Differences In Inflammatory Response Research Articles

Differential inflammatory responses of the native left and right ventricle associated with donor heart preservation.

BackgroundDysfunction and inflammation of hearts subjected to cold ischemic preservation may differ between left and right ventricles, suggesting distinct strategies for amelioration.Methods and ResultsExplanted murine hearts subjected to cold ischemia for 0, 4, or 8 h in preservation solution were assessed for function during 60 min of warm perfusion and then analyzed for cell death and inflammation by immunohistochemistry and western blotting and total RNA sequencing. Increased cold ischemic times led to greater left ventricle (LV) dysfunction compared to right ventricle (RV). The LV experienced greater cell death assessed by TUNEL+ cells and cleaved caspase‐3 expression (n = 4). While IL‐6 protein levels were upregulated in both LV and RV, IL‐1β, TNFα, IL‐10, and MyD88 were disproportionately increased in the LV. Inflammasome components (NOD‐, LRR‐, and pyrin domain‐containing protein 3 (NLRP3), adaptor molecule apoptosis‐associated speck‐like protein containing a CARD (ASC), cleaved caspase‐1) and products (cleaved IL‐1β and gasdermin D) were also more upregulated in the LV. Pathway analysis of RNA sequencing showed increased signaling related to tumor necrosis factor, interferon, and innate immunity with ex‐vivo ischemia, but no significant differences were found between the LV and RV. Human donor hearts showed comparable inflammatory responses to cold ischemia with greater LV increases of TNFα, IL‐10, and inflammasomes (n = 3).ConclusionsMouse hearts subjected to cold ischemia showed time‐dependent contractile dysfunction and increased cell death, inflammatory cytokine expression and inflammasome expression that are greater in the LV than RV. However, IL‐6 protein elevations and altered transcriptional profiles were similar in both ventricles. Similar changes are observed in human hearts.

Nanocellulose Length Determines the Differential Cytotoxic Effects and Inflammatory Responses in Macrophages and Hepatocytes.

Nanocellulose including cellulose nanocrystal (CNC) and cellulose nanofiber (CNF) has attracted much attention due to its exceptional mechanical, chemical, and rheological properties. Although considered biocompatible, recent reports have demonstrated nanocellulose can be hazardous, including serving as drug carriers that accumulate in the liver. However, the nanocellulose effects on liver cells, including Kupffer cells (KCs) and hepatocytes are unclear. Here, the toxicity of nanocellulose with different lengths is compared, including the shorter CNCs (CNC-1, CNC-2, and CNC-3) and longer CNF (CNF-1 and CNF-2), to liver cells. While all CNCs triggered significant cytotoxicity in KCs and only CNC-2 induced toxicity to hepatocytes, CNFs failed to induce significant cytotoxicity due to their minimal cellular uptake. The phagocytosis of CNCs by KCs induced mitochondria ROS generation, caspase-3/7 activation, and apoptotic cell death as well as lysosomal damage, cathepsin B release, NLRP3 inflammasome and caspase-1 activation, and IL-1β production. The cellular uptake of CNC-2 by hepatocytes is through clathrin-mediated endocytosis, and it induced the caspase-3/7-mediated apoptosis. CNC-2 shows the highest levels of uptake and cytotoxicity among CNCs. These results demonstrate the length-dependent mechanisms of toxicity on liver cells in a cell type-dependent fashion, providing information to safely use nanocellulose for biomedical applications.

Neutrophil to lymphocyte ratio and adverse clinical outcomes in patients with ischemic stroke.

BackgroundNeutrophils and lymphocytes mediate differential inflammatory responses after ischemic stroke and have different effects on patients’ clinical outcomes. Several studies have used the neutrophil to lymphocyte ratio (NLR) as a prognostic indicator for ischemic stroke; however, some limitations remain.MethodsWe derived data from the Third China National Stroke Registry. The NLR is defined as neutrophil count/lymphocyte count. Patients included in the study were classified into four groups by NLR quartiles. Odds ratios (ORs) of adverse clinical outcomes were calculated with the lowest quartile group as the reference category. We plotted receiver operating characteristic (ROC) curves of NLR for adverse clinical outcomes and calculated area under the curve (AUC) values and cutoff values. Under different TOAST classifications, medians of NLR and ORs of adverse clinical outcomes were also calculated. Furthermore, interaction tests between NLR and etiology were performed.ResultsA total of 13,018 patients were enrolled. At both 3- and 12-month follow-ups, higher quartile groups were associated with increased risks of death and poor functional outcomes, even after adjustments. For death, the cutoff values of NLR were 3.872 at 3-month follow-up and 3.180 at 12-month follow-up. For poor functional outcomes, the cutoff value of NLR was 2.846 at both 3- and 12-month follow-ups. The association between NLR and stroke recurrence was significant only at 3-month follow-up before adjustments. There was no correlation between NLR and hemorrhagic transformation during hospitalization. Under different TOAST classifications, the medians of NLR were different; conversely, the correlations of NLR with adverse clinical outcomes had no differences.ConclusionsHigh level of NLR within the first 24 h after admission was associated with increased risks of both short- and long-term adverse clinical outcomes in patients with ischemic stroke, regardless of etiology.

Differential Inflammatory Responses in the Healing of Oral Mucosa and Skin Wounds

Differential Inflammatory Responses in the Healing of Oral Mucosa and Skin Wounds

Differential inflammatory responses associated with Leishmania major and Ltropica in culture.

Anthroponotic cutaneous leishmaniasis (ACL) due to Leishmania tropica and zoonotic CL (ZCL) due to Lmajor have different clinical and epidemiological features. To determine whether pro-inflammatory cytokines are involved in diverse pathogenicity of Leishmania species causing CL. The capacity of Lmajor/Ltropica to modulate expression of IL-1β, IL-8 (CXCL8), IFN-γ, TNF-α and MCP-1 (CCL2) in peripheral blood mononuclear cells (PBMCs) and monocyte-derived macrophages (MDMs) was evaluated by real-time RT-PCR technique. PBMCs from both ZCL and ACL cases expressed significantly higher IFN-γ (P<.001) and TNF-α (P<.05) compared with healthy controls (HC). PBMCs from ACL patients expressed significantly higher IL-1β and IL-8 compared with ZCL patients and HC when stimulated with live Lmajor or Ltropica promastigotes (P<.001). After 4 and 10hours, Lmajor-infected MDMs expressed significantly higher IFN-γ (P<.05), and after 10hours, Ltropica-infected MDMs expressed significantly higher IL-1β, IFN-γ and IL-8 compared with noninfected cells (P<.05). This study shows differential parasite-mediated stimulations of the inflammatory response with Lmajor vs Ltropica ex vivo. Pro-inflammatory cytokines particularly IL-8 (CXCL8) and IL-1β might contribute in diverse clinical features of CL such as longer duration of lesion persistence in ACL patients.

No Sex Differences in Inflammatory Response and Vascular Function During Low Estrogen Phase

Premenopausal women experience a lower incidence of cardiovascular disease as compared to age-matched men, partially due to the cardioprotective effects of estrogen. Acute inflammation increases pro-inflammatory cytokine production and reduces nitric oxide bioavailability, resulting in impaired vascular function. Estrogen has both anti-inflammatory and pro-vasodilatory properties. During the high estrogen mid-luteal phase of the menstrual cycle, sex differences in vascular function and inflammatory responses to induced inflammation have been observed in premenopausal women as compared with age-matched men. However, limited knowledge exists regarding sex differences in estrogen independent responses to acute inflammation. Thus, our objective was to evaluate sex differences in vascular responses to induced inflammation- with a reduced influence of estrogen concentration. Due to the dampening of estrogen-related anti-inflammatory and pro-vasodilatory effects when estrogen concentrations are low, we hypothesized that women would experience a greater inflammatory response as compared with men and that women and men would exhibit similar vascular function. In a double-blind crossover sham-controlled study of 15 women (21 ± 3y) and 15 men (21 ± 2y), we assessed pro-inflammatory interleukin-6 concentration ([IL-6]) and vascular function via brachial artery flow-mediated dilation (FMD) at baseline (BL), 24 hours (24H), and 48 hours (48H) after influenza vaccine administration. Women were studied when circulating estrogen levels are typically low (early follicular phase or placebo pill phase of oral contraceptive pills) and similar between sexes. Following induced inflammation, both sexes exhibited an increase in [IL-6] at 24H (BL v 24H: women p=0.0049; men p=0.0015) that returned to near baseline levels by 48H (BL v 48H: women p=0.43; men p=0.29; [IL-6] (pg/mL): women BL: 0.56 ± 0.5, 24H: 1.14 ± 0.7, 48H: 0.59 ± 0.4; men BL: 0.38 ± 0.2, 24H: 1.05 ± 0.7, 48H: 0.56 ± 0.2). There were no sex differences in FMD or [IL-6] at any time point (FMD: sex p=0.062, time p=0.30, interaction p=0.25; FMD (%): women BL: 8.33 ± 4.8, 24H: 6.40 ± 4.4, 48H: 9.05 ± 5.5; men: BL: 5.25 ± 2.9, 24H: 6.14 ± 2.1, 48H: 6.71 ± 1.1; [IL-6]: BL: p=0.38, 24H: p=0.57, 48H: p=0.99). Notably, women exhibited significantly lower resting brachial artery diameter (BL D; p<0.0001 all time points) than men (BL D (mm): women BL: 3.22 ± 0.4, 24H: 3.18 ± 0.4, 48H: 3.19 ± 0.4; men BL: 4.07 ± 0.6, 24H: 4.24 ± 0.4, 48H, 4.22 ± 0.4). Due to sex difference in baseline diameter, we allometrically scaled FMD to normalize the flow-mediated response to variability in baseline diameter between the sexes. However, there were no sex differences in allometrically scaled FMD (sex p=0.11, time p=0.20, interaction p= 0.072; Scaled FMD (%): women BL: 6.39 ± 3.7, 24H: 4.32 ± 3.8, 48H: 6.92 ± 3.7; men BL: 7.03 ± 3.9, 24H: 8.71 ± 4.2, 48H: 8.69 ± 4.3). When estrogen concentrations are low, women show similar inflammatory responses and vascular function as compared with age-matched men, suggesting a primary role of estrogen in previously observed sex differences in inflammatory responses and vascular function.

Sex-specific cytokine responses and neurocognitive outcome after blood transfusions in preterm infants.

BACKGROUND:The objective of this study was to determine sex-specific differences in inflammatory cytokine responses to RBC transfusion in preterm infants in the neonatal period and their relationship to later neurocognitive status.METHODS:Infants with a birth weight <1000 grams and gestational age 22–29 weeks were enrolled in the Transfusion of Prematures (TOP) trial. The total number of transfusions was used as a marker of transfusion status. 19 cytokines and biomarkers were analyzed from 71 infants longitudinally during neonatal period. 26 infants completed the Bayley Scales of Infant & Toddler Development, 3rd Edition (Bayley-III) at 12 months’ corrected age.RESULTS:Nine cytokine levels were significantly elevated in proportion to the number of transfusions received. Of those, one cytokine showed a sex-specific finding (p=0.004): monocyte chemoattractant protein, MCP-1, rose substantially in females (8.9% change per additional transfusion), but not males (−0.8% change). Higher concentrations of MCP-1 exclusively were associated with worse Bayley-III scores: decreased cognitive raw scores (p=0.0005) and motor scaled scores (p<0.0001).CONCLUSION:This study provides evidence of a sex-specific difference in the inflammatory response to RBC transfusions during neonatal life, with MCP-1 levels rising only in females and inversely correlating with neurocognitive status at 12 months old.

Comparison of Host Cytokine Response in Piglets Infected With Toxigenic and Non-toxigenic Staphylococcus hyicus.

Staphylococcus hyicus is the most common causative agent of exudative epidermitis (EE) in piglets. Staphylococcus hyicus can be grouped into toxigenic and non-toxigenic strains based on its ability to cause EE in pigs. However, the inflammatory response of piglets infected with toxigenic and non-toxigenic S. hyicus has not been elucidated. In this study, we evaluated the serum cytokine profile in piglets inoculated with toxigenic and non-toxigenic S. hyicus strains and recorded the clinical signs in piglets. Fifteen piglets were divided into three groups (n = 5) and inoculated with a toxigenic strain (ZC-4), a non-toxigenic strain (CF-1), and PBS (control), respectively. The changes in serum levels of cytokines (interleukin [IL]-1β, IL-4, IL-6, IL-8, IL-10, IL-12, granulocyte-macrophage colony-stimulating factor, interferon-γ, transforming growth factor-β1, and tumor necrosis factor-α) were evaluated using a cytokine array at 6, 24, 48, and 72 h post inoculation. The results showed that piglets infected with the toxigenic strain exhibited more severe clinical signs and higher mortality than those infected with the non-toxigenic strain. The serum levels of pro-inflammatory cytokine IL-1β were significantly increased in toxigenic-and non-toxigenic-strain-infected piglets compared to those in the control group (p < 0.05), while the anti-inflammatory cytokine IL-10 was significantly up-regulated only in toxigenic group than in control group (p < 0.05). These results indicated that piglets infected with toxigenic and non-toxigenic S. hyicus showed differential infection status and inflammatory responses. Both toxigenic- and non-toxigenic- S. hyicus infection could induce a pro-inflammatory reaction in piglets. In addition, the toxigenic strain induced a strong anti-inflammatory response in piglets as indicated by the increased serum level of IL-10, which may be associated with the severe clinical signs and increased mortality and may be the key cytokine response responsible for pathogenic mechanisms of S. hyicus.

Tissue-specific endothelial cell heterogeneity contributes to unequal inflammatory responses

Endothelial cells (EC) coordinate vascular homeostasis and inflammation. In organ transplantation, EC are a direct alloimmune target. We posited that tissue specific heterogeneity of vascular EC may partly underlie the disparate organ-specific alloimmune risk. We examined the vascular endothelial response to inflammation across six primary endothelial beds from four major transplanted organs: the heart, lung, kidney and liver. First, we reanalyzed a public dataset of cardiac allograft rejection and found that endothelial inflammatory response genes were elevated in human cardiac allograft biopsies undergoing rejection compared with stable grafts. Next, the inducible inflammatory phenotypes of EC from heart, lung, kidney, and liver were characterized in vitro, focused on expression of adhesion molecules and chemokines, and recruitment of allogeneic peripheral blood mononuclear immune cells. Large vessel cardiac EC most highly upregulated VCAM-1, particularly compared with hepatic EC, supported greater leukocyte adhesion and had distinct chemokine profiles after stimulation with cytokines and complement. Differentially expressed gene candidates that are known regulators of cytokine signaling and inflammatory programming were verified in publicly available datasets of organ-specific endothelial transcriptomes. In summary, differential baseline expression of immune regulating genes may contribute to differential vascular inflammatory responses depending on organ.

The effects of foods available through the Food Distribution Program on Indian Reservations (FDPIR) on inflammation response, appetite and energy intake.

To compare the effects of a typical Food Distribution Program on Indian Reservations (FDPIR) diet with an FDPIR diet that meets Dietary Guidelines for Americans (DGA) on inflammation response, appetite and energy intake on a combination of American Indian (AI) and non-AI individuals. A within-subjects, randomised, crossover design was used to compare two dietary conditions: (1) a FDPIR diet that met DGA and (2) a FDPIR diet that did not meet DGA. Each participant served as their own control and was exposed to both dietary conditions. Repeated-measures ANOVA and t tests assessed significance between the two dietary conditions. This took place in the Montana State University Nutrition Research Laboratory in the USA. Female and male participants (n 13) aged 18-55 years from the university and local community. There were no significant differences in inflammatory response and appetite sensations between the two dietary conditions. Findings indicated that participants ate 14 % more (P < 0·01) kcal on a typical FDPIR diet compared with a FDPIR diet that met DGA. Higher energy intake during a typical FDPIR diet compared with a FDPIR diet that meets DGA may increase risk for obesity and nutrition-related diseases, including type 2 diabetes and other chronic inflammatory conditions.

Differences in inflammatory responses of rainbow trout infected by two genotypes of the myxozoan parasite Ceratonova shasta

Two genotypes of the intestinal parasite Ceratonova shasta infect Oncorhynchus mykiss: genotype 0 results in a chronic infection with low mortality while genotype IIR causes disease with high mortality. We determined parasite load and the relative expression of six immune factors (IgT, IgM, IL-6, IL-8, IL-10, IFNG) in fish infected with either genotype over 29 days post-exposure. In genotype IIR infections the host responded with upregulation of inflammatory and regulatory cytokines. In contrast, genotype 0 infection did not elicit an inflammatory response and expression of IFNG and IL-10 was lower. Antibody expression was upregulated in both infections but appeared to have limited efficacy in the virulent genotype IIR infections. Histologically, in genotype 0 infections the parasite migrated through the tissue layers causing inflammation but minimal damage to the mucosal epithelium, which contrasts with the severe pathology found in genotype IIR infections.

Histopathologic Lesion Scoring and Histomorphometric Methods for Measuring Vaccine Reactions in the Trachea of Broiler Chickens.

Severity of the tracheal histologic inflammatory response induced in broilers by ocular inoculation of two infectious bronchitis (IBV) and three Newcastle disease virus (NDV) commercial vaccines were evaluated. The vaccine was delivered by eye drop with a coarse spray to day-old chicks. The vaccines were given individually or in various combinations and were evaluated relative to nonvaccinated controls. Evaluations were performed on postvaccination (PV) days 7 and 14. Histologic endpoints included semiquantitative severity scoring of inflammatory components and quantitative morphometric determinations of inflammatory cell concentration, mucosal thickness, and percentage of ciliated mucosal surface. Strong positive correlations were observed between routine severity scoring and morphometric inflammatory parameters, whereas a negative correlation was present between inflammation severity and the percentage of mucosal ciliation. Variable, sometimes extensive, and often statistically significant differences in inflammatory responses were observed between the various vaccines. One IBV Massachusetts strain vaccine (IBV-A) produced the greatest overall inflammatory response when given alone or in combination with the NDV vaccines. Enhancement of tracheitis was seen on PV day 14 by covaccination of IBV-A with the NDV vaccines, but not by covaccination of another IBV Massachusetts strain vaccine (IBV-B) with NDV. Reduction in cilia percentage was observed for all vaccine groups relative to controls on PV day 7. However, although reactive cilia regeneration occurred on PV day 14 for most vaccine groups, a cilia regenerative response was not apparent for individual or NDV combination vaccination for IBV-A. The study also demonstrates that substantial microscopic trachea pathology may be present in vaccinated birds not exhibiting apparent clinical respiratory signs.

Rabies viruses of different virulence regulates inflammatory responses both in vivo and in vitro via MAPK and NF-κB pathway

Immune responses and central nervous system dysfunction are two main factors to be considered during rabies virus (RABV) infection. However, the mechanisms by which RABV strains of different virulence regulate with chemokine expression and the signaling pathways responsible for the immune responses in the terminal stage of infection both in vivo and in vitro have not been fully elucidated. In this study, we found low expression levels of proinflammatory chemokines in the mouse brain upon infection with street RABV strains (CXZ17 and HN10) at the late stage of infection. We also examined the difference in inflammatory response upon infection with RABV strains of different virulence in a mouse model. We found that the expression of proinflammatory chemokines increased to a varying degree upon infection with street RABV (CXZ17 and HN10) or laboratory-fixed RABV (CVS-11, aG, and CTN); CXCL10, CCL5, and CCL2 were the most significantly upregulated chemokines in brain tissue and microglial BV-2 cells in response to infection with RABV strains of different virulence. Our data also demonstrate significant activation of the MAPK and NF-κB pathways in mouse brain tissue at the late stage of RABV infection. We also found (i) low phosphorylation signals of MAPK and NF-κB p65 in neuronal cells upon infection with CXZ17 and HN10 in the mouse brain and (ii) strong phosphorylation signals in cerebrovascular endothelial cells and neuronal cells upon CTN or aG infection. Moreover, we quantified the nuclear localization status of MAPK signals and NF-κB p65 upon infection with CVS-11, aG, and CTN in BV-2 cells in vitro. We also found (i) that the activation of the p38, ERK1/2, and NF-κB p65 pathway, which stimulates CXCL10, CCL5, and CCL2 expression upon infection with RABV strains of different virulence (aG, CTN, and CVS-11), is triggered after virus entry into BV-2 cells and (ii) that the expression of CXCL10, CCL5, and CCL2 is required for the activation of NF-κB, p38, and ERK1/2, but not JNK. Overall, our study provides insight into the regulation of inflammatory responses mediated by MAPK and NF-κB in the mouse brain and in microglial cells upon RABV infection of different virulence.

Chikungunya virus infection induces differential inflammatory and antiviral responses in human monocytes and monocyte-derived macrophages

Chikungunya virus (CHIKV) is a zoonotic arthropod-borne virus that has caused several outbreaks in tropical and subtropical areas worldwide during the last 50 years. The virus is known to target different human cell types throughout the course of infection including epithelial and endothelial cells, fibroblasts, primary monocytes and monocyte-derived macrophages (MDMs). The two latter are phagocytic cell populations of the innate immune system which are involved in some aspects of CHIKV pathogenesis. However, monocytes and macrophages also potentially contribute to the control of viral replication through the expression of different pattern recognition receptors sensing viral pathogens and subsequently, inducing an type I interferone (IFN-I)-dependent antiviral immune response. The aim of this study was to determine the modulation of the expression of Toll-like receptors (TLRs), cytokine secretion capabilities and antiviral factor production in monocytes and MDMs following infection with CHIKV. Moreover, we sought to determine the replication kinetics of CHIKV in these two cell populations. We found that the maximum peak of CHIKV replication was observed between 18- and 24-hours post-infection (hpi), while after that the is strongly reduced. Furthermore, CHIKV infection induced the pro-inflammatory cytokine production starting from the first 6 hpi in both monocytes and MDMs, with similar kinetics but different protein levels. In contrast, the kinetics of transcriptional expression of some TLRs were different between both cell types. In addition, IFN-I, 2′,5′-oligoadenylate synthetase 1 (OAS1), and double-stranded RNA-activated protein kinase R (PKR) mRNA levels were detected in response to CHIKV infection of monocytes and MDMs, resulting the highest expression levels at 48 hpi. In conclusion, our data provides evidence that CHIKV infection activates the TLR pathways in primary monocytes and MDMs, which play a crucial role in CHIKV pathogenesis and/or host defense, differentially. However, additional studies are required to determine the functional role of TLRs in monocytes and MDMs.

Single-Cell Analyses of Colon and Blood Reveal Distinct Immune Cell Signatures of Ulcerative Colitis and Crohn’s Disease

Studies are needed to determine the mechanisms of mucosal dysregulation in patients with inflammatory bowel diseases (IBDs) and differences in inflammatory responses of patients with ulcerative colitis (UC) vs Crohn's disease (CD). We used mass cytometry (CyTOF) to characterize and compare immune cell populations in the mucosa and blood from patients with IBD and without IBD (controls) at single-cell resolution. We performed CyTOF analysis of colonic mucosa samples (n= 87) and peripheral blood mononuclear cells (n= 85) from patients with active or inactive UC or CD and controls. We also performed single-cell RNA sequencing, flow cytometry, and RNA in situ hybridization analyses to validate key findings. We used random forest modeling to identify differences in signatures across subject groups. Compared with controls, colonic mucosa samples from patients with IBD had increased abundances of HLA-DR+CD38+ T cells, including T-regulatory cells that produce inflammatory cytokines; CXCR3+ plasmablasts; and IL1B+ macrophages and monocytes. Colonic mucosa samples from patients with UC were characterized by expansion of IL17A+ CD161+ effector memory T cells and IL17A+ T-regulatory cells; expansion of HLA-DR+CD56+ granulocytes; and reductions in type 3 innate lymphoid cells. Mucosal samples from patients with active CD were characterized by IL1B+HLA-DR+CD38+ T cells, IL1B+TNF+IFNG+ naïve B cells, IL1B+ dendritic cells (DCs), and IL1B+ plasmacytoid DCs. Peripheral blood mononuclear cells from patients with active CD differed from those of active UC in that the peripheral blood mononuclear cells from patients with CD had increased IL1B+ T-regulatory cells, IL1B+ DCs and IL1B+ plasmacytoid DCs, IL1B+ monocytes, and fewer group 1 innate lymphoid cells. Random forest modeling differentiated active UC from active CD in colonic mucosa and blood samples; top discriminating features included many of the cellular populations identified above. We used single-cell technologies to identify immune cell populations specific to mucosa and blood samples from patients with active or inactive CD and UC and controls. This information might be used to develop therapies that target specific cell populations in patients with different types of IBD.

A Pro- and Anti-inflammatory Axis Modulates the Macrophage Circadian Clock.

The circadian clock broadly governs immune cell function, leading to time-of-day differences in inflammatory responses and subsequently, pathogen clearance. However, the effect of inflammatory signals on circadian machinery is poorly understood. We found that in bone marrow-derived macrophages, some host-derived pro-inflammatory cytokines, e.g., IFN-γ or TNF-α, and pathogen-associated molecular patterns, e.g., LPS or Pam3Csk4, suppress the amplitude in oscillations of circadian negative feedback arm clock components such as PER2, and when examined, specific combinations of these immune-related signals suppressed the amplitude of these oscillations to a greater degree in both bone marrow-derived and peritoneal macrophages. At the transcript level, multiple components of the circadian clock were affected in different ways by pro-inflammatory stimulus, including Per2 and Nr1d1. This suppressive effect on PER2 did not arise from nor correlate with cell death or clock resetting. Suppression of the clock by IFN-γ was dependent on its cognate receptor; however, pharmacological inhibition of the canonical JAK/STAT and MEK pathways did not hinder suppression, suggesting a mechanism involving a non-canonical pathway. In contrast, anti-inflammatory signals such as IL-4 and dexamethasone enhanced the expression of PER2 protein and Per2 mRNA. Our results suggest that the circadian system in macrophages can differentially respond to pro- and anti-inflammatory signals in their microenvironments.

Sex Differences in the Oxidative Stress and Inflammation Response During and After Simulated Hemorrhage in Humans

IntroductionHemorrhage (i.e., massive blood loss) induces an oxidative stress and inflammatory response that can persist even following hemostasis and resuscitation. These responses can result in tissue and organ damage if therapeutic inventions are delayed. Pre‐menopausal females exhibit a survival advantage following hemorrhage compared to males of a similar age. The role of oxidative stress and inflammation on increased survival from blood loss in young females is under investigation. Examining the potential mechanisms underpinning this survival advantage following hemorrhage may facilitate the development of sex‐specific therapies. In our laboratory, we utilize lower body negative pressure (LBNP) to simulate blood loss in conscious human subjects. In this study, we hypothesized that young males would elicit a greater oxidative stress and inflammatory response compared to young females, both during and after a simulated hemorrhage via LBNP.MethodsYoung, healthy human subjects (10F; 10M) participated in a stepwise‐LBNP protocol to presyncope. Stroke volume was estimated via finger photoplethysmography as a marker of central hypovolemia (indexed to body surface area). Venous blood samples were collected at baseline, at the onset of presyncope, and 60‐min into recovery (i.e., following “resuscitation”). The oxidative stress response was assessed via measurement of circulating F2‐Isoprostanes (F2‐IsoP) using gas chromatography‐negative ion chemical ionization‐mass spectrometry. The inflammatory response was assessed via measurement of circulating interleukin (IL)‐6 and IL‐10 using a MSD® Multiplex assay. Unpaired t‐tests were used to compare LBNP tolerance and stroke volume responses between male and female subjects. Two factor (time and sex) linear mixed model analyses with repeated measures were performed for all other comparisons, followed by Holm‐corrected post‐hoc tests. All data are represented as mean ± SE.ResultsLBNP tolerance time was similar between male and female subjects (Males, 1592 ± 124 s vs. Females, 1437 ± 113 s; P=0.37), and stroke volume index decreased by a similar magnitude at presyncope (Males, −50.2 ± 6.3% vs. Females, −49.4 ± 3.2%; P = 0.87). There was no effect of time or sex on [F2‐IsoP] or on the %Δ [F2‐IsoP] during or after LBNP (P ≥ 0.12). However, male subjects exhibited a greater increase in both the %Δ [IL‐6] and %Δ [IL‐10] compared to female subjects at the 60‐min recovery time point (IL‐6: Males, 101.4 ± 138.9% vs. Females, 12.3 ± 34.0%; P = 0.06. IL‐10: Males, 71.1 ± 133.3% vs. Females, −2.2 ± 11.8%; P = 0.06).ConclusionThese data suggest that there may be a sex difference in the inflammatory response to blood loss and subsequent fluid resuscitation. Future clinical studies should continue to explore sex differences in inflammatory responses to actual blood loss, and tailor therapeutic interventions accordingly.Support or Funding InformationWilliam and Ella Owens Medical Research Foundation

The effect of quantity and quality of dietary fat intake on subcutaneous white adipose tissue inflammatory responses.

The global prevalence of obesity and obesity-associated cardiometabolic diseases is a significant public health burden. Chronic low-grade inflammation in metabolic tissues such as white adipose tissue (WAT) is linked to obesity and may play a role in disease progression. The overconsumption of dietary fat has been suggested to modulate the WAT inflammatory environment. It is also recognised that fats varying in degree of fatty acid saturation may elicit differential WAT inflammatory responses. This information has originated predominantly from animal or cell models and translation into human participants in vivo remains limited. This review will summarise human intervention studies investigating the effect of dietary fat quantity and quality on subcutaneous WAT inflammation, with a specific focus on the toll-like receptor 4 (TLR4)/NF-κB and nucleotide-binding and oligomerisation domain-like receptor, leucine-rich repeat and pyrin domain-containing 3 (NLRP3) inflammasome molecular signalling pathways. Overall, firm conclusions are hard to draw regarding the effect of dietary fat quantity and quality on WAT inflammatory responses due to the heterogeneity of study designs, diet composition and participant cohorts recruited. Previous studies have predominantly focused on measures of WAT gene expression. It is suggested that future work includes measures of WAT total content and phosphorylation of proteins involved in TLR4/NF-κB and NLRP3 signalling as this is more representative of alterations in WAT physiological function. Understanding pathways linking the intake of total fat and specific fatty acids with WAT metabolic-inflammatory responses may have important implications for public health by informing dietary guidelines aimed at cardiometabolic risk reduction.

M. tuberculosis infection of human iPSC-derived macrophages reveals complex membrane dynamics during xenophagy evasion.

ABSTRACTXenophagy is an important cellular defence mechanism against cytosol-invading pathogens, such as Mycobacterium tuberculosis (Mtb). Activation of xenophagy in macrophages targets Mtb to autophagosomes; however, how Mtb is targeted to autophagosomes in human macrophages at a high spatial and temporal resolution is unknown. Here, we use human induced pluripotent stem cell-derived macrophages (iPSDMs) to study the human macrophage response to Mtb infection and the role of the ESX-1 type VII secretion system. Using RNA-seq, we identify ESX-1-dependent transcriptional responses in iPSDMs after infection with Mtb. This analysis revealed differential inflammatory responses and dysregulated pathways such as eukaryotic initiation factor 2 (eIF2) signalling and protein ubiquitylation. Moreover, live-cell imaging revealed that Mtb infection in human macrophages induces dynamic ESX-1-dependent, LC3B-positive tubulovesicular autophagosomes (LC3-TVS). Through a correlative live-cell and focused ion beam scanning electron microscopy (FIB SEM) approach, we show that upon phagosomal rupture, Mtb induces the formation of LC3-TVS, from which the bacterium is able to escape to reside in the cytosol. Thus, iPSDMs represent a valuable model for studying spatiotemporal dynamics of human macrophage–Mtb interactions, and Mtb is able to evade capture by autophagic compartments.

Enteroids Derived From Inflammatory Bowel Disease Patients Display Dysregulated Endoplasmic Reticulum Stress Pathways, Leading to Differential Inflammatory Responses and Dendritic Cell Maturation.

Endoplasmic reticulum [ER] stress in intestinal epithelial cells [IECs] contributes to the pathogenesis of inflammatory bowel disease [IBD]. We hypothesized that ER stress changes innate signalling in human IECs, augmenting toll-like receptor [TLR] responses and inducing pro-inflammatory changes in underlying dendritic cells [DCs]. Caco-2 cells and primary human colon-derived enteroid monolayers were exposed to ATP [control stressor] or thapsigargin [Tg] [ER stress inducer], and were stimulated with the TLR5 agonist flagellin. Cytokine release was measured by an enzyme immunoassay. ER stress markers CHOP, GRP78 and XBP1s/u were measured via quantitative PCR and Western blot. Monocyte-derived DCs [moDCs] were cultured with the IEC supernatants and their activation state was measured. Responses from enteroids derived from IBD patients and healthy control participants were compared. ER stress enhanced flagellin-induced IL-8 release from Caco-2 cells and enteroids. Moreover, conditioned media activated DCs to become pro-inflammatory, with increased expression of CD80, CD86, MHCII, IL-6, IL-15 and IL-12p70 and decreased expression of CD103 and IL-10. Flagellin-induced IL-8 production correlated with DC activation, suggesting a common stress pathway. Moreover, there were distinct differences in cytokine expression and basal ER stress between IBD and healthy subject-derived enteroid monolayers, suggesting a dysregulated ER stress pathway in IBD-derived enteroids. Cellular stress enhances TLR5 responses in IECs, leading to increased DC activation, indicating a previously unknown mechanistic link between epithelial ER stress and immune activation in IBD. Furthermore, dysregulated ER stress may be propagated from the intestinal epithelial stem cell niche in IBD patients.