Rationale: Pulmonary arterial hypertension (PAH) often results in death from right ventricular failure (RVF). NLRP3 (nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3)-macrophage activation may promote RVF in PAH. Objectives: Evaluating the contribution of the NLRP3 inflammasome in RV macrophages to PAH RVF. Methods: Rats with decompensated RV hypertrophy (monocrotaline [MCT] and Sugen-5416 hypoxia [SuHx]) were compared with compensated RV hypertrophy rats (pulmonary artery banding). Echocardiography and right heart catheterization were performed. Macrophages, atrial natriuretic peptides, and fibrosis were evaluated by microscopy or flow cytometry. NLRP3 inflammasome activation and cardiotoxicity were confirmed by immunoblot and in vitro strategies. MCT rats were treated with SC-144 (a GP130 antagonist) or MCC950 (an NLRP3 inhibitor). Macrophage–NLRP3 activity was evaluated in patients with PAH RVF. Measurements and Main Results: Macrophages, fibrosis, and atrial natriuretic peptides were increased in MCT and SuHx RVs but not in left ventricles or pulmonary artery banding rats. Although MCT RV macrophages were inflammatory, lung macrophages were antiinflammatory. CCR2⁺ macrophages (monocyte-derived) were increased in MCT and SuHx RVs and highly expressed NLRP3. The macrophage–NLRP3 pathway was upregulated in patients with PAH with decompensated RVs. Cultured MCT monocytes showed NLRP3 activation, and in coculture experiments resulted in cardiomyocyte mitochondrial damage, which MCC950 prevented. In vivo, MCC950 reduced NLRP3 activation and regressed pulmonary vascular disease and RVF. SC-144 reduced RV macrophages and NLRP3 content, prevented STAT3 (signal transducer and activator of transcription 3) activation, and improved RV function without regressing pulmonary vascular disease. Conclusions: NLRP3–macrophage activation occurs in the decompensated RV in preclinical PAH models and patients with PAH. Inhibiting GP130 or NLRP3 signaling improves RV function. The concept that PAH RVF results from RV inflammation rather than solely from elevated RV afterload suggests a new therapeutic paradigm.

Polycystic ovary syndrome (PCOS) is a continuum of endocrine and reproductive disorders characterized by hyperandrogenism, antral follicle growth arrest, and chronic inflammation. Macrophages play key role in inflammation, and the balance between M1 (inflammatory) and M2 (anti-inflammatory) macrophages determines physiological/pathological outcomes. Here, we investigated if hyperandrogenism increases ovarian chemerin altering the balance of M1 and M2 macrophages and the granulosa cell death. Ovarian chemerin was upregulated by 5α-dihydrotestosterone (DHT) in lean and overweight rats; while increased serum chemerin levels were only evident in overweight rats, suggesting that the serum chemerin may be reflective of a systemic response and associated with obesity, whereas increased ovarian chemerin expression is a localized response independent of the metabolic status. DHT altered follicle dynamics while increased the M1: M2 macrophages ratio in antral and pre-ovulatory follicles. While ovarian M1 macrophages expressing chemokine-like receptor 1 (CMKLR1) were increased, CMKLR1+ monocytes, which migrated toward chemerin-rich environment, were markedly decreased after 15 days of DHT. Androgen-induced granulosa cell apoptosis was dependent on the presence of macrophages. In humans, chemerin levels in follicular fluid, but not in serum, were higher in lean PCOS patients compared to BMI-matched controls and were associated with increased M1: M2 ratio. Our results support the concept that in PCOS, hyperandrogenemia increases chemerin expression while promotes CMKLR1+ monocytes recruitment and deregulates the immunological niche of ovaries. This study established a new immunological perspective in PCOS at the ovarian level. Hyperandrogenism is associated with upregulation of chemerin and macrophage unbalance in the ovaries.

BACKGROUND: Drp1 (dynamin-related protein 1), a large GTPase, mediates the increased mitochondrial fission, which contributes to hyperproliferation of pulmonary artery smooth muscle cells in pulmonary arterial hypertension (PAH). We developed a potent Drp1 GTPase inhibitor, Drpitor1a, but its specificity, pharmacokinetics, and efficacy in PAH are unknown. METHODS: Drpitor1a’s ability to inhibit recombinant and endogenous Drp1 GTPase was assessed. Drpitor1a’s effects on fission were studied in control and PAH human pulmonary artery smooth muscle cells (hPASMC) and blood outgrowth endothelial cells (BOEC). Cell proliferation and apoptosis were studied in hPASMC. Pharmacokinetics and tissue concentrations were measured following intravenous and oral drug administration. Drpitor1a’s efficacy in regressing monocrotaline-PAH was assessed in rats. In a pilot study, Drpitor1a reduced PA remodeling only in females. Subsequently, we compared Drpitor1a to vehicles in control and monocrotaline-PAH females. RESULTS: Drp1 GTPase activity was increased in PAH hPASMC. Drpitor1a inhibited the GTPase activity of recombinant and endogenous Drp1 and reversed the increased fission, seen in PAH hPASMC and PAH BOEC. Drpitor1a inhibited proliferation and induced apoptosis in PAH hPASMC without affecting electron transport chain activity, respiration, fission/fusion mediator expression, or mitochondrial Drp1 translocation. Drpitor1a did not inhibit proliferation or alter mitochondrial dynamics in normal hPASMC. Drpitor1a regressed monocrotaline-PAH without systemic vascular effects or toxicity. CONCLUSIONS: Drpitor1a is a specific Drp1 GTPase inhibitor that reduces mitochondrial fission in PAH hPASMC and PAH BOEC. Drpitor1a reduces proliferation and induces apoptosis in PAH hPASMC and regresses monocrotaline-PAH. Drp1 is a therapeutic target in PAH, and Drpitor1a is a potential therapy with an interesting therapeutic sexual dimorphism.

Type 1 Diabetes Mellitus (T1DM) is characterized by an augmented pro-inflammatory immune state. This contributes to the increased risk for gestational complications observed in T1DM mothers. In normal pregnancies, critical immunological changes occur, including the massive recruitment of lymphocytes, particularly CD56^bright NK cells, into early decidua basalis and a 2^nd trimester shift towards Type 2 immunity. Decidual CD56^bright NK cells arise at least partly from circulating progenitors expressing adhesion molecules SELL and ITGA4 and the chemokine receptors CXCR3 and CXCR4. In vitro studies show that T1DM reduces interactions between blood CD56⁺ NK cells and decidual endothelial cells by reducing SELL and ITGA4-based interactions. To address the mechanisms by which specific lymphocyte subsets may be recruited from the circulation during pregnancy and whether these mechanisms are altered in T1DM, flow cytometry was used to examine eight peripheral blood lymphocyte subsets (Type 1 (IL18R1⁺) and Type 2 (IL1RL1⁺) CD56^bright NK, CD56^dim NK, NKT and T cells) from control and T1DM women. Blood was collected serially over pregnancy and postpartum, and lymphocytes were compared for expression of homing receptors SELL, ITGA4, CXCR3, and CXCR4. The decline of Type 1/Type 2 immune cells in normal pregnancy was driven by an increase in Type 2 cells that did not occur in T1DM. CD56^bright NK cells from control women had the highest expression of all four receptors with greatest expression in 2^nd trimester. At this time, these receptors were expressed at very low levels by CD56^bright NK cells from TIDM patients. Type 1/Type 2 NKT cell ratios were not influenced by either pregnancy or TIDM. Our results suggest that T1DM alters immunological balances during pregnancy with its greatest impact on CD56^bright NK cells. This implicates CD56^bright NK cells in diabetic pregnancy complications.

Background: As partial pressure of oxygen (pO ₂ ) rises with the first breath, the ductus arteriosus (DA) constricts, diverting blood flow to the pulmonary circulation. The DA's O ₂ sensor resides within smooth muscle cells. The DA smooth muscle cells’ mitochondrial electron transport chain (ETC) produces reactive oxygen species (ROS) in proportion to oxygen tension, causing vasoconstriction by regulating redox‐sensitive ion channels and enzymes. To identify which ETC complex contributes most to DA O ₂ sensing and determine whether ROS mediate O ₂ sensing independent of metabolism, we used electron leak suppressors, S1QEL (suppressor of site I _Q electron leak) and S3QEL (suppressor of site III _Qo electron leak), which decrease ROS production by inhibiting electron leak from quinone sites I _Q and III _Qo , respectively. Methods and Results: The effects of S1QEL, S3QEL, and ETC inhibitors (rotenone and antimycin A) on DA tone, mitochondrial metabolism, O ₂ ‐induced changes in intracellular calcium, and ROS were studied in rabbit DA rings, and human and rabbit DA smooth muscle cells. S1QEL's effects on DA patency were assessed in rabbit kits, using micro computed tomography. In DA rings, S1QEL, but not S3QEL, reversed O ₂ ‐induced constriction ( P =0.0034) without reducing phenylephrine‐induced constriction. S1QEL did not inhibit mitochondrial metabolism or ETC‐I activity. In human DA smooth muscle cells, S1QEL and rotenone inhibited O ₂ ‐induced increases in intracellular calcium ( P =0.02 and 0.001, respectively), a surrogate for DA constriction. S1QEL inhibited O ₂ ‐induced ROS generation ( P =0.02). In vivo, S1QEL prevented O ₂ ‐induced DA closure ( P <0.0001). Conclusions: S1QEL, but not S3QEL, inhibited O ₂ ‐induced rises in ROS and DA constriction ex vivo and in vivo. DA O ₂ sensing relies on pO ₂ ‐dependent changes in electron leak at site I _Q in ETC‐I, independent of metabolism. S1QEL offers a therapeutic means to maintain DA patency.