Early-life bisphenol A exposure causes neuronal pyroptosis in juvenile and adult male rats through the NF-κB/IL-1ß/NLRP3/caspase-1 signaling pathway: exploration of age and dose as effective covariates using an in vivo and in silico modeling approach.

Bisphenol A (BPA), a common endocrine-disrupting chemical, is found in a wide range of home plastics. Early-life BPA exposure has been linked to neurodevelopmental disorders; however, the link between neuroinflammation, pyroptosis, and the development of psychiatric disorders is rarely studied. The current study attempted to investigate the toxic effect of BPA on inflammatory and microglial activation markers, as well as behavioral responses, in the brains of male rats in a dose- and age-dependent manner. Early BPA exposure began on postnatal day (PND) 18 at dosages of 50 and 125 mg/kg/day. We started with a battery of behavioral activities, including open field, elevated plus- and Y-maze tests, performed on young PND 60 rats and adult PND 95 rats. BPA causes anxiogenic-related behaviors, as well as cognitive and memory deficits. The in vivo and in silico analyses revealed for the first time that BPA is a substantial activator of nuclear factor kappa B (NF-κB), interleukin (IL)-1ß, -2, -12, cyclooxygenase-2, and the NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome, with higher beclin-1 and LC3B levels in BPA rats' PFC and hippocampus. Furthermore, BPA increased the co-localization of caspase-1 immunoreactive neurons, as well as unique neurodegenerative histopathological hallmarks. In conclusion, our results support the hypothesis that neuroinflammation and microglial activation are involved with changes in the brain after postnatal BPA exposure and that these alterations may be linked to the development of psychiatric conditions later in life. Collectively, our findings indicate that BPA triggers anxiety-like behaviors and pyroptotic death of nerve cells via the NF-κB/IL-1ß/NLRP3/Caspase-1 pathway.

Reactive Oxygen Species Modulate Th17/Treg Balance in Chlamydia psittaci Pneumonia via NLRP3/IL-1ß/Caspase-1 Pathway Differentiation.

Chlamydia psittaci pneumonia (CPP) is a lung disease caused by the infection with the Chla-mydia psittaci bacterium, which can lead to severe acute respiratory distress syndrome and systemic symptoms. This study explored the specific mechanisms underlying the impact of reactive oxygen species (ROS) on the Th17/Treg balance in CPP. The levels of ROS and the differentiation ratio of Th17/Treg in the peripheral blood of healthy individuals and CPP patients were measured using ELISA and flow cytometry, respectively. The association between the ROS levels and Th17/Treg was assessed using Pearson correlation analysis. The ROS levels and the Th17/Treg ratio were measured in CD4+ T cells following H2O2 treatment and NLRP3 inhibition. The effects of H2O2 treatment and NLRP3 inhibition on the NLRP3/IL-1ß/caspase-1 pathway were observed using immunoblotting. Compared to the healthy group, the CPP group exhibited increased levels of ROS in the peripheral blood, an elevated ratio of Th17 differentiation, and a decreased ratio of Treg differentiation. ROS levels were positively correlated with the Th17 cell proportion but negatively correlated with the Treg cell proportion. The ROS levels and NLRP3/IL-1ß/caspase-1 expression were up-regulated in CD4+ T cells after H2O2 treatment. Furthermore, there was an increase in Th17 differentiation and a decrease in Treg differentiation. Conversely, the NLRP3/IL-1ß/caspase-1 pathway inhibition reversed the effects of H2O2 treatment, with no significant change in the ROS levels. ROS regulates the Th17/Treg balance in CPP, possibly through the NLRP3/IL-1ß/caspase-1 pathway. This study provides a new perspective on the development of immunotherapy for CPP.

[Mongolian medicine Naru-3 reduces neuroinflammation in maintenance stage of neuropathic pain by inhibiting astrocyte activation].

Neuropathic pain(NP) is difficult to be treated since it has similar phenotypes but different pathogenesis in different pathological stages. Targeted intervention of the core regulatory elements at different pathological stages of NP has become a new direction of drug research and development in recent years and provides the possibility for the treatment of NP. The Mongolian medicine Naru-3(NR-3) is effective in the treatment of sciatica and trigeminal neuralgia, the mechanisms of which remain unknown. On the basis of the previous study of the priming stage, this study established the mouse model of spinal nerve ligation(SNL) and measured the changes of pain thresholds by behavioral tests. The network analysis, Western blot, immunofluorescence assay, ELISA, and agonist/antagonist were employed to decipher the mechanism of NR-3 in the treatment of NP in the maintenance stage. The results showed that NR-3 increased the mechanical and thermal pain thresholds of SNL mice, while it had no significant effect on the basal pain threshold of normal mice. NR-3 may relieve the pain in the maintenance stage of NP by blocking the matrix metalloproteinase 2(MMP2)/interleukin-1ß(IL-1ß) pathway in the astrocytes of the dorsal root ganglion(DRG) and spinal cord. The findings have enriched the biological connotation of NR-3 in the treatment of the maintenance stage of NP and provide reference for the rational use of this medicine in clinical practice.

Diminished vasculogenesis under inflammatory conditions is mediated by Activin A.

Severe inflammatory stress often leads to vessel rarefaction and fibrosis, resulting in limited tissue recovery. However, signaling pathways mediating these processes are not completely understood. Patients with ischemic and inflammatory conditions have increased systemic Activin A level, which frequently correlates with the severity of pathology. Yet, Activin A's contribution to disease progression, specifically to vascular homeostasis and remodeling, is not well defined. This study investigated vasculogenesis in an inflammatory environment with an emphasis on Activin A's role. Exposure of endothelial cells (EC) and perivascular cells (adipose stromal cells, ASC) to inflammatory stimuli (represented by blood mononuclear cells from healthy donors activated with lipopolysaccharide, aPBMC) dramatically decreased EC tubulogenesis or caused vessel rarefaction compared to control co-cultures, concurrent with increased Activin A secretion. Both EC and ASC upregulated Inhibin Ba mRNA and Activin A secretion in response to aPBMC or their secretome. We identified TNFα (in EC) and IL-1ß (in EC and ASC) as the exclusive inflammatory factors, present in aPBMC secretome, responsible for induction of Activin A. Similar to ASC, brain and placental pericytes upregulated Activin A in response to aPBMC and IL-1ß, but not TNFα. Both these cytokines individually diminished EC tubulogenesis. Blocking Activin A with neutralizing IgG mitigated detrimental effects of aPBMC or TNFα/IL-1ß on tubulogenesis in vitro and vessel formation in vivo. This study delineates the signaling pathway through which inflammatory cells have a detrimental effect on vessel formation and homeostasis, and highlights the central role of Activin A in this process. Transitory interference with Activin A during early phases of inflammatory or ischemic insult, with neutralizing antibodies or scavengers, may benefit vasculature preservation and overall tissue recovery.

TET2 deficiency promotes anxiety and depression-like behaviors by activating NLRP3/IL-1ß pathway in microglia of allergic rhinitis mice.

BACKGROUND: Anxiety and depression-like behaviors in allergic rhinitis (AR) are attracting attention, while the precise mechanism has not been clearly elucidated. Recent evidence shows that neuroinflammation in anterior cingulate cortex (ACC) may be the core of these neuropsychiatric symptoms in AR. Here, we investigated the molecular link between the anxiety and depression-like behaviors and neuroinflammation in ACC. METHODS: Mice were sensitized and challenged with ovalbumin (OVA) to induce AR. Nasal inflammation levels were assessed by H&E staining and PAS staining. Anxiety and depression-like behaviors were evaluated by behavioral experiments including open field test, forced swimming test, and sucrose preference test. Neuronal impairment was characterized via Nissl staining and 18FDG-PET. The role of ten-eleven translocation 2 (TET2) in AR-related anxiety and depression was assessed by Tet2-/- mice. In addition, the murine BV2 microglial cell line was utilized to explore the molecular mechanisms by which TET2 mediates neuroinflammation. The levels of TET2, NLRP3 and their downstream molecules were detected by immunohistochemistry, Western blot, Dot blot and ELISA. The effects of metformin on depression-like behaviors in AR mice were also evaluated. RESULTS: AR mice showed significant anxiety and depression-like behaviors, which associated with the activation of ACC. Loss of TET2 activated the NLRP3/IL-1ß pathway of microglia in AR mice, further accelerating the anxiety and depression-like behaviors. In addition, knockdown of TET2 activated the NLRP3/IL-1ß pathway in BV2 cells. Metformin improved the neuropsychiatric symptoms of AR mice by reducing the activation of NLRP3/IL-1ß pathway after upregulating TET2. CONCLUSION: TET2 deficiency activates the NLRP3/IL-1ß pathway of microglia in the ACC, promoting the pathological process of anxiety and depression-like behavior in AR. Metformin could be effective in treating neuroinflammation by regulating microglia via TET2 up-regulation, indicating that metformin is a potential way to treat anxiety and depression-like behaviors in AR.

BC-1215 inhibits ATP-induced IL-1ß secretion via the FBXL2-mediated ubiquitination and degradation of not only NLRP3, but also pro-IL-1ß in LPS-primed THP-1 cells.

BC-1215, bis-pyridinyl benzyl ethanediamine, is an inhibitor of F-box only protein 3 (FBXO3) and exerts anti-inflammatory effects. BC-1215 inhibits interactions between FBXO3-F-box and the leucine rich repeat protein 2 (FBXL2), leading to the upregulation of FBXL2 expression, FBXL2-mediated ubiquitination and the degradation of tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6) or NOD-, LRR- and the pyrin domain-containing protein 3 (NLRP3), which subsequently results in the down-regulation of inflammatory cytokine production. In the current study, we investigated the issue of whether or how BC-1215 suppresses the ATP-induced secretion of IL-1ß in LPS-primed human macrophage-like cells, THP-1 cells. Our result show that pre-treatment with BC-1215 attenuated the ATP-induced secretion of IL-1ß in LPS-primed THP-1 cells. Treatment of the LPS-primed THP-1 cells with BC-1215 resulted in a decrease in the level of NLRP3 and pro-IL-1ß at the protein level, but not at the mRNA level. In addition, treatment with MG-132, but not leupeptin, inhibited the BC-1215-induced degradation of NLRP3 and pro-IL-1ß proteins, and restored their levels, suggesting that BC-1215 decreases the stability of NLRP3 and pro-IL-1ß at the protein level via proteasome-dependent degradation. Our results also show that FBXL2, which is increased by BC-1215, bound to and ubiquitinated NLRP3 and pro-IL-1ß, but not pro-caspase-1. These collective results indicate that treatment with BC-1215, an inhibitor of FBXO3, inhibits ATP-induced IL-1ß secretion via the FBXL2-mediated ubiquitination and degradation of pro-IL-1ß as well as NLRP3 in LPS-primed THP-1 cells, suggesting that FBXO3 is a potential therapeutic target for developing agents against inflammatory diseases.

The effect of secukinumab treatment for psoriasis on serum cytokines and correlation with disease severity.

OBJECTIVE: To investigate the effects of secukinumab treatment for psoriasis on different functional cytokines and inflammatory mediators in patients' serum METHODS: Enzyme-linked immunosorbent assay was used to detect interleukin (IL)-1ß and IL-1RA associated with intrinsic immunity; IL-6, IL-18, and growth regulated oncogene alpha (GROα) associated with neutrophils; IL-12, tumour necrosis factor (TNF)-α, and interferon (IFN)-γ associated with Th1; IL-23, IL-17A, and IL-22 associated with Th17; Thymus activation regulated chemokine (TARC), IL-13, and defensin beta 2 (DEFB2) associated with Th2; Vascular endothelial growth factor (VEGF)-A and IL-10 associated with angiogenesis; and IFN-γ associated with sepsis in the peripheral blood of 12 patients with common psoriasis treated with secukinumab and 15 healthy controls. IL-23, IL-17A, IL-22 associated with Th17; TARC, IL-13, DEFB2 associated with Th2; VEGF-A, IL-10 associated with angiogenesis and procalcitonin (PCT) associated with sepsis. The differences in expression of the above cytokines before and after treatment and the correlation with psoriasis disease severityï¼»Psoriasis Area Severity Index(PASI) scoreï¼½, age, and disease duration were analyzed. RESULTS: The mean PASI score of the enrolled patients with moderate to severe psoriasis was 21.6 ± 11.0 before treatment and decreased to below 1 after treatment. Serum IL-6; IL-18, GROα, IFN-γ, TNF-α, VEGF-A, and IL-17A were significantly higher than normal. And IL-17A and IFN-γ were positively correlated with disease duration and age, and IL-18 was positively correlated with PASI score. The expression levels of IL-6, GROα, VEGF-A, IFN-γ, TNF-α, IL-17A and IL-23 were significantly lower after secukinumab treatment compared with those before treatment, but the expression levels of IFN-γ, VEGF-A, TARC, IL-13, and DEFB2 were still significantly higher than those of normal subjects after treatment CONCLUSIONS: secukinumab clears skin lesions by antagonizing IL-17A and simultaneously decreasing the expression levels of IL-6, GRO α, VEGF-A, IFN-γ, TNF-α, IL-17A, and IL-23.

NLRP3 Inflammasome Involvement in Heart, Liver, and Lung Diseases-A Lesson from Cytokine Storm Syndrome.

Inflammation and inflammasomes have been proposed as important regulators of the host-microorganism interaction, playing a key role in morbidity and mortality due to the coronavirus disease 2019 (COVID-19) in subjects with chronic conditions and compromised immune system. The inflammasome consists of a multiprotein complex that finely regulates the activation of caspase-1 and the production and secretion of potent pro-inflammatory cytokines such as IL-1ß and IL-18. The pyrin containing NOD (nucleotide-binding oligomerization domain) like receptor (NLRP) is a family of intracellular receptors, sensing patterns associated to pathogens or danger signals and NLRP3 inflammasome is the most deeply analyzed for its involvement in the innate and adaptive immune system as well as its contribution to several autoinflammatory and autoimmune diseases. It is highly expressed in leukocytes and up-regulated in sentinel cells upon inflammatory stimuli. NLRP3 expression has also been reported in B and T lymphocytes, in epithelial cells of oral and genital mucosa, in specific parenchymal cells as cardiomyocytes, and keratinocytes, and chondrocytes. It is well known that a dysregulated activation of the inflammasome is involved in the pathogenesis of different disorders that share the common red line of inflammation in their pathogenetic fingerprint. Here, we review the potential roles of the NLRP3 inflammasome in cardiovascular events, liver damage, pulmonary diseases, and in that wide range of systemic inflammatory syndromes named as a cytokine storm.

[Mechanism of Mongolian drug Naru-3 in initiation of neuroinflammation of neuropathic pain from MMP9/IL-1ß signaling pathway].

Neuropathic pain(NP) has similar phenotypes but different sequential neuroinflammatory mechanisms in the pathological process. It is of great significance to inhibit the initiation of neuroinflammation, which has become a new direction of NP treatment and drug development in recent years. Mongolian drug Naru-3 is clinically effective in the treatment of trigeminal neuralgia, sciatica, and other NPs in a short time, but its pharmacodynamic characteristics and mechanism of analgesia are still unclear. In this study, a spinal nerve ligation(SNL) model simulating clinical peripheral nerve injury was established and the efficacy and mechanism of Naru-3 in the treatment of NPs was discussed by means of behavioral detection, side effect evaluation, network analysis, and experimental verification. Pharmacodynamic results showed that Naru-3 increased the basic pain sensitivity threshold(mechanical hyperalgesia and thermal radiation hyperalgesia) in the initiation of SNL in animals and relieved spontaneous pain, however, there was no significant effect on the basic pain sensitivity threshold and motor coordination function of normal animals under physiological and pathological conditions. Meanwhile, the results of primary screening of target tissues showed that Naru-3 inhibited the second phase of injury-induced nociceptive response of formalin test in mice and reduced the expression of inflammatory factors in the spinal cord. Network analysis discovered that Naru-3 had synergy in the treatment of NP, and its mechanism was associated with core targets such as matrix metalloproteinase-9(MMP9) and interleukin-1ß(IL-1ß). The experiment further took the dorsal root ganglion(DRG) and the stage of patho-logical spinal cord as the research objects, focusing on the core targets of inducing microglial neuroinflammation. By means of Western blot, immunofluorescence, agonists, antagonists, behavior, etc., the mechanism of Naru-3 in exerting NP analgesia may be related to the negative regulation of the MMP9/IL-1ß signaling pathway-mediated microglia p38/IL-1ß inflammatory loop in the activation phase. The relevant research enriches the biological connotation of Naru-3 in the treatment of NP and provides references for clinical rational drug use.

Past Exposure to Cigarette Smoke Aggravates Cognitive Impairment in a Rat Model of Vascular Dementia via Neuroinflammation.

Smoking is a risk factor for dementia. Cognitive function can be partially restored after quitting smoking, but still lower than never smoked group. The underlying mechanisms still remain unclear. The effects of smoking cessation combined with cerebral chronic hypoperfusion (CCH) on cognitive function have never been described. Here, we established a cigarette smoking cessation model, a CCH model, and a cigarette smoking cessation plus CCH model. We investigated cognitive function in these models and the mechanisms of the neuroinflammation, nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3(NLRP3)/cysteine aspartate-specific proteinase (caspase-1)/interleukin- 1ß (IL-1ß) pathway, and eucaryotic initiation factor 2α (eIF2α) /autophagy pathway. We used morris water maze (MWM) and novel object recognition (NOR) test to evaluate cognitive function in rats. Nissl staining was performed to observe cell morphology in the hippocampal CA1 area. A neuroinflammatory marker (glial fibrillary acidic protein, GFAP) was assessed by Western blot analysis and immunohistochemistry staining. IL-1ß levels were detected by ELISA. The protein levels of NLRP3/caspase-1/ IL-1ß and eIF2α/autophagy pathway were evaluated by Western blot analysis. LC3 was assessed by immunofluorescence staining. CCH can affect cognitive function by influencing neuroinflammation, NLRP3/caspase-1/IL-1ß pathway, and eIF2α/autophagy pathway. Past exposure to cigarette smoke can also affect cognitive function by influencing neuroinflammation and NLRP3/caspase-1/IL-1ß pathway, which may be induced by smoking and may not be alleviated after smoking cessation. Past exposure to cigarette smoke does not influence autophagy, which may be increased by smoking and then decrease to normal levels after smoking cessation. Past exposure to smoking can further aggravate cognitive impairment and neuroinflammation in VaD animals: cognitive impairment induced by CCH via neuroinflammation, NLRP3/caspase-1/IL-1ß, and eIF2α/autophagy pathway and cognitive impairment induced by past exposure to cigarette smoke via neuroinflammation and NLRP3/caspase-1/IL-1ß pathway. The combined group had the worst cognitive impairment because of harmful reasons.

Knockdown of DAPK1 attenuates IL-1ß-induced extracellular matrix degradation and inflammatory response in osteoarthritis chondrocytes via regulating the p38 MAPK-signaling pathway.

OBJECTIVE: To reveal the possible effects of death-associated protein kinase 1 (DAPK1) on the progression of osteoarthritis (OA) and the potential underlying mechanism. METHODS: : The expression of DAPK1 in OA and normal samples and interleukin (IL)-1ß-stimulated chondrocytes was analyzed by quantitative real-time polymerase chain reaction and Immunoblot assay. Cell viability, proliferation, and apoptosis in DAPK1-knockdown cells stimulated with IL-1ß were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) solution, 5-ethynyl-2ß-deoxyuridine staining and flow cytometry. The chondrocyte degradation and inflammatory response in IL-1ß-induced chondrocytes were investigated by Immunoblot analysis and enzyme-linked-immunosorbent serologic assay. In addition, the effect of DAPK1 on p38 mitogen-activated protein kinase (MAPK) activation was analyzed by immunoblot assay. RESULTS: : This study revealed that DAPK1 was highly expressed in OA patients and IL-1ß-induced chondrocytes. Down-regulation of DAPK1 enhanced IL-1ß-induced chondrocyte proliferation. DAPK1 knockdown inhibited IL-1ß-induced chondrocyte degradation. In addition, DAPK1 depletion inhibited IL-1ß-induced chondrocyte inflammation. Mechanically, it was revealed that down--regulation of DAPK1 could inhibit the p38 MAPK pathway, and therefore affected progression of OA. CONCLUSION: : DAPK1 knockdown attenuates IL-1ß-induced extracellular matrix degradation and inflammatory response in OA chondrocytes by regulating the p38 MAPK pathway.

Microbe-Derived Antioxidants Reduce Lipopolysaccharide-Induced Inflammatory Responses by Activating the Nrf2 Pathway to Inhibit the ROS/NLRP3/IL-1ß Signaling Pathway.

Inflammation plays an important role in the innate immune response, yet overproduction of inflammation can lead to a variety of chronic diseases associated with the innate immune system; therefore, modulation of the excessive inflammatory response has been considered a major strategy in the treatment of inflammatory diseases. Activation of the ROS/NLRP3/IL-1ß signaling axis has been suggested to be a key initiating phase of inflammation. Our previous study found that microbe-derived antioxidants (MA) are shown to have excellent antioxidant and anti-inflammatory properties; however, the mechanism of action of MA remains unclear. The current study aims to investigate whether MA could protect cells from LPS-induced oxidative stress and inflammatory responses by modulating the Nrf2-ROS-NLRP3-IL-1ß signaling pathway. In this study, we find that MA treatment significantly alleviates LPS-induced oxidative stress and inflammatory responses in RAW264.7 cells. MA significantly reduce the accumulation of ROS in RAW264.7 cells, down-regulate the levels of pro-inflammatory factors (TNF-α and IL-6), inhibit NLRP3, ASC, caspase-1 mRNA, and protein levels, and reduce the mRNA, protein levels, and content of inflammatory factors (IL-1ß and IL-18). The protective effect of MA is significantly reduced after the siRNA knockdown of the NLRP3 gene, presumably related to the ability of MA to inhibit the ROS-NLRP3-IL-1ß signaling pathway. MA is able to reduce the accumulation of ROS and alleviate oxidative stress by increasing the content of antioxidant enzymes, such as SOD, GSH-Px, and CAT. The protective effect of MA may be due to its ability of MA to induce Nrf2 to enter the nucleus and initiate the expression of antioxidant enzymes. The antioxidant properties of MA are further enhanced in the presence of the Nrf2 activator SFN. After the siRNA knockdown of the Nrf2 gene, the antioxidant and anti-inflammatory properties of MA are significantly affected. These findings suggest that MA may inhibit the LPS-stimulated ROS/NLRP3/IL-1ß signaling axis by activating Nrf2-antioxidant signaling in RAW264.7 cells. As a result of this study, MA has been found to alleviate inflammatory responses and holds promise as a therapeutic agent for inflammation-related diseases.

A single dose of anti-IL-1ß antibodies prevents Western diet-induced immune activation during early stage collagenase-induced osteoarthritis, but does not ameliorate end-stage pathology.

OBJECTIVE: Metabolic dysfunction can cause IL-1ß mediated activation of the innate immune system, which could have important implications for the therapeutic efficacy of IL-1ß neutralizing drugs as treatment for OA in the context of metabolic syndrome (MetS). In the present study, we investigated whether early treatment with a single dose of IL-1ß blocking antibodies could prevent Western diet (WD) induced changes to systemic monocyte populations and their cytokine secretion profile and herewith modulate collagenase induced osteoarthritis (CiOA) pathology. METHODS: CiOA was induced in female C57Bl/6 mice fed either a standard diet (SD) or WD and treated with a single dose of either polyclonal anti-IL-1ß antibodies or control. Monocyte subsets and granulocytes in bone marrow and blood were analyzed with flow cytometry, and cytokine expression by bone marrow cells was analyzed using qPCR. Synovial cellularity, cartilage damage and osteophyte formation were assessed on histology. RESULTS: WD feeding of C57Bl/6 mice led to increased serum levels of low-density lipoprotein (LDL) and innate immune activation in the form of an increased number of Ly6Chigh cells in bone marrow and blood and increased cytokine expression of IL-6 and TNF-α by bone marrow cells. The increase in monocyte number and activity was ameliorated by anti-IL-1ß treatment. However, anti-IL-1ß treatment did not significantly affect synovial lining thickness, cartilage damage and ectopic bone formation during WD feeding. CONCLUSIONS: Single-dose systemic anti-IL-1ß treatment prevented WD-induced innate immune activation during early stage CiOA in C57Bl/6 mice, but did not ameliorate joint pathology.

E3 ubiquitin ligase TRIM24 deficiency promotes NLRP3/caspase-1/IL-1ß-mediated pyroptosis in endometriosis.

Endometriosis is an inflammation-dependent disease that shares similarities with malignant tumors including attachment and infiltration. Tripartite motif-containing 24 (TRIM24) has been illustrated in inflammatory responses and gynecological tumors, and Nod-like receptor protein 3 (NLRP3) inflammasome has been implicated in endometriosis. However, the involvement of TRIM24 and the role of NLRP3/caspase-1/interleukin-1ß (IL-1ß)-mediated pyroptosis in endometriosis remain obscure. In this study, we originally detected the decreased expression of TRIM24 in the ectopic endometrium of endometriosis compared with the normal endometrium. Then we measured the promoted protein expression of pyroptotic biomarkers (NLRP3, procaspase-1, caspase-1, pro-IL-1ß, and IL-1ß) using Western blot analysis and the stimulated secretion of IL-1ß and IL-18 by enzyme-linked immunosorbent assay in ectopic human endometrial stromal cells (hESC) compared with normal hESC. TRIM24-small-interfering RNA (siTRIM24) was used to silence TRIM24, whereas TRIM24-pcDNA3.1 was used for overexpressing TRIM24. The migration of hESC was determined by a Transwell migration assay. Coimmunoprecipitation and ubiquitination analyses were conducted to explore the interaction between TRIM24 and NLRP3. Subsequently, we found that TRIM24 negatively regulated NLRP3/caspase-1/IL-1ß-mediated pyroptosis and cell migration of hESC, and CY-09, the specific inhibitor of NLRP3, could reverse the promoted pyroptosis and cell migration induced by siTRIM24. Furthermore, TRIM24 interacted with NLRP3 and the upregulation of TRIM24 facilitated the ubiquitination of NLRP3 in ectopic hESC. Our findings suggest that TRIM24 may participate in the progression of endometriosis through the NLRP3/caspase-1/IL-1ß-mediated pyroptotic pathway via ubiquitination of NLRP3, which reveals the significant molecular mechanism underlying endometriosis.

Protective effects of NLRP3 inhibitor MCC950 on sepsis-induced myocardial dysfunction.

Sepsis-induced myocardial dysfunction (SIMD) leads to poor prognosis or even death in severe sepsis cases, therefore, exploring its pathogenesis and new therapeutic targets has become the focus of current research. Specifically, an SIMD rat model was constructed by cecal ligation and puncture (CLP) method. At 24 h after intraperitoneal injection of the NLRP3 selective inhibitor MCC950, the levels of serum cardiac troponin I (cTnI) and Lactate dehydrogenase (LDH) in serum were detected, and the cardiac function of rats was examined via echocardiography. In addition, the pathological changes of myocardial tissues were observed by histological method, and the expression changes of inflammatory factors were detected in the tissue and serum. At the same time, H9C2 cells were treated with lipopolysaccharide (LPS) to simulate the in vitro model, and the expressions of inflammation and pyroptosis-related factors were detected. The results manifested that in the CLP group, the levels of serum cTnI and LDH were obviously increased, the myocardial tissue structure was disordered, the cell edema was severe, and the cardiac function was markedly reduced. Meanwhile, the expressions of inflammatory factors IL-6, IL-8 and TNF-α rose remarkably. On the contrary, MCC950 effectively reversed the above situation. Moreover, MCC950 inhibited LPS-induced inflammation and pyroptosis of H9C2 cells. In conclusion, the NLRP3 inhibitor MCC950 can reduce the release of LDH and other cellular inflammatory factors in the cytoplasm, thereby improving the cardiac function and slowing down the apoptosis of cardiomyocytes, which may be related to the inhibition of NLRP3/Caspase-1/IL-1ß pathway.

Cellular FADD-like IL-1ß-converting enzyme-inhibitory protein attenuates myocardial ischemia/reperfusion injury via suppressing apoptosis and autophagy simultaneously.

BACKGROUND AND AIMS: Myocardial ischemia/reperfusion injury (MI/RI) is a result of coronary revascularization, and often increases cell apoptosis and autophagy. Downregulated cellular FADD-like-IL-1ß-converting enzyme-inhibitory protein (cFLIP) was associated with development of several myocardial diseases, whether overexpression of cFLIP can attenuate MI/RI remains unclear. This study aimed to determine the effects of cFLIP on apoptosis and autophagy in MI/RI. METHODS AND RESULTS: Ischemia/reperfusion (I/R) rat model and hypoxia/reoxygenation (H/R) cardiomyocytes model were established. Both I/R injury and H/R injury down-regulated expression of two cFLIP isoforms (cFLIPL and cFLIPS), and instigated apoptosis and autophagy simultaneously. Overexpression of cFLIPL and/or cFLIPS led to a significant increase in cardiomyocytes viability in vitro, and also reduced the myocardial infarct volume in vivo, these changes were associated with suppressed apoptosis and autophagy. Mechanistically, overexpression of cFLIP significantly downregulated pro-apoptotic molecules (Caspase-3, -8, -9), and pro-autophagic molecules (Beclin-1 and LC3-II). Moreover, cFLIP significantly suppressed activity of NF-κB pathway to upregulate the expression of Bcl-2, which is the molecular of interplay of apoptosis and autophagy. CONCLUSION: Overexpression of cFLIP significantly attenuated MI/RI both in vivo and vitro via suppression of apoptosis and lethal autophagy. cFLIP can suppress activity of NF-κB pathway, and further upregulated expression of Bcl-2.

Palonosetron/Methyllycaconitine Deactivate Hippocampal Microglia 1, Inflammasome Assembly and Pyroptosis to Enhance Cognition in a Novel Model of Neuroinflammation.

Since westernized diet-induced insulin resistance is a risk factor in Alzheimer's disease (AD) development, and lipopolysaccharide (LPS) coexists with amyloid ß (Aß)1-42 in these patients, our AD novel model was developed to resemble sporadic AD by injecting LPS into high fat/fructose diet (HFFD)-fed rats. The neuroprotective potential of palonosetron and/or methyllycaconitine, 5-HT3 receptor and α7 nAChR blockers, respectively, was evaluated after 8 days of daily administration in HFFD/LPS rats. All regimens improved histopathological findings and enhanced spatial memory (Morris Water Maze); however, palonosetron alone or with methyllycaconitine promoted animal performance during novel object recognition tests. In the hippocampus, all regimens reduced the expression of glial fibrillary acidic protein and skewed microglia M1 to M2 phenotype, indicated by the decreased M1 markers and the enhanced M2 related parameters. Additionally, palonosetron and its combination regimen downregulated the expression of ASC/TMS1, as well as levels of inflammasome downstream molecules and abated cleaved caspase-1, interleukin (IL)-1ß, IL-18 and caspase-11. Furthermore, ACh and 5-HT were augmented after being hampered by the insult. Our study speculates that blocking 5-HT3 receptor using palonosetron overrides methyllycaconitine to combat AD-induced neuroinflammation and inflammasome cascade, as well as to restore microglial function in a HFFD/LPS novel model for sporadic AD.

Changes in serum inflammatory factors in acute gouty arthritis patients treated using ultrashort wave combined with loxoprofen sodium.

OBJECTIVES: To study the effect of ultrashort wave combined with loxoprofen sodium on serum inflammatory factors in patients with acute gouty arthritis. METHODS: Records of patients with acute gouty arthritis who were treated in The Fourth Hospital of Changsha from May 2018 to September 2020, were reviewed. Of them, 77 cases were selected and divided into two groups based on the received treatment. The control group (n=39) was treated with loxoprofen sodium, and the treatment group (n=38) was treated with an ultrashort wave combined with loxoprofen sodium, for 10 continuous days. The clinical efficacy of the treatment in two groups was analyzed. RESULTS: After treatment, the quality of life of patients in both groups was improved (P < 0.05), but there was no significant difference in the degree of improvement between the two groups (P > 0.05). After treatment, the VAS score of the treatment group was lower than that of the control group (P < 0.05), the improvement of symptoms and signs of the treatment group was better than that of the control group (P < 0.05). Serum CRP and ESR levels in the treatment group were lower than those in the control group (P < 0.05), and the serum IL-1 ß, IL-8, TNF-a and MMP-3 levels of the treatment group were lower than those of the control group (P < 0.05). The total effective rate of the treatment group (94.87%) was higher than that of the control group (87.18%), the difference was statistically significant (P < 0.05). No adverse reactions occurred in all patients during the treatment. CONCLUSION: An ultrashort wave combined with loxoprofen sodium in the treatment of acute gouty arthritis can reduce the inflammatory reaction, improve the degree of joint pain and swelling, improve the curative effect, and do not increase the adverse reactions. The results may be related to the regulation of IL-1 ß, IL-8, TNF-a and MMP-3.

Serelaxin inhibits the profibrotic TGF-ß1/IL-1ß axis by targeting TLR-4 and the NLRP3 inflammasome in cardiac myofibroblasts.

The recombinant form of the peptide hormone relaxin, serelaxin (RLX), mediates its anti-fibrotic actions by impeding the profibrotic activity of cytokines including TGF-ß1 and IL-1ß. As IL-1ß can be produced by the nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domains-containing protein 3 (NLRP3) inflammasome, this study determined whether RLX targeted the inflammasome to inhibit the profibrotic TGF-ß1/IL-1ß axis in primary human cardiac myofibroblasts (HCMFs) in vitro and in mice with isoproterenol (ISO)-induced cardiomyopathy in vivo. HCMFs stimulated with TGF-ß1 (5 ng/ml), LPS (100 ng/ml), and ATP (5 mM) (T+L+A) for 8 h, to induce the NLRP3 inflammasome, demonstrated significantly increased protein expression of markers of NLRP3 priming (NLRP3, apoptosis-associated speck-like protein containing a C-terminal caspase-recruitment domain, procaspase-1) and activity (IL-1ß, IL-18). After 72 h, there was significantly increased neuronal NOS (nNOS), TLR-4, procaspase-1, myofibroblast differentiation, and collagen-I deposition. These measures, along with interstitial TGF-ß1 expression and collagen deposition, were also increased in the left ventricle (LV) of ISO-injured mice 14 d postinjury. RLX [16.8 nM (100 ng/ml) in vitro; 0.5 mg/kg per day in vivo] inhibited T+L+A- and ISO-induced TLR-4 expression, NLRP3 priming, IL-1ß, IL-18, myofibroblast differentiation, and interstitial collagen deposition at the time points studied, via the promotion of nNOS; with the NLRP3- and IL-1ß-inhibitory effects of RLX in HCMFs being abrogated by pharmacological blockade of nNOS or TLR-4. Comparatively, the small molecule NLRP3 inhibitor, N-{[(1,2,3,5,6,7-hexahydro-s-indacen-4-yl)amino]carbonyl}-4-(1-hydroxy-1-methylethyl)-2-furansulfonamide (1 µM in vitro, 10 mg/kg/d in vivo), inhibited components of the NLRP3 inflammasome in vitro and in vivo and ISO-induced interstitial LV fibrosis in vivo but did not affect nNOS, TLR-4, myofibroblast differentiation, or myofibroblast-induced collagen deposition. Hence, RLX can inhibit the TGF-ß1/IL-1ß axis via a nNOS-TLR-4-NLRP3 inflammasome-dependent mechanism on cardiac myofibroblasts.-Cáceres, F. T., Gaspari, T. A., Samuel, C. S., Pinar, A. A. Serelaxin inhibits the profibrotic TGF-ß1/IL-1ß axis by targeting TLR-4 and the NLRP3 inflammasome in cardiac myofibroblasts.

Protective Effect of Esculetin, Natural Coumarin in Mice Model of Fibromyalgia: Targeting Pro-Inflammatory Cytokines and MAO-A.

Fibromyalgia is a refractory syndrome characterized by chronic wayward pain and complex co-morbid psychological trepidation. The current treatments have a limited role and proper clinical benefits are far from satisfactory. Naturally occurring coumarins such as osthole are known to have analgesic and anti-inflammatory activities. Therefore, the current investigation was designed to explore the potential of natural coumarin esculetin (2.5, 5, and 10 mg/kg) in mitigating reserpine-induced fibromyalgia in Swiss albino mice. Esculetin is a 6,7 dihydroxy-coumarin obtained from various plant sources such as Aesculus hippocastanum L, Ceratostigma willmottianum, Citrus limonia, etc. Reserpine (0.5 mg/kg/day s.c.) treatment for first 3 days, significantly altered the behavior of mice as evidenced by reduced paw withdrawal threshold in pressure application measurement (PAM) test and electronic von-Frey (eVF) test, increased immobility time in forced swim test (FST), increased latency to reach the platform in Morris water maze (MWM) test and reduced number of square crossed in the open field test (OFT). These behavioral deficits with reserpine treatment were integrated with a reduced level of serotonin (5-HT), reduced glutathione (GSH), along with an increase in monoamine oxidase-A (MAO-A) activity, pro-inflammatory cytokines (IL-1ß, TNF-α), thiobarbituric acid reactive substances (TBARS) and glutamate level. Esculetin (10 mg/kg/day i.p) treatment for 5 days, significantly abrogated reserpine induced behavioral and biochemical alterations. Whereas, no significant improvement was observed with lower doses of esculetin i.e. 2.5 and 5 mg/kg.