Kazuhiro Iwasaki

Nuclear Import Mechanism for Myocardin Family Members and Their Correlation with Vascular Smooth Muscle Cell Phenotype

Myocardin (Mycd), which is essential for the differentiation of the smooth muscle cell lineage, is constitutively located in the nucleus, although its family members, myocardin-related transcription factors A and B (MRTF-A/B), mostly reside in the cytoplasm and translocate to the nucleus in response to Rho signaling. The mechanism for their nuclear import is unclear. Here we investigated the mechanism for the nuclear import of Mycd family members and demonstrated any correlation between such mechanism and the phenotype of vascular smooth muscle cells (VSMCs). In cultured VSMCs, the knockdown of importin β1 inhibited the nuclear import of Mycd and MRTF-A/B. Their NH2-terminal basic domain was identified as a binding site for importin α/β1 by in vitro analyses. However, Mycd had a higher affinity for importin α/β1 than did MRTF-A/B, even in the absence of G-actin, and Mycd affinity for importin α1/β1 was stronger than for any other importin α/β1 heterodimers. The binding of Mycd to importin α/β1 was insensitive to G-actin, whereas that of MRTF-A/B was differently inhibited by G-actin. In dedifferentiated VSMCs, the levels of importins α1 and β1 were reduced concomitant with down-regulation of Mycd, serum response factor, and smooth muscle cell markers. By contrast, in differentiated VSMCs, their expressions were up-regulated. Thus, the nuclear import of Mycd family members in VSMCs depends on importin α/β1, and their relative affinities for importin α/β1 heterodimers determine Mycd nuclear import. The expression of Mycd nuclear import machineries is related to the expression levels of VSMC phenotype-dependent smooth muscle cell markers.

Rho/Rho-associated kinase signal regulates myogenic differentiation via myocardin-related transcription factor-A/Smad-dependent transcription of the Id3 gene.

RhoA is known to be involved in myogenic differentiation, but whether it acts as a positive or negative regulator is controversial. To resolve this issue, we investigated the differentiation stage-specific roles of RhoA and its effector, Rho-associated kinase, using C2C12 myoblasts. We found that proliferating myoblasts show high levels of RhoA and serum-response factor activities and strong expression of the downstream target of RhoA, myocardin-related transcription factor-A (MRTF-A or MAL); these activities and expression are markedly lower in differentiating myocytes. We further demonstrated that, in proliferating myoblasts, an increase in MRTF-A, which forms a complex with Smad1/4, strikingly activates the expression level of the Id3 gene; the Id3 gene product is a potent inhibitor of myogenic differentiation. Finally, we found that during differentiation, one of the forkhead transcription factors translocates into the nucleus and suppresses Id3 expression by preventing the association of the MRTF-A-Smad complex with the Id3 promoter, which leads to the enhancement of myogenic differentiation. We conclude that RhoA/Rho-associated kinase signaling plays positive and negative roles in myogenic differentiation, mediated by MRTF-A/Smad-dependent transcription of the Id3 gene in a differentiation stage-specific manner.

Partial Bladder Boost Using Lipiodol Marking During Image-guided Radiotherapy for Bladder Cancer

Background/Aim: Secure dose escalation is required to compensate avoidance of concurrent chemotherapy in radiotherapy for increasing elderly bladder cancer. We aimed to evaluate the efficacy of lipiodol submucosally injected as a fiducial marker during image-guided radiotherapy (Lip-IGRT) for muscle invasive bladder cancer (BC). Patients and Methods: Twenty-three patients with T2a-4aN0-1M0 BC underwent whole-bladder irradiation of 46 Gy and Lip-IGRT of 20 Gy, conventionally. The bladder volume exposed to 19 Gy (bV19:%) on Lip-IGRT was referred as an index predicting cystitis. Results: Lipiodol consistently highlighted the boundaries of 20 tumors (88%) on planning and portal verification images. Three of 4 patients under oral anticoagulant agents usage were complicated with grade ≥2 hematuria for 3 days (a patient with a bV19 of >50%) or more than a year (2 patients with bV19 of <50%) after the injection. The 3-year overall survival and disease-free survival rates were 70.4% and 71.1%, respectively. Conclusion: Lipiodol marking is an effective way of demarcating BC. However, it is necessary to address the comorbidities of elderly patients.

Relationship between nocturnal polyuria and non-dipping blood pressure in male patients with lower urinary tract symptoms

The aim of the present study was to examine factors of nocturnal polyuria and blood pressure variability in male patients with lower urinary tract symptoms (LUTS) who were treated.

MP49-13 SELECTIVE NEOADJUVANT CHEMOTHERAPY BY PREDICTION SYSTEMS IMPROVES THE CHEMO-SENSITIVITY OF THE MUSCLE INVASIVE BLADDER CANCERS

INTRODUCTION AND OBJECTIVES: To compare the rate of chemo-sensitivity in neoadjuvant chemotherapy for muscle invasive bladder cancer between historical control and this interventional prospective study by using our M-VAC and CaG prediction systems. METHODS: Muscle invasive bladder cancer patients (Stage II-III) were engaged in this study. They were assigned to M-VAC, CaG, operation or radiation therapy according to the result of the prediction system which we had already established. According to their responses to the neoadjuvant chemotherapy, we categorized the patients into two groups: 0responders0 who achieved significant tumor shrinking (>60%), and 0non-responders0 ( 60%). Patients were evaluated either 0accurate0 or 0 inaccurate0 according to a tumor shrinkage rate cut off line of a 60% decrease after chemotherapies. The primary endpoint of this study was comparison with the historical controls in terms of the rate of chemo-sensitivity and efficacy. The secondary endpoint was down staging ( pT1) and the overall survival. RESULTS: From March 2011 to July 2013, an intention-to-treat 35 cancer samples from patients were entered in this study. Seven patients M-VAC, 22 CaG, 2 operation and 4 radiation therapy were received respectively. The predicted responder rate for M-VAC was 42.9% (15/35) and for CaG was 57.1% (20/35). Moreover, 74.3% (26/35) of all patients could be expected to respond to both or either of these two regimens by applying our two prediction systems. While, in previous historical control studies (M-VAC 39, CaG 37, total 76 cases), the total predicted response rate of the 76 cases was 57.9% (44/76) (p1⁄40.09). And in the predicted 0responder0 cases, the response rate of M-VAC neoadjuvant chemotherapy was 85.7% (6/7) and CaG was 89.5% (17/19). Therefore, the combined response rate in this group was 88.5% (23/26). In the historical control cases, the observed response rate of M-VAC/CaG therapy was 59.0% (23/39)/54.1% (20/37), and combined observed response rate was 56.6% (43/76) (p1⁄40.004). Down staging and overall survival between this prospective study and the historical control were not significant statistical differences, however in overall survival this prospective study was better tendency than that of the historical control (p1⁄40.233). CONCLUSIONS: Combining the M-VAC and CaG prediction systems would improve the chemo-sensitivity for muscle invasive bladder cancers.