinase-3beta (GSK3β) activity is an important therapeutic strategy for the treatment of pancreatic cancer because GSK3β is one of the key factors involved in the onset, progression as well as in the acquisition of chemoresistance in pancreatic cancer. Here, we report the identification of MJ34 as a potent GSK3β inhibitor that significantly reduced growth and survival of human mutant KRas dependent pancreatic tumors. MJ34 mediated GSK3β inhibition was seen to induce apoptosis in a β-catenin dependent manner and downregulate NF-kB activity in MiaPaCa-2 cells thereby impeding cell survival and anti-apoptotic processes in these cells as well as in the xenograft model of pancreatic cancer. In vivo acute toxicity and in vitro cardiotoxicity studies indicate that MJ34 is well tolerated without any adverse effects. Taken together, we report the discovery of MJ34 as a potential drug candidate for the therapeutic treatment of mutant KRas-dependent human cancers through pharmacological inhibition of GSK3β.

ACTN4-dependent regulation of double-strand DNA break repair is independent of NF-Kb activity

α-Actinin-4 is an actin-binding protein that is involved in a wide range of cellular processes. Along with actin and other proteins of the actin cytoskeleton, α-actinin-4 was found not only in the cytoplasm, but also in the nucleus of various cells. As a nuclear protein, it is involved in regulation of certain transcription factors. In particular, it can regulate transcriptional activity of NF-kB, which largely determines the resistance of cancer cells to apoptosis and anticancer therapy. During our previous studies, it was found that α-actinin-4 can influence resistance of cancer cells to topoisomerase II inhibitors and determine the efficiency of DNA double-strand break repair. We have demonstrated that α-actinin-4 interferes with the assembly of complexes involved in DNA repair via NHEJ and HRR, which in turn leads to an imbalance between these pathways. In this study, we were answering to the question of how α-actinin-4 is involved in the regulation of the DNA double-strand breaks repair following genotoxic stress. Our results indicate that the effect of α-actinin-4 on repair progression in H1299 non-small cell lung cancer cells does not depend on the transcription factor NF-kB activity. We found that in the nucleus of H1299 cells, α-actinin-4 is localized not only in the nucleoplasm, but also reveals close association with chromatin.

Toll-Like Receptor Activation and B Cell Maturation Via MyD88-Dependent Pathway Under Hyperglycemia Condition

hyperglycemia causes a degenerative syndrome that involves an inflammatory process with an increase in certain proinflammatory cytokines and chemokines which in the process will activate B cells to produce immunoglobulins through several mechanisms. One of the interesting mechanisms is the mechanism via the MyD88 pathway. Objectives: to explore the role of MyD88 adapter protein in Toll-like receptor activation and B cell maturation under hyperglycemia conditions. Methods: a literature review was done to answer the study objectives. Results: Class switching process under hyperglycemia conditions involves activation of NFkB through the inflammatory MyD88-dependent pathway to trigger the expression of TLR and B cell maturation and proliferation as well as antibody production. The MyD88 adapter protein is a protein formed by stimulation of pro-inflammatory cytokines IL-6 and plays a role in the continuation of signals from the TLR and IL-1 pathways. Mature B cell stimulation induces 2 genetic changes in the Ig gene locus, called somatic hypermutation (SHM) and class switch recombination (CSR) to produce antibodies. Changes in immunoglobulin genes occur related to changes in certain DNA segments at the locus of genes where CSR occurs, this gene change requires the role of AID (activated-induced cytidine deaminase) in DNA cleavage. AID in mature B cells is activated by proinflammatory cytokines via induction of NfKB activation via the inflammatory MyD88-dependent pathway. Conclusion: Toll-like receptor activation plays a crucial role in B cell maturation activated by pro-inflammatory cytokine via MyD88 dependent-NFkB activation.

Mechanisms of mammalian orthoreovirus-induced cell death and NF-kB activation in ER+ breast cancer.

e13171 Background: Estrogen receptor positive (ER+) breast cancer is one of the most commonly diagnosed types of breast cancer in the United States. Although the five-year survival rate of ER+ breast cancer is 90% or higher, ER+ breast cancer can recur and metastasize 10-20+ years after initial diagnosis. Novel therapeutic strategies are needed to prevent and treat breast cancer recurrence and metastasis. A possible solution is mammalian orthoreovirus (MRV), a replication-competent natural oncolytic virus. The strain T3D (Reolysin) is in current clinical trials to treat metastatic breast cancer; however, the mechanisms of MRV-induced death are not well elucidated. Furthermore, other MRV strains have not been well studied for their oncolytic potential in ER+ breast cancer. Our studies aim to explore the different cell death pathways elicited by MRV. Prior studies have reported that MRV induces cell death in L929 cells (murine fibroblasts, permissive to MRV infection) by activating apoptosis, necroptosis, and inhibiting the NFκB pathway to enhance viral replication. Methods: Our goal is to compare the effects of MRV in L929 cells and ER+ breast cancer cell line MCF7 using chemical inhibitors of necroptosis, apoptosis, and the NFκB pathway. Results: Our results indicate that necroptosis inhibitors do not impact cell viability in response to MRV infection, but apoptosis inhibitors increase cell viability, suggesting that MRV-induces apoptotic cell death in ER+ breast cancer cells. Previous work has shown that the NFκB pathway may be activated or inhibited by MRV. Our cell viability studies in MCF7 cells with the IKK inhibitor BAY-11-7082 indicate that several MRV strains likely activate the NFκB pathway to potentially enhance viral replication. Conclusions: Future studies are aimed at further defining mechanisms of MRV-induced cell death in ER+ breast cancer. We anticipate future studies will lead to an improved MRV virotherapeutic that may be combined with clinically relevant treatments, such as CDK4/6 inhibitors and anti-estrogens, to prevent and treat therapy-resistant ER+ breast cancer.

NFKBIA deletions reshape the epigenome antithetical to the IDH mutation and indication of prognosis in diffuse gliomas.

2083 Background: Genetic alterations help predict the clinical behavior of diffuse gliomas, but some variability remains uncorrelated. The NFKBIA gene at 14q13.2 encodes IkBa, a protein that regulates the activity of NF-kB in the cytoplasm. Evidence that, in chromatin, nuclear NFKBIA dynamically interacts with histones H2A and H4 to regulate polycomb-dependent transcriptional repression and, thus, stem cell maturation and lineage specification and our characterization of NFKBIA as a tumor suppressor in glioblastoma prompted our investigation of the relationship of NFKBIA deletions to other genetic markers, alterations in the methylome, and the clinical course of gliomas. Methods: We analyzed the genetic profiles of 2,343 WHO grade 2 to 4 diffuse gliomas in multiple clinically well-characterized patient populations, including national randomized phase III consortium trial RTOG 9802, to determine whether incorporation of NFKBIA deletions could enhance the prognostic value of current molecular descriptions. Results: We demonstrate that haploinsufficient deletions of NFKBIA display distinct patterns of occurrence in relation to other genetic markers, including driver mutations IDH, TERT, ATRX, and PTEN, 1p19q codeletion, and CDKN2A/B deletion, and are disproportionally present at recurrence. NFKBIA haploinsufficiency is associated with unfavorable patient outcomes, independent of genetic and clinicopathologic predictors. NFKBIA deletions reshape the DNA and histone methylome antipodal to the IDH mutation. The methylome changes facilitated through NFKBIA deletions engender a transcriptome landscape partly reminiscent of H3K27M mutant pediatric gliomas and are highly enriched for neural stem cell and neurogenesis genes bearing repressive H3K27 marks. Our findings imply that the NFKBIA deletion in adult gliomas and the H3K27M mutation in pediatric diffuse midline gliomas define aggressive disease subtypes that may share an epigenetic state reflecting a common PRC2 loss-of-function phenotype antipodal to the quasi PRC2 gain-of-function phenotype of IDH mutant gliomas and their favorable clinical course. Lower-grade gliomas harboring NFKBIA deletions behave much like high-grade gliomas. In IDH mutant gliomas, NFKBIA deletions are common in tumors with a clinical course similar to that of IDH wildtype tumors. A sparse, easily interpretable, and externally validated nomogram model for estimating individual patient survival in IDH mutant gliomas, incorporating NFKBIA deletion and current best-established genetic and clinicopathologic factors, confirms that NFKBIA deletions predict comparatively brief survival. Conclusions: NFKBIA haploinsufficiency aligns with distinct epigenome changes, portends a poor prognosis, and should be incorporated into clinical models predicting the disease fate of diffuse gliomas.

Effect of phospholipid curcumin Meriva on liver histology and kidney disease in nonalcoholic steatohepatitis: A randomized, double-blind, placebo-controlled trial.

NASH confers an increased liver-related and kidney morbidity. Phospholipid curcumin (Meriva) is a phospholipid formulation with ameliorated systemic curcumin absorption and delivery. We assessed the safety and efficacy of Meriva in NASH. In this double-blind trial, 52 patients with biopsy-proven NASH (71% with stage ≥F2 fibrosis, 58% with stage A2-G2/A2-G3a chronic kidney disease) were randomized 1:1 to receive Meriva 2g/d or placebo for 72 weeks. The primary endpoint was NASH resolution with no worsening of fibrosis. The secondary endpoints included a ≥1 stage liver fibrosis improvement with no NASH worsening; regression of significant (ie, stage ≥F2) fibrosis and CKD; and improvement in renal, glucose, lipid, and inflammatory parameters. We also explored the treatment effect on hepatic activation of NF-kB, a key proinflammatory transcription factor and a major target of curcumin. Fifty-one patients (26 on Meriva and 25 on placebo) completed the trial. Sixteen (62%) patients on Meriva versus 3 (12%) patients on placebo had NASH resolution (RR = 5.33 [95% CI = 1.76-12.13]; p = 0.003). Thirteen (50%) patients on Meriva versus 2 (8%) patients on placebo had ≥1 stage fibrosis improvement (RR = 6.50 [1.63-21.20]; p = 0.008). Eleven (42%) patients on Meriva versus 0 (0%) on placebo had regression of significant liver fibrosis (RR = 18.01 [1.43-36.07]; p = 0.02). Hepatic NF-kB inhibition predicted NASH resolution (AUC = 0.90, 95% CI = 0.84-0.95) and fibrosis improvement (AUC = 0.89, 95% CI = 0.82-0.96). Thirteen (50%) patients on Meriva versus 0 (0%) on placebo had chronic kidney disease regression (RR = 10.71 [1.94-17.99)]; p = 0.004). Compared with placebo, Meriva improved eGFR (difference in adjusted eGFR change: +3.59 [2.96-4.11] mL/min/1.73m 2 /y, p = 0.009), fasting glucose(-17mg/dL; 95% CI = -22, -12), HbA1c (-0.62%; 95% CI = -0.87%, -0.37%), LDL-C (-39mg/dL; 95% CI = -45, -33), triglycerides (-36mg/dL, 95% CI = -46, -26), HDL-C (+10mg/dL; 95% CI = +8, +11), and inflammatory markers. Adverse events were rare, mild, and evenly distributed. In patients with NASH, Meriva administration for 72 weeks was safe, well-tolerated, and improved liver histology, possibly through NF-kB inhibition, kidney disease, and metabolic profile.

Daidzein alleviates isoproterenol-induced cardiac injury in rats via NF-kB pathway inhibition

The current study set out to explore the anti-inflammatory effects of Daidzein (Dz) on the isoproterenol (Iso)-induced myocardial infarction (MI) in male Wistar rats. Iso(80 mg/kg) was injected subcutaneously into rats to cause MI for two days at intervals of 24 hours. ELISA kits were used to measure the amounts of TNF-α & IL-6 in cardiac tissues. The quantity of NF-kB in the cytosol and nuclei was measured using a western blot approach. H&E staining was used to identify histological cardiac changes. In rats with Iso-induced myocardial infarction, Dz therapy shrunk the size of the infarct and reduced histological anomalies in the myocardium. Dz significantly lowered cardiac pro-inflammatory cytokine levels and prevented NF-kB nuclear translocation in MI-damaged rats, substantially reversing Iso-induced damage. This research shows that daidzein treatment in vivo reduces Iso-induced myocardial damage by preventing NF-kB activation, which may therefore reduce the release of inflammatory cytokines.

Synergistic cytotoxicity effect by combination of N-hexane fraction of the herbs (Peperomia pellucida) with doxorubicin against breast cancer cells (MCF-7)

Doxorubicin is the first-line chemotherapy agent for breast cancer treatment. However, it's reported to cause cardiotoxicity and induce chemoresistance in breast cancer cells. Doxorubicin is able to induce Pgp (P-glycoprotein) overexpression through NF-kB activation resulting in efflux of chemotherapeutic agents in MCF-7 cells. One of the approaches being developed for cancer treatment is combination therapy by combining two or more anticancer agents that may reduce side effects and provide better effects than single therapy. Peperomia pellucida is reported to have anticancer activity and is widely used as a traditional medicine. This study aimed to investigate the cytotoxic activity of the n-hexane fraction of Peperomia pellucida (NFP) and its combination with doxorubicin (D) against MCF-7 cells. The selectivity of NFP was evaluated using vero cells as a comparator. The phytochemical properties of NFP were evaluated using TLC and LC-MS/MS methods. The cytotoxic activity was determined using the MTT assay. Combination Index (CI) calculations were performed to determine the effect of the combination. This study found that NFP contained about 30 identified compounds and had an IC50 of 96.68 ppm, while doxorubicin had an IC50 of 2.10 ppm. The combination of 24.2 ppm (1/4 IC50) of NFP with 0.26 ppm (1/8 IC50) of doxorubicin had a synergistic cytotoxic effect (CI=0.6) against MCF-7 cells. NFP was selective for MCF-7 cells with a selectivity index of 21.83. The results suggest that NFP may have the potential to enhance the therapeutic efficacy of doxorubicin in breast cancer treatment.

Unraveling the Intricacies of OPG/RANKL/RANK Biology and Its Implications in Neurological Disorders-A Comprehensive Literature Review.

The OPG/RANKL/RANK framework, along with its specific receptors, plays a crucial role in bone remodeling and the functioning of the central nervous system (CNS) and associated disorders. Recent research and investigations provide evidence that the components of osteoprotegerin (OPG), receptor activator of NF-kB ligand (RANKL), and receptor activator of NF-kB (RANK) are expressed in the CNS. The CNS structure encompasses cells involved in neuroinflammation, including local macrophages, inflammatory cells, and microglia that cross the blood-brain barrier. The OPG/RANKL/RANK trio modulates the neuroinflammatory response based on the molecular context. The levels of OPG/RANKL/RANK components can serve as biomarkers in the blood and cerebrospinal fluid. They act as neuroprotectants following brain injuries and also participate in the regulation of body weight, internal body temperature, brain ischemia, autoimmune encephalopathy, and energy metabolism. Although the OPG/RANKL/RANK system is primarily known for its role in bone remodeling, further exploring deeper into its multifunctional nature can uncover new functions and novel drug targets for diseases not previously associated with OPG/RANKL/RANK signaling.

Targeted Therapies in the Treatment of Mantle Cell Lymphoma.

Mantle cell lymphoma (MCL) is a rare, heterogeneous B-cell non-Hodgkin's lymphoma. The standard front-line treatment utilizes chemotherapy, often followed by consolidation with an autologous hematopoietic cell transplant; however, in most patients, the lymphoma will recur and require subsequent treatments. Additionally, mantle cell lymphoma primarily affects older patients and is frequently chemotherapy-resistant, which has further fostered the necessity for new, chemotherapy-free treatment options. In the past decade, targeted therapies in mantle cell lymphoma have been practice-changing as the treatment paradigm shifts further away from relying primarily on cytotoxic agents. Here, we will review the pathophysiology of mantle cell lymphoma and discuss the emergence of targeted, chemotherapy-free treatments aimed at disrupting the abnormal biology driving its lymphomagenesis. Treatments targeting the constitutive activation of NF-kB, Bruton's Tyrosine Kinase signaling, and anti-apoptosis will be the primary focus as we discuss their clinical data and toxicities. Our review will also focus primarily on the emergence and use of targeted therapies in the relapsed/refractory setting but will also discuss the emergence of their use in front-line therapy and in combination with other agents.

Bioactives of Melipona rufiventris Propolis: Exploring its Antimicrobial, Anti-Inflammatory, and Antioxidant Activities.

This study explores the potential of propolis, a resinous substance produced by bees, from Melipona rufiventris species. With its composition encompassing resin, wax, pollen, and soil, propolis holds historical significance in traditional medicine within tropical regions. This research is driven by the scarcity of information surrounding M. rufiventris propolis, prompting an investigation into its chemical constituents, in vivo toxicity, and antimicrobial, antioxidant, and anti-inflammatory properties. This exploration could potentially uncover novel applications for this natural product, bolstering both meliponiculture practices and the preservation of native bee populations. The propolis was sampled in Cabo Verde-MG and underwent ethanolic extraction to yield an extract (EEP) for analysis. Chemical assessments (Folin-Ciocalteau, and UHPLC-HRMS) revealed the presence of polyphenols, including flavonoids. The EEP demonstrated higher antimicrobial activity against Gram-positive bacteria and exhibited efficacy against multiresistant strains isolated from complex wounds. Synergistic interactions with commercial antibiotics were also observed. Furthermore, anti-inflammatory evaluations showcased the EEP's potential in reducing NF-kB activation and TNF-α release at non-toxic concentrations. Despite these promising biological activities, the EEP exhibited no antiproliferative effects and demonstrated safety in both the MTS assay and the G. mellonella model. Collectively, these findings highlight the M. rufiventris propolis extract as a valuable reservoir of bioactive compounds with multifaceted potential.

Molecular Mechanisms and Therapeutic Potential of Gabapentin with a Focus on Topical Formulations to Treat Ocular Surface Diseases.

Gabapentin (GBP) was originally developed as a potential agonist for Gamma-Amino-Butyric-Acid (GABA) receptors, aiming to inhibit the activation of pain-signaling neurons. Contrary to initial expectations, it does not bind to GABA receptors. Instead, it exhibits several distinct pharmacological activities, including: (1) binding to the alpha-2-delta protein subunit of voltage-gated calcium channels in the central nervous system, thereby blocking the excitatory influx of calcium; (2) reducing the expression and phosphorylation of CaMKII via modulation of ERK1/2 phosphorylation; (3) inhibiting glutamate release and interfering with the activation of NMDA receptors; (4) enhancing GABA synthesis; (5) increasing cell-surface expression of δGABA_A receptors, contributing to its antinociceptive, anticonvulsant, and anxiolytic-like effects. Additionally, GBP displays (6) inhibition of NF-kB activation and subsequent production of inflammatory cytokines, and (7) stimulation of the purinergic adenosine A1 receptor, which supports its anti-inflammatory and wound-healing properties. Initially approved for treating seizures and postherpetic neuralgia, GBP is now broadly used for various conditions, including psychiatric disorders, acute and chronic neuropathic pain, and sleep disturbances. Recently, as an eye drop formulation, it has also been explored as a therapeutic option for ocular surface discomfort in conditions such as dry eye, neurotrophic keratitis, corneal ulcers, and neuropathic ocular pain. This review aims to summarize the evidence supporting the molecular effects of GBP, with a special emphasis on its applications in ocular surface diseases.

Clinicoradiological Parameters and Biochemical and Molecular Alterations Predicting Remission and Recurrence After Surgical Treatment of Corticotroph Adenomas—Cushing Disease

ObjectivesEndonasal endoscopic transsphenoidal surgery (TSS) and resection of pituitary adenomas is considered to be the gold standard treatment for Cushing’s Disease (CD). Even with various recent advances in management, disease persistence and recurrences are common in these patients. The remission rate in the global population after surgery has been reported to vary widely from 64 to 93%. This study aims to determine various clinical, biochemical, radiological and histological factors that correlate with persistence and recurrence in patients with CD. This study also aims to understand the clinicopathological significance of EGFR-MAPK, NF-kB and SHH pathway activation and to study the protein expression levels of activation markers of these pathways viz. c-Fos , c-Jun, GLI-1 , pMEK, NR4A1 and p44 in functioning corticotroph pituitary adenomas. MethodsFrom January 2009 to September 2022, the clinical data of 167 patients who underwent surgical treatment (n=174 surgeries) for Cushing’s disease was ambispectively analysed with a median follow up of 8.1 years (1-13.29 years). The preoperative clinical, biochemical, radiological features, operative findings, postoperative clinical and biochemical data, histopathological and molecular profile, were retrieved from electronic records. The patients were followed up to assess the remission status. ResultsAmong the 174 surgeries performed, 140 were primary surgeries, 22 were revision surgeries, 24 surgeries were for paediatric patients and 12 surgeries were performed in patients with Nelson syndrome. In the primary surgery cohort, 74.3% were females and the average age was 28.73 years (SD 10.15). 75% of the patients experienced remission after primary surgery compared to 47.4% in revision surgery. The remission rate in paediatric patients was 55.5%. Post-operative day 1 plasma cortisol (p<0.001; AUC 0.8894 [0.8087-0.9701]) and ACTH (p<0.001; AUC 0.9 [0.7386-1]) levels were seen to be strong independent predictors of remission in the primary surgery cohort. Remission rates after Endoscopic TSS were more than that after microscopic TSS, in patients undergoing primary surgery (81.08% vs 57.14%; p=0.008). Presence of adenoma on histopathological examination (HPE) was also a strong predictor of disease remission (p=0.020). On categorising based on surgical approach and HPE, microscopically operated patients without histopathological evidence of adenoma had significantly higher odds of non-remission (OR 38.1 95% CI 4.2-348.3) compared to endoscopically operated patients with adenoma on HPE. Lower immune reactivity score (IRS) of NR4A1 was found to correlate with higher remission rates (p=0.074). However, none of the molecular markers studied viz. c-Fos , c-Jun, GLI-1, pMEK and p44 showed a significant correlation with the preoperative cortisol values. ConclusionRemission rate after primary surgery is higher as compared to that after revision surgery, and that in paediatric patients is lower as compared to adults. Post-operative day 1 plasma cortisol and ACTH levels are strong independent predictors of remission in the primary surgery cohort. Endoscopic approach with histopathological evidence of adenoma is associated with a higher remission rate and thus endoscopy should be the approach of choice in these patients with the goal of identification of an adenoma on histopathological analysis.

Molecular mechanism of CD14+ human monocyte/macrophage activation by Ebola virus protein VP40

Abstract Impaired functions of many innate immune cells have been implicated in Ebola virus disease (EVD) pathogenesis. Previously, we reported that CD14+ accessory cells are required for the optimal Ebola virus protein VP40-induced activation of human natural killer (NK) cells. In the current study, we explored the molecular mechanism(s) of VP40-induced activation of CD14+ accessory cells and their role in triggering NK cell activation. Human PBMCs, purified CD14+ myeloid cells or CD14+ cell and CD56+ NK cell co-cultures were incubated with VP40, mCherry-VP40 fusion protein or virus-like particles (VLPs) containing it and binding, internalization, and distribution of VP40 analyzed by flow and confocal microscopy. Molecular pathway(s) triggered in CD14+ cells and the resulting gene expression changes were analyzed by flow, western blotting, luminex and RNA-Seq. Ebola VP40/VLPs bound to and were internalized by classical, intermediate and non-classical monocytes but to varying degrees. VP40 triggered activation of NF-kB and inflammatory pathways in CD14+ cells resulting in their enhanced secretion of multiple cytokines and factors. IL-12 or IL-18 blockade partially abrogated VP40-induced, CD14+ cell dependent activation of NK cells. Taken together, our data suggests that activation of CD14+ cell by Ebola VP40/VLPs can rapidly trigger NK cells that, in turn, may eliminate Ebola virus infected host cells, findings that may help in designing better anti-Ebola countermeasures.

Siglec-E is a novel co-inhibitory receptor in the modulation of innate activation after transplantation

Abstract Early after transplantation, inflammation and tissue injury are associated with significant activation of the innate immune cells, such as DCs. This activation drives adaptive immune responses, triggering acute rejection while hindering the development of transplant tolerance. Current immunosuppressive drugs primarily target T cells, while innate immunity is not efficiently inhibited in transplantation. The sialic acid–binding immunoglobulin-like lectin E (Siglec-E, or SigE) is an innate receptor that binds to ligands carrying sialic acids and inhibits TLRs-triggered inflammatory responses. Here, we demonstrate the role of SigE in experimental solid organ transplantation mouse models. Upon heart transplantation in mice, SigE is upregulated in graft-infiltrating APCs, including DCs. In humans, Siglec-7 and -9, the human homologs of SigE, are also upregulated in human heart biopsies following transplantation. Genetic ablation of recipient SigE leads to accelerated rejection of heart, kidney, and skin allografts. Mechanistically, SigE-deficient DCs are more susceptible to activation by DAMPS with an enhanced NF-kB activation and production of TNF-a. Moreover, DCs lacking SigE have a higher ability to induce allo-T cell responses in vitro and in vivo. In contrast, overexpression of SigE on DCs decreases their activation by DAMPs and reduces their T cell allostimulatory capacity. Thus, Siglec-E is a major inhibitory receptor in transplantation by controlling APC activation.

PM2.5 toxicity in blood impairs cardiac redox balance and promotes mitochondrial dysfunction in rat heart that further aggravates ischemia reperfusion injury by modulating PI3K/AKT/mTOR/NF-kB signaling axis.

According to the pathophysiological mechanisms linking particulate matter (PM2.5) exposure and cardiovascular diseases, PM2.5 may directly translocate into the blood stream and remote target organs and thereby induce cardiovascular effects. The toxicity of PM2.5 is known to induce oxidative stress in pulmonary tissue, but its impact on the redox state in heart (distant organ) is unknown and how it modulates the cardiac response to ischemia reperfusion (IR) remains unclear. In the present study, we evaluated the toxic effect of PM2.5 on cardiac physiology in the presence and absence of IR after introducing PM2.5 into the blood. Female Wistar rats were injected with diesel particulate matter (DPM) via i.p & i.v routes at a concentration of 10 µg/ml. The toxic impact of PM2.5 not only adversely affects the cardiac ultra-structure (leading to nuclear infiltration, edema, irregularities in heart muscle and nuclear infiltration), but also altered the cellular redox balance, elevated inflammation and promoted the upregulation of proapoptotic mediator genes at the basal level of myocardium. The results showed alterations in cardiac ultrastructure, elevated oxidative stress and significant redox imbalance, increased inflammation and proapoptotic mediators at the basal level of myocardium. Moreover, the cardioprotective pro survival signaling axis was declined along with an increased NF-kB activation at the basal level. IR inflicted further injury with deterioration of cardiac hemodynamic indices (Heart rate [HR], Left ventricular developed pressure [LVDP], Left ventricular end-diastolic pressure [LVEDP] and rate pressure product [RPP]) along with prominent inactivation of signaling pathways. Furthermore, the levels of GSH/GSSG, NADH/NAD, NADPH/NADP were significantly low along with increased lipid peroxidation in mitochondria of PM2.5 treated IR rat hearts. This observation was supported by downregulation of glutaredoxin and peroxiredoxin genes in the myocardium. Similarly the presence of oxidative stress inducing metals was found at a higher concentration in cardiac mitochondria. Thus, the toxic impact of PM2.5 in heart augment the IR associated pathological changes by altering the physiological response, initiating cellular metabolic alterations in mitochondria and modifying the signaling molecules.

Study of individual domains contributing to MALT1 dimerization in BCL10-independent and dependent assembly

The CARMA-BCL10-MALT1 (CBM) signalosome functions as a pivotal supramolecular module, integrating diverse receptor-induced signaling pathways to regulate BCL10-dependent NF-kB activation in innate and adaptive immunity. Conversely, the API2-MALT1 fusion protein in t(11; 18)(q21; q21) MALT lymphoma constitutively induces BCL10-independent NF-kB activation. MALT1 dimer formation is indispensable for the requisite proteolytic activity and is critical for NF-kB activation regulation in both scenarios. However, the molecular assembly of MALT1 individual domains in CBM activation remains elusive. Here we report the crystal structure of the MALT1 death domain (DD) at a resolution of 2.1 Å, incorporating reconstructed residues in previously disordered loops 1 and 2. Additionally, we observe a conformational regulation element (CRE) regulating stem-helix formation in NLRPs pyrin (PYD) within the MALT1 DD structure. The structure reveals a stem-helix-mediated dimer further corroborated in solution. To elucidate how the BCL10 filament facilitates MALT1 dimerization, we reconstitute a BCL10-CARD-MALT1-DD-IG1-IG2 complex model. We propose a N+7 rule for BCL10-dependent MALT1 dimerization via the IG1-IG2 domain and for MALT1-dependent cleavage in trans. Biochemical data further indicates concentration-dependent dimerization of the MALT1 IG1-IG2 domain, facilitating MALT1 dimerization in BCL10-independent manner. Our findings provide a structural and biochemical foundation for understanding MALT1 dimeric mechanisms, shedding light on potential BCL10-independent MALT1 dimer formation and high-order BCL10-MALT1 assembly.