RESUMO
BACKGROUND: Black women experience a disproportionate impact of uterine fibroids compared to White women, including earlier diagnosis, higher frequency, and more severe symptoms. The etiology underlying this racial disparity remains elusive. OBJECTIVE: The aim of this study was to evaluate the molecular differences in normal myometrium (fibroid-free uteri) and at-risk myometrium (fibroid-containing uteri) tissues in Black and White women. STUDY DESIGN: We conducted whole-genome RNA-seq on normal and at-risk myometrium tissues obtained from both self-identified Black and White women (not Hispanic or Latino) to determine global gene expression profiles and to conduct enriched pathway analyses (n=3 per group). We initially assessed the differences within the same type of tissue (normal or at-risk myometrium) between races. Subsequently, we analyzed the transcriptome of normal myometrium compared to at-risk myometrium in each race and determined the differences between them. We validated our findings through real-time PCR (sample size range=5-12), western blot (sample size range=5-6), and immunohistochemistry techniques (sample size range=9-16). RESULTS: The transcriptomic analysis revealed distinct profiles between Black and White women in normal and at-risk myometrium tissues. Interestingly, genes and pathways related to extracellular matrix and mechanosensing were more enriched in normal myometrium from Black than White women. Transcription factor enrichment analysis detected greater activity of the serum response transcription factor positional motif in normal myometrium from Black compared to White women. Furthermore, we observed increased expression levels of myocardin-related transcription factor-serum response factor and the serum response factor in the same comparison. In addition, we noted increased expression of both mRNA and protein levels of vinculin, a target gene of the serum response factor, in normal myometrium tissues from Black women as compared to White women. Importantly, the transcriptomic profile of normal to at-risk myometrium conversion differs between Black and White women. Specifically, we observed that extracellular matrix-related pathways are involved in the transition from normal to at-risk myometrium and that these processes are exacerbated in Black women. We found increased levels of Tenascin C, type I collagen alpha 1 chain, fibronectin, and phospho-p38 MAPK (Thr180/Tyr182, active) protein levels in at-risk over normal myometrium tissues from Black women, whereas such differences were not observed in samples from White women. CONCLUSION: These findings indicate that the racial disparities in uterine fibroids may be attributed to heightened production of extracellular matrix in the myometrium in Black women, even before the tumors appear. Future research is needed to understand early life determinants of the observed racial differences.
Assuntos
Negro ou Afro-Americano , Matriz Extracelular , Fibronectinas , Leiomioma , Miométrio , Fator de Resposta Sérica , Transativadores , População Branca , Feminino , Humanos , Miométrio/metabolismo , População Branca/genética , Negro ou Afro-Americano/genética , Matriz Extracelular/metabolismo , Leiomioma/genética , Leiomioma/metabolismo , Leiomioma/etnologia , Adulto , Transativadores/metabolismo , Transativadores/genética , Fibronectinas/metabolismo , Fibronectinas/genética , Fator de Resposta Sérica/metabolismo , Fator de Resposta Sérica/genética , Neoplasias Uterinas/genética , Neoplasias Uterinas/etnologia , Neoplasias Uterinas/metabolismo , Pessoa de Meia-Idade , Transcriptoma , Tenascina/genética , Tenascina/metabolismo , Colágeno Tipo I/metabolismo , Colágeno Tipo I/genética , RNA-Seq , Proteínas Nucleares , VersicanasRESUMO
Environmental exposure to endocrine-disrupting chemicals (EDCs) is linked to the development of uterine fibroids (UFs) in women. UFs, non-cancerous tumors, are thought to originate from abnormal myometrial stem cells (MMSCs). Defective DNA repair capacity may contribute to the emergence of mutations that promote tumor growth. The multifunctional cytokine TGFß1 is associated with UF progression and DNA damage repair pathways. To investigate the impact of EDC exposure on TGFß1 and nucleotide excision repair (NER) pathways, we isolated MMSCs from 5-month-old Eker rats exposed neonatally to diethylstilbestrol (DES), an EDC, or to vehicle (VEH). EDC-MMSCs exhibited overactivated TGFß1 signaling and reduced mRNA and protein levels of NER pathway components compared to VEH-MMSCs. EDC-MMSCs also demonstrated impaired NER capacity. Exposing VEH-MMSCs to TGFß1 decreased NER capacity while inhibiting TGFß signaling in EDC-MMSCs restored it. RNA-seq analysis and further validation revealed decreased expression of Uvrag, a tumor suppressor gene involved in DNA damage recognition, in VEH-MMSCs treated with TGFß1, but increased expression in EDC-MMSCs after TGFß signaling inhibition. Overall, we demonstrated that the overactivation of the TGFß pathway links early life exposure to EDCs with impaired NER capacity, which would lead to increased genetic instability, arise of mutations, and fibroid tumorigenesis. We demonstrated that the overactivation of the TGFß pathway links early life exposure to EDCs with impaired NER capacity, which would lead to increased fibroid incidence.
Assuntos
Disruptores Endócrinos , Leiomioma , Feminino , Animais , Ratos , Reparo do DNA/genética , Dano ao DNA , Fator de Crescimento Transformador beta/genética , Carcinogênese , Disruptores Endócrinos/toxicidade , Leiomioma/induzido quimicamente , Leiomioma/genéticaRESUMO
Bromodomain-containing proteins (BRDs) are involved in many biological processes, most notably epigenetic regulation of transcription, and BRD dysfunction has been linked to many diseases, including tumorigenesis. However, the role of BRDs in the pathogenesis of uterine fibroids (UFs) is entirely unknown. The present study aimed to determine the expression pattern of BRD9 in UFs and matched myometrium and further assess the impact of a BRD9 inhibitor on UF phenotype and epigenetic/epitranscriptomic changes. Our studies demonstrated that the levels of BRD9 were significantly upregulated in UFs compared to matched myometrium, suggesting that the aberrant BRD expression may contribute to the pathogenesis of UFs. We then evaluated the potential roles of BRD9 using its specific inhibitor, I-BRD9. Targeted inhibition of BRD9 suppressed UF tumorigenesis with increased apoptosis and cell cycle arrest, decreased cell proliferation, and extracellular matrix deposition in UF cells. The latter is the key hallmark of UFs. Unbiased transcriptomic profiling coupled with downstream bioinformatics analysis further and extensively demonstrated that targeted inhibition of BRD9 impacted the cell cycle- and ECM-related biological pathways and reprogrammed the UF cell epigenome and epitranscriptome in UFs. Taken together, our studies support the critical role of BRD9 in UF cells and the strong interconnection between BRD9 and other pathways controlling the UF progression. Targeted inhibition of BRDs might provide a non-hormonal treatment option for this most common benign tumor in women of reproductive age.
Assuntos
Epigenoma , Leiomioma , Humanos , Feminino , Epigênese Genética , Proteínas que Contêm Bromodomínio , Leiomioma/genética , Carcinogênese/genética , Transformação Celular Neoplásica , Fatores de Transcrição , Transdução de SinaisRESUMO
STUDY QUESTION: Is resveratrol able to prevent the lipopolysaccharide (LPS)-induced preterm labor in 15-day pregnant BALB/c mice? SUMMARY ANSWER: Resveratrol prevented the LPS-induced onset of preterm labor in 64% of the cases and showed anti-inflammatory and tocolytic effects by downregulating COX-2 and iNOS expression and NOS activity, and by changing the uterine prostaglandin and endocannabinoid profiling. WHAT IS KNOWN ALREADY: Genital tract infections by Gram-negative bacteria are a common complication in human pregnancy and have been shown to increase risk of preterm delivery. Bacterial LPS elicits a strong maternal inflammatory response that results in preterm delivery and fetal death in a murine model endotoxin-induced preterm labor. STUDY DESIGN, SIZE, DURATION: An in vivo animal study was conducted. On Day 15 of pregnancy, mice received at 8:00 h a dose of vehicle (40% ethanol in saline solution) or resveratrol (3 mg/kg in vehicle) via oral gavage followed by two doses of LPS or vehicle administered intraperitoneally (i.p.), the first one at 10:00 h (0.17 mg/kg in 0.1 ml of sterile saline solution) and the second at 13:00 h (0.5 mg/kg in 0.1 ml of sterile saline solution). The mice were closely observed for any signs of morbidity (piloerection, decreased movement, and diarrhea), vaginal bleeding or preterm delivery. The beginning of preterm delivery was defined by early delivery of the first pup. Normal term labor occurs on Day 19 of gestation. PARTICIPANTS/MATERIALS, SETTING, METHODS: Time of labor, pregnancy outcome and morphological features were evaluated after LPS and/or resveratrol administration. Uterine stripes were collected 5 h after the last LPS injection and prostaglandin and endocannabinoid profiling was analyzed by mass spectrometry. Nitric oxide synthase (NOS) activity was measured by radioconversion assay. Cyclooxygenase-2 (Cox-2) and 15-hydroxyprostaglandin dehydrogenase (15-Pgdh) mRNA levels were analyzed by RT-PCR whilst the protein expression of inducible nitric oxide synthase (iNOS), COX-1 and COX-2 were studied by western blot. MAIN RESULTS AND THE ROLE OF CHANCE: In vivo treatment of 15-day pregnant BALB/c mice with resveratrol prevented the LPS-induced preterm birth in 64% of the cases, whereas only 15% of mice with LPS alone escaped preterm birth. Treatment with resveratrol resulted in a reduced NOS activity (P < 0.05) in the uterus of LPS-treated mice. Similarly, resveratrol reduced the expression of LPS-induced pro-inflammatory agents such as iNOS (P < 0.05), COX-2 (P < 0.05), prostaglandin E2 (PGE2) (P < 0.05) and anandamide (AEA) (P < 0.05). Moreover, resveratrol administration resulted in changes in the uterine endocannabinoid profiling altered by LPS. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Since our experimental design involves the use of mice, the extrapolation of the results presented here to humans is limited. WIDER IMPLICATIONS OF THE FINDINGS: Our findings provide evidence for the tocolytic effects of resveratrol. STUDY FUNDING AND COMPETING INTEREST(S): Dr Ana María Franchi was funded by Agencia Nacional para la Promoción Científica y Tecnológica (PICT 2013/0097) and by Consejo Nacional de Investigaciones Científicas y Técnicas (PIP 2012/0061). Dr Heather B. Bradshaw was funded by NIH (DA006668). The authors have no competing interests.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Inflamação/prevenção & controle , Trabalho de Parto Prematuro/prevenção & controle , Estilbenos/farmacologia , Útero/efeitos dos fármacos , Animais , Endocanabinoides/biossíntese , Feminino , Inflamação/induzido quimicamente , Lipopolissacarídeos , Camundongos Endogâmicos BALB C , Gravidez , Prostaglandinas/biossíntese , Substâncias Protetoras/farmacologia , Resveratrol , Útero/metabolismo , Útero/patologiaRESUMO
Preterm birth is a major contributor to early and delayed physical and cognitive impairment. Epidemiological and experimental data indicate that maternal infections are a significant and preventable cause of preterm birth. Recently, melatonin has been suggested to exert neuroprotective effects in several models of brain injury. Here, we sought to investigate whether the administration of melatonin is able to prevent lipopolysaccharide (LPS)-induced fetal brain damage in a model of LPS-induced preterm labor. For this purpose, 15-day pregnant BALB/c mice received intraperitoneally 2 doses of LPS or vehicle: the first one at 10:00 hours (0.26 mg/kg) and the second at 13:00 hours (0.52 mg/kg). On day 14 of pregnancy, a group of mice was subcutaneously implanted with a pellet of 25 mg melatonin. This experimental protocol resulted in 100% of preterm birth and pup death in the LPS group and a 50% of term birth and pup survival in the melatonin + LPS group. In the absence of melatonin, fetuses from LPS-treated mothers showed histological signs of brain damage, microglial/macrophage activation, and higher levels of IL-1ß, inducible nitric oxide synthase (NOS), and neuronal NOS mRNAs as well as increased histone acetyltransferase activity and histone H3 hyperacetylation. In contrast, antenatal administration of melatonin prevented LPS-induced fetal brain damage. Moreover, when behavioral traits were analyzed in the offspring from control, melatonin, and melatonin + LPS, no significant differences were found, suggesting that melatonin prevented LPS-induced long-term neurodevelopmental impairments. Collectively, our results suggest that melatonin could be a new therapeutic tool to prevent fetal brain damage and its long-term consequences induced by maternal inflammation.
Assuntos
Traumatismos do Nascimento/prevenção & controle , Lesões Encefálicas/prevenção & controle , Melatonina/farmacologia , Fármacos Neuroprotetores/farmacologia , Nascimento Prematuro , Animais , Traumatismos do Nascimento/etiologia , Lesões Encefálicas/etiologia , Feminino , Inflamação/induzido quimicamente , Lipopolissacarídeos/toxicidade , Camundongos , Camundongos Endogâmicos BALB C , Trabalho de Parto Prematuro/induzido quimicamente , Gravidez , Nascimento Prematuro/induzido quimicamenteRESUMO
STUDY QUESTION: What is the role of the endocannabinoid system (eCS) in the alterations of the endocrine system in a murine model of lipopolysaccharide (LPS)-induced miscarriage? SUMMARY ANSWER: In 7-days pregnant wild type, but not cannabinoid receptor type 1 knockout (CB1-KO) mice, LPS increased COX-2 expression and prostaglandin F2α (PGF2α) production in the uterus leading to lower expression of prolactin receptor in the ovary and a marked regression of corpora lutea (CL), suggesting that the eCS mediates the deleterious effects of LPS on reproductive events. WHAT IS KNOWN ALREADY: Appropriate systemic progesterone levels are critical for a successful pregnancy outcome. Precocious loss of luteal progesterone (P4) secretion leads to miscarriage in rodents. We have previously shown that LPS administration to pregnant mice induces embryonic resorption accompanied by a dramatic decrease in systemic progesterone levels in a murine model of inflammatory miscarriage, with the eCS mediating these LPS-induced deleterious effects. STUDY DESIGN SAMPLES/MATERIALS, METHODS: CD1 wild-type (WT) and CB1-KO mice were randomly allocated to Vehicle (saline; i.p.) or LPS (0.5 µg/g body weight; i.p.) treated groups: (WT-Vehicle; WT-LPS; CB1-KO-Vehicle and CB1-KO-LPS). A single injection was given on day 7 of pregnancy and tissues (blood, ovary, uterus) were collected 6, 12, 24 and 48 h later. P4 and PGF2α plasma levels were determined by radioimmunoassay. Cyclooxygenase-2 (COX-2) mRNA (RT-PCR) and protein (Western blot) content in uterus was assayed. COX-2 and prolactin receptor (PrlR) mRNA levels in the ovary were assayed by RT-PCR. Tissue morphology of the CL was assessed by haematoxylin-eosin staining. MAIN RESULTS AND THE ROLE OF CHANCE: Treatment of 7-day pregnant WT mice with LPS induced a P4 withdrawal (p < 0.05), increased in uterine COX-2 mRNA and protein expression (p < 0.05) as well as an increase in uterine PGF2α production (p < 0.05). These changes were absent in LPS-treated 7-day pregnant CB1-KO mice. In ovarian tissues, LPS treatment to 7-day pregnant WT mice induced a downregulation of PrlR mRNA expression (p < 0.05) together with an increase in COX-2 mRNA expression (p < 0.05) and PGF2α content (p < 0.05). These effects were absent in the CB1-KO mice. Collectively, our results suggest a role for the eCS mediating LPS-induced deleterious effects on reproductive tissues. LIMITATIONS, REASONS FOR CAUTION: An important caveat of this study is the endocrine differences between mice and humans during pregnancy (e.g. P4 is produced by the CL throughout pregnancy in mice, whereas this is not the case in humans), which limits the extrapolation of the results presented here. WIDER IMPLICATIONS OF THE FINDINGS: Our findings provide new insights in the role of the endocannabinoid system in the physiopathology of reproduction as well as the role of this endogenous system as a mediator of LPS deleterious effects on reproductive tissues. LARGE SCALE DATA: None. STUDY FUNDING AND COMPETING INTERESTS: Dr Ana María Franchi was funded by Agencia Nacional para la Promoción Científica y Tecnológica (PICT 2010/0813 and PICT 2013/0097) and by Consejo Nacional de Investigaciones Científicas y Técnicas (PIP 2012/0061). The authors have no competing interests.
Assuntos
Aborto Espontâneo/tratamento farmacológico , Aborto Espontâneo/metabolismo , Endocanabinoides/metabolismo , Lipopolissacarídeos/toxicidade , Fase Luteal/metabolismo , Progesterona/metabolismo , Animais , Corpo Lúteo/metabolismo , Feminino , Luteólise/metabolismo , Camundongos , Camundongos Knockout , Gravidez , RadioimunoensaioRESUMO
Prematurity is the leading cause of perinatal morbidity and mortality worldwide. There is a strong causal relationship between infection and preterm births. Intrauterine infection elicits an immune response involving the release of inflammatory mediators like cytokines and prostaglandins (PG) that trigger uterine contractions and parturition events. Anandamide (AEA) is an endogenous ligand for the cannabinoid receptors CB1 and CB2. Similarly to PG, endocannabinoids are implicated in different aspects of reproduction, such as maintenance of pregnancy and parturition. Little is known about the involvement of endocannabinoids on the onset of labor in an infectious milieu. Here, using a mouse model of preterm labor induced by lipopolysaccharide (LPS), we explored changes on the expression of components of endocannabinoid system (ECS). We have also determined whether AEA and CB antagonists alter PG production that induces labor. We observed an increase in uterine N-acylphosphatidylethanolamine-specific phospholipase D expression (NAPE-PLD, the enzyme that synthesizes AEA) upon LPS treatment. Activity of catabolic enzyme fatty acid amide hydrolase (FAAH) did not change significantly. In addition, we also found that LPS modulated uterine cannabinoid receptors expression by downregulating Cb2 mRNA levels and upregulating CB1 protein expression. Furthermore, LPS and AEA induced PGF2a augmentation, and this was reversed by antagonizing CB1 receptor. Collectively, our results suggest that ECS may be involved in the mechanism by which infection causes preterm birth.
Assuntos
Endocanabinoides/metabolismo , Lipopolissacarídeos , Trabalho de Parto Prematuro/metabolismo , Útero/metabolismo , Amidoidrolases/genética , Amidoidrolases/metabolismo , Animais , Ácidos Araquidônicos/metabolismo , Antagonistas de Receptores de Canabinoides/farmacologia , Dinoprosta/metabolismo , Modelos Animais de Doenças , Endocanabinoides/genética , Feminino , Regulação da Expressão Gênica , Idade Gestacional , Camundongos Endogâmicos BALB C , Trabalho de Parto Prematuro/induzido quimicamente , Trabalho de Parto Prematuro/genética , Trabalho de Parto Prematuro/fisiopatologia , Fosfolipase D/genética , Fosfolipase D/metabolismo , Alcamidas Poli-Insaturadas/metabolismo , Gravidez , Progesterona/sangue , Receptor CB1 de Canabinoide/genética , Receptor CB1 de Canabinoide/metabolismo , Receptor CB2 de Canabinoide/genética , Receptor CB2 de Canabinoide/metabolismo , Fatores de Tempo , Útero/efeitos dos fármacos , Útero/fisiopatologiaRESUMO
Endocrine-disrupting chemicals (EDCs) are a class of exogenous substances that mimic the effects of hormones in the body, inducing hormonal dysregulation and contributing to various disorders. Epigenome regulation has emerged as an important mechanism for maintaining organ function in health and disease. Dissecting epigenomic and resultant gene expression changes provides unprecedented insight into the chromatin state, which underlines disease development and shapes risk and phenotypic plasticity in response to the environment and internal cues. The cutting-edge, high throughput technologies provide new routes to understanding the etiology of disease and new footholds on the promising path to better treatment and disease prevention. We have recently revealed that myometrial stem cells (MMSCs), the cell origin of UFs, are the target of developmental EDC exposure. The EDC-induced epigenetic changes in MMSCs identified by multi-omics approaches include DNA methylation and histone modification modulated by DNA methyltransferases and MLL1, which characterized the molecular mechanism underlying EDC-related risk in hormone-dependent UFs. Future studies are needed to determine the link between real-life exposures to EDCs and their impact on the development of human diseases and transgenerational epigenetic inheritance, which can help explore strategies that may prevent adverse outcomes linked to EDC exposure.
Assuntos
Disruptores Endócrinos , Leiomioma , Humanos , Disruptores Endócrinos/toxicidade , Epigenômica , Leiomioma/induzido quimicamente , Leiomioma/genética , Epigênese GenéticaRESUMO
Uterine fibroids (UFs; leiomyoma) are the most common benign neoplastic threat to women worldwide, exacting an immense personal burden on female health and a monetary expense to the healthcare system estimated in the hundreds of billions of dollars every year globally. With no long-term non-invasive treatment option currently available to treat UFs, deeper insights regarding tumor etiology are the key for developing newer therapies. Accordingly, in this review, we discuss new mechanistic paradigm to explain UF tumor development through an exquisite model involving developmental reprogramming of myometrial stem cells due to early life endocrine disruptors exposure, inflammation, fibrosis, DNA damage, and eventually tissue stiffness. Further, we propose to utilize shear wave elastography as a potential screening tool for the early identification of women at risk for developing UFs who can benefit from several simple preventive strategies, including the consumption of natural compounds such as vitamin D and green tea as a safe fertility friendly non-hormonal modality to delay or even arrest or reverse UF progression.
RESUMO
Environmental exposure to endocrine-disrupting chemicals (EDCs) is linked to the development of uterine fibroids (UFs) in women. UFs, non-cancerous tumors, are thought to originate from abnormal myometrial stem cells (MMSCs). Defective DNA repair capacity may contribute to the emergence of mutations that promote tumor growth. The multifunctional cytokine TGFß1 is associated with UF progression and DNA damage repair pathways. To investigate the impact of EDC exposure on TGFß1 and nucleotide excision repair (NER) pathways, we isolated MMSCs from 5-months old Eker rats exposed neonatally to Diethylstilbestrol (DES), an EDC, or to vehicle (VEH). EDC-MMSCs exhibited overactivated TGFß1 signaling and reduced mRNA and protein levels of NER pathway components compared to VEH-MMSCs. EDC-MMSCs also demonstrated impaired NER capacity. Exposing VEH-MMSCs to TGFß1 decreased NER capacity while inhibiting TGFß signaling in EDC-MMSCs restored it. RNA-seq analysis and further validation revealed decreased expression of Uvrag, a tumor suppressor gene involved in DNA damage recognition, in VEH-MMSCs treated with TGFß1, but increased expression in EDC-MMSCs after TGFß signaling inhibition. Overall, we demonstrated that the overactivation of the TGFß pathway links early-life exposure to EDCs with impaired NER capacity, which would lead to increased genetic instability, arise of mutations, and fibroid tumorigenesis. We demonstrated that the overactivation of the TGFß pathway links early-life exposure to EDCs with impaired NER capacity, which would lead to increased fibroid incidence.
RESUMO
Uterine leiomyomas are the most common pelvic tumor in women of reproductive age; they cause irregular heavy menstrual bleeding leading to anemia and subsequent negative effects on quality of life. Exosomes have arisen as main players of disease progression in several illnesses, including a range of benign and malignant conditions; however, their role in leiomyomas' pathophysiology remains unknown. We investigated the effect of exosomes derived from human uterine leiomyoma tumor cells (HULM) and human myometrial cells (UTSM) on the behavior of human endometrial microvascular endothelial cells (HEMEC). HULM- and UTSM-derived exosomes were isolated and cocultured with HEMECs. Then, cell proliferation, mRNA expression, tube formation assay, and RNA-seq were performed. Treatment of HEMEC with HULM-derived exosomes increased cell proliferation by 60% compared to control untreated cells, upregulated C-MYC and VEGFA expression levels, and increased tube formation, length, and branching (markers of angiogenesis). Profiling of miRNA revealed that 84 miRNAs were significantly downregulated and 71 were upregulated in HULM-derived exosomes compared to UTSM-derived exosomes. These findings suggest that HULM-derived exosomes might have effects on HEMEC function, containing factors that enhance endometrial proliferation and angiogenesis, which may contribute to heavy menstrual bleeding. Further research on exosomes in uterine leiomyoma may identify possible novel biomarkers for treatment.
RESUMO
Uterine leiomyosarcoma (uLMS) is the most frequent subtype of uterine sarcoma that presents a poor prognosis, high rates of recurrence, and metastasis. Currently, the molecular mechanism of the origin and development of uLMS is unknown. Class I histone deacetylases (including HDAC1, 2, 3, and 8) are one of the major classes of the HDAC family and catalyze the removal of acetyl groups from lysine residues in histones and cellular proteins. Class I HDACs exhibit distinct cellular and subcellular expression patterns and are involved in many biological processes and diseases through diverse signaling pathways. However, the link between class I HDACs and uLMS is still being determined. In this study, we assessed the expression panel of Class I HDACs in uLMS and characterized the role and mechanism of class I HDACs in the pathogenesis of uLMS. Immunohistochemistry analysis revealed that HDAC1, 2, and 3 are aberrantly upregulated in uLMS tissues compared to adjacent myometrium. Immunoblot analysis demonstrated that the expression levels of HDAC 1, 2, and 3 exhibited a graded increase from normal and benign to malignant uterine tumor cells. Furthermore, inhibition of HDACs with Class I HDACs inhibitor (Tucidinostat) decreased the uLMS proliferation in a dose-dependent manner. Notably, gene set enrichment analysis of differentially expressed genes (DEGs) revealed that inhibition of HDACs with Tucidinostat altered several critical pathways. Moreover, multiple epigenetic analyses suggested that Tucidinostat may alter the transcriptome via reprogramming the oncogenic epigenome and inducing the changes in microRNA-target interaction in uLMS cells. In the parallel study, we also determined the effect of DL-sulforaphane on the uLMS. Our study demonstrated the relevance of class I HDACs proteins in the pathogenesis of malignant uLMS. Further understanding the role and mechanism of HDACs in uLMS may provide a promising and novel strategy for treating patients with this aggressive uterine cancer.
Assuntos
Leiomiossarcoma , Neoplasias Uterinas , Feminino , Humanos , Histona Desacetilases/metabolismo , Leiomiossarcoma/tratamento farmacológico , Leiomiossarcoma/genética , Leiomiossarcoma/patologia , Neoplasias Uterinas/tratamento farmacológico , Neoplasias Uterinas/genética , Neoplasias Uterinas/patologia , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/uso terapêutico , Miométrio/metabolismoRESUMO
Uterine fibroids are benign monoclonal neoplasms of the myometrium, representing the most common tumors in women worldwide. To date, no long-term or noninvasive treatment option exists for hormone-dependent uterine fibroids, due to the limited knowledge about the molecular mechanisms underlying the initiation and development of uterine fibroids. This paper comprehensively summarizes the recent research advances on uterine fibroids, focusing on risk factors, development origin, pathogenetic mechanisms, and treatment options. Additionally, we describe the current treatment interventions for uterine fibroids. Finally, future perspectives on uterine fibroids studies are summarized. Deeper mechanistic insights into tumor etiology and the complexity of uterine fibroids can contribute to the progress of newer targeted therapies.
Assuntos
Leiomioma , Neoplasias Uterinas , Feminino , Humanos , Leiomioma/etiologia , Leiomioma/terapia , Miométrio/patologia , Fatores de Risco , Neoplasias Uterinas/etiologia , Neoplasias Uterinas/patologia , Neoplasias Uterinas/terapiaRESUMO
Uterine leiomyosarcoma (uLMS) is the most common type of uterine sarcoma associated with poor prognosis, high rates of recurrence, and metastasis. There is currently limited information about uLMS molecular mechanisms of origin and development. Bromodomain (BRD)-containing proteins are involved in many biological processes, most notably epigenetic regulation of transcription, and BRD protein dysfunction has been linked to many diseases including tumorigenesis. However, the role of BRD proteins in the pathogenesis of uLMS is unknown. Here, we show for the first time that BRD9 is aberrantly overexpressed in uLMS tissues compared to adjacent myometrium. BRD9 expression is also upregulated in uLMS cell lines compared to benign uterine fibroid and myometrium cell lines. Inhibition of BRD9 using the specific inhibitor (TP-472) suppressed uLMS cell proliferation via inducing apoptosis and cell cycle arrest. To further characterize the mechanistic basis for TP-472 inhibition of uLMS cell growth, we performed a comparative RNA-seq analysis of vehicle-treated and TP-472-treated uLMS cells (n = 4 each). Bioinformatics analysis revealed that TP-472 treatment distinctly altered the uLMS cell transcriptome. Gene set enrichment analysis identified critical pathways altered by BRD9 inhibition, including interferon-alpha response, KRAS signaling, MYC targets, TNF-a signaling via NFkB, and MTORC1 signaling. Parsimonious gene correlation network analysis identified nine enriched modules, including cell cycle and apoptosis modules. Moreover, the ENCODE Histone Modifications gene set and TargetScan microRNA analysis in Enrichr suggested that TP-472-induced BRD9 inhibition may alter the uLMS cell transcriptome by reprograming the oncogenic epigenome and inducing miRNA-mediated gene regulation. Therefore, BRD9 constitutes a specific vulnerability in malignant uLMS, and targeting non-BET BRD proteins in uLMS may provide a promising and novel strategy for treating patients with this aggressive uterine cancer.
Assuntos
Leiomioma , Leiomiossarcoma , MicroRNAs , Fatores de Transcrição , Neoplasias Uterinas , Carcinogênese/genética , Epigênese Genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Leiomiossarcoma/patologia , MicroRNAs/genética , Fatores de Transcrição/metabolismo , Neoplasias Uterinas/patologiaRESUMO
Uterine fibroids (leiomyomas) are the most common benign gynecological tumors in women of reproductive age worldwide. They cause heavy menstrual bleeding, usually leading to severe anemia, pelvic pain/pressure, infertility, and other debilitating morbidities. Fibroids are believed to be monoclonal tumors arising from the myometrium, and recent studies have demonstrated that fibroids actively influence the endometrium globally. Studies suggest a direct relationship between the number of fibroids removed and fertility problems. In this review, our objective was to provide a complete overview of the origin of uterine fibroids and the molecular pathways and processes implicated in their development and growth, which can directly affect the function of a healthy endometrium. One of the most common characteristics of fibroids is the excessive production of extracellular matrix (ECM) components, which contributes to the stiffness and expansion of fibroids. ECM may serve as a reservoir of profibrotic growth factors such as the transforming growth factor ß (TGF-ß) and a modulator of their availability and actions. Fibroids also elicit mechanotransduction changes that result in decreased uterine wall contractility and increased myometrium rigidity, which affect normal biological uterine functions such as menstrual bleeding, receptivity, and implantation. Changes in the microRNA (miRNA) expression in fibroids and myometrial cells appear to modulate the TGF-ß pathways and the expression of regulators of ECM production. Taken together, these findings demonstrate an interaction among the ECM components, TGF-ß family signaling, miRNAs, and the endometrial vascular system. Targeting these components will be fundamental to developing novel pharmacotherapies that not only treat uterine fibroids but also restore normal endometrial function.
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Maternal overnutrition negatively impacts the offspring's health leading to an increased risk of developing chronic diseases or metabolic syndrome in adulthood. What we eat affects the endocannabinoid system (eCS) activity, which in turn modulates lipogenesis and fatty acids utilization in hepatic, muscle, and adipose tissues. This study aimed to evaluate the transgenerational effect of maternal obesity on cannabinoid receptor 1 knock-out (CB1 KO) animals in combination with a postnatal obesogenic diet on the development of metabolic disturbances on their offspring. CB1 KO mice were fed a control diet (CD) or a high-fat diet (HFD; 33% more energy from fat) for 3 months. Offspring born to control and obese mothers were also fed with CD or HFD. We observed that pups born to an HFD-fed mother presented higher postnatal weight, lower hepatic fatty acid amide hydrolase activity, and increased blood cholesterol levels when compared to the offspring born to CD-fed mothers. When female mice born to HFD-fed CB1 KO mothers were exposed to an HFD, they gained more weight, presented elevated blood cholesterol levels, and more abdominal adipose tissue accumulation than control-fed adult offspring. The eCS is involved in several reproductive physiological processes. Interestingly, we showed that CB1 KO mice in gestational day 15 presented resistance to LPS-induced deleterious effects on pregnancy outcome, which was overcome when these mice were obese. Our results suggest that an HFD in CB1 receptor-deficient mice contributes to a "nutritional programming" of the offspring resulting in increased susceptibility to metabolic challenges both perinatally and during adulthood.
Assuntos
Lipopolissacarídeos/efeitos adversos , Obesidade Materna/genética , Receptor CB1 de Canabinoide/genética , Animais , Animais Recém-Nascidos , Dieta Hiperlipídica/efeitos adversos , Feminino , Fenômenos Fisiológicos da Nutrição Materna , Doenças Metabólicas/etiologia , Doenças Metabólicas/genética , Doenças Metabólicas/metabolismo , Camundongos , Camundongos Knockout , Obesidade , Obesidade Materna/metabolismo , Gravidez , Resultado da Gravidez , Efeitos Tardios da Exposição Pré-Natal/etiologia , Efeitos Tardios da Exposição Pré-Natal/genética , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Receptor CB1 de Canabinoide/metabolismoRESUMO
Cryopreservation by negatively affecting sperm quality decreases the efficiency of assisted reproduction techniques (ARTs). Thus, we first evaluated sperm motility at different conditions for the manipulation of equine cryopreserved spermatozoa. Higher motility was observed when spermatozoa were incubated for 30 min at 30 × 106/mL compared to lower concentrations (p < 0.05) and when a short centrifugation at 200× g was performed (p < 0.05). Moreover, because sperm suitable for oocyte fertilization is released from oviduct epithelial cells (OECs), in response to the capacitation process, we established an in vitro OEC culture model to select a sperm population with potential fertilizing capacity in this species. We demonstrated E-cadherin and cytokeratin expression in cultures of OECs obtained. When sperm-OEC cocultures were performed, the attached spermatozoa were motile and presented an intact acrosome, suggesting a selection by the oviductal model. When co-cultures were incubated in capacitating conditions a greater number of alive (p < 0.05), capacitated (p < 0.05), with progressive motility (p < 0.05) and with the intact acrosome sperm population was observed (p < 0.05) suggesting that the sperm population released from OECs in vitro presents potential fertilizing capacity. Improvements in handling and selection of cryopreserved sperm would improve efficiencies in ARTs allowing the use of a population of higher-quality sperm.
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PURPOSE OF REVIEW: Uterine leiomyoma (fibroids) is a gynecologic disorder impacting the majority of women in the United States. When symptomatic, these noncancerous tumors can cause severe morbidity including pelvic pain, menorrhagia, and infertility. Endocrine-disrupting chemicals (EDCs) may represent a modifiable risk factor. The aim of this review is to summarize recent human and experimental evidence on EDCs exposures and fibroids. RECENT FINDINGS: Multiple EDCs are associated with fibroid outcomes and/or processes including phthalates, parabens, environmental phenols, alternate plasticizers, Diethylstilbestrol, organophosphate esters, and tributyltin. Epidemiologic studies suggest exposure to certain EDCs, such as di-(2-ethylhxyl)-phthalate (DEHP), are associated with increased fibroid risk and severity. Both human and experimental studies indicate that epigenetic processes may play an important role in linking EDCs to fibroid pathogenesis. In-vitro and in-vivo studies show that DEHP, bisphenol A, and diethylstilbestrol can impact biological pathways critical to fibroid pathogenesis. SUMMARY: While research on EDCs and fibroids is still evolving, recent evidence suggests EDC exposures may contribute to fibroid risk and progression. Further research is needed to examine the impacts of EDC mixtures and to identify critical biological pathways and windows of exposure. These results could open the door to new prevention strategies for fibroids.
Assuntos
Disruptores Endócrinos/toxicidade , Leiomioma/induzido quimicamente , Neoplasias Uterinas/induzido quimicamente , Animais , Exposição Ambiental/efeitos adversos , Exposição Ambiental/estatística & dados numéricos , Epigênese Genética/efeitos dos fármacos , Feminino , Interação Gene-Ambiente , Humanos , Leiomioma/epidemiologia , Leiomioma/genética , Fatores de Risco , Estados Unidos/epidemiologia , Neoplasias Uterinas/epidemiologia , Neoplasias Uterinas/genéticaRESUMO
It is not easy to separate frozen-thawed South American camelid sperm from seminal plasma (SP) and diluents to be used for in vitro embryo production. The objective of this study was to evaluate Androcoll-E™ (AE) efficiency to separate llama sperm from SP and freezing extender in frozen-thawed semen. A total of 22 ejaculates from five Lama glama males were collected using electroejaculation. After performing semen analysis (sperm motility, concentration, viability, membrane function, and acrosome integrity), samples were cryopreserved with a diluent containing lactose, ethylenediaminetetraacetic acid (EDTA), egg yolk, and 7% dimethylformamide. After thawing, samples were divided in aliquots, one of which was used as a control and the others processed by AE. Experiment 1 (12 ejaculates): 100 µl of frozen-thawed semen was placed on top of 1,000 µl AE column and centrifuged at 800 g for 10 min. Experiment 2 (10 ejaculates): two samples of 100 µl of frozen-thawed semen were placed on two columns of 500 µl AE each, and both were centrifuged at 800 g for 10 and 20 min, respectively. Pellets were resuspended in Tyrode's albumin lactate pyruvate (TALP) medium, and sperm parameters were evaluated. A significant decrease in all sperm parameters was observed in thawed samples compared to raw semen. AE allowed the separation of frozen-thawed sperm from SP and freezing extender independently from the height of the column used and time of centrifugation assayed. Although no significant differences were found between AE columns, higher sperm recovery was observed with 500 µl of AE coupled with 20 min of centrifugation. Despite the significant decrease observed in sperm motility in AE samples, no changes in sperm viability, membrane function, and acrosome integrity were observed when comparing control thawed semen with the sperm recovered after AE (p > 0.05). The use of AE columns, either 500 or 1,000 µl, allows the separation of frozen-thawed llama sperm from SP and freezing extender, preserving the viability, membrane function, and acrosome integrity. Of the protocols studied, 800 g centrifugation during 20 min using a 500 µl column of AE would be the method of choice to process frozen-thawed llama semen destined for reproductive biotechnologies.
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Genital tract infections caused by Gram-negative bacteria induce miscarriage and are one of the most common complications of human pregnancy. LPS administration to 7-day pregnant mice induces embryo resorption after 24h, with nitric oxide playing a fundamental role in this process. We have previously shown that progesterone exerts protective effects on the embryo by modulating the inflammatory reaction triggered by LPS. Here we sought to investigate whether the in vivo administration of progesterone modulated the LPS-induced nitric oxide production from peripheral blood mononuclear cells from pregnant and non-pregnant mice. We found that progesterone downregulated LPS-induced nitric oxide production by a progesterone receptor-independent mechanism. Moreover, our results suggest a possible participation of glucocorticoid receptors in at least some of the anti-inflammatory effects of progesterone.