Reduced IL-6 Production Research Articles

The Anti-inflammatory Potential of Tricyclic Antidepressants (TCAs): A Novel Therapeutic Approach to Atherosclerosis Pathophysiology

Atherosclerosis, a chronic inflammatory disease, is driven by complex molecular mechanisms involving inflammatory cytokines and immune pathways. According to recent research, tricyclic antidepressants (TCAs), which are typically prescribed to treat depressive disorders, have strong anti-inflammatory effects. TCAs, including imipramine and amitriptyline, alter inflammatory signaling cascades, which include lowering the levels pro-inflammatory cytokines like TNF-α, IL-1β, and IL-6 and inhibiting NF-κB activation. By inhibiting the NLRP3 inflammasome and suppressing pathways including JAK/STAT, MAPK, and PI3K, these effects are produced, improving endothelial function and reducing oxidative stress. The intricacy of TCAs’ anti-inflammatory actions has demonstrated by the existence of contradictory findings about how they alter IL-6 levels. The dependence of the heterogeneity of the reaction on the use of particular TCAs and experimental settings is shown by the fact that some studies show reduced IL-6 production, while others indicate increases or no changes. This review explores the multifaceted mechanisms through which TCAs modulate inflammatory pathways. TCAs inhibit NF-κB activation, reduce oxidative stress, and suppress the production of key inflammatory mediators, including IL-6 and TNF-α. They also regulate Toll-like receptor (TLR) signaling and NOD-, LRR-, and NLR family pyrin domain-containing protein 3 (NLRP3) inflammasome activation, reducing the release of IL-1β and IL-18, critical drivers of endothelial dysfunction and plaque instability. Given their capacity to target critical inflammatory molecules and pathways, TCAs provide great potential in the therapy of atherosclerosis, particularly for individuals with associated depression and cardiovascular risk factors. Nevertheless, further research is essential to clarify the precise molecular mechanisms, resolve inconsistencies in current findings, and establish the clinical applicability of TCAs as anti-inflammatory agents in atherosclerosis management.

Leptin/LPS-treated dendritic cells reduce the expression of genes involved in tumor tissue metastasis and angiogenesis in an animal model of breast cancer.

Leptin, an immune-regulating protein, enhances the maturation of dendritic cells (DCs). We previously demonstrated that leptin and lipopolysaccharide (LPS) promote the expression of co-stimulatory molecules on the surface of DCs. Leptin/LPS-treated DCs increased T cell responses against 4T1 breast cancer in mice. Therefore, in the present study, we investigate the effects of a DC vaccine treated with leptin and LPS on the genes involved in tumor metastasis, angiogenesis, and related cytokines in a mouse model of breast cancer. Tumor induction was achieved through subcutaneous injection of 4T1 cells into syngeneic mice. On days 12 and 19, the mouse groups received the DC vaccine treated with leptin and a combination of leptin and LPS. After sacrificing the mice on day 26, the levels of IL-6 and IL-33 in the serum were assayed using the ELISA technique, and the expression levels of the VEGF, CCL2, MMP9, and CCL5 genes in the tumors were measured by Real-Time PCR. Compared to untreated tumor-bearing mice, the leptin-treated mature DC (mDC) group exhibited a significant reduction in the expression of MMP9 (0.33-fold, p = 0.01) and CCL5 (0.81-fold, p = 0.02). The leptin-LPS-treated mDC group showed decreased expression of genes involved in metastasis and tumor growth, including VEGF (0.72-fold, p = 0.03), MMP9 (0.26-fold, p = 0.001), and CCL5 (0.3-fold, p = 0.006), indicating more efficient prevention of metastasis. The CCL2 gene expression levels in both treatment groups showed a slight decreasing trend, but these changes were not statistically significant. The leptin-treated mDC group reduced IL-6 production by approximately 16% (p = 0.02), while treatment with the leptin-LPS-treated mDC significantly decreased IL-6 production by approximately 22% (p = 0.01) and increased IL-33 production by approximately 42% (p = 0.03). The findings of the present study indicate that the leptin-LPS-treated mDC vaccine group reduced the expression of genes and cytokines involved in metastasis and angiogenesis, demonstrating greater efficacy compared to the leptin-treated mDC vaccine group.

EXTH-80. A NOVEL BISAMINOQUINOLINE DERIVATIVE NANOPARTICLE (BAQ13-NP) FOR THE TREATMENT OF GLIOBLASTOMA

Abstract Aminoquinolines, such as hydroxycholoquine, are known to inhibit autophagy and slow cancer cell growth, improving survival in preclinical models of glioblastoma. Unfortunately, they failed to demonstrate benefit in clinical trials due to a lack of sufficient potency and BBB penetration. We have developed a novel bisaminoquinoline derivative packaged in a nanoparticle (BAQ13-NP) with improved potency and intratumoral accumulation after systemic administration, confirmed by distribution and toxicity studies in animals. In this study, we evaluated the impact of BAQ13-NP on tumor cell autophagy and survival using patient-derived glioma cell lines and murine models. GBM patient-derived cell lines (n=4) and murine glioma cells (GL261, CT-2A) were treated with BAQ13-NP (1-4 µM) in culture, demonstrating growth restriction to less than 50% of control at 5-7 days (p<0.01) by MTT assay. Cells treated with BAQ13-NP also demonstrated reduced autophagy with significant increases in accumulation of LC3B and p62 (p<0.01) and reduced IL-6 dependent STAT3 phosphorylation in tumor cells, T cells, and myeloid cells (p<0.01). In addition, treatment of tumor cells with BAQ13-NP reduced IL-6 production over 50% (p<0.01). Mice with orthotopic GL261 or CT2A tumors were treated with BAQ13-NP by tail vein injection (25 mg/kg q2 days) starting 7 days post implantation. Significantly increased survival was observed in BAQ13-NP treated GL261 (HR 0.75, p=0.030) and CT2A (HR 0.78, p=0.033) bearing mice compared to vehicle control treated animals. Reduced tumor size at specific intervals was also observed with BAQ13-NP treatment, corelating with improved survival. BAQ13-NP can inhibit autophagy and slow glioma tumor growth, improving survival in preclinical models. Given the excellent distribution and low toxicity of this novel agent when delivered intravenously, we believe it has great potential to improve outcomes for GBM patients. Our drug has recently received IND approval and will soon be tested in a phase I clinical trial.

Cohesin-mediated chromatin remodeling controls the differentiation and function of conventional dendritic cells.

The cohesin protein complex extrudes chromatin loops, stopping at CTCF-bound sites, to organize chromosomes into topologically associated domains, yet the biological implications of this process are poorly understood. We show that cohesin is required for the post-mitotic differentiation and function of antigen-presenting dendritic cells (DCs), particularly for antigen cross-presentation and IL-12 secretion by type 1 conventional DCs (cDC1s) in vivo. The chromatin organization of DCs was shaped by cohesin and the DC-specifying transcription factor IRF8, which controlled chromatin looping and chromosome compartmentalization, respectively. Notably, optimal expression of IRF8 itself required CTCF/cohesin-binding sites demarcating the Irf8 gene. During DC activation, cohesin was required for the induction of a subset of genes with distal enhancers. Accordingly, the deletion of CTCF sites flanking the Il12b gene reduced IL-12 production by cDC1s. Our data reveal an essential role of cohesin-mediated chromatin regulation in cell differentiation and function in vivo, and its bi-directional crosstalk with lineage-specifying transcription factors.

Renal protective and immunoregulatory effects of Lactobacillus casei strain Shirota in nephropathy-prone mice.

The incidence of severe acute kidney injury (AKI) is considerably high worldwide. A previous study showed that gut microbial dysbiosis was a hallmark of AKI in mice. Whether the probiotic Lactobacillus casei strain Shirota (LcS) plays a role in kidney disease, particularly AKI, remains unclear. To investigate the effects of LcS on kidney injury, tubule-specific conditional von Hippel-Lindau gene-knockout C57BL/6 mice (Vhlhdel/del mice) were supplemented without (Ctrl) or with probiotics (LcS) in Experiment 1, and their lifespan was monitored. Additionally, the Vhlhdel/+ mice were supplemented without (Ctrl and AA) or with probiotics (LcS and LcS + AA) in Experiment 2. Probiotic LcS (1 × 109 colony-forming units) was supplemented once daily. After 4 weeks of LcS supplementation, AA and LcS + AA mice were administered aristolochic acid (AA; 4 mg/kg body weight/day)-containing purified diet for 2 weeks to induce AA nephropathy before sacrifice. Supplementation of LcS significantly prolonged the lifespan of Vhlhdel/del mice, suggesting a potential renal protective effect. AA induced-nephropathy increased not only the indicators of renal dysfunction and injury, including urinary protein and kidney injury molecule (KIM)-1, serum blood urea nitrogen (BUN) and creatinine, but also serum interleukin (IL)-6 levels, renal macrophage infiltrations, and pathological lesions in Vhlhdel/+ mice. LcS supplementation significantly reduced urinary protein and KIM-1 levels, serum BUN and IL-6 levels, and renal M1 macrophages, tissue lesions, and injury scores. We also found that LcS maintained gut integrity under AA induction and increased intestinal lamina propria dendritic cells. Furthermore, LcS significantly reduced pro-inflammatory IL-17A and upregulated anti-inflammatory IL-10 production by immune cells from intestinal Peyer's patches (PP) or mesenteric lymph nodes (MLN), and significantly increased IL-10 and reduced IL-6 production by splenocytes. Prior supplementation with probiotic LcS significantly alleviated the severity of renal injury. This renal protective effect was partially associated with the enhancements of intestinal and systemic anti-inflammatory immune responses, suggesting that LcS-induced immunoregulation might contribute to its renal protective effects.

MPoMA protects against lung epithelial cell injury via p65 degradation

Numerous cases of lung injury caused by viral infection were reported during the coronavirus disease-19 pandemic. While there have been significant efforts to develop drugs that block viral infection and spread, the development of drugs to reduce or reverse lung injury has been a lower priority. This study aimed to identify compounds from a library of compounds that prevent viral infection that could reduce and prevent lung epithelial cell damage. We investigated the cytotoxicity of the compounds, their activity in inhibiting viral spike protein binding to cells, and their activity in reducing IL-8 production in lung epithelial cells damaged by amodiaquine (AQ). We identified N-(4-(4-methoxyphenoxy)-3-methylphenyl)-N-methylacetamide (MPoMA) as a non-cytotoxic inhibitor against viral infection and AQ-induced cell damage. MPoMA inhibited the expression of IL-8, IL-6, IL-1β, and fibronectin induced by AQ and protected against AQ-induced morphological changes. However, MPoMA did not affect basal IL-8 expression in lung epithelial cells in the absence of AQ. Further mechanistic analysis confirmed that MPoMA selectively promoted the proteasomal degradation of inflammatory mediator p65, thereby reducing intracellular p65 expression and p65-mediated inflammatory responses. MPoMA exerted potent anti-inflammatory and protective functions in epithelial cells against LPS-induced acute lung injury in vivo. These findings suggest that MPoMA may have beneficial effects in suppressing viral infection and preventing lung epithelial cell damage through the degradation of p65 and inhibition of the production of inflammatory cytokines.

Ginseng-Epimedii formula ameliorated experimental Sjögren’s syndrome via reducing IL-6 production

Medicinal herbs have been recently investigated to explore their therapeutic potential on autoimmune disorders, as an alternative treatment to relieve disease symptoms. Here, we sought to determine the immunomodulatory function of a Chinese medicine formula Ginseng-Epimedii (GE), constituted by Lycium barbarum, Ginseng, Eleutherococcus senticosus and Epimedium brevicornum, on mice with experimental Sjögren’s syndrome (ESS). We found that GE-treated ESS mice exhibited improved salivary gland function and decreased serum levels of autoantibodies. In line with the network pharmacology analysis, GE treatment suppressed IL-6 production by B cells and conventional dendritic cells, associated with decreased effector T cell subsets. Using co-culture assay and adoptive transfer model, we demonstrated that splenoctyes from ESS mice receiving GE treatment showed impaired capacity to drive pathogenic Th17 and T follicular helper cell differentiation. Thus, this study suggested the therapeutic potential of GE formula in treating SS, by skewing the microenvironment that unfavored effector T cell responses.

In-vitro, In-vivo Study of Jatropha curcas Leaves Extract, and Preparation of a Nanoemulsion by a Low-energy/Solvent-free Method, In-silico Study of β-Sitosterol in Ulcerative Colitis

Introduction: Ulcerative colitis (UC) is characterized by recurring colon inflammation, leading to severe symptoms and decreased quality of life. Jatropha curcas, a traditional remedy for UC, particularly its active component β-sitosterol, is investigated here for its potential therapeutic benefits. Methods: The study evaluated J. curcas leaf extract’s effects on RAW 264.7 cell lines in-vitro, demonstrating a significant reduction in IL-6 production and promising anti-UC properties. In-vivo experiments on BALB/C mice with DSS-induced colitis confirmed dose-dependent reductions in disease severity, colon tissue damage, and pro-inflammatory cytokine levels. Additionally, a solvent-free nanoemulsion of J. curcas leaf extract was developed, exhibiting enhanced stability, encapsulation efficiency, and sustained release of active components, offering a promising drug delivery strategy for UC therapy. This interdisciplinary research contributes valuable insights into the therapeutic potential of J. curcas leaf extract and its nanoemulsion for UC treatment. Molecular docking simulations suggest strong binding affinities between β-sitosterol and inflammatory mediators, shedding light on its anti-inflammatory mechanisms. Pharmacokinetic and toxicity profiling further support the development of J. curcas-based UC therapeutics, laying a solid foundation for future studies and clinical applications.

Cuban Policosanol (Raydel®) Exerts Higher Antioxidant and Anti-Glycation Activities than Chinese Policosanol (BOC Sciences) in Reconstituted High-Density Lipoproteins: In Vivo Anti-Inflammatory Activities in Zebrafish and Its Embryos.

The present study compares sugarcane-wax purified policosanols sourced from Cuba (Raydel®) and China (BOC Sciences) and utilized following the synthesis of reconstituted high-density lipoproteins (rHDL). The two policosanols exhibited distinctly different ingredient ratios of long-chain aliphatic alcohols, particularly 1-octacosanol (C28) and 1-tetratriacotanol (C34). After synthesizing rHDL with apolipoprotein A-I (apoA-I), the two policosanols bound well with phospholipid and apoA-I to form the discoidal rHDL. Notably, rHDL-1, containing Cuban policosanol, displayed the largest particle diameter at approximately 78 ± 3 nm. In contrast, both control rHDL (rHDL-0) and rHDL containing Chinese policosanol (rHDL-2) exhibited smaller particles, with diameters of approximately 58 ± 3 nm and 61 ± 2 nm, respectively. Furthermore, rHDL-1 demonstrated enhanced anti-glycation activity, safeguarding apoA-I from degradation within HDL, and displayed the antioxidant ability to inhibit LDL oxidation. A microinjection of each rHDL into zebrafish embryos in the presence of carboxymethyllysine (CML) revealed rHDL-1 to have the strongest antioxidant activity with the highest embryo survivability and normal developmental morphology. Dermal application to recover the wound revealed rHDL-1 to have the highest wound-healing activity (75%) and survivability (92%) in the cutaneous wound area in the presence of CML. In adult zebrafish, injecting CML (250 μg) caused acute death and hyperinflammation, marked by heightened neutrophil infiltration and interleukin (IL)-6 production in liver. However, co-administering rHDL-1 notably increased survival (85%) and exhibited strong anti-inflammatory properties, reducing IL-6 production while improving the blood lipid profile. However, a co-injection of rHDL-2 resulted in the lowest survivability (47%) with more hepatic inflammation. In conclusion, Cuban policosanol (Raydel®) has more desirable properties for the in vitro synthesis of rHDL with stronger anti-glycation and antioxidant activities than those of Chinese policosanol (BOC Sciences). Moreover, Raydel-policosanol-integrated rHDL demonstrates a noteworthy effect on accelerated wound healing and robust anti-inflammatory properties, leading to increased survivability in zebrafish embryos and adults by effectively suppressing CML-induced hyperinflammation.

The NLRP3 molecule influences the therapeutic effects of mesenchymal stem cells through Glut1-mediated energy metabolic reprogramming

IntroductionNumerous studies demonstrated that NLRP3 has been implicated in the pathogenesis of inflammatory bowel disease (IBD). Mesenchymal stem cells (MSCs) regulated the NLRP3 inflammasome, which has emerged as a novel therapeutic approach for treating IBD. ObjectivesThe exact role of NLRP3 in regulating MSCs’ function is unclear. Our study aimed to explore how NLRP3 affects the therapeutic effects of MSCs in colitis. MethodsWe extracted MSCs from the bone marrow of C57BL/6 mice and Nlrp3 KO mice, and identified them using differentiation assays and flow cytometry. In vitro, Both WT MSCs and Nlrp3 KO MSCs were stimulated with inflammatory factor Lipopolysaccharide (LPS), and only WT MSCs were stimulated with varying concentrations of the NLRP3 inhibitor MCC950, then, quantified IL-10 levels in the supernatant. RNA-seq was performed to examine gene expression patterns and Seahorse was used to assess oxidative phosphorylation (OXPHOS) and glycolysis levels. Western blot was used to evaluate protein expression. In vivo, we treated DSS-induced colitis with either WT or Nlrp3 KO MSCs, monitoring weight, measuring colon length, and further evaluation. We also treated DSS-induced colitis with pretreated MSCs (BAY876, oe-Glut1, or oe-NLRP3), following the same experimental procedures as described above. ResultsOur results demonstrate that Nlrp3 deletion did not affect MSC phenotypes, but rather promoted osteogenic differentiation. However, the absence of Nlrp3 reduced IL-10 production in MSCs in the presence of LPS, leading to impaired protection on DSS-induced colitis. Conversely, overexpression of NLRP3 promotes the production of IL-10, enhancing therapeutic effects. Further investigation revealed that Nlrp3 deficiency downregulated Glut1 expression and glycolysis activation in MSCs, resulting in decreased IL-10 production. Notably, overexpressing Glut1 in Nlrp3 KO MSCs restored their therapeutic effect that was previously dampened due to Nlrp3 deletion. ConclusionOur findings demonstrate that NLRP3 heightens the therapeutic effects of MSC treatment on DSS-induced colitis.

Ligilactobacillus salivarius CNCM I-4866, a potential probiotic candidate, shows anti-inflammatory properties in vitro and in vivo

IntroductionThe aim of this work was to characterize a new strain of Ligilactobacillus salivarius (CNCM I-4866) (CNCM I-4866) to address its potential as probiotic with a special focus on intestinal inflammation. Potential anti-inflammatory abilities of this strain were evaluated through in vivo and in vitro experiments.MethodsFirstly, the strain was tested in a murine acute inflammation colitis model induced by DNBS. In vitro characterization was then performed with diverse tests: modulation capability of intestinal permeability; study of the impact on immunity profile through cytokines dosage; capacity to inhibit pathogens and adhere to intestinal cells lines. Production of metabolites, antibiotic resistance and survival to gastro-intestinal tract conditions were also tested.ResultsIn vitro assay has shown a reduction of colonic damage and markers of inflammation after treatment with CNCM I-4866. Transcriptomic analysis performed on colons showed the capacity of the strain to down-regulate pro-inflammatory cytokines. L. salivarius CNCM I-4866 exerted anti-inflammatory profile by reducing IL-8 production by TNF-α stimulated cell and modulated cytokines profile on peripheral blood mononuclear cells (PBMC). It protected intestinal integrity by increasing trans-epithelial electrical resistance (TEER) on Caco-2 TNF-α inflamed cells. Additionally, L. salivarius CNCM I-4866 displayed inhibition capacity on several intestinal pathogens and adhered to eukaryotic cells. Regarding safety and technical concerns, CNCM I-4866 was highly resistant to 0.3% of bile salts and produced mainly L-lactate. Finally, strain genomic characterization allowed us to confirm safety aspect of our strain, with no antibiotic gene resistance found.DiscussionTaken together, these results indicate that L. salivarius CNCM I-4866 could be a good probiotic candidate for intestinal inflammation, especially with its steady anti-inflammatory profile.

ER-α36 is involved in calycosin inhibition of IL-6 production in macrophages.

The tumour microenvironment (TME) is crucial for tumour development and progression. Tumour-associated macrophages (TAMs) in the TME can promote tumour progression and metastasis by releasing cytokines, such as IL-6. Calycosin, a phytoestrogen that is one of the active compounds in Radix Astragali, has been shown to inhibit tumour growth and metastasis. However, the underlying mechanism by which calycosin inhibits tumour growth remains unclear. Thus, this study aimed to investigate the effect of calycosin on IL-6 production in peripheral blood mononuclear cell (PBMC)- and THP-1-derived macrophages and explore its potential mechanisms using co-immunoprecipitation, western blotting, immunofluorescence, chromatin immunoprecipitation and luciferase assays. We found that calycosin treatment substantially upregulated the expression of ER-α36, a variant of the ER, and reduced IL-6 production in macrophages. Mechanistically, ER-α36 physically interacted with NF-κBp65 and retained p65 in the cytoplasm to attenuate NF-κB function as an IL-6 transcriptional inducer. In conclusion, our result indicated that calycosin inhibited IL-6 production by enhancing ER-α36 expression and its interaction with p65, which attenuated NF-κB function as an IL-6 inducer. Therefore, calycosin can be developed as an effective agent for cancer therapy by targeting TAMs.

Human HLA prolongs the host inflammatory response in Streptococcus suis serotype 2 infection compared to mouse H2 molecules.

Streptococcus suis (S. suis) is widely acknowledged as a significant zoonotic pathogen in Southeast Asia and China, which has led to a substantial number of fatalities in both swine and humans. Despite the prevalent use of mice as the primary animal model to study S. suis pathogenesis, the substantial differences in the major histocompatibility complex (MHC) between humans and mice underscore the ongoing exploration for a more suitable and effective animal model. In this study, humanized transgenic HLA-A11/DR1 genotypes mice were used to evaluate the differences between humanized HLA and murine H2 in S. suis infection. Following intravenous administration of S. suis suspensions, we investigated bacterial load, cytokine profiles, pathological alterations, and immune cell recruitment in both Wild-type (WT) and humanized mice across different post-infection time points. Relative to WT mice, humanized mice exhibited heightened pro-inflammatory cytokines, exacerbated tissue damage, increased granulocyte recruitment with impaired resolution, notably more pronounced during the late infection stage. Additionally, our examination of bacterial clearance rates suggests that HLA-A11/DR1 primarily influences cell recruitment and mitochondrial reactive oxygen species (ROS) production, which affects the bacterial killing capacity of macrophages in the late stage of infection. The reduced IL-10 production and lower levels of regulatory T cells in humanized mice could underlie their compromised resolution ability. Intervention with IL-10 promotes bacterial clearance and inflammatory regression in the late stages of infection in transgenic mice. Our findings underscore the heightened sensitivity of HLA-A11/DR1 mice with impaired resolution to S. suis infection, effectively mirroring the immune response seen in humans during infection. The humanized HLA-A11/DR1 mice could serve as an optimal animal model for investigating the pathogenic and therapeutic mechanisms associated with sepsis and other infectious diseases.

Optimizing extraction of Pelargonium sidoides roots: Impact of ethanol concentration on biological activity of extracts

Pelargonium sidoides DC. (Geraniaceae) is an important indigenous medicinal plant in South Africa, historically employed by various ethnic groups to treat respiratory and gastrointestinal ailments. The proprietary herbal tincture, Umckaloabo®, utilizes an ethanolic extract (EPs® 7630) from the roots of P. sidoides, and has demonstrated effectiveness in alleviating symptoms of respiratory infections.P. sidoides roots contain numerous highly oxygenated coumarins and phenolic metabolites, notably the marker compound umckalin. While Umckaloabo® is prepared using 11 % ethanol extraction, the South African commercial market predominantly employs 60 % ethanol extraction due to its consistent umckalin yield and antimicrobial activity. The effects of these extraction methods on chemical composition and in vitro pharmacological activity remain poorly understood.This study confirmed superior antibacterial and antifungal activity in the 60 % ethanol extracts prepared from six root samples compared to the 11 % ethanol extracts. MIC values for both Gram-negative and Gram-positive bacteria ranged from 0.078 to 2.5 mg/mL in the 60 % extracts. Remarkably, two of the root extracts exhibited excellent to very good activity against C. albicans and C. neoformans with MIC values of 0.039 and 0.078 mg/mL, respectively.A positive correlation was identified between total minimum inhibitory concentration (MIC) levels and polyphenol content, rather than umckalin levels. The influence of these extracts on the pro-inflammatory cytokine IL-6 was assessed. Both 60 % and 11 % root extracts of P. sidoides at 100 µg/mL significantly reduced IL-6 production, with the 60 % extracts demonstrating a more pronounced effect (p = 0.008).Statistical analysis revealed significant differences between 11 % and 60 % ethanol extractions in terms of average MIC overall, average Gram-negative MIC, average Gram-positive MIC, fungal MIC, and IL-6 levels. Extracts with higher polyphenol values exhibited superior antimicrobial activity and antioxidant potential, suggesting that polyphenol content may serve as a more reliable indicator of antimicrobial activity than umckalin levels. Additionally, polyphenol levels in the roots may vary with altitude and other environmental factors, warranting further research.

Alleviating psoriatic skin inflammation through augmentation of Treg cells via CTLA-4 signaling peptide.

Psoriasis is a chronic inflammatory skin disease characterized by hyperplasia of keratinocytes and immune cell infiltration. The IL-17-producing T cells play a key role in psoriasis pathogenesis, while regulatory T (Treg) cells are diminished during psoriatic inflammation. Current psoriasis treatments largely focus on IL-17 and IL-23, however, few studies have explored therapeutic drugs targeting an increase of Treg cells to control immune homeostasis. In this study, we investigated the effects of a cytotoxic T lymphocyte antigen-4 (CTLA-4) signaling peptide (dNP2-ctCTLA-4) in Th17, Tc17, γδ T cells, Treg cells in vitro and a mouse model of psoriasis. Treatment with dNP2-ctCTLA-4 peptide showed a significant reduction of psoriatic skin inflammation with increased Treg cell proportion and reduced IL-17 production by T cells, indicating a potential role in modulating psoriatic skin disease. We compared dNP2-ctCTLA-4 with CTLA-4-Ig and found that only dNP2-ctCTLA-4 ameliorated the psoriasis progression, with increased Treg cells and inhibited IL-17 production from γδ T cells. In vitro experiments using a T cell-antigen presenting cell co-culture system demonstrated the distinct mechanisms of dNP2-ctCTLA-4 compared to CTLA-4-Ig in the induction of Treg cells. These findings highlight the therapeutic potential of dNP2-ctCTLA-4 peptide in psoriasis by augmenting Treg/Teff ratio, offering a new approach to modulating the disease.

Citri Reticulatae Pericarpium Limits TLR-4-Triggered Inflammatory Response in Raw264.7 Macrophages by Activating RasGRP3.

Inflammation is an important immune response to pathogen invasion, but excessive inflammation leads to tissue injury and even cytokine storm. Therefore, proper response is needed depending on the intensity of the infection. Ras guanine nucleotide releasing protein 3 (RasGRP3) is a regulator of the TLR-mediated response. In low-intensity inflammation, it negatively regulates production of pro-inflammatory cytokines, especially IL-6. Citri Reticulatae Pericarpium, the peel of Citrus reticulata Blanco, is a major medicinal herb in Korean medicine. The present study aims to investigate whether the Citri Reticulatae Pericarpium extract (CRE) has immunomodulatory activity using the Raw264.7 macrophage. Also, we investigated the effect of CRE on RasGRP3 expression. In the present study, CRE reduced IL-6 production in the low-LPS environment (1 ng/mL) and did not in the high-LPS environment (100 ng/mL). The suppression of IL-6 production in the low-LPS environment (1 ng/mL) was abolished after the pretreatment of RasGRP3 siRNA. The reduced RasGRP3 protein content by 100 ng/mL LPS treatment was increased by CRE treatment. Additionally, nobiletin, a major component of CRE showed a suppressive effect on IL-6 production in the low-LPS environment (1 ng/mL). The present results suggest that CRE alleviates inflammatory response via activating RasGRP3 expression in low-intensity inflammation.

Dupilumab strengthens herpes simplex virus type 1–specific immune responses in atopic dermatitis

Impaired virus clearance in a subgroup of atopic dermatitis (AD) patients can lead to severe herpes simplex virus (HSV) infections called eczema herpeticum (EH). We recently identified a type 2 skewed viral immune response in EH patients. Clinical data suggest a reduced incidence of EH in AD patients treated with dupilumab, although immunologic investigations of this phenomenon are still lacking. We examined the impact of dupilumab on the HSV type 1 (HSV-1) specific immune response in AD, focusing on patients with (ADEH+) and without (ADEH-) a history of EH. Sera and peripheral blood mononuclear cells were collected from ADEH+ and ADEH- patients, a subgroup of whom was receiving dupilumab treatment, and healthy controls. Serum samples were tested for IgE against HSV-1 glycoprotein D (n= 85). Peripheral blood mononuclear cells were stimulated with HSV peptides, and activated CD4+ and CD8+ cells were characterized by flow cytometry after magnetic enrichment via CD154 or CD137 (n= 60). Cytokine production of HSV-1-reactive T-cell lines (n= 33) and MHC-I tetramer+ (HSV-1-UL25) CD8+ T cells was investigated by bead assay and intracellular cytokine staining (n= 21). We confirmed that HSV-1-specific IgE is elevated in ADEH+ patients. During dupilumab treatment, the IgE levels were significantly decreased, reaching levels of healthy controls. HSV-1-specific TC1 frequencies were elevated in ADEH- patients treated with dupilumab compared to dupilumab-negative patients. There were no changes in the frequencies of HSV-1-specific TH cells while receiving dupilumab therapy. AD patients receiving dupilumab exhibited elevated IFN-γ and reduced IL-4 production in HSV-1-UL25-epitope-specific T cells compared to dupilumab-negative patients. Dupilumab may improve the HSV-1-specific immune response in AD as a result of an increased type I immune response and a reduction of HSV-1-specific IgE.

Bacitracin Supplementation as a Growth Promoter Down-Regulates Innate and Adaptive Cytokines in Broilers’ Intestines

In the past decade, the withdrawal of antibiotics used as growth promoters (AGP) has increased some poultry industry challenges, such as the rise of intestinal diseases. Experts advocate that AGPs improve performance due to the modulation of the intestinal microbiota, with resulting anti-inflammatory effects. However, the impact and interactions of AGPs with the host intestinal immune system are still unknown, which represents issues in developing effective alternatives for AGPs. Therefore, this study was aimed at better understanding the potential mechanism of action of bacitracin used as AGP and its impacts on the intestinal immune system. Ninety day-of-hatch chickens were randomly assigned to two treatments with three repetitions of fifteen birds, a control (CNT) group with a corn/soybean meal standard diet, and a control diet supplemented with 50 g/ton of feed of bacitracin (BMD). The cytokines’ and chemokines’ production (IFN-α, IFN-γ, IL-16, IL-10, IL-21, IL-6, M-CSF, MIP-3α, MIP-1β, VEGF and CCL-5) were assessed in the jejunum and ileum at 14, 21, 28 and 36 days of age by using a chicken-specific cytokine/chemokine peptide ELISA array. Broilers with BMD supplementation were found to have a lower production of IL-16, IFN-γ, M-CSF, IL-21, MIP-1β and VEGF in the jejunum at 14 d. However, from 21 through 36 days, the effect of BMD on cytokine production in the jejunum was negligible except for CCL-5, which was reduced at D36. In the ileum, BMD effects on the cytokine profile started at 28 d, when BMD-supplemented broilers showed a reduced IL-6 production level. At day 36, BMD reduced IL-16 and MIP-3α production but increased VEGF concentration in the ileum tissue. The present study demonstrated that the use of bacitracin as an AGP modulates the small intestine immune system, especially in the first phase of the broiler’s life (up to 14 days). Moreover, BMD anti-inflammatory effects include not only innate immunity but also seemed to influence the development of the adaptive immune response as seen by the decreased production of IL-21 and IL-16. These results demonstrate that a commonly used AGP in broiler feed had a local anti-inflammatory effect.

Effects of Lidocaine-Derived Organic Compounds on Eosinophil Activation and Survival.

Lidocaine, a local anesthetic, is known to possess anti-inflammatory properties. However, its clinical use is limited by inconveniences, such as its local synesthetic effects. This study evaluated lidocaine analogs designed and synthesized to overcome the disadvantages of lidocaine, having anti-inflammatory properties. Interleukin 5 (IL-5)-induced eosinophil activation and survival were evaluated using 36 lidocaine analogs with modified lidocaine structure on the aromatic or the acyl moiety or both. Eosinophil survival was evaluated using a CellTiter 96® aqueous cell proliferation assay kit. Superoxide production was determined using the superoxide dismutase-inhibitable reduction of cytochrome C method. Eosinophil cationic protein (ECP), IL-8, and transcription factor expression were determined using enzyme-linked immunosorbent assay. The platelet-activating factor (PAF)-induced migration assay was performed using a Transwell insert system. Compounds EI137 and EI341 inhibited IL-5-induced eosinophil survival and superoxide and ECP production in a concentration-dependent manner. These compounds also significantly reduced IL-8 production. Although compounds EI137 and EI341 significantly reduced phosphorylated ERK 1/2 expression, they did not influence other total and phosphorylated transcription factors. Moreover, 1000 µM of compound EI341 only inhibited PAF-induced migration of eosinophils. Lidocaine analogs EI137 and EI341 inhibited IL-5-mediated activation and survival of eosinophils. These compounds could be new therapeutic agents to treat eosinophilic inflammatory diseases.

Stimulation of ectopically expressed muscarinic receptors induces IFN-γ but suppresses IL-2 production by inhibiting activation of pAKT pathways in primary T cells

T cell antigen receptor stimulation induces tyrosine phosphorylation of downstream signaling molecules and the phosphatidylinositol, Ras, MAPK, and PI3 kinase pathways, leading to T cell activation. Previously, we reported that the G-protein-coupled human muscarinic receptor could bypass tyrosine kinases to activate the phosphatidylinositol pathway and induce interleukin-2 production in Jurkat leukemic T cells. Here, we demonstrate that stimulating G-protein-coupled muscarinic receptors (M1 and synthetic hM3Dq) can activate primary mouse T cells if PLCβ1 is coexpressed. Resting peripheral hM3Dq+PLCβ1 (hM3Dq/β1) T cells did not respond to clozapine, an hM3Dq agonist, unless they were preactivated by TCR and CD28 stimulation which increased hM3Dq and PLCβ1 expression. This permitted large calcium and phosphorylated ERK responses to clozapine. Clozapine treatment induced high IFN-γ, CD69, and CD25 expression, but surprisingly did not induce substantial IL-2 in hM3Dq/β1 T cells. Importantly, costimulation of both muscarinic receptors plus the TCR even led to reduced IL-2 expression, suggesting a selective inhibitory effect of muscarinic receptor costimulation. Stimulation of muscarinic receptors induced strong nuclear translocation of NFAT and NFκB and activated AP-1. However, stimulation of hM3Dq led to reduced IL-2 mRNA stability which correlated with an effect on the IL-2 3'UTR activity. Interestingly, stimulation of hM3Dq resulted in reduced pAKT and its downstream pathway. This may explain the inhibitory impact on IL-2 production in hM3Dq/β1T cells. Moreover, an inhibitor of PI3K reduced IL-2 production in TCR-stimulated hM3Dq/β1 CD4 T cells, suggesting that activating the pAKT pathway is critical for IL-2 production in T cells.