Inflammatory Signaling Research Articles

Circulating small extracellular vesicles as blood-based biomarkers of muscle health in aging nonhuman primates.

Age-associated loss of muscle mass and function and subsequent mobility decline define poor health outcomes, reduced quality of life, and mortality risk. The rate and extent of aging-related muscle loss varies across older adults. It is challenging to understand the molecular pathogenesis of mobility decline, as anthropometric and imaging techniques, primarily used in muscle function assessment, do not offer much molecular information. Small extracellular vesicles (sEV) are lipid membrane-bound, nano-sized (≤ 200nm) vesicles which carry a wide array of biomolecules as their cargo. sEV contain cell/tissue-specific signatures on their surface and can be isolated from biofluids. These properties pose sEV as a minimally invasive means to monitor the functional and biological health of difficult-to-access tissues, establishing them as a promising liquid biopsy tool. Here, we first isolated skeletal muscle-derived sEV (sEVSKM) from the serum of vervet monkeys (16 to < 25years old) using alpha sarcoglycan (SGCA) as a muscle-specific sEV surface marker. sEVSKM were extensively characterized for size, concentration, purity, and specificity. Further, sEVSKM isolated from young (11-15years) and old (25-29years) monkeys' serum were characterized for oxidized proteins by mass spectrometry and miRNAs by small-RNAseq. Notably, the analysis of oxidized proteins indicated perturbation of metabolic pathways, actin cytoskeleton, muscle cytoskeleton regulation, and HIF-1 signaling in older monkeys. Furthermore, small-RNAseq analysis identified differential expression of several miRNAs regulating metabolic pathways, inflammation, and stress signaling. Altogether, these results suggest that it is feasible to isolate sEVSKM and use them to identify molecular biomarkers that reflect the physiological state of muscle tissue.

Comparative liver transcriptome analysis in hamsters infected with food-borne trematodes Opisthorchis felineus, Opisthorchis viverrini, or Clonorchis sinensis.

Epidemiologically important food-borne trematodes Opisthorchis viverrini and Clonorchis sinensis are recognized as biological carcinogens of Group 1A, while Opisthorchis felineus is in Group 3 as noncarcinogenic to humans. Mechanisms of the biological carcinogenesis are still elusive. Some studies highlight chronic inflammation as a key factor and common pathway for cancer initiation and progression. Nonetheless, the chronic inflammation alone does not explain why these three species differ in carcinogenicity. We focused this study on genome-wide landscapes of liver gene expression and activation of cellular pathways in Mesocricetus auratus golden hamsters infected with C. sinensis (South Korea), O. viverrini (Thailand), or O. felineus (Russia) at 1 and 3 months after infection initiation. Liver transcriptomes of golden hamsters (HiSeq Illumina, 2X150 bp) were sequenced at 1 and 3 months postinfection. Data processing was carried out using the following bioinformatic and experimental approaches: analysis of differential gene expression, estimates of proportions of affected liver cell types, liver histopathology, and examination of weighted gene coexpression networks. All infections caused enrichment with inflammatory response signaling pathways, fibrogenesis and cell proliferation, and IL2-STAT5, TNF-NF-κB, TGF-β, Hippo, MAPK, and PI3K-Akt signaling pathways. Nevertheless, species-specific responses to each infection were noted too. We also identified species-specific responses of liver cell types, differentially expressed gene clusters, and cellular pathways associated with structural liver damage (such as periductal fibrosis, epithelial neoplasia, and inflammation). This is the first comparative analysis of gene expression landscapes in the liver of experimental animals infected with O. viverrini, O. felineus, or C. sinensis. The trematodes have species-specific effects on the hepatobiliary system by triggering signaling pathways, thereby leading to differences in the severity of hepatobiliary structural lesions and contributing to the pathogenicity of closely related foodborne trematodes.

Abstract A017: The FES tyrosine kinase as an emerging target for cancer immunotherapy

Abstract Immunotherapies are a promising emerging pillar of cancer treatment, but they still face many barriers due to the immunosuppressive nature of cancer. Cancer immunotherapy relies on the interplay between innate and adaptive immune responses. One way of stimulating such responses, known as immunogenic cell death (ICD), involves the release of tumour-associated antigens and damage associated molecular patterns (DAMPs). These DAMPs function to recruit and activate innate immune cells, including antigen-presenting cells (APCs), through engagement of pattern recognition receptors (PRRs), subsequently leading to production of the pro-inflammatory Signal 3 cytokines required for activation of adaptive immune cells (e.g., cytotoxic T lymphocytes [CTLs] and natural killer [NK] cells). The tyrosine kinase Fes suppresses innate immune responses in APCs by inhibiting components of the PRR signaling cascade. In non-cancer contexts, the negative regulation of APCs by Fes may guard against consequences of overactive innate immunity, including endotoxic shock or autoimmune disease. However, this same inhibitory effect on APC function may also serve as a checkpoint to successful anti-cancer immunotherapy, by obstructing efficient priming of cancer specific CTLs by APCs. Therefore, by inhibiting Fes, we hypothesize there will be greater Signal 3 cytokine production, resulting in greater CTL activation, and therefore improved tumor control. Using bone marrow derived APCs, including macrophages (BMDMs) and dendritic cells (BMDCs), from wildtype (WT) or Fes knockout (fes-/-) mice, we have shown through both Western blotting and flow cytometry analysis, that PRR signal transduction cascades are suppressed by Fes and increase levels of Signal 3 cytokines produced by fes-/- APCs. This includes higher levels of cell associated IL-12 in fes-/- APCs. Using syngeneic orthotopic mouse engraftment models of triple negative breast cancer (EO771) and melanoma (B16-F10) we showed that treatment with doxorubicin (which induces ICD) or anti-PD-1 (immune checkpoint inhibitor) plus doxorubicin controls tumor growth and prolongs survival to a greater extent in fes-/- mice. Immunophenotyping of tumors and spleens from these mice showed higher levels of activated CTLs and skewing of macrophages to a M1 state in fes-/- mice. SIINFEKL peptide loaded-BMDM/BMDCs from fes-/- mice were also more effective at priming CTLs from OT-1 mice (which express a T cell receptor that recognizes the SIINFEKL peptide) in antigen cross-presentation co-culture assays. These results implicate Fes as a potential novel immune checkpoint whose inhibition may enhance anti-cancer immunotherapy by suppressing its role in dampening inflammatory Signal 3 cytokine production by APCs. I will present recent data exploring the role of Fes in regulating the expression and trafficking of IL-12 in APCs to better understand the molecular basis of improved CTL activation by fes-/- APCs. Citation Format: Julian Simonetti, Brian J. Laight, Natasha Dmytryk, Danielle Harper, Yan Gao, Changnian Shi, Madhuri Koti, Sameh Basta, Peter A. Greer. The FES tyrosine kinase as an emerging target for cancer immunotherapy [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Optimizing Therapeutic Efficacy and Tolerability through Cancer Chemistry; 2024 Dec 9-11; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(12_Suppl):Abstract nr A017.

Imperfect wound healing sets the stage for chronic diseases.

Although the age of the genome gave us much insight about how our organs fail with disease, it also suggested that diseases do not arise from mutations alone; rather, they develop as we age. In this Review, we examine how wound healing might act to ignite disease. Wound healing works well when we are younger, repairing damage from accidents, environmental assaults, and battles with pathogens. Yet, with age and accumulation of mutations and tissue damage, the repair process can devolve, leading to inflammation, fibrosis, and neoplastic signaling. We discuss healthy wound responses and how our bodies might misappropriate these pathways in disease. Although we focus predominantly on epithelial-based (lung and skin) diseases, similar pathways might operate in cardiac, muscle, and neuronal diseases.

The DERMIS Study: Methodologies, Results, and Implications for the Future.

Ongoing innovation in diabetes technologies has led to the development of advanced tools such as automated insulin delivery (AID) systems that adjust insulin delivery in response to current and predicted glucose levels, residual insulin action, and other inputs (eg, meal and exercise announcements). However, infusion sets continue to be the "Achilles heel" of accurate and precise insulin delivery and continued device use. A recent study by Kalus et al (DERMIS Study) revealed higher vessel density and signals of inflammation by optical coherence tomography (OCT), in addition to increased inflammation, fat necrosis, fibrosis, and eosinophilic infiltration by histopathology. Although the study provided a comprehensive description of what was happening, the results raise important questions that require additional research. On February 29, 2024, the Leona M. and Harry B. Helmsley Charitable Trust sponsored a conference to begin addressing these issues. This article summarizes the DERMIS study findings and testing methodologies discussed at the conference and proposes the next steps for developing insulin infusion sets that reduce the variability in insulin delivery and extend wear.

The CCL5/CCR5/SHP2 axis sustains Stat1 phosphorylation and activates NF-κB signaling promoting M1 macrophage polarization and exacerbating chronic prostatic inflammation

Background and objectiveChronic prostatitis (CP) is a condition markered by persistent prostate inflammation, yet the specific cytokines driving its progression remain largely undefined. This study aims to identify key cytokines involved in CP and investigate their role in driving inflammatory responses through mechanistic and therapeutic exploration.MethodsA 48-cytokine panel test was conducted to compare the plasma cytokine profiles between participants with CP-like symptoms (CP-LS) and healthy controls. Experimental autoimmune prostatitis (EAP) models were used for functional validation, with further mechanistic studies performed through in vivo and in vitro assays. Pharmacological inhibition was applied using maraviroc, and pathway inhibitors to assess therapeutic potential.ResultsOur analysis identified CCL5 as one of the most prominently elevated cytokines in CP-LS patients. Further validation in the EAP model mice confirmed elevated CCL5 levels, highlighting its role in driving prostatic inflammation. Mechanistic studies revealed that CCL5 interacts with the CCR5 receptor, promoting M1 macrophage polarization and activating key inflammatory signaling pathways, including Stat1 and NF-κB, as indicated by increased phosphorylation of Stat1 and p65. In vitro, CCL5 combined with LPS stimulation amplified these effects, further promoting M1 polarization. CCL5 also sustained Stat1 activation by inhibiting its dephosphorylation through reduced interaction with SHP2, leading to prolonged inflammatory signaling. Single-cell transcriptomics confirmed high CCR5 expression in macrophages, correlating with inflammatory pathways. Pharmacological inhibition of CCR5, or its downstream signaling, significantly reduced macrophage-driven inflammation both in vivo and in vitro.ConclusionThese findings establish the CCL5/CCR5 axis as a critical driver of persistant prostatic inflammation and present it as a potential therapeutic target for CP.Graphic The graphic abstract illustrates the central role of the CCL5/CCR5 axis in promoting chronic prostatitis progression. CCL5 binds to the CCR5 receptor on macrophages, enhancing M1 polarization and activating key inflammatory pathways, including Stat1 and NF-κB, as shown by the increased phosphorylation of Stat1 and p65. The interaction also inhibits SHP2-mediated Stat1 dephosphorylation, sustaining prolonged inflammatory signaling. Pharmacological inhibition of CCR5 or its downstream pathways attenuates macrophage-driven inflammation, highlighting this axis as a critical driver and potential therapeutic target for chronic prostatitis.

Mechanisms of ferroptotic and non-ferroptotic organ toxicity of chemotherapy: protective and therapeutic effects of ginger, 6-gingerol and zingerone in preclinical studies.

Chemotherapy (CT) is one of the flagship options for the treatment of cancers worldwide. It involves the use of cytotoxic anticancer agents to kill or inhibit the proliferation of cancer cells. However, despite its clinical efficacy, CT triggers side effect toxicities in several organs, which may impact cancer patient's quality of life and treatment outcomes. While the side effect toxicity is consistent with non-ferroptotic mechanisms involving oxidative stress, inflammation, mitochondrial impairment and other aberrant signalling leading to apoptosis and necroptosis, recent studies show that ferroptosis, a non-apoptotic, iron-dependent cell death pathway, is also involved in the pathophysiology of CT organ toxicity. CT provokes organ ferroptosis via system Xc-/GPX-4/GSH/SLC7A11 axis depletion, ferritinophagy, iron overload, lipid peroxidation and upregulation of ferritin-related proteins. Cisplatin (CP) and doxorubicin (DOX) are common CT drugs indicated to induce ferroptosis in vitro and in vivo. Studies have explored natural preventive and therapeutic strategies using ginger rhizome and its major bioactive compounds, 6-gingerol (6G) and zingerone (ZG), to combat mechanisms of CT side effect toxicity. Ginger extract, 6G and ZG mitigate non-ferroptotic oxidative inflammation, apoptosis and mitochondrial dysfunction mechanisms of CT side effect toxicity, but their effects on CT-induced ferroptosis remain unclear. Systematic investigations are, therefore, needed to unfold the roles of ginger, 6G and ZG on ferroptosis involved in CT side effect toxicity, as they are potential natural agents for the prevention of CT toxicity. This review reveals the ferroptotic and non-ferroptotic toxicity mechanisms of CT and the protective mechanisms of ginger, 6G and ZG against CT-induced, ferroptotic and non-ferroptotic organ toxicities.

Selectively Antagonizing the NOD1-Mediated Inflammatory Signaling Pathway Mitigates the Gastric Inflammation Induced by Helicobacter pylori Infection.

Helicobacter pylori (H. pylori) infection is characterized by the complex interplay between H. pylori and gastric disorders. It has been established that NOD1 can be activated by the peptidoglycan (PGN) present in the cell wall of H. pylori, serving as a key mediator of inflammation and initiating the RIP2/NF-κB and MAPK inflammatory signaling pathways. In this article, we reported on the development of a 2-chloroquinazolin-4-ol derivative 66 as a potent and selective antagonist of both human and mouse NOD1, which effectively inhibited the expression of inflammatory cytokines (IL-6, TNF-α) and chemokines (CXCL1, CXCL8) in immune and epithelial cells, as well as inflammatory cytokines (KC, IL-6) in a H. pylori-induced murine model of gastritis following oral administration. This study laid a foundation for treating gastritis induced by H. pylori infection.

The Anti-Inflammatory Properties of Polysaccharides Extracted from Moringa oleifera Leaves on IEC6 Cells Stimulated with Lipopolysaccharide In Vitro

Moringa oleifera (M. oleifera) is a plant with significant medicinal and nutritional value and contains various bioactive compounds, particularly in its leaves (MOL). This study sought to explore the impact of M. oleifera leaf polysaccharides (MOLPs) on lipopolysaccharide (LPS)-activated intestinal epithelial cells (IEC6) and to uncover the mechanisms involved. The cytotoxicity of MOLP on IEC6 cells was assessed using the Cell Counting Kit-8 (CCK-8) assay, which demonstrated a safe concentration range of 0–1280 µg/mL. The impact of MOLP on cell viability was further evaluated over 12 to 48 h. IEC6 cells were treated with three concentrations of MOLP low (25 µg/mL), medium (50 µg/mL), and high (100 µg/mL) alongside LPS (50 µg/mL) stimulation for one day. The findings revealed that treatment with MOLP significantly promoted cell migration and increased the production of interleukin-10 (IL-10), while it simultaneously decreased cell apoptosis and the levels of pro-inflammatory cytokines, such as tumour necrosis factor alpha (TNF-α), interleukin 1β (IL-1β), and interleukin 6 (IL-6). Additionally, MOLP treatments across all concentrations significantly reduced the expression of Toll-like receptor 4 (TLR-4), myeloid differentiation primary response 88 (MyD88), phosphorylated nuclear factor kappa B-alpha (pIκB-α), and phosphorylated NF-κB p65 signalling pathways. Moreover, MOLP restored the expression of tight junction proteins, such as zonula occludens-1 (ZO-1) and occludin, which had been disrupted by LPS. These results indicate that MOLP exhibits anti-inflammatory properties by inhibiting inflammatory signalling pathways and maintaining intestinal barrier integrity through the upregulation of tight junction proteins in IEC6 cells. This study enhances our understanding of the anti-inflammatory capabilities of MOLP.

Necroptosis contributes to deoxynivalenol-induced liver injury and inflammation in weaned piglets

BackgroundThe aim of this study was to investigate the role of necroptosis in deoxynivalenol (DON)-induced liver injury and inflammation in weaned piglets.MethodsIn Exp. 1, 12 weaned piglets were divided into 2 groups including pigs fed basal diet and pigs fed diet contaminated with 4 mg/kg DON for 21 d. In Exp. 2, 12 weaned piglets were divided into 2 groups including control piglets and piglets given a gavage of 2 mg/kg body weight (BW) DON. In Exp. 3, 24 weaned piglets were used in a 2 × 2 factorial design and the main factors including necrostatin-1 (Nec-1) (DMSO or 0.5 mg/kg BW Nec-1) and DON challenge (saline or 2 mg/kg BW DON gavage). On 21 d in Exp. 1, or at 6 h post DON gavage in Exp. 2 and 3, pigs were killed for blood samples and liver tissues. Liver histology, blood biochemical indicators, and liver inflammation and necroptosis signals were tested.ResultsDietary or oral gavage with DON caused liver morphological damage in piglets. Dietary DON led to hepatocyte damage indicated by increased aspartate transaminase (AST) activity and AST/alanine aminotransferase (ALT) ratio, and DON gavage also caused hepatocyte damage and cholestasis indicated by increased AST and alkaline phosphatase (AKP) activities. Dietary DON caused liver necroptosis indicated by increased protein abundance of total receptor interacting protein kinase 3 (t-RIP3) and total mixed lineage kinase domain-like protein (t-MLKL). Moreover, DON gavage increased mRNA expression of interleukin (IL)-6 and IL-1β in liver. DON gavage also induced liver necroptosis demonstrated by increased protein abundance of t-RIP3, phosphorylated-RIP3 (p-RIP3), t-MLKL and p-MLKL. However, pretreatment with Nec-1, a specific inhibitor of necroptosis, inhibited liver necroptosis indicated by decreased protein expression of t-RIP3, p-RIP3, t-MLKL and p-MLKL. Nec-1 pretreatment reduced liver morphological damage after DON gavage. Pretreatment with Nec-1 also attenuated liver damage induced by DON indicated by decreased activities of AST and AKP. Furthermore, Nec-1 pretreatment inhibited liver mRNA expression of IL-6 and IL-1β after DON challenge.ConclusionsOur data demonstrate for the first time that necroptosis contributes to DON-induced liver injury and inflammation in piglets.

Mechanistic Study of Purple Sweet Potato Anthocyanins: Multifaceted Anti-Fibrotic Effects and Targeting of PDGFRβ in Liver Fibrosis.

The purple sweet potato anthocyanins (PSPA) are known for their diverse health benefits, yet their hepatoprotective effects and the mechanisms by which they combat liver fibrosis have not been thoroughly investigated. This study aimed to elucidate these effects by employing a carbon tetrachloride (CCl4)-induced mouse model of liver fibrosis. We conducted a comprehensive analysis of the effects of PSPA on liver injury, oxidative stress, inflammation, and fibrosis-related signaling pathways. Our results demonstrate that PSPA can mitigate liver damage in mice, regulate key antioxidant enzymes such as catalase and SOD, and reduce oxidative stress as indicated by lowered MDA levels. PSPA also decrease the expression of inflammatory proteins, including CD3, CD4, CD45, IL-1β, TNF-α, and IL-17A, and reduce the accumulation of fibrotic markers like type I and III collagens and α-SMA. Additionally, PSPA have demonstrated the ability to inhibit key fibrogenic signaling proteins, including TGFβR2, p-Smad2, p-Smad3, p-PDGFRβ, p-AKT, p-ERK1/2, p-JNK1/2, and p-p38. Furthermore, we identified two potent monomers, PSPA-1 and PSPA-2, which directly target the PDGFRβ, a key player in fibrosis. The mechanism of action involves the inhibition of PDGF-B binding to PDGFRβ, thus disrupting the PDGF-B/PDGFRβ signaling pathway. These findings suggest that the hepatoprotective and antifibrotic effects of PSPA are due to their multifunctional bioactivities and the presence of specific active components that can effectively target fibrogenic protein.

Chitooligosaccharide-modified PLGA-loaded PPD nanoparticles ameliorated sepsis-associated acute kidney injury via the NF-κB signaling pathway

Objectives Sepsis-associated acute kidney injury (SA-AKI) is a significant clinical challenge with high morbidity and mortality. Low bioavailability of protopanaxadiol (PPD) limits its clinical application. In this study, PPD was encapsulated with chitooligosaccharide (COS) modified polylactic-co-glycolic acid (PLGA) to develop novel nanomedicines for the treatment of SA-AKI. Methods COS-PLGA-PPD nanoparticles were prepared by emulsified solvent evaporation method, and their properties were evaluated. In vitro, the anti-inflammatory and protective effects of COS-PLGA-PPD NPs were investigated in a cellular model of LPS-induced NRK-52E cells and their uptake in Caco-2 cells. Indicators of renal injury, inflammation, and NF-κB signaling pathway were evaluated by injecting LPS into SD rats and inducing SA-AKI model in vivo. The oral bioavailability of nanoparticles was evaluated by pharmacokinetics. Results Compared with PPD and unmodified nanoparticles, COS-PLGA-PPD NPs were more stable, with a particle size of 139.69 nm, which enhanced the viability of NRK-52E cells, increased the uptake of Caco-2 cells, alleviated the symptoms of SA-AKI in rats, inhibited the NF-κB signaling pathway, reduced the expression of inflammatory factors, and had a bioavailability 1.7-fold that of PPD. Conclusion COS-PLGA-PPD NPs ameliorate LPS-induced SA-AKI in rats by inhibiting the NF-κB signaling pathway, providing a basis for the treatment of SA-AKI.

High glucose potentiates Zika virus induced-astroglial dysfunctions.

Zika virus (ZIKV) is a neurotropic flavivirus that induces congenital Zika syndrome and neurodevelopmental disorders. Given that ZIKV can infect and replicate in neural cells, neurological complications in adult brain are also observed. Glial cells may emerge to delay and/or prevent the development of ZIKV-induced neurodegeneration. These cells actively participate in metabolic, inflammatory and redox processes, and consequently, in the pathophysiology of neurodegenerative diseases, including diabetic encephalopathy. In this sense, changes in glucose metabolism can support the inflammatory activity of astroglial cells; however, the effects of increased glucose concentration during ZIKV infection have not yet been explored in astroglial cells. Here, we evaluated functional parameters of astroglial cells exposed to ZIKV upon normal and high glucose concentrations, focusing on inflammatory profile, oxidative stress, and expression of critical genes for astroglial functions. High glucose potentiated the pro-inflammatory and oxidative effects of ZIKV, as well as potentiated the downregulation of signaling pathways, such as Nrf-2 (nuclear factor erythroid derived 2 like 2), sirtuin 1 (SIRT1), peroxisome proliferator activated receptor gamma coactivator 1-alpha (PGC-1α), and poly (ADP-ribose) polymerase (PARP). In summary, our results suggest that high glucose can favor the activation of inflammatory signaling while impairing cytoprotective pathways in astroglial cells exposed to ZIKV and reinforce the hypothesis that this virus is highly neurotrophic, with significant impact in glial cells.

Evaluation of the effect of phellodendrin application on rats creating an experimental model of non-compression lumbar disc herniation on the NF-κB-related inflammatory signaling pathway

ObjectiveTo explore the therapeutic effects of phellodendrine on non-compression lumbar disc herniation (NCLDH).MethodsThe Sprague Dawley rat model of NCLDH was established via autologous caudal nucleus pulposus transplantation. Behavioral observations and neurological function scoring were conducted in Sprague Dawley rats, and the serum levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were measured via enzyme-linked immunosorbent assay (ELISA). Real-time quantitative polymerase chain reaction (RT‒qPCR) was used to detect the expression of nuclear factor kappa-B (NF-κB) p65 mRNA in L5 nerve roots and surrounding tissues. Western blotting was used to assess the protein expression of NF-κB p65 and TNF-α. Immunofluorescence and immunohistochemical analyses were performed to investigate the distribution and expression of the NF-κB p65 protein in the L5 nerve and its surrounding tissues.ResultsIn this animal study, phellodendrine was found to downregulate the expression of p65 mRNA, decrease the release of inflammatory factors, and alleviate motor dysfunction caused by lumbar disc herniation(LDH). Therefore, the phellodendrine technique has potential value for the treatment of NCLDH.ConclusionIn this animal experiment, phellodendrine was found to significantly reduce the expression level of p65 mRNA, decrease the release of inflammatory cytokines, and alleviate lumbar disc pain.Clinical trial numberNot applicable.

Effect of cigarette smoke on the proliferation, viability, gene expression, and cellular functions of adipose-derived mesenchymal stem cells from smoking and non-smoking donors.

Cigarette smoking negatively impacts mesenchymal stem cell functionality, including proliferation, viability, and differentiation potential. Adipose-derived mesenchymal stem cells (ADMSCs) are increasingly used for therapeutic purposes, but the specific effects of smoking in vivo on these cells are poorly understood. This study investigates the effects of cigarette smoke on the proliferation, viability, gene expression, and cellular functions of ADMSCs from smoking and non-smoking donors. In this study, ADMSCs were isolated from healthy smokers and non-smokers, and cell proliferation was assessed using the MTT assay, viability with apoptosis assays, mitochondrial membrane potential (MMP), and gene expression related to oxidative stress and cellular functions. Cell cycle analysis was also conducted. Our findings reveal a significant decrease in the proliferation of ADMSCs from smokers. Apoptosis assays showed reduced viable cells in smokers without a significant change in MMP, suggesting alternative pathways contributing to decreased viability. Gene expression analysis indicated the upregulation of genes associated with oxidative stress response and cellular defense mechanisms and the downregulation of genes related to inflammatory signaling, detoxification, and cellular metabolism. Cell cycle analysis indicates cycle arrest or delay in smokers, possibly due to stress and potential DNA damage. Smoking negatively affects ADMSCs' proliferation, viability, and function through oxidative stress and gene expression alterations. These findings highlight the importance of considering smoking status in ADMSC therapies and the need for further research to mitigate the effect of smoking on stem cells.

The ameliorative effect of pioglitazone against colistin-induced nephrotoxicity is mediated by inhibition of NF-κB and restoration of Nrf2 signaling: An integrative bioinformatics prediction-guided in vitro study.

Pioglitazone, an anti-diabetic drug, has been previously shown to ameliorate kidney damage through anti-inflammatory and antioxidant effects. In this study, we employed an integrative bioinformatics approach to study the possible mechanisms involved in the mitigative effect of pioglitazone against colistin-induced nephrotoxicity. Next, we validated the results obtained from the bioinformatics study by pre-treating human kidney-2 (HK-2) cell line with pioglitazone 100 μM for 30 minutes and then treating the cells with colistin sulfate 1200 μM for 24 hours. Inflammatory signaling by cytokines and the nuclear factor erythroid 2 related factor 2 (Nrf2) signaling pathways were predicted to be involved in the ameliorative effect of pioglitazone against colistin-induced nephrotoxicity. The nuclear factor kappa B subunit p65 (NF-κB p65) and Nrf2 were among the predicted transcription factors regulating the hub genes. Moreover, miR-24, miR-16, and miR-21 were identified as potential pathogenic miRNAs regulating the hub genes. In contrast, miR-17, miR-27a, and miR-146a were identified as potential protective miRNAs. The in vitro study indicated that pioglitazone pre-treatment increased cell viability in HK-2 cells exposed to colistin. Pioglitazone pre-treatment reduced the expression of pro-inflammatory cytokine genes (IL6 and TNF). Moreover, pioglitazone reduced the protein expression of NF-κB p65 and increased the protein expression of Nrf2. The protective effect of pioglitazone against colistin-induced toxicity in HK-2 cells is related to its anti-inflammatory and antioxidant activity through modulating NF-κB-mediated inflammatory signaling and Nrf2-mediated antioxidative stress signaling.

Dorzolamide Intermediates with Potential Anti-Inflammatory Activity

Dorzolamide (DZD), a Carbonic anhydrase (CA) inhibitor clinically used to lower intraocular pressure, exhibits anti-inflammatory effects owing to the drug’s ability to inhibit the TIR domain-containing adaptor protein (TIRAP)-mediated signalling in macrophages. Here, we investigated whether DZD intermediates also demonstrate any anti-inflammatory property like DZD but with a reduced inhibition of CA. We found that several intermediates of DZD show increased binding to TIRAP at the common interface of kinases, such as Protein kinase C-delta (PKCδ) and Bruton's tyrosine kinase (BTK). Such binding results in a decreased activity of TIRAP, p38 Mitogen-activating protein kinases (MAPK), and p65, which are essential for major inflammatory signaling pathways. Remarkably, the DZD intermediates were more effective than DZD in decreasing the mRNA expression levels of pro-inflammatory cytokines in Lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. The DZD intermediates also exhibit a reduced binding energy to CA II and CA IV, highlighting their improved specificity as anti-inflammatory compounds with decreased unwanted biological effects. Furthermore, we validated the anti-inflammatory effect of the most efficient DZD intermediate, DRZ V, in a model of mouse sepsis. DRZ V-treated septic mice exhibited improved survival compared to DZD-treated septic mice. Our data indicate that the tested DZD intermediates are more effectual in dampening TIRAP-mediated inflammatory signaling as compared to DZD. Thus, DZD intermediates may be a promising option for developing novel anti-inflammatory therapeutics.

Stat3 mediates Fyn kinase-driven dopaminergic neurodegeneration and microglia activation.

The Alzheimer's disease and Parkinson's disease risk locus FYN kinase is implicated in neurodegeneration and inflammatory signaling. To investigate in vivo mechanisms of Fyn-driven neurodegeneration, we built a zebrafish neural-specific Gal4:UAS model of constitutively active FynY531F signaling. Using in vivo live imaging, we demonstrated that neural FynY531F expression leads to dopaminergic neuron loss and mitochondrial aggregation in 5 day larval brain. Dopaminergic loss coincided with microglia activation and induction of tnfa, il1b and il12a inflammatory cytokine expression. Transcriptome analysis revealed Stat3 signaling as a potential Fyn target. Chemical inhibition experiments confirmed Fyn-driven dopaminergic neuron loss, and the inflammatory response was dependent upon activation of Stat3 and NF-κB pathways. Dual chemical inhibition demonstrated that Stat3 acts synergistically with NF-κB in dopaminergic neuron degeneration. These results identify Stat3 as a novel downstream effector of Fyn signaling in neurodegeneration and inflammation.

Protective Effects of NRF2 Activator Sulforaphane in Polyinosinic:Polycytidylic Acid-Induced In Vitro and In Vivo Model.

NRF2 is a nuclear transcription factor involved in the cellular protection against oxidative stress and inflammatory signaling. Sulforaphane is a known NRF2 activator used for its strong antioxidant and anti-inflammatory activity through regulation of Keap-1-HO-1 pathway. However, there is a limited exploration about the role of NRF2 activator, sulforaphane in regulation of poly(I:C)-induced oxidative stress, inflammation and injury in lung. Therefore, we aimed to evaluate the therapeutic effect of sulforaphane in poly(I:C)-induced responses using in vitro as well as in vivo model. We evaluated oxidative stress and inflammatory cytokines in poly(I:C)-induced RAW264.7 cells. We also employed in vivo animal study to evaluate tissue oxidative-antioxidative balance along with expression of NRF2, Keap-1, histopathological assessment by hematoxylin-eosin staining and picrosirius red staining to explore the protective mechanisms of sulforaphane in poly(I:C)-induced mouse model. Our results indicated that sulforaphane increased the expression of NRF2 and its downstream proteins. In addition, sulforaphane alleviated poly(I:C)-induced activation of the oxidative and pro-inflammatory pathways, histopathological changes, depleted expression of GSH and superoxide dismutase in lung tissue. This study suggested that sulforaphane may be one of the useful therapeutic alternatives for poly(I:C) induced lung injury and inflammation.

Imbalanced and Unchecked: The Role of Metal Dyshomeostasis in Driving COPD Progression.

Chronic obstructive pulmonary disease (COPD) is a chronic respiratory condition characterized by persistent inflammation and oxidative stress, which ultimately leads to progressive restriction of airflow. Extensive research findings have cogently suggested that the dysregulation of essential transition metal ions, notably iron, copper, and zinc, stands as a critical nexus in the perpetuation of inflammatory processes and oxidative damage within the lungs of COPD patients. Unraveling the intricate interplay between metal homeostasis, oxidative stress, and inflammatory signaling is of paramount importance in unraveling the intricacies of COPD pathogenesis. This comprehensive review aims to examine the current literature on the sources, regulation, and mechanisms by which metal dyshomeostasis contributes to COPD progression. We specifically focus on iron, copper, and zinc, given their well-characterized roles in orchestrating cytokine production, immune cell function, antioxidant depletion, and matrix remodeling. Despite the limited number of clinical trials investigating metal modulation in COPD, the advent of emerging methodologies tailored to monitor metal fluxes and gauge responses to chelation and supplementation hold great promise in unlocking the potential of metal-based interventions. We conclude that targeted restoration of metal homeostasis represents a promising frontier for ameliorating pathological processes driving COPD progression.