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1.
Artif Organs ; 2020 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-33355401

RESUMO

The clinical application of most materials used to fill severe bone defects is limited owing to the insufficient ability of such materials to induce bone regeneration over a long repair period. The purpose of this study was to establish a model for the alveolar process cleft in rabbits to evaluate the effect of active bone material in bone defect repair. The active bone material used in this study is a new bone repair material composed of a heterogeneous collagen membrane implanted with modified recombinant human bone morphogenetic protein 2. This proposed active bone material can specifically bind to collagen. Twenty-four young Japanese white rabbits (JWRs) were selected and randomly divided into four groups (normal, control, material, and bone morphogenetic protein groups). The alveolar process cleft model was established by removing an equal volume bone at the left maxillary position. Blood samples were collected from the JWRs 3 and 6 months after the surgery to evaluate the biocompatibility of the active bone materials. Subsequently, the skull model was established, and the appearance was observed. Imaging methods (including X-ray examination and micro-computerized tomography scanning), tissue staining, and immunohistochemistry were employed for the evaluation. The bone collagen material and active bone material exhibited high biocompatibility. In addition, the ability of the active bone material to induce bone repair and regeneration was higher than that of the bone collagen material. The active bone material exhibited satisfactory bone regeneration performance in rabbits, indicating its potential as an active material for repairing congenital alveolar process clefts in humans.

2.
Toxicol Res (Camb) ; 9(5): 622-631, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33178422

RESUMO

Diethylhexyl phthalate (DEHP) is known as a persistent environmental pollutant. However, the possible effects of DEHP on human neural tube defects (NTDs) remain elusive. We set out to investigate the exposure of DEHP in human and explore the association of DEHP and NTDs. The level of DEHP in maternal urine was measured and analyzed by GC-MS. To further validate the results in human NTDs, chick embryos were used as animal models. Viability, reactive oxygen species (ROS) level, oxidative stress indicators and apoptosis were detected in DEHP-treated chick embryos. Our research revealed that the detection ratio of positive DEHP and its metabolites in maternal urine were observed dramatically higher in NTDs population than that in normal controls (P < 0.01, P < 0.05, respectively). Moreover, DEHP treatment (10-6 M) led to developmental toxicity in chick embryos via accelerating oxidative stress response and cell apoptosis, and changing the level of oxidative stress-related indicators. Moreover, high dose choline (100 µg/µl) could partially restrain the toxicity effects induced by DEHP. Our data collectively imply that the incidence of NTDs may closely associate with DEHP exposure, which disturbs the development of neural tubes by enhancing oxidative stress.

3.
Diabetes Metab Syndr Obes ; 13: 3009-3034, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32943895

RESUMO

Purpose: This study aimed to investigate the role of miR-21 expression in the reduction of placental function in GDM patients. Materials and Methods: qRT-PCR was used to detect the differential expression of miR-21 in the serum of gestational diabetes mellitus (GDM) and normal pregnant women, and to verify the functional target gene PPAR-α of miR-21 by double fluorescence experiments. Cellular experiments were performed to verify the effect of PPAR-α on cell function. Results: miR-21 is down-regulated in the serum and placenta of GDM patients compared to normal pregnant women. In the case of insulin resistance, miR-21-5p knockdown promoted glucose uptake, but no significant effect was found under physiological condition. Functional studies have shown that reduced PPAR-α expression can restore miR-21 knockdown-mediated cell growth and metastasis inhibition. Additionally, decreased expression of miR-21 but increased expression of -PPAR-α was observed in patients with GDM and GDM rats. Conclusion: The expression of the placental miR-21-5p, which inhibits cell growth and infiltration by up-regulating PPAR-α, is downregulated in pregnant GDM patients, which in turn may affect the placental function.

4.
Biomed Eng Online ; 19(1): 62, 2020 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-32746926

RESUMO

BACKGROUND: Alveolar cleft is a type of cleft lip and palate that seriously affects the physical and mental health of patients. In this study, a model of the alveolar cleft phenotype was established in rabbits to evaluate the effect of bone collagen particles combined with human umbilical cord mesenchymal stem cells (HUC-MSCs) on the repair of alveolar cleft bone defects. METHODS: A model of alveolar clefts in rabbits was established by removing the incisors on the left side of the upper jaw bone collagen particles combined with HUC-MSCs that were then implanted in the defect area. Blood biochemical analysis was performed 3 months after surgery. Skull tissues were harvested for gross observation, and micro-focus computerised tomography (micro-CT) analysis. Tissues were harvested for histological and immunohistochemical staining. The experiments were repeated 6 months after surgery. RESULTS: Bone collagen particles and HUC-MSCs showed good biocompatibility. Bone collagen particles combined with HUC-MSCs were markedly better at inducing bone repair and regeneration than bone collagen particles alone. CONCLUSIONS: Combining HUC-MSCs with bone collagen particles provides a simple, rapid and suitable method to fill a bone defect site and treat of alveolar cleft bone defects.

5.
Toxicol Res (Camb) ; 9(3): 222-229, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32670553

RESUMO

Benzyl butyl phthalate (BBP) is a persistent environmental pollutant. BBP exposure and the possible effects on human neural tube defects (NTDs) remain elusive. In this study, we found that the detection ratio of positive BBP and its metabolites in maternal urine was obviously higher in NTDs' population than that in normal controls by GC-MS (P < 0.01, P < 0.05, respectively). Animal experiments showed that BBP treatment induced developmental toxicity in chick embryo by enhancing the levels of oxidative stress and cell apoptosis (P < 0.01). More interestingly, the supplement of high-dose choline (CHO, 10 5  µg/mL) could partially restore the teratogenic effects of BBP by inhibiting the occurrence of oxidative stress. Our data collectively suggest that BBP exposure may disturb neural tube development by strengthening oxidative stress. CHO can partially restore the toxicity effects of BBP. This study may provide new insight for NTD prevention.

6.
Artigo em Inglês | MEDLINE | ID: mdl-32296392

RESUMO

Gestational diabetes mellitus (GDM) is a disease that changes the function of microvascular of placenta. MicroRNA (miRNA) expression in placenta may contribute to the pathogenesis of GDM. Here, we evaluate the role and function of miR-29b in the development of GDM. This study discovered that miR-29b expression was lower in placentas derived from patients with GDM than that in control placentas. MiR-29b over-expression inhibited cell growth and migration, and miR-29b knockdown promoted cell migration. Then we predicted and confirmed that HIF3A was a direct target of miR-29b with two specific binding sites at the recognition sequences of miR-29b in 3'-UTR of HIF3A mRNA, which was negatively correlated with miR-29b expression level. The up-regulation of HIF3A partially antagonized the inhibitory effect of miR-29b over-expression on cell growth and migration. The enhancement of cell migration induced by miR-29b knockdown was attenuated by down-regulating HIF3A. These results imply that down-regulation of miR-29b may be related with the development of GDM partially via increasing the expression of HIF3A, which may provide a new insight for the mechanism of GDM.

7.
Reprod Sci ; 27(1): 152-162, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-32046397

RESUMO

Recurrent spontaneous abortion (RSA) is a common health problem that affects 1-5% of women in reproductive age. Plenty of studies have indicated that microRNAs (miRNAs) are involved in the occurrence of miscarriage. MiR-93 has a wide range of functions in mammalian tissues and plays an important role in many diseases especially for cancers. However, it remains unknown whether miR-93 is associated with human RSA. In this report, clinical samples revealed that miR-93 expression was significantly elevated in the villi tissues of RSA patients. Upregulation of miR-93 inhibited human trophoblast cells HTR-8/SVneo cell proliferation, migration, and invasiveness, but promoted cell apoptosis in vitro. Conversely, the downregulation of miR-93 reversed these effects. Bcl-2 like protein 2 (BCL2L2), a potential target gene of miR-93, was inversely correlated with miR-93 expression in the villi of clinical samples. Furthermore, the luciferase reporter system demonstrated that miR-93 directly downregulated the expression of BCL2L2 by binding a specific sequence of its 3'-untranslated region (3'UTR). Collectively, these data strongly suggest that miR-93 regulates trophoblast cell proliferation, migration, invasive, and apoptosis by targeting BCL2L2 expression and is involved in the pathogenesis of RSA.


Assuntos
Aborto Habitual/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Apoptose/fisiologia , Proliferação de Células/fisiologia , MicroRNAs/metabolismo , Trofoblastos/metabolismo , Adulto , Linhagem Celular , Feminino , Humanos , Gravidez , Trofoblastos/citologia , Regulação para Cima
8.
Reproduction ; 159(5): 525-537, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32045359

RESUMO

Missed abortion (MA) is a common disease in obstetrics and gynecology. More and more studies have focused on the relationship between miRNAs and pregnancy maintenance and its related diseases. The aim of this article is to explore the relationship between miRNA and MA. The expression of miR-98 were detected by in situ hybridization and real-time PCR. Cell proliferation, activity and migration were measured via Edu, MTT, and transwell assays. The target genes of miR-98 are identified by dual-luciferase activity assay. And the expression levels of target genes were determined by Western blot, real-time PCR and immunohistochemistry. miR-98 was significantly up-regulated in placental villi from over 35 years old MA patients compared with the age-matched normal pregnant women. Up-regulation of miR-98 suppressed the proliferation, activity and migration of the human trophoblast HTR-8/SVneo cell in vitro. miR-98 could bind to GDF6 and FAPP2 mRNA 3'-UTR and negatively regulate their expression. The downregulation of miR-98 promoted cell proliferation, then knockdown of GDF6 or FAPP2 inhibited miR-98-mediated cell proliferation. GDF6 and FAPP2 expression in the placental villi from MA patients were decreased compared to normal placental tissues. The expression of miR-98 in MA had an opposite relationship with the expression of GDF6 and FAPP2. Overexpression of miR-98 is associated with the occurrence of MA. miR-98 prevents proliferation, viability and migration of trophoblast cells partially through targeting GDF6 and FAPP2.

9.
Environ Sci Pollut Res Int ; 26(29): 29763-29779, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31407264

RESUMO

Dibutyl phthalate (DBP), a persistent environmental pollutant, can induce neural tube abnormal development in animals. The possible effects of DBP exposure on human neural tube defects (NTDs) remain elusive. In this study, the distribution of DBP in the body fluid of human NTDs was detected by GC-MS. Then, chick embryos were used to investigate the effects of DBP on early embryonic development. Oxidative stress indicators in chick embryos and the body fluid of human NTDs were detected by ELISA. The cell apoptosis and total reactive oxygen species (ROS) level in chick embryos were detected by whole-mount TUNEL and oxidized DCFDA, respectively. The study found that the detection ratio of positive DBP and its metabolites in maternal urine was higher in the NTD population than that in normal controls. 8-hydroxy-2 deoxyguanosine (8-OHDG) and malondialdehyde (MDA) were evidently upregulated and superoxide dismutase (SOD) was observably downregulated in amniotic fluid and urine. Animal experiments indicated that DBP treatment induced developmental toxicity in chick embryos by enhancing the levels of oxidative stress and cell apoptosis. MDA was increased and SOD was decreased in DBP-treated embryos. Interestingly, the supplement of high-dose choline (100 µg/µL), not folic acid, could partially restore the teratogenic effects of DBP. Our data collectively suggest that the incidence of NTDs is closely associated with DBP exposure. This study may provide new insight for NTD prevention.


Assuntos
Galinhas/metabolismo , Colina/metabolismo , Dibutilftalato/toxicidade , Desenvolvimento Embrionário/efeitos dos fármacos , Poluentes Ambientais/toxicidade , Defeitos do Tubo Neural/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Líquidos Corporais/metabolismo , Embrião de Galinha , Galinhas/crescimento & desenvolvimento , Dibutilftalato/urina , Poluentes Ambientais/urina , Feminino , Ácido Fólico/metabolismo , Humanos , Exposição Materna/efeitos adversos , Teratogênese/efeitos dos fármacos
10.
Int J Pediatr Otorhinolaryngol ; 124: 164-172, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31200319

RESUMO

OBJECTIVE: Cleft palate is one of the most common craniofacial birth defects in the maxillofacial region. There is an urgent need in tissue regeneration research to establish animal models that faithfully mimic human diseases. Here, we compared three surgical models of bone tissue defects in cleft palate in rabbits in order to screen for the biomaterials that induced optimal bone regeneration. DESIGN: Rabbits were used to establish the models of hard palate cleft, alveolar cleft, and alveolar process cleft. Eight weeks following surgery, bone tissue self-healing capacity was estimated by macroscopic appearance and calculating the area of defective bone tissue. The dimensions of the upper jaw in left and right sides were measured at zero and eight weeks. RESULTS: Bone defects in three types of cleft palate models were made at the positions of the hard palate, alveoli and alveolar process. After 8 weeks, when the hard palate was partially excised, it underwent self-healing. When the hard palate was completely excised, it underwent partial self-healing. However, in the models of alveolar cleft and alveolar process cleft, there was no significant self-healing in the bone tissues. The dimensions of the upper jaw in left and right sides were no significant differences in three types of cleft palate models. CONCLUSIONS: Bone defects in the alveolar and alveolar process clefts exhibit a diminished capability for self-healing. This study may provide valuable information for the screening of materials that induce bone regeneration.


Assuntos
Fissura Palatina/cirurgia , Modelos Animais de Doenças , Modelos Anatômicos , Processo Alveolar/cirurgia , Animais , Regeneração Óssea , Fissura Palatina/etiologia , Feminino , Masculino , Maxila/cirurgia , Palato Duro/cirurgia , Coelhos , Cicatrização
11.
Exp Cell Res ; 374(1): 210-220, 2019 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-30503865

RESUMO

MicroRNAs (miRNAs) regulate diverse cellular processes such as cell differentiation, proliferation and apoptosis. Mutation in miRNAs results in various pathological conditions such as inflammation, viral infections, neurodegeneration, and autoimmunity. We have evaluated the association of miR-423 rs6505162C>A and rs8067576 A>T among patients with recurrent pregnancy loss (RPL) and controls from North China. Our study found that one SNP rs6505162C>A in miR-423 coding region was associated with the increase risk of humanunexplained RPL (URPL), but no differences were found in another SNP rs8067576 A>T. However, in two-locus haplotype analysis, miR-423-CC/TT haplotype was associated with an increased risk of URPL. The level of mature miR-423 was obviously down-regulated in cells transfected with miR-423-CC/TT haplotype. miR-423-CC/TT haplotype inhibited HTR-8/SVneo cells proliferation and migration and promoted cells apoptosis. Further experiments identified that mesoderm development candidate 1 (MESDC1) was a functionally relevant target of miR-423, and its expression was reversely regulated by miR-423. More importantly, dual-luciferase assay indicated miR-423-CC/TT haplotype decreasing miR-423 expression, could up-regulate MESDC1 expression. Collectively, our data suggest that miR-423-CC/TT haplotype in pre-miR-423 may aggravate the risk of developing URPL by influencing the level of mature miR-423 and its target gene MESDC1.


Assuntos
Aborto Habitual/genética , Grupo com Ancestrais do Continente Asiático/genética , Grupos Étnicos/genética , Estudos de Associação Genética , Predisposição Genética para Doença , Haplótipos/genética , MicroRNAs/genética , Polimorfismo de Nucleotídeo Único/genética , Regiões 3' não Traduzidas/genética , Apoptose/genética , Sequência de Bases , Linhagem Celular , Movimento Celular/genética , Proliferação de Células , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , MicroRNAs/química , MicroRNAs/metabolismo , Mifepristona/farmacologia , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Conformação de Ácido Nucleico , Gravidez , Progesterona/farmacologia , Fatores de Risco
12.
Stem Cell Res Ther ; 9(1): 36, 2018 02 13.
Artigo em Inglês | MEDLINE | ID: mdl-29433563

RESUMO

BACKGROUND: Repair deficiency after endometrial injury is an important reason for intra-uterine adhesions, amenorrhea, and infertility in females. Bone marrow-derived mesenchymal stem cell (BMSC) transplantation is effective in repairing the damaged endometrium. However, the possibility of using umbilical cord-derived MSCs (UC-MSCs) to treat endometrial injury is rarely reported. METHODS: Ethanol (95%) was injected into rat uterus to establish a model of endometrial injury. UC-MSCs were injected through the tail vein, either as a single, twice, or thrice administration. Functional restoration of the uterus was assessed by testing embryo implantation rates. Endometrial morphological alteration was observed by hematoxylin and eosin staining. Endometrial fibrosis, markers of epithelial and stromal cells of endometrium, cell proliferation and angiogenesis, and inflammatory factors were detected using immunohistochemistry, Western blotting, and quantitative reverse-transcription polymerase chain reaction. RESULTS: Endometrial morphology and embryo implantation rates were significantly improved on day 8 of transplantation among single-, twice-, or thrice-administered rats. Moreover, UC-MSCs could alleviate fibrosis in general, and reduced the expression of fibrosis markers, α-smooth muscle actin (α-SMA) and transforming growth factor (TGF)-ß. The cell proliferation marker Ki-67 had a positive expression in the injured endometrium after UC-MSC transplantation. The endometrial stromal marker vimentin and epithelial marker cytokeratin-19 (CK-19) expressions were visibly increased. The expression of vascular markers CD31, vascular endothelial growth factor (VEGF)A, and matrix metalloprotein (MMP)9 was generally upregulated. Proinflammatory factors interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and interleukin (IL)-2 were significantly downregulated in the rats administered UC-MSCs twice and thrice. CONCLUSIONS: UC-MSC transplantation contributed to the repair of endometrial injury and restoration of fertility, likely through the suppression of excessive fibrosis and inflammation, and enhancement of endometrial cell proliferation and vascular remodeling.


Assuntos
Endométrio , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/metabolismo , Cordão Umbilical/metabolismo , Doenças Uterinas , Animais , Antígenos de Diferenciação/metabolismo , Doença Crônica , Endométrio/lesões , Endométrio/metabolismo , Endométrio/patologia , Feminino , Xenoenxertos , Humanos , Células-Tronco Mesenquimais/patologia , Ratos , Ratos Sprague-Dawley , Cordão Umbilical/patologia , Doenças Uterinas/metabolismo , Doenças Uterinas/patologia , Doenças Uterinas/terapia
13.
Sci Rep ; 6: 32268, 2016 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-27573367

RESUMO

MiR-98 expression was up-regulated in kidney in response to early diabetic nephropathy in mouse and down-regulated in muscle in type 2 diabetes in human. However, the expression prolife and functional role of miR-98 in human gestational diabetes mellitus (GDM) remained unclear. Here, we investigated its expression and function in placental tissues from GDM patients and the possible molecular mechanisms. The results showed that miR-98 was up-regulated in placentas from GDM patients compared with normal placentas. MiR-98 over-expression increased global DNA methylational level and miR-98 knockdown reduced global DNA methylational level. Further investigation revealed that miR-98 could inhibit Mecp2 expression by binding the 3'-untranslated region (UTR) of methyl CpG binding protein 2 (Mecp2), and then led to the expression dysregulation of canonical transient receptor potential 3 (Trpc3), a glucose uptake related gene. More importantly, in vivo analysis found that the expression level of Mecp2 and Trpc3 in placental tissues from GDM patients, relative to the increase of miR-98, was diminished, especially for GDM patients over the age of 35 years. Collectively, up-regulation of miR-98 in the placental tissues of human GDM is linked to the global DNA methylation via targeting Mecp2, which may imply a novel regulatory mechanism in GDM.


Assuntos
Diabetes Gestacional/genética , Regulação da Expressão Gênica , Proteína 2 de Ligação a Metil-CpG/genética , MicroRNAs/genética , Regulação para Cima , Regiões 3' não Traduzidas/genética , Adulto , Metilação de DNA , Feminino , Humanos , Recém-Nascido , Placenta/metabolismo , Gravidez , Canais de Cátion TRPC/genética
14.
Oncotarget ; 7(7): 8208-22, 2016 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-26824181

RESUMO

MicroRNA-10a (miR-10a) has a wide range of functions in nearly all mammalian tissues and is involved in the occurrence of many diseases. However, it remains unknown whether miR-10a is associated with human recurrent spontaneous abortion (RSA). In this study, we found that rs3809783 A > T in miR-10a coding region was significantly associated with the increase of the risk of human unexplained RSA (URSA) acquisition in a Han-Chinese population. The T allele of rs3809783 hindered the production of mature miR-10a. A to T substitution in miR-10a rs3809783 repressed cell proliferation and migratory capacity. Further investigation discovered that Bcl-2-interacting mediator (Bim) was the functional target of miR-10a and inversely regulated Bim expression. Dual-luciferase assay indicated that A allele in miR-10a rs3809783 could more effectively suppress Bim expression than T allele. In addition, A to T substitution in miR-10a rs3809783 attenuated the sensibility of cells to progesterone and its antagonist mifepristone. Collectively, our data suggest that rs3809783 A > T in pri-miR-10a may be conductive to the genetic predisposition to RSA by disrupting the production of mature miR-10a and reinforcing the expression of Bim.


Assuntos
Aborto Espontâneo/epidemiologia , Aborto Espontâneo/genética , Proteína 11 Semelhante a Bcl-2/metabolismo , Biomarcadores/análise , MicroRNAs/genética , Polimorfismo de Nucleotídeo Único/genética , Apoptose , Proteína 11 Semelhante a Bcl-2/genética , Western Blotting , Movimento Celular , Proliferação de Células , Células Cultivadas , China/epidemiologia , Feminino , Citometria de Fluxo , Predisposição Genética para Doença , Humanos , Técnicas Imunoenzimáticas , Gravidez , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Recidiva , Reação em Cadeia da Polimerase Via Transcriptase Reversa
15.
Reproduction ; 150(1): 65-76, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25926693

RESUMO

Although the relationship between polymorphisms in microRNAs (miRNAs) and recurrent pregnancy loss (RPL) has been studied, there is very little data available in the literature. In the present study, we scanned 55 potentially functional polymorphisms in the miRNA coding region in Chinese women with unexplained RPL (URPL; no. 2011-10). The rs6505162 C>A in the MIR423 coding region was found to be significantly associated with the occurrence of human URPL. The rare A allele contributed to an increase in the expression of mature MIR423. C to A substitution in the polymorphism rs6505162 in pre-MIR423 repressed cell proliferation and migratory capacity. Further investigations showed that MIR423 could inversely regulate the expression of proliferation-associated 2 group 4 (PA2G4) by binding the 3'-UTR of PA2G4. Dual-luciferase assay indicated that the A allele in the polymorphism rs6505162 could more effectively suppress the expression of PA2G4 than the C allele could. Collectively, the present data suggest that rs6505162 C>A in pre-MIR423 may contribute to the genetic predisposition to RPL by disrupting the production of mature MIR423 and its target gene, which consequently interferes with MIR423 functioning.


Assuntos
Aborto Habitual/genética , MicroRNAs/genética , Polimorfismo de Nucleotídeo Único , Alelos , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Frequência do Gene , Predisposição Genética para Doença , Humanos , Gravidez
16.
Biochim Biophys Acta ; 1850(4): 708-21, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25486623

RESUMO

BACKGROUND: To study the role of miR-143 during embryo implantation in rat. METHODS: MiR-143 expression in rat early pregnancy was detected by Northern blot. The relation between miR-143 and Lifr predicted and confirmed by bioinformatics method, dual-luciferase activity assay, Western blot and immunohistochemistry. The role of miR-143 was detected by MTS, Edu and ranswell chamber assays. RESULTS: The expression level of miR-143 on gestation day 5-8 (g.d. 5-8) was higher than on g.d. 3-4 in uteri of pregnant rat. MiR-143 was mainly localized in the superficial stroma/primary decidual zone, luminal and glandular epithelia. The expression of miR-143 was not significantly influenced by pseudopregnancy, but the activation of delayed implantation and experimentally induced decidualization significantly promoted miR-143 expression. Over-expression of miR-143 in human endometrial stromal cells (ESCs) inhibited cell proliferation, migration and invasion. Knockdown of miR-143 promoted cell proliferation and invasion. The results of recombinant luciferase reporters showed that miR-143 could bind to the 3¢-untranslated region (UTR) of leukemia inhibitory factor receptor (Lifr) to inhibit Lifr translation. CONCLUSIONS: Uterine miR-143 may be involved in the successful pregnancy, especially during the process of blastocyst implantation through regulating Lifr. GENERAL SIGNIFICANCE: This study may have the potential to provide new insights into the understanding of miR-143 function during embryo implantation.


Assuntos
Implantação do Embrião , MicroRNAs/fisiologia , Animais , Movimento Celular , Proliferação de Células , Estradiol/farmacologia , Feminino , Humanos , MicroRNAs/análise , Gravidez , Ratos , Ratos Sprague-Dawley , Receptores de OSM-LIF/genética , Útero/metabolismo
17.
FEBS Lett ; 588(23): 4438-47, 2014 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-25448984

RESUMO

MiR-185 expression has been associated with many cancers. However, the roles of miR-185 in human breast cancer remain elusive. Here, we found that miR-185 expression was decreased in human breast cancer tissues compared with healthy tissue controls. Up-regulation of miR-185 inhibited breast cancer cell proliferation and invasion and vice versa. MiR-185 was shown to bind to the 3'-untranslated region (UTR) of vascular endothelial growth factor a (Vegfa), and a significant inverse correlation was found between miR-185 and Vegfa. Vegfa overexpression partially restored the inhibition of cell proliferation and invasion that was induced by miR-185, and vice versa. Additionally, Vegfa expression was found to be high in human breast cancer tissues. Thus, miR-185-mediated Vegfa targeting may be involved in breast cancer formation.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Carcinogênese/genética , MicroRNAs/genética , Fator A de Crescimento do Endotélio Vascular/genética , Apoptose/genética , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células , Sobrevivência Celular/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Invasividade Neoplásica , Metástase Neoplásica
18.
PLoS One ; 9(12): e114781, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25479352

RESUMO

MicroRNAs (miRNAs) are short non-coding RNAs which modulate gene expression by binding to complementary segments present in the 3'UTR of the mRNAs of protein coding genes. MiRNAs play very important roles in maintaining normal human body physiology conditions, meanwhile, abnormal miRNA expressions have been found related to many human diseases spanning from psychiatric disorders to malignant cancers. Recently, emerging reports have indicated that disturbed miRNAs expression contributed to the pathogenesis of recurrent pregnancy loss (RPL). In this study, we identified a new mutation site (+29A>G, position relative to pre-miR-125a) by scanning pri-miR-125a coding region in 389 Chinese Han RPL patients. This site was co-existed with two polymorphisms (rs12976445 and rs41275794) in patients heterogeneously and changed the predicted secondary structures of pri-miR-125a. Subsequent in vitro analysis indicated that the A>G mutation reduced mature miR-125a expression, and further led to less efficient inhibition of verified target genes. Functional analysis showed that mutant pri-mir-125a can enhance endometrial stromal cells (ESCs) invasive capacity and increase the sensitivity of ESCs cells to mifepristone. Moreover, we further analyzed the possible molecular mechanism by RIP-chip assay and found that mutant pri-mir-125a disturbed the expression of miR-125a targetome, the functions of which includes embryonic development, cell proliferation, migration and invasion. These data suggest that A>G mutation in pri-miR-125a coding region contributes to the genetic predisposition to RPL by disordering the production of miR-125a, which consequently meddled in gene regulatory network between mir-125a and mRNA.


Assuntos
Aborto Habitual/genética , MicroRNAs/genética , Mutação , Regiões 3' não Traduzidas , Grupo com Ancestrais do Continente Asiático/genética , Estudos de Casos e Controles , Células Cultivadas , Endométrio/citologia , Feminino , Regulação da Expressão Gênica , Predisposição Genética para Doença , Haplótipos , Humanos , MicroRNAs/química , Mifepristona/farmacologia , Polimorfismo de Nucleotídeo Único , Gravidez , Estabilidade de RNA , Células Estromais/efeitos dos fármacos , Células Estromais/patologia
19.
PLoS One ; 9(8): e103695, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25084349

RESUMO

MicroRNAs (miRNAs) are small, noncoding RNA molecules that regulate post-transcriptional gene expression by base pairing with partially complementary sequences within target messenger RNAs (mRNAs). Although the target genes and the precise biological functions of individual miRNAs remain largely unknown, miRNAs have been implicated in diverse biological processes, including both normal and pathological states. As a single stranded mRNA can be directly targeted by multiple miRNAs, and as the target sites may exist in the 3'-untranslated region (UTR), 5'-UTR, or the coding regions, it is essential to develop an effective method to identify the full-scale miRNA regulatory pattern of each particular gene. In this study, we employed a biochemical approach to identify the miRNA profiles that regulate the expression of embryonic ectoderm development (EED) protein by using anti-PABPC1 ribonucleoprotein (RNP) co-immunoprecipitation (Co-IP). The full length EED mRNA was subcloned into an expression vector and transiently transfected into a Flag-PABPC1 stable expression cell line. Subsequent to cross-linking and an anti-Flag Co-IP, the miRNAs that directly targeted EED were identified. We found that the best time point to distinguish the positive miRNAs from the background was 18 hours after the plasmid transfection. As expected, the miRNAs that directly target EED were found to interact with EED mRNA through the miRNA-induced silencing complex (miRISC). Meanwhile, the EED mRNA was bound by Flag-PABPC1. This method depends on the integrity of the miRISC complex and achieves greater efficiency when ultraviolet irradiation is used for the process of cross-linking. By using anti-PABPC1 RIP, we identified EED to be a new target gene of miR-16; a finding further confirmed using a dual-luciferase assay. In summary, our data indicate that anti-PABPC1 RIP is a validated and direct biochemical method to provide data about specific miRNA-mRNA interactions, as well as global miRNA patterns regulating the mRNAs.


Assuntos
Regiões 3' não Traduzidas/genética , MicroRNAs/genética , Proteína I de Ligação a Poli(A)/metabolismo , Complexo Repressor Polycomb 2/genética , RNA Mensageiro/genética , Ribonucleoproteínas/metabolismo , Células HEK293 , Humanos , Immunoblotting , Imunoprecipitação , Proteína I de Ligação a Poli(A)/genética , Reação em Cadeia da Polimerase em Tempo Real , Ribonucleoproteínas/genética
20.
FEBS J ; 281(7): 1872-91, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24528955

RESUMO

Embryo implantation is a complex initial step in establishment of a successful pregnancy. Many mRNAs have been shown to be differentially expressed in the rat uterus during embryo implantation. However, the expression profiles of microRNAs (miRNAs), a key post-transcriptional regulator of gene expression, in the rat uterus between the pre-receptive and receptive phases are still unknown. Here, an miRNA microarray was used to examine differential expression of miRNAs in the rat uterus between the pre-receptive and receptive phases. Twenty-eight miRNAs were up-regulated and 29 miRNAs were down-regulated at least twofold during the receptive phase in rat uterus; these results were confirmed by Northern blotting. miR-29a was only highly expressed in rat uterus during the implantation period, and activation of delayed implantation and artificial decidualization enhanced the miR-29a level. Further investigation revealed that both the pro-apoptotic factor genes Bak1 and Bmf and the anti-apoptotic factor gene Bcl-w are targets of miR-29a. There was weak binding between miR-29a and the 3' UTR of the anti-apoptotic factor gene Mcl1. Over-expression of miR-29a inhibited the late apoptosis of endometrial stromal cells, which may be due to the stronger binding capacity between miR-29a and the 3' UTR of pro-apoptotic factors than that between miR-29a and the 3' UTR of anti-apoptotic factors. Collectively, miR-29a plays an important role during embryo implantation by regulating both pro-apoptotic and anti-apoptotic factors. miR-29a may predominantly bind pro-apoptotic factors, leading to inhibition of cell apoptosis.


Assuntos
Implantação do Embrião , Endométrio/metabolismo , MicroRNAs/genética , Regiões 3' não Traduzidas , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Apoptose , Feminino , MicroRNAs/metabolismo , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Gravidez , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Ratos Sprague-Dawley , Proteína Killer-Antagonista Homóloga a bcl-2/genética , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo
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