Bone morphogenetic proteins（BMPs） play critical roles in follicle growth and development.BMPs initiate signaling by assembling BMP receptors and activating Smads,which in turn alter expression of target genes.The me...Bone morphogenetic proteins（BMPs） play critical roles in follicle growth and development.BMPs initiate signaling by assembling BMP receptors and activating Smads,which in turn alter expression of target genes.The mechanism underlying the regulation of the expression of BMP receptors and Smads during follicle development in pigs is still unknown.By quantitative real-time PCR,the mRNA expression of BMP receptors and Smads in granulosa cells（GC） was investigated.Cells were obtained from small porcine follicles（SF;3 mm diameter） and dominant follicles（DF;6 mm diameter）;ActR1A and BMPR2 mRNA levels in DF were significantly higher（P0.05） than that in SF,whereas BMPR1B,Smad4 and Smad7 expression tended to decrease（P0.05）.The levels of BMPR1A,ActR2,Smad1,Smad5,and Smad8 mRNA did not differ between DFs and SFs.To investigate the effect of LH on BMP receptors in GC,cells obtained from porcine DFs were cultured in medium supplemented with different doses of luteinizing hormone（LH）.High doses of LH（4 IU mL-1） significantly decreased the concentration of estradiol（E2） and progesterone（P4） in medium and the expression of Cyp19a1（P450 aromatase,P450arom） and Cyp11a1（cholesterol side-chain cleavage enzyme P450,P450scc）,while significantly increased viable cell numbers and up-regulated expression of cyclin dependent kinase-4（CDK4） and cyclin D2.However,LH had no effect on the expression of BMP receptor genes.Thus,the present study indicates that the expression of members of the BMP signaling pathway in porcine GC is regulated during follicle development and the expression of BMP receptors are not regulated by LH in porcine GCs.展开更多
Exposure to a growth factor abundant milieu has remarkable regenerative and rejuvenating effects on organ diseases,tissue damage,and regeneration,including skeletal system defects and bone regeneration.Although the in...Exposure to a growth factor abundant milieu has remarkable regenerative and rejuvenating effects on organ diseases,tissue damage,and regeneration,including skeletal system defects and bone regeneration.Although the introduction of candidate growth factors into relevant fields has been reported,their regenerative effects remain unsatisfactory,mainly because of the experimental challenges with limited types of growth factors,elusive dosage adjustment,and asynchronous stem cell activation with cytokine secretion.Here,an innovative hydrogel recapitulating a growth factor-enriched microenvironment(GEM)for regenerative advantage,is reported.This sulfated hydrogel includes bone morphogenetic protein-2(BMP-2),an essential growth factor in osteogenesis,to direct mesenchymal stem cell(MSC)differentiation,stimulate cell proliferation,and improve bone formation.The semi-synthetic hydrogel,sulfonated gelatin(S-Gelatin),can amplify BMP-2 signaling in mouse MSCs by enhancing the binding between BMP-2 and BMP-2 type II receptors(BMPR2),which are located on MSC nuclei and activated by the hydrogel.Importantly,the dramatically improved cytokine secretion of MSCs throughout regeneration confirms the growth factor-acquiring potential of S-Gelatin/rhBMP-2 hydrogel,leading to the vascularization enhancement.These findings provide a new strategy to achieve an in situ GEM and accelerated bone regeneration by amplifying the regenerative capacity of rhBMP-2 and capturing endogenous growth factors.展开更多
The objective was to investigate the expression of bone morphogenetic protein （BMP） family members in the mouse uterus during the estrous cycle by real-time polymerase chain reaction （PCR） and immunohistochemistry...The objective was to investigate the expression of bone morphogenetic protein （BMP） family members in the mouse uterus during the estrous cycle by real-time polymerase chain reaction （PCR） and immunohistochemistry. Uterine samples from Swiss ICR mice were collected and dissected free of surrounding tissue. One uterine horn was snap frozen in liquid nitrogen immediately after collection and stored at -80 ℃for RNA extraction, and the other was fixed in 40 mg/ml paraformaldehyde at room temperature for immunolocalization of BMP2 protein. Real-time PCR analysis showed that the expression level of Bmp2 was significantly higher at proestrus than at estrus and metestrus （P〈0.05）. The relative abundance of Bmp4 exhibited significant fluctuations, but there were no statistically significant differences between the expression levels of Bmp2 and Bmp4 （P〉0.05）. The expression levels of Bmprla and Bmpr2 remained unchanged during estrous cycles. However, the level of Bmprlb mRNA decreased significantly at estrus （P〈0.05）, increasing subsequently at metestrus. Furthermore, the level of Bmprlb mRNA was significantly lower than those of Bmprla and Bmpr2 mRNA at the corresponding stages （P〈0.05）. All three receptor-regulated Smads （R-Smads） detected were differentially expressed in the mouse uterus and the expression levels of Smadl and Smad5 were significantly higher than that of Smad8 （P〈0.05）. In addition, the expression level of Smad4 did not change substantially throughout the estrous cycle. Immunohistochemical experiments revealed that BMP2 protein was differentially expressed and localized mainly in the uterine luminal and glandular epithelial cells throughout the estrous cycle. In conclusion, our results provide information about the variation in the mRNA levels of Bmp2 and Bmp4 and related components of the BMP signaling pathway. The data provide quantitative and useful information about the roles of endometrial BMP proposed and demonstrated by others, such as the degradation and remodeling of the endometrium.展开更多
基金supported by the National High Tech-nology Research & Development Program of China(2006AA10Z136)
文摘Bone morphogenetic proteins（BMPs） play critical roles in follicle growth and development.BMPs initiate signaling by assembling BMP receptors and activating Smads,which in turn alter expression of target genes.The mechanism underlying the regulation of the expression of BMP receptors and Smads during follicle development in pigs is still unknown.By quantitative real-time PCR,the mRNA expression of BMP receptors and Smads in granulosa cells（GC） was investigated.Cells were obtained from small porcine follicles（SF;3 mm diameter） and dominant follicles（DF;6 mm diameter）;ActR1A and BMPR2 mRNA levels in DF were significantly higher（P0.05） than that in SF,whereas BMPR1B,Smad4 and Smad7 expression tended to decrease（P0.05）.The levels of BMPR1A,ActR2,Smad1,Smad5,and Smad8 mRNA did not differ between DFs and SFs.To investigate the effect of LH on BMP receptors in GC,cells obtained from porcine DFs were cultured in medium supplemented with different doses of luteinizing hormone（LH）.High doses of LH（4 IU mL-1） significantly decreased the concentration of estradiol（E2） and progesterone（P4） in medium and the expression of Cyp19a1（P450 aromatase,P450arom） and Cyp11a1（cholesterol side-chain cleavage enzyme P450,P450scc）,while significantly increased viable cell numbers and up-regulated expression of cyclin dependent kinase-4（CDK4） and cyclin D2.However,LH had no effect on the expression of BMP receptor genes.Thus,the present study indicates that the expression of members of the BMP signaling pathway in porcine GC is regulated during follicle development and the expression of BMP receptors are not regulated by LH in porcine GCs.
基金supported by the National Natural Science Foundation of China for Innovative Research Groups(Grant No.51621002)the Fundamental Research Funds for the Central Universities,State Administration of Foreign Experts Affairs P.R.China(Grant No.B14018)+1 种基金National Natural Science Foundation of China(No.31870953)National Key R&D Program of China(2018YFE0201500).
文摘Exposure to a growth factor abundant milieu has remarkable regenerative and rejuvenating effects on organ diseases,tissue damage,and regeneration,including skeletal system defects and bone regeneration.Although the introduction of candidate growth factors into relevant fields has been reported,their regenerative effects remain unsatisfactory,mainly because of the experimental challenges with limited types of growth factors,elusive dosage adjustment,and asynchronous stem cell activation with cytokine secretion.Here,an innovative hydrogel recapitulating a growth factor-enriched microenvironment(GEM)for regenerative advantage,is reported.This sulfated hydrogel includes bone morphogenetic protein-2(BMP-2),an essential growth factor in osteogenesis,to direct mesenchymal stem cell(MSC)differentiation,stimulate cell proliferation,and improve bone formation.The semi-synthetic hydrogel,sulfonated gelatin(S-Gelatin),can amplify BMP-2 signaling in mouse MSCs by enhancing the binding between BMP-2 and BMP-2 type II receptors(BMPR2),which are located on MSC nuclei and activated by the hydrogel.Importantly,the dramatically improved cytokine secretion of MSCs throughout regeneration confirms the growth factor-acquiring potential of S-Gelatin/rhBMP-2 hydrogel,leading to the vascularization enhancement.These findings provide a new strategy to achieve an in situ GEM and accelerated bone regeneration by amplifying the regenerative capacity of rhBMP-2 and capturing endogenous growth factors.
基金Project supported by the National Natural Science Foundation of China(No.31172206)the Grant-in-Aid for Innovative Training of Doctoral Students in Jiangsu Province(No.CXLX13-287)
文摘The objective was to investigate the expression of bone morphogenetic protein （BMP） family members in the mouse uterus during the estrous cycle by real-time polymerase chain reaction （PCR） and immunohistochemistry. Uterine samples from Swiss ICR mice were collected and dissected free of surrounding tissue. One uterine horn was snap frozen in liquid nitrogen immediately after collection and stored at -80 ℃for RNA extraction, and the other was fixed in 40 mg/ml paraformaldehyde at room temperature for immunolocalization of BMP2 protein. Real-time PCR analysis showed that the expression level of Bmp2 was significantly higher at proestrus than at estrus and metestrus （P〈0.05）. The relative abundance of Bmp4 exhibited significant fluctuations, but there were no statistically significant differences between the expression levels of Bmp2 and Bmp4 （P〉0.05）. The expression levels of Bmprla and Bmpr2 remained unchanged during estrous cycles. However, the level of Bmprlb mRNA decreased significantly at estrus （P〈0.05）, increasing subsequently at metestrus. Furthermore, the level of Bmprlb mRNA was significantly lower than those of Bmprla and Bmpr2 mRNA at the corresponding stages （P〈0.05）. All three receptor-regulated Smads （R-Smads） detected were differentially expressed in the mouse uterus and the expression levels of Smadl and Smad5 were significantly higher than that of Smad8 （P〈0.05）. In addition, the expression level of Smad4 did not change substantially throughout the estrous cycle. Immunohistochemical experiments revealed that BMP2 protein was differentially expressed and localized mainly in the uterine luminal and glandular epithelial cells throughout the estrous cycle. In conclusion, our results provide information about the variation in the mRNA levels of Bmp2 and Bmp4 and related components of the BMP signaling pathway. The data provide quantitative and useful information about the roles of endometrial BMP proposed and demonstrated by others, such as the degradation and remodeling of the endometrium.
