THP-1 Macrophages Research Articles

Synthesis of Nitrostyrylthiazolidine-2,4-dione Derivatives Displaying Antileishmanial Potential.

A series of 61 thiazolidine-2,4-diones bearing a styryl group at position 5 was synthesized in 2-5 steps and their structure was proved by elemental and spectral analyses. The compounds obtained were evaluated in vitro against the promastigote stage of the kinetoplastid parasite Leishmania infantum and the human HepG2 cell line, to determine selectivity indices and to compare their activities with those of antileishmanial reference drugs. The study of structure-activity relationships indicated the potential of some derivatives bearing a nitro group on the phenyl ring, especially when located at the meta position. Thus, among the tested series, compound 14c appeared as a hit compound with good antileishmanial activity (EC50 = 7 µM) and low cytotoxicity against both the hepatic HepG2 and macrophage THP-1 human cell lines (CC50 = 101 and 121 µM, respectively), leading to good selectivity indices (respectively, 14 and 17), in comparison with the reference antileishmanial drug compound miltefosine (EC50 = 3.3 µM, CC50 = 85 and 30 µM, SI = 26 and 9). Regarding its mechanism of action, among several possibilities, it was demonstrated that compound 14c is a prodrug bioactivated, predominantly by L. donovani nitroreductase 1, likely leading to the formation of cytotoxic metabolites that form covalent adducts in the parasite. Finally, compound 14c is lipophilic (measured CHI LogD7.7 = 2.85) but remains soluble in water (measured PBS solubility at pH7.4 = 16 µM), highlighting the antileishmanial potential of the nitrostyrylthiazolidine-2,4-dione scaffold.

LncRNA DANCR promotes macrophage lipid accumulation through modulation of membrane cholesterol transporters.

The progression of atherosclerosis (AS), the pathological foundation of coronary artery disease (CAD), is featured by massive lipid deposition in the vessel wall. LncRNAs are implicated in lipid disorder and AS, whereas the specific role of lncRNA DANCR in atherogenesis remains unknown. Here, we demonstrated that DANCR promotes macrophage lipid accumulation by regulating the expression of membrane cholesterol transport proteins. qPCR showed that compared to control groups, CAD patients and atherosclerotic mice had higher DANCR levels. Treating human THP-1 macrophages and mouse RAW264.7 macrophages with ox-LDL significantly upregulated the expression levels of DANCR. Oil Red O staining showed that the silence of DANCR robustly reduced, while overexpression of DANCR significantly increased the numbers and size of lipid droplets in ox-LDL-treated THP-1 macrophages. In contrast, the opposite phenomena were observed in DANCR overexpressing cells. The expression of ABCA1, ABCG1, SR-BI, and NBD-cholesterol efflux was increased obviously by DANCR inhibition and decreased by DANCR overexpression, respectively. Furthermore, transfection with DANCR siRNA induced a robust decrease in the levels of CD36, SR-A, and Dil-ox-LDL uptake, while DANCR overexpression amplified the expression of CD36, SR-A and the uptake of Dil-ox-LDL in lipid-laden macrophages. Lastly, we found that the effects of DANCR on macrophage lipid accumulation and the expression of membrane cholesterol transport proteins were not likely related to miR-33a. The present study unraveled the adverse role of DANCR in foam cell formation and its relationship with cholesterol transport proteins. However, the competing endogenous RNA network underlying these phenomena warrants further exploration.

Impact of refining on phytochemicals and anti-inflammatory activity of papaya (Carica papaya L.) seed oil in LPS-stimulated THP-1 cells

This study investigated the changes in phytochemical composition and inflammatory response of crude papaya (Carica papaya L.) seed oil (CPO) and its refined forms (degummed, PDG; deacidified, PDA; decolorized, PDC; deodorized, PDO). Oils were analyzed for their phytochemical composition, oil quality parameters, antioxidant activity, and their inflammatory response in LPS-stimulated THP-1 macrophages. At higher refining degrees, particularly after deacidification, the contents of phytochemicals (sterols, tocopherols, and polyphenols) decreased while oxidation products increased. Both CPO (0.1–1.0 mg/mL) and PDG reduced the secretion and mRNA expression of LPS-stimulated inflammatory cytokines and mediators and also blocked the activation of the NF-κB pathway. PDA, PDC, and PDO showed low anti-inflammatory or even pro-inflammatory activity. Correlation analysis showed that 4 polyphenols and 2 phytosterols were responsible for the oil's anti-inflammatory effects. These findings indicated that moderate refining is suggested for papaya seed oil processing for retaining bioactive ingredients and anti-inflammatory ability.

Ribonuclease inhibitor 1 emerges as a potential biomarker and modulates inflammation and iron homeostasis in sepsis

Sepsis, marked by organ dysfunction, necessitates reliable biomarkers. Ribonuclease inhibitor 1 (RNH1), a ribonuclease (RNase) inhibitor, emerged as a potential biomarker for acute kidney injury and mortality in thoracoabdominal aortic aneurysm patients. Our study investigates RNH1 dynamics in sepsis, its links to mortality and organ dysfunction, and the interplay with RNase 1 and RNase 5. Furthermore, we explore RNH1 as a therapeutic target in sepsis-related processes like inflammation, non-canonical inflammasome activation, and iron homeostasis. We showed that RNH1 levels are significantly higher in deceased patients compared to sepsis survivors and correlate with creatine kinase, aspartate and alanine transaminase, bilirubin, serum creatinine and RNase 5, but not RNase 1. RNH1 mitigated LPS-induced TNFα and RNase 5 secretion, and relative mRNA expression of ferroptosis-associated genes HMOX1, FTH1 and HAMP in PBMCs. Monocytes were identified as the predominant type of LPS-positive PBMCs. Exogenous RNH1 attenuated LPS-induced CASP5 expression, while increasing IL-1β secretion in PBMCs and THP-1 macrophages. As RNH1 has contradictory effects on inflammation and non-canonical inflammasome activation, its use as a therapeutic agent is limited. However, RNH1 levels may play a central role in iron homeostasis during sepsis, supporting our clinical observations. Hence, RNH1 shows promise as biomarkers for renal and hepatic dysfunction and hepatocyte injury, and may be useful in predicting the outcome of septic patients.

Broncho-Vaxom Attenuates Lipopolysaccharide-Induced Inflammation in a Mouse Model of Acute Lung Injury.

Acute lung injury (ALI) is a condition associated with acute respiratory failure, resulting in significant morbidity and mortality. It involves cellular changes such as disruption of the alveolar-capillary membrane, excessive neutrophil migration, and release of inflammatory mediators. Broncho-Vaxom® (BV), a lyophilized product containing cell membrane components derived from eight bacteria commonly found in the respiratory tract, is known for its potential to reduce viral and bacterial lung infections. However, the specific effect of BV on ALI has not been clearly defined. This study explored the preventive effects of BV and its underlying mechanisms in a lipopolysaccharide (LPS)-induced ALI mouse model. Oral BV (1 mg/kg) gavage was administered one hour before the intratracheal injection of LPS to evaluate its preventive effect on the ALI model. The pre-administration of BV significantly mitigates inflammatory parameters, including the production of inflammatory mediators, macrophage infiltration, and NF-κB activation in lung tissue, and the increase in inflammatory cells in bronchoalveolar lavage fluid (BALF). Moreover, BV (3 μg/mL) pretreatment reduced the expression of M1 macrophage markers, interleukins (IL-1β, IL-6), tumor necrosis factor α, and cyclooxygenase-2, which are activated by LPS, in both mouse alveolar macrophage MH-S cells and human macrophage THP-1 cells. These findings showed that BV exhibits anti-inflammatory effects by suppressing inflammatory mediators through the NF-κB pathway, suggesting its potential to attenuate bronchial and pulmonary inflammation.

Fragment-Based Anti-inflammatory Agent Design and Target Identification: Discovery of AF-45 as an IRAK4 Inhibitor to Treat Ulcerative Colitis and Acute Lung Injury.

UC and ALI are inflammatory diseases with limited treatment in the clinic. Herein, fragment-based anti-inflammatory agent designs were carried out deriving from cyclohexylamine/cyclobutylamine and several fragments from anti-inflammatory agents in our lab. AF-45 (IC50 = 0.53/0.60 μM on IL-6/TNF-α in THP-1 macrophages) was identified as the optimal molecule using ELISA and MTT assays from the 33 synthesized compounds. Through mechanistic studies and a systematic target search process, AF-45 was found to block the NF-κB/MAPK pathway and target IRAK4, a promising target for inflammation and autoimmune diseases. The selectivity of AF-45 targeting IRAK4 was validated by comparing its effects on other kinase/nonkinase proteins. In vivo, AF-45 exhibited a good therapeutic effect on UC and ALI, and favorable PK proprieties. Since there are currently no clinical or preclinical trials for IRAK4 inhibitors to treat UC and ALI, AF-45 provides a new lead compound or candidate targeting IRAK4 for the treatment of these diseases.

BMAL1 plays a crucial role in immune homeostasis during sepsis-induced acute lung injury

Sepsis is a widespread and life-threatening disease characterised by infection-triggered immune hyperactivation and cytokine storms, culminating in tissue damage and multiple organ dysfunction syndrome. BMAL1 is a pivotal transcription factor in the circadian clock that plays a crucial role in maintaining immune homeostasis. BMAL1 dysregulation has been implicated in inflammatory diseases and immunodeficiency. However, the mechanisms underlying BMAL1 disruption in sepsis-induced acute lung injury (ALI) remain poorly understood. In vitro, we used THP1 and mouse peritoneal macrophages to elucidate the potential mechanism of BMAL1 function in sepsis. In vivo, an endotoxemia model was used to investigate the effect of BMAL1 on sepsis and the therapeutic role of targeting CXCR2. We showed that BMAL1 significantly affected the regulation of innate immunity in sepsis-induced ALI. BMAL1 deficiency in the macrophages exacerbated systemic inflammation and sepsis-induced ALI. Mechanistically, BMAL1 acted as a transcriptional suppressor and regulated the expression of CXCL2. BMAL1 deficiency in macrophages upregulated CXCL2 expression, increasing the recruitment of polymorphonuclear neutrophils and the formation of neutrophil extracellular traps (NETs) by binding to the chemokine receptor CXCR2, thereby intensifying lung injury in a sepsis model. Furthermore, a selective inhibitor of CXCR2, SB225002, exerted promising therapeutic effects by markedly reducing neutrophil infiltration and NETs formation and alleviating lung injury. Importantly, CXCR2 blockade mitigated multiple organ dysfunction. Collectively, these findings suggest that BMAL1 controls the CXCL2/CXCR2 pathway, and the therapeutic efficacy of targeting CXCR2 in sepsis has been validated, presenting BMAL1 as a potential therapeutic target for lethal infections.

Identification of effective components in Flos Chrysanthemi Indici for alleviating gouty inflammation through integrated absorption and metabolism studies

Flos Chrysanthemi Indici (FCI) is a widely used edible flower and functional food resource, whose extract demonstrates a significant inhibitory effect on monosodium urate (MSU)-induced inflammation in mice. However, the main active components responsible for this effect have remained elusive. Understanding the bioavailability of these components is essential for ensuring the efficacy of functional foods. In this study, we aimed to identify the active components of FCI by examining their in vivo exposure, including absorbed compounds and their metabolites. Initially, we investigated the profiles of FCI extract and absorption and metabolism in rat plasma. Subsequently, through cellular experiments, we found that luteolin, apigenin, luteolin-3′-O-glucuronide, luteolin-7-O-glucuronide, acacetin-7-O-glucuronide, and apigenin-7-O-glucuronide exhibited anti-inflammatory effects against MSU-induced inflammation in THP-1 macrophage cells. The active-related parent compounds in FCI (linarin and luteolin-7-O-β-d-glucoside) were considered as quality control standards for the development of functional products targeting gout. This study characterized the active polyphenolics of FCI in vivo, which may contribute to its reported anti-gout properties, providing valuable insights into the health benefits of FCI polyphenols.

Tracking Chlamydia – Host interactions and antichlamydial activity in Caenorhabditis elegans

The fading efficacy of antibiotics is a growing global health concern due to its life-threatening consequences and increased healthcare costs. Non-genetic mechanisms of antimicrobial resistance, such as those employed by Chlamydia pneumoniae and Chlamydia trachomatis, complicate treatment as these bacteria can enter a non-replicative, persistent state under stress, evading antibiotics and linking to inflammatory conditions. Understanding chlamydial persistence at the molecular level is challenging, and new models for studying Chlamydia-host interactions in vivo are urgently needed. Caenorhabditis elegans offers an alternative given its immune system and numerous orthologues of human genes. This study established C. elegans as an in vivo model for chlamydial infection. Both Chlamydia species reduced the worm's lifespan, their DNA being detectable at three- and six-days post-infection. Azithromycin at its MIC (25 nM) failed to prevent the infection-induced lifespan reduction, indicating a persister phenotype. In contrast, the methanolic extract of Schisandra chinensis berries showed anti-chlamydial activity both in vitro (in THP-1 macrophages) and in vivo, significantly extending the lifespan of infected C. elegans and reducing the bacterial load. Moreover, S. chinensis increased the transcriptional activity of SKN-1 in the worms, but was unable to impact the bacterial load or lifespan in a sek-1 defective C. elegans strain. In summary, this study validated C. elegans as a chlamydial infection model and showcased S. chinensis berries' in vivo anti-chlamydial potential, possibly through SEK/SKN-1 signaling modulation.

Engineered CD147-Deficient THP-1 Impairs Monocytic Myeloid-Derived Suppressor Cell Differentiation but Maintains Antibody-Dependent Cellular Phagocytosis Function for Jurkat T-ALL Cells with Humanized Anti-CD147 Antibody.

CD147 is upregulated in cancers, including aggressive T-ALL. Traditional treatments for T-ALL often entail severe side effects and the risk of relapse, highlighting the need for more efficacious therapies. ADCP contributes to the antitumor response by enhancing the ability of phagocytic cells to engulf cancer cells upon antibody binding. We aimed to engineer CD147KO THP-1 cells and evaluated their differentiation properties compared to the wild type. A humanized anti-CD147 antibody, HuM6-1B9, was also constructed for investing the phagocytic function of CD147KO THP-1 cells mediated by HuM6-1B9 in the phagocytosis of Jurkat T cells. The CD147KO THP-1 was generated by CRISPR/Cas9 and maintained polarization profiles. HuM6-1B9 was produced in CHO-K1 cells and effectively bound to CD147 with high binding affinity (KD: 2.05 ± 0.30 × 10-9 M). Additionally, HuM6-1B9 enhanced the phagocytosis of Jurkat T cells by CD147KO THP-1-derived LPS-activated macrophages (M-LPS), without self-ADCP. The formation of THP-1-derived mMDSC was limited in CD147KO THP-1 cells, highlighting the significant impact of CD147 deletion. Maintaining expression markers and phagocytic function in CD147KO THP-1 macrophages supports future engineering and the application of induced pluripotent stem cell-derived macrophages. The combination of HuM6-1B9 and CD147KO monocyte-derived macrophages holds promise as an alternative strategy for T-ALL.

TNFAIP1 promotes macrophage lipid accumulation and accelerates the development of atherosclerosis through the LEENE/FoxO1/ABCA1 pathway.

Macrophage lipid accumulation is a critical contributor to foam cell formation and atherosclerosis. Tumor necrosis factor-α-induced protein 1 (TNFAIP1) is closely associated with cardiovascular disease. However, its role and molecular mechanisms in atherogenesis remain unclear. TNFAIP1 was knocked down in THP-1 macrophage-derived foam cells and apolipoprotein-deficient (apoE-/-) mice using lentiviral vector. The expression of lncRNA enhancing endothelial nitric oxide synthase expression (LEENE), Forkhead box O1 (FoxO1) and ATP binding cassette transporter A1 (ABCA1) was evaluated by qRT-PCR and/or western blot. Lipid accumulation in macrophage was assessed by high-performance liquid chromatography and Oil red O staining. RNA immunoprecipitation and RNA pull-down assay were performed to verify the interaction between LEENE and FoxO1 protein. Atherosclerotic lesions were analyzed using HE, Oil red O and Masson staining. Our results showed that TNFAIP1 was significantly increased in THP-1 macrophages loaded with oxidized low-density lipoprotein. Knockdown of TNFAIP1 enhanced LEENE expression, promoted the direct interaction of LEENE with FoxO1 protein, stimulated FoxO1 protein degradation through the proteasome pathway, induced ABCA1 transcription, and finally suppressed lipid accumulation in THP-1 macrophage-derived foam cells. TNFAIP1 knockdown also up-regulated ABCA1 expression, improved plasma lipid profiles, enhanced the efficiency of reverse cholesterol transport and attenuated lesion area in apoE-/- mice. Taken together, these results provide the first direct evidence that TNFAIP1 aggravates atherosclerosis by promoting macrophage lipid accumulation via the LEENE/FoxO1/ABCA1 signaling pathway. TNFAIP1 may represent a promising therapeutic target for atherosclerotic cardiovascular disease.

Lysophosphatidylcholines Promote Influenza Virus Reproduction through the MAPK/JNK Pathway in PMA-Differentiated THP-1 Macrophages.

Obesity and metabolic syndrome alter serum lipid profiles. They also increase vulnerability to viral infections and worsen the survival rate and symptoms after infection. How serum lipids affect influenza virus proliferation is unclear. Here, we investigated the effects of lysophosphatidylcholines on influenza A virus (IAV) proliferation. IAV particles in the culture medium were titrated using extraction-free quantitative PCR, and viral RNA and protein levels were assessed using real-time PCR and Western blot, respectively. RNA sequencing data were analyzed using PCA and heatmap analysis, and pathway analysis was performed using the KEGG mapper and PathIN tools. Statistical analysis was conducted using SPSS21.0. LPC treatment of THP-1 cells significantly increased IAV proliferation and IAV RNA and protein levels, and saturated LPC was more active in IAV RNA expression than unsaturated LPC was. The functional analysis of genes affected by LPCs showed that the expression of genes involved in IAV signaling, such as suppressor of cytokine signaling 3 (SOCS3), phosphoinositide-3-kinase regulatory subunit 3 (PI3K) and AKT serine/threonine kinase 3 (AKT3), Toll-like receptor 7 (TKR7), and interferon gamma receptor 1 (IFNGR1), was changed by LPC. Altered influenza A pathways were linked with MAPK and PI3K/AKT signaling. Treatment with inhibitors of MAPK or PI3K attenuated viral gene expression changes induced by LPCs. The present study shows that LPCs stimulated virus reproduction by modifying the cellular environment to one in which viruses proliferated better. This was mediated by the MAPK, JNK, and PI3K/AKT pathways. Further animal studies are needed to confirm the link between LPCs from serum or the respiratory system and IAV proliferation.

Antileishmania, Immune Modulation and Apoptosis Induction by Astragalus ecbatanus Extract Against Leishmania tropica

Background: Due to the unique properties of Astragalus in treating diseases and strengthening the immune system, this study aimed to investigate, for the first time, the immune modulation and apoptosis induction by Astragalus ecbatanus extract against Leishmania tropica. Methods: The study assessed the in vitro efficacy of the Astragalus ecbatanus extract against both the promastigote and amastigote stages of L. tropica (MHOM/AF/88/KK27). Additionally, the effects of the extract on inducing nitric oxide (NO) release and its cytotoxicity on human macrophage cells were determined by calculating the 50% cytotoxic concentrations (CC50). The study also evaluated the Caspase-3-like activity in extract-treated parasites and conducted quantitative real-time PCR to assess the expression of genes associated with T lymphocytes. Results: Our study demonstrated that the A. ecbatanus extract significantly (P < 0.001) decreased the viability of both L. tropica promastigote and amastigote forms compared to the negative control. Moreover, the extract exhibited a high selectivity index (> 10), indicating its strong specificity towards intracellular parasites while showing low cytotoxicity to host cells. Our results indicated a dose-dependent upregulation of the expression levels of genes for inducible nitric oxide synthase (iNOS), interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNFα), and interleukin-10 (IL-10) in macrophages after exposure to A. ecbatanus ethyl acetate extract (P < 0.01). In contrast, the gene expression level of IL-10 exhibited a dose-dependent downregulation after exposure to A. ecbatanus ethyl acetate extract. We also found that the THP-1 macrophages exposed to the A. ecbatanus ethyl acetate extract exhibited enhanced production of nitric oxide (P < 0.001). Additionally, the ethyl acetate extract of A. ecbatanus significantly enhanced caspase-3 activation in Leishmania parasites (P < 0.01). Conclusions: The results demonstrated the significant impact of A. ecbatanus ethyl acetate extract on inhibiting and eradicating Leishmania parasites in laboratory settings. While some cellular mechanisms of action were identified, such as immune modulation and apoptosis induction against Leishmania parasites, further investigation is essential to elucidate the specific mechanisms of action and assess the efficacy in animal and human populations.

Contribution of Extracellular Particles Isolated from Morus sp. (Mulberry) Fruit to Their Reported Protective Health Benefits: An In Vitro Study.

Morus sp. (mulberry) has a long tradition of use as a medicinal treatment, including for cardiovascular disease and type 2 diabetes, being shown to have antioxidant properties and to promote wound healing. Extracellular vesicles (EVs) are sub-micron, membrane-enclosed particles that were first identified in mammalian bodily fluids. EV-like particles have been described in plants (PDVs) and shown to have similar characteristics to mammalian EVs. We hypothesised that some of the health benefits previously attributed to the fruit of Morus sp. could be due to the release of PDVs. We isolated PDVs from Morus nigra and Morus alba via ultracentrifugation and incubated THP-1 monocytes, differentiated THP-1 macrophages, or HMEC-1 endothelial cells with pro-oxidant compounds DMNQ (THP-1) and glucose oxidase (HMEC-1) or lipopolysaccharide (LPS) in the presence of different fractions of mulberry EVs. Mulberry EVs augmented ROS production with DMNQ in THP-1 and caused the downregulation of ROS in HMEC-1. Mulberry EVs increased LPS-induced IL-1β secretion but reduced CCL2 and TGF-β secretion in THP-1 macrophages. In scratch wound assays, mulberry EVs inhibited HMEC-1 migration but increased proliferation in both low and high serum conditions, suggesting that they have opposing effects in these two important aspects of wound healing. One of the limitations of plant-derived therapeutics has been overcoming the low bioavailability of isolated compounds. We propose that PDVs could provide the link between physiological dose and therapeutic benefit by protecting plant active compounds in the GIT as well as potentially delivering genetic material or proteins that contribute to previously observed health benefits.

The Anti-Inflammatory Effect of SDF-1 Derived Peptide on Porphyromonas gingivalis Infection via Regulation of NLRP3 and AIM2 Inflammasome.

(1) Background: Peptides are appealing as pharmacological materials because they are easily produced, safe, and tolerable. Despite increasing gum-care awareness, periodontitis is still prevalent and is influenced by factors like high sugar consumption, smoking, and aging. Porphyromonas gingivalis is considered a major etiologic agent of periodontitis and activates the NLR family pyrin domain containing 3 (NLRP3) but is absent in melanoma 2 (AIM2) inflammasomes, resulting in pro-inflammatory cytokine release. (2) Methods: We examined the anti-inflammatory effects of 18 peptides derived from human stromal cell-derived factor-1 (SDF-1) on THP-1 macrophages. Inflammation was induced by P. gingivalis, and the anti-inflammatory effects were analyzed using molecular biological techniques. In a mouse periodontitis model, alveolar bone resorption was assessed using micro-CT. (3) Results: Of the 18 SDF-1-derived peptides, S10 notably reduced IL-1β and TNF-α secretion. S10 also diminished the P. gingivalis-induced expression of NLRP3, AIM2, ASC (apoptosis-associated speck-like protein), caspase-1, and IL-1β. Furthermore, S10 attenuated the enhanced TLR (toll-like receptor) signaling pathway and decreased the phosphorylation of nuclear factor-κB (NF-κB) and mitogen-activated protein kinases (MAPKs). In addition, S10 mitigated alveolar bone loss in our P. gingivalis-induced mouse model of periodontitis. (4) Conclusions: S10 suppressed TLR/NF-κB/NLRP3 inflammasome signaling and the AIM2 inflammasome in our P. gingivalis-induced murine periodontitis model, which suggests that it has potential use as a therapeutic treatment for periodontitis.

Resveratrol and piceid enhance efferocytosis by increasing the secretion of MFG-E8 in human THP-1 macrophages.

The process of apoptotic cell clearance by phagocytes, known as efferocytosis, plays an essential role in maintaining homeostasis. Defects in efferocytosis can lead to inflammatory diseases such as atherosclerosis and autoimmune disorders. Therefore, the maintenance and promotion of efferocytosis are considered crucial for preventing these diseases. In this study, we observed that resveratrol, a representative functional food ingredient, and its glycoside, piceid, promoted efferocytosis in both human THP-1 macrophages differentiated with phorbol 12-myristate 13-acetate and peritoneal macrophages from thioglycolate-elicited mice. Resveratrol and piceid significantly increased mRNA expression and protein secretion of MFG-E8 in THP-1 macrophages. Furthermore, the activation of efferocytosis and the increment in MFG-E8 protein secretion caused by resveratrol or piceid treatment were canceled by MFG-E8 knockdown in THP-1 macrophages. In conclusion, we have demonstrated for the first time that resveratrol and piceid promote efferocytosis through the upregulation of MFG-E8 excretion in human THP-1 macrophages.

Type-I interferon shapes peritoneal immunity in cirrhosis and drives caspase-5-mediated progranulin release upon infection

Gut bacterial translocation contributes to immune dysfunction and spontaneous bacterial peritonitis (SBP) in cirrhosis. We hypothesized that exposure of peritoneal macrophages (PMs) to bacterial DNA results in type-I interferon (IFN) production, shaping subsequent immune responses, inflammasome activation, and the release of damage-associated molecular patterns (DAMPs). PMs from patients with cirrhosis were stimulated with E.coli single-stranded DNA (ssDNA), lipopolysaccharide and IFN, or infected with E.coli, S.aureus, and Group B streptococcus invitro. Cytokine release, inflammasome activation, and DAMP release were quantified by quantitative-PCR, ELISA, western blots, and reporter cells employing primary PMs, monocytes, and caspase-deficient THP-1 macrophages. Serum progranulin concentration was correlated with transplant-free survival in 77 patients with SBP. E.coli ssDNA induced strong type-I IFN activity in PMs and monocytes, priming them for enhanced lipopolysaccharide-mediated tumor necrosis factor production without inducing toll-like receptor 4 tolerance. During invitro macrophage bacterial infection, type-I IFN release aligned with upregulated expression of IFN-regulatory factors (IRF)1/2 and guanylate binding proteins (GBP)2/5. PMs upregulated inflammasome-associated proteins and type-I IFN upon E.coli ssDNA exposure and released interleukin-1β upon bacterial infection. Proteomic screening in mouse macrophages revealed progranulin release as being caspase-11-dependent during E.coli infection. PMs and THP-1 macrophages released significant amounts of progranulin when infected with S.aureus or E.coli via gasdermin D in a type-I IFN- and caspase-5-dependent manner. During SBP, PMs upregulated IRF1, GBP2/5 and caspase-5 and higher serum progranulin concentrations were indicative of lower 90-day transplant-free survival after SBP. Type-I IFN shapes peritoneal immune responses and regulates caspase-5-mediated progranulin release during SBP. Patients with cirrhosis exhibit impaired immune responses and increased susceptibility to bacterial infections. This study reveals that type-I interferon responses, triggered by pathogen-associated molecular patterns, are crucial in regulating macrophage activation and priming them for inflammatory responses. Additionally, we elucidate the mechanisms by which type-I interferons promote the release of progranulin from macrophages during spontaneous bacterial peritonitis. Our findings enhance understanding of how bacterial translocation affects immune responses, identify novel biomarkers for inflammasome activation during infections, and point to potential therapeutic targets.

Oxymatrine attenuates sepsis-induced inflammation and organ injury via inhibition of HMGB1/RAGE/NF-κB signaling pathway.

Sepsis is a life-threatening organ dysfunction that endangers patient lives and is caused by an imbalance in the host defense against infection. Sepsis continues to be a significant cause of morbidity and mortality in critically sick patients. Oxymatrine (OMT), a quinolizidine alkaloid derived from the traditional Chinese herb Sophora flavescens Aiton, has been shown to have anti-inflammatory effects on a number of inflammatory illnesses according to research. In this study, we aimed to evaluate the therapeutic effects of OMT on sepsis and explore the underlying mechanisms. We differentiated THP-1 cells into THP-1 macrophages and studied the anti-inflammatory mechanism of OMT in a lipopolysaccharide (LPS)-induced THP-1 macrophage sepsis model. Activation of the receptor for advanced glycation end products (RAGE), as well as NF-κB, was assessed by Western blot analysis and immunofluorescence staining. ELISA was used to measure the levels of inflammatory factors. We found that OMT significantly inhibited HMGB1-mediated RAGE/NF-κB activation and downstream inflammatory cytokine production in response to LPS stimulation. Finally, an in vivo experiment was performed on septic mice to further study the effect of OMT on injured organs. The animal experiments showed that OMT significantly inhibited HMGB1-mediated RAGE/NF-κB activation, protected against the inflammatory response and organ injury induced by CLP, and prolonged the survival rate of septic mice. Herein, we provide evidence that OMT exerts a significant therapeutic effect on sepsis by inhibiting the HMGB1/RAGE/NF-κB signaling pathway.

Bovine DDX3X Restrains Bovine SP110c-Mediated Activation of Inflammasome in Macrophages.

The inflammasome is a vital part of the host's innate immunity activated by cellular infection or stress. Our previous research identified the bovine SP110c isoform (bSP110c) as a novel activator of the inflammasome that promoted the secretion of proinflammatory cytokines IL-1β and IL-18 in macrophages infected with Listeria monocytogenes or stimulated with lipopolysaccharide (LPS). However, the exact molecular mechanism for inhibiting bSP110c-induced inflammasome activation requires further clarification. Here, the researchers identified bovine DDX3X (bDDX3X) as an NLRP3-associated protein and an inhibitor of the bSP110c-induced inflammasome in the human THP1 macrophage cell line. Immunoprecipitation showed that bDDX3X interacted with the bSP110c CARD domain via its helicase domain. The co-expression of bSP110c and bDDX3X in THP1 macrophages significantly prevented the bSP110c-induced activation of inflammasomes. In addition, both bDDX3X and bSP110c interacted with bovine NLRP3 (bNLRP3), and bDDX3X enhanced the interaction between bSP110c and bNLRP3. The expression of bDDX3X in nigericin-stimulated THP1 macrophages significantly suppressed NLRP3 inflammasome activation, ASC speck formation, and pyroptosis. These findings demonstrate that bDDX3X negatively regulates the bSP110c-mediated inflammatory response by restricting the activation of the NLRP3 inflammasome. This discovery unveils a novel regulatory mechanism involving bDDX3X and bSP110c in coordinating inflammasome activation and subsequent cell-fate decisions in LPS-treated macrophages and, in turn, constitutes a step forward toward the implementation of marker-assisted selection in breeding programs aimed at utilizing cattle's immune defenses.

Bile promotes Lactobacillus johnsonii N6.2 extracellular vesicle production with conserved immunomodulatory properties

Recently, Lactobacillus johnsonii N6.2-derived extracellular vesicles (EVs) were shown to reduce apoptosis in human beta cell lines and stimulate insulin secretion in human islets. Our goal was to identify a physiologically relevant environmental condition that induces a hypervesiculation phenotype in L. johnsonii N6.2 and to evaluate if transcriptional changes are involved in this process. Culturing this strain in the presence of 0.2% bovine bile, which mimics a stressor encountered by the bacterium in the small intestine, resulted in approximately a 100-fold increase in EVs relative to cells grown in media without bile. Whole transcriptome analysis of cells grown with bile revealed upregulation of several peptidoglycan hydrolases as well as several genes involved in fatty acid utilization. These results suggest that the hypervesiculation phenotype may be the result of increased cell wall turnover combined with increased accumulation of phospholipids, in agreement with our previous proteomic and lipidomics results. Additionally, EVs isolated from L. johnsonii N6.2 grown in presence of bile maintained their immunomodulatory properties in host-derived βlox5 pancreatic and THP-1 macrophage cell lines. Our findings suggest that in L. johnsonii N6.2 vesiculogenesis is significantly impacted by the expression of cell wall modifying enzymes and proteins utilized for exogenous fatty acid uptake that are regulated at the transcriptional level. Furthermore, this data suggests that vesiculogenesis could be stimulated in vivo using small molecules thereby maximizing the beneficial interactions between bacteria and their hosts.