Purpose: TGF-b promotes tumor invasion and metastasis by inducing an epithelial-mesenchymal transition (EMT). However, the underlying mechanisms causing this are not entirely clear. Long noncoding RNAs (lncRNA) have been shown to play important regulatory roles in cancer progression. The lncRNA malat1 (metastasis associated lung adenocarcinoma transcript 1) is a critical regulator of the metastasis phenotype of lung cancer cells.Experimental Design: We, therefore, investigated whether TGF-b regulates malat1 expression to promote tumor metastasis of bladder cancer. The expression levels of malat1 and EMT markers were assayed in specimens of bladder cancer. The role of malat1 in regulating bladder cancer metastasis was evaluated in cell and animal models.Results: TGF-b induces malat1 expression and EMT in bladder cancer cells. malat1 overexpression is significantly correlated with poor survival in patients with bladder cancer. malat1 and E-cadherin expression is negatively correlated in vitro and in vivo. malat1 knockdown inhibits TGF-b-induced EMT. malat1 is associated with suppressor of zeste 12 (suz12), and this association results in decrease of E-cadherin expression and increase of N-cadherin and fibronectin expression. Furthermore, targeted inhibition of malat1 or suz12 suppresses the migratory and invasive properties induced by TGF-b. Finally, we demonstrated that malat1 or suz12 knockdown inhibits tumor metastasis in animal models.Conclusion: These data suggest that malat1 is an important mediator of TGF-b-induced EMT, and suggest that malat1 inhibition may represent a promising therapeutic option for suppressing bladder cancer progression. Clin Cancer Res; 20(6); 1531-41. Ó2014 AACR.

Chemotherapy is a reasonable alternative to cystectomy in patients with invasive and advanced bladder cancer. However, bladder cancer cells often develop drug resistance to these therapies, and~50% of patients with advanced bladder cancer do not respond to chemotherapy. Recent studies have shown that long non-coding RNA (lncRNA) is involved in the development of chemoresistance. Here we investigated the role of the urothelial cancer-associated 1 (UCA1) lncRNA in cisplatin resistance during chemotherapy for bladder cancer. We showed that cisplatin-based chemotherapy results in up-regulation of UCA1 expression in patients with bladder cancer. Similarly, UCA1 levels are increased in cisplatin-resistant bladder cancer cells. Over-expression of UCA1 significantly increases the cell viability during cisplatin treatment, whereas UCA1 knockdown reduces the cell viability during cisplatin treatment. UCA1 inhibition also partially overcomes drug resistance in cisplatin-resistant T24 cells. Furthermore, we showed that UCA1 positively regulates expression of wingless-type MMTV integration site family member 6 (Wnt6) in human bladder cancer cell lines. UCA1 and Wnt6 expression is also positively correlated in vivo. Up-regulation of UCA1 activates Wnt signaling in a Wnt6-dependent manner. We finally demonstrate that UCA1 increases the cisplatin resistance of bladder cancer cells by enhancing the expression of Wnt6, and thus represents a potential target to overcome chemoresistance in bladder cancer.

Mesenchymal stem cells (MSCs) derived exosomes have been shown to have protective effects on the kidney in ischemia/reperfusion-induced renal injury. However, the key components in the exosomes and their potential mechanisms for the kidney protective effects are not well understood. In our current study, we focused on the abundant proteins in exosomes derived from MSCs (MSC-exo) and found that the C-C motif chemokine receptor-2 (CCR2) was expressed on MSC-exo with a high ability to bind to its ligand CCL2. We also proved that CCR2 high-expressed MSC-exo could reduce the concentration of free CCL2 and suppress its functions to recruit or activate macrophage. Further, knockdown of CCR2 expression on the MSC-exo greatly abolished these effects. Finally, we also found that CCR2 knockdown impaired the protective effects of MSC-exo for the renal ischemia/reperfusion injury in mouse. The results indicate that CCR2 expressed on MSC-exo may play a key role in inflammation regulation and renal injury repair by acting as a decoy to suppress CCL2 activity. Our study may cast new light on understanding the functions of the MSC-exo and these receptor proteins expressed on exosomes.

Recent studies have suggested a role for aldosterone in the pathogenesis of renal injury. This study investigated the potential contributions of Rho-kinase and TGF-␤ pathways to aldosterone-induced renal injury. Rats were uninephrectomized and then treated for 5 wk with 1% NaCl in a drinking solution and one of the following: Vehicle (2% ethanol, subcutaneously; n ‫؍‬ 9); aldosterone (0.75 g/h, subcutaneously; n ‫؍‬ 9); or aldosterone ؉ fasudil, a specific Rho-kinase inhibitor (10 mg/kg per d, subcutaneously; n ‫؍‬ 8). Phosphorylation of myosin phosphate target subunit-1 (MYPT1) and Smad2/3 in renal cortical tissue was measured by Western blotting with anti-phospho MYPT1 and Smad2/3 antibodies, respectively. Rats that received aldosterone infusion exhibited hypertension and severe renal injury characterized by proteinuria, glomerular sclerosis, and tubulointerstitial fibrosis with increases in ␣-smooth muscle actin staining and numbers of monocytes/macrophages in the interstitium. Renal cortical mRNA levels of types I and III collagen, TGF-␤, connective tissue growth factor, and monocyte chemoattractant protein-1 as well as Smad2/3 phosphorylation were significantly increased in rats that received aldosterone infusion. All of these changes were associated with an increase in renal tissue MYPT1 phosphorylation. Treatment with fasudil did not alter BP but significantly ameliorated proteinuria and renal injury in rats that received aldosterone infusion. Severe glomerular injury and interstitial fibrosis were observed in rats that were treated long term with aldosterone/ salt (3,4). Moreover, exogenous infusion of aldosterone reversed the renoprotective effects of angiotensin II blockade in remnant kidney hypertensive rats (5). In stroke-prone spontaneously hypertensive rats (6); Dahl salt-sensitive hypertensive rats (7); and rats that were treated with angiotensin II and a nitric oxide synthase inhibitor (8), cyclosporine A (9), or radiation (10), treatment with mineralocorticoid receptor (MR) antagonists had no effect on systemic BP (SBP) but significantly ameliorated renal injury. In patients with chronic renal failure (11) and early diabetic nephropathy (12), the addition of MR antagonists to angiotensin-converting enzyme (ACE) inhibitors had no hemodynamic effects but markedly reduced proteinuria. Further studies also showed that monotherapy with MR antagonists was more effective than ACE inhibitors in reducing proteinuria in hypertensive patients (13,14). We recently found that aldosterone stimulates reactive oxygen species production (15) and induces cell proliferation (16) or deformation (16) in cultured glomerular mesangial cells. In addition, aldosterone stimulated collagen synthesis in cultured renal fibroblasts (17). Collectively, these data suggest that aldosterone has deleterious effects on the kidney that cannot be explained simply by BP changes. However, the precise mechanisms that are responsible for aldosterone-induced renal injury remain undetermined.Rho-kinase, an effector of the small G protein ...

Renal fibrosis is the fundamental pathway leading to end-stage renal disease, while its exact molecular basis remains incompletely elucidated. Previous studies have demonstrated that transforming growth factor beta 1 (TGFβ1) is an inducer of the epithelial-to-mesenchymal transition (EMT) of renal tubular epithelial cells, while bone morphogenic protein 7 (BMP7) counteracts TGFβ1-induced EMT and reverses chronic renal injury. Although macrophage recruitment is believed to play an important role during the whole pathogenesis, the mechanism underlying their activate involvement in the formation of renal fibrosis besides phagocytosizing extracellular matrix and apoptotic cells is largely unknown. Here, in a mouse unilateral ureteral obstruction (UUO) model, we show that the recruited macrophages are mainly M1 macrophages at early stage. However, these F4/80-positive and CD301-negative M1 macrophages were shortly polarized into F4/80-positive and CD301-positive M2 macrophages, respectively, which released high levels TGFβ1, to contradict the local expression of BMP7 to facilitate EMT-induced renal fibrosis. M2 macrophages depletion specifically inhibited EMT, and subsequently the renal fibrosis. Adoptive transplantation of M2 macrophages increased the features of renal fibrosis. Our study thus identified double-edged effects of macrophages in the formation of renal fibrosis, which suggest that modulation of macrophage polarization may substantially improve the treatment of renal fibrosis.