Therapeutic effects of the extract of Sancao Formula, a Chinese herbal compound, on imiquimod-induced psoriasis via cysteine-rich protein 61.

OBJECTIVE: Psoriasis is a common chronic inflammatory skin disease that is prone to recurrence, and the proinflammatory factor, cysteine-rich protein 61 (Cyr61), is important in its pathophysiology. Long-term clinical practice has shown that Sancao Formula (SC), a Chinese herbal compound, is effective in the treatment of psoriasis, but the precise mechanism remains unknown. In this study, we investigate the mechanism by which SC extract alleviates imiquimod (IMQ)-induced psoriasis. METHODS: The expression of Cyr61 in psoriatic lesions and normal healthy skin was detected using immunohistochemical analysis to investigate the biological role of Cyr61 in models of psoriatic inflammation. A psoriatic mouse model was established by topical application of IMQ, and the effect of topical application of SC extract was evaluated using the psoriasis area and severity index (PASI) score, hematoxylin-eosin staining, and histopathological features of the skin. Next, a HaCaT cell inflammation model was established using interferon-γ (IFN-γ), and the effect of SC extract on the mRNA and protein levels of Cyr61 and intercellular cell adhesion molecule-1 (ICAM-1) was confirmed using Western blot and quantitative real-time polymerase chain reaction analyses. RESULTS: Immunohistochemical staining showed that the expression of Cyr61 in psoriatic lesions was higher than that in normal skin samples (78.26% vs 41.18%, P < 0.05), and the number of Cyr61-positive cells in psoriatic lesions was also significantly higher than in normal skin (18.66 ± 2.51 vs 4.33 ± 1.52, P < 0.05). Treatment in mice with IMQ-induced psoriasis showed that SC extract could significantly improve the inflammatory phenotype, PASI score (10.875 ± 0.744 vs 3.875 ± 0.582, P < 0.05), and pathological features compared with those in IMQ model group; SC treatment was also associated with decreased levels of Cyr61 and ICAM-1. In the IFN-γ-induced inflammatory cell model, the mRNA and protein levels of Cyr61 and ICAM-1 were upregulated, while the SC extract downregulated the levels of Cyr61 and ICAM-1. CONCLUSION: The results provide a theoretical basis for the involvement of Cyr61 in the pathogenesis of psoriasis, and suggest that SC should be used to target Cyr61 for the prevention of psoriasis recurrence.

Trehangelins ameliorate inflammation-induced skin senescence by suppressing the epidermal YAP-CCN1 axis.

Trehangelins (THG) are newly identified trehalose compounds derived from broth cultures of an endophytic actinomycete, Polymorphospora rubra. THG are known to suppress Cellular Communication Network factor 1 (CCN1), which regulates collagen homeostasis in the dermis. Although the physical properties of THG suggest a high penetration of the stratum corneum, the effect of THG on the epidermis has not been reported. Here we describe a possible mechanism involved in skin aging focusing on the effect of THG on epidermal CCN1. This study shows that: (1) THG suppress epidermal CCN1 expression by inhibiting the translocation of Yes-Associated Protein (YAP) to nuclei. (2) Epidermal CCN1, localized at the basement membrane, regulates the balance between the growth and differentiation of keratinocytes. (3) Keratinocytes secrete more CCN1 than fibroblasts, which leads to disruption of the basement membrane and extracellular matrix components. (4) The secretion of CCN1 from keratinocytes is increased by ultraviolet B exposure, especially in aged keratinocytes, and deteriorates the elastic fiber structures in the underlying dermis. (5) Topical application of THG ameliorates the structure of the basement membrane in ex vivo human skin explants. Taken together, THG might be a promising treatment for aged skin by suppressing the aberrant YAP-CCN1 axis.

Therapeutic effects of the extract of Sancao Formula, a Chinese herbal compound, on imiquimod-induced psoriasis via cysteine-rich protein 61 / 中西医结合学报

OBJECTIVE@#Psoriasis is a common chronic inflammatory skin disease that is prone to recurrence, and the proinflammatory factor, cysteine-rich protein 61 (Cyr61), is important in its pathophysiology. Long-term clinical practice has shown that Sancao Formula (SC), a Chinese herbal compound, is effective in the treatment of psoriasis, but the precise mechanism remains unknown. In this study, we investigate the mechanism by which SC extract alleviates imiquimod (IMQ)-induced psoriasis.@*METHODS@#The expression of Cyr61 in psoriatic lesions and normal healthy skin was detected using immunohistochemical analysis to investigate the biological role of Cyr61 in models of psoriatic inflammation. A psoriatic mouse model was established by topical application of IMQ, and the effect of topical application of SC extract was evaluated using the psoriasis area and severity index (PASI) score, hematoxylin-eosin staining, and histopathological features of the skin. Next, a HaCaT cell inflammation model was established using interferon-γ (IFN-γ), and the effect of SC extract on the mRNA and protein levels of Cyr61 and intercellular cell adhesion molecule-1 (ICAM-1) was confirmed using Western blot and quantitative real-time polymerase chain reaction analyses.@*RESULTS@#Immunohistochemical staining showed that the expression of Cyr61 in psoriatic lesions was higher than that in normal skin samples (78.26% vs 41.18%, P < 0.05), and the number of Cyr61-positive cells in psoriatic lesions was also significantly higher than in normal skin (18.66 ± 2.51 vs 4.33 ± 1.52, P < 0.05). Treatment in mice with IMQ-induced psoriasis showed that SC extract could significantly improve the inflammatory phenotype, PASI score (10.875 ± 0.744 vs 3.875 ± 0.582, P < 0.05), and pathological features compared with those in IMQ model group; SC treatment was also associated with decreased levels of Cyr61 and ICAM-1. In the IFN-γ-induced inflammatory cell model, the mRNA and protein levels of Cyr61 and ICAM-1 were upregulated, while the SC extract downregulated the levels of Cyr61 and ICAM-1.@*CONCLUSION@#The results provide a theoretical basis for the involvement of Cyr61 in the pathogenesis of psoriasis, and suggest that SC should be used to target Cyr61 for the prevention of psoriasis recurrence.

RASSF1A-Hippo pathway link in patients with urothelial carcinoma of bladder: plausible therapeutic target.

RASSF1A is a tumor suppressor gene, and its hypermethylation has been observed in cancers. RASSF1A acts as an upstream regulator of Hippo pathway and modulates its function. The aim of this study was to analyze expression of RASSF1A, Hippo pathway molecules (YAP, MST) and downstream targets (CTGF, Cyr61 and AREG) in bladder cancer patients. Later, the link between RASSF1A and Hippo pathway and a potential therapeutic scope of this link in UBC were also studied. MSPCR was performed to study methylation of RASSF1A promoter. Expression of molecules was studied using qPCR, Western blot and IHC. The link between RASSF1A and Hippo pathway was studied using Spearman's correlation in patients and validated by overexpressing RASSF1A in HT1376 cells and its effect on Hippo pathway was observed using qPCR and Western blot. Further therapeutic potential of this link was studied using MTT and PI assays. The expression of RASSF1A was lower, whereas the expression of YAP, CTGF and CYR61 was higher. The expression of RASSF1A protein gradually decreased, while the expression of YAP, CTGF and CYR61 increased with severity of disease. Based on Spearman's correlation, RASSF1A showed a negative correlation with YAP, CTGF and CYR61. YAP showed a positive correlation with CTGF and CYR61. To validate this link, RASSF1A was overexpressed in HT1376 cells. Overexpressed RASSF1A activated Hippo pathway, followed by a decrease in CTGF and CYR61 at mRNA, and enhanced cytotoxicity to chemotherapeutic drugs. This study finds a previously unrecognized role of RASSF1A in the regulation of CTGF and CYR61 through mediation of Hippo pathway in UBC and supports the significance of this link as a potential therapeutic target for UBC.

RNA Expression Profile and Alternative Splicing Signatures of Genistein-Treated Breeder Hens Revealed by Hepatic Transcriptomic Analysis.

Little information has been available about the influence of dietary genistein (GEN) on hepatic transcriptome of laying broiler breeder (LBB) hens. The study is aimed at broadening the understanding of RNA expression profiles and alternative splicing (AS) signatures of GEN-treated breeder hens and thereby improving laying performance and immune function of hens during the late egg-laying period. 720 LBB hens were randomly allocated into three groups with supplemental dietary GEN doses (0, 40 mg/kg, and 400 mg/kg). Each treatment has 8 replicates of 30 birds. Dietary GEN enhanced the antioxidative capability of livers, along with the increased activities of glutathione peroxidase and catalase. Furthermore, it improved lipid metabolic status and apoptotic process in the liver of hens. 40 mg/kg dietary GEN had the better effects on improving immune function and laying performance. However, transcriptome data indicated that 400 mg/kg dietary GEN did negative regulation of hormone biosynthetic process. Also, it upregulated the expressions of EDA2R and CYR61 by the Cis regulation of neighbouring genes (lncRNA_XLOC_018890 and XLOC_024242), which might activate NF-κB and immune-related signaling pathway. Furthermore, dietary GEN induced AS events in the liver, which also enriched into immune and metabolic process. Therefore, the application of 40 mg/kg GEN in the diet of breeder hens during the late egg-laying period can improve lipid metabolism and immune function. We need to pay attention to the side-effects of high-dose GEN on the immune function.

Yin/Yang expression of CCN family members: Transforming growth factor beta 1, via ALK5/FAK/MEK, induces CCN1 and CCN2, yet suppresses CCN3, expression in human dermal fibroblasts.

The role of the microenvironment in driving connective tissue disease is being increasingly appreciated. Matricellular proteins of the CCN family are signaling modifiers that are secreted by cells into the extracellular matrix microenvironment where they have profound, context-dependent effects on organ development, homeostasis and disease. Indeed, CCN proteins are emergent targets for therapeutic intervention. Recent evidence suggests that, in vivo, CCN3 has effects opposing CCN2. Moreover, when CCN3 expression is high, CCN2 expression is low. That is, they appear to be regulated in a yin/yang fashion, leading to the hypothesis that the CCN2:CCN3 ratio is important to control tissue homeostasis. To begin to test the hypothesis that alterations in CCN2:CCN3 expression might be important in skin biology in vivo, we evaluated the relative ex vivo effects of the profibrotic protein TGFbeta1 on dermal fibroblasts on protein and RNA expression of CCN3 and CCN2, as well as the related protein CCN1. We also used signal transduction inhibitors to begin to identify the signal transduction pathways controlling the ability of fibroblasts to respond to TGFbeta1. As anticipated, CCN1 and CCN2 protein and mRNA were induced by TGFbeta1 in human dermal fibroblasts. This induction was blocked by TAK1, FAK, YAP1 and MEK inhibition. Conversely, TGFbeta1 suppressed CCN3 mRNA expression in a fashion insensitive to FAK, MEK, TAK1 or YAP1 inhibition. Unexpectedly, CCN3 protein was not detected in human dermal fibroblasts basally. These data suggest that, in dermal fibroblasts, the profibrotic protein TGFbeta1 has a divergent effect on CCN3 relative to CCN2 and CCN1, both at the mRNA and protein level. Given that the major source in skin in vivo of CCN proteins are fibroblasts, our data are consistent that alterations in CCN2/CCN1: CCN3 ratios in response to profibrotic agents such as TGFbeta1 may play a role in connective tissue pathologies including fibrosis.

CCN1 Promotes VEGF Production in Osteoblasts and Induces Endothelial Progenitor Cell Angiogenesis by Inhibiting miR-126 Expression in Rheumatoid Arthritis.

Angiogenesis is the formation of new capillaries from preexisting vasculature. The perpetuation of angiogenesis plays a critical role in the pathogenesis of various disease states including rheumatoid arthritis (RA). Cysteine-rich 61 (Cyr61 or CCN1) is an important proinflammatory cytokine in RA. Here, we investigated the role of CCN1 in angiogenesis associated with vascular endothelial growth factor (VEGF) production and osteoblasts. We found higher expression of CCN1 and VEGF in synovial fluid from RA patients compared with healthy controls. CCN1 induced VEGF expression in osteoblasts and increased endothelial progenitor cells (EPCs) angiogenesis by inhibiting miR-126 via the protein kinase C-alpha (PKC-α) signaling pathway. CCN1 knockdown inhibited angiogenesis in both in vitro and in vivo models. Inhibition of CCN1 expression with lentiviral vectors expressing short hairpin RNA (shRNA) ameliorated articular swelling, cartilage erosion, and angiogenesis in the ankle joint of mice with collagen-induced arthritis (CIA). Our study is the first to describe how CCN1 promotes VEGF expression in osteoblasts and increased EPCs angiogenesis in RA disease. CCN1 may serve as a potential target for RA treatment. © 2016 American Society for Bone and Mineral Research.

DDR2-CYR61-MMP1 Signaling Pathway Promotes Bone Erosion in Rheumatoid Arthritis Through Regulating Migration and Invasion of Fibroblast-Like Synoviocytes.

Regulation of matrix metalloproteinases (MMPs) by collagen in the fibroblast-like synoviocytes (FLSs) plays a critical role in joint destruction in rheumatoid arthritis (RA). Our previous study indicated that discoidin receptor 2 (DDR2) mediated collagen upregulation of MMPs. However, the precise underlying mechanism remains unclear. We report here that CYR61, a secreted, extracellular matrix-associated signaling protein which is capable of regulating a broad range of cellular activities, including cell adhesion, migration, proliferation, and apoptosis, is significantly upregulated in collagen II-stimulated RA FLS. Further studies found that collagen II-activated phosphorylated-DDR2 induces CYR61 through activation of transcription factor activator protein 1 (AP-1). The elevated CYR61, in turn, accelerates MMP1 production via ETS1 (ETS proto-oncogene 1). In addition, CYR61 significantly promotes FLS invasion and migration. Blockade of CYR61 by an adenovirus expressing CYR61 shRNA (Ad-shCYR61) in vivo remarkably ameliorated the severity of arthritis, reduced inflammatory cytokine secretion, and attenuated bone erosion as detected by micro-computed tomography (µCT), in collagen-induced arthritis (CIA) rats. Taken together, we uncovered the Collagen II-DDR2-AP-1-CYR61-ETS1-MMP1 loop in RA FLS. In which, CYR61 acts as a hinge to promote cartilage damage through regulating FLS invasion, migration, and MMP1 production and the inflammatory cascade in RA. Thus, CYR61 may be a promising diagnostic and therapeutic target for RA treatment. © 2016 American Society for Bone and Mineral Research.

CCN1 secretion induced by cigarette smoking extracts augments IL-8 release from bronchial epithelial cells.

Inflammation involves in many cigarette smoke (CS) related diseases including the chronic obstructive pulmonary disease (COPD). Lung epithelial cell released IL-8 plays a crucial role in CS induced lung inflammation. CS and cigarette smoke extracts (CSE) both induce IL-8 secretion and subsequently, IL-8 recruits inflammatory cells into the lung parenchyma. However, the molecular and cellular mechanisms by which CSE triggers IL-8 release remain not completely understood. In this study, we identified a novel extracellular matrix (ECM) molecule, CCN1, which mediated CSE induced IL-8 secretion by lung epithelial cells. We first found that CS and CSE up-regulated CCN1 expression and secretion in lung epithelial cells in vivo and in vitro. CSE up-regulated CCN1 via induction of reactive oxygen spices (ROS) and endoplasmic reticulum (ER) stress. p38 MAPK and JNK activation were also found to mediate the signal pathways in CSE induced CCN1. CCN1 was secreted into ECM via Golgi and membrane channel receptor aquaporin4. After CSE exposure, elevated ECM CCN1 functioned via an autocrine or paracrine manner. Importantly, CCN1 activated Wnt pathway receptor LRP6, subsequently stimulated Wnt pathway component Dvl2 and triggered beta-catenin translocation from cell membrane to cytosol and nucleus. Treatment of Wnt pathway inhibitor suppressed CCN1 induced IL-8 secretion from lung epithelial cells. Taken together, CSE increased CCN1 expression and secretion in lung epithelial cells via induction of ROS and ER stress. Increased ECM CCN1 resulted in augmented IL-8 release through the activation of Wnt pathway.

Epigallocatechin-3-gallate diminishes cytokine-stimulated Cyr61 expression in human osteoblastic cells: a therapeutic potential for arthritis.

OBJECTIVE: To assess the effects of epigallocatechin-3-gallate (EGCG) on cytokine-induced Cyr61 synthesis in human osteoblastic cells and the associated signalling pathways. The therapeutic effect of EGCG on CIA in rats was also studied. METHODS: The expression of Cyr61 and NF-κB pathway molecules was examined by western blotting. CCL2 expression was assessed by northern blotting and ELISA. Interaction between NF-κB and Cyr61 promoter was evaluated by electrophoretic mobility shift assay. In rat CIA, osteoblastic expression of Cyr61 was examined by immunohistochemistry and disease progression was assessed by clinical, radiographic and histological examinations. RESULTS: EGCG inhibited Cyr61 expression stimulated by cytokines in primary human osteoblasts and human osteoblastic cell line U2OS. In U2OS, oncostatin M (OSM) induced IκB-α degradation through the mTOR/rictor/Akt pathway, and EGCG attenuated the action. Electrophoretic mobility shift assay revealed that the OSM-enhanced NF-κB/DNA binding was reduced by EGCG, possibly through abrogating nucleus localization of p65 and p50. Cyr61 enhanced OSM-induced expression of CCL2. Moreover, EGCG diminished OSM-stimulated CCL2 expression at least partially via suppressing Cyr61 induction. Co-distribution of CD68(+) macrophages and Cyr61(+) osteoblasts in osteolytic areas was obvious in the CIA model. Clinical, radiographic and immunohistochemical analyses revealed that administration of EGCG markedly diminished the severity of CIA, macrophage infiltration, and the number of Cyr61-synthesizing osteoblasts. CONCLUSION: By modulating the mTOR/rictor/Akt/NF-κB pathway, EGCG attenuated Cyr61 production in osteoblastic cells and in turn diminished macrophage chemotaxis. Our data support the therapeutic potential of EGCG on arthritis.

Epidermal growth factor upregulates endometrial CYR61 expression via activation of the JAK2/STAT3 pathway.

Endometrial cysteine-rich protein 61 (CYR61, CCN1) is a growth factor-inducible gene whose expression is elevated during the proliferative phase of the menstrual cycle and which has been implicated in the pathogenesis of endometriosis. This study aimed to define the mediators of epidermal growth factor (EGF) signalling on CYR61 expression in spontaneously immortalised human endometrial epithelial cells (HES) as a model system. After 30 min of EGF treatment, the receptor was phosphorylated and internalised as well as mRNA CYR61 increased in HES cells. However, neither inhibition of C-terminal EGF receptor (EGFR)-phosphorylation nor blockage of the mitogen-activated proteinkinase/extracellular signal-regulated kinase (MAPK/ERK) pathway was able to reduce CYR61 levels. Surprisingly, the HES cells showed upregulation of CYR61 mRNA expression after inhibition of the MAPK/ERK pathway when treated with EGF. Specific inhibitor studies identified the contribution of Janus kinase 2 (JAK2) and the signal transducer and activator of transcription protein STAT3 to the regulation of CYR61 expression. The JAK2/STAT3 interaction contributed to the basal expression of CYR61 and mediated EGF-driven regulation of CYR61 after 30 and 120 min of treatment. In summary, EGF-mediated CYR61 upregulation in HES cells involves STAT3 and is counter-regulated by the EGFR/MAPK/ERK pathway.

Qingyihuaji formula inhibits progress of liver metastases from advanced pancreatic cancer xenograft by targeting to decrease expression of Cyr61 and VEGF.

OBJECTIVE: To observe the effects of Qingyihuaji formula (QYHJ) on the progression of liver metastases from human pancreatic cancer and to detect the expression changes of some biological factors associated with angiogenesis and metastasis during the development of advanced pancreatic cancer. METHODS: Nude mice were inoculated intrasplenically with human pancreatic cancer cell line SW1990 and then randomly assigned into 4 groups: a control group and groups QYHJ-A, QYHJ-B, and QYHJ-C. Following this, the mice were treated with or without QYHJ formula for 4 weeks and were sacrificed at the end of the sixth week. The changes in body weight were observed, followed by the livers being excised and weighed. Then, both the numbers and the volume of metastatic nodules per liver were evaluated. Subsequently, the expressions of MMPs, VEGF, and Cyr61 in the tissue of liver metastases were detected by reverse transcription polymerase chain reaction, immunohistochemistry, or Western blot. Finally, the correlation was evaluated between the expressions of the factors associated with metastasis and the growth of liver metastasis. RESULTS: Liver metastases were identified in 11 of 15 mice (73%) in the control group, 9 of 15 mice (60%) in group QYHJ-A, 6 of 14 mice (43%) in group QYHJ-B, and 8 of 14 mice (57%) in group QYHJ-C both the number and the volume of metastatic nodules per liver same as the ratio of liver-body weight in QYHJ groups were significantly less than the controlled group (P < 0.05). The expressions of Cyr61, MMP-2, and VEGF at the levels of mRNA and protein were decreased in the QYHJ groups when compared with the control, as confirmed by immunohistochemistry detection (P < .05). However, no significant difference was observed in the mRNA expression of MMP-1 and MMP-9 between the QYHJ groups and the control group (P > .05). Regression analysis indicated that QYHJ possessed an evident inhibition against the progression of liver metastasis by downregulating the expression of VEGF and Cyr61 rather than MMP-2. CONCLUSIONS: The QYHJ formula exerted an inhibitory effect on the growth of liver metastasis from pancreatic cancer, perhaps by targeting VEGF and Cyr61 to some extent.

Protective effects of Chlorella-derived peptide on UVB-induced production of MMP-1 and degradation of procollagen genes in human skin fibroblasts.

UV exposure is known to induce premature aging, which is mediated by matrix metalloproteinase-1 (MMP-1) activity. MMP-1 mRNA expression is up-regulated by elevated cysteine-rich 61 (CYR61) and monocyte chemoattractant protein-1 (MCP-1) via action of transcription factor AP-1. Collagen is degraded by MMP-1 activity but synthesized by transforming growth factor-ß (TGF-ß) signal. Chlorella has been shown to inhibit UVB-induced MMP-1 level, however its regulatory molecular mechanisms have not been studied. In this study, Chlorella derived peptide (CDP) was added to skin fibroblasts after UVB irradiation and the expression of MMP-1, CYR61, procollagen, c-fos, c-jun, and TGF-ß receptor (TbRII) mRNA and MCP-1 production were investigated. CDP (10 or 5mg/ml) diminished UVB-induced MMP-1 and CYR61 mRNA expression and MCP-1 production, whereas, UVB-suppressed procollagen and TbRII mRNA was restored by CDP treatment. UVB-induced c-fos and c-jun expressions were also inhibited by the CDP treatment. Taken together, CDP inhibits UVB-induced MMP-1 expression in skin fibroblasts by suppressing expression of AP-1 and CYR61 and MCP-1 production.

Arecoline stimulated Cyr61 production in human gingival epithelial cells: inhibition by lovastatin.

Cyr61 is associated with growth and progression of many types of tumors and is an independent poor prognostic indicator for oral cancer patients. Areca nut (AN) chewing is the most important etiological factor in the pathogenesis of oral cancer in India and many Southeast Asian countries. Yet, the molecular mechanisms involved in the AN-induced oral cancer remain largely unknown. In this study, we show that arecoline, a main alkaloid found in AN, stimulated Cyr61 synthesis in human gingival epithelial S-G cells. Constitutive overexpression of Cyr61 protein in oral epithelial cells during AN chewing may play a role in the pathogenesis of oral cancer. ERK inhibitor PD98059, N-acetyl-L-cysteine, Rho-associated protein kinase (ROCK) selective inhibitor Y-27632 and a geranylgeranyltransferase inhibitor reduced the arecoline-stimulated levels of Cyr61 protein by ∼31%, 47%, 65% and 100%, respectively. Lovastatin also completely inhibited arecoline-induced Cyr61 synthesis and the inhibition is dose-dependent. Decreased of geranylgeranylated proteins could be the mechanism that lovastatin regulates Cyr61 synthesis and lovastatin could serve as a useful agent in controlling AN-induced oral cancer.

An extract of green tea, epigallocatechin-3-gallate, reduces periapical lesions by inhibiting cysteine-rich 61 expression in osteoblasts.

Recent investigations indicate that epigallocatechin-3-gallate (EGCG), the major polyphenol of green tea, has anti-inflammatory properties. This study assessed the effect of EGCG on oncostatin M (OSM)-induced synthesis of cysteine-rich 61 (Cyr61), a potential osteolytic mediator, in MG-63 human osteoblastic cells. The therapeutic effect of EGCG in apical periodontitis in rats was also examined. Western blot analysis showed that OSM stimulated Cyr61 synthesis in MG-63 in a time-dependent manner, whereas EGCG readily attenuated this effect. On the other hand, Cyr61 treatment of MG-63 cells induced the release of CCL2, a chemokine responsible for macrophage chemotaxis. In a rat model of induced apical periodontitis, radiography and histopathology revealed that administration of EGCG markedly diminished the severity of periapical lesions. The numbers of Cyr61-synthesizing osteoblasts and infiltrating macrophages were also decreased. Thus, EGCG suppresses the progression of apical periodontitis, possibly by diminishing Cyr61 expression in osteoblasts and, subsequently, macrophage chemotaxis into the lesions.

[Bushen Rougan Recipe in prevention of hepatic fibrosis in rats induced by dimethylnitrosamine: a study on its preliminary mechanism].

OBJECTIVE: To study the effects of Bushen Rougan Recipe (BSRGR), a compound traditional Chinese herbal medicine, on hepatic fibrosis in rats induced by dimethylnitrosamine (DMN), and to explore its preliminary mechanism. METHODS: A total of 40 male Wistar rats were randomly divided into normal control group (n=10), untreated group (n=15), and BSRGR group (n=15). Except for the rats in normal control group, hepatic fibrosis in rats was induced by peritoneal injection of DMN for 4 weeks. And the rats in the BSRGR group were also intragastrically administered BSRGR within the 4-week course. At the end of the 4-week course, rats were all sacrificed. The liver functions were determined by automatic biochemistry analyzer, including serum total bilirubin (Tbil), alanine aminotransferase (ALT), aspartate aminotransferase (AST), and albumin. Expressions of connective tissue growth factor (CTGF) and collagen type I mRNA in liver tissue were detected by reverse transcription polymerase chain reaction. RESULTS: It was found that the serum Tbil level and the activities of AST, ALT were declined in the BSRGR group as compared with those in the untreated group (P<0.01). The serum albumin content in the BSRGR group was increased as compared with that in the untreated group (P<0.01). The expressions of collagen type I and CTGF mRNAs in the untreated group were higher than those in the BSRGR group (P<0.01). CONCLUSION: BSRGR can decrease the expressions of collagen type I and CTGF mRNAs in the rats with hepatic fibrosis, which may be one of possible mechanisms in treating hepatic fibrosis.

Stretch-inducible expression of the angiogenic factor CCN1 in vascular smooth muscle cells is mediated by Egr-1.

CCN1 is an angiogenic factor that promotes cell adhesion, proliferation, and differentiation. CCN1-deficient mice suffer embryonic death because of vascular defects, demonstrating that CCN1 is required for vessel development. Because mechanical stretch may act as a trigger for vessel development, we investigated the impact of mechanical stretch on the regulatory mechanism of CCN1 expression. Mechanical stretch rapidly enhances CCN1 expression and release in vascular smooth muscle cells (VSMC) in vitro and CCN1 expression in murine aortic segments in vivo. Transfection experiments of VSMC with deletion constructs of the CCN1 promoter revealed the regulatory region responsible for the stretch-induced CCN1 expression in the approximately 200-bp promoter region upstream of the TATA-box containing potential binding sites for early growth response-1 (Egr-1), nuclear factor of activated T-cells and cAMP response element binding protein. Decoy oligonucleotides to Egr-1, but not to nuclear factor of activated T-cells or cAMP response element binding protein, abolished the stretch-induced transcription of CCN1. In addition, mutagenesis of the Egr-1 binding site within the CCN1 promoter completely blunted the stretch-induced activation of the promoter. Furthermore, mechanical stretch induced the expression and DNA-binding activity of Egr-1 in VSMC as demonstrated by Western blot and electromobility shift assay. Moreover, a pressure overload-dependent de novo synthesis of Egr-1 was observed after aortic banding. These findings indicate that mechanical stretch leads to enhanced expression of CCN1 via the mechanosensitive transcription factor Egr-1, suggesting a central role for mechanical stretch in the regulation of CCN1-dependent pro-angiogenic potency.