FOXA1 Reprogramming Dictates Retinoid X Receptor Response in ESR1-Mutant Breast Cancer.

Estrogen receptor alpha (ER/ESR1) mutations occur in 30% to 40% of endocrine resistant ER-positive (ER+) breast cancer. Forkhead box A1 (FOXA1) is a key pioneer factor mediating ER-chromatin interactions and endocrine response in ER+ breast cancer, but its role in ESR1-mutant breast cancer remains unclear. Our previous FOXA1 chromatin immunoprecipitation sequencing (ChIP-seq) identified a large portion of redistributed binding sites in T47D genome-edited Y537S and D538G ESR1-mutant cells. Here, we further integrated FOXA1 genomic binding profile with the isogenic ER cistrome, accessible genome, and transcriptome data of T47D cell model. FOXA1 redistribution was significantly associated with transcriptomic alterations caused by ESR1 mutations. Furthermore, in ESR1-mutant cells, FOXA1-binding sites less frequently overlapped with ER, and differential gene expression was less associated with the canonical FOXA1-ER axis. Motif analysis revealed a unique enrichment of retinoid X receptor (RXR) motifs in FOXA1-binding sites of ESR1-mutant cells. Consistently, ESR1-mutant cells were more sensitive to growth stimulation with the RXR agonist LG268. The mutant-specific response was dependent on two RXR isoforms, RXR-α and RXR-ß, with a stronger dependency on the latter. In addition, T3, the agonist of thyroid receptor (TR) also showed a similar growth-promoting effect in ESR1-mutant cells. Importantly, RXR antagonist HX531 blocked growth of ESR1-mutant cells and a patient-derived xenograft (PDX)-derived organoid with an ESR1 D538G mutation. Collectively, our data support the evidence for a stronger RXR response associated with FOXA1 reprograming in ESR1-mutant cells, suggesting development of therapeutic strategies targeting RXR pathways in breast tumors with ESR1 mutation. IMPLICATIONS: It provides comprehensive characterization of the role of FOXA1 in ESR1-mutant breast cancer and potential therapeutic strategy through blocking RXR activation.

Modulation of preeclampsia by the cholinergic anti-inflammatory pathway: Therapeutic perspectives.

The cholinergic anti-inflammatory pathway (CAP) is vital for the orchestration of the immune and inflammatory responses under normal and challenged conditions. Over the past two decades, peripheral and central circuits of CAP have been shown to be critically involved in dampening the inflammatory reaction in a wide array of inflammatory disorders. Additionally, emerging evidence supports a key role for CAP in the regulation of the female reproductive system during gestation as well as in the advent of serious pregnancy-related inflammatory insults such as preeclampsia (PE). Within this framework, the modulatory action of CAP encompasses the perinatal maternal and fetal adverse consequences that surface due to antenatal PE programming. Albeit, a considerable gap still exists in our knowledge of the precise cellular and molecular underpinnings of PE/CAP interaction, which hampered global efforts in safeguarding effective preventive or therapeutic measures against PE complications. Here, we summarize reports in the literature regarding the roles of peripheral and reflex cholinergic neuroinflammatory pathways of nicotinic acetylcholine receptors (nAChRs) in reprogramming PE complications in mothers and their progenies. The possible contributions of α7-nAChRs, cholinesterases, immune cells, adhesion molecules, angiogenesis, and endothelial dysfunction to the interaction have also been reviewed.

Pre-eclamptic Fetal Programming Alters Neuroinflammatory and Cardiovascular Consequences of Endotoxemia in Sex-Specific Manners.

Pre-eclampsia (PE)-induced fetal programming predisposes offspring to health hazards in adult life. Here, we tested the hypothesis that pre-eclamptic fetal programming elicits sexually dimorphic inflammatory and cardiovascular complications to endotoxemia in adult rat offspring. PE was induced by oral administration of L-NAME (50 mg/kg per day for seven consecutive days) starting from day 14 of conception. Cardiovascular studies were performed in conscious adult male and female offspring preinstrumented with femoral indwelling catheters. Compared with non-PE male counterparts, intravenous administration of lipopolysaccharide (LPS, 5 mg/kg) to PE male offspring caused significantly greater 1) falls in blood pressure, 2) increases in heart rate, 3) rises in arterial dP/dtmax, a correlate of left ventricular contractility, and 4) decreases in time- and frequency-domain indices of heart rate variability (HRV). By contrast, the hypotensive and tachycardic actions of LPS in female offspring were independent of the pre-eclamptic state and no clear changes in HRV or dP/dtmax were noted. Measurement of arterial baroreflex activity by vasoactive method revealed no sex specificity in baroreflex dysfunction induced by LPS. Immunohistochemical studies showed increased protein expression of toll-like receptor 4 in heart as well as in brainstem neuronal pools of the nucleus of solitary tract and rostral ventrolateral medulla in endotoxic PE male, but not female, offspring. Enhanced myocardial, but not neuronal, expression of monocyte chemoattractant protein-1 was also demonstrated in LPS-treated male offspring. Together, pre-eclamptic fetal programming aggravates endotoxic manifestations of hypotension and autonomic dysfunction in male offspring via exacerbating myocardial and neuromedullary inflammatory pathways. SIGNIFICANCE STATEMENT: Current molecular and neuroanatomical evidence highlights a key role for pre-eclamptic fetal programming in offspring predisposition to health hazards induced by endotoxemia in adult life. Pre-eclampsia accentuates endotoxic manifestations of hypotension, tachycardia, and cardiac autonomic dysfunction in male offspring via exacerbating myocardial and central inflammatory pathways. The absence of such detrimental effects in female littermates suggests sexual dimorphism in the interaction of pre-eclamptic fetal programming with endotoxemia.

Time and sex dependency of hemodynamic, renal, and survivability effects of endotoxemia in rats.

Widely different exposure times to endotoxic insults have been employed in reported studies. The current experimental study systematically evaluated the time-course and sex influences of endotoxic insult on survivability and cardiovascular and renal functions. Rats received i.p. lipopolysaccharide (LPS, 5 mg/kg) once or twice (over 2 successive days). Systolic blood pressure (SBP), biomarkers of renal function and inflammation, and vasodilator responsiveness of isolated perfused kidneys to acetylcholine (ACh) or N-ethylcarboxamidoadenosine (NECA) were evaluated 6 hr after first LPS injection or 1, 2, or 6 days later. A single 6-hr LPS challenge caused (i) sex-unrelated elevations in serum urea and creatinine and reductions in NECA, but not ACh, vasodilations, (ii) more increases in renal NF-κB/iNOS expressions in male than in female rats, and (iii) hypotension and tachycardia only in male rats. These parameters, except for hemodynamic changes, were restored to near-control levels 1 day after single LPS dosing. The 2-days dosing with LPS had no effects on renal function biomarkers, but caused hypotension, tachycardia, and increases in renal NF-κB/iNOS expression and NECA and ACh vasodilations in both rat sexes. None of these parameters were different from control values when measured 6 days after the endotoxic insult. Alternatively, the rat mortality was observed during first 2 days of the study and was notably higher in male than in female rats. Our data suggest that the frequency and time elapsed after LPS exposure as well as rat sex are important determinants of the magnitude and direction of detrimental effects of endotoxemia.

The α7-nAChR/heme oxygenase-1/carbon monoxide pathway mediates the nicotine counteraction of renal inflammation and vasoconstrictor hyporeactivity in endotoxic male rats.

OBJECTIVE: The objective of the study was to test the hypothesis that nicotine guards against endotoxemia-associated renal inflammation and vasoconstrictor dysfunction via the activation of α7-nicotinic acetylcholine receptors (α7-nAChRs)/heme oxygenase-1 (HO-1) cascade. MATERIALS: 91 male and female rats were included in the study. TREATMENTS: Lipopolysaccharide (LPS, 5 mg kg-1), nicotine (0.5-2 mg kg-1), pentoxifylline (PTX, TNFα inhibitor, 3 mg kg-1), methyllycaconitine (MLA, α7-nAChR blocker), zinc protoporphyrin (ZnPP, HO-1 inhibitor), hemin (HO-1 inducer), tricarbonyldichlororuthenium (carbon monoxide-releasing molecule, CORM-2) or bilirubin was administered before LPS. METHODS: Isolated perfused kidney was used to evaluate renal vasoconstriction and immunohistochemistry to assess inflammatory cytokines. RESULTS: LPS reduced renal vasoconstrictions induced by phenylephrine or vasopressin in perfused kidneys of male, but not female, rats. Higher elevations in serum interleukin-1ß and renal expressions of inducible nitric oxide synthase (iNOS) and nuclear factor-κB (NF-κB) were observed in LPS-treated male rats, whereas greater HO-1 expression was evident in endotoxic female rats. LPS effects were reversed by nicotine or PTX. Further, the favorable nicotine actions were (i) diminished by MLA or ZnPP and (ii) replicated by hemin or CORM-2, but not bilirubin, and (iii) associated with exaggerated and MLA-sensitive increases in HO-1 expression. CONCLUSIONS: α7-nAChR/HO-1/CO signaling mediates nicotine protection against renal inflammation and vasoconstrictor hyporeactivity in endotoxic male rats.

Nicotine Improves Survivability, Hypotension, and Impaired Adenosinergic Renal Vasodilations in Endotoxic Rats: Role of α7-nAChRs/HO-1 Pathway.

The nicotinic/cholinergic antiinflammatory pathway protects against acute kidney injury and other end-organ damages induced by endotoxemia. In this study, we tested the hypothesis that functional α7-nAChRs/heme oxygenase-1 (HO-1) pathway is imperative for the nicotine counteraction of hemodynamic and renovascular dysfunction caused by acute endotoxemia in rats. Renal vasodilations were induced by cumulative bolus injections of acetylcholine (ACh, 0.01 nmol-7.29 nmol) or ethylcarboxamidoadenosine (NECA, adenosine receptor agonist, 1.6 nmol-100 nmol) in isolated phenylephrine-preconstricted perfused kidneys. The data showed that 6-h treatment with lipopolysaccharide (LPS, 5 mg/kg i.p.) decreased systolic blood pressure and renal vasodilations caused by NECA but not Ach. The endotoxic insult also increased the mortality rate and elevated serum urea and creatinine. These LPS effects were sex-unrelated, except hypotension, and enhanced mortality which were more evident in male rodents, and abrogated after co-administration of nicotine (0.5, 1 mg/kg and 2 mg/kg) in a dose-dependent fashion. The advantageous effects of nicotine on NECA vasodilations, survivability, and kidney biomarkers in endotoxic male rats disappeared upon concurrent exposure to methyllycaconitine citrate (α7-nAChR blocker) or zinc protoporphyrin (HO-1 inhibitor) and were reproduced after treatment with bilirubin, but not hemin (HO-1 inducer) or tricarbonyldichlororuthenium (II) dimer (carbon monoxide-releasing molecule). Together, current biochemical and pharmacological evidence suggests key roles for α7-nAChRs and the bilirubin byproduct of the HO-1 signaling in the nicotine counteraction of renal dysfunction and reduced adenosinergic renal vasodilator capacity in endotoxic rats.

Nicotine reverses the enhanced renal vasodilator capacity in endotoxic rats: Role of α7/α4ß2 nAChRs and HSP70.

BACKGROUND: Nicotine alleviates renal inflammation and injury induced by endotoxemia. This study investigated (i) the nicotine modulation of hemodynamic and renal vasodilatory responses to endotoxemia in rats, and (ii) roles of α7 or α4ß2-nAChRs and related HSP70/TNFα/iNOS signaling in the interaction. METHODS: Endotoxemia was induced by ip lipopolysaccharide (5 mg/kg/day, for 2 days) and changes in systolic blood pressure and vasodilator responsiveness of isolated perfused kidney to acetylcholine or 5'-N-ethylcarboxamidoadenosine (NECA, adenosine receptor agonist) were evaluated. RESULTS: Lipopolysaccharide had no effect on serum creatinine, reduced blood pressure, and increased renal vasodilations induced by acetylcholine or NECA in male and female preparations. Immunohistochemical analyses showed that lipopolysaccharide reduced renal HSP70 expression, but increased α7-nAChRs, α4ß2-nAChRs and iNOS expressions. The co-administration of aminoguanidine (iNOS inhibitor), pentoxifylline (TNFα inhibitor), or nicotine attenuated lipopolysaccharide mediation of renal vasodilations and elevations in α7/α4ß2-nAChR and iNOS expressions. Nicotine also reversed the downregulating effect of lipopolysaccharide on HSP70 expression. α7-nAChRs (methyllycaconitine citrate, MLA) or α4ß2-nAChRs (dihydro-ß-erythroidine, DHßE) blockade potentiated the lipopolysaccharide enhancement of renal vasodilations, and abolished the depressant effect of nicotine on lipopolysaccharide responses. A similar abolition of nicotine effects was seen after HSP70 inhibition by quercetin. Alternatively, lipopolysaccharide hypotension was eliminated in rats treated with DHßE/nicotine or quercetin/nicotine regimen in contrast to no effect for nicotine alone or combined with MLA. CONCLUSIONS: These findings establish that nicotine offsets lipopolysaccharide facilitation of renal vasodilations possibly through a crosstalk between HSP70 and nAChRs of the α7 and α4ß2 types.