Bladder cancer is the fourth most common type of cancer in men (ninth in women) in the United States. Cisplatin is an effective agent against the most common subtype, urothelial carcinoma. However, the development of chemotherapy resistance is a severe clinical problem for the successful treatment of this and other cancers. A better understanding of the cellular and molecular events in response to cisplatin treatment and the development of resistance are critical to improve the therapeutic options for patients. Here, we report that expression of the CCAAT/enhancer binding protein delta (CEBPD, C/EBPδ, NF-IL6β) is induced by cisplatin in the human bladder urothelial carcinoma NTUB1 cell line and is specifically elevated in a cisplatin resistant subline. Expression of CEBPD reduced cisplatin-induced reactive oxygen species (ROS) and apoptosis in NTUB1 cells by inducing the expression of Cu/Zn-superoxide dismutase (SOD1) via direct promoter transactivation. Several reports have implicated CEBPD as a tumor suppressor gene. This study reveals a novel role for CEBPD in conferring drug resistance, suggesting that it can also be pro-oncogenic. Furthermore, our data suggest that SOD inhibitors, which are already used as anti-angiogenic agents, may be suitable for combinatorial chemotherapy to prevent or treat cisplatin resistance in bladder and possibly other cancers.

Heme oxygenase (HO)-1 is an inducible stress response protein and well known to protect cells and tissues against injury. Despite its important function in cytoprotection against physiological stress, the role of HO-1 in embryonic stem cell (ESC) differentiation remains largely unknown. We showed previously that induced pluripotent stem (iPS) cells that lack HO-1 are more sensitive to oxidant stress-induced cell death and more prone to lose pluripotent markers upon LIF withdrawal. To elucidate the role of HO-1 in ESC differentiation and to rule out the controversy of potential gene flaws in iPS cells, we derived and established mouse HO-1 knockout ESC lines from HO-1 knockout blastocysts. Using wild type D3 and HO-1 knockout ESCs in the 3-dimensional embryoid body (EB) differentiation model, we showed that at an early time point during EB development, an absence of HO-1 led to enhanced ROS level, concomitant with increased expressions of master mesodermal regulator brachyury and endodermal marker GATA6. In addition, critical smooth muscle cell (SMC) transcription factor serum response factor and its coactivator myocardin were enhanced. Furthermore, HO-1 deficiency increased Smad2 in ESCs and EBs, revealing a role of HO-1 in controlling Smad2 level. Smad2 not only mediates mesendoderm differentiation of mouse ESCs but also SMC development. Collectively, loss of HO-1 resulted in higher level of mesodermal and SMC regulators, leading to accelerated and enhanced SMC marker SM α-actin expression. Our results reveal a previously unrecognized function of HO-1 in regulating SMC gene expressions during ESC-EB development. More importantly, our findings may provide a novel strategy in enhancing ESC differentiation toward SMC lineage.

Bladder cancer is the fourth most common type of cancer in men (ninth in women) in the United States. Cisplatin is an effective agent against the most common subtype, urothelial carcinoma. However, the development of chemotherapy resistance is a severe clinical problem for the successful treatment of cancers. A better understanding of the cellular and molecular events in response to cisplatin treatment and the development of resistance are critical to improve the therapeutic options for patients. Here, we report that expression of the CCAAT/enhancer binding protein delta (CEBPD, C/EBPδ, NFIL-6β) is induced by cisplatin in the human bladder urothelial carcinoma NTUB1 cell line and is specifically elevated in a cisplatin resistant subline. Expression of CEBPD reduces cisplatin-induced reactive oxygen species (ROS) and apoptosis in NTUB1 cells by inducing the expression of Cu/Zn-superoxide dismutase (SOD1) via direct promoter trans-activation. Several reports have implicated CEBPD as a tumor suppressor gene. This study reveals a novel role for CEBPD in conferring drug resistance, suggesting that it can also be pro-oncogenic. Furthermore, our data suggest that SOD inhibitors, which are already used as antiangiogenic agents, may be suitable for combinatorial chemotherapy to prevent or treat cisplatin resistance in bladder and possibly other cancers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2558.

Heme oxygenase (HO)‐1 is well known as a stress response protein, playing an important role for protecting cells and tissues from injury with its antioxidative and anti‐inflammatory properties. Although it possess crucial cytoprotective ability against physiological stress, the functions of HO‐1 in embryonic stem cell (ESC) differentiation remain to be elucidated. In previous study, we indicated that in the absence of HO‐1, induced pluripotent stem (iPS) cells are more sensitive to oxidant stress‐induced cell death and more prone to lose pluripotent markers without LIF. To investigate the role of HO‐1 in ESC differentiation and to eliminate the controversy of potential gene defects in iPS cells, we attempted to set up mouse HO‐1 knockout ESC lines from HO‐1 knockout blastocysts. Under the 3‐dimensional embryoid body (EB) formation from wild type D3 and HO‐1 knockout ESCs, we showed that at an early time point after EB differentiation, a lack of HO‐1 led to enhanced ROS level, accompanied with upregulations of master mesodermal regulator brachyury and endodermal marker GATA6. Furthermore, critical smooth muscle cell (SMC) transcription factor serum response factor and its coactivator myocardin were increased. Moreover, HO‐1 deficiency augmented Smad2 expression in ESCs and EBs, indicating that Smad2 not only mediates mesendoderm differentiation of mouse ESCs but also SMC development. Taken together, absence of HO‐1 contributed to higher level of mesodermal and SMC regulators, leading to accelerated and enhanced SMC marker expressions. Our results demonstrated a previously indeterminate function of HO‐1 in regulating SMC gene expressions during ESC‐EB development. More significantly, this finding may provide a novel approach to enhance SMC differentiation from ESCs.Support or Funding InformationNational Health Research Institutes and Central Government S&T grant, Taiwan (106‐0324‐01‐10‐07), Ministry of Science and Technology of Taiwan (MOST 104‐2320‐B‐400‐006‐MY3 and MOST 106‐3114‐Y‐043‐021), and National Health Research Institutes of Taiwan (CS‐106‐PP‐05).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.