Xenografted adult human mesenchymal stem cells provide a platform for sustained biological pacemaker function in canine heart.

BACKGROUND: Biological pacemaking has been performed with viral vectors, human embryonic stem cells, and adult human mesenchymal stem cells (hMSCs) as delivery systems. Only with human embryonic stem cells are data available regarding stability for >2 to 3 weeks, and here, immunosuppression has been used to facilitate survival of xenografts. The purpose of the present study was to determine whether hMSCs provide stable impulse initiation over 6 weeks without the use of immunosuppression, the "dose" of hMSCs that ensures function over this period, and the catecholamine responsiveness of hMSC-packaged pacemakers. METHODS AND RESULTS: A full-length mHCN2 cDNA subcloned in a pIRES2-EGFP vector was electroporated into hMSCs. Transfection efficiency was estimated by GFP expression. I(HCN2) was measured with patch clamp, and cells were administered into the left ventricular anterior wall of adult dogs in complete heart block and with backup electronic pacemakers. Studies encompassed 6 weeks. I(HCN2) for all cells was 32.1+/-1.3 pA/pF (mean+/-SE) at -150 mV. Pacemaker function in intact dogs required 10 to 12 days to fully stabilize and persisted consistently through day 42 in dogs receiving > or =700,000 hMSCs (approximately 40% of which carried current). Rhythms were catecholamine responsive. Tissues from animals killed at 42 days manifested neither apoptosis nor humoral or cellular rejection. CONCLUSIONS: hMSCs provide a means for administering catecholamine-responsive biological pacemakers that function stably for 6 weeks and manifest no cellular or humoral rejection at that time. Cell doses >700,000 are sufficient for pacemaking when administered to left ventricular myocardium.

Treating tobacco dependence in clinically depressed smokers: effect of smoking cessation on mental health functioning.

We analyzed data from a randomized trial of 322 actively depressed smokers and examined the effect of smoking cessation on their mental health functioning. Only 1 of 10 measures at 4 follow-up time points was significant: participants who successfully stopped smoking reported less alcohol use than did participants who continued smoking. Depressive symptoms declined significantly over time for participants who stopped smoking and those who continued smoking; there were no group differences. Individuals in treatment for clinical depression can be helped to stop smoking without adversely affecting their mental health functioning.

Regenerative therapies in electrophysiology and pacing.

The prevention and treatment of cardiac arrhythmias conferring major morbidity and mortality is far from optimal, and relies heavily on devices and drugs for the partial successes that have been seen. The greatest success has been in the use of electronic pacemakers to drive the hearts of patients having high degree heart block. Recent years have seen the beginnings of attempts to use novel approaches available through gene and cell therapies to treat both brady- and tachyarrhythmias. By far the most successful approaches to date have been seen in the development of biological pacemakers. However, the far more difficult problems posed by atrial fibrillation and ventricular tachycardia are now being addressed. In the following pages we review the approaches now in progress as well as the specific methodologic demands that must be met if these therapies are to be successful.

Replacing damaged myocardium.

Heart failure survival after diagnosis has barely changed for more than half a century. Recently, investigation has focused on differentiation of stem cells in vitro and their delivery for use in vivo as replacement cardiac contractile elements. Here we report preliminary results using mesenchymal stem cells partially differentiated to a cardiac lineage in vitro. When delivered to the canine heart on an extracellular matrix patch to replace a full-thickness ventricular defect in vivo, they improve regional mechanical function. The delivered cells were also tracked, and some became myocytes with mature sarcomeres.

Treatment for cigarette smoking among depressed mental health outpatients: a randomized clinical trial.

OBJECTIVES: Using a brief contact control, we tested the efficacy of a staged care intervention to reduce cigarette smoking among psychiatric patients in outpatient treatment for depression. METHODS: We conducted a randomized clinical trial that included assessments at baseline and at months 3, 6, 12, and 18. Three hundred twenty-two patients in mental health outpatient treatment who were diagnosed with depression and smoked > or =1 cigarette per day participated. The desire to quit smoking was not a prerequisite for participation. Staged care intervention participants received computerized motivational feedback at baseline and at 3, 6, and 12 months and were offered a 6-session psychological counseling and pharmacological cessation treatment program. Brief contact control participants received a self-help guide and referral list of local smoking-treatment providers. RESULTS: As we hypothesized, abstinence rates among staged care intervention participants exceeded those of brief contact control participants at months 12 and 18. Significant differences favoring staged care intervention also were found in occurrence of a quit attempt and stringency of abstinence goal. CONCLUSION: The data suggest that individuals in psychiatric treatment for depression can be aided in quitting smoking through use of staged care interventions and that smoking cessation interventions used in the general population can be implemented in psychiatric outpatient settings.

Predictors of health functioning in two high-risk groups of smokers.

The relative and combined health effects of cigarette smoking, heroin use, and depression were examined in 322 clinically depressed smokers and 117 opioid-dependent smokers participating in two studies of the San Francisco Treatment Research Center. Opioid-dependent smokers averaged 16 years (S.D.=9) of heroin use; 3% of depressed smokers used opiates in the past 6 months. Cigarettes per day (M=15, S.D.=10) and Beck Depression (BDI-II) scores (M=21, S.D.=11) were comparable between the two groups. Health functioning was assessed using the Medical Outcomes Study Short Form (SF-36). Adjusting for demographic differences, depressed smokers reported better physical but poorer emotional health relative to opioid-dependent smokers. Both groups scored significantly lower than published norms (p<.05). Within groups, severity of depressive symptoms, tobacco use, and opiate use were independent predictors of lower health functioning (p<.05). Examining risk-related subgroups based on depression scores (BDI-II> or =20), cigarettes per day (> or =1 pack), and opiate use, number of risk factors was monotonically related to health functioning in both samples. Individuals with two or more risk factors scored the lowest (p<.05). Severity of depressive symptoms, tobacco use, and opiate use contributed individually and collectively to lower health functioning. Blended treatments that target multiple risk factors are needed to improve health outcomes.

Depressed smokers and stage of change: implications for treatment interventions.

Tobacco Dependence among smokers with psychiatric disorders has been under-addressed by the mental health, addictions, and tobacco control communities. This study examined depressed smokers' readiness to quit and the applicability of the Stages of Change framework to a psychiatric sample. Currently depressed smokers (N=322) were recruited from four outpatient psychiatric clinics. Participants averaged 16 cigarettes per day (S.D.=10) and 24 years (S.D.=13) of smoking. The majority (79%) reported intention to quit smoking with 24% ready to take action in the next 30 days. Individuals in the preparation stage reported more prior quit attempts, a greater commitment to abstinence, increased recognition of the cons of smoking, and greater use of the processes of change. Precontemplators were least likely to identify a goal related to their smoking behavior. Depressive symptom severity and history of recurrent depressive episodes were unrelated to readiness to quit. This study is one of the first to examine the smoking behaviors of currently depressed psychiatric outpatients. The level and longevity of their tobacco use underscore the need for cessation interventions. The consistency in hypothesized patterns among theoretical constructs of the Stages of Change model supports the transfer of stage-tailored interventions to this clinical population.

Increased myocyte content and mechanical function within a tissue-engineered myocardial patch following implantation.

During the past few years, studies involving the implantation of stem cells, chemical factors, and scaffolds have demonstrated the ability to augment the mammalian heart's native regenerative capacity. Scaffolds comprised of extracellular matrix (ECM) have been used to repair myocardial defects. These scaffolds become populated with myocytes and provide regional contractile function, but quantification of the myocyte population has not yet been conducted. The purpose of this study was to quantitate the myocyte content within the ECM bioscaffold and to correlate this cell population with the regional mechanical function over time. Xenogenic ECM scaffolds derived from porcine urinary bladder were implanted into a full-thickness, surgically induced, right ventricular-free wall defect in a dog model. Zero, 2, and 8 weeks following implantation, regional function and myocyte content were determined in each patch region. Regional function did not significantly increase from 0 to 2 weeks. At 8 weeks, however, regional stroke work increased to 3.7 +/- 0.7% and systolic contraction increased to 4.4 +/- 1.2%. The myocyte content also significantly increased during that period generating a linear relationship between regional function and myocyte content. In conclusion, ECM used as a myocardial patch increases both the regional function and the myocyte content over time. The mechanical function generated in the patch region is correlated with the quantity of local tissue myocytes.

Enhanced recovery of mechanical function in the canine heart by seeding an extracellular matrix patch with mesenchymal stem cells committed to a cardiac lineage.

The need to regenerate tissue is paramount, especially for the heart that lacks the ability to regenerate after injury. The urinary bladder extracellular matrix (ECM), when used to repair a right ventricular defect, successfully regenerated some mechanical function. The objective of the current study was to determine whether the regenerative effect of ECM could be improved by seeding the patch with human mesenchymal stem cells (hMSCs) enhanced to differentiate down a cardiac linage. hMSCs were used to form three-dimensional spheroids. The expression of cardiac proteins was determined in cells exposed to the spheroid formation and compared with nonmanipulated hMSCs. To determine whether functional calcium channels were present, the cells were patch clamped. To evaluate the ability of these cells to regenerate mechanical function, the spheroids were seeded on ECM and then implanted into the canine heart to repair a full-thickness right ventricular defect. As a result, many of the cells spreading from the spheroids expressed cardiac-specific proteins, including sarcomeric alpha-actinin, cardiotin, and atrial natriuretic peptide, as well as the cell cycle markers cyclin D1 and proliferating cell nuclear antigen. A calcium current similar in amplitude to that of ventricular myocytes was present in 16% of the cells. The cardiogenic cell-seeded scaffolds increased the regional mechanical function in the canine heart compared with the unmanipulated hMSC-seeded scaffolds. In addition, the cells prelabeled with fluorescent markers demonstrated myocyte-specific actinin staining with sarcomere spacing similar to that of normal myocytes. In conclusion, the spheroid-derived cells express cardiac-specific proteins and demonstrate a calcium current similar to adult ventricular myocytes. When these cells are implanted into the canine heart, some of these cells appear striated and mechanical function is improved compared with the unmanipulated hMSCs. Further investigation will be required to determine whether the increased mechanical function is due to a differentiation of the cardiogenic cells to myocytes or to other effects.

Finding fluorescent needles in the cardiac haystack: tracking human mesenchymal stem cells labeled with quantum dots for quantitative in vivo three-dimensional fluorescence analysis.

Stem cells show promise for repair of damaged cardiac tissue. Little is known with certainty, however, about the distribution of these cells once introduced in vivo. Previous attempts at tracking delivered stem cells have been hampered by the autofluorescence of host tissue and limitations of existing labeling techniques. We have developed a novel loading approach to stably label human mesenchymal stem cells with quantum dot (QD) nanoparticles. We report the optimization and validation of this long-term tracking technique and highlight several important biological applications by delivering labeled cells to the mammalian heart. The bright QD crystals illuminate exogenous stem cells in histologic sections for at least 8 weeks following delivery and permit, for the first time, the complete three-dimensional reconstruction of the locations of all stem cells following injection into the heart. Disclosure of potential conflicts of interest is found at the end of this article.

Acceptance of nicotine dependence treatment among currently depressed smokers.

This study reports on baseline characteristics associated with acceptance and refusal of available smoking treatment among currently depressed smokers in a psychiatric outpatient clinic who were enrolled in a larger clinical trial. The sample (N=154) was 68% female and 72% White, with a mean age of 41.4 years and average smoking rate of 17 cigarettes/day. All participants were assigned to a repeated contact experimental condition; received a stage-based expert system program to facilitate treatment acceptance; and were then offered smoking treatment, consisting of behavioral counseling, nicotine patch, and bupropion. Acceptors (n=53) were defined as those accepting behavioral counseling and pharmacological treatment at some point during the 18-month study, whereas refusers (n=101) received only the expert system. The number of days to treatment acceptance was significantly predicted by stage of change, with those in preparation entering treatment more quickly than contemplators or precontemplators. In a logistic regression, the variables most strongly associated with accepting treatment were current use of psychiatric medication and perceived success for quitting. Severity of depressive symptoms, duration of depression history, and history of recurrent depression were not related to treatment acceptance. Findings have implications for the psychiatric assessment and treatment of smokers in clinical settings. Psychiatric medication may play a significant role in smoking cessation treatment acceptance by currently depressed smokers.

Mapping mechanical strain of an endogenous cytoskeletal network in living endothelial cells.

A central aspect of cellular mechanochemical signaling is a change of cytoskeletal tension upon the imposition of exogenous forces. Here we report measurements of the spatiotemporal distribution of mechanical strain in the intermediate filament cytoskeleton of endothelial cells computed from the relative displacement of endogenous green fluorescent protein (GFP)-vimentin before and after onset of shear stress. Quantitative image analysis permitted computation of the principal values and orientations of Lagrangian strain from 3-D high-resolution fluorescence intensity distributions that described intermediate filament positions. Spatially localized peaks in intermediate filament strain were repositioned after onset of shear stress. The orientation of principal strain indicated that mechanical stretching was induced across cell boundaries. This novel approach for intracellular strain mapping using an endogenous reporter demonstrates force transfer from the lumenal surface throughout the cell.