The Experts below are selected from a list of 528 Experts worldwide ranked by ideXlab platform
Mary Macdougall - One of the best experts on this subject based on the ideXlab platform.
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Fluoride Alters Signaling Pathways Associated with the Initiation of Dentin Mineralization in Enamel Fluorosis Susceptible Mice
Biological Trace Element Research, 2020Co-Authors: Yu-hsing Kao, Hayato Ohshima, Mary Macdougall, Nanase Igarashi, Dawud Abduweli Uyghurturk, Yan Zhang, Yoshiro Takano, Pamela Den Besten, Yukiko NakanoAbstract:Fluoride can alter the formation of mineralized tissues, including enamel, Dentin, and bone. Dentin fluorosis occurs in tandem with enamel fluorosis. However, the pathogenesis of Dentin fluorosis and its mechanisms are poorly understood. In this study, we report the effects of fluoride on the initiation of Dentin matrix formation and odontoblast function. Mice from two enamel fluorosis susceptible strains (A/J and C57BL/6J) were given either 0 or 50 ppm fluoride in drinking water for 4 weeks. In both mouse strains, there was no overall change in Dentin thickness, but fluoride treatment resulted in a significant increase in the thickness of the preDentin layer. The lightly mineralized layer (LL), which lies at the border between preDentin and fully mineralized Dentin and is associated with Dentin Phosphoprotein (DPP), was absent in fluoride exposed mice. Consistent with a possible reduction of DPP, fluoride-treated mice showed reduced immunostaining for Dentin sialoprotein (DSP). Fluoride reduced RUNX2, the transcription regulator of Dentin sialoPhosphoprotein (DSPP), that is cleaved to form both DPP and DSP. In fluoride-treated mouse odontoblasts, the effect of fluoride was further seen in the upstream of RUNX2 as the reduced nuclear translocation of β-catenin and phosphorylated p65/NFκB. In vitro, MD10-F2 pre-odontoblast cells showed inhibition of the Dspp mRNA level in the presence of 10 μM fluoride, and qPCR analysis showed a significantly downregulated level of mRNAs for RUNX2, β-catenin, and Wnt10b. These findings indicate that in mice, systemic exposure to excess fluoride resulted in reduced Wnt/β-catenin signaling in differentiating odontoblasts to downregulate DSPP production via RUNX2.
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differential regulation of Dentin sialoPhosphoprotein expression by runx2 during odontoblast cytodifferentiation
Journal of Biological Chemistry, 2005Co-Authors: Shuo Chen, Ting Ting Gu, Sheela Rani, Yimin Wu, Aaron Unterbrink, Jelica Gluhakheinrich, Hui Hsiu Chuang, Mary MacdougallAbstract:Abstract Dentin sialoPhosphoprotein (DSPP) consists of Dentin sialoprotein (DSP) and Dentin Phosphoprotein (DPP). The spatial-temporal expression of DSPP is largely restricted during differentiational stages of dental cells. DSPP plays a vital role in tooth development. It is known that an osteoblast-specific transcription factor, Runx2, is essential for osteoblast differentiation. However, effects of Runx2 on DSPP transcription remain unknown. Here, we studied different roles of Runx2 in controlling DSPP expression in mouse preodontoblast (MD10-F2) and odontoblast (MO6-G3) cells. Two Runx2 isoforms were expressed in preodontoblast and odontoblast cells, and in situ hybridization assay showed that DSPP expression increased, whereas Runx2 was down-regulated during odontoblast differentiation and maturation. Three potential Runx2 sites are present in promoters of mouse and rat DSPP genes. Runx2 binds to these sites as demonstrated by electrophoretic mobility shift assay and supershift experiments. Mutations of Runx2 sites in mouse DSPP promoter resulted in a decline of promoter activity in MD10-F2 cells compared with an increase of its activity in MO6-G3 cells. Multiple Runx2 sites were more active than a single site in regulating the DSPP promoter. Furthermore, forced overexpression of Runx2 isoforms induced increases of endogenous DSPP protein levels in MD10-F2 cells but reduced its expression in MO6-G3 cells consistent with the DSPP promoter analysis. Thus, our results suggest that differential positive and negative regulation of DSPP by Runx2 is dependent on use of cytodifferentiation of dental ectomesenchymal-derived cells that may contribute to the spatial-temporal expression of DSPP during tooth development.
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Dentin Sialoprotein and Dentin Phosphoprotein Overexpression during Amelogenesis
Journal of Biological Chemistry, 2005Co-Authors: Michael L. Paine, Wen Luo, Hongjun Wang, Pablo Bringas, Amanda Y. W. Ngan, Vetea G. Miklus, Dan-hong Zhu, Mary Macdougall, Shane N. White, Malcolm L. SneadAbstract:Abstract The gene for Dentin sialoPhosphoprotein produces a single protein that is post-translationally modified to generate two distinct extracellular proteins: Dentin sialoprotein and Dentin Phosphoprotein. In teeth, Dentin sialoPhosphoprotein is expressed primarily by odontoblast cells, but is also transiently expressed by presecretory ameloblasts. Because of this expression profile it appears that Dentin sialoPhosphoprotein contributes to the early events of amelogenesis, and in particular to those events that result in the formation of the Dentino-enamel junction and the adjacent “aprismatic” enamel. Using a transgenic animal approach we have extended Dentin sialoprotein or Dentin Phosphoprotein expression throughout the developmental stages of amelogenesis. Overexpression of Dentin sialoprotein results in an increased rate of enamel mineralization, however, the enamel morphology is not significantly altered. In wild-type animals, the inclusion of Dentin sialoprotein in the forming aprismatic enamel may account for its increased hardness properties, when compared with bulk enamel. In contrast, the overexpression of Dentin Phosphoprotein creates “pitted” and “chalky” enamel of non-uniform thickness that is more prone to wear. Disruptions to the prismatic enamel structure are also a characteristic of the Dentin Phosphoprotein overexpressing animals. These data support the previous suggestion that Dentin sialoprotein and Dentin Phosphoprotein have distinct functions related to tooth formation, and that the Dentino-enamel junction should be viewed as a unique transition zone between enamel and the underlying Dentin. These results support the notion that the Dentin proteins expressed by presecretory ameloblasts contribute to the unique properties of the Dentino-enamel junction.
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Dentin Phosphoprotein compound mutation in Dentin sialoPhosphoprotein causes Dentinogenesis imperfecta type III.
American Journal of Medical Genetics Part A, 2004Co-Authors: Juan Dong, Leticia Jeffords, Mary MacdougallAbstract:A rare compound mutation involving a 36 bp deletion and 18 bp insertion within exon 5 of the Dentin sialoPhosphoprotein (DSPP) gene has been identified in a family with Dentinogenesis imperfecta type III (DGI-III). The DSPP gene encodes two major tooth matrix proteins Dentin sialoprotein (DSP) and Dentin Phosphoprotein (DPP). DSPP mutations associated with DGI-III results in an in frame truncation of the serine aspartic acid triplet repeat found in DPP near the highly conserved carboxyl terminal region shortening the protein by six amino acids. Clinically this family presents with discolored amber opalescent teeth and severe attrition of the tooth structure. This study is the first report of a mutation within DPP associated with a genetic Dentin disease. Our study indicates that DGI-III is allelic with some forms of DGI-II with and without progressive hearing loss and Dentin dysplasia type II that have been shown to be caused by mutations within the DSP coding or signal peptide regions.
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Spatial and temporal activity of the Dentin sialoPhosphoprotein gene promoter: differential regulation in odontoblasts and ameloblasts
The International journal of developmental biology, 1999Co-Authors: Taduru Sreenath, Mary Macdougall, Andrew Cho, Ashok B KulkarniAbstract:Dentin sialoprotein and Dentin Phosphoprotein are non-collagenous proteins that are cleavage products of Dentin sialoPhosphoprotein (DSPP). Although these two protein products are believed to have a crucial role in the process of tooth mineralization, their precise biological functions and the molecular mechanisms of gene regulation are not clearly understood. To understand such functions, we have developed a transgenic mouse model expressing a reporter gene (lacZ) under the control of approximately 6 kb upstream sequences of Dspp. The transgenic fusion protein was designed to reside within the cells to facilitate the precise identification of cell type and developmental stages at which the Dspp-lacZ gene is expressed. The results presented in this report demonstrate: (a) the 6 kb upstream sequences of Dspp have the necessary regulatory elements to direct the tissue specific expression of the transgene similar to endogenous Dspp, (b) both odontoblasts and ameloblasts exhibit transgene expression in a differentiation dependent manner, and (c) a differential regulation of the transgene in odontoblasts and ameloblasts occurs during tooth development and mineralization.
Chunlin Qin - One of the best experts on this subject based on the ideXlab platform.
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Transgenic expression of Dentin Phosphoprotein (DPP) partially rescued the Dentin defects of DSPP-null mice.
PLOS ONE, 2018Co-Authors: Hua Zhang, P. Liu, Xiaohua Xie, Tian Liang, Chunlin QinAbstract:Mutations in the Dentin sialoPhosphoprotein (DSPP) gene cause Dentinogenesis imperfecta. After synthesis, DSPP is proteolytically processed into NH2- and COOH-terminal fragments. The NH2-terminal fragment of DSPP is highly glycosylated but not phosphorylated, whereas the COOH-terminal fragment (named "Dentin Phosphoprotein" or "DPP") is highly phosphorylated but not glycosylated. These two fragments are believed to perform distinct roles in Dentin formation. To analyze the functions of DPP in Dentinogenesis, we created "Dspp-/-;DPP Tg mice", which expressed transgenic DPP driven by a Type I collagen promoter but lacked the endogenous Dspp gene. We characterized the Dentin of the Dspp-/-;DPP Tg mice using X-ray radiography, histology, scanning electron microscopy, double fluorochrome labeling, immunohistochemistry and in situ hybridization. Micro-computed tomography analyses revealed that at postnatal 6 months, the transgenic expression of DPP increased the Dentin thickness of the Dspp-null mice by 97.1% and restored the Dentin material density by 29.5%. Histological analyses showed that the Dspp-null mice manifested an abnormal widening of the preDentin while the preDentin in Dspp-/-;DPP Tg mice was narrower than in the Dspp-null mice. Scanning electron microscopy analyses showed that the Dentinal tubules in the Dspp-/-;DPP Tg mice were better organized than in the Dspp-null mice. The double fluorochrome labeling analyses demonstrated that the Dentin mineral deposition rate in the Dspp-/-;DPP Tg mice was significantly improved compared to that in the Dspp-null mice. These findings indicate that the transgenic expression of DPP partially rescued the Dentin defects of the DSPP-null mice, suggesting that DPP may promote Dentin formation and that the coordinated actions between DPP and the NH2-terminal fragment of DSPP may be necessary for Dentinogenesis.
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Transgenic Expression of Dentin Phosphoprotein Inhibits Skeletal Development
European Journal of Histochemistry, 2016Co-Authors: Hua Zhang, P. Liu, S. Wang, C. Liu, Priyam Jani, Chunlin QinAbstract:Dentin sialoPhosphoprotein (DSPP) is proteolytically processed into an NH2-terminal fragment called Dentin sialoprotein (DSP) and a COOH-terminal fragment known as Dentin Phosphoprotein (DPP). These two fragments are believed to perform distinct roles in formation of bone and Dentin. To investigate the functions of DPP in skeletal development, we generated transgenic mice to overexpress hemagglutinin (HA)-tagged DPP under the control of a 3.6 kb type I collagen (Col1a1) promoter (designated as Col1a1-HA-DPP). The Col1a1-HA-DPP transgenic mice were significantly smaller by weight, had smaller skeletons and shorter long bones than their wild type littermates, as demonstrated by X-ray radiography. They displayed reduced trabecular bone formation and narrower zones of proliferative and hypertrophic chondrocytes in the growth plates of the long bones. Histological analyses showed that the transgenic mice had reduced cell proliferation in the proliferating zone, but lacked obvious defects in the chondrocyte differentiation. In addition, the transgenic mice with a high level of transgene expression developed spontaneous long bone fractures. In conclusion, overexpressing DPP inhibited skeletal development, suggesting that the balanced actions between the NH2- and COOH-terminal fragments of DSPP may be required for normal skeletal development.
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Mineralization induction effects of osteopontin, bone sialoprotein, and Dentin Phosphoprotein on a biomimetic collagen substrate
Journal of biomedical materials research. Part A, 2012Co-Authors: Kevin M. Zurick, Chunlin Qin, Matthew T. BernardsAbstract:Native bone tissue is composed of a matrix of collagen, noncollagenous proteins, and calcium phosphate minerals, which are primarily hydroxyapatite. The SIBLING (small integrin-binding ligand, N-linked glycoprotein) family of proteins is the primary noncollagenous protein group found in mineralized tissues. In this work, the mineralization induction capabilities of three of the SIBLING members, bone sialoprotein (BSP), osteopontin (OPN), and the calcium-binding subdomain of Dentin sialoPhosphoprotein, Dentin Phosphoprotein (DPP), are directly compared on a biomimetic collagen substrate. A self-assembled, loosely aligned collagen fibril substrate was prepared, and then 125I-radiolabeled adsorption isotherms were developed for BSP, OPN, and DPP. The results showed that BSP exhibited the highest binding capacity for collagen at lower concentrations, followed by DPP and OPN. However, at the highest concentrations, all three proteins had similar adsorption levels. The adsorption isotherms were then used to identify conditions that resulted in identical amounts of adsorbed protein. These substrates were prepared and placed in simulated body fluid for 5, 10, and 24 h at 37°C. The resulting mineral morphology was assessed by atomic force microscopy, and the composition was determined using photochemical assays. Mineralization was seen in the presence of all the proteins. However, DPP was seen to be the only protein that formed individual mineral nodules similar to those seen in developing bone. This suggests that DPP plays a significant role in the biomineralization process and that the incorporation of DPP into tissue engineering constructs may facilitate the induction of biomimetic mineral formation. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.
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adhesion of mc3t3 e1 cells bound to Dentin Phosphoprotein specifically bound to collagen type i
Journal of Biomedical Materials Research Part A, 2012Co-Authors: Kevin M. Zurick, Chunlin Qin, Matthew T. BernardsAbstract:Dentin sialoPhosphoprotein (DSPP) is a member of the SIBLING (small integrin binding N-linked glycoprotein) family of proteins commonly found in mineralized tissues. Dentin Phosphoprotein (DPP) is a naturally occurring subdomain of DSPP that contains the cell binding RGD sequence. Previously, the orientation and conformation of other SIBLING family members specifically bound to collagen I have been investigated with respect to their cell adhesion properties. In this study, the orientation of DPP under similar circumstances is examined, and the results are discussed relative to the previous investigations. Radiolabeled adsorption isotherms were developed for DPP adsorbing to both tissue culture polystyrene (TCPS) and collagen coated TCPS. Then, a MC3T3-E1 cell adhesion assay was performed on TCPS and collagen coated TCPS in the presence of identical amounts of adsorbed DPP. It was discovered that there was a significant difference in the number of bound cells on the TCPS and collagen coated TCPS, with a preference for TCPS. Furthermore, a cell inhibition assay was conducted to confirm that the cell binding that occurred was due to specific integrin interactions with the RGD sequence of DPP. These results suggest that the orientation of DPP, rather than its conformation, dictates the accessibility of the cell binding RGD domains of DPP and that the RGD sequence in DPP is less accessible when DPP is specifically bound to collagen. The results obtained in this study are in stark contrast to previous studies with related SIBLING proteins, and suggest that DPP does not play a key role in cell binding to the collagen matrix of developing bone.
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Adhesion of MC3T3‐E1 cells bound to Dentin Phosphoprotein specifically bound to collagen type I
Journal of biomedical materials research. Part A, 2012Co-Authors: Kevin M. Zurick, Chunlin Qin, Matthew T. BernardsAbstract:Dentin sialoPhosphoprotein (DSPP) is a member of the SIBLING (small integrin binding N-linked glycoprotein) family of proteins commonly found in mineralized tissues. Dentin Phosphoprotein (DPP) is a naturally occurring subdomain of DSPP that contains the cell binding RGD sequence. Previously, the orientation and conformation of other SIBLING family members specifically bound to collagen I have been investigated with respect to their cell adhesion properties. In this study, the orientation of DPP under similar circumstances is examined, and the results are discussed relative to the previous investigations. Radiolabeled adsorption isotherms were developed for DPP adsorbing to both tissue culture polystyrene (TCPS) and collagen coated TCPS. Then, a MC3T3-E1 cell adhesion assay was performed on TCPS and collagen coated TCPS in the presence of identical amounts of adsorbed DPP. It was discovered that there was a significant difference in the number of bound cells on the TCPS and collagen coated TCPS, with a preference for TCPS. Furthermore, a cell inhibition assay was conducted to confirm that the cell binding that occurred was due to specific integrin interactions with the RGD sequence of DPP. These results suggest that the orientation of DPP, rather than its conformation, dictates the accessibility of the cell binding RGD domains of DPP and that the RGD sequence in DPP is less accessible when DPP is specifically bound to collagen. The results obtained in this study are in stark contrast to previous studies with related SIBLING proteins, and suggest that DPP does not play a key role in cell binding to the collagen matrix of developing bone.
Matthew T. Bernards - One of the best experts on this subject based on the ideXlab platform.
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Probing the influence of SIBLING proteins on collagen-I fibrillogenesis and denaturation.
Connective Tissue Research, 2017Co-Authors: Chengyu Jiang, Kevin M. Zurick, Matthew T. BernardsAbstract:ABSTRACTBone tissue is comprised of collagen, non-collagenous proteins, and hydroxyapatite and the SIBLING (small integrin binding, N-linked glycoprotein) family of proteins is the primary group of non-collagenous proteins. By replicating the native interactions between collagen and the SIBLING proteins at the interface of an implant, it is believed that a bone scaffold will more easily integrate with the surrounding tissue. In this work, bone sialoprotein, osteopontin (OPN), Dentin sialoprotein (DSP), Dentin Phosphoprotein (DPP), C-terminal fragment of Dentin matrix protein 1 (DMP1-C), and proteoglycan versions of DSP (DSP-PG) and DMP1 (DMP1-PG) were tested individually to determine their roles in collagen fibrillogenesis and the prevention of denaturation. It was shown that DSP and DPP slowed down fibrillogenesis, while other SIBLINGs had limited impact. In addition, the denaturation time was faster in the presence of DSP and OPN, indicating a negative impact. The role of calcium ions in these processes...
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Mineralization induction effects of osteopontin, bone sialoprotein, and Dentin Phosphoprotein on a biomimetic collagen substrate
Journal of biomedical materials research. Part A, 2012Co-Authors: Kevin M. Zurick, Chunlin Qin, Matthew T. BernardsAbstract:Native bone tissue is composed of a matrix of collagen, noncollagenous proteins, and calcium phosphate minerals, which are primarily hydroxyapatite. The SIBLING (small integrin-binding ligand, N-linked glycoprotein) family of proteins is the primary noncollagenous protein group found in mineralized tissues. In this work, the mineralization induction capabilities of three of the SIBLING members, bone sialoprotein (BSP), osteopontin (OPN), and the calcium-binding subdomain of Dentin sialoPhosphoprotein, Dentin Phosphoprotein (DPP), are directly compared on a biomimetic collagen substrate. A self-assembled, loosely aligned collagen fibril substrate was prepared, and then 125I-radiolabeled adsorption isotherms were developed for BSP, OPN, and DPP. The results showed that BSP exhibited the highest binding capacity for collagen at lower concentrations, followed by DPP and OPN. However, at the highest concentrations, all three proteins had similar adsorption levels. The adsorption isotherms were then used to identify conditions that resulted in identical amounts of adsorbed protein. These substrates were prepared and placed in simulated body fluid for 5, 10, and 24 h at 37°C. The resulting mineral morphology was assessed by atomic force microscopy, and the composition was determined using photochemical assays. Mineralization was seen in the presence of all the proteins. However, DPP was seen to be the only protein that formed individual mineral nodules similar to those seen in developing bone. This suggests that DPP plays a significant role in the biomineralization process and that the incorporation of DPP into tissue engineering constructs may facilitate the induction of biomimetic mineral formation. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.
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adhesion of mc3t3 e1 cells bound to Dentin Phosphoprotein specifically bound to collagen type i
Journal of Biomedical Materials Research Part A, 2012Co-Authors: Kevin M. Zurick, Chunlin Qin, Matthew T. BernardsAbstract:Dentin sialoPhosphoprotein (DSPP) is a member of the SIBLING (small integrin binding N-linked glycoprotein) family of proteins commonly found in mineralized tissues. Dentin Phosphoprotein (DPP) is a naturally occurring subdomain of DSPP that contains the cell binding RGD sequence. Previously, the orientation and conformation of other SIBLING family members specifically bound to collagen I have been investigated with respect to their cell adhesion properties. In this study, the orientation of DPP under similar circumstances is examined, and the results are discussed relative to the previous investigations. Radiolabeled adsorption isotherms were developed for DPP adsorbing to both tissue culture polystyrene (TCPS) and collagen coated TCPS. Then, a MC3T3-E1 cell adhesion assay was performed on TCPS and collagen coated TCPS in the presence of identical amounts of adsorbed DPP. It was discovered that there was a significant difference in the number of bound cells on the TCPS and collagen coated TCPS, with a preference for TCPS. Furthermore, a cell inhibition assay was conducted to confirm that the cell binding that occurred was due to specific integrin interactions with the RGD sequence of DPP. These results suggest that the orientation of DPP, rather than its conformation, dictates the accessibility of the cell binding RGD domains of DPP and that the RGD sequence in DPP is less accessible when DPP is specifically bound to collagen. The results obtained in this study are in stark contrast to previous studies with related SIBLING proteins, and suggest that DPP does not play a key role in cell binding to the collagen matrix of developing bone.
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Adhesion of MC3T3‐E1 cells bound to Dentin Phosphoprotein specifically bound to collagen type I
Journal of biomedical materials research. Part A, 2012Co-Authors: Kevin M. Zurick, Chunlin Qin, Matthew T. BernardsAbstract:Dentin sialoPhosphoprotein (DSPP) is a member of the SIBLING (small integrin binding N-linked glycoprotein) family of proteins commonly found in mineralized tissues. Dentin Phosphoprotein (DPP) is a naturally occurring subdomain of DSPP that contains the cell binding RGD sequence. Previously, the orientation and conformation of other SIBLING family members specifically bound to collagen I have been investigated with respect to their cell adhesion properties. In this study, the orientation of DPP under similar circumstances is examined, and the results are discussed relative to the previous investigations. Radiolabeled adsorption isotherms were developed for DPP adsorbing to both tissue culture polystyrene (TCPS) and collagen coated TCPS. Then, a MC3T3-E1 cell adhesion assay was performed on TCPS and collagen coated TCPS in the presence of identical amounts of adsorbed DPP. It was discovered that there was a significant difference in the number of bound cells on the TCPS and collagen coated TCPS, with a preference for TCPS. Furthermore, a cell inhibition assay was conducted to confirm that the cell binding that occurred was due to specific integrin interactions with the RGD sequence of DPP. These results suggest that the orientation of DPP, rather than its conformation, dictates the accessibility of the cell binding RGD domains of DPP and that the RGD sequence in DPP is less accessible when DPP is specifically bound to collagen. The results obtained in this study are in stark contrast to previous studies with related SIBLING proteins, and suggest that DPP does not play a key role in cell binding to the collagen matrix of developing bone.
Shigeki Suzuki - One of the best experts on this subject based on the ideXlab platform.
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Dentin Phosphoprotein inhibits lipopolysaccharide induced macrophage activation independent of its serine aspartic acid rich repeats
Archives of Oral Biology, 2020Co-Authors: Jun Nakanishi, Shigeki Suzuki, Kazuma Yoshida, Naoto Haruyama, Satoru Yamada, Shizu Hiratatsuchiya, Hideki ShibaAbstract:Abstract Objective The objective of this study was to investigate the effects of Dentin Phosphoprotein (DPP) on lipopolysaccharide-induced inflammatory responses of macrophages in vitro. Design Wildtype and mutant recombinant Dentin Phosphoprotein (rDPP) proteins were generated using a mammalian expression system. Macrophages, phorbol 12-myristate 13-acetate-differentiated THP-1 cells, were stimulated with lipopolysaccharide in the absence or presence of rDPP proteins. After the 24-hr incubation, the inflammatory gene expression levels were examined by quantitative reverse-transcription polymerase chain reaction and the amount of secreted TNF-α protein was evaluated by enzyme-linked immunosorbent assay. Furthermore, the subcellular localization of exogenously added rDPP was examined by immunocytochemistry, and the direct binding of rDPP to lipopolysaccharide was quantified by solid-phase binding assay. Results rDPP dose-dependently reduced the expression of lipopolysaccharide-induced inflammatory genes, such as TNFα, IL-1β, and IL-8, and TNF-α protein secretion from the macrophages. Furthermore, mutant rDPP having a shortened serine/aspartic acid-rich repeats (SDrr) was also able to inhibit lipopolysaccharide-induced inflammatory responses of macrophages. rDPP was localized adjacent to the cellular membrane rather than in the cytoplasm, and rDPP was able to bind to lipopolysaccharide. These results suggested that rDPP inhibited lipopolysaccharide-induced inflammatory responses by binding to lipopolysaccharide. Conclusions In addition to the well-known functions of DPP for Dentin mineralization that depend on the SDrr, we demonstrated that DPP possesses anti-inflammatory effects on lipopolysaccharide-stimulated macrophages that are independent of the SDrr.
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Dentin Phosphoprotein inhibits lipopolysaccharide-induced macrophage activation independent of its serine/aspartic acid-rich repeats
Archives of Oral Biology, 2020Co-Authors: Jun Nakanishi, Shigeki Suzuki, Kazuma Yoshida, Shizu Hirata-tsuchiya, Naoto Haruyama, Satoru Yamada, Hideki ShibaAbstract:Abstract Objective The objective of this study was to investigate the effects of Dentin Phosphoprotein (DPP) on lipopolysaccharide-induced inflammatory responses of macrophages in vitro. Design Wildtype and mutant recombinant Dentin Phosphoprotein (rDPP) proteins were generated using a mammalian expression system. Macrophages, phorbol 12-myristate 13-acetate-differentiated THP-1 cells, were stimulated with lipopolysaccharide in the absence or presence of rDPP proteins. After the 24-hr incubation, the inflammatory gene expression levels were examined by quantitative reverse-transcription polymerase chain reaction and the amount of secreted TNF-α protein was evaluated by enzyme-linked immunosorbent assay. Furthermore, the subcellular localization of exogenously added rDPP was examined by immunocytochemistry, and the direct binding of rDPP to lipopolysaccharide was quantified by solid-phase binding assay. Results rDPP dose-dependently reduced the expression of lipopolysaccharide-induced inflammatory genes, such as TNFα, IL-1β, and IL-8, and TNF-α protein secretion from the macrophages. Furthermore, mutant rDPP having a shortened serine/aspartic acid-rich repeats (SDrr) was also able to inhibit lipopolysaccharide-induced inflammatory responses of macrophages. rDPP was localized adjacent to the cellular membrane rather than in the cytoplasm, and rDPP was able to bind to lipopolysaccharide. These results suggested that rDPP inhibited lipopolysaccharide-induced inflammatory responses by binding to lipopolysaccharide. Conclusions In addition to the well-known functions of DPP for Dentin mineralization that depend on the SDrr, we demonstrated that DPP possesses anti-inflammatory effects on lipopolysaccharide-stimulated macrophages that are independent of the SDrr.
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Dentin sialoprotein and Dentin Phosphoprotein have distinct roles in Dentin mineralization
Matrix Biology, 2009Co-Authors: Shigeki Suzuki, Naoto Haruyama, Taduru Sreenath, Cherlita Honeycutt, Anita Terse, Thomas Kohler, Ralph Muller, Michel Goldberg, Ashok B KulkarniAbstract:Dentin sialoPhosphoprotein (DSPP), a major non-collagenous matrix protein of odontoblasts, is proteolytically cleaved into Dentin sialoprotein (DSP) and Dentin Phosphoprotein (DPP). Our previous studies revealed that DSPP null mice display a phenotype similar to human autosomal dominant Dentinogenesis imperfecta, in which teeth have widened preDentin and irregular Dentin mineralization resulting in sporadic unmineralized areas in Dentin and frequent pulp exposure. Earlier in vitro studies suggested that DPP, but not DSP, plays a significant role in initiation and maturation of Dentin mineralization. However, the precise in vivo roles of DSP and DPP are far from clear. Here we report the generation of DPPcKO mice, in which only DSP is expressed in a DSPP null background, resulting in a conditional DPP knockout. DPPcKO teeth show a partial rescue of the DSPP null phenotype with the restored preDentin width, an absence of irregular unmineralized areas in Dentin, and less frequent pulp exposure. Micro-computed tomography (micro-CT) analysis of DPPcKO molars further confirmed this partial rescue with a significant recovery in the Dentin volume, but not in the Dentin mineral density. These results indicate distinct roles of DSP and DPP in Dentin mineralization, with DSP regulating initiation of Dentin mineralization, and DPP being involved in the maturation of mineralized Dentin.
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Dentin sialoprotein and Dentin Phosphoprotein have distinct roles in Dentin mineralization
Matrix Biology, 2009Co-Authors: Shigeki Suzuki, Naoto Haruyama, Taduru Sreenath, Cherlita Honeycutt, Anita Terse, Thomas Kohler, Ralph Muller, Michel Goldberg, Andrew Cho, Ashok B KulkarniAbstract:Dentin sialoPhosphoprotein (DSPP), a major non-collagenous matrix protein of odontoblasts, is proteolytically cleaved into Dentin sialoprotein (DSP) and Dentin Phosphoprotein (DPP). Our previous studies revealed that DSPP null mice display a phenotype similar to human autosomal dominant Dentinogenesis imperfecta, in which teeth have widened preDentin and irregular Dentin mineralization resulting in sporadic unmineralized areas in Dentin and frequent pulp exposure. Earlier in vitro studies suggested that DPP, but not DSP, plays a significant role in initiation and maturation of Dentin mineralization. However, the precise in vivo roles of DSP and DPP are far from clear. Here we report the generation of DPPcKO mice, in which only DSP is expressed in a DSPP null background, resulting in a conditional DPP knockout. DPPcKO teeth show a partial rescue of the DSPP null phenotype with the restored preDentin width, an absence of irregular unmineralized areas in Dentin, and less frequent pulp exposure. Micro-computed tomography (micro-CT) analysis of DPPcKO molars further confirmed this partial rescue with a significant recovery in the Dentin volume, but not in the Dentin mineral density. These results indicate distinct roles of DSP and DPP in Dentin mineralization, with DSP regulating initiation of Dentin mineralization, and DPP being involved in the maturation of mineralized Dentin.
Ashok B Kulkarni - One of the best experts on this subject based on the ideXlab platform.
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Expression of Dentin SialoPhosphoprotein in Non-mineralized Tissues
Journal of Histochemistry and Cytochemistry, 2011Co-Authors: Monica Prasad, Adele L Boskey, Ashok B Kulkarni, Xiaofang Wang, Jian Q FengAbstract:Dentin sialoPhosphoprotein (DSPP) and its cleaved products, Dentin Phosphoprotein (DPP) and Dentin sialoprotein (DSP), play important roles in biomineralization. Believed to be tooth specific, the authors’ group revealed its expression in bone, and more recently, they and other groups also showed its expression in a few types of soft tissues. In this study, the authors systematically examined the expression of DSPP in a variety of non-mineralized tissues using reverse-transcription polymerase chain reaction (RT-PCR), real-time PCR, Western immunoblotting, and immunohistochemistry analyses in wild-type mice as well as β-galactosidase assays in the Dspp lacZ knock-in mice. These approaches showed the presence of DSPP in the salivary glands, cartilage, liver, kidney, and brain and its absence in the heart and spleen. Real-time PCR showed that the expression levels of DSPP mRNA in salivary glands, cartilage, liver, and kidney were higher than in the bone. Interestingly, DSPP was observed in the pericytes of blood vessels in the dental pulp, which are believed to be able to differentiate into odontoblasts. On the basis of these observations, the authors conclude that DSPP and/or its cleaved products may fulfill important functions in certain non-mineralized tissues in addition to its role in biomineralization.
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Dentin sialoprotein and Dentin Phosphoprotein have distinct roles in Dentin mineralization
Matrix Biology, 2009Co-Authors: Shigeki Suzuki, Naoto Haruyama, Taduru Sreenath, Cherlita Honeycutt, Anita Terse, Thomas Kohler, Ralph Muller, Michel Goldberg, Ashok B KulkarniAbstract:Dentin sialoPhosphoprotein (DSPP), a major non-collagenous matrix protein of odontoblasts, is proteolytically cleaved into Dentin sialoprotein (DSP) and Dentin Phosphoprotein (DPP). Our previous studies revealed that DSPP null mice display a phenotype similar to human autosomal dominant Dentinogenesis imperfecta, in which teeth have widened preDentin and irregular Dentin mineralization resulting in sporadic unmineralized areas in Dentin and frequent pulp exposure. Earlier in vitro studies suggested that DPP, but not DSP, plays a significant role in initiation and maturation of Dentin mineralization. However, the precise in vivo roles of DSP and DPP are far from clear. Here we report the generation of DPPcKO mice, in which only DSP is expressed in a DSPP null background, resulting in a conditional DPP knockout. DPPcKO teeth show a partial rescue of the DSPP null phenotype with the restored preDentin width, an absence of irregular unmineralized areas in Dentin, and less frequent pulp exposure. Micro-computed tomography (micro-CT) analysis of DPPcKO molars further confirmed this partial rescue with a significant recovery in the Dentin volume, but not in the Dentin mineral density. These results indicate distinct roles of DSP and DPP in Dentin mineralization, with DSP regulating initiation of Dentin mineralization, and DPP being involved in the maturation of mineralized Dentin.
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Dentin sialoprotein and Dentin Phosphoprotein have distinct roles in Dentin mineralization
Matrix Biology, 2009Co-Authors: Shigeki Suzuki, Naoto Haruyama, Taduru Sreenath, Cherlita Honeycutt, Anita Terse, Thomas Kohler, Ralph Muller, Michel Goldberg, Andrew Cho, Ashok B KulkarniAbstract:Dentin sialoPhosphoprotein (DSPP), a major non-collagenous matrix protein of odontoblasts, is proteolytically cleaved into Dentin sialoprotein (DSP) and Dentin Phosphoprotein (DPP). Our previous studies revealed that DSPP null mice display a phenotype similar to human autosomal dominant Dentinogenesis imperfecta, in which teeth have widened preDentin and irregular Dentin mineralization resulting in sporadic unmineralized areas in Dentin and frequent pulp exposure. Earlier in vitro studies suggested that DPP, but not DSP, plays a significant role in initiation and maturation of Dentin mineralization. However, the precise in vivo roles of DSP and DPP are far from clear. Here we report the generation of DPPcKO mice, in which only DSP is expressed in a DSPP null background, resulting in a conditional DPP knockout. DPPcKO teeth show a partial rescue of the DSPP null phenotype with the restored preDentin width, an absence of irregular unmineralized areas in Dentin, and less frequent pulp exposure. Micro-computed tomography (micro-CT) analysis of DPPcKO molars further confirmed this partial rescue with a significant recovery in the Dentin volume, but not in the Dentin mineral density. These results indicate distinct roles of DSP and DPP in Dentin mineralization, with DSP regulating initiation of Dentin mineralization, and DPP being involved in the maturation of mineralized Dentin.
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Spatial and temporal activity of the Dentin sialoPhosphoprotein gene promoter: differential regulation in odontoblasts and ameloblasts
The International journal of developmental biology, 1999Co-Authors: Taduru Sreenath, Mary Macdougall, Andrew Cho, Ashok B KulkarniAbstract:Dentin sialoprotein and Dentin Phosphoprotein are non-collagenous proteins that are cleavage products of Dentin sialoPhosphoprotein (DSPP). Although these two protein products are believed to have a crucial role in the process of tooth mineralization, their precise biological functions and the molecular mechanisms of gene regulation are not clearly understood. To understand such functions, we have developed a transgenic mouse model expressing a reporter gene (lacZ) under the control of approximately 6 kb upstream sequences of Dspp. The transgenic fusion protein was designed to reside within the cells to facilitate the precise identification of cell type and developmental stages at which the Dspp-lacZ gene is expressed. The results presented in this report demonstrate: (a) the 6 kb upstream sequences of Dspp have the necessary regulatory elements to direct the tissue specific expression of the transgene similar to endogenous Dspp, (b) both odontoblasts and ameloblasts exhibit transgene expression in a differentiation dependent manner, and (c) a differential regulation of the transgene in odontoblasts and ameloblasts occurs during tooth development and mineralization.
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genomic organization chromosomal mapping and promoter analysis of the mouse Dentin sialoPhosphoprotein dspp gene which codes for both Dentin sialoprotein and Dentin Phosphoprotein
Journal of Biological Chemistry, 1998Co-Authors: Jian Q Feng, Xianghong Luan, Ashok B Kulkarni, John Wallace, Dai Jing, Toshio Ohshima, Rena N Dsouza, Christine A Kozak, Mary MacdougallAbstract:Abstract Our laboratory has reported that two major noncollagenous Dentin proteins, Dentin sialoprotein and Dentin Phosphoprotein, are specific cleavage products of a larger precursor protein termed Dentin sialoPhosphoprotein (MacDougall, M., Simmons, D., Luan, X., Nydegger, J., Feng, J. Q., and Gu, T. T. (1997) J. Biol. Chem. 272:835–842). To confirm our single gene hypothesis and initiate in vitropromoter studies, we have characterized the structural organization of the mouse Dentin sialoPhosphoprotein gene. This gene has a transcription unit of ∼9.4 kilobase pairs and is organized into 5 exons and 4 introns. Exon 1 contains a noncoding 5′ sequence, and exon 2 contains the transcriptional start site, signal peptide, and first two amino acids of the NH2 terminus. Exons 3 and 4 contain coding information for 29 and 314 amino acids, respectively. The remainder of the coding information and the untranslated 3′ region are contained in exon 5. Chromosomal mapping localized the gene to mouse chromosome 5q21 in close proximity to other Dentin/bone matrix genes. Computer analysis of the promoter proximal 1.6-kilobase pair sequence revealed a number of potentially important cis-regulatory sequences; these include the recognition elements of AP-1, AP-2, Msx-1, serum response elements, SP-1, and TCF-1. In vitro studies showed that the DSPP promoter is active in an odontoblast cell line, MO6-G3, with basal activity mapped to −95 bp. Two potential enhancer and suppresser elements were identified in the regions between −1447 and −791 bp and −791 and −95 bp, respectively. The structural organization of the Dentin sialoPhosphoprotein gene confirms our finding that both Dentin sialoprotein and Dentin Phosphoprotein are encoded by a single gene with a continuous open reading frame.