scholarly journals Real-time monitoring of specific oxygen uptake rates of embryonic stem cells in a microfluidic cell culture device

2016 ◽  
Vol 11 (9) ◽  
pp. 1179-1189 ◽  
Author(s):  
Alexandre Super ◽  
Nicolas Jaccard ◽  
Marco Paulo Cardoso Marques ◽  
Rhys Jarred Macown ◽  
Lewis Donald Griffin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Embryonic Stem Cells ◽  
Oxygen Uptake ◽  
Real Time ◽  
Embryonic Stem ◽  
Real Time Monitoring ◽  
Uptake Rates ◽  
Microfluidic Cell Culture

scholarly journals Human Embryonic Stem Cell Lines Model Experimental Human Cytomegalovirus Latency

mBio ◽  
2013 ◽  
Vol 4 (3) ◽  
Author(s):  
Rhiannon R. Penkert ◽  
Robert F. Kalejta
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Embryonic Stem Cells ◽  
Human Cytomegalovirus ◽  
Embryonic Stem ◽  
Immune Defense ◽  
Model Systems ◽  
Productive Infection ◽  
Latent Infections

ABSTRACTHerpesviruses are highly successful pathogens that persist for the lifetime of their hosts primarily because of their ability to establish and maintain latent infections from which the virus is capable of productively reactivating. Human cytomegalovirus (HCMV), a betaherpesvirus, establishes latency in CD34+hematopoietic progenitor cells during natural infections in the body. Experimental infection of CD34+cellsex vivohas demonstrated that expression of the viral gene products that drive productive infection is silenced by an intrinsic immune defense mediated by Daxx and histone deacetylases through heterochromatinization of the viral genome during the establishment of latency. Additional mechanistic details about the establishment, let alone maintenance and reactivation, of HCMV latency remain scarce. This is partly due to the technical challenges of CD34+cell culture, most notably, the difficulty in preventing spontaneous differentiation that drives reactivation and renders them permissive for productive infection. Here we demonstrate that HCMV can establish, maintain, and reactivatein vitrofrom experimental latency in cultures of human embryonic stem cells (ESCs), for which spurious differentiation can be prevented or controlled. Furthermore, we show that known molecular aspects of HCMV latency are faithfully recapitulated in these cells. In total, we present ESCs as a novel, tractable model for studies of HCMV latency.IMPORTANCEHuman cytomegalovirus (HCMV) is a significant human pathogen that is known for causing birth defects, blindness in AIDS patients, and organ transplant rejection. The ability of HCMV to cause disease is dependent upon its capacity to establish and maintain latent infections. Very few of the molecular mechanisms of latency have been elucidated, due in part to the lack of a tractable cell culture model. Here we present embryonic stem cells (ESCs) as a model for HCMV latency, one in which genome maintenance and reactivation could be closely monitored. HCMV establishes latency in ESCs in the same fashion as it does in CD34+cells, the currently favoredin vitromodel. Hence, ESCs represent a novel model with unique properties, such as the ability to be genetically manipulated and cultured indefinitely in an undifferentiated state, that will facilitate the mechanistic examination of certain aspects of HCMV latency that have proven technically challenging in other model systems.

scholarly journals Overexpression of Cardiac-Specific Kinase TNNI3K Promotes Mouse Embryonic Stem Cells Differentiation into Cardiomyocytes

2017 ◽  
Vol 41 (1) ◽  
pp. 381-398 ◽  
Author(s):  
Yin Wang ◽  
Shi-qiang Wang ◽  
Li-peng Wang ◽  
Yu-hong Yao ◽  
Chun-yan Ma ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Real Time ◽  
Cardiac Troponin ◽  
Troponin I ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Cardiac Troponin I ◽  
Control Group ◽  
Knock Down

Backgroud/Aims: The biological function of cardiac troponin I-interacting kinase (TNNI3K), a cardiac-specific functional kinase, is largely unknown. We investigated the effect of human TNNI3K (hTNNI3K) on the differentiation of mouse embryonic stem cells (mESCs) into cardiomyocytes. Methods: First, the time-space expression of endogenous Tnni3k was detected by real-time polymerase chain reaction (PCR) and western blotting at 16 different time-points over a period of 28 days. Further, action potentials and calcium current with/without 5 µM nifedipine were measured by patch clamp for mESC-derived cardiomyocytes. HTNNI3K and mouse-derived siRNA were transfected into mESC using lentivirus vector to induce hTNNI3K overexpression and knock-down, respectively. Results: The number of troponin-T (cTnT) positive cells was greater in the group with TNNI3K overexpression as compared to that in control group, while less such cells were detected in the mTnni3k knock-down group as evaluated on flow cytometry (FCM) and ImageXpress Micro system. After upregulation of connexin43, cardiac troponin-I (Ctni), Ctni, Gata4 were detected in mESCs with TNNI3K overexpression; however, overexpression of α-Actinin and Mlc2v was not detected. Interestingly, Ctnt, connexin40 and connexin45, the markers of ventricular, atrial, and pacemaker cells, respectively, were detected in by real-time PCR in TNNI3K overexpression group. Conclusion: our study indicated that TNNI3K overexpression promoted mESC differentiating into beating cardiomyocytes and induced up-regulating expression of cTnT by PKCε signal pathway, which suggested a modulation of TNNI3K activity as a potential therapeutic approach for ischemic cardiac disease.

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