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.

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.

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.