Uterine Sensitization-associated Gene-1 Research Articles

MO425: Rosuvastatin Activates Hox13-Usag-1 Pathway and Prevents Renal Fibrosis

Abstract BACKGROUND AND AIMS Statins, 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors, are primarily cholesterol-lowering drugs that have become standard of care in the primary and secondary prevention of cardiovascular diseases. Apart from lipid-lowering, statins may act beneficially through anti-fibrotic mechanisms to protect the diseased kidney. This study evaluated anti-fibrotic effects of rosuvastatin (RSV) in a chronic kidney fibrosis model and against the TGF-ß1 stimulated Madin-Darby canine kidney (MDCK) cells in vitro. METHOD Mice subjected to unilateral ischemic reperfusion injury with contralateral nephrectomy (uIRIx) were treated with vehicle or RSV (10 mg/kg, by oral gavage) daily for 4 weeks and kidneys were analyzed for markers of fibrosis, bone morphogenetic protein-7 (BMP-7), uterine sensitization-associated gene-1 (USAG-1), and SMAD signaling. Control and homeobox protein Hox-A13 (HOXA13) knocked down MDCK cells were stimulated with TGF- ß1 (5 ng/ml) and then treated with RSV. RESULTS Kidneys from uIRIx mice showed increased expression of α-SMA, Collagen 1 and decrease in BMP-7 (20.35 ± 2.37 versus 1.00 ± 0.27, P < 0.05; 8.43 ± 1.55 versus 1.00 ± 0.35, P < 0.05; 0.75 ± 0.06 versus 1.00 ± 0.15, P < 0.05, respectively). In contrast, expression of USAG-1, a BMP-7 antagonist, was markedly increased in fibrotic kidney (6.60 ± 1.11 versus 1.00 ± 0.02, P < 0.05). Interestingly, RSV treatment not only attenuated expression of USAG-1 (0.72 ± 0.08 versus 1.36 ± 0.14, P < 0.05) but also showed a tendency to activate expression of HOXA13 (0.36 ± 0.14 versus 0.51 ± 0.15, P > 0.05) and improved other markers of fibrosis. Moreover, RSV treatment significantly reduced phosphorylated Smad3 (3.94 ± 0.81 versus 7.17 ± 1.50, P < 0.05) and increased phosphorylation levels of Smad 1/5/9 (0.67 ± 0.10 versus 0.30 ± 0.09, P < 0.05) that is associated with BMP-7 signaling in the fibrotic kidney. MDCK cells stimulated with TGF-β1 in vitro showed increased expression of α-SMA, fibronectin, vimentin, collagen 1, USAG-1, and phosphorylation of Smad3 as well as decreased expression of phosphorylated Smad 1/5/9. RSV treatment significantly reversed these changes as well as increased level of transcriptional factor HOXA13 (0.89 ± 0.12 versus 0.33 ± 0.08, P < 0.05), which negatively regulates USAG-1, without changes in BMP-7 expression. In addition, effect of RSV treatment on USAG-1 expression was significantly decreased in HOXA13 gene knocked down MDCK cells (1.01 ± 0.19 versus 2.04 ± 0.38, P < 0.05; 1.66 ± 0.13 versus 2.04 ± 0.38, P > 0.05; TGF- ß1 + RSV versus TGF- ß1 and TGF- ß1 + RSV + siRNA vs TGF- ß1, respectively). These in vitro data suggest that RSV enhanced anti-fibrotic pathway by downregulating a dominant BMP-7 antagonist USAG-1 via HOXA13 upregulation and not by enhancing BMP-7 production. CONCLUSION The present results demonstrate that RSV inhibits the progression of kidney fibrosis in part by upregulating BMP-7-mediated signaling via HOXA13 expression and down regulation of USAG-1.

Uterine Sensitization-Associated Gene-1 in the Progression of Kidney Diseases.

Uterine sensitization-associated gene-1 (USAG-1), originally identified as a secretory protein preferentially expressed in the sensitized rat endometrium, has been determined to modulate bone morphogenetic protein (BMP) and Wnt expression to play important roles in kidney disease. USAG-1 affects the progression of acute and chronic kidney damage and the recovery of allograft kidney function by regulating the BMP and Wnt signaling pathways. Moreover, USAG-1 has been found to be involved in the process of T cell immune response, and its ability to inhibit germinal center activity and reduce humoral immunity is of great significance for the treatment of autoimmune nephropathy and antibody-mediated rejection (AMR) after renal transplantation. This article summarizes the many advances made regarding the roles of USAG-1 in the progression of kidney disease and outlines potential treatments.

Local application of Usag-1 siRNA can promote tooth regeneration in Runx2-deficient mice

Runt-related transcription factor 2 (Runx2)-deficient mice can be used to model congenital tooth agenesis in humans. Conversely, uterine sensitization-associated gene-1 (Usag-1)-deficient mice exhibit supernumerary tooth formation. Arrested tooth formation can be restored by crossing both knockout-mouse strains; however, it remains unclear whether topical inhibition of Usag-1 expression can enable the recovery of tooth formation in Runx2-deficient mice. Here, we tested whether inhibiting the topical expression of Usag-1 can reverse arrested tooth formation after Runx2 abrogation. The results showed that local application of Usag-1 Stealth small interfering RNA (siRNA) promoted tooth development following Runx2 siRNA-induced agenesis. Additionally, renal capsule transplantation of siRNA-loaded cationized, gelatin-treated mouse mandibles confirmed that cationized gelatin can serve as an effective drug-delivery system. We then performed renal capsule transplantation of wild-type and Runx2-knockout (KO) mouse mandibles, treated with Usag-1 siRNA, revealing that hindered tooth formation was rescued by Usag-1 knockdown. Furthermore, topically applied Usag-1 siRNA partially rescued arrested tooth development in Runx2-KO mice, demonstrating its potential for regenerating teeth in Runx2-deficient mice. Our findings have implications for developing topical treatments for congenital tooth agenesis.

Anti-USAG-1 therapy for tooth regeneration through enhanced BMP signaling.

Uterine sensitization-associated gene-1 (USAG-1) deficiency leads to enhanced bone morphogenetic protein (BMP) signaling, leading to supernumerary teeth formation. Furthermore, antibodies interfering with binding of USAG-1 to BMP, but not lipoprotein receptor-related protein 5/6 (LRP5/6), accelerate tooth development. Since USAG-1 inhibits Wnt and BMP signals, the essential factors for tooth development, via direct binding to BMP and Wnt coreceptor LRP5/6, we hypothesized that USAG-1 plays key regulatory roles in suppressing tooth development. However, the involvement of USAG-1 in various types of congenital tooth agenesis remains unknown. Here, we show that blocking USAG-1 function through USAG-1 knockout or anti-USAG-1 antibody administration relieves congenital tooth agenesis caused by various genetic abnormalities in mice. Our results demonstrate that USAG-1 controls the number of teeth by inhibiting development of potential tooth germs in wild-type or mutant mice missing teeth. Anti-USAG-1 antibody administration is, therefore, a promising approach for tooth regeneration therapy.

Febuxostat inhibits TGF‑β1‑induced epithelial‑mesenchymal transition via downregulation of USAG‑1 expression in Madin‑Darby canine kidney cells invitro.

Our previous study demonstrated that febuxostat, a xanthine oxidase inhibitor, can alleviate kidney dysfunction and ameliorate renal tubulointerstitial fibrosis in a rat unilateral ureteral obstruction (UUO) model; however, the underlying mechanisms remain unknown. Increasing evidence has revealed that epithelial-mesenchymal transition (EMT) is one of the key mechanisms mediating the progression of renal tubulointerstitial fibrosis in chronic kidney disease (CKD). Uterine sensitization-associated gene-1 (USAG-1), a kidney-specific bone morphogenetic protein antagonist, is involved in the development of numerous types of CKDs. The present study aimed to investigate the role of febuxostat in the process of EMT in Madin-Darby canine kidney (MDCK) cells in vitro. Western blotting, reverse transcription-semiquantitative polymerase chain reaction analysis and immunofluorescence staining were used to evaluate the expression levels of bone morphogenetic protein 7, USAG-1, α-smooth muscle actin (α-SMA) and E-cadherin, respectively. The results demonstrated that the expression of USAG-1 and α-SMA increased, and that of E-cadherin decreased significantly in MDCK cells following treatment with transforming growth factor-β1 (TGF-β1). The application of small interfering RNA-USAG-1 potently inhibited TGF-β1-induced EMT. Subsequently, the effects of febuxostat on TGF-β1-induced EMT was investigated. The results demonstrated that febuxostat downregulated the expression of USAG-1, and reversed TGF-β1-induced EMT in MDCK cells. Furthermore, pretreatment with febuxostat significantly restored the decreased expression levels of phosphorylated Smad1/5/8 induced by TGF-β1 in MDCK cells. The results of the present study suggested that USAG-1 may be involved in the EMT process of MDCK cells induced by TGF-β1, and febuxostat inhibited EMT by activating the Smad1/5/8 signaling pathway via downregulating the expression of USAG-1 in MDCK cells.

Artesunate attenuates unilateral ureteral obstruction-induced renal fibrosis by regulating the expressions of bone morphogenetic protein-7 and uterine sensitization-associated gene-1 in rats.

Kidney fibrosis is the most common final stage of progressive renal disease. Bone morphogenetic protein-7 (BMP-7) has been shown to be important in both preservation of kidney function and resistance to injury. Recently, it has been realized that uterine sensitization-associated gene-1 (USAG-1) functions as a kidney-specific BMP antagonist. Because of the reported anti-fibrotic effects of artesunate (Art), this study was designed to investigate the effects of Art on renal fibrosis in unilateral ureteral obstruction (UUO) rats and to explore the underlying mechanisms. Thirty male Sprague-Dawley rats were randomly divided into five groups: Sham group, UUO group, low-dose Art-treated (Art-L), middle-dose Art-treated (Art-M), and high-dose Art-treated (Art-H) groups. The UUO rat model was established by ligating the left ureter. Fourteen days later, interstitial collagen deposition, expression of USAG-1, BMP-7, E-cadherin, α-smooth muscle actin (α-SMA), fibronectin (FN), collagen I, as well as the inflammatory infiltration levels in the kidneys were assessed. Art treatment significantly attenuated the deposition of interstitial collagens in the UUO rats' kidneys and exhibited the ability to improve renal function, followed by the up-regulated expression of BMP-7 and E-cadherin and the down-regulated expression of USAG-1 and α-SMA. In addition, increased macrophages infiltration in the kidneys of the UUO rats were also attenuated by the administration of Art. These results indicate that Art is able to improve the renal function decline and renal fibrosis induced by UUO, which may be associated with the up-regulation of BMP-7 and down-regulation of USAG-1. Accordingly, Art may become a potential preventive or therapeutic agent for chronic kidney diseases.

Febuxostat Prevents Renal Interstitial Fibrosis by the Activation of BMP-7 Signaling and Inhibition of USAG-1 Expression in Rats

Background: Renal interstitial fibrosis (RIF) is a common pathology associated with end-stage renal diseases. The activation of bone morphogenetic protein-7 (BMP-7)-Smad1/5/8 pathway seems to alleviate RIF. Uterine sensitization-associated gene-1 (USAG-1), a kidney-specific BMPs antagonist, is associated with the development and prognosis of several renal diseases. Febuxostat is a xanthine oxidase inhibitor that can attenuate the renal dysfunction of patients. The purpose of this study was to investigate the effects of febuxostat on renal fibrosis and to clarify the mechanisms underlying these effects. Methods: Rats were randomly divided into 6 groups termed a sham-operated group, a unilateral ureteral obstruction (UUO) group, 3 doses of febuxostat groups (low, intermediate and high doses) and a sham group treated with high-dose febuxostat. After 14 days, renal function, relative kidney weight, accumulation of glycogen and collagens were examined by different methods. Expression of α-SMA, transforming growth factor-β1 (TGF-β1), BMP-7 and USAG-1 was detected by western blotting and RT-PCR, respectively. The phosphorylation level of Smad1/5/8 was also quantified by western blotting. Results: The renal function was declined, and large amounts of glycogen and collagens were deposited in the kidneys of UUO rats compared with the rats in the sham group. Besides, expression of α-SMA and USAG-1 in these kidneys was elevated, and the TGF-β1 was also activated, while the BMP-7-Smad1/5/8 pathway was inhibited. Febuxostat reversed the changes stated earlier, exhibiting protective effects on RIF induced by UUO. Conclusion: Febuxostat was able to attenuate RIF caused by UUO, which was associated with the activation of BMP-7-Smad1/5/8 pathway and the inhibition of USAG-1 expression in the kidneys of UUO rats.

Interactions between BMP-7 and USAG-1 (uterine sensitization-associated GENE-1) regulate formations of supernumerary maxillary incisors

Interactions between BMP-7 and USAG-1 (uterine sensitization-associated GENE-1) regulate formations of supernumerary maxillary incisors

Interactions between BMP-7 and USAG-1 (uterine sensitization-associated gene-1) regulate supernumerary organ formations.

Bone morphogenetic proteins (BMPs) are highly conserved signaling molecules that are part of the transforming growth factor (TGF)-beta superfamily, and function in the patterning and morphogenesis of many organs including development of the dentition. The functions of the BMPs are controlled by certain classes of molecules that are recognized as BMP antagonists that inhibit BMP binding to their cognate receptors. In this study we tested the hypothesis that USAG-1 (uterine sensitization-associated gene-1) suppresses deciduous incisors by inhibition of BMP-7 function. We learned that USAG-1 and BMP-7 were expressed within odontogenic epithelium as well as mesenchyme during the late bud and early cap stages of tooth development. USAG-1 is a BMP antagonist, and also modulates Wnt signaling. USAG-1 abrogation rescued apoptotic elimination of odontogenic mesenchymal cells. BMP signaling in the rudimentary maxillary incisor, assessed by expressions of Msx1 and Dlx2 and the phosphorylation of Smad protein, was significantly enhanced. Using explant culture and subsequent subrenal capsule transplantation of E15 USAG-1 mutant maxillary incisor tooth primordia supplemented with BMP-7 demonstrated in USAG-1+/− as well as USAG-1−/− rescue and supernumerary tooth development. Based upon these results, we conclude that USAG-1 functions as an antagonist of BMP-7 in this model system. These results further suggest that the phenotypes of USAG-1 and BMP-7 mutant mice reported provide opportunities for regenerative medicine and dentistry.

3-Hydroxy-3-methylglutaryl-coenzyme A reductase inhibitor simvastatin ameliorates renal fibrosis through HOXA13–USAG-1 pathway

Epidemiological data have suggested that 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors (statins) prevent the progression of chronic kidney diseases (CKDs), whereas the precise mechanism explaining in vitro to in vivo is missing. This study is aimed at exploring a new mechanism of action by statins on renal fibrosis, a hallmark of CKD, using mouse renal fibrosis model in vivo and Madin–Darby canine kidney (MDCK) cells expressing USAG-1 in vitro. C57/BL6 mice fed a 0.2% adenine-containing diet for 4 weeks developed renal dysfunction accompanied with severe tubulointerstitial fibrosis. Subsequent simvastatin (SIM) treatment (50 mg/kg per day) for 2 weeks significantly suppressed fibrosis progression. We found that SIM enhanced bone morphogenetic protein-7 (BMP-7)-mediated anti-fibrotic signaling with the reduced expression of uterine sensitization-associated gene-1 (USAG-1), a BMP-7 antagonist produced by renal distal tubular epithelial cells. Therefore, MDCK cells were incubated with transforming growth factor-β1 and showed increased expression of USAG-1 and α-smooth muscle actin; SIM significantly reduced them. SIM significantly increased E-cadherin expression. Gene knockdown experiments using MDCK suggested that homeobox protein Hox-A13 (HOXA13) played a suppressive role in the USAG-1 gene and thus SIM reduced USAG-1 by increasing HOXA13 expression. The data from our study demonstrate that SIM, one of statins, contributes to prevent the progression of renal fibrosis by upregulating BMP-7-mediated anti-fibrotic signaling and that one aspect of crucial efficacies is achieved by regulating HOXA13 and USAG-1. HOXA13–USAG-1 pathway is a newly identified mechanism in renal fibrosis and will be a new therapeutic target for preventing renal fibrosis progression in CKDs.

Simvastatin ameliorates renal fibrosis via suppressing uterine sensitization‐associated gene‐1 (USAG‐1), a BMP‐7 antagonist, in mouse adenine‐induced tubulointerstitial injury model

Background & PurposeIt is known that BMP‐7 counteracts with TGF‐β, the dominant molecule promoting renal fibrosis, and that uterine sensitization‐associated gene‐1(USAG‐1), the dominant BMP‐7 antagonist expressing in distal tubular epithelial cells, promotes renal fibrosis. In this study, we investigated whether simvastatin (SIM), one of the HMG‐CoA reductase inhibitors, attenuates renal fibrosis by modulating BMP‐7 and USAG‐1 mediated pathway.Method & ResultsC57/BL6 mice fed by 0.2% adenine‐containing diet for 4 weeks showed significant increase of blood urea nitrogen (BUN) accompanied with severe tubulointerstitial fibrosis. SIM treatment for two weeks (50mg/kg/day) significantly reduced BUN compared with the control group (p<0.01). In pathological analysis, the area of fibrosis and the expression of alpha smooth muscle actin were significantly attenuated by SIM (p<0.05). In quantitative real‐time PCR, USGA‐1 expression was significantly lower in the SIM group (p<0.05). MDCK cells (distal epithelial tubular cells) incubated with TGF‐β (5ng/ml) for 72 hours showed increased expression of USAG‐1. SIM significantly reduced USAG‐1 expression (p<0.01). Several database analyses predicted the binding sites of HOXA‐13, a transcriptional factor, in the promoter region of USAG‐1 gene. The expression assay in vitro suggested that HOXA13 functions as a suppressor of the USAG‐1 gene and SIM decreased USAG‐1 expression by enhancing HOXA13.ConclusionOur data showed that SIM ameliorated renal fibrosis by suppressing USAG‐1. Decreased USAG‐1 levels may enhance the counteract effect of BMP‐7 against TGF‐β. We also demonstrated that HOXA13 plays a crucial role in the mechanism of this action by SIM.

Loss of the BMP antagonist USAG-1 ameliorates disease in a mouse model of the progressive hereditary kidney disease Alport syndrome

The glomerular basement membrane (GBM) is a key component of the filtering unit in the kidney. Mutations involving any of the collagen IV genes (COL4A3, COL4A4, and COL4A5) affect GBM assembly and cause Alport syndrome, a progressive hereditary kidney disease with no definitive therapy. Previously, we have demonstrated that the bone morphogenetic protein (BMP) antagonist uterine sensitization-associated gene-1 (USAG-1) negatively regulates the renoprotective action of BMP-7 in a mouse model of tubular injury during acute renal failure. Here, we investigated the role of USAG-1 in renal function in Col4a3-/- mice, which model Alport syndrome. Ablation of Usag1 in Col4a3-/- mice led to substantial attenuation of disease progression, normalization of GBM ultrastructure, preservation of renal function, and extension of life span. Immunohistochemical analysis revealed that USAG-1 and BMP-7 colocalized in the macula densa in the distal tubules, lying in direct contact with glomerular mesangial cells. Furthermore, in cultured mesangial cells, BMP-7 attenuated and USAG-1 enhanced the expression of MMP-12, a protease that may contribute to GBM degradation. These data suggest that the pathogenetic role of USAG-1 in Col4a3-/- mice might involve crosstalk between kidney tubules and the glomerulus and that inhibition of USAG-1 may be a promising therapeutic approach for the treatment of Alport syndrome.

Reversal of experimental renal fibrosis by BMP7 provides insights into novel therapeutic strategies for chronic kidney disease

Bone morphogenic protein-7 (BMP7) is a morphogen that is important for kidney development and which is also an integral part of the kidney's physiological response to repair of acute kidney injury. Several studies demonstrate that preexisting renal BMP7 pathways can be utilized by administering recombinant BMP7 to protect the kidney in experimental models of chronic kidney disease (CKD). Effectiveness of recombinant BMP7 in animal studies raises the possibility that the BMP7 pathway could be equally utilized to treat patients with CKD and interstitial fibrosis. However, regulation of BMP7 activity in the kidney is complex. BMP7 activity in the kidney is not only determined by availability of BMP7 itself, but also by a balance of agonists, such as Kielin/chordin-like protein (KCP) or BMP receptors, and antagonists including gremlin, noggin, or uterine sensitization-associated gene-1 (USAG-1). Presence of BMP7 agonists and antagonists has to be considered when recombinant BMP7 is supplemented to treat injured kidneys. Here we summarize recent insights into the role of BMP7 in acute and chronic kidney injury and discuss the implications for future directions of antifibrotic therapies.

Enhanced BMP signaling results in supernumerary tooth formation in USAG-1 deficient mouse

Uterine sensitization associated gene-1 (USAG-1) is a BMP antagonist, and also modulates Wnt signaling. We previously reported that USAG-1 deficient mice have supernumerary teeth. The supernumerary maxillary incisor appears to form as a result of the successive development of the rudimentary upper incisor. USAG-1 abrogation rescued apoptotic elimination of odontogenic mesenchymal cells. We confirmed that BMPs were expressed in both the epithelium and mesenchyme of the rudimentary incisor at E14 and E15. BMP signaling in the rudimentary maxillary incisor, assessed by expressions of Msx1 and Dlx2 and the phosphorylation of Smad protein, was significantly enhanced. Wnt signaling as demonstrated by the nuclear localization of β-catenin was also up-regulated. Inhibition of BMP signaling rescues supernumerary tooth formation in E15 incisor explant culture. Based upon these results, we conclude that enhanced BMP signaling results in supernumerary teeth and BMP signaling was modulated by Wnt signaling in the USAG-1 deficient mouse model.

Expression of BMP-7 and USAG-1 (a BMP antagonist) in kidney development and injury

Once developed, end-stage renal disease cannot be reversed by any current therapy. Bone morphogenetic protein-7 (BMP-7), however, is a possible treatment for reversing end-stage renal disease. Previously, we showed that the BMP antagonist uterine sensitization-associated gene-1 (USAG-1, also known as ectodin and sclerostin domain-containing 1) negatively regulates the renoprotective action of BMP-7. Here, we show that the ratio between USAG-1 and BMP-7 expression increased dramatically in the later stage of kidney development, with USAG-1 expression overlapping BMP-7 only in differentiated distal tubules. Examination of USAG-1 expression in developing kidney indicated that a mosaic of proximal and distal tubule marker-positive cells reside side by side in the immature nephron. This suggests that each cell controls its own fate for becoming a proximal or distal tubule cell. In kidney injury models, the ratio of USAG-1 to BMP-7 expression decreased with kidney damage but increased after subsequent kidney regeneration. Our study suggests that USAG-1 expression in a kidney biopsy could be useful in predicting outcome.

Balance Between Bone Morphogenetic Proteins and Their Antagonists in Kidney Injury

Dialysis dependency is one of the leading causes of morbidity and mortality in the world, and once end-stage renal disease develops, it cannot be reversed by currently available therapy. Although the administration of large dose of bone morphogenetic protein-7 (BMP-7) has been shown to repair established renal injuries and improves renal function, pathophysiological role of endogenous BMP-7 and regulatory mechanism of its activities remain elusive. Here we show that uterine sensitization-associated gene-1 (USAG-1), novel BMP antagonist abundantly expressed in the kidney, is the central negative regulator of BMP function in the kidney, and that mice lacking USAG-1 (USAG-1(-/-) mice) are resistant to kidney injuries. USAG-1(-/-) mice exhibited prolonged survival and preserved renal function in acute and chronic renal injuries. Renal BMP signaling, assessed by phosphorylation of Smad proteins, is significantly enhanced in USAG-1(-/-) mice during renal injury, indicating that the preservation of renal function is attributed to enhancement of endogenous BMP signaling. Furthermore, the administration of neutralizing antibody against BMP-7 abolished reno-protection in USAG-1(-/-) mice, indicating that USAG-1 plays a critical role in the modulation of reno-protective action of BMP, and that inhibition of USAG-1 will be promising means of development of novel treatment for kidney diseases.

Rudiment incisors survive and erupt as supernumerary teeth as a result of USAG-1 abrogation

The term “supernumerary teeth” describes production of more than the normal number of teeth in the primary or permanent dentitions. Their aetiology is not understood. Uterine sensitization associated gene-1 (USAG-1) is a BMP antagonist that plays important roles in the local regulation of BMP signaling by binding and neutralizing BMP activities [M. Yanagita, BMP antagonists: their roles in development and involvement in pathophysiology, Cytokine Growth Factor Rev. 16 (2005) 309–317], and also serves as a modulator of Wnt signaling [N. Itasaki, C.M. Jones, S. Mercurio, A. Rowe, P.M. Domingos, J.C. Smith, R. Krumlauf, Wise, a context-dependent activator and inhibitor of Wnt signaling, Development 130 (2003) 4295–4305]. We report here that USAG-1 deficient mice have supernumerary teeth. The supernumerary maxillary incisor appears to form as a result of the successive development of the rudimentary upper incisor tooth. We confirmed that the USAG-1 expression is localized to the epithelium and mesenchyme of the rudimentary maxillary incisor tooth organ formation. USAG-1 abrogation rescued apoptotic elimination of odontogenic mesenchymal cells. Based upon these results, we conclude that USAG-1 controls the number of teeth in the maxillary incisor region by regulating apoptosis.

Expression of Uterine Sensitization-Associated Gene-1 (USAG-1) in the Mouse Uterus During the Peri-Implantation Period

Rat uterine sensitization-associated gene-1 (USAG-1) mRNA is expressed in the uterus during the peri-implantation period, and its mRNA expression in uterine epithelial cells is highest on day 5 of pregnancy. On the other hand, since changes in USAG-1 mRNA expression in the mouse uterus are not seen during the estrous cycle, USAG-1 expression might be specifically regulated by embryonic factors rather than by the maternal environment. However, the expression pattern and function of USAG-1 in the mouse uterus have not been determined. Thus, we examined the tissue-specific USAG-1 mRNA expression in the uteri of ICR mice during peri-implantation using real-time quantitative PCR. Uterine tissues, such as the myometrium, luminal epithelium, and stroma, were collected by laser capture microdissection at 3.5-6.5 dpc. USAG-1 mRNA was expressed in the uteri of pregnant mice from 3.5 dpc to 6.5 dpc, and the highest level of expression was seen at 4.5 dpc (P<0.01). Significantly high USAG-1 mRNA expression was detected in the luminal epithelium at 4.5 dpc (P<0.05). The stroma and myometrium exhibited unchanged expression levels of USAG-1 mRNA at 3.5-5.5 dpc. USAG-1 mRNA was undetectable in blastocysts and implanting embryos. Expression of USAG-1 mRNA appears to be associated with blastocyst implantation to the luminal epithelium, suggesting that physiological or biochemical contact of the blastocyst to the uterus is required for USAG-1 expression.

Modulator of bone morphogenetic protein activity in the progression of kidney diseases

Tubular damage and interstitial fibrosis is a final common pathway leading to end-stage renal disease, and once tubular damage is established, it cannot be reversed by currently available treatment. The administration of bone morphogenetic protein-7 (BMP-7) in pharmacological doses repairs established tubular damages and improves renal function in several kidney disease models; however, pathophysiological role of endogenous BMP-7 and regulatory mechanism of its activities remain elusive. The activity of BMP is precisely regulated by certain classes of molecules termed BMP agonist/antagonist. In this review, roles of BMP agonist/antagonists possibly modulating the activity of BMP in kidney diseases are discussed. Our group demonstrated that uterine sensitization-associated gene-1 (USAG-1), a novel BMP antagonist abundantly expressed in the kidney, is the central negative regulator of BMP-7 in the kidney, and that mice lacking USAG-1 (USAG-1(-/-) mice) are resistant to kidney injuries. USAG-1(-/-) mice exhibited markedly prolonged survival and preserved renal function in acute and chronic renal injuries. Renal BMP signaling, assessed by phosphorylation of Smad proteins, is significantly enhanced in USAG-1(-/-) mice during renal injury, indicating that the preservation of renal function is attributed to enhancement of endogenous BMP-7 signaling. Furthermore, the administration of neutralizing antibody against BMP-7 abolished renoprotection in USAG-1(-/-) mice, indicating that USAG-1 plays a critical role in the modulation of renoprotective action of BMP, and that inhibition of USAG-1 will be promising means of development of novel treatment for kidney diseases.

Uterine sensitization-associated gene-1 (USAG-1), a novel BMP antagonist expressed in the kidney, accelerates tubular injury

Dialysis dependency is one of the leading causes of morbidity and mortality in the world, and once end-stage renal disease develops, it cannot be reversed by currently available therapy. Although administration of large doses of bone morphogenetic protein-7 (BMP-7) has been shown to repair established renal injury and improve renal function, the pathophysiological role of endogenous BMP-7 and regulatory mechanism of its activities remain elusive. Here we show that the product of uterine sensitization-associated gene-1 (USAG1), a novel BMP antagonist abundantly expressed in the kidney, is the central negative regulator of BMP function in the kidney and that mice lacking USAG-1 (USAG1 mice) are resistant to renal injury. USAG1 mice exhibited prolonged survival and preserved renal function in acute and chronic renal injury models. Renal BMP signaling, assessed by phosphorylation of Smad proteins, was significantly enhanced in USAG1 mice with renal injury, indicating that the preservation of renal function is attributable to enhancement of endogenous BMP signaling. Furthermore, the administration of neutralizing antibody against BMP-7 abolished renoprotection in USAG1 mice, indicating that USAG-1 plays a critical role in the modulation of renoprotective action of BMP and that inhibition of USAG-1 is a promising means of development of novel treatment for renal diseases.