Prevalence and determinants of defensive medicine among physicians: a systematic review and meta-analysis.

Defensive medicine, characterized by physicians' inclination toward excessive diagnostic tests and procedures, has emerged as a significant concern in modern healthcare due to its high prevalence and detrimental effects. Despite the growing concerns among healthcare providers, policymakers, and physicians, comprehensive synthesis of the literature on the prevalence and determinants of defensive medicine among physicians has yet been reported. A comprehensive literature search was conducted to identify eligible studies published between 1 January 2000 and 31 December 2022, utilizing six databases (i.e. Web of Science, PubMed, Embase, Scopus, PsycINFO, and Cochrane Library). A meta-analysis was conducted to determine the prevalence and determinants of defensive medicine. Of the 8892 identified articles, 64 eligible studies involving 35.9 thousand physicians across 23 countries were included. The overall pooled prevalence of defense medications was 75.8%. Physicians engaged in both assurance and avoidance behaviors, with the most prevalent subitems being increasing follow-up and avoidance of high-complication treatment protocols. The prevalence of defensive medicine was higher in the African region [88.1%; 95% confidence interval (CI): 80.4%-95.8%] and lower-middle-income countries (89.0%; 95% CI: 78.2%-99.8%). Among the medical specialties, anesthesiologists (92.2%; 95% CI: 89.2%-95.3%) exhibited the highest prevalence. Further, the pooled odds ratios (ORs) of the nine factors at the individual, relational, and organizational levels were calculated, and the influence of previous experience in medical-legal litigation (OR: 1.65; 95% CI: 1.13-2.18) should be considered. The results of this study indicate a high global prevalence of defensive medicine among physicians, underscoring the necessity of implementing targeted interventions to reduce its use, especially in certain regions and specialties. Policymakers should implement measures to improve physicians' medical skills, enhance physician-patient communication, address physicians' medical-legal litigation fears, and reform the medical liability system. Future research should focus on devising and assessing interventions to reduce the use of defensive medicine and to improve the quality of patient care.

Bacterial Quorum-Sensing Signal DSF Inhibits LPS-Induced Inflammations by Suppressing Toll-like Receptor Signaling and Preventing Lysosome-Mediated Apoptosis in Zebrafish.

Bacteria and their eukaryotic hosts have co-evolved for millions of years, and the former can intercept eukaryotic signaling systems for the successful colonization of the host. The diffusible signal factor (DSF) family represents a type of quorum-sensing signals found in diverse Gram-negative bacterial pathogens. Recent evidence shows that the DSF is involved in interkingdom communications between the bacterial pathogen and the host plant. In this study, we explored the anti-inflammatory effect of the DSF and its underlying molecular mechanism in a zebrafish model. We found that the DSF treatment exhibited a strong protective effect on the inflammatory response of zebrafish induced by lipopolysaccharide (LPS). In the LPS-induced inflammation zebrafish model, the DSF could significantly ameliorate the intestinal pathological injury, reduce abnormal migration and the aggregation of inflammatory cells, inhibit the excessive production of inflammatory mediator reactive oxygen species (ROS) content, and prevent apoptosis. Through an RNA-Seq analysis, a total of 938 differentially expressed genes (DEGs) was screened between LPS and LPS + DSF treatment zebrafish embryos. A further bioinformatics analysis and validation revealed that the DSF might inhibit the LPS-induced zebrafish inflammatory response by preventing the activation of signaling in the Toll-like receptor pathway, attenuating the expression of pro-inflammatory cytokines and chemokines, and regulating the activation of the caspase cascade through restoring the expression of lysosomal cathepsins and apoptosis signaling. This study, for the first time, demonstrates the anti-inflammatory role and a potential pharmaceutical application of the bacterial signal DSF. These findings also suggest that the interkingdom communication between DSF-producing bacteria and zebrafish might occur in nature.

Constitutive expression of spliced XBP1 causes perinatal lethality in mice.

Upon endoplasmic reticulum (ER) stress, inositol-requiring enzyme 1 (IRE1) is activated and catalyzes nonconventional splicing of an unspliced X-box binding protein 1 (XBP1U) mRNA to yield a spliced XBP1 (XBP1S) mRNA that encodes a potent XBP1S transcription factor. XBP1S is a key mediator of the IRE1 branch that is essential for alleviating ER stress. We generated a novel mouse strain (referred to as "Xbp1CS/+ " mice) that constitutively expressed XBP1S after Cre recombinase-mediated recombination. Further breeding of these mice with Twist2 Cre recombinase (Twist2-Cre) knock-in mice generated Twist2-Cre;Xbp1CS/+ mice. Most Twist2-Cre;Xbp1CS/+ mice died shortly after birth. Reverse-transcription polymerase chain reaction (RT-PCR) showed that constitutive expression of XBP1S occurred in various mouse tissues examined, but not in the brain. Immunohistochemistry confirmed that although the immunostaining signals for total XBP1 (XBP1U and XBP1S) were found in the calvarial bones in both Twist2-Cre;Xbp1CS/+ and control mice, the signals for XBP1S were only detected in the Twist2-Cre;Xbp1CS/+ mice, but not in the control mice. These results suggest that a precise control of XBP1S production is essential for normal mouse development.

Effect of high phosphate diet on the formation of dentin in Fam20c-deficient mice.

FAM20C (family with sequence similarity 20-member C), a kinase that phosphorylates secretory proteins, plays essential roles in various biological processes. In humans, mutations in FAM20C gene cause Raine syndrome, an autosomal recessive hereditary disease manifesting a broad spectrum of developmental defects including skeletal and craniofacial deformities. Our previous studies revealed that inactivation of Fam20c in mice led to hypophosphatemic rickets and that high phosphate (hPi) diet significantly improved the development of the skeleton in Fam20c-deficient mice. In this study, we evaluated the effects of hPi diet on the formation of dentin in Fam20c-deficient mice, using plain x-ray radiography, micro-computed tomography (µCT), histology, and immunohistochemistry. Plain x-ray radiography and µCT analyses showed that the hPi diet improved the dentin volume fraction and dentin mineral density of the Fam20c-deficient mice. Histology analyses further demonstrated that the hPi diet dramatically improved the integrity of the mandibular first molars and prevented pulp infection and dental abscesses in Fam20c-deficient mice. Our results support that the hPi diet significantly increased the formation and mineralization of dentin in Fam20c-deficient mice, implying that hypophosphatemia is a significant contributor to the dentin defects in Fam20c-deficient subjects.

Live imaging of collagen deposition during skin development and repair in a collagen I - GFP fusion transgenic zebrafish line.

Fibrillar collagen is a major component of many tissues but has been difficult to image in vivo using transgenic approaches because of problems associated with establishing cells and organisms that generate GFP-fusion collagens that can polymerise into functional fibrils. Here we have developed and characterised GFP and mCherry collagen-I fusion zebrafish lines with basal epidermal-specific expression. We use these lines to reveal the dynamic nature of collagen-I fibril deposition beneath the developing embryonic epidermis, as well as the repair of this collagen meshwork following wounding. Transmission electron microscope studies show that these transgenic lines faithfully reproduce the collagen ultrastructure present in wild type larval skin. During skin development we show that collagen I is deposited by basal epidermal cells initially in fine filaments that are largely randomly orientated but are subsequently aligned into a cross-hatch, orthogonal sub-epithelial network by embryonic day 4. Following skin wounding, we see that sub-epidermal collagen is re-established in the denuded domain, initially as randomly orientated wisps that subsequently become bonded to the undamaged collagen and aligned in a way that recapitulates developmental deposition of sub-epidermal collagen. Crossing our GFP-collagen line against one with tdTomato marking basal epidermal cell membranes reveals how much more rapidly wound re-epithelialisation occurs compared to the re-deposition of collagen beneath the healed epidermis. By use of other tissue specific drivers it will be possible to establish zebrafish lines to enable live imaging of collagen deposition and its remodelling in various other organs in health and disease.

High-Phosphate Diet Improved the Skeletal Development of Fam20c-Deficient Mice.

FAM20C (family with sequence similarity 20 - member C) is a protein kinase that phosphorylates secretory proteins, including the proteins that are essential to the formation and mineralization of calcified tissues. Previously, we reported that inactivation of Fam20c in mice led to hypophosphatemic rickets/osteomalacia along with increased circulating fibroblast growth factor 23 (FGF23) levels and dental defects. In this study, we examined whether a high-phosphate (hPi) diet could rescue the skeletal defects in Fam20c-deficient mice. Fam20c conditional knockout (cKO) mice were generated by crossing female Fam20c-floxed mice (Fam20cfl/fl) with male Sox2-Cre;Fam20cfl/+ mice. The pregnant female Fam20cfi/fl mice were fed either a normal or hPi diet until the litters were weaned. The cKO and control offspring were continuously given a normal or hPi diet for 4 weeks after weaning. Plain X-ray radiography, micro-CT, histology, immunohistochemistry (FGF23, DMP1, OPN, and SOX9), and in situ hybridization (type II and type X collagen) analyses were performed to evaluate the effects of an hPi diet on the mouse skeleton. Plain X-ray radiography and micro-CT radiography analyses showed that the hPi diet improved the shape and mineral density of the Fam20c-deficient femurs/tibiae, and rescued the growth plate defects in the long bone. Histology analyses further demonstrated that an hPi diet nearly completely rescued the growth plate-widening defects in the long bone and restored the expanded hypertrophic zone to nearly normal width. These results suggested that the hPi diet significantly improved the skeletal development of the Fam20c-deficient mice, implying that hypophosphatemia partially contributed to the skeletal defects in Fam20c-deficient subjects.

Abrogation of Fam20c altered cell behaviors and BMP signaling of immortalized dental mesenchymal cells.

FAM20C mutations compromise the mineralization of skeleton and tooth in both human and mouse. Putatively, the mineralization disorder is attributed to the elevated fibroblast growth factor 23 (FGF23), which reduced the serum phosphorus by suppressing the reabsorption of phosphorus in kidney. Besides the regulation on systemic phosphorus homeostasis, FAM20C was also implicated to regulate cell behaviors and gene expression through a cell-autonomous manner. To identify the primary effects of Fam20c on dental mesenchymal cells, mouse Fam20c-deficient dental mesenchymal cells were generated by removing the floxed alleles from the immortalized mouse Fam20cf/f dental mesenchymal cells with Cre-expressing lentivirus. The removal of Fam20c exerted no impact on cell morphology, but suppressed the proliferation and mobility of the dental mesenchymal cells. Fam20c deficiency also significantly reduced the expression of Osterix, Runx2, type I Collagen a 1 (Col1a1), Alkaline phosphatase (Alpl) and the members of the small integrin-binding ligand, N-linked glycoprotein (SIBLING) family, but increased Fgf23 expression. Consistently, the in vitro mineralization of Fam20c-deficient dental mesenchymal cells was severely disabled. However, supplements of the non-collagenous proteins from wild type rat dentin failed to rescue the compromised mineralization, suggesting that the roles of FAM20C in tooth mineralization are more than phosphorylating local matrices and regulating systemic phosphorus metabolism. Moreover, the down-regulated BMP signaling pathways in the Fam20c deficient dental mesenchymal cells revealed that the kinase activity of FAM20C might be required to maintain BMP signaling. In summary, our study discloses that Fam20c indeed regulates cell behaviors and cell signaling pathway in a cell-autonomous manner.

Development of a new diaper dermatitis-like reconstructed skin equivalent for testing children atopic dermatitis relieving cosmetics.

BACKGROUND: Diaper dermatitis (DD) is the most common acute inflammatory skin disease. It has a serious effect on children's and families' quality of life. We aimed to screen and evaluate the efficacy of different formulas for relieving the diaper dermatitis symptoms by developing a kind of diaper dermatitis-like reconstructed human skin equivalent in vitro. MATERIALS AND METHOD: We developed the human skin equivalent for diaper dermatitis with 0.2% Sodium lauryl sulfate (SLS). The diaper dermatitis-like human skin equivalent was characterized by high level of inflammation, such as overexpression of interleukin-1α (IL-1α), and impaired skin barrier. Four formulas with potential of anti-inflammation and promotion of skin barrier function were topically applied on the diaper dermatitis-like human skin equivalent surface. The afterward protection efficacy was evaluated by endpoints of IL-1α, tissue viability, and skin barrier function. RESULTS: The chemical irritant induced high release of IL-1α, impaired tissue viability, and skin barrier function. The cream prepared with potential of anti-inflammation and skin protection could effectively decrease and relive the impact of irritant with decreased level of IL-1α and the higher tissue viability than the placebo exposure. CONCLUSION: The results showed that diaper dermatitis-like human skin equivalent induced by SLS can mimic the skin irritation response of the diaper rash.

FAM20C regulates osteoblast behaviors and intracellular signaling pathways in a cell-autonomous manner.

Recent studies indicate that Family with sequence similarity 20 member C (FAM20C) catalyzes the phosphorylation of secreted proteins, and participates in a variety of biological processes, including cell proliferation, migration, mineralization, and phosphate homeostasis. To explore the local influences of FAM20C on osteoblast, Fam20c-deficient osteoblasts were generated by treating the immortalized Fam20cf/f osteoblasts with CMV-Cre-IRES-EGFP lentivirus. Compared with the normal Fam20cf/f osteoblasts, the expression of Bone sialoprotein (Bsp), Osteocalcin (Ocn), Fibroblast growth factor 23 (Fgf23), and transcription factors that promote osteoblast maturation were up-regulated in the Fam20c-deficient osteoblasts. In contrast, the expression of Dental matrix protein 1 (Dmp1), Dentin sialophosphoprotein (Dspp), Osteopontin (Opn), type I Collagen a 1 (Col1a1), and Alkine phosphatase (Alp) were down-regulated in the Fam20c-deficient cells. These alterations disclosed the primary regulation of Fam20c on gene expression. The Fam20c-deficient osteoblasts showed a remarkable reduction in the ability of forming mineralized nodules. However, supplements of extracellular matrix proteins extracted from the normal bone failed to rescue the reduced mineralization, suggesting that FAM20C may affect the biomineralization by the means more than local phosphorylation of extracellular matrix proteins and systemic phosphorus homeostasis. Moreover, although Fam20c deficiency had little impact on cell proliferation, it significantly reduced cell migration and lowered the levels of p-Smad1/5/8, p-Erk and p-p38, suggesting that the kinase activity of FAM20C might be essential to cell mobility and the activity of BMP ligands. In summary, these findings provide evidences that FAM20C may regulate osteoblast maturation, migration, mineralization, and BMP signaling pathways in a cell-autonomous manner.

Inactivation of Fam20b in the neural crest-derived mesenchyme of mouse causes multiple craniofacial defects.

The glycosaminoglycan (GAG) chains attached to the core proteins of proteoglycans exert multiple roles, such as enriching signal molecules and regulating the binding of ligands to the corresponding receptors. A newly identified kinase - family with sequence similarity 20 member B (FAM20B) - is essential for the formation of GAG chains. The FAM20B protein phosphorylates the initial xylose on the side chain of a serine residue in the protein. Although the GAG chains of proteoglycans are believed to be indispensable during craniofacial development, there are few reports on their exact functions in craniofacial organogenesis. In this study, by mating Wnt1-cre mice with Fam20b-floxed mice (Fam20bflox/flox), we created Wnt1-Cre;Fam20bflox/flox mice in which Fam20b is ablated in the neural crest-derived mesenchyme. The Wnt1-Cre;Fam20bflox/flox mice died immediately after birth because of complete cleft palates. In addition to cleft palate, Wnt1-Cre;Fam20bflox/flox mice also manifested tongue elevation, micrognathia, microcephaly, suture widening, and reduced mineralization in the calvaria, facial bones, and temporomandibular joint. These findings indicate that the proteoglycans formed through the catalysis of FAM20B are essential for the morphogenesis and mineralization of the craniofacial complex.

Twist1 Is Essential for Tooth Morphogenesis and Odontoblast Differentiation.

Twist1 is a basic helix-loop-helix-containing transcription factor that is expressed in the dental mesenchyme during the early stages of tooth development. To better delineate its roles in tooth development, we generated Twist1 conditional knockout embryos (Twist2(Cre) (/+);Twist1(fl/fl)) by breeding Twist1 floxed mice (Twist1(fl/fl)) with Twist2-Cre recombinase knockin mice (Twist2(Cre) (/+)). The Twist2(Cre) (/+);Twist1(fl/fl) embryos formed smaller tooth germs and abnormal cusps during early tooth morphogenesis. Molecular and histological analyses showed that the developing molars of the Twist2(Cre) (/+);Twist1(fl/fl) embryos had reduced cell proliferation and expression of fibroblast growth factors 3, 4, 9, and 10 and FGF receptors 1 and 2 in the dental epithelium and mesenchyme. In addition, 3-week-old renal capsular transplants of embryonic day 18.5 Twist2(Cre) (/+);Twist1(fl/fl) molars showed malformed crowns and cusps with defective crown dentin and enamel. Immunohistochemical analyses revealed that the implanted mutant molars had defects in odontoblast differentiation and delayed ameloblast differentiation. Furthermore, in vitro ChIP assays demonstrated that Twist1 was able to bind to a specific region of the Fgf10 promoter. In conclusion, our findings suggest that Twist1 plays crucial roles in regulating tooth development and that it may exert its functions through the FGF signaling pathway.

The LPV Motif Is Essential for the Efficient Export of Secretory DMP1 From the Endoplasmic Reticulum.

Dentin matrix protein 1 (DMP1) is found abundantly in the extracellular matrices of bone and dentin. Secretory DMP1 begins with a tripeptide of leucine-proline-valine (LPV) after the endoplasmic reticulum (ER)-entry signal peptide is cleaved. The goal of this study was to determine the role of the LPV motif in the secretion of DMP1. A series of DNA constructs was generated to express various forms of DMP1 with or without the LPV motif. These constructs were transfected into a preosteoblast cell line, the MC3T3-E1 cells, and the subcellular localization and secretion of various forms of DMP1 were examined by immunofluorescent staining and Western-blotting analyses. Immunofluorescent staining showed that the LPV-containing DMP1 variants were primarily localized in the Golgi complex, whereas the LPV-lacking DMP1 variants were found abundantly within the ER. Western-blotting analyses demonstrated that the LPV-containing DMP1 variants were rapidly secreted from the transfected cells, as they did not accumulate within the cells, and the amounts increased in the conditioned media over time. In contrast, the LPV-lacking DMP1 variants were predominantly retained within the cells, and only small amounts were secreted out of the cells over time. These results suggest that the LPV motif is essential for the efficient export of secretory DMP1 from the ER to the Golgi complex.

Constitutive nuclear expression of dentin matrix protein 1 fails to rescue the Dmp1-null phenotype.

Dentin matrix protein 1 (DMP1) plays multiple roles in bone, tooth, phosphate homeostasis, kidney, salivary gland, reproductive cycles, and the development of cancer. In vitro studies have indicated two different biological mechanisms: 1) as a matrix protein, DMP1 interacts with αvß3 integrin and activates MAP kinase signaling; and 2) DMP1 serves as a transcription co-factor. In vivo studies have demonstrated its key role in osteocytes. This study attempted to determine whether DMP1 functions as a transcription co-factor and regulates osteoblast functions. For gene expression comparisons using adenovirus constructs, we targeted the expression of DMP1 either to the nucleus only by replacing the endogenous signal peptide with a nuclear localization signal (NLS) sequence (referred to as (NLS)DMP1) or to the extracellular matrix as the WT type (referred to as (SP)DMP1) in MC3T3 osteoblasts. High levels of DMP1 in either form greatly increased osteogenic gene expression in an identical manner. However, the targeted (NLS)DMP1 transgene driven by a 3.6-kb rat Col 1α1 promoter in the nucleus of osteoblasts and osteocytes failed to rescue the phenotyope of Dmp1-null mice, whereas the (SP)DMP1 transgene rescued the rickets defect. These studies support the notion that DMP1 functions as an extracellular matrix protein, rather than as a transcription co-factor in vivo. We also show that DMP1 continues its expression in osteoblasts during postnatal development and that the deletion of Dmp1 leads to an increase in osteoblast proliferation. However, poor mineralization in the metaphysis indicates a critical role for DMP1 in both osteoblasts and osteocytes.

Immortalized Mouse Floxed Fam20c Dental Papillar Mesenchymal and Osteoblast Cell Lines Retain Their Primary Characteristics.

Fam20c is essential for the normal mineralization of dentin and bone. The generation of odontoblast and osteoblast cell lines carrying floxed Fam20c allele can offer valuable tools for the study of the roles of Fam20c in the mineralization of dentin and bone. The limited capability of the primary odontoblasts and osteoblasts to proliferate necessitates the development of odontoblast and osteoblast cell lines serving as substitutes for the study of differentiation and mineralization of the odontoblasts and osteoblasts. In this study, we established and characterized immortalized mouse floxed Fam20c dental papilla mesenchymal and osteoblast cell lines. The isolated primary mouse floxed Fam20c dental papilla mesenchymal cells and osteoblasts were immortalized by the infection of lentivirus containing Simian Virus 40 T-antigen (SV40 T-Ag). The immortalization of floxed Fam20c dental papilla mesenchymal cells and osteoblasts was verified by the long-term passages and genomic integration of SV40 T-Ag. The immortalized floxed Fam20c dental papilla mesenchymal and osteoblast cell lines not only proliferated at a high rate and retained the morphology of their primary counterparts, but also preserved the dentin and bone specific gene expression as the primary dental papilla mesenchymal cells and osteoblasts did. Consistently, the capability of the primary floxed Fam20c dental papilla mesenchymal cells and osteoblasts to mineralize was also inherited by the immortalized dental papilla mesenchymal and osteoblast cell lines. Thus, we have successfully generated the immortalized mouse floxed Fam20c dental papilla mesenchymal and osteoblast cell lines.

Inactivation of a novel FGF23 regulator, FAM20C, leads to hypophosphatemic rickets in mice.

Family with sequence similarity 20,-member C (FAM20C) is highly expressed in the mineralized tissues of mammals. Genetic studies showed that the loss-of-function mutations in FAM20C were associated with human lethal osteosclerotic bone dysplasia (Raine Syndrome), implying an inhibitory role of this molecule in bone formation. However, in vitro gain- and loss-of-function studies suggested that FAM20C promotes the differentiation and mineralization of mouse mesenchymal cells and odontoblasts. Recently, we generated Fam20c conditional knockout (cKO) mice in which Fam20c was globally inactivated (by crossbreeding with Sox2-Cre mice) or inactivated specifically in the mineralized tissues (by crossbreeding with 3.6 kb Col 1a1-Cre mice). Fam20c transgenic mice were also generated and crossbred with Fam20c cKO mice to introduce the transgene in the knockout background. In vitro gain- and loss-of-function were examined by adding recombinant FAM20C to MC3T3-E1 cells and by lentiviral shRNA-mediated knockdown of FAM20C in human and mouse osteogenic cell lines. Surprisingly, both the global and mineralized tissue-specific cKO mice developed hypophosphatemic rickets (but not osteosclerosis), along with a significant downregulation of osteoblast differentiation markers and a dramatic elevation of fibroblast growth factor 23 (FGF23) in the serum and bone. The mice expressing the Fam20c transgene in the wild-type background showed no abnormalities, while the expression of the Fam20c transgene fully rescued the skeletal defects in the cKO mice. Recombinant FAM20C promoted the differentiation and mineralization of MC3T3-E1 cells. Knockdown of FAM20C led to a remarkable downregulation of DMP1, along with a significant upregulation of FGF23 in both human and mouse osteogenic cell lines. These results indicate that FAM20C is a bone formation "promoter" but not an "inhibitor" in mouse osteogenesis. We conclude that FAM20C may regulate osteogenesis through its direct role in facilitating osteoblast differentiation and its systemic regulation of phosphate homeostasis via the mediation of FGF23.

Loss of DMP1 causes rickets and osteomalacia and identifies a role for osteocytes in mineral metabolism.

The osteocyte, a terminally differentiated cell comprising 90%-95% of all bone cells, may have multiple functions, including acting as a mechanosensor in bone (re)modeling. Dentin matrix protein 1 (encoded by DMP1) is highly expressed in osteocytes and, when deleted in mice, results in a hypomineralized bone phenotype. We investigated the potential for this gene not only to direct skeletal mineralization but also to regulate phosphate (P(i)) homeostasis. Both Dmp1-null mice and individuals with a newly identified disorder, autosomal recessive hypophosphatemic rickets, manifest rickets and osteomalacia with isolated renal phosphate-wasting associated with elevated fibroblast growth factor 23 (FGF23) levels and normocalciuria. Mutational analyses showed that autosomal recessive hypophosphatemic rickets family carried a mutation affecting the DMP1 start codon, and a second family carried a 7-bp deletion disrupting the highly conserved DMP1 C terminus. Mechanistic studies using Dmp1-null mice demonstrated that absence of DMP1 results in defective osteocyte maturation and increased FGF23 expression, leading to pathological changes in bone mineralization. Our findings suggest a bone-renal axis that is central to guiding proper mineral metabolism.

The rescue of dentin matrix protein 1 (DMP1)-deficient tooth defects by the transgenic expression of dentin sialophosphoprotein (DSPP) indicates that DSPP is a downstream effector molecule of DMP1 in dentinogenesis.

Dentin matrix protein 1 (DMP1) and dentin sialophosphoprotein (DSPP) are essential for the formation of dentin. Previous in vitro studies have indicated that DMP1 might regulate the expression of DSPP during dentinogenesis. To examine whether DMP1 controls dentinogenesis through the regulation of DSPP in vivo, we cross-bred transgenic mice expressing normal DSPP driven by a 3.6-kb rat Col1a1 promoter with Dmp1 KO mice to generate mice expressing the DSPP transgene in the Dmp1 KO genetic background (referred to as "Dmp1 KO/DSPP Tg mice"). We used morphological, histological, and biochemical techniques to characterize the dentin and alveolar bone of Dmp1 KO/DSPP Tg mice compared with Dmp1 KO and wild-type mice. Our analyses showed that the expression of endogenous DSPP was remarkably reduced in the Dmp1 KO mice. Furthermore, the transgenic expression of DSPP rescued the tooth and alveolar bone defects of the Dmp1 KO mice. In addition, our in vitro analyses showed that DMP1 and its 57-kDa C-terminal fragment significantly up-regulated the Dspp promoter activities in a mesenchymal cell line. In contrast, the expression of DMP1 was not altered in the Dspp KO mice. These results provide strong evidence that DSPP is a downstream effector molecule that mediates the roles of DMP1 in dentinogenesis.

Overexpression of Dmp1 fails to rescue the bone and dentin defects in Fam20C knockout mice.

FAM20C is a kinase phosphorylating the small-integrin-binding ligand, N-linked glycoproteins (SIBLINGs), a group of extracellular matrix proteins that are essential for bone and dentin formation. Previously, we showed that Sox2-Cre;Fam20Cfl/fl mice had bone and dentin defects, along with hypophosphatemia and significant downregulation of dentin matrix protein 1 (DMP1). While the assumed phosphorylation failure of the SIBLINGs is likely associated with the defects in the Fam20C-deficient mice, it remains unclear if the downregulation of Dmp1 contributes to these phenotypes. In this study, we crossed 3.6 kb Col1-Dmp1 transgenic mice with 3.6 kb Col1-Cre;Fam20Cfl/fl mice to overexpress Dmp1 in the mineralized tissues of Fam20C conditional knockout (cKO) mice. X-ray, micro-computed tomography, serum biochemistry and histology analyses showed that expressing the Dmp1 transgene failed to rescue the bone and dentin defects, as well as the serum levels of FGF23 and phosphate in the Fam20C-cKO mice. These results indicated that the downregulation of Dmp1 may not directly associate with, or significantly contribute to the bone and dentin defects in the Fam20C-cKO mice.

FAM20A is a golgi-localized Type II transmembrane protein.

Family with sequence similarity 20, member A (FAM20A) is a pseudo-kinase in the secretory pathway and is essential for enamel formation in humans. Here we examine if FAM20A is a membrane-associated protein. We show that the full-length FAM20A can be purified from HEK293 cells transfected with a FAM20A-expresing construct. Further, it is only found in the membrane fraction, but not in the soluble fraction, of cell lysate. Consistently, it is not secreted out of the expressing cells. Moreover, it is co-localized with GM130, a cis-Golgi network marker, and membrane topology analysis indicates that it has its C-terminus oriented towards the lumen of the organelle. Our results support that FAM20A is a Type II transmembrane protein within the secretory compartments.

Quorum-Sensing Signal DSF Inhibits the Proliferation of Intestinal Pathogenic Bacteria and Alleviates Inflammatory Response to Suppress DSS-Induced Colitis in Zebrafish.

The imbalance of gut microbiota is an important factor leading to inflammatory bowel disease (IBD). Diffusible signal factor (DSF) is a novel quorum-sensing signal that regulates bacterial growth, metabolism, pathogenicity, and host immune response. This study aimed to explore the therapeutic effect and underlying mechanisms of DSF in a zebrafish colitis model induced by sodium dextran sulfate (DSS). The results showed that intake of DSF can significantly improve intestinal symptoms in the zebrafish colitis model, including ameliorating the shortening of the intestine, reducing the increase in the goblet cell number, and restoring intestinal pathological damage. DSF inhibited the upregulation of inflammation-related genes and promoted the expression of claudin1 and occludin1 to protect the tightness of intestinal tissue. The gut microbiome analysis demonstrated that DSF treatment helped the gut microbiota of the zebrafish colitis model recover to normal at the phylum and genus levels, especially in terms of pathogenic bacteria; DSF treatment downregulated the relative abundance of Aeromonas hydrophila and Staphylococcus aureus, and it was confirmed in microbiological experiments that DSF could effectively inhibit the colonization and infection of these two pathogens in the intestine. This study suggests that DSF can alleviate colitis by inhibiting the proliferation of intestinal pathogens and inflammatory responses in the intestine. Therefore, DSF has the potential to become a dietary supplement that assists in the antibiotic and nutritional treatment of IBD.