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1.
Artigo em Inglês | MEDLINE | ID: mdl-31910029

RESUMO

Excess androgen-induced obesity has become a public health problem, and its prevalence has increased substantially in recent years. Chemokine-like receptor 1 (CMKLR1), a receptor of Chemerin secreted by adipose tissue, is linked to adipocyte differentiation, adipose tissue development and obesity. However, the effect of CMKLR1 signaling on androgen-mediated adiposity in vivo remains unclear. Here, using pharmacological method, orchidectomized model and CMKLR1 knockout mice, we demonstrated that androgen excess in female mice resulted in a larger cell size in white adipose tissue (WAT) and brown adipose tissue (BAT), whereas androgen deprivation of male mice induced a smaller cell size. Both effects relating to the adipocytes size were both attenuated in CMKLR1 knockout mice. CMKLR1 deficiency influenced the effect of androgen on adipose tissue by regulating the mRNA expression of androgen receptor (AR) and adipocyte markers (such as Fabp4 and Cidea). Moreover, in vivo suppression of CMKLR1 by its antagonist α-NETA can also weaken the enlargement of the adipocyte cell size caused by 5α-dihydrotestosterone (DHT). Furthermore, using in vitro BAT explants culture and PI3K signaling antagonist wortmannin, we found DHT could reduce the phosphorylated ERK (pERK) in BAT, whereas CMKLR1 inactivation inhibited this effect caused by DHT through PI3K signaling pathway. Taken together, these findings reveal an anti-obesity role of CMKLR1 deficiency in regulating lipid accumulation, highlighting the scientific importance for further development of small molecule CMKLR1 antagonists as fundamental scientific tools or a potential drug for the treatment of adiposity.

2.
Artigo em Inglês | MEDLINE | ID: mdl-31851527

RESUMO

R-Spondin3 (RSPO3), an activator of Wnt/ß-catenin signaling, plays a key role in tumorigenesis of various cancers, but its role in choriocarcinoma remains unknown. To investigate the effect of RSPO3 on the tumor growth of choriocarcinoma JEG-3 cells, the expression of RSPO3 in human term placenta was detected, and a stable RSPO3-overexpressing JEG-3 cell line was established via lentivirus-mediated transduction. The expressions of biomarkers involving in tumorigenicity were detected in the RSPO3-overexpressing JEG-3 cells, and cell proliferation, invasion, migration and apoptosis were investigated. Moreover, soft agar clonogenicity and xenograft tumorigenicity were performed to assess the effect of RSPO3 on tumor growth in vitro and in vivo. The results showed that RSPO3 was widely expressed in human term placenta, and overexpression of RSPO3 promoted the proliferation, inhibited the migration, invasion and apoptosis of the JEG-3 cells. Meanwhile, RSPO3 overexpression promoted tumor growth both in vivo and in vitro. Further investigation showed that the phosphorylation levels of AKT, PI3K and ERK as well the expression of ß-catenin and PCNA were increased in the RSPO3-overexpressing JEG-3 cells and tumor xenograft. Taken together, these data indicate that RSPO3 promotes the tumor growth of choriocarcinoma via AKT/PI3K/ERK signaling, which supports RSPO3 as an oncogenic driver promoting the progression of choriocarcinoma.

3.
Int J Mol Sci ; 20(21)2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31683965

RESUMO

The available and effective therapeutic means to treat choriocarcinoma is seriously lacking, mainly due to the toxic effects caused by chemotherapy and radiotherapy. Accordingly, we developed a method for targeting delivery of chemotherapeutical drugs only to cancer cells, not normal cells, in vivo, by using a synthetic placental chondroitin sulfate (CSA)-binding peptide (plCSA-BP) derived from malarial protein VAR2CSA. A 28 amino acids placental CSA-binding peptide (plCSA-BP) from the VAR2CSA was synthesized as a guiding peptide for tumor-targeting delivery, dendrigraft poly-L-lysines (DGL) was modified with plCSA-BP and served as a novel targeted delivery carrier. Choriocarcinoma was selected to test the effect of targeted delivery carrier, and prodigiosin isolated from Serratia marcescens subsp. lawsoniana was selected as a chemotherapeutical drug and encapsulated in the DGL modified by the plCSA-BP nanoparticles (DGL/CSA-PNPs). DGL/CSA-PNPs had a sustained slow-release feature at pH 7.4, which could specifically bind to the JEG3 cells and exhibited better anticancer activity than that of the controls. The DGL/CSA-PNPs induced the apoptosis of JEG3 cells through caspase-3 and the P53 signaling pathway. DGL/CSA-PNPs can be used as an excellent targeted delivery carrier for anticancer drugs, and the prodigiosin could be an alternative chemotherapeutical drug for choriocarcinoma.

4.
Am J Physiol Cell Physiol ; 317(3): C556-C565, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31241986

RESUMO

Choriocarcinoma is characterized by malignant proliferation and transformation of trophoblasts and is currently treated with systemic chemotherapeutic agents. The lack of specific targets for chemotherapeutic agents results in indiscriminate drug distribution. In our study, we aimed to delineate the mechanism by which G protein-coupled receptor 1 (GPR1) regulates the development of choriocarcinoma and thus investigated GPR1 as a prospective chemotherapeutic target. In this study, GPR1 expression levels were examined in several trophoblast cell lines. We found significantly higher GPR1 expression in choriocarcinoma cells (JEG3 and BeWo) than in normal trophoblast cells (HTR-8/SVneo). Additionally, we studied the role of GPR1 in choriocarcinoma in vitro and in vivo. GPR1 knockdown suppressed proliferation, invasion, and Akt and ERK phosphorylation in vitro and slowed tumor growth in vivo. Interestingly, GPR1 overexpression promoted increased proliferation, invasion, and Akt and ERK phosphorylation in vitro. Furthermore, we identified a specific GPR1-binding seven-amino acid peptide, LRH7-G3, that might also suppress choriocarcinoma in vitro and in vivo through phage display. Our study is the first to report that GPR1 may play a role in regulating choriocarcinoma progression through the Akt and ERK pathways. GPR1 could be a promising potential pharmaceutical target for choriocarcinoma.

5.
Enzyme Microb Technol ; 125: 45-52, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30885324

RESUMO

Xanthophyllomyces dendrorhous is an excellent industrial source for production of natural astaxanthin, but the yield of astaxanthin is relative low due to the contradiction between biomass weight and astaxanthin accumulation. Glutamate, a metabolite connecting nitrogen and carbon metabolisms, is probably a promising entry point to interfere cellular metabolisms. Thus, the effect of glutamate on cell growth and astaxanthin accumulation in X. dendrorhous was investigated. Results showed that glutamate feeding facilitated glucose consumption and further led to the increment of astaxanthin content with little influence of cell growth. A comparative proteomics study was applied to decipher the regulatory mechanisms of enhanced astaxanthin biosynthesis in X. dendrorhous as a response to the glutamate feeding. The expressions of proteins with the highest degree of fold change were involved in carbohydrate, amino acids, and carotenogenesis metabolisms as well as redox and stress-associated metabolisms. In addition, a possible regulatory model of enhanced astaxanthin accumulation in response to glutamate feeding in X. dendrorhous is also proposed.


Assuntos
Basidiomycota/metabolismo , Ácido Glutâmico/metabolismo , Aminoácidos/metabolismo , Basidiomycota/crescimento & desenvolvimento , Biomassa , Metabolismo dos Carboidratos/efeitos dos fármacos , Carotenoides/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Perfilação da Expressão Gênica , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Ácido Glutâmico/farmacologia , Redes e Vias Metabólicas , Modelos Biológicos , Oxirredução/efeitos dos fármacos , Proteômica , Estresse Fisiológico/efeitos dos fármacos , Xantofilas/biossíntese , Xantofilas/metabolismo
6.
Am J Physiol Endocrinol Metab ; 316(6): E987-E997, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30835511

RESUMO

Chemerin and G protein-coupled receptor 1 (GPR1) are increased in serum and placenta in mice during pregnancy. Interestingly, we observed increased serum chemerin levels and decreased GPR1 expression in placenta of high-fat-diet-fed mice compared with chow-fed mice at gestational day 18. GPR1 protein and gene levels were significantly decreased in gestational diabetes mellitus (GDM) patient placentas. Therefore, we hypothesized that chemerin/GPR1 signaling might participate in the pathogenic mechanism of GDM. We investigated the role of GPR1 in carbohydrate homeostasis during pregnancy using pregnant mice transfected with small interfering RNA for GPR1 or a negative control. GPR1 knockdown exacerbated glucose intolerance, disrupted lipid metabolism, and decreased ß-cell proliferation and insulin levels. Glucose transport protein-3 and fatty acid binding protein-4 were downregulated with reducing GPR1 in vivo and in vitro via phosphorylated AKT pathway. Taken together, our findings first demonstrate the expression of GPR1, the characterization of its direct biological effects in humans and mice, as well as the molecular mechanism that indicates the role of GPR1 signaling in maternal metabolism during pregnancy, suggesting a novel feedback mechanism to regulate glucose balance during pregnancy, and GPR1 could be a potential target for the detection and therapy of GDM.

7.
Cell Mol Life Sci ; 76(2): 355-367, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30374519

RESUMO

The adipokine Chemerin and its receptor, chemokine-like receptor 1 (CMKLR1), are associated with osteoblastogenic differentiation of mesenchymal stem cells (MSCs) and osteoclastogenic differentiation of osteoclast precursors in vitro, suggesting that CMKLR1 would affect the bone mineral density (BMD). However, the role of CMKLR1 on BMD in vivo remains unknown. Here, using CMKLR1 knockout mouse model, we unveiled that CMKLR1 effected the amount of Leydig cells in testis and regulated androgen-dependent bone maintenance in male mice, which exhibited lower serum testosterone levels, thereby reducing the trabecular bone mass. Correspondingly, the mRNA expression of testosterone synthesis enzymes in testis decreased. The bone tissue also showed decreased mRNAs expression of osteogenic markers and increased mRNA levels for osteoclast markers. Furthermore, by in vitro differentiation models, we found CMKLR1-deficiency could break the balance between osteoblastogenesis and osteoclastogenesis that caused a shift from osteogenic to adipogenic differentiation in MSCs and enhanced osteoclast formation. In addition, bone mass increase in CMKLR1 KO male mice can be promoted by treatment with 5α-dihydrotestosterone (DHT), and the inactivation of CMKLR1 in male wild-type (WT) mice with antagonist treatment can lead to low bone mass. Taken together, these data indicate that CMKLR1 positively regulates bone metabolism through mediating testosterone production and the balance between osteoblast and osteoclast formation.


Assuntos
Densidade Óssea , Receptores Acoplados a Proteínas-G/genética , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Diferenciação Celular , Proteínas de Ligação a Ácido Graxo/genética , Proteínas de Ligação a Ácido Graxo/metabolismo , Fêmur/diagnóstico por imagem , Fêmur/fisiologia , Interleucina-1beta/análise , Interleucina-6/análise , Células Intersticiais do Testículo/citologia , Células Intersticiais do Testículo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteoclastos/citologia , Osteoclastos/metabolismo , Osteogênese , PPAR gama/genética , PPAR gama/metabolismo , Receptores Acoplados a Proteínas-G/deficiência , Testículo/metabolismo , Testículo/patologia , Testosterona/biossíntese , Testosterona/sangue , Tíbia/diagnóstico por imagem , Tíbia/fisiologia
8.
Reprod Biol Endocrinol ; 16(1): 50, 2018 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-29793502

RESUMO

BACKGROUND: Polycystic ovary syndrome (PCOS) is a complex genetic disease with multifarious phenotypes. Many researches use dehydroepiandrosterone (DHEA) to induce PCOS in pubertal mouse models. The aim of this study was to investigate the role of GPR1 in dehydroepiandrosterone (DHEA)-induced hyperandrogenized mice. METHODS: Prepubertal C57BL/6 mice (25 days of age) and Gpr1-deficient mice were each divided into two groups and injected daily with sesame oil with or without DHEA (6 mg/100 g) for 21 consecutive days. Hematoxylin and eosin (H&E) staining was performed to determine the characteristics of the DHEA-treated ovaries. Real-time PCR was used to examine steroid synthesis enzymes gene expression. Granulosa cell was cultured to explore the mechanism of DHEA-induced, GPR1-mediated estradiol secretion. RESULTS: DHEA treatment induced some aspects of PCOS in wild-type mice, such as increased body weight, elevated serum testosterone, increased number of small, cystic, atretic follicles, and absence of corpus luteum in ovaries. However, Gpr1 deficiency significantly attenuated the DHEA-induced weight gain and ovarian phenotype, improving steroidogenesis in ovaries and estradiol synthesis in cultured granulosa cells, partially through mTOR signaling. CONCLUSIONS: In conclusion, Gpr1 deficiency leads to the improvement of steroid synthesis in mice hyperandrogenized with DHEA, indicating that GPR1 may be a therapeutic target for DHEA-induced hyperandrogenism.


Assuntos
Hiperandrogenismo/sangue , Hiperandrogenismo/genética , Receptores Acoplados a Proteínas-G/genética , Testosterona/sangue , Animais , Células Cultivadas , Desidroepiandrosterona , Modelos Animais de Doenças , Estradiol/sangue , Feminino , Hiperandrogenismo/induzido quimicamente , Hiperandrogenismo/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Síndrome do Ovário Policístico/sangue , Síndrome do Ovário Policístico/induzido quimicamente , Síndrome do Ovário Policístico/genética , Síndrome do Ovário Policístico/patologia
9.
Theranostics ; 8(10): 2765-2781, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29774074

RESUMO

Rationale: The availability of therapeutics to treat pregnancy complications is severely lacking, mainly due to the risk of harm to the fetus. In placental malaria, Plasmodium falciparum-infected erythrocytes (IEs) accumulate in the placenta by adhering to chondroitin sulfate A (CSA) on the surfaces of trophoblasts. Based on this principle, we have developed a method for targeted delivery of payloads to the placenta using a synthetic placental CSA-binding peptide (plCSA-BP) derived from VAR2CSA, a CSA-binding protein expressed on IEs. Methods: A biotinylated plCSA-BP was used to examine the specificity of plCSA-BP binding to mouse and human placental tissue in tissue sections in vitro. Different nanoparticles, including plCSA-BP-conjugated nanoparticles loaded with indocyanine green (plCSA-INPs) or methotrexate (plCSA-MNPs), were administered intravenously to pregnant mice to test their efficiency at drug delivery to the placenta in vivo. The tissue distribution and localization of the plCSA-INPs were monitored in live animals using an IVIS imaging system. The effect of plCSA-MNPs on fetal and placental development and pregnancy outcome were examined using a small-animal high-frequency ultrasound (HFUS) imaging system, and the concentrations of methotrexate in fetal and placental tissues were measured using high-performance liquid chromatography (HPLC). Results: plCSA-BP binds specifically to trophoblasts and not to other cell types in the placenta or to CSA-expressing cells in other tissues. Moreover, we found that intravenously administered plCSA-INPs accumulate in the mouse placenta, and ex vivo analysis of the fetuses and placentas confirmed placenta-specific delivery of these nanoparticles. We also demonstrate successful delivery of methotrexate specifically to placental cells by plCSA-BP-conjugated nanoparticles, resulting in dramatic impairment of placental and fetal development. Importantly, plCSA-MNPs treatment had no apparent adverse effects on maternal tissues. Conclusion: These results demonstrate that plCSA-BP-guided nanoparticles could be used for the targeted delivery of payloads to the placenta and serve as a novel placenta-specific drug delivery option.


Assuntos
Nanopartículas/metabolismo , Trofoblastos/metabolismo , Animais , Antígenos de Protozoários/metabolismo , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/farmacocinética , Feminino , Humanos , Metotrexato/administração & dosagem , Metotrexato/farmacocinética , Camundongos , Nanopartículas/efeitos adversos , Gravidez
10.
Drug Deliv ; 25(1): 461-471, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29426237

RESUMO

Gestational trophoblastic neoplasia (GTN) can result from the over-proliferation of trophoblasts. Treatment of choriocarcinoma, the most aggressive GTN, currently requires high doses of systemic chemotherapeutic agents, which result in indiscriminate drug distribution and severe toxicity. To overcome these disadvantages and enhance the chemotherapeutic efficacy, chondroitin sulfate A (CSA)-binding nanoparticles were developed for the targeted delivery of doxorubicin (DOX) to choriocarcinoma cells using a synthetic CSA-binding peptide (CSA-BP), derived from malarial protein, which specifically binds to the CSA exclusively expressed in the placental trophoblast. CSA-BP-conjugated nanoparticles rapidly bonded to choriocarcinoma (JEG3) cells and were efficiently internalized into the lysosomes. Moreover, CSA-BP modification significantly increased the anti-cancer activity of the DOX-loaded nanoparticles in vitro. Intravenous injections of CSA-BP-conjugated nanoparticles loaded with indocyanine green (CSA-INPs) were rapidly localized to the tumor. The CSA-targeted nanoparticles loaded with DOX (CSA-DNPs) strongly inhibited primary tumor growth and, more importantly, significantly suppressed metastasis in vivo. Collectively, our results highlight the potential of the CSA-BP-decorated nanoparticles as an alternative targeted delivery system of chemotherapeutic agents for treating choriocarcinoma and for developing new GTN therapies based on drug targeting.


Assuntos
Sulfatos de Condroitina/administração & dosagem , Coriocarcinoma/tratamento farmacológico , Doxorrubicina/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Nanopartículas/administração & dosagem , Neoplasias Uterinas/tratamento farmacológico , Animais , Antibióticos Antineoplásicos/administração & dosagem , Antibióticos Antineoplásicos/metabolismo , Sítios de Ligação/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Sulfatos de Condroitina/metabolismo , Coriocarcinoma/metabolismo , Relação Dose-Resposta a Droga , Doxorrubicina/metabolismo , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanopartículas/metabolismo , Gravidez , Resultado do Tratamento , Neoplasias Uterinas/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
11.
Bioresour Technol ; 244(Pt 1): 664-671, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28813692

RESUMO

The type and concentration of inorganic carbon and nitrogen sources were manipulated to improve cell growth and lutein productivity of Desmodesmus sp. F51. Using nitrate as nitrogen source, the better cell growth and lutein accumulation were obtained under 2.5% CO2 supply when compared to the addition of NaHCO3 or Na2CO3. To solve the pH variation problem of ammonium consumption, the strategy of using dual carbon sources (NaHCO3 and CO2) was explored. A lower bicarbonate-C: ammonium-N ratio led to a lower culture pH as well as lower lutein productivity, but significantly enhanced the auto-flocculation efficiency of the microalgal cells. The highest biomass productivity (939mg/L/d) and lutein productivity (5.22mg/L/d) were obtained when the bicarbonate-C/ammonium-N ratio and ammonium-N concentration were 1:1 and 150mg/L, respectively. The lutein productivity of 5.22mg/L/d is the highest value ever reported in the literature using batch phototrophic cultivation.


Assuntos
Carbono , Clorófitas , Luteína , Compostos de Amônio , Biomassa , Microalgas
12.
Bioprocess Biosyst Eng ; 40(7): 1091-1100, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28455664

RESUMO

The ratio of carbon to nitrogen (C/N) in media plays a crucial role in the production of microbial carotenoids. However, the effects of a high C/N ratio on carotenoid production are ambiguous, and the mechanism of how C/N ratio affects astaxanthin accumulation in X. dendrorhous is unclear. In this study, the influence of C/N ratio on astaxanthin biosynthesis in X. dendrorhous at a fixed nitrogen concentration was investigated, and comparative proteomics were applied to address how C/N ratio affects cell growth and astaxanthin accumulation in X. dendrorhous. The results showed that cell growth and astaxanthin accumulation in X. dendrorhous were strongly related to the ratio of carbon to nitrogen with increasing C/N ratio in medium. However, the astaxanthin content per cell showed an inverse relationship, decreasing with an increasing C/N ratio. Differential proteomics showed the proteins with highest degree of change in expression under varying C/N ratios were mainly involved in carbohydrate metabolic pathways and carotenogenesis metabolism. In addition, several redox- and stress-associated proteins were up-regulated along with the carotenogenesis proteins, implying the environmental stress may affect metabolism and astaxanthin synthesis. A possible regulatory mechanism in response to glucose in X. dendrorhous is discussed.


Assuntos
Basidiomycota , Carbono , Proteômica , Xantofilas
13.
Biotechnol Bioeng ; 113(10): 2088-99, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27563850

RESUMO

The unicellular green alga Haematococcus pluvialis has been exploited as a cell factory to produce the high-value antioxidant astaxanthin for over two decades, due to its superior ability to synthesize astaxanthin under adverse culture conditions. However, slow vegetative growth under favorable culture conditions and cell deterioration or death under stress conditions (e.g., high light, nitrogen starvation) has limited the astaxanthin production. In this study, a new paradigm that integrated heterotrophic cultivation, acclimation of heterotrophically grown cells to specific light/nutrient regimes, followed by induction of astaxanthin accumulation under photoautotrophic conditions was developed. First, the environmental conditions such as pH, carbon source, nitrogen regime, and light intensity, were optimized to induce astaxanthin accumulation in the dark-grown cells. Although moderate astaxanthin content (e.g., 1% of dry weight) and astaxanthin productivity (2.5 mg L(-1) day(-1) ) were obtained under the optimized conditions, a considerable number of cells died off when subjected to stress for astaxanthin induction. To minimize the susceptibility of dark-grown cells to light stress, the algal cells were acclimated, prior to light induction of astaxanthin biosynthesis, under moderate illumination in the presence of nitrogen. Introduction of this strategy significantly reduced the cell mortality rate under high-light and resulted in increased cellular astaxanthin content and astaxanthin productivity. The productivity of astaxanthin was further improved to 10.5 mg L(-1) day(-1) by implementation of such a strategy in a bubbling column photobioreactor. Biochemical and physiological analyses suggested that rebuilding of photosynthetic apparatus including D1 protein and PsbO, and recovery of PSII activities, are essential for acclimation of dark-grown cells under photo-induction conditions. Biotechnol. Bioeng. 2016;113: 2088-2099. © 2016 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.


Assuntos
Clorófitas/metabolismo , Nitrogênio/metabolismo , Fotobiorreatores/microbiologia , Fotossíntese/fisiologia , Técnicas de Cultura de Células/métodos , Relação Dose-Resposta à Radiação , Luz , Estimulação Luminosa/métodos , Fotossíntese/efeitos da radiação , Doses de Radiação , Xantofilas/biossíntese , Xantofilas/isolamento & purificação , Xantofilas/efeitos da radiação
14.
J Endocrinol ; 230(1): 55-65, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27149986

RESUMO

Chemerin, a chemokine, plays important roles in immune responses, inflammation, adipogenesis, and carbohydrate metabolism. Our recent research has shown that chemerin has an inhibitory effect on hormone secretion from the testis and ovary. However, whether G protein-coupled receptor 1 (GPR1), the active receptor for chemerin, regulates steroidogenesis and luteolysis in the corpus luteum is still unknown. In this study, we established a pregnant mare serum gonadotropin-human chorionic gonadotropin (PMSG-hCG) superovulation model, a prostaglandin F2α (PGF2α) luteolysis model, and follicle and corpus luteum culture models to analyze the role of chemerin signaling through GPR1 in the synthesis and secretion of gonadal hormones during follicular/luteal development and luteolysis. Our results, for the first time, show that chemerin and GPR1 are both differentially expressed in the ovary over the course of the estrous cycle, with highest levels in estrus and metestrus. GPR1 has been localized to granulosa cells, cumulus cells, and the corpus luteum by immunohistochemistry (IHC). In vitro, we found that chemerin suppresses hCG-induced progesterone production in cultured follicle and corpus luteum and that this effect is attenuated significantly by anti-GPR1 MAB treatment. Furthermore, when the phosphoinositide 3-kinase (PI3K) pathway was blocked, the attenuating effect of GPR1 MAB was abrogated. Interestingly, PGF2α induces luteolysis through activation of caspase-3, leading to a reduction in progesterone secretion. Treatment with GPR1 MAB blocked the PGF2α effect on caspase-3 expression and progesterone secretion. This study indicates that chemerin/GPR1 signaling directly or indirectly regulates progesterone synthesis and secretion during the processes of follicular development, corpus luteum formation, and PGF2α-induced luteolysis.


Assuntos
Quimiocinas/metabolismo , Corpo Lúteo/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Luteólise/fisiologia , Receptores Acoplados a Proteínas-G/metabolismo , Transdução de Sinais/fisiologia , Superovulação/fisiologia , Animais , Caspase 3/metabolismo , Gonadotropina Coriônica/farmacologia , Corpo Lúteo/efeitos dos fármacos , Dinoprosta/farmacologia , Estradiol/metabolismo , Feminino , Luteólise/efeitos dos fármacos , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Progesterona/metabolismo , Transdução de Sinais/efeitos dos fármacos
15.
Sci Rep ; 6: 21328, 2016 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-26893072

RESUMO

Elevated serum chemerin levels correlate with increased severity of polycystic ovary syndrome (PCOS). However, the role of CMKLR1 signaling in ovarian biology under conditions of excess DHT remains unclear. In this study we compared the effects of continuous 90-day high dose DHT exposure (83.3 □g/day) on wild type and CMKLR1-deficient mice. DHT induced PCOS-like clinical signs in wild type mice as well as significant changes in the expression of hormone receptors, steroid synthesis enzymes, and BMPs and their receptors. In contrast, CMKLR1-deficient mice significantly attenuated DHT-induced clinical signs of PCOS and alterations in ovarian gene expression. To determine whether the BMP4 signaling pathway was involved in the pathogenic effects of CMKLR1 signaling in DHT-induced ovarian steroidogenesis, antral follicles were isolated from wild type and CMKLR1 knockout (KO) mice and treated in vitro with combinations of hCG, DHT, and BMP4 inhibitors. BMP4 inhibition attenuated the induction effects of hCG and DHT on estrogen and progesterone secretion in CMKLR1 KO mice, but not in WT mice, implicating the BMP4 signaling pathway in the CMKLR1-dependent response to DHT. In conclusion, CMKLR1 gene deletion attenuates the effects of chronic DHT treatment on ovarian function in experimental PCOS, likely via BMP4 signaling.


Assuntos
Di-Hidrotestosterona/metabolismo , Ovário/metabolismo , Receptores Acoplados a Proteínas-G/deficiência , Esteroides/biossíntese , Animais , Apoptose/genética , Di-Hidrotestosterona/farmacologia , Ciclo Estral , Feminino , Regulação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Hormônios Esteroides Gonadais/biossíntese , Camundongos , Camundongos Knockout , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Folículo Ovariano/patologia , Ovário/efeitos dos fármacos , Ovário/patologia , Síndrome do Ovário Policístico/genética , Síndrome do Ovário Policístico/metabolismo , Síndrome do Ovário Policístico/patologia , RNA Mensageiro/genética , Receptores de Esteroides/metabolismo , Esteroides/sangue
16.
Plant J ; 81(1): 95-107, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25353310

RESUMO

Astaxanthin, a red ketocarotenoid with strong antioxidant activity and high commercial value, possesses important physiological functions in astaxanthin-producing microalgae. The green microalga Haematococcus pluvialis accumulates up to 4% fatty acid-esterified astaxanthin (by dry weight), and is used as a model species for exploring astaxanthin biosynthesis in unicellular photosynthetic organisms. Although coordination of astaxanthin and fatty acid biosynthesis in a stoichiometric fashion was observed in H. pluvialis, the interaction mechanism is unclear. Here we dissected the molecular mechanism underlying coordination between the two pathways in H. pluvialis. Our results eliminated possible coordination of this inter-dependence at the transcriptional level, and showed that this interaction was feedback-coordinated at the metabolite level. In vivo and in vitro experiments indicated that astaxanthin esterification drove the formation and accumulation of astaxanthin. We further showed that both free astaxanthin biosynthesis and esterification occurred in the endoplasmic reticulum, and that certain diacylglycerol acyltransferases may be the candidate enzymes catalyzing astaxanthin esterification. A model of astaxanthin biosynthesis in H. pluvialis was subsequently proposed. These findings provide further insights into astaxanthin biosynthesis in H. pluvialis.


Assuntos
Clorófitas/metabolismo , Ácidos Graxos/biossíntese , Microalgas/metabolismo , Proteínas de Algas/metabolismo , Proteínas de Algas/fisiologia , Clorófitas/genética , Diacilglicerol O-Aciltransferase/metabolismo , Diacilglicerol O-Aciltransferase/fisiologia , Retículo Endoplasmático/metabolismo , Esterificação , Redes e Vias Metabólicas , Transcrição Genética , Xantofilas/biossíntese
17.
PLoS One ; 9(9): e106679, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25221928

RESUMO

The unicellular microalga Haematococcus pluvialis has emerged as a promising biomass feedstock for the ketocarotenoid astaxanthin and neutral lipid triacylglycerol. Motile flagellates, resting palmella cells, and cysts are the major life cycle stages of H. pluvialis. Fast-growing motile cells are usually used to induce astaxanthin and triacylglycerol biosynthesis under stress conditions (high light or nutrient starvation); however, productivity of biomass and bioproducts are compromised due to the susceptibility of motile cells to stress. This study revealed that the Photosystem II (PSII) reaction center D1 protein, the manganese-stabilizing protein PsbO, and several major membrane glycerolipids (particularly for chloroplast membrane lipids monogalactosyldiacylglycerol and phosphatidylglycerol), decreased dramatically in motile cells under high light (HL). In contrast, palmella cells, which are transformed from motile cells after an extended period of time under favorable growth conditions, have developed multiple protective mechanisms--including reduction in chloroplast membrane lipids content, downplay of linear photosynthetic electron transport, and activating nonphotochemical quenching mechanisms--while accumulating triacylglycerol. Consequently, the membrane lipids and PSII proteins (D1 and PsbO) remained relatively stable in palmella cells subjected to HL. Introducing palmella instead of motile cells to stress conditions may greatly increase astaxanthin and lipid production in H. pluvialis culture.


Assuntos
Clorófitas/citologia , Luz , Metabolismo dos Lipídeos , Aclimatação , Proteínas de Algas/metabolismo , Proteínas de Algas/fisiologia , Movimento Celular , Clorófitas/metabolismo , Clorófitas/efeitos da radiação , Cromatografia Líquida de Alta Pressão , Transporte de Elétrons , Fotossíntese/fisiologia , Complexo de Proteína do Fotossistema II/metabolismo , Complexo de Proteína do Fotossistema II/fisiologia , Pigmentos Biológicos/química
18.
J Exp Bot ; 65(15): 4317-34, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24821952

RESUMO

Haematococcus pluvialis cells predominantly remain in the macrozooid stage under favourable environmental conditions but are rapidly differentiated into haematocysts upon exposure to various environmental stresses. Haematocysts are characterized by massive accumulations of astaxanthin sequestered in cytosolic oil globules. Lipidomic analyses revealed that synthesis of the storage lipid triacylglycerol (TAG) was substantially stimulated under high irradiance. Simultaneously, remodelling of membrane glycerolipids occurred as a result of dramatic reductions in chloroplast membrane glycolipids but remained unchanged or declined slightly in extraplastidic membrane glycerolipids. De novo assembly of transcriptomes revealed the genomic and metabolic features of this unsequenced microalga. Comparative transcriptomic analysis showed that so-called resting cells (haematocysts) may be more active than fast-growing vegetative cells (macrozooids) regarding metabolic pathways and functions. Comparative transcriptomic analyses of astaxanthin biosynthesis suggested that the non-mevalonate pathway mediated the synthesis of isopentenyl diphosphate, as the majority of genes involved in subsequent astaxanthin biosynthesis were substantially up-regulated under high irradiance, with the genes encoding phytoene synthase, phytoene desaturase, and ß-carotene hydroxylase identified as the most prominent regulatory components. Accumulation of TAG under high irradiance was attributed to moderate up-regulation of de novo fatty acid biosynthesis at the gene level as well as to moderate elevation of the TAG assembly pathways. Additionally, inferred from transcriptomic differentiation, an increase in reactive oxygen species (ROS) scavenging activity, a decrease in ROS production, and the relaxation of over-reduction of the photosynthetic electron transport chain will work together to protect against photooxidative stress in H. pluvialis under high irradiance.


Assuntos
Clorófitas/metabolismo , Metabolismo dos Lipídeos , Estresse Oxidativo , Transcriptoma , Clorófitas/efeitos da radiação , Anotação de Sequência Molecular , Análise de Sequência de DNA , Luz Solar , Xantofilas/biossíntese
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