Pseudomonas aeruginosa pulmonary infection results in S100A8/A9-dependent cardiac dysfunction.

Pseudomonas aeruginosa (P.a.) infection accounts for nearly 20% of all cases of hospital acquired pneumonia with mortality rates >30%. P.a. infection induces a robust inflammatory response, which ideally enhances bacterial clearance. Unfortunately, excessive inflammation can also have negative effects, and often leads to cardiac dysfunction with associated morbidity and mortality. However, it remains unclear how P.a. lung infection causes cardiac dysfunction. Using a murine pneumonia model, we found that P.a. infection of the lungs led to severe cardiac left ventricular dysfunction and electrical abnormalities. More specifically, we found that neutrophil recruitment and release of S100A8/A9 in the lungs activates the TLR4/RAGE signaling pathways, which in turn enhance systemic inflammation and subsequent cardiac dysfunction. Paradoxically, global deletion of S100A8/A9 did not improve but aggravated cardiac dysfunction and mortality likely due to uncontrolled bacterial burden in the lungs and heart. Our results indicate that P.a. infection induced release of S100A8/9 is double-edged, providing increased risk for cardiac dysfunction yet limiting P.a. growth.

Impact of age and sex on myelopoiesis and inflammation during myocardial infarction.

Of all the different risk factors known to cause cardiovascular disease (CVD), age and sex are considered to play a crucial role. Aging follows a continuum from birth to death, and therefore it inevitably acts as a risk for CVD. Along with age, sex differences have also been shown to demonstrate variations in immune system responses to pathological insults. It has been widely perceived that females are protected against myocardial infarction (MI) and the protection is quite apparent in young vs. old women. Acute MI leads to changes in the population of myeloid and lymphoid cells at the injury site with myeloid bias being observed in the initial inflammation and the lymphoid in the late-resolution phases of the pathology. Multiple evidence demonstrates that aging enhances damage to various cellular processes through inflamm-aging, an inflammatory process identified to increase pro-inflammatory markers in circulation and tissues. Following MI, marked changes were observed in different sub-sets of major myeloid cell types viz., neutrophils, monocytes, and macrophages. There is a paucity of information regarding the tissue and site-specific functions of these sub-sets. In this review, we highlight the importance of age and sex as crucial risk factors by discussing their role during MI-induced myelopoiesis while emphasizing the current status of myeloid cell sub-sets. We further put forth the need for designing and executing age and sex interaction studies aimed to determine the appropriate age and sex to develop personalized therapeutic strategies post-MI.

Retention of the NLRP3 Inflammasome-Primed Neutrophils in the Bone Marrow Is Essential for Myocardial Infarction-Induced Granulopoiesis.

BACKGROUND: Acute myocardial infarction (MI) results in overzealous production and infiltration of neutrophils to the ischemic heart. This is mediated in part by granulopoiesis induced by the S100A8/A9-NLRP3-IL-1ß signaling axis in injury-exposed neutrophils. Despite the transcriptional upregulation of the NLRP3 (Nod Like Receptor Family Pyrin Domain-Containing 3) inflammasome and associated signaling components in neutrophils, the serum levels of IL-1ß (interleukin-1ß), the effector molecule in granulopoiesis, were not affected by MI, suggesting that IL-1ß is not released systemically. We hypothesize that IL-1ß is released locally within the bone marrow (BM) by inflammasome-primed and reverse-migrating neutrophils. METHODS: Using a combination of time-dependent parabiosis and flow cytometry techniques, we first characterized the migration patterns of different blood cell types across the parabiotic barrier. We next induced MI in parabiotic mice by permanent ligation of the left anterior descending artery and examined the ability of injury-exposed neutrophils to permeate the parabiotic barrier and induce granulopoiesis in noninfarcted parabionts. Last, using multiple neutrophil adoptive and BM transplant studies, we studied the molecular mechanisms that govern reverse migration and retention of the primed neutrophils, IL-1ß secretion, and granulopoiesis. Cardiac function was assessed by echocardiography. RESULTS: MI promoted greater accumulation of the inflammasome-primed neutrophils in the BM. Introducing a time-dependent parabiotic barrier to the free movement of neutrophils inhibited their ability to stimulate granulopoiesis in the noninfarcted parabionts. Previous priming of the NLRP3 inflammasome is not a prerequisite, but the presence of a functional CXCR4 (C-X-C-motif chemokine receptor 4) on the primed-neutrophils and elevated serum S100A8/A9 levels are necessary for homing and retention of the reverse-migrating neutrophils. In the BM, the primed-neutrophils secrete IL-1ß through formation of gasdermin D pores and promote granulopoiesis. Pharmacological and genetic strategies aimed at the inhibition of neutrophil homing or release of IL-1ß in the BM markedly suppressed MI-induced granulopoiesis and improved cardiac function. CONCLUSIONS: Our data reveal a new paradigm of how circulatory cells establish a direct communication between organs by delivering signaling molecules (eg, IL-1ß) directly at the sites of action rather through systemic release. We suggest that this pathway may exist to limit the off-target effects of systemic IL-1ß release.

Transient Intermittent Hyperglycemia Accelerates Atherosclerosis by Promoting Myelopoiesis.

RATIONALE: Treatment efficacy for diabetes mellitus is largely determined by assessment of HbA1c (glycated hemoglobin A1c) levels, which poorly reflects direct glucose variation. People with prediabetes and diabetes mellitus spend >50% of their time outside the optimal glucose range. These glucose variations, termed transient intermittent hyperglycemia (TIH), appear to be an independent risk factor for cardiovascular disease, but the pathological basis for this association is unclear. OBJECTIVE: To determine whether TIH per se promotes myelopoiesis to produce more monocytes and consequently adversely affects atherosclerosis. METHODS AND RESULTS: To create a mouse model of TIH, we administered 4 bolus doses of glucose at 2-hour intervals intraperitoneally once to WT (wild type) or once weekly to atherosclerotic prone mice. TIH accelerated atherogenesis without an increase in plasma cholesterol, seen in traditional models of diabetes mellitus. TIH promoted myelopoiesis in the bone marrow, resulting in increased circulating monocytes, particularly the inflammatory Ly6-Chi subset, and neutrophils. Hematopoietic-restricted deletion of S100a9, S100a8, or its cognate receptor Rage prevented monocytosis. Mechanistically, glucose uptake via GLUT (glucose transporter)-1 and enhanced glycolysis in neutrophils promoted the production of S100A8/A9. Myeloid-restricted deletion of Slc2a1 (GLUT-1) or pharmacological inhibition of S100A8/A9 reduced TIH-induced myelopoiesis and atherosclerosis. CONCLUSIONS: Together, these data provide a mechanism as to how TIH, prevalent in people with impaired glucose metabolism, contributes to cardiovascular disease. These findings provide a rationale for continual glucose control in these patients and may also suggest that strategies aimed at targeting the S100A8/A9-RAGE (receptor for advanced glycation end products) axis could represent a viable approach to protect the vulnerable blood vessels in diabetes mellitus. Graphic Abstract: A graphic abstract is available for this article.

Apoptotic Ablation of Platelets Reduces Atherosclerosis in Mice With Diabetes.

OBJECTIVE: People with diabetes are at a significantly higher risk of cardiovascular disease, in part, due to accelerated atherosclerosis. Diabetic subjects have increased number of platelets that are activated, more reactive, and respond suboptimally to antiplatelet therapies. We hypothesized that reducing platelet numbers by inducing their premature apoptotic death would decrease atherosclerosis. Approach and Results: This was achieved by targeting the antiapoptotic protein Bcl-xL (B-cell lymphoma-extra large; which is essential for platelet viability) via distinct genetic and pharmacological approaches. In the former, we transplanted bone marrow from mice carrying the Tyr15 to Cys loss of function allele of Bcl-x (known as Bcl-xPlt20) or wild-type littermate controls into atherosclerotic-prone Ldlr+/- mice made diabetic with streptozotocin and fed a Western diet. Reduced Bcl-xL function in hematopoietic cells significantly decreased platelet numbers, exclusive of other hematologic changes. This led to a significant reduction in atherosclerotic lesion formation in Bcl-xPlt20 bone marrow transplanted Ldlr+/- mice. To assess the potential therapeutic relevance of reducing platelets in atherosclerosis, we next targeted Bcl-xL with a pharmacological strategy. This was achieved by low-dose administration of the BH3 (B-cell lymphoma-2 homology domain 3) mimetic, ABT-737 triweekly, in diabetic Apoe-/- mice for the final 6 weeks of a 12-week study. ABT-737 normalized platelet numbers along with platelet and leukocyte activation to that of nondiabetic controls, significantly reducing atherosclerosis while promoting a more stable plaque phenotype. CONCLUSIONS: These studies suggest that selectively reducing circulating platelets, by targeting Bcl-xL to promote platelet apoptosis, can reduce atherosclerosis and lower cardiovascular disease risk in diabetes. Graphic Abstract: A graphic abstract is available for this article.

Neutrophil-Derived S100A8/A9 Amplify Granulopoiesis After Myocardial Infarction.

BACKGROUND: Myocardial infarction (MI) triggers myelopoiesis, resulting in heightened production of neutrophils. However, the mechanisms that sustain their production and recruitment to the injured heart are unclear. METHODS: Using a mouse model of the permanent ligation of the left anterior descending artery and flow cytometry, we first characterized the temporal and spatial effects of MI on different myeloid cell types. We next performed global transcriptome analysis of different cardiac cell types within the infarct to identify the drivers of the acute inflammatory response and the underlying signaling pathways. Using a combination of genetic and pharmacological strategies, we identified the sequelae of events that led to MI-induced myelopoiesis. Cardiac function was assessed by echocardiography. The association of early indexes of neutrophilia with major adverse cardiovascular events was studied in a cohort of patients with acute MI. RESULTS: Induction of MI results in rapid recruitment of neutrophils to the infarct, where they release specific alarmins, S100A8 and S100A9. These alarmins bind to the Toll-like receptor 4 and prime the nod-like receptor family pyrin domain-containing 3 inflammasome in naïve neutrophils and promote interleukin-1ß secretion. The released interleukin-1ß interacts with its receptor (interleukin 1 receptor type 1) on hematopoietic stem and progenitor cells in the bone marrow and stimulates granulopoiesis in a cell-autonomous manner. Genetic or pharmacological strategies aimed at disruption of S100A8/A9 and their downstream signaling cascade suppress MI-induced granulopoiesis and improve cardiac function. Furthermore, in patients with acute coronary syndrome, higher neutrophil count on admission and after revascularization correlates positively with major adverse cardiovascular disease outcomes. CONCLUSIONS: Our study provides novel evidence for the primary role of neutrophil-derived alarmins (S100A8/A9) in dictating the nature of the ensuing inflammatory response after myocardial injury. Therapeutic strategies aimed at disruption of S100A8/A9 signaling or their downstream mediators (eg, nod-like receptor family pyrin domain-containing 3 inflammasome, interleukin-1ß) in neutrophils suppress granulopoiesis and may improve cardiac function in patients with acute coronary syndrome.

Bone Marrow-Derived Cells Restore Functional Integrity of the Gut Epithelial and Vascular Barriers in a Model of Diabetes and ACE2 Deficiency.

RATIONALE: There is incomplete knowledge of the impact of bone marrow cells on the gut microbiome and gut barrier function. OBJECTIVE: We postulated that diabetes mellitus and systemic ACE2 (angiotensin-converting enzyme 2) deficiency would synergize to adversely impact both the microbiome and gut barrier function. METHODS AND RESULTS: Bacterial 16S rRNA sequencing and metatranscriptomic analysis were performed on fecal samples from wild-type, ACE2-/y, Akita (type 1 diabetes mellitus), and ACE2-/y-Akita mice. Gut barrier integrity was assessed by immunofluorescence, and bone marrow cell extravasation into the small intestine was evaluated by flow cytometry. In the ACE2-/y-Akita or Akita mice, the disrupted barrier was associated with reduced levels of myeloid angiogenic cells, but no increase in inflammatory monocytes was observed within the gut parenchyma. Genomic and metatranscriptomic analysis of the microbiome of ACE2-/y-Akita mice demonstrated a marked increase in peptidoglycan-producing bacteria. When compared with control cohorts treated with saline, intraperitoneal administration of myeloid angiogenic cells significantly decreased the microbiome gene expression associated with peptidoglycan biosynthesis and restored epithelial and endothelial gut barrier integrity. Also indicative of diabetic gut barrier dysfunction, increased levels of peptidoglycan and FABP-2 (intestinal fatty acid-binding protein 2) were observed in plasma of human subjects with type 1 diabetes mellitus (n=21) and type 2 diabetes mellitus (n=23) compared with nondiabetic controls (n=23). Using human retinal endothelial cells, we determined that peptidoglycan activates a noncanonical TLR-2 (Toll-like receptor 2) associated MyD88 (myeloid differentiation primary response protein 88)-ARNO (ADP-ribosylation factor nucleotide-binding site opener)-ARF6 (ADP-ribosylation factor 6) signaling cascade, resulting in destabilization of p120-catenin and internalization of VE-cadherin as a mechanism of deleterious impact of peptidoglycan on the endothelium. CONCLUSIONS: We demonstrate for the first time that the defect in gut barrier function and dysbiosis in ACE2-/y-Akita mice can be favorably impacted by exogenous administration of myeloid angiogenic cells.

Attack of the NETs! NETosis primes IL-1ß-mediated inflammation in diabetic foot ulcers.

In volume 133 issue 4 of Clinical Science, Liu et al. showed that neutrophils release extracellular traps (NETs) in the setting of diabetes which acts as a stimulus for NLRP3 inflammasome activation in macrophages to promote IL1ß-dependent exacerbation of inflammation. They also provide evidence to show that degrading NETs improves the wound healing process. These findings provide an insight into how NETs communicate with other cells in the vicinity (e.g. macrophages) to exacerbate the inflammatory response. Most importantly, they provide novel avenues to improve wound healing process such as diabetic foot ulcers (DFUs) by targeting NETs.

Emerging roles of neutrophil-borne S100A8/A9 in cardiovascular inflammation.

Elevated neutrophil count is associated with higher risk of major adverse cardiac events including myocardial infarction and early development of heart failure. Neutrophils contribute to cardiac damage through a number of mechanisms, including attraction of other immune cells and release of inflammatory mediators. Recently, a number of independent studies have reported a causal role for neutrophil-derived alarmins (i.e. S100A8/A9) in inducing inflammation and cardiac injury following myocardial infarction (MI). Furthermore, a positive correlation between serum S100A8/A9 levels and major adverse cardiac events (MACE) in MI patients was also observed implying that targeting neutrophils or their inflammatory cargo could be beneficial in reducing heart failure. However, contradictory to this idea, neutrophils and neutrophil-derived S100A8/A9 also seem to play a vital role in the resolution of inflammation. Thus, a better understanding of how neutrophils balance these seemingly contrasting functions would allow us to develop effective therapies that preserve the inflammation-resolving function while restricting the damage caused by inflammation. In this review, we specifically discuss the mechanisms behind neutrophil-derived S100A8/A9 in promoting inflammation and resolution in the context of MI. We also provide a perspective on how neutrophils could be potentially targeted to ameliorate cardiac inflammation and the ensuing damage.

Chronic sympathetic driven hypertension promotes atherosclerosis by enhancing hematopoiesis.

Hypertension is a major, independent risk factor for atherosclerotic cardiovascular disease. However, this pathology can arise through multiple pathways, which could influence vascular disease through distinct mechanisms. An overactive sympathetic nervous system is a dominant pathway that can precipitate in elevated blood pressure. We aimed to determine how the sympathetic nervous system directly promotes atherosclerosis in the setting of hypertension. We used a mouse model of sympathetic nervous system-driven hypertension on the atherosclerotic-prone apolipoprotein E-deficient background. When mice were placed on a western type diet for 16 weeks, we showed the evolution of unstable atherosclerotic lesions. Fortuitously, the changes in lesion composition were independent of endothelial dysfunction, allowing for the discovery of alternative mechanisms. With the use of flow cytometry and bone marrow imaging, we found that sympathetic activation caused deterioration of the hematopoietic stem and progenitor cell niche in the bone marrow, promoting the liberation of these cells into the circulation and extramedullary hematopoiesis in the spleen. Specifically, sympathetic activation reduced the abundance of key hematopoietic stem and progenitor cell niche cells, sinusoidal endothelial cells and osteoblasts. Additionally, sympathetic bone marrow activity prompted neutrophils to secrete proteases to cleave the hematopoietic stem and progenitor cell surface receptor CXCR4. All these effects could be reversed using the ß-blocker propranolol during the feeding period. These findings suggest that elevated blood pressure driven by the sympathetic nervous system can influence mechanisms that modulate the hematopoietic system to promote atherosclerosis and contribute to cardiovascular events.

Defective cholesterol metabolism in haematopoietic stem cells promotes monocyte-driven atherosclerosis in rheumatoid arthritis.

Aim: Rheumatoid arthritis (RA) is associated with an approximately two-fold elevated risk of cardiovascular (CV)-related mortality. Patients with RA present with systemic inflammation including raised circulating myeloid cells, but fail to display traditional CV risk-factors, particularly dyslipidaemia. We aimed to explore if increased circulating myeloid cells is associated with impaired atherosclerotic lesion regression or altered progression in RA. Methods and results: Using flow cytometry, we noted prominent monocytosis, neutrophilia, and thrombocytosis in two mouse models of RA. This was due to enhanced proliferation of the haematopoietic stem and progenitor cells (HSPCs) in the bone marrow and the spleen. HSPCs expansion was associated with an increase in the cholesterol content, due to a down-regulation of cholesterol efflux genes, Apoe, Abca1, and Abcg1. The HSPCs also had enhanced expression of key myeloid promoting growth factor receptors. Systemic inflammation was found to cause defective cellular cholesterol metabolism. Increased myeloid cells in mice with RA were associated with a significant impairment in lesion regression, even though cholesterol levels were equivalent to non-arthritic mice. Lesions from arthritic mice exhibited a less stable phenotype as demonstrated by increased immune cell infiltration, lipid accumulation, and decreased collagen formation. In a progression model, we noted monocytosis, enhanced monocytes recruitment to lesions, and increased plaque macrophages. This was reversed with administration of reconstituted high-density lipoprotein (rHDL). Furthermore, RA patients have expanded CD16+ monocyte subsets and a down-regulation of ABCA1 and ABCG1. Conclusion: Rheumatoid arthritis impairs atherosclerotic regression and alters progression, which is associated with an expansion of myeloid cells and disturbed cellular cholesterol handling, independent of plasma cholesterol levels. Infusion of rHDL prevented enhanced myelopoiesis and monocyte entry into lesions. Targeting cellular cholesterol defects in people with RA, even if plasma cholesterol is within the normal range, may limit vascular disease.

Deficiency of ATP-binding cassette transporters A1 and G1 in macrophages increases inflammation and accelerates atherosclerosis in mice.

RATIONALE: Plasma high-density lipoprotein levels are inversely correlated with atherosclerosis. Although it is widely assumed that this is attributable to the ability of high-density lipoprotein to promote cholesterol efflux from macrophage foam cells, direct experimental support for this hypothesis is lacking. OBJECTIVE: To assess the role of macrophage cholesterol efflux pathways in atherogenesis. METHODS AND RESULTS: We developed mice with efficient deletion of the ATP-binding cassette transporters A1 and G1 (ABCA1 and ABCG1) in macrophages (MAC-ABC(DKO) mice) but not in hematopoietic stem or progenitor populations. MAC-ABC(DKO) bone marrow (BM) was transplanted into Ldlr(-/-) recipients. On the chow diet, these mice had similar plasma cholesterol and blood monocyte levels but increased atherosclerosis compared with controls. On the Western-type diet, MAC-ABC(DKO) BM-transplanted Ldlr(-/-) mice had disproportionate atherosclerosis, considering they also had lower very low-density lipoprotein/low-density lipoprotein cholesterol levels than controls. ABCA1/G1-deficient macrophages in lesions showed increased inflammatory gene expression. Unexpectedly, Western-type diet-fed MAC-ABC(DKO) BM-transplanted Ldlr(-/-) mice displayed monocytosis and neutrophilia in the absence of hematopoietic stem and multipotential progenitor cells proliferation. Mechanistic studies revealed increased expressions of machrophage colony stimulating factor and granulocyte colony stimulating factor in splenic macrophage foam cells, driving BM monocyte and neutrophil production. CONCLUSIONS: These studies show that macrophage deficiency of ABCA1/G1 is proatherogenic likely by promoting plaque inflammation and uncover a novel positive feedback loop in which cholesterol-laden splenic macrophages signal BM progenitors to produce monocytes, with suppression by macrophage cholesterol efflux pathways.

A novel role for bioactive lipids in stem cell mobilization during cardiac ischemia: new paradigms in thrombosis: novel mediators and biomarkers.

Despite major advances in pharmacological and reperfusion therapies, regenerating and/or replacing the infarcted myocardial tissue is an enormous challenge and therefore ischemic heart disease (IHD) remains a major cause of mortality and morbidity worldwide. Adult bone marrow is home for a variety of hematopoietic and non-hematopoietic stem cells including a small subset of primitive cells that carry a promising regenerative potential. It is now well established that myocardial ischemia (MI) induces mobilization of bone marrow-derived cells including differentiated lineage as well as undifferentiated stem cells. While the numbers of stem cells carrying pluripotent features among the mobilized stem cells is small, their regenerative capacity appears immense. Therapies aimed at selective mobilization of these pluripotent stem cells during myocardial ischemia have a promising potential to regenerate the injured myocardium. Emerging evidence suggest that bioactive sphingolipids such as sphingosine-1-phosphate and ceramide-1-phosphate hold a great promise in selective mobilization of pluripotent stem cells to the infarcted region during MI. This review highlights the recent advances in the mechanisms of stem cell mobilization and provides newer evidence in support of bioactive lipids as potential therapeutic agents in the treatment of ischemic heart disease.

Disulfiram Reduces Atherosclerosis and Enhances Efferocytosis, Autophagy, and Atheroprotective Gut Microbiota in Hyperlipidemic Mice.

BACKGROUND: Pyroptosis executor GsdmD (gasdermin D) promotes atherosclerosis in mice and humans. Disulfiram was recently shown to potently inhibit GsdmD, but the in vivo efficacy and mechanism of disulfiram's antiatherosclerotic activity is yet to be explored. METHODS AND RESULTS: We used human/mouse macrophages, endothelial cells, and smooth muscle cells and a hyperlipidemic mouse model of atherosclerosis to determine disulfiram antiatherosclerotic efficacy and mechanism. The effects of disulfiram on several atheroprotective pathways such as autophagy, efferocytosis, phagocytosis, and gut microbiota were determined. Atomic force microscopy was used to determine the effects of disulfiram on the biophysical properties of the plasma membrane of macrophages. Disulfiram-fed hyperlipidemic apolipoprotein E-/- mice showed significantly reduced interleukin-1ß release upon in vivo Nlrp3 (NLR family pyrin domain containing 3) inflammasome activation. Disulfiram-fed mice showed smaller atherosclerotic lesions (~27% and 29% reduction in males and females, respectively) and necrotic core areas (~50% and 46% reduction in males and females, respectively). Disulfiram induced autophagy in macrophages, smooth muscle cells, endothelial cells, hepatocytes/liver, and atherosclerotic plaques. Disulfiram modulated other atheroprotective pathways (eg, efferocytosis, phagocytosis) and gut microbiota. Disulfiram-treated macrophages showed enhanced phagocytosis/efferocytosis, with the mechanism being a marked increase in cell-surface expression of efferocytic receptor MerTK. Atomic force microscopy analysis revealed altered biophysical properties of disulfiram-treated macrophages, showing increased order-state of plasma membrane and increased adhesion strength. Furthermore, 16sRNA sequencing of disulfiram-fed hyperlipidemic mice showed highly significant enrichment in atheroprotective gut microbiota Akkermansia and a reduction in atherogenic Romboutsia species. CONCLUSIONS: Taken together, our data show that disulfiram can simultaneously modulate several atheroprotective pathways in a GsdmD-dependent as well as GsdmD-independent manner.

Syndecan-4 as a genetic determinant of the metabolic syndrome.

BACKGROUND: Syndecan-4 (SDC4) is a member of the heparan sulfate proteoglycan family of cell-surface receptors. We and others previously reported that variation in the SDC4 gene was associated with several components of the metabolic syndrome, including intra-abdominal fat, fasting glucose and triglyceride levels, and hypertension, in human cohorts. Additionally, we demonstrated that high fat diet (HFD)-induced obese female mice with a Sdc4 genetic deletion had higher visceral adiposity and a worse metabolic profile than control mice. Here, we aimed to first investigate whether the mouse Sdc4 null mutation impacts metabolic phenotypes in a sex- and diet-dependent manner. We then tested whether SDC4 polymorphisms are related to the metabolic syndrome (MetS) in humans. METHODS: For the mouse experiment, Sdc4-deficient (Sdc4-/-) and wild-type (WT) mice were treated with 14-weeks of low-fat diet (LFD). Body composition, energy balance, and selected metabolic phenotypes were assessed. For the human genetic study, we used logistic regression models to test 11 SDC4 SNPs for association with the MetS and its components in a cohort of 274 (113 with MetS) elderly subjects from southern Italy. RESULTS: Following the dietary intervention in mice, we observed that the effects of the Sdc4 null mutation on several phenotypes were different from those previously reported in the mice kept on an HFD. Nonetheless, LFD-fed female Sdc4-/- mice, but not males, displayed higher levels of triglycerides and lower insulin sensitivity at fasting than WT mice, as seen earlier in the HFD conditions. In the parallel human study, we found that carriers of SDC4 rs2228384 allele C and rs2072785 allele T had reduced risk of MetS. The opposite was true for carriers of the SDC4 rs1981429 allele G. Additionally, the SNPs were found related to fasting triglyceride levels and triglyceride glucose (TyG) index, a reliable indicator of insulin resistance, with sex-stratified analysis detecting the association of rs1981429 with these phenotypes only in females. CONCLUSIONS: Altogether, our results suggest that SDC4 is an evolutionary conserved genetic determinant of MetS and that its genetic variation is associated with fasting triglyceride levels in a female-specific manner.