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1.
Biophys J ; 121(19): 3651-3662, 2022 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-35778844

RESUMO

Mutations of the intracellular estrogen receptor alpha (ERα) is implicated in 70% of breast cancers. Therefore, it is of considerable interest to image various mutants (L536S, Y537S, D538G) in living cancer cell lines, particularly as a function of various anticancer drugs. We therefore developed a small (13 kDa) Affimer, which, after fluorescent labeling, is able to efficiently label ERα by traveling through temporary pores in the cell membrane, created by the toxin streptolysin O. The Affimer, selected by a phage display, predominantly labels the Y537S mutant and can tell the difference between L536S and D538G mutants. The vast majority of Affimer-ERαY537S is in the nucleus and is capable of an efficient, unrestricted navigation to its target DNA sequence, as visualized by single-molecule fluorescence. The Affimer can also differentiate the effect of selective estrogen receptor modulators. More generally, this is an example of a small binding reagent-an Affimer protein-that can be inserted into living cells with minimal perturbation and high efficiency, to image an endogenous protein.


Assuntos
Antineoplásicos , Neoplasias da Mama , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Receptor alfa de Estrogênio/química , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Células MCF-7 , Mutação , Receptores de Estrogênio/genética , Receptores de Estrogênio/uso terapêutico , Moduladores Seletivos de Receptor Estrogênico/uso terapêutico
2.
BMC Gastroenterol ; 21(1): 62, 2021 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-33573601

RESUMO

BACKGROUND: Cholecystectomy (XGB) is the most common abdominal surgery performed in the United States and is associated with an increased post-surgery incidence of metabolic and gastrointestinal (GI) diseases. Two main risk factors for XGB are sex (female) and age (40-50 yr), corresponding with onset of menopause. Post-menopausal estrogen loss alone facilitates metabolic dysfunction, but the effects of XGB on metabolic and GI health have yet to be investigated in this population. Study objectives were to (1) identify possible short-term effects of XGB and (2) develop a novel murine model of XGB in human menopause via subsequent ovariectomy (OVX) and assess longitudinal effects of OVX on metabolism, GI physiology, and GI microbiota in XGB mice. METHODS: Female C57BL/6 mice were utilized in two parallel studies (S1&S2). In S1, XGB mice were compared to a non-XGB baseline group after six wk. In S2, mice were XGB at wk0, either sham (SHM) or OVX at wk6, and sacrificed at wk12, wk18, and wk24. Body composition assessment and fresh fecal collections were conducted periodically. Serum and tissues were collected at sacrifice for metabolic and GI health endpoints. RESULTS: Compared to baseline, XGB increased hepatic CYP7A1 and decreased HMGCR relative expression, but did not influence BW, fat mass, or hepatic triglycerides after six wk. In S2, XGB/OVX mice had greater BW and fat mass than XGB/SHM. Cecal microbiota alpha diversity metrics were lower in XGB/OVX mice at wk24 compared the XGB/SHM. No consistent longitudinal patterns in fasting serum lipids, fecal microbial diversity, and GI gene expression were observed between S2 groups. CONCLUSIONS: In addition to developing a novel, clinically-representative model of XGB and subsequent OVX, our results suggest that OVX resulted in the expected phenotype to some extent, but that XGB may modify or mask some responses and requires further investigation.


Assuntos
Colecistectomia , Animais , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Fenótipo , Projetos Piloto , Triglicerídeos
3.
Breast Cancer Res Treat ; 181(2): 297-307, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32277377

RESUMO

PURPOSE: Many human breast tumors become resistant to endocrine therapies and recur due to estrogen receptor (ERα) mutations that convey constitutive activity and a more aggressive phenotype. Here, we examined the effectiveness of a novel adamantyl antiestrogen, K-07, in suppressing the growth of breast cancer metastases containing the two most frequent ER-activating mutations, Y537S and D538G, and in extending survival in a preclinical metastatic cancer model. METHODS: MCF7 breast cancer cells expressing luciferase and Y537S or D538G ER were injected into NOD-SCID-gamma female mice, and animals were treated orally with the antiestrogen K-07 or control vehicle. Comparisons were also made with the antiestrogen Fulvestrant. The development of metastases was monitored by in vivo bioluminescence imaging with phenotypic characterization of the metastases in liver and lung by immunohistochemical and biochemical analyses. RESULTS: These breast cancer cells established metastases in liver and lung, and K-07 treatment reduced the metastatic burden. Mice treated with K-07 also survived much longer. By day 70, only 28% of vehicle-treated mice with mutant ER metastases were alive, whereas all K-07-treated D538G and Y537S mice were still alive. K-07 also markedly reduced the level of metastatic cell ER and the expression of ER-regulated genes. CONCLUSION: The antiestrogen K-07 can reduce in vivo metastasis of breast cancers and extend host survival in this preclinical model driven by constitutively active mutant ERs, suggesting that this compound may be suitable for further translational examination of its efficacy in suppression of metastasis in breast cancers containing constitutively active mutant ERs.


Assuntos
Adamantano/análogos & derivados , Adamantano/farmacologia , Neoplasias da Mama/tratamento farmacológico , Moduladores de Receptor Estrogênico/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Mutação , Receptores de Estrogênio/genética , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Cetonas/farmacologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/secundário , Células MCF-7 , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Taxa de Sobrevida , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Gen Comp Endocrinol ; 289: 113382, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-31917150

RESUMO

Seasonally spawning fish rely on a dynamic and complex hormonal interplay to regulate cycles of gonadal development and the regression. Thyroid hormones have been shown to be a key player during gonadal development, and can regulate the activity of a number of essential reproductive hormones. Apoptosis is a vital cellular process that contributes to the hormonal control of gonadal development and regression, but the roles of thyroid hormones on gonadal apoptosis in goldfish have not been explored. The present study examines the role of acute T3 exposure on caspase 3-dependent apoptosis in dispersed goldfish gonadal tissue in vitro. We examined the levels of caspase 3 activity in early, mid, and late recrudescent gonadal tissue after exposure to physiological doses of T3 for up to 24 h. Acute treatment with T3 did not alter basal caspase 3 activity in goldfish gonads in vitro in these reproductive stages. This initial study suggests that transient increases in T3 levels are unlikely to directly contribute to basal caspase 3-dependent apoptosis in the gonadal tissue of goldfish, although we cannot rule out an interaction of T3 with other hormones involved in the control of apoptosis in the testis and ovary.


Assuntos
Caspase 3/efeitos dos fármacos , Carpa Dourada/fisiologia , Gônadas/efeitos dos fármacos , Tri-Iodotironina/efeitos adversos , Animais , Feminino , Masculino
5.
Proc Natl Acad Sci U S A ; 108(45): 18348-53, 2011 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-22042850

RESUMO

Stimulation of resting CD4(+) T lymphocytes leads to rapid proliferation and differentiation into effector (Teff) or inducible regulatory (Treg) subsets with specific functions to promote or suppress immunity. Importantly, Teff and Treg use distinct metabolic programs to support subset specification, survival, and function. Here, we describe that the orphan nuclear receptor estrogen-related receptor-α (ERRα) regulates metabolic pathways critical for Teff. Resting CD4(+) T cells expressed low levels of ERRα protein that increased on activation. ERRα deficiency reduced activated T-cell numbers in vivo and cytokine production in vitro but did not seem to modulate immunity through inhibition of activating signals or viability. Rather, ERRα broadly affected metabolic gene expression and glucose metabolism essential for Teff. In particular, up-regulation of Glut1 protein, glucose uptake, and mitochondrial processes were suppressed in activated ERRα(-/-) T cells and T cells treated with two chemically independent ERRα inhibitors or by shRNAi. Acute ERRα inhibition also blocked T-cell growth and proliferation. This defect appeared as a result of inadequate glucose metabolism, because provision of lipids, but not increased glucose uptake or pyruvate, rescued ATP levels and cell division. Additionally, we have shown that Treg requires lipid oxidation, whereas Teff uses glucose metabolism, and lipid addition selectively restored Treg--but not Teff--generation after acute ERRα inhibition. Furthermore, in vivo inhibition of ERRα reduced T-cell proliferation and Teff generation in both immunization and experimental autoimmune encephalomyelitis models. Thus, ERRα is a selective transcriptional regulator of Teff metabolism that may provide a metabolic means to modulate immunity.


Assuntos
Diferenciação Celular , Ativação Linfocitária , Receptores de Estrogênio/fisiologia , Linfócitos T/imunologia , Animais , Proliferação de Células , Glucose/metabolismo , Homeostase , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Reação em Cadeia da Polimerase , Interferência de RNA , Receptores de Estrogênio/genética , Receptor ERRalfa Relacionado ao Estrogênio
6.
bioRxiv ; 2024 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-38746134

RESUMO

Extracellular vesicles (EVs) serve as crucial mediators of cell-to-cell communication in normal physiology as well as in diseased states, and have been largely studied in regard to their role in cancer progression. However, the mechanisms by which their biogenesis and secretion are regulated by metabolic or endocrine factors remain unknown. Here, we delineate a mechanism by which EV secretion is regulated by a cholesterol metabolite, 27-Hydroxycholesterol (27HC), where treatment of myeloid immune cells (RAW 264.7 and J774A.1) with 27HC impairs lysosomal homeostasis, leading to shunting of multivesicular bodies (MVBs) away from lysosomal degradation, towards secretion as EVs. This impairment of lysosomal function is caused by mitochondrial dysfunction and subsequent increase in reactive oxygen species (ROS). Interestingly, cotreatment with a mitochondria-targeted antioxidant rescued the lysosomal impairment and attenuated the 27HC-mediated increase in EV secretion. Overall, our findings establish how a cholesterol metabolite regulates EV secretion and paves the way for the development of strategies to regulate cancer progression by controlling EV secretion.

7.
Gut Microbes ; 16(1): 2295429, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38153260

RESUMO

Women are at significantly greater risk of metabolic dysfunction after menopause, which subsequently leads to numerous chronic illnesses. The gut microbiome is associated with obesity and metabolic dysfunction, but its interaction with female sex hormone status and the resulting impact on host metabolism remains unclear. Herein, we characterized inflammatory and metabolic phenotypes as well as the gut microbiome associated with ovariectomy and high-fat diet feeding, compared to gonadal intact and low-fat diet controls. We then performed fecal microbiota transplantation (FMT) using gnotobiotic mice to identify the impact of ovariectomy-associated gut microbiome on inflammatory and metabolic outcomes. We demonstrated that ovariectomy led to greater gastrointestinal permeability and inflammation of the gut and metabolic organs, and that a high-fat diet exacerbated these phenotypes. Ovariectomy also led to alteration of the gut microbiome, including greater fecal ß-glucuronidase activity. However, differential changes in the gut microbiome only occurred when fed a low-fat diet, not the high-fat diet. Gnotobiotic mice that received the gut microbiome from ovariectomized mice fed the low-fat diet had greater weight gain and hepatic gene expression related to metabolic dysfunction and inflammation than those that received intact sham control-associated microbiome. These results indicate that the gut microbiome responds to alterations in female sex hormone status and contributes to metabolic dysfunction. Identifying and developing gut microbiome-targeted modulators to regulate sex hormones may be useful therapeutically in remediating menopause-related diseases.


Assuntos
Microbioma Gastrointestinal , Humanos , Feminino , Camundongos , Animais , Microbioma Gastrointestinal/fisiologia , Obesidade/metabolismo , Fígado/metabolismo , Dieta Hiperlipídica/efeitos adversos , Inflamação/metabolismo , Hormônios Esteroides Gonadais/metabolismo , Camundongos Endogâmicos C57BL
8.
bioRxiv ; 2024 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-39131340

RESUMO

Small extracellular vesicles (sEVs) are emerging as critical mediators of intercellular communication in the tumor microenvironment (TME). Here, we investigate the mechanisms by which sEVs derived from neutrophils treated with the cholesterol metabolite, 27-hydroxycholesterol (27HC), influence breast cancer progression. sEVs released from 27HC treated neutrophils enhance epithelial-mesenchymal transition (EMT) and stem-like properties in breast cancer cells, resulting in loss of adherence, increased migratory capacity and resistance to cytotoxic chemotherapy. Decreased microRNAs (miRs) within the sEVs resulted in activation of the WNT/ß-catenin signaling pathway in recipient cells and suggest that this may be a predominant pathway for stem-like phenotype and EMT. Our findings underscore a novel mechanism by which 27HC-modulated neutrophils contribute to breast cancer pathophysiology through EV-mediated intercellular communication, suggesting potential therapeutic targets in cancer treatment.

9.
Mol Cell Biol ; : 1-19, 2024 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-39133105

RESUMO

A significant number of the genetic alterations observed in cancer patients lie within nonprotein-coding segments of the genome, including regions coding for long noncoding RNAs (lncRNAs). LncRNAs display aberrant expression in breast cancer (BrCa), but the functional implications of this altered expression remain to be elucidated. By performing transcriptome screen in a triple negative BrCa (TNBC) isogenic 2D and 3D spheroid model, we observed aberrant expression of >1000 lncRNAs during BrCa progression. The chromatin-associated lncRNA MANCR shows elevated expression in metastatic TNBC. MANCR is upregulated in response to cellular stress and modulates DNA repair and cell proliferation. MANCR promotes metastasis as MANCR-depleted cells show reduced cell migration, invasion, and wound healing in vitro, and reduced metastatic lung colonization in xenograft experiments in vivo. Transcriptome analyses reveal that MANCR modulates expression and pre-mRNA splicing of genes, controlling DNA repair and checkpoint response. MANCR promotes the transcription of NET1A, a Rho-GEF that regulates DNA damage checkpoint and metastatic processes in cis, by differential promoter usage. Experiments suggest that MANCR regulates the expression of cancer-associated genes by modulating the association of various transcription factors and RNA-binding proteins. Our results identified the metastasis-promoting activities of MANCR in TNBC by cis-regulation of gene expression.

10.
Cancer Lett ; 597: 217042, 2024 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-38908543

RESUMO

Although survival from breast cancer has dramatically increased, many will develop recurrent, metastatic disease. Unfortunately, survival for this stage of disease remains very low. Activating the immune system has incredible promise since it has the potential to be curative. However, immune checkpoint blockade (ICB) which works through T cells has been largely disappointing for metastatic breast cancer. One reason for this is a suppressive myeloid immune compartment that is unaffected by ICB. Cholesterol metabolism and proteins involved in cholesterol homeostasis play important regulatory roles in myeloid cells. Here, we demonstrate that NR0B2, a nuclear receptor involved in negative feedback of cholesterol metabolism, works in several myeloid cell types to impair subsequent expansion of regulatory T cells (Tregs); Tregs being a subset known to be highly immune suppressive and associated with poor therapeutic response. Within myeloid cells, NR0B2 serves to decrease many aspects of the inflammasome, ultimately resulting in decreased IL1ß; IL1ß driving Treg expansion. Importantly, mice lacking NR0B2 exhibit accelerated tumor growth. Thus, NR0B2 represents an important node in myeloid cells dictating ensuing Treg expansion and tumor growth, thereby representing a novel therapeutic target to re-educate these cells, having impact across different solid tumor types. Indeed, a paper co-published in this issue demonstrates the therapeutic utility of targeting NR0B2.


Assuntos
Neoplasias da Mama , Progressão da Doença , Células Mieloides , Linfócitos T Reguladores , Linfócitos T Reguladores/imunologia , Animais , Feminino , Camundongos , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Humanos , Células Mieloides/imunologia , Células Mieloides/metabolismo , Camundongos Knockout , Interleucina-1beta/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Inflamassomos/metabolismo , Inflamassomos/imunologia
11.
Cancer Lett ; 597: 217086, 2024 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-38944231

RESUMO

Immune checkpoint blockade (ICB) has had limited utility in several solid tumors such as breast cancer, a major cause of cancer-related mortality in women. Therefore, there is considerable interest in alternate strategies to promote an anti-cancer immune response. A paper co-published in this issue describes how NR0B2, a protein involved in cholesterol homeostasis, functions within myeloid immune cells to modulate the inflammasome and reduce the expansion of immune-suppressive regulatory T cells (Treg). Here, we develop NR0B2 as a potential therapeutic target. NR0B2 in tumors is associated with improved survival for several cancer types including breast. Importantly, NR0B2 expression is also prognostic of ICB success. Within breast tumors, NR0B2 expression is inversely associated with FOXP3, a marker of Tregs. While a described agonist (DSHN) had some efficacy, it required high doses and long treatment times. Therefore, we designed and screened several derivatives. A methyl ester derivative (DSHN-OMe) emerged as superior in terms of (1) cellular uptake, (2) ability to regulate expected expression of genes, (3) suppression of Treg expansion using in vitro co-culture systems, and (4) efficacy against the growth of primary and metastatic tumors. This work identifies NR0B2 as a target to re-educate myeloid immune cells and a novel ligand with significant anti-tumor efficacy in preclinical models.


Assuntos
Células Mieloides , Linfócitos T Reguladores , Humanos , Células Mieloides/imunologia , Células Mieloides/metabolismo , Células Mieloides/efeitos dos fármacos , Feminino , Animais , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Neoplasias da Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/imunologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Camundongos , Linhagem Celular Tumoral , Microambiente Tumoral , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico
12.
Gen Comp Endocrinol ; 192: 15-24, 2013 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-23583769

RESUMO

Estrogens, steroid hormones critically involved in reproductive processes of vertebrates, signal primarily through their intracellular estrogen receptors (ERs). The ERs belong to a superfamily of nuclear receptors that act as ligand inducible transcription factors. Herein, we review what is known about ER structure, subtypes, mechanism(s) of action and auto-regulation by estrogens. Focus is placed on the ER in fish but comparisons are made to mammals and other vertebrates. Finally, we provide context and a proposed model integrating our knowledge on autoregulation of the receptor and its functions in the liver. Future areas of study are suggested, along with cautions when designing experiments, especially for the detection of endocrine disruptors.


Assuntos
Peixes/metabolismo , Receptores de Estrogênio/metabolismo , Vertebrados/metabolismo , Animais , Receptores de Estrogênio/classificação
13.
Cancer Res ; 83(18): 3115-3130, 2023 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-37522838

RESUMO

Several emerging therapies kill cancer cells primarily by inducing necrosis. As necrosis activates immune cells, potentially, uncovering the molecular drivers of anticancer therapy-induced necrosis could reveal approaches for enhancing immunotherapy efficacy. To identify necrosis-associated genes, we performed a genome-wide CRISPR-Cas9 screen with negative selection against necrosis-inducing preclinical agents BHPI and conducted follow-on experiments with ErSO. The screen identified transient receptor potential melastatin member 4 (TRPM4), a calcium-activated, ATP-inhibited, sodium-selective plasma membrane channel. Cancer cells selected for resistance to BHPI and ErSO exhibited robust TRPM4 downregulation, and TRPM4 reexpression restored sensitivity to ErSO. Notably, TRPM4 knockout (TKO) abolished ErSO-induced regression of breast tumors in mice. Supporting a broad role for TRPM4 in necrosis, knockout of TRPM4 reversed cell death induced by four additional diverse necrosis-inducing cancer therapies. ErSO induced anticipatory unfolded protein response (a-UPR) hyperactivation, long-term necrotic cell death, and release of damage-associated molecular patterns that activated macrophages and increased monocyte migration, all of which was abolished by TKO. Furthermore, loss of TRPM4 suppressed the ErSO-induced increase in cell volume and depletion of ATP. These data suggest that ErSO triggers initial activation of the a-UPR but that it is TRPM4-mediated sodium influx and cell swelling, resulting in osmotic stress, which sustains and propagates lethal a-UPR hyperactivation. Thus, TRPM4 plays a pivotal role in sustaining lethal a-UPR hyperactivation that mediates the anticancer activity of diverse necrosis-inducing therapies. SIGNIFICANCE: A genome-wide CRISPR screen reveals a pivotal role for TRPM4 in cell death and immune activation following treatment with diverse necrosis-inducing anticancer therapies, which could facilitate development of necrosis-based cancer immunotherapies.


Assuntos
Trifosfato de Adenosina , Canais de Cátion TRPM , Camundongos , Animais , Necrose/metabolismo , Morte Celular , Membrana Celular/metabolismo , Trifosfato de Adenosina/metabolismo , Sódio/metabolismo , Canais de Cátion TRPM/genética , Canais de Cátion TRPM/metabolismo
14.
Transl Oncol ; 33: 101676, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37086619

RESUMO

G0/G1 switch gene 2 (G0S2) is known to inhibit lipolysis by inhibiting adipose triglyceride lipase (ATGL). In this report, we dissect the role of G0S2 in ER+ versus ER- breast cancer. Overexpression of G0S2 in ER- cells increased cell proliferation, while G0S2 overexpression in ER+ cells decreased cell proliferation. Transcriptome analysis revealed that G0S2 mediated distinct but overlapping transcriptional responses in ER- and ER+ cells. G0S2 reduced genes associated with an epithelial phenotype, especially in ER- cells, including CDH1, ELF3, STEAP4 and TACSTD2, suggesting promotion of the epithelial-mesenchymal transition (EMT). G0S2 also repressed estrogen signaling and estrogen receptor target gene signatures, especially in ER+ cells, including TFF1 and TFF3. In addition, G0S2 overexpression increased cell migration in ER- cells and increased estrogen deprivation sensitivity in ER+ cells. Interestingly, two genes downstream of ATGL in fat utilization and very important in steroid hormone biosynthesis, HMGCS1 and HMGCS2, were downregulated in G0S2 overexpressing ER+ cells. In addition, HSD17B11, a gene that converts estradiol to its less estrogenic derivative, estrone, was highly upregulated in G0S2 overexpressing ER+ cells, suggesting G0S2 overexpression has a negative effect on estradiol production and maintenance. High expression of G0S2 and HSD17B11 was associated with improved relapse-free survival in breast cancer patients while high expression of HMGSC1 was associated with poor survival. Finally, we deleted G0S2 in breast cancer-prone MMTV-PyMT mice. Our data indicates a complex role for G0S2 in breast cancer, dependent on ER status, that may be partially mediated by suppression of the estrogen signaling pathway.

15.
Nat Commun ; 14(1): 4101, 2023 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-37491347

RESUMO

Hypercholesterolemia and vascular inflammation are key interconnected contributors to the pathogenesis of atherosclerosis. How hypercholesterolemia initiates vascular inflammation is poorly understood. Here we show in male mice that hypercholesterolemia-driven endothelial activation, monocyte recruitment and atherosclerotic lesion formation are promoted by a crosstalk between macrophages and endothelial cells mediated by the cholesterol metabolite 27-hydroxycholesterol (27HC). The pro-atherogenic actions of macrophage-derived 27HC require endothelial estrogen receptor alpha (ERα) and disassociation of the cytoplasmic scaffolding protein septin 11 from ERα, leading to extranuclear ERα- and septin 11-dependent activation of NF-κB. Furthermore, pharmacologic inhibition of cyp27a1, which generates 27HC, affords atheroprotection by reducing endothelial activation and monocyte recruitment. These findings demonstrate cell-to-cell communication by 27HC, and identify a major causal linkage between the hypercholesterolemia and vascular inflammation that partner to promote atherosclerosis. Interventions interrupting this linkage may provide the means to blunt vascular inflammation without impairing host defense to combat the risk of atherosclerotic cardiovascular disease that remains despite lipid-lowering therapies.


Assuntos
Aterosclerose , Hipercolesterolemia , Masculino , Camundongos , Animais , Receptor alfa de Estrogênio/metabolismo , Hipercolesterolemia/complicações , Hipercolesterolemia/metabolismo , Células Endoteliais/metabolismo , Septinas/metabolismo , Colesterol/metabolismo , Aterosclerose/metabolismo , Macrófagos/metabolismo , Transdução de Sinais , Inflamação/patologia
16.
bioRxiv ; 2023 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-37645737

RESUMO

Immune checkpoint blockade (ICB) has revolutionized cancer therapy but has had limited utility in several solid tumors such as breast cancer, a major cause of cancer-related mortality in women. Therefore, there is considerable interest in alternate strategies to promote an anti-cancer immune response. We demonstrate that NR0B2, a protein involved in cholesterol homeostasis, functions within myeloid immune cells to modulate the NLRP3 inflammasome and reduce the expansion of immune-suppressive regulatory T cells (Treg). Loss of NR0B2 increased mammary tumor growth and metastasis. Small molecule agonists, including one developed here, reduced Treg expansion, reduced metastatic growth and improved the efficacy of ICB. This work identifies NR0B2 as a target to re-educate myeloid immune cells providing proof-of-principle that this cholesterol-homeostasis axis may have utility in enhancing ICB.

17.
Biochem Biophys Res Commun ; 425(2): 407-12, 2012 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-22846567

RESUMO

G protein-coupled receptor kinase interacting protein 2 (GIT2) is a signaling scaffold protein involved in the regulation of cytoskeletal structure, membrane trafficking, and G protein-coupled receptor internalization. Since dynamic cytoskeletal reorganization plays key roles both in osteoblast differentiation and in the maintenance of osteoclast polarity during bone resorption, we hypothesized that skeletal physiology would be altered in GIT2(-/-) mice. We found that adult GIT2(-/-) mice have decreased bone mineral density and bone volume in both the trabecular and cortical compartments. This osteopenia was associated with decreased numbers of mature osteoblasts, diminished osteoblastic activity, and increased marrow adiposity, suggesting a defect in osteoblast maturation. In vitro, mesenchymal stem cells derived from GIT2(-/-) mice exhibited impaired differentiation into osteoblasts and increased adipocyte differentiation, consistent with a role for GIT2 in mesenchymal stem cell fate determination. Despite elevated osteoclast inducing cytokines and osteoclast numbers, GIT2(-/-) mice also exhibit impaired bone resorption, consistent with a further role for GIT2 in regulating osteoclast function. Collectively, these findings underscore the importance of the cytoskeleton in both osteoblast and osteoclast function and demonstrate that GIT2 plays essential roles in skeletal metabolism, affecting both bone formation and bone resorption in vivo.


Assuntos
Reabsorção Óssea/genética , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular , Células-Tronco Mesenquimais/citologia , Osteoblastos/citologia , Osteogênese/genética , Fosfoproteínas/metabolismo , Animais , Densidade Óssea/genética , Contagem de Células , Proteínas de Ciclo Celular/genética , Citoesqueleto/metabolismo , Feminino , Proteínas Ativadoras de GTPase , Peptídeos e Proteínas de Sinalização Intercelular , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosfoproteínas/genética
18.
Biochem Pharmacol ; 196: 114621, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34043965

RESUMO

Cholesterol has been implicated in the pathophysiology and progression of several cancers now, although the mechanisms by which it influences cancer biology are just emerging. Two likely contributing mechanisms are the ability for cholesterol to directly regulate signaling molecules within the membrane, and certain metabolites acting as signaling molecules. One such metabolite is the oxysterol 27-hydroxycholesterol (27HC), which is a primary metabolite of cholesterol synthesized by the enzyme Cytochrome P450 27A1 (CYP27A1). Physiologically, 27HC is involved in the regulation of cholesterol homeostasis and contributes to cholesterol efflux through liver X receptor (LXR) and inhibition of de novo cholesterol synthesis through the insulin-induced proteins (INSIGs). 27HC is also a selective modulator of the estrogen receptors. An increasing number of studies have identified its importance in cancer progression of various origins, especially in breast cancer. In this review, we discuss the physiological roles of 27HC targeting these two nuclear receptors and the subsequent contribution to cancer progression. We describe how 27HC promotes tumor growth directly through cancer-intrinsic factors, and indirectly through its immunomodulatory roles which lead to decreased immune surveillance and increased tumor invasion. This review underscores the importance of the cholesterol metabolic pathway in cancer progression and the potential therapeutic utility of targeting this metabolic pathway.


Assuntos
Biomarcadores Tumorais/metabolismo , Hidroxicolesteróis/metabolismo , Neoplasias/metabolismo , Animais , Anticolesterolemiantes/uso terapêutico , Colestanotriol 26-Mono-Oxigenase/metabolismo , Colesterol na Dieta/efeitos adversos , Colesterol na Dieta/metabolismo , Humanos , Neoplasias/induzido quimicamente , Neoplasias/tratamento farmacológico
19.
J Exp Nanosci ; 17(1): 599-616, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36968097

RESUMO

The incidence of inflammatory bowel disease (IBD) is increasing worldwide. Although current diagnostic and disease monitoring tests for IBD sensitively detect gut inflammation, they lack the molecular and cellular specificity of positron emission tomography (PET). In this proof-of-concept study, we use a radiolabeled macrophage-targeted nanocarrier probe (64Cu-NOTA-D500) administered by oral, enema, and intraperitoneal routes to evaluate the delivery route dependence of biodistribution across healthy and diseased tissues in a murine model of dextran sodium sulfate (DSS)-induced colitis. High inter-subject variability of probe uptake in intestinal tissue was reduced by normalization to uptake in liver or total intestines. Differences in normalized uptake between healthy and DSS colitis animal intestines were highest for oral and IP routes. Differences in absolute liver uptake reflected a possible secondary diagnostic metric of IBD pathology. These results should inform the preclinical development of inflammation-targeted contrast agents for IBD and related gut disorders to improve diagnostic accuracy.

20.
Sci Adv ; 8(28): eabn5295, 2022 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-35857506

RESUMO

27-Hydroxycholesterol (27-HC) is the most abundant oxysterol that increases the risk of breast cancer progression. However, little is known about epigenetic regulation of 27-HC metabolism and its role in breast tumor initiation. Using genetic mouse mammary tumor and human breast cancer models, we showed here that the histone reader ZMYND8 was selectively expressed in breast cancer stem cells (BCSCs) and promoted epithelial-mesenchymal transition (EMT), BCSC maintenance and self-renewal, and oncogenic transformation through its epigenetic functions, leading to breast tumor initiation. Mechanistically, ZMYND8 was a master transcriptional regulator of 27-HC metabolism. It increased cholesterol biosynthesis and oxidation but blocked cholesterol efflux and 27-HC catabolism, leading to accumulation of 27-HC in BCSCs. Consequently, 27-HC promoted EMT, oncogenic transformation, and tumor initiation through activation of liver X receptor. These findings reveal that ZMYND8 is an epigenetic booster that drives breast tumor initiation through metabolic reprogramming.


Assuntos
Neoplasias da Mama , Animais , Neoplasias da Mama/patologia , Carcinogênese/patologia , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Colesterol/metabolismo , Epigênese Genética , Feminino , Humanos , Hidroxicolesteróis , Camundongos , Células-Tronco Neoplásicas/metabolismo
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