Noninvasive in vivo tracking of mesenchymal stem cells and evaluation of cell therapeutic effects in a murine model using a clinical 3.0 T MRI

Cardiac cell therapy with mesenchymal stem cells (MSCs) represents a promising treatment approach for endstage heart failure. However, little is known about the underlying mechanisms and the fate of the transplanted cells. The objective of the presented work is to determine the feasibility of magnetic resonance imaging (MRI) and in vivo monitoring after transplantation into infarcted mouse hearts using a clinical 3.0 T MRI device. The labeling procedure of bone marrow-derived MSCs with micron-sized paramagnetic iron oxide particles (MPIOs) did not affect the viability of the cells and their cell type-defining properties when compared to unlabeled cells. Using a clinical 3.0 T MRI scanner equipped with a dedicated small animal solenoid coil, 105 labeled MSCs could be detected and localized in the mouse hearts for up to 4 weeks after intramyocardial transplantation. Weekly ECG-gated scans using T1-weighted sequences were performed, and left ventricular function was assessed. Histological analysis of hearts confirmed the survival of labeled MSCs in the target area up to 4 weeks after transplantation. In conclusion, in vivo tracking of labeled MSCs using a clinical 3.0 T MRI scanner is feasible. In combination with assessment of heart function, this technology allows the monitoring of the therapeutic efficacy of regenerative therapies in a small animal model. </jats:p

Maximal oxygen uptake and fatty acid oxidation in athletic older men and women and healthy control

Introduction: Cardiopulmonary and musculoskeletal systems deteriorate through middle and into older age. This has a negative impact on physical capability and energy metabolism. The purpose of the present study was to determine the effects of ageing and exercise on peak rates of oxygen uptake (VO2peak) and fatty acid oxidation (PFO).Methods: All participants provided written, informed consent. Masters Athletes (MA: n=40, aged 37-90) specialised in endurance (n=10) or sprint running (n=30) were recruited during the 2012 European MA Championships in Zittau, Germany. Untrained (n=42, aged 18-67; 23 men and 16 women) were recruited from the general Manchester population (UK). The untrained participants also completed 12 weeks very high intensity sprint cycle training (4* 20s at 170% VO2max, 3/wk). VO2max and PFO were assessed using indirect calorimetry and incremental cycle ergometry. Statistical significance was gained by independent samples t-tests using IBM SPSS v.20.Results: The endurance and sprint trained MA were a similar age and had similar VO2max (Endurance MA: 47.22 ml/kg/min (±4.15) vs Sprint MA: 43.52 ml/kg/min (±2.21) p=0.416). Both MA groups were significantly higher than untrained people (38.86 ml/kg/min). MA sprinters and endurance runners had a VO2max similar to 19 years younger untrained, healthy people. Regression analysis showed that VO2max decreased by around 11% per decade after the age of 40 yrs in the MA group and 5% per decade after the age of 40 yrs in the untrained group. PFO was similar in endurance and sprint trained MA (Endurance: 8.09 mg/kg/min (±0.95) vs Sprint: 6.91 mg/kg/min (±0.53) p=0.284). In the untrained group, PFO was significantly lower than MA (p=0.006). Regression showed that PFO of MAs was similar to that of an untrained, healthy person 19 years younger. The sprint-training programme caused VO2max to increase by 10% (Pre: 38.86 ml/kg/min (±1.31) vs Post: 42.84 ml/kg/min (±1.24) p&lt;0.001) and PFO to increase by 18% (Pre: 5.57 mg/kg/min (±0.33) vs Post: 6.58 mg/kg/min (±0.41) p=0.050).Conclusion: These results show that MAs have a cardiopulmonary and metabolic fitness at levels equivalent to someone almost 20 yrs younger. Previously untrained middle-aged people can achieve substantial gains in fitness by completing relatively short duration, but high intensity sprint training and reach levels similar to those observed in the master athletes

Dynamic Support Culture of Murine Skeletal Muscle-derived Stem Cells Improves Their Cardiogenic Potential In Vitro

Ischemic heart disease is the main cause of death in western countries and its burden is increasing worldwide. It typically involves irreversible degeneration and loss of myocardial tissue leading to poor prognosis and fatal outcome. Autologous cells with the potential to regenerate damaged heart tissue would be an ideal source for cell therapeutic approaches. Here, we compared different methods of conditional culture for increasing the yield and cardiogenic potential of murine skeletal muscle-derived stem cells. A subpopulation of nonadherent cells was isolated from skeletal muscle by preplating and applying cell culture conditions differing in support of cluster formation. In contrast to static culture conditions, dynamic culture with or without previous hanging drop preculture led to significantly increased cluster diameters and the expression of cardiac specific markers on the protein and mRNA level. Whole-cell patch-clamp studies revealed similarities to pacemaker action potentials and responsiveness to cardiac specific pharmacological stimuli. This data indicates that skeletal muscle-derived stem cells are capable of adopting enhanced cardiac muscle cell-like properties by applying specific culture conditions. Choosing this route for the establishment of a sustainable, autologous source of cells for cardiac therapies holds the potential of being clinically more acceptable than transgenic manipulation of cells

Plastizitätstheoretische Untersuchungen von Umformvorgängen mit der Methode der Finiten Elemente

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The Fried Frailty Criteria as Inclusion Criteria for a Randomized Controlled Trial: Personal Experience and Literature Review

Background: Among current operational definitions of frailty, the criteria proposed by Fried and colleagues have attracted great scientific interest. However, these criteria have usually been applied in epidemiological and only rarely in interventional studies. Objective: The present paper aims at testing the applicability of the Fried frailty criteria in the context of the recruitment process of a randomized controlled trial in prefrail older persons, and it discusses the respective scientific literature. Methods: Recruitment was promoted by newspaper articles as well as by targeted mail to customers of a local health insurance company and to recently treated patients of a geriatric day clinic. Furthermore, presentations were given in assisted living facilities. Potential candidates were screened for prefrailty, i.e. to see whether they met 1 or 2 of the Fried criteria (weight loss, handgrip strength, gait speed, exhaustion, physical activity). Results: A total of 298 people were screened. Among them 181 were not frail, 116 were prefrail and 1 was diagnosed as frail. The most prevalent criterion was exhaustion (24% of those screened). The second most prevalent criterion was low handgrip strength (20%). Low gait speed (8%), low physical activity (2%) and weight loss (2%) had a lower prevalence. According to the Geriatric Depression Scale, 14% of those who met the criterion ‘exhaustion’ were depressed. With regard to the Minnesota Leisure Time Physical Activity Questionnaire, used for the evaluation of ‘physical activity’, only 3 activities among the 18 selected by Fried were applicable to our cohort. Conclusions: Under the study conditions, good applicability of the Fried criteria was observed. Nevertheless, further refinement may be expedient in several criteria, especially exhaustion and physical activity, to enhance clinical usefulness. It may be helpful to adapt the cutoffs when applying the criteria to a European population

Co-transplantation of Mesenchymal Stromal Cells and Induced Pluripotent Stem Cell-Derived Cardiomyocytes Improves Cardiac Function After Myocardial Damage

Induced pluripotent stem cell-derived cardiomyocytes (iPS-CMs) represent an attractive resource for cardiac regeneration. However, survival and functional integration of transplanted iPS-CM is poor and remains a major challenge for the development of effective therapies. We hypothesized that paracrine effects of co-transplanted mesenchymal stromal cells (MSCs) augment the retention and therapeutic efficacy of iPS-CM in a mouse model of myocardial infarction (MI). To test this, either iPS-CM, MSC, or both cell types were transplanted into the cryoinfarction border zone of syngeneic mice immediately after injury. Bioluminescence imaging (BLI) of iPS-CM did not confirm enhanced retention by co-application of MSC during the 28-day follow-up period. However, histological analyses of hearts 28 days after cell transplantation showed that MSC increased the fraction of animals with detectable iPS-CM by 2-fold. Cardiac MRI analyses showed that from day 14 after transplantation on, the animals that have received cells had a significantly higher left ventricular ejection fraction (LVEF) compared to the placebo group. There was no statistically significant difference in LVEF between animals transplanted only with iPS-CM or only with MSC. However, combined iPS-CM and MSC transplantation resulted in higher LVEF compared to transplantation of single-cell populations during the whole observation period. Histological analyses revealed that MSC increased the capillarization in the myocardium when transplanted alone or with iPS-CM and decreased the infarct scar area only when transplanted in combination with iPS-CM. These results indicate that co-transplantation of iPS-CM and MSC improves cardiac regeneration after cardiac damage, demonstrating the potential of combining multiple cell types for increasing the efficacy of future cardiac cell therapies

Residual effects of muscle strength and muscle power training and detraining on physical function in community-dwelling prefrail older adults: a randomized controlled trial

Abstract Background Although resistance exercise interventions have been shown to be beneficial in prefrail or frail older adults it remains unclear whether there are residual effects when the training is followed by a period of detraining. The aim of this study was to establish the sustainability of a muscle power or muscle strength training effect in prefrail older adults following training and detraining. Methods 69 prefrail community-dwelling older adults, aged 65–94 years were randomly assigned into three groups: muscle strength training (ST), muscle power training (PT) or controls. The exercise interventions were performed for 60 minutes, twice a week over 12 weeks. Physical function (Short Physical Performance Battery=SPPB), muscle power (sit-to-stand transfer=STS), self-reported function (SF-LLFDI) and appendicular lean mass (aLM) were measured at baseline and at 12, 24 and 36 weeks after the start of the intervention. Results For the SPPB, significant intervention effects were found at 12 weeks in both exercise groups (ST: p = 0.0047; PT: p = 0.0043). There were no statistically significant effects at 24 and 36 weeks. In the ST group, the SPPB declined continuously after stop of exercising whereas the PT group and controls remained unchanged. No effects were found for muscle power, SF-LLFDI and aLM. Conclusions The results showed that both intervention types are equally effective at 12 weeks but did not result in statistically significant residual effects when the training is followed by a period of detraining. The unchanged SPPB score at 24 and 36 weeks in the PT group indicates that muscle power training might be more beneficial than muscle strength training. However, more research is needed on the residual effects of both interventions. Taken the drop-out rates (PT: 33%, ST: 21%) into account, muscle power training should also be used more carefully in prefrail older adults. Trial registration This trial has been registered with clinicaltrials.gov (NCT00783159)</p

Effects of Strength Training versus Power Training on Physical Performance in Prefrail Community-Dwelling Older Adults

Background: It has been unclear which training mode is most effective and feasible for improving physical performance in the risk group of prefrail community-dwelling older adults. Objective: The purpose of the present study was to compare the effects of strength training (ST) versus power training (PT) on functional performance in prefrail older adults. This study was registered at clinicaltrials.gov as NCT00783159. Methods: 69 community-dwelling older adults ( 1 65 years) who were prefrail according to the definition of Fried were included in a 12-week exercise program. The participants were randomized into an ST group, a PT group and a control group. All participants were supplemented with vitamin D 3 orally before entering the intervention period. The primary outcome was the global score on the Short Physical Performance Battery (SPPB). Secondary outcomes were muscle power, appendicular lean mass (aLM) measured by dual energy X-ray absorptiometry and self-reported functional deficits (Short Form of the Late-Life Func- tion and Disability Instrument, SF-LLFDI). Results: Regarding changes in the SPPB score during the intervention, significant heterogeneity between the groups was observed (p = 0.023). In pair-wise comparisons, participants in both training groups significantly (PT: p = 0.012, ST: 0.009) increased their SPPB score (PT: _ mean = 0.8, ST: _m ean = 1.0) compared to the control group, with no statistical difference among training groups (p = 0.301). No statistical differences were found in changes in aLM (p = 0.769), muscle power (p = 0.308) and SF-LLFDI (p = 0.623) between the groups. Muscle power significantly increased (p = 0.017) under vitamin D 3 intake. Conclusions: In prefrail community-dwelling adults, PT is not superior to ST, although both training modes resulted in significant improvements in physical performance. With regard to dropout rates, ST appears to be advantageous compared to PT. The high prevalence of vitamin D 3 deficiency and the slight improvement of physical performance under vitamin D 3 supplementation among study participants underline the relevance of this approach in physical exercise interventions

Bioluminescent Imaging of Genetically Selected Induced Pluripotent Stem Cell-Derived Cardiomyocytes after Transplantation into Infarcted Heart of Syngeneic Recipients

<div><p>Cell loss after transplantation is a major limitation for cell replacement approaches in regenerative medicine. To assess the survival kinetics of induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CM) we generated transgenic murine iPSC lines which, in addition to CM-specific expression of puromycin <i>N</i>-acetyl-transferase and enhanced green fluorescent protein (EGFP), also constitutively express firefly luciferase (FLuc) for bioluminescence (BL) <i>in vivo</i> imaging. While undifferentiated iPSC lines generated by random integration of the transgene into the genome retained stable FLuc activity over many passages, the BL signal intensity was strongly decreased in purified iPS-CM compared to undifferentiated iPSC. Targeted integration of FLuc-expression cassette into the ROSA26 genomic locus using zinc finger nuclease (ZFN) technology strongly reduced transgene silencing in iPS-CM, leading to a several-fold higher BL compared to iPS-CM expressing FLuc from random genomic loci. To investigate the survival kinetics of iPS-CM <i>in vivo</i>, purified CM obtained from iPSC lines expressing FLuc from a random or the ROSA26 locus were transplanted into cryoinfarcted hearts of syngeneic mice. Engraftment of viable cells was monitored by BL imaging over 4 weeks. Transplanted iPS-CM were poorly retained in the myocardium independently of the cell line used. However, up to 8% of cells survived for 28 days at the site of injection, which was confirmed by immunohistological detection of EGFP-positive iPS-CM in the host tissue. Transplantation of iPS-CM did not affect the scar formation or capillary density in the periinfarct region of host myocardium. This report is the first to determine the survival kinetics of drug-selected iPS-CM in the infarcted heart using BL imaging and demonstrates that transgene silencing in the course of iPSC differentiation can be greatly reduced by employing genome editing technology. FLuc-expressing iPS-CM generated in this study will enable further studies to reduce their loss, increase long-term survival and functional integration upon transplantation.</p></div

Luciferase activity and CD4 coreceptor expression in stable pCAG-CD4/FLuc iPSC clones decrease during differentiation <i>in vitro</i>.

<p><b>A.</b> Schematic representation of the plasmid used to generate transgenic cell line expressing FLuc and extracellular domain of CD4 receptor under the control of CAG-promoter. <b>B.</b> FLuc-activity in 17 pCAG-CD4/FLuc αPIG-iPSC clones after hygromycin selection. BL was measured in iPSC lysates in a luminescence plate reader (n = 3). Values are given as mean relative luminescent units (RLU) ± SD per µg protein. <b>C.</b> FLuc-activity of the two pCAG-CD4/FLuc αPIG-iPSC clones was measured in cell lysates before and after spontaneous <i>in vitro</i> differentiation and selection of cardiomyocytes (d16 CM). Values are given as mean RLU ± SD per µg protein (n = 3). *p<0.05; ***p<0.001. <b>D.</b> Flow cytometric analysis of CD4 expression on the surface of pCAG-CD4/FLuc iPSC clone C4 in pluripotent state, day 8 EB, day 16 EB and puromycin selected CM at day 16 of differentiation. <b>E.</b> Quantification of CD4 expression levels in pCAG-CD4/FLuc iPSC clones A1 and C4 at different stages of differentiation. Values are given as mean fluorescent intensity (MFI) ± SD of triplicate measurements. *p<0.05; **p<0.01, ***p<0.001, compared to MFI of CD4 expression in iPSC.</p