Deletion or mutation of dentin matrix protein 1 （DMP1） leads to hypophosphatemic rickets and defects within the dentin. However, it is largely unknown if this pathological change is a direct role of DMP1 or an indir...Deletion or mutation of dentin matrix protein 1 （DMP1） leads to hypophosphatemic rickets and defects within the dentin. However, it is largely unknown if this pathological change is a direct role of DMP1 or an indirect role of phosphate （Pi） or both. It has also been previously shown that Klotho-deficient mice, which displayed a high Pi level due to a failure of Pi excretion, causes mild defects in the dentinal structure. This study was to address the distinct roles of DMP1 and Pi homeostasis in cell differentiation, apoptosis and mineralization of dentin and enamel. Our working hypothesis was that a stable Pi homeostasis is critical for postnatal tooth formation, and that DMP1 has an antiapoptotic role in both amelogenesis and dentinogenesis. To test this hypothesis, Dmpl-null （Dmpl-/-）, Klotho-deficient （kl/kl）, Dmpl/Klotho-double-deficient （Dmpl-/-/kl/kl） and wild-type （WT） mice were killed at the age of 6 weeks. Combinations of X-ray, microcomputed tomography （I^CT）, scanning electron microscopy （SEM）, histology, apoptosis and immunohistochemical methods were used for characterization of dentin, enamel and pulp structures in these mutant mice. Our results showed that Dmpl-/- （a low Pi level） or kl/kl（a high Pi level） mice displayed mild dentin defects such as thin dentin and a reduction of dentin tubules. Neither deficient mouse line exhibited any apparent changes in enamel or pulp structure. However, the double-deficient mice （a high Pi level） displayed severe defects in dentin and enamel structures, including loss of dentinal tubules and enamel prisms, as well as unexpected ectopic ossification within the pulp root canal. TUNEL assay showed a sharp increase in apoptotic cells in ameloblasts and odontoblasts. Based on the above findings, we conclude that DMP1 has a protective role for odontoblasts and ameloblasts in a pro-apoptotic environment （a high Pi level）.展开更多
It is still a huge challenge for bone regenerative biomaterial to balance its mechanical,biological and biodegradable properties.In the present study,a new composite material including treated dentin matrix(TDM)andα-...It is still a huge challenge for bone regenerative biomaterial to balance its mechanical,biological and biodegradable properties.In the present study,a new composite material including treated dentin matrix(TDM)andα-calcium sulphate hemihydrate(α-CSH)was prepared.The optimal composition ratio between TDM andα-CSH was explored.The results indicate that both components were physically mixed and structurally stable.Its compressive strength reaches up to 5.027±0.035 MPa for 50%TDM/α-CSH group,similar to human cancellous bone tissues.Biological experiments results show that TDM/α-CSH composite exhibits excellent biocompatibility and the expression of osteogenic related genes and proteins(ALP,RUNX2,OPN)is significantly increased.In vivo experiments suggest that the addition of TDM for each group(10%,30%,50%)effectively promotes cell proliferation and osteomalacia.In addition,50%of the TDM/α-CSH combination displays optimal osteoconductivity.The novel TDM/α-CSH composite is a good candidate for certain applications in bone tissue engineering.展开更多
The blood-brain barrier （BBB） is a tight boundary formed between endothelial cells and astrocytes, which separates and protects brain from most pathogens as well as neural toxins in circulation. However, detailed mo...The blood-brain barrier （BBB） is a tight boundary formed between endothelial cells and astrocytes, which separates and protects brain from most pathogens as well as neural toxins in circulation. However, detailed molecular players involved in formation of BBB are not completely known. Dentin matrix protein I （DMP1）-proteoglycan （PG）, which is known to be involved in mineralization of bones and dentin, is also expressed in soft tissues including brain with unknown functions. In the present study, we reported that DMPI-PG was expressed in brain astrocytes and enriched in BBB units. The only glycosylation site of DMP1 is serine89 （S89） in the N-terminal domain of the protein in mouse. Mutant mice with DMP1 point mutations changing S89 to glycine （S89G）, which completely eradicated glycosylation of the protein, demonstrated severe BBB disruption. Another breed of DMP1 mutant mice, which lacked the C-terminal domain of DMP1, manifested normal BBB function. The polarity of S89G-DMP1 astrocytes was disrupted and cell-cell adhesion was loosened. Through a battery of analyses, we found that DMP1 glycosylation was critically required for astrocyte maturation both in vitro and in vivo. S89G-DMP1 mutant astrocytes failed to express aquaporin 4 and had reduced laminin and ZO1 expression, which resulted in disruption of BBB. Interestingly, overexpression of wild-type DMP1-PG in mouse brain driven by the nestin promoter elevated laminin and ZO1 expression beyond wild type levels and could effectively resisted intravenous mannitol-induced BBB reversible opening. Taken together, our study not only revealed a novel element, i.e., DMP1-PG, that reg- ulated BBB formation, but also assigned a new function to DMP1-PG.展开更多
Treated dentin matrix(TDM)is an ideal scaffold material containing multiple extracellular matrix factors.The canonical Wnt signaling pathway is necessary for tooth regeneration.Thus,this study investigated whether the...Treated dentin matrix(TDM)is an ideal scaffold material containing multiple extracellular matrix factors.The canonical Wnt signaling pathway is necessary for tooth regeneration.Thus,this study investigated whether the TDM can promote the odontogenic differentiation of human dental pulp stem cells(hDPSCs)and determined the potential role of Wnt/β-catenin signaling in this process.Different concentrations of TDM promoted the dental differentiation of the hDPSCs and meanwhile,the expression of GSK3βwas decreased.Of note,the expression of the Wnt/β-catenin pathway-related genes changed significantly in the context of TDM induction,as per RNA sequencing(RNA seq)data.In addition,the experiment showed that new dentin was visible in rat mandible cultured with TDM,and the thickness was significantly thicker than that of the control group.In addition,immunohistochemical staining showed lower GSK3βexpression in new dentin.Consistently,the GSK3βknockdown hDPSCs performed enhanced odotogenesis compared with the control groups.However,GSK3βoverexpressing could decrease odotogenesis of TDM-induced hDPSCs.These results were confirmed in immunodeficient mice and Wistar rats.These suggest that TDM promotes odontogenic differentiation of hDPSCs by directly targeting GSK3βand activating the canonical Wnt/β-catenin signaling pathway and provide a theoretical basis for tooth regeneration engineering.展开更多
Background： Ankylosing spondylitis （AS） is the most common rheumatic condition that is slowly progressive and predominantly affects adolescents. Pathological bone formation associated with AS is an important cause ...Background： Ankylosing spondylitis （AS） is the most common rheumatic condition that is slowly progressive and predominantly affects adolescents. Pathological bone formation associated with AS is an important cause of disability. The aim of the study was to investigate the possible involvement of the genes related to endochondral ossification and ectopia ossification in genetic susceptibility to AS in a Chinese Han population. Methods： Sixty-eight single nucleotide polymorphisms （SNPs） from 13 genes were genotyped in discovery cohorts including 300 AS patients and 180 healthy controls. The rs10019009 in dentin matrix protein 1 （DMP1） gene shown as association with AS after multiple testing corrections in discovery cohorts was replicated in a validation independent cohort of 620 AS patients and 683 healthy controls. The rs 10019009 was assessed with bioin fomlatics including phylogenetic context, F-SNP and FastSNP functional predictions, secondary structure prediction, and molecular modeling. We performed a functional analysis of rs10019009 via reverse transcription-polymerase chain reaction, alkaline phosphatase （ALP） activity in human osteosarcoma U2OS cells. Results： Interestingly, the SNP rs10019009 was associated with AS in both the discovery cohort （P = 0.0012） and validation cohort （P - 0.0349）, as well as overall （P = 0.0004） in genetic case-control association analysis. After a multivariate logistic regression analysis, the effect of this genetic variant was observed to be independent of linkage disequilibrium. Via bioinformatics analysis, it was found that the amino acid change of the rs 10019009 led to changes of SNP function, secondary structure, tertiary confomlation, and splice mode. Finally, functional analysis ofrsl0019009 in U2OS cells demonstrated that the risk T allele of the rsl0019009 increased enzymatic activity of ALP, compared to that of the nonrisk allele （P = 0.0080）. Conclusions： These results suggested that the DMP1 gene seems to be involved in genetic predisposition to AS, which may contribute to the ectopic mineralization or ossification in AS. In addition, DMP1 gene may be a promising intervention target for AS in the future.展开更多
The presence of matrix metalloproteinase-2 （MMP-2） in dentin has been reported, but its distribution and activity level in mature human coronal dentin are not well understood. The purpose of this study was to determ...The presence of matrix metalloproteinase-2 （MMP-2） in dentin has been reported, but its distribution and activity level in mature human coronal dentin are not well understood. The purpose of this study was to determine the MMP-2 distribution and relative activity in demineralized dentin. Crowns of twenty eight human molars were sectioned into inner （ID）, middle （MD）, and outer dentin （OD） regions and demineralized. MMP-2 was extracted with 0.33 mol·L-1 EDTA/2 mol·L-1 guanidine-HCl, pH 7.4, and MMP-2 concentration was estimated with enzyme-linked immunoabsorbant assay （ELISA）. Further characterization was accomplished by Western blotting analysis and gelatin zymography. The mean concentrations of MMP-2 per mg dentin protein in the dentin regions were significantly different （P=0.043）： 0.9 ng （ID）, 0.4 ng （MD）, and 2.2 ng （OD）, respectively. The pattern of MMP-2 concentration was OD〉ID〉MD. Western blotting analysis detected -66 and -72 kDa immunopositive proteins corresponding to pro- and mature MMP-2, respectively, in the ID and MD, and a -66 kDa protein in the OD. Gelatinolytic activity consistent with MMP-2 was detected in all regions. Interestingly, the pattern of levels of Western blot immunodetection and gelatinolytic activity was MD〉ID〉OD. The eoneentration of MMP-2 in human coronal dentin was highest in the region of dentin that contains the dentinoenamel junction and least in the middle region of dentin. However, levels of Western blot immunodetection and gelatinolytic activity did not correlate with the estimated regional concentrations of MMP-2, potentially indicating region specific protein interactions.展开更多
多种生物因子参与着慢性肾脏病矿物质和骨代谢异常(chronic kidney disease–mineral and bone disorder,CKD-MBD)的发生发展。除了经典的甲状旁腺激素(PTH)/维生素D轴理论,最近有研究发现成纤维细胞生长因子23(fibroblast growth facto...多种生物因子参与着慢性肾脏病矿物质和骨代谢异常(chronic kidney disease–mineral and bone disorder,CKD-MBD)的发生发展。除了经典的甲状旁腺激素(PTH)/维生素D轴理论,最近有研究发现成纤维细胞生长因子23(fibroblast growth factor23,FGF23)/Klotho、牙基质蛋白1(dentin matrix protein 1,DMP1)、分泌型蛋白Dickkopf-1(secretory protein Dickkopf 1,DKK1)和硬化蛋白(sclerostin)等因子也直接或间接参与着CKD-MBD的进展。本文将对上述因子的最新研究做一综述。展开更多
基金supported by NIH grants Jian-Quan Feng (DE018486) and to Chun-Lin Qin (DE005092)State Key Laboratory of Oral Diseases Open Funding (SKLODOF2010-03) to Jian-Quan Feng
文摘Deletion or mutation of dentin matrix protein 1 （DMP1） leads to hypophosphatemic rickets and defects within the dentin. However, it is largely unknown if this pathological change is a direct role of DMP1 or an indirect role of phosphate （Pi） or both. It has also been previously shown that Klotho-deficient mice, which displayed a high Pi level due to a failure of Pi excretion, causes mild defects in the dentinal structure. This study was to address the distinct roles of DMP1 and Pi homeostasis in cell differentiation, apoptosis and mineralization of dentin and enamel. Our working hypothesis was that a stable Pi homeostasis is critical for postnatal tooth formation, and that DMP1 has an antiapoptotic role in both amelogenesis and dentinogenesis. To test this hypothesis, Dmpl-null （Dmpl-/-）, Klotho-deficient （kl/kl）, Dmpl/Klotho-double-deficient （Dmpl-/-/kl/kl） and wild-type （WT） mice were killed at the age of 6 weeks. Combinations of X-ray, microcomputed tomography （I^CT）, scanning electron microscopy （SEM）, histology, apoptosis and immunohistochemical methods were used for characterization of dentin, enamel and pulp structures in these mutant mice. Our results showed that Dmpl-/- （a low Pi level） or kl/kl（a high Pi level） mice displayed mild dentin defects such as thin dentin and a reduction of dentin tubules. Neither deficient mouse line exhibited any apparent changes in enamel or pulp structure. However, the double-deficient mice （a high Pi level） displayed severe defects in dentin and enamel structures, including loss of dentinal tubules and enamel prisms, as well as unexpected ectopic ossification within the pulp root canal. TUNEL assay showed a sharp increase in apoptotic cells in ameloblasts and odontoblasts. Based on the above findings, we conclude that DMP1 has a protective role for odontoblasts and ameloblasts in a pro-apoptotic environment （a high Pi level）.
基金the National Natural Science Foundation of China(grant numbers 31670994,U1904145 and 81901039)Nature Science Foundation of Henan Province(grant numbers 182300410340,222300420569)+4 种基金Henan Medical Science and Technology Research Project(grant number SBGJ202002073,SBGJ202002075)Science and Technology Department of Science and Technology Research(grant number 182102310456)Henan Province Middle-Aged And Young Health Science and Technology Innovation Leader Training Project(grant number YXKC2021014)Scientific Research and Innovation Team of The First Affiliated Hospital of Zhengzhou University(grant number QNCXTD2023021)Union project of Medical and Technology Research Program of Henan Province(grant number LHGJ20190191).
文摘It is still a huge challenge for bone regenerative biomaterial to balance its mechanical,biological and biodegradable properties.In the present study,a new composite material including treated dentin matrix(TDM)andα-calcium sulphate hemihydrate(α-CSH)was prepared.The optimal composition ratio between TDM andα-CSH was explored.The results indicate that both components were physically mixed and structurally stable.Its compressive strength reaches up to 5.027±0.035 MPa for 50%TDM/α-CSH group,similar to human cancellous bone tissues.Biological experiments results show that TDM/α-CSH composite exhibits excellent biocompatibility and the expression of osteogenic related genes and proteins(ALP,RUNX2,OPN)is significantly increased.In vivo experiments suggest that the addition of TDM for each group(10%,30%,50%)effectively promotes cell proliferation and osteomalacia.In addition,50%of the TDM/α-CSH combination displays optimal osteoconductivity.The novel TDM/α-CSH composite is a good candidate for certain applications in bone tissue engineering.
基金We thank Dr. Chunlin Qin （Bayler colleage of dentistry, Texas A＆M University） for providing us DMP1 antibody. This study was supported by China National Key Research and Development Program （2016YFA0100801 YS）, and the National Natural Science Foundation of China （Grant Nos. 8133030 YS and 31620103904 YS）, and grants： 2016YFC102705 YS 2014BAI04B07 WZL+1 种基金 81470715 YS TJ1504219036 WZL： 2017BR009 YS.
文摘The blood-brain barrier （BBB） is a tight boundary formed between endothelial cells and astrocytes, which separates and protects brain from most pathogens as well as neural toxins in circulation. However, detailed molecular players involved in formation of BBB are not completely known. Dentin matrix protein I （DMP1）-proteoglycan （PG）, which is known to be involved in mineralization of bones and dentin, is also expressed in soft tissues including brain with unknown functions. In the present study, we reported that DMPI-PG was expressed in brain astrocytes and enriched in BBB units. The only glycosylation site of DMP1 is serine89 （S89） in the N-terminal domain of the protein in mouse. Mutant mice with DMP1 point mutations changing S89 to glycine （S89G）, which completely eradicated glycosylation of the protein, demonstrated severe BBB disruption. Another breed of DMP1 mutant mice, which lacked the C-terminal domain of DMP1, manifested normal BBB function. The polarity of S89G-DMP1 astrocytes was disrupted and cell-cell adhesion was loosened. Through a battery of analyses, we found that DMP1 glycosylation was critically required for astrocyte maturation both in vitro and in vivo. S89G-DMP1 mutant astrocytes failed to express aquaporin 4 and had reduced laminin and ZO1 expression, which resulted in disruption of BBB. Interestingly, overexpression of wild-type DMP1-PG in mouse brain driven by the nestin promoter elevated laminin and ZO1 expression beyond wild type levels and could effectively resisted intravenous mannitol-induced BBB reversible opening. Taken together, our study not only revealed a novel element, i.e., DMP1-PG, that reg- ulated BBB formation, but also assigned a new function to DMP1-PG.
基金This study was funded by the National Natural Science Foundation of China(grant numbers 31670994,U1904145,and 81901039)Nature Science Foundation of Henan province(grant number 182300410340)and Union project of Medical and Technology Research Program of Henan Province(grant number LHGJ20190191).
文摘Treated dentin matrix(TDM)is an ideal scaffold material containing multiple extracellular matrix factors.The canonical Wnt signaling pathway is necessary for tooth regeneration.Thus,this study investigated whether the TDM can promote the odontogenic differentiation of human dental pulp stem cells(hDPSCs)and determined the potential role of Wnt/β-catenin signaling in this process.Different concentrations of TDM promoted the dental differentiation of the hDPSCs and meanwhile,the expression of GSK3βwas decreased.Of note,the expression of the Wnt/β-catenin pathway-related genes changed significantly in the context of TDM induction,as per RNA sequencing(RNA seq)data.In addition,the experiment showed that new dentin was visible in rat mandible cultured with TDM,and the thickness was significantly thicker than that of the control group.In addition,immunohistochemical staining showed lower GSK3βexpression in new dentin.Consistently,the GSK3βknockdown hDPSCs performed enhanced odotogenesis compared with the control groups.However,GSK3βoverexpressing could decrease odotogenesis of TDM-induced hDPSCs.These results were confirmed in immunodeficient mice and Wistar rats.These suggest that TDM promotes odontogenic differentiation of hDPSCs by directly targeting GSK3βand activating the canonical Wnt/β-catenin signaling pathway and provide a theoretical basis for tooth regeneration engineering.
基金This study was supported by grants from the National Natural Science Foundation of China，the Natural Science Foundation of Liaoning Province
文摘Background： Ankylosing spondylitis （AS） is the most common rheumatic condition that is slowly progressive and predominantly affects adolescents. Pathological bone formation associated with AS is an important cause of disability. The aim of the study was to investigate the possible involvement of the genes related to endochondral ossification and ectopia ossification in genetic susceptibility to AS in a Chinese Han population. Methods： Sixty-eight single nucleotide polymorphisms （SNPs） from 13 genes were genotyped in discovery cohorts including 300 AS patients and 180 healthy controls. The rs10019009 in dentin matrix protein 1 （DMP1） gene shown as association with AS after multiple testing corrections in discovery cohorts was replicated in a validation independent cohort of 620 AS patients and 683 healthy controls. The rs 10019009 was assessed with bioin fomlatics including phylogenetic context, F-SNP and FastSNP functional predictions, secondary structure prediction, and molecular modeling. We performed a functional analysis of rs10019009 via reverse transcription-polymerase chain reaction, alkaline phosphatase （ALP） activity in human osteosarcoma U2OS cells. Results： Interestingly, the SNP rs10019009 was associated with AS in both the discovery cohort （P = 0.0012） and validation cohort （P - 0.0349）, as well as overall （P = 0.0004） in genetic case-control association analysis. After a multivariate logistic regression analysis, the effect of this genetic variant was observed to be independent of linkage disequilibrium. Via bioinformatics analysis, it was found that the amino acid change of the rs 10019009 led to changes of SNP function, secondary structure, tertiary confomlation, and splice mode. Finally, functional analysis ofrsl0019009 in U2OS cells demonstrated that the risk T allele of the rsl0019009 increased enzymatic activity of ALP, compared to that of the nonrisk allele （P = 0.0080）. Conclusions： These results suggested that the DMP1 gene seems to be involved in genetic predisposition to AS, which may contribute to the ectopic mineralization or ossification in AS. In addition, DMP1 gene may be a promising intervention target for AS in the future.
文摘The presence of matrix metalloproteinase-2 （MMP-2） in dentin has been reported, but its distribution and activity level in mature human coronal dentin are not well understood. The purpose of this study was to determine the MMP-2 distribution and relative activity in demineralized dentin. Crowns of twenty eight human molars were sectioned into inner （ID）, middle （MD）, and outer dentin （OD） regions and demineralized. MMP-2 was extracted with 0.33 mol·L-1 EDTA/2 mol·L-1 guanidine-HCl, pH 7.4, and MMP-2 concentration was estimated with enzyme-linked immunoabsorbant assay （ELISA）. Further characterization was accomplished by Western blotting analysis and gelatin zymography. The mean concentrations of MMP-2 per mg dentin protein in the dentin regions were significantly different （P=0.043）： 0.9 ng （ID）, 0.4 ng （MD）, and 2.2 ng （OD）, respectively. The pattern of MMP-2 concentration was OD〉ID〉MD. Western blotting analysis detected -66 and -72 kDa immunopositive proteins corresponding to pro- and mature MMP-2, respectively, in the ID and MD, and a -66 kDa protein in the OD. Gelatinolytic activity consistent with MMP-2 was detected in all regions. Interestingly, the pattern of levels of Western blot immunodetection and gelatinolytic activity was MD〉ID〉OD. The eoneentration of MMP-2 in human coronal dentin was highest in the region of dentin that contains the dentinoenamel junction and least in the middle region of dentin. However, levels of Western blot immunodetection and gelatinolytic activity did not correlate with the estimated regional concentrations of MMP-2, potentially indicating region specific protein interactions.
文摘多种生物因子参与着慢性肾脏病矿物质和骨代谢异常(chronic kidney disease–mineral and bone disorder,CKD-MBD)的发生发展。除了经典的甲状旁腺激素(PTH)/维生素D轴理论,最近有研究发现成纤维细胞生长因子23(fibroblast growth factor23,FGF23)/Klotho、牙基质蛋白1(dentin matrix protein 1,DMP1)、分泌型蛋白Dickkopf-1(secretory protein Dickkopf 1,DKK1)和硬化蛋白(sclerostin)等因子也直接或间接参与着CKD-MBD的进展。本文将对上述因子的最新研究做一综述。
