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Obesity, a widespread global health issue affecting millions, is characterized by excess fat deposition and metabolic dysfunction, significantly elevating the risk of comorbidities like type 2 diabetes, cardiovascular disease, and certain cancers, all of which contribute to rising rates of preventable morbidity and mortality. Current approaches to obesity, including lifestyle modifications, and pharmacotherapy, often face limitations such as poor long-term adherence, side effects, and insufficient targeting of the complex, multifactorial pathways underlying the disease. Herein we report a dual, RNA-mediated combinatorial approach using targeting lipid nanoparticles (LNP) for the treatment of obesity. LNPs were co-encapsulated with mRNA encoding Interleukin-27 (mIL-27) to coactivate PGC-1α, PPARα, and UCP-1, thereby promoting adipocyte differentiation and enhancing adaptive thermogenesis within adipocytes, and siRNA targeting Dipeptidyl peptidase-4 (siDPP-4) to silence the primary inhibitory enzyme of GLP-1, and GIP within the incretin system, effectively restoring glucose homeostasis. Following post translational silencing of DPP-4 and upregulation of IL-27 in a diet-induced obesity (DIO) mice model, increased expression of thermogenic biomarkers PGC-1α, PPARα, and UCP-1 was observed at the molecular, protein, and tissue level, and insulin sensitivity was restored. Importantly, this gene modulation led to a 21.1 % reduction of bodyweight after treatment in the DIO model. These findings demonstrate for the first time a dual RNA-mediated combinatorial approach, leveraging liver targeting LNP delivery with synergistic effects from incretin system regulation and induction of adipocyte differentiation and thermogenesis after codelivery of siDPP-4 and mIL-27. This innovative strategy provides a promising alternate framework for addressing obesity and its associated metabolic dysfunction. Show less

The newly generated CD4 single-positive (SP) T lymphocytes are featured by enhanced IL-4 but repressed IFN-γ production. The mechanisms underlying this functional bias remain elusive. Previous studies have reported that CD4 Show less

GATA2 establishes transcriptomes governing hematopoietic stem/progenitor cell development. In progenitors, GATA2 represses inflammatory genes (Il6st and Il6ra) encoding IL6ST/GP130 and IL6RA receptor subunits mediating IL-6 signaling. While IL6ST heterodimerizes with IL6RA, IL-11, IL-27, oncostatin M, and leukemia inhibitory factor receptors, IL6RA heterodimerizes exclusively with IL6ST to confer IL-6 signaling. As GATA2-dependent repression is not well understood, we devised a multi-omics strategy to elucidate mechanisms underlying repression and applied the approach to the cytokine/chemokine receptor gene family. Identifying accessible distal and intronic chromatin sites in GATA2-deficient (GATA2 Show less

Hepatitis C virus (HCV) infection is a significant global health concern, as both acute and chronic hepatitis caused by HCV can lead to liver cancer and long-term liver damage. Thymoquinone (TQ), the active compound found in the remarkable herb Nigella sativa, has various anti-inflammatory and antiproliferative effects. In this study, we investigated the effect of TQ on the interferon-alpha (IFN-α) pathway and its ability to prevent HCV replication in the HepG2 cell line. Our findings showed no significant alterations in cell viability or lactate dehydrogenase (LDH) production in TQ-treated cells, while significant alteration in both factors was detected in cells treated with Sovaldi, the most commonly used drug for treatment of HCV infection. Interestingly, the level of the HCV NS5A protein was significantly reduced in infected cells treated with either TQ or Sovaldi in a dose-dependent manner. The expression of phosphorylated Raf-1 (phospho-Raf-1) and phospho-Mek-1 in infected cells was inhibited by TQ treatment, and the potential interaction between TQ and Ref-1 was confirmed by a molecular docking simulation. Unlike autophagy-related 12 (Atg12), the expression of LC3B in infected cells was also inhibited in a dose-dependent manner by TQ treatment. Conversely, the levels of interleukin-27 (IL-27) and interferon-alpha (IFN-α) in infected cells were significantly increased in a time- and dose-dependent manner by TQ treatment. These data suggest that TQ exerts antiviral effects in HepG2 cells by disrupting HCV replication through targeting of the Raf-1 signaling pathway and promoting the overproduction of IL-27 and IFN-α in infected cells. Show less

Neuroinflammation is an inflammatory response occurring within the central nervous system (CNS). The process is marked by the production of pro-inflammatory cytokines, chemokines, small-molecule messengers, and reactive oxygen species. Microglia and astrocytes are primarily involved in this process, while endothelial cells and infiltrating blood cells contribute to neuroinflammation when the blood-brain barrier (BBB) is damaged. Neuroinflammation is increasingly recognized as a pathological hallmark of several neurological diseases, including amyotrophic lateral sclerosis (ALS), and is closely linked to neurodegeneration, another key feature of ALS. In fact, neurodegeneration is a pathological trigger for inflammation, and neuroinflammation, in turn, contributes to motor neuron (MN) degeneration through the induction of synaptic dysfunction, neuronal death, and inhibition of neurogenesis. Importantly, resolution of acute inflammation is crucial for avoiding chronic inflammation and tissue destruction. Inflammatory processes are mediated by soluble factors known as cytokines, which are involved in both promoting and inhibiting inflammation. Cytokines with anti-inflammatory properties may exert protective roles in neuroinflammatory diseases, including ALS. In particular, interleukin (IL)-10, transforming growth factor (TGF)-β, IL-4, IL-13, and IL-9 have been shown to exert an anti-inflammatory role in the CNS. Other recently emerging immune regulatory cytokines in the CNS include IL-35, IL-25, IL-37, and IL-27. This review describes the current understanding of neuroinflammation in ALS and highlights recent advances in the role of anti-inflammatory cytokines within CNS with a particular focus on their potential therapeutic applications in ALS. Furthermore, we discuss current therapeutic strategies aimed at enhancing the anti-inflammatory response to modulate neuroinflammation in this disease. Show less

Acute respiratory distress syndrome (ARDS) is a lung inflammatory condition associated with the accumulation of fluid edema and cell infiltrates into the alveolar space along with dysregulation of the immune response. Current therapeutics are limited to palliative care, i.e., mechanical ventilators, thus highlighting the need to develop targeted therapeutic for ARDS. Interleukin-27 (IL-27) is a multifunctional cytokine with the capability for immune modulation. Our interest lies in exploring the properties of IL-27, particularly as an anti-inflammatory cytokine that functions as an antagonist of IL-6 signaling, as an inducer of anti-viral genes, as a promoter of tissue repair, and as a regulator of both the innate and adaptive immune responses, possessing promising potential as a therapeutic for ARDS. To overcome the challenge of repeated administration due to the short half-life of cytokines, we utilized a cell-based gene therapy approach. An IL-27-expressing plasmid was transfected into adipose mesenchymal stromal cells (ASC) that serve as the gene therapy carriers. For in vitro studies, we treated mono- and co-culture lung lipopolysaccharide (LPS)-induced lung epithelial and monocytes/macrophages cell line with IL-27-expressing ASC (IL-27 ASC) conditioned media (CM) to determine the effects on pro-inflammatory gene expression. For in vivo studies, male C57BL/6 mice were intratracheally injected with LPS (5 mg/kg) and treated either PBS, ASC, or IL-27 ASC (5 × 10 IL-27 ASC CM reduced pro-inflammatory gene expression of lung epithelial and macrophages cultured in both mono- and co-culture systems. Additionally, IL-27 ASC were able to reduce pro-inflammatory markers, decrease cell infiltration into the lungs, promote genes and immune cells involved in tissue repair, and rebalance innate and adaptive immunity in an LPS-induced in vivo model. Collectively, our in vitro and in vivo results show promising potential for IL-27 cell-based gene therapy as a treatment for ARDS. Show less

Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) are life-threatening conditions marked by extensive epidermal necrolysis and skin sloughing. The study aimed to assess the serum levels of specific proinflammatory interleukins (IL-18, -21, 22, -23, -27, and -31) and their relationship with the severity of SJS/TEN within the Vietnamese population. This descriptive cross-sectional study was conducted from 2018 to 2020. Serum levels of IL-18, -21, -22, -23, -27, and -31 were measured using the fluorescence covalent microbead immunosorbent assay. The study included 61 patients (29 males and 32 females; 21 with SJS and 40 with TEN), with a median age of 51 years (interquartile range: 37-58), and 20 healthy controls. The median lesional area covered 45% of the body surface area (interquartile range: 8-70%). The most frequently identified medications were traditional medicine (19 patients; 31.15%), allopurinol (9 patients; 14.75%), and carbamazepine (8 patients; 13.11%). In the TEN group, the serum level of IL-18 was significantly elevated compared to the healthy control group. A correlation was found between serum levels of IL-18 and IL-27 and the lesional area in SJS/TEN patients, as well as between serum levels of IL-18 and IL-31 and the lesional area in TEN patients. Serum levels of IL-18 were increased in TEN group. Additionally, serum concentrations of IL-18, IL-27, and IL-31 were associated with disease severity as indicated by the lesional area. These interleukins may play an important role in the pathogenesis of SJS/TEN. Show less

Biomarker profiling from biofluids such as blood are widely measured in clinical research, using for example Olink proteomics panels. One such research focus area is cardiovascular disease (CVD), for which chronic sleep restriction (SR) is a risk factor. However, it remains unclear whether blood levels of commonly measured CVD biomarkers are sensitive to acute dynamic factors such as SR, physical exercise (PEx), and time of day. In this crossover design, 16 normal-weight, healthy men underwent three highly standardized in-lab nights of SR (4.25 h/night) and normal sleep (NS, 8.5 h/night) in randomized order, with 88 CVD blood protein biomarkers quantified using the Olink technology (and selected validation using ELISA) in the morning, evening, and immediately before and repeatedly after 30 min of high-intensity exercise. We found significant time-of-day-dependent changes in several CVD biomarkers. Whereas several proteins were exercise-induced across sleep conditions (such as the canonical exerkines IL- 6 and BDNF), exercise-induced proteomic dynamics differed in response to recurrent SR, compared with following NS. Moreover, SR compared with NS resulted in a biomarker profile previously associated with increased prospective risk of several CVDs across large-scale cohorts (such as higher circulating levels of IL-27 and LGALS9). Our findings highlight how dynamic physiology can modulate CVD biomarker levels. These results also underscore the need to consider sleep duration as a key determinant of cardiovascular health-an emphasis reflected in recent American Heart Association guidelines. Further studies in women, older individuals, and patients with prior CVD, and across different chronotypes and dietary schedules are warranted. Show less

Intrahepatic immune responses are often insufficient to control hepatitis virus infections. A recent study by Venzin and colleagues demonstrates a detailed mechanism by which an intrahepatic tricellular network and the cytokine IL-27 can augment virus-specific immunity. Show less

Strong T cell receptor (TCR) and interleukin (IL)-27 signaling influence type 1 regulatory (Tr1) T cell development, but whether other signals determine their differentiation is unclear. Utilizing Tg4 TCR transgenic mice, we established a model for rapid Tr1 cell induction. A single high dose of [4Y]-MBP peptide drove the differentiation of Show less

Obesity is turning into a more critical problem for public health. Vitamin D The study aims to examine the influence of VD Firstly, a small sample population study was conducted to compare the disparities in serum 25(OH)D A correlation was identified between serum 25(OH)D The study shows that VD Show less

Mayaro virus (MAYV) belongs to the Togaviridae family and is the etiologic agent of Mayaro fever, a disease in which inflammatory responses play a critical role in viral pathogenesis. Macrophages are targets of viral infection and key components of innate immunity and antiviral response. This study analyzed an RNA-sequencing (RNA-seq) dataset to gain insights into inflammatory and antiviral responses in monocyte-derived macrophages (MDMs) infected with the MAYV strain (Venezuelan 2010) at a multiplicity of infection (MOI) of 10. The RNA-seq results were validated by real-time quantitative polymerase chain reaction in MDMs infected with the MAYV strain from Brazil (MOI of 2). In addition, the replication capacity of MAYV and the resulting cell death in infected MDMs were assessed using plaque assays and flow cytometry. At 72 hours post-infection, transcriptomic analysis revealed that MAYV promotes a robust proinflammatory response by upregulating the expression of Toll-like receptors, RIG-I-like receptors, and the nuclear factor-κB complex. This strong inflammatory response was accompanied by a robust antiviral response dependent on type I/III interferon and interleukin-27. Both antiviral responses are mediated through the Janus kinase/signal transducer and activator of transcription signaling pathway, leading to the expression of interferon-stimulated genes. Moreover, MAYV-infected MDMs expressed markers of programmed cell death. These findings highlight the inflammatory response and antiviral activity of MDMs at a late stage in MAYV infection, suggesting a critical role of macrophages in MAYV pathogenesis. Show less

Hyperactivation of cutaneous macrophages promotes the development of chronic pain. Stimulation of nociceptive regions promotes neuroplasticity, which affects pain perception and related physiological responses. However, the specific mechanisms by which cutaneous macrophages sense and elicit nociceptive responses are unknown. Here, we exacerbated the reduction of systemic pain threshold after chronic heart failure (CHF) by silencing follistatin-like 1 (FSTL1), especially the abnormal cutaneous nociceptive sensation at PC6 acupoint, the site associated with cardiac involvement pain. The upregulation of P2Y6 and interleukin-27 expression is intimately linked to the activation of skin macrophages. Hyperactivation of P2Y6 receptor (P2Y6R) may be associated with MHC II M1 Show less

Age-related macular degeneration (AMD) is a leading cause of visual impairment and blindness in the elderly. Aging is the most important risk factor for AMD, and the aging immune system seems to be involved in pathogenesis. This study investigates the systemic aging immune profile in relation to AMD stage and treatment response. Treatment-naïve patients with neovascular AMD (nAMD), intermediate AMD and healthy controls were included in this prospective study. Participants were examined for systemic aging immune profiles and compared to AMD stage, as well as initial and one-year treatment response in nAMD patients. Flowcytometry was performed to determine T cell differentiation (naïve, central memory and effector memory) and expression of costimulatory markers (CD27, CD28, CD56). Cytokine assays were performed to measure the concentrations of plasma cytokines IFN-γ, IL-1β, IL-2, IL-6, IL-10, IL-12, IL-17, IL-22, IL-27, TNF-α. Polymorphisms of CFH and ARMS2 genes were compared in nAMD patients. Patients with nAMD had significantly higher proportions of central and effector memory CD8+ T cells compared to controls (both P < 0.036). nAMD patients had significantly elevated concentrations of IFN-γ, IL-1β, IL-2, and IL-10 (all P < 0.05). nAMD patients with poor initial treatment response had a significantly higher concentration of plasma IFN-γ compared to good responders (P =0.026). Patients with nAMD had a more advanced systemic aging immune profile with higher levels of T cell differentiation and plasma cytokines compared to controls. Poor initial response had elevated levels of plasma IFN-γ compared to good responders in nAMD. Show less

The refrigeration (cold storage) of platelet components provides several benefits over room-temperature (RT) storage, extending the shelf-life up to 21 days. However, the effect of storage conditions on platelet activation in response to stimulation remains unclear. A paired study was conducted where buffy-coat platelet concentrates were pooled, split, and allocated to RT or cold storage (n = 6 in each group). Platelet samples were taken on days 1, 7, 14, and 21, which were tested without stimulation or following activation with TRAP-6, A23187, lipopolysaccharides, or Histone-H4. Imaging flow cytometry was used to assess the surface characteristics of platelets and extracellular vesicles (EVs). The supernatant concentration of EGF, RANTES, PF4, CD62P, IL-27, CD40L, TNF-α, and OX40L was examined using ELISA. Cold-stored platelets generated a greater proportion of procoagulant platelets and EVs than RT-stored platelets in response to stimulation. The supernatant of cold-stored components contained lower concentrations of soluble factors under basal conditions, suggesting that platelet granules were better retained. Cold-stored platelets released higher concentrations of soluble factors following stimulation with TRAP-6, A23187, or Histone-H4. Only cold-stored platelets responded to lipopolysaccharides. These data demonstrate that cold-stored platelets retain the capacity to respond to stimuli after 21 days of storage, which may facilitate improved functional post-transfusion. Show less

Cytokines influence cell-fate decisions of naïve lymphocytes and determine outcome of immune responses by transducing signals that regulate the initiation, intensity and duration of immune responses. However, aberrant regulation of physiological levels of cytokines contribute to the development of autoimmune and other inflammatory diseases. The Interleukin 6 (IL-6)/IL-12 superfamily of cytokines have a profound influence on all aspects of host immunity and our focus in this review is on the signaling pathways that mediate their functions, with emphasis on how this enigmatic family of cytokines promote or suppress inflammation depending on the physiological context. We also describe regulatory lymphocyte populations that suppress neuroinflammatory diseases by producing cytokines, such as IL-27 (i27-Breg) or IL-35 (i35-Breg and iT Show less

Diabetic foot ulcer (DFU), a serious complication of diabetes, is a life-threatening disease that often leads to lower limb amputation and a shortened lifespan. Interleukin-27 (IL-27) is a member of the IL-12 family and has the potential to exert dual effects on the immune response. The role of IL-27 in diabetic skin wound healing is unknown. The aim of this study was to investigate whether there is abnormal expression of IL-27 in diabetic skin and whether supplementation with IL-27 can promote diabetic wound healing by modulating macrophage polarization. We established a streptozotocin (STZ)-induced diabetic mouse model and constructed diabetic wounds. We assessed protein expression by western blotting (WB) and immunohistochemical (IHC) staining. We also performed hematoxylin-eosin (H&E) staining and Masson's trichrome staining. In the presence of lipopolysaccharide (LPS) and high glucose (HG), we treated the mononuclear macrophage line RAW264.7 and bone marrow-derived macrophages (BMDMs) with IL-27. To assess macrophage polarization, we examined the expression of inducible nitric oxide synthase (iNOS), IL-1β and arginase-1 (Arg-1). To understand the underlying mechanisms, we used macrophage IL-27ra knockout mice to knockout macrophage IL-27 receptors. Our in vivo experiments revealed that the expression of IL-27 in the skin of diabetic mice was significantly decreased and that supplementation with IL-27 promoted diabetic wound healing. In vitro, compared with the LPS group, supplementation with IL-27 alleviated the suppression of multiple cellular functions, such as iNOS and IL-1β expression, cell migration, and phagocytosis, in macrophages after HG exposure. Mechanistically, we found that IL-27 expression was decreased and that the activation of signal transducer and activator of transcription 3 (STAT3) by phosphorylation was inhibited in diabetic skin, leading to an inability of wound macrophages to polarize to an M1 phenotype effectively, which in turn blocked M1-to-M2 polarization of wound macrophages and ultimately delayed wound healing. The present study revealed that supplementation with IL-27 promoted M1-to-M2 polarization of wound macrophages and diabetic wound healing through the IL-27-IL-27Rα-p-STAT3 axis. These findings suggest that IL-27 may be a potential therapeutic target for DFU. Show less

Mutations in the Leucine-rich repeat kinase 2 ( We investigated the levels of soluble immune regulators in the serum (n=651) and cerebrospinal fluid (CSF, n=129) of In this extensive discovery cohort, we identified several elevated serum immune regulatory factors associated with This study highlights distinct immune profiles associated with LRRK2 mutations and PD in the periphery and CNS. Serum levels of SDF-1alpha and TNF-RII were elevated in LRRK2 mutation carriers, while CSF immune markers were reduced. In PD, irrespective of LRRK2 status, reduced CSF inflammatory analytes and weak serum signals were observed. These results provide insight into immune dysregulation linked to LRRK2 mutations. If replicable in independent datasets, they offer potential avenues for biomarker and therapeutic exploration. Show less

Hundreds of genetic associations for asthma have been identified, yet translating these findings into mechanistic insights remains challenging. We leveraged plasma proteomics from the UK Biobank Pharma Proteomics Project (UKB-PPP) to identify biomarkers and effectors of asthma risk or heterogeneity using genetic causal inference approaches. We identified 609 proteins associated with asthma status (269 proteins after controlling for body mass index [BMI] and smoking). Analysis of genetically predicted protein levels identified 70 proteins with putative causal roles in asthma risk, including known drug targets and proteins without prior genetic evidence in asthma (e.g., GCHFR, TDRKH, and CLEC7A). The genetic architecture of causally associated proteins provided evidence for a Toll-like receptor (TLR)1-interleukin (IL)-27 asthma axis. Lastly, we identified evidence of causal relationships between proteins and heterogeneous aspects of asthma biology, including between TSPAN8 and neutrophil counts. These findings illustrate that integrating biobank-scale genetics and plasma proteomics can provide a framework to identify therapeutic targets and mechanisms underlying disease risk and heterogeneity. Show less

Cytokines are essential for regulating immune cell activity during pregnancy. Research shows that CD4+ T-cells exhibit specific cytokine secretion patterns, resulting in polarized immune responses. This study aims to compare the gene expression levels of Th1, Th2, and Th17 cytokines in women with normal pregnancies versus those with a history of recurrent spontaneous abortion (RSA). In this case-control study, 20 patients with RSA within 24 h of their last abortion were compared to 20 pregnant women with no history of abortion (Control Group). Cytokine levels of IL-2, IL-17, and IL-27 were quantified using real-time polymerase chain reaction (RT-PCR). Overall cytokine levels were similar between the groups, but the cytokine levels in both groups were generally similar. However, higher IL-17 and IL-2 levels were observed in the healthy pregnancy group ( Elevated IL-2 levels may be a risk factor for RSA. Consistent with recent studies, our findings emphasize the role of IL-17 and IL-27 as crucial regulatory cytokines for maintaining a successful pregnancy. Show less

Sepsis is associated with high morbidity and high mortality and has strongly motivated intense studies into its mechanisms. Antibiotics, aimed to eradicate bacteria, have some impact on the immune system due to anti-inflammatory properties. Tigecycline, an antibiotic of the glycylcycline class, is commonly used for severe infections. This study aimed to investigate tigecycline's mechanism on the inflammatory response of sepsis to find new targets for sepsis treatment. The objective included (i) to observe the changes in inflammatory factors in LPS (lipopolysaccharide) induced septic mice after tigecycline administration, (ii) to detect the effect of tigecycline on macrophages NF-κB (nuclear factor kappa B) signalling. For LPS-induced sepsis in mice and intervention with tigecycline, mice were first injected with tigecycline (6.5 mg/kg) via tail vein followed by LPS (15 mg/kg). Luminex analysis was performed on 16 mediators. NF-κB signalling pathway antibody chip detected the expression of target sites in macrophages of the LPS group and tigecycline + LPS group. Tigecycline has inhibitory effects on LPS-induced inflammatory response in septic mice, decreasing the concentrations of IL (interleukin)-6, IL-27, TNF-α (tumour necrosis factor-α), TNF RII, IFN-γ (interferon-gamma), CCL5/RANTES (CC Motif Chemokine Ligand) while increasing IL-6Rα, IL-10, and TWEAK (TNF-related weak inducer of apoptosis). Tigecycline downregulated phosphorylation levels of key sites JNK (c-Jun N-terminal kinase)1/2/3, p-p65 (s468) and p-p105/p50 (s907) in NF-κB signalling. Tigecycline may inhibit the excessive immune response induced by LPS in sepsis, which may cause a potential protective effect on the host through immune regulation. Show less

Interleukin (IL)-27 is a heterodimeric immunoregulatory cytokine expressed at elevated levels early in life that compromises bacterial clearance and promotes severe outcomes during neonatal sepsis. In turn, IL-27Rα-deficient neonatal mice exhibit better control of bacteria, reduced systemic inflammation, and improved outcomes. IL-27 primarily activates and signals through either Signal transducer and activator of transcription (STAT)-1 or STAT-3 in macrophages. Targeted deletion of STAT-3 in macrophages has been reported to improve responsiveness to Lipopolysaccharide (LPS) and promote Th1 activity. As such, in the present study, we investigated the role of STAT-3 signaling in IL-27-mediated suppression of bacterial clearance and lysosomal activity in neonatal macrophages during The neonatal period is a time in which newborns have increased vulnerability and the highest risk of death from infection. This includes sepsis for which there is a considerable global burden of disease. We have determined that the cytokine interleukin (IL)-27 is expressed at elevated levels in the first days of life and continues to rise during experimental bacterial neonatal sepsis. Neonatal mice that cannot respond to IL-27 exhibit improved outcomes. In this work, we have investigated the influence of STAT-3 on control of bacteria and inflammation during IL-27 signaling in neonates. It is critical that we understand mechanisms that underlie neonatal susceptibility to infection so that we can identify new targets for therapeutic intervention. Here, we define the value of STAT-3 in approaches to targeted therapies for bacterial neonatal sepsis. Show less

Interleukin (IL)-27 is an anti-viral cytokine. IL-27-treated monocyte-derived macrophages (27-Mac) suppressed HIV replication. Macrophages are generally divided into two subtypes, M1 and M2 macrophages. M2 macrophages can be polarized into M2a, M2b, M2c, and M2d by various stimuli. IL-6 and adenosine induce M2d macrophages. Since IL-27 is a member of the IL-6 family of cytokines, 27-Mac was considered M2d macrophages. In the current study, we compared biological function and gene expression profiles between 27-Mac and M2d subtypes. Monocytes derived from health donors were differentiated to M2 using macrophage colony-stimulating factor. Then, the resulting M2 was polarized into different subtypes using IL-27, IL-6, or BAY60-658 (an adenosine analog). HIV replication was monitored using a p24 antigen capture assay, and the production of reactive oxygen species (ROS) was determined using a Hydrogen Peroxide Assay. Phagocytosis assay was run using GFP-labeled opsonized E. coli. Cytokine production was detected by the IsoPlexis system, and the gene expression profiles were analyzed using single-cell RNA sequencing (scRNA-seq). 27-Mac and BAY60-658-polarized M2d (BAY-M2d) resisted HIV infection, but IL-6-polarized M2d (6-M2d) lacked the anti-viral effect. Although phagocytosis activity was comparable among the three macrophages, only 27-Mac, but neither 6-M2d nor BAY-M2d, enhanced the generation of ROS. The cytokine-producing profile of 27-Mac did not resemble that of the two subtypes. The scRNA-seq revealed that 27-Mac exhibited a different clustering pattern compared to other M2ds, and each 27-Mac expressed a distinct combination of anti-viral genes. Furthermore, 27-Mac did not express the biomarkers of M2a, M2b, and M2c. However, it significantly expressed CD38 (p<0.01) and secreted CXCL9 (p<0.001), which are biomarkers of M1. These data suggest that 27-Mac may be classified as either an M1-like subtype or a novel subset of M2, which resists HIV infection mediated by a different mechanism in individual cells using different anti-viral gene products. Our results provide a new insight into the function of IL-27 and macrophages. Show less

Acute pulmonary inflammation is a severe lower respiratory tract infection. Sinensetin (SIN), a polymethoxyflavone with strong anti-inflammatory properties, is known to ameliorate LPS-induced acute inflammatory lung injury, but its molecular mechanisms are not fully understood. This study aimed to provide insight into the pharmacological mechanisms of SIN in attenuating acute pulmonary inflammation. In LPS-induced inflammation assays in vivo and in vitro, SIN significantly reduced the mRNA levels of inflammatory genes including MCP-1, ICAM1, Ccl3, Ccl4, Ccl5, Ccl7, Cxcl9, Cxcl10, IL1α, IL1β, IL6, IL11, IL18, IL27, TNF-α, IFN-γ, TLR4, MyD88, F4/80, COX2, iNOS, NLRP3, ASC, JAK2, STAT3, STAT4, and Bcl2l1, as well as increased the mRNA levels of anti-inflammatory genes such as IL4, IL10, and IL12α. Besides, SIN markedly decreased the expression of CD68, TLR4, MyD88, phospho-IκBα (S32/S36), phospho-NF-κB p65 (S536), MCP-1, ICAM1, phospho-JAK2 (Tyr1008), phospho-STAT1 (S727), phospho-STAT3 (Y705), and phospho-STAT4 (Y693), inhibited NF-κB p65 translocation into the nucleus, thereby blocking in combination with STAT transcription factors to induce target gene expression. Further GC-MS/MS and LC-MS/MS metabolomic analysis revealed that SIN significantly increased the abundance of anti-inflammatory metabolites, such as L-alanine, L-carnitine, L-glutamic acid, Glycine, and L-cysteine. In conclusion, the results indicated that SIN attenuated LPS-induced acute pulmonary inflammation by modulating NF-κB p65-mediated immune resistance and STAT3-mediated tissue resilience. All these favorable findings presented critical insights into the remarkable abilities and health benefits of SIN in ameliorating inflammatory lung disease. Show less