Takeshi Hatani

MicroRNA-responsive ON and OFF switches provide a safe and time-saving method for purifying a wide range of cell types.

One of the earliest maturation steps in cardiomyocytes (CMs) is the sarcomere protein isoform switch between TNNI1 and TNNI3 (fetal and neonatal/adult troponin I). Here, we generate human induced pluripotent stem cells (hiPSCs) carrying a TNNI1^EmGFP and TNNI3^mCherry double reporter to monitor and isolate mature sub-populations during cardiac differentiation. Extensive drug screening identifies two compounds, an estrogen-related receptor gamma (ERRγ) agonist and an S-phase kinase-associated protein 2 inhibitor, that enhances cardiac maturation and a significant change to TNNI3 expression. Expression, morphological, functional, and molecular analyses indicate that hiPSC-CMs treated with the ERRγ agonist show a larger cell size, longer sarcomere length, the presence of transverse tubules, and enhanced metabolic function and contractile and electrical properties. Here, we show that ERRγ-treated hiPSC-CMs have a mature cellular property consistent with neonatal CMs and are useful for disease modeling and regenerative medicine.

Specific cell lineages differentiated from human induced pluripotent stem cells (iPSCs) are promising sources for cell replacement therapy and are useful biomedical research tools for research on disease mechanisms and drug discovery. Among the different lineages, cardiac lineage has been one of the most efficiently differentiated through several established protocols. In this chapter, we describe our reproducible and highly efficient methodology for differentiating iPSCs into cardiomyocytes using embryoid bodies. We also describe methods to dissociate iPSC-derived cardiomyocytes and to evaluate iPSC-derived cardiomyocytes.

Human pluripotent stem cell-derived cardiomyocytes (CMs) are a promising tool for cardiac cell therapy. Although transplantation of induced pluripotent stem cell (iPSC)-derived CMs have been reported in several animal models, the treatment effect was limited, probably due to poor optimization of the injected cells. To optimize graft cells for cardiac reconstruction, we compared the engraftment efficiency of intramyocardially-injected undifferentiated-iPSCs, day4 mesodermal cells and day8, day20 and day30 purified iPSC-CMs after initial differentiation by tracing the engraftment ratio (ER) using in vivo bioluminescence imaging. This analysis revealed the ER of day20 CMs was significantly higher compared to other cells. Transplantation of day20 CMs into the infarcted hearts of immunodeficient mice showed good engraftment and echocardiography showed significant functional improvement by cell therapy. Moreover, the imaging signal and ratio of Ki67-positive CMs at 3 months post injection indicated engrafted CMs proliferated in the host heart. Although this graft growth reached a plateau at 3 months, histological analysis confirmed progressive maturation from 3 to 6 months. These results suggested that day20 CMs had very high engraftment, proliferation and therapeutic potential in host mouse hearts. They also demonstrate this model can be used to track the fate of transplanted cells over a long time.

Background and objective:  Septic pulmonary embolism due to periodontal disease (SPE‐PD) is rarely reported and little is known about its clinical features. The purpose of this study was to evaluate the clinical and radiological features, as well as outcome, in SPE‐PD.Methods:  Patients' records were retrospectively reviewed and 12 patients with SPE‐PD were identified (10 men, mean age 60.5 years). The patients' demographic features, laboratory data, physical and radiological findings, and clinical outcomes were evaluated.Results:  All but one patient were smokers. Eight of the 12 patients had comorbidities including hypertension (58%) and/or diabetes mellitus (17%). Prevalent symptoms were fever (67%) and chest pain (58%). Only two patients fulfilled the criteria of systemic inflammatory response syndrome; most of the subjects were not clinically severely ill. Blood cultures were negative in all cases. Contrast‐enhanced chest computed tomography (CT) showed multiple peripheral nodules in all 12 patients, wedge‐shaped peripheral lesions abutting on the pleura in 10 (83%) and a feeding‐vessel sign in 9 (75%). All patients recovered from their illness after antimicrobial therapy concomitant with tooth extraction or periodontal care. The median duration of antibiotic administration was 51 days.Conclusions:  Most patients with SPE‐PD were not seriously ill. Contrast‐enhanced chest CT appeared to be useful to diagnose SPE‐PD.