Thiamine increases resident endoglin positive cardiac progenitor cells and atrial contractile force in humans: A randomised controlled trial.

BACKGROUND: The heart has an intrinsic ability to regenerate, orchestrated by progenitor or stem cells. However, the relative complexity of non-resident cardiac progenitor cell (CPC) therapy makes modulation of resident CPCs a more attractive treatment target. Thiamine analogues improve resident CPC function in pre-clinical models. In this double blinded randomised controlled trial (identifier: ACTRN12614000755639), we examined whether thiamine would improve CPC function in humans. METHODS AND RESULTS: High dose oral thiamine (one gram twice daily) or matching placebo was administered 3-5 days prior to coronary artery bypass surgery (CABG). Right atrial appendages were collected at the time of CABG, and CPCs isolated. There was no difference in the primary outcome (proliferation ability of CPCs) between treatment groups. Older age was not associated with decreased proliferation ability. In exploratory analyses, isolated CPCs in the thiamine group showed an increase in the proportion of CD34-/CD105+ (endoglin) cells, but no difference in CD34-/CD90+ or CD34+ cells. Thiamine increased maximum force developed by isolated trabeculae, with no difference in relaxation time or beta-adrenergic responsiveness. CONCLUSION: Thiamine does not improve proliferation ability of CPC in patients undergoing CABG, but increases the proportion of CD34-/CD105+ cells. Having not met its primary endpoint, this study provides the impetus to re-examine CPC biology prior to any clinical outcome-based trial examining potential beneficial cardiovascular effects of thiamine.

Frailty in Elderly Patients Undergoing Cardiac Surgery Increases Hospital Stay and 12-Month Readmission Rate.

BACKGROUND: Cardiac surgery risk scoring systems predict operative mortality but not outcomes related to preoperative frailty. The aim of this study was to assess frailty in a cohort of older cardiac surgery patients as a predictor of postoperative outcomes. METHODS: Prospective data was collected on patients 65 years of age and older undergoing cardiac surgery between September 2015 and October 2016 at Dunedin Hospital. Frailty was assessed with the Edmonton frail scale and five-metre gait speed. The primary endpoint was length of hospital stay. Secondary outcomes included postoperative complications, major adverse events, death and 12-month readmission rate. RESULTS: Among the 96 patients, median age was 74 (interquartile range 10.5) and 65 (68%) were males. Of the sample 64 (67%) were scored as not frail, 22 (23%) as vulnerable, and 10 (10%) as frail. The median (interquartile range) postoperative days' stay were: not frail 6 (2), vulnerable 9.5 (8), and frail 15 (13). Survival analysis adjusting for EuroSCORE II, age, sex and surgery type showed that greater Edmonton Frail Scale scores were independently predictive of longer post-surgery hospital stay with a hazard ratio for discharge of 0.83 (95% confidence interval 0.76-0.91, p<0.001) per point. The Edmonton Frail Scale score was associated with the 12-month post discharge number of readmissions (adjusted incidence rate ratio 1.24 (95% confidence interval 1.13-1.37, p<0.001) per point. CONCLUSIONS: The Edmonton Frail Scale score predicts length of hospital stay post cardiac surgery and 12-month readmission rate in patients older than 65 years of age.

Early dysregulation of cardiac-specific microRNA-208a is linked to maladaptive cardiac remodelling in diabetic myocardium.

BACKGROUND: The diabetic heart undergoes remodelling contributing to an increased incidence of heart failure in individuals with diabetes at a later stage. The molecular regulators that drive this process in the diabetic heart are still unknown. METHODS: Real-time (RT) PCR analysis was performed to determine the expression of cardiac specific microRNA-208a in right atrial appendage (RAA) and left ventricular (LV) biopsy tissues collected from diabetic and non-diabetic patients undergoing coronary artery bypass graft surgery. To determine the time-dependent changes, cardiac tissue were collected from type 2 diabetic mice at different age groups. A western blotting analysis was conducted to determine the expression of contractile proteins α- and ß-myosin heavy chain (MHC) and thyroid hormone receptor-α (TR-α), the negative regulator of ß-MHC. To determine the beneficial effects of therapeutic modulation of miR-208a, high glucose treated adult mouse HL-1 cardiomyocytes were transfected with anti-miR-208a. RESULTS: RT-PCR analysis showed marked upregulation of miR-208a from early stages of diabetes in type 2 diabetic mouse heart, which was associated with a marked increase in the expression of pro-hypertrophic ß-MHC and downregulation of TR-α. Interestingly, upregulation of miR-208a preceded the switch of α-/ß-MHC isoforms and the development of diastolic and systolic dysfunction. We also observed significant upregulation of miR-208a and modulation of miR-208a associated proteins in the type 2 human diabetic heart. Therapeutic inhibition of miR-208a activity in high glucose treated HL-1 cardiomyocytes prevented the activation of ß-MHC and hence the hypertrophic response. CONCLUSION: Our results provide the first evidence that early modulation of miR-208a in the diabetic heart induces alterations in the downstream signaling pathway leading to cardiac remodelling and that therapeutic inhibition of miR-208a may be beneficial in preventing diabetes-induced adverse remodelling of the heart.

Down-regulation of miR-15a/b accelerates fibrotic remodelling in the Type 2 diabetic human and mouse heart.

Aim: Myocardial fibrosis is a well-established cause of increased myocardial stiffness and subsequent diastolic dysfunction in the diabetic heart. The molecular regulators that drive the process of fibrotic events in the diabetic heart are still unknown. We determined the role of the microRNA (miR)-15 family in fibrotic remodelling of the diabetic heart.Methods and results: Right atrial appendage (RAA) and left ventricular (LV) biopsy tissues collected from diabetic and non-diabetic (ND) patients undergoing coronary artery bypass graft surgery showed significant down-regulation of miR-15a and -15b. This was associated with marked up-regulation of pro-fibrotic transforming growth factor-ß receptor-1 (TGFßR1) and connective tissue growth factor (CTGF), direct targets for miR-15a/b and pro-senescence p53 protein. Interestingly, down-regulation of miR-15a/b preceded the development of diastolic dysfunction and fibrosis in Type 2 diabetic mouse heart. Therapeutic restoration of miR-15a and -15b in HL-1 cardiomyocytes reduced the activation of pro-fibrotic TGFßR1 and CTGF, and the pro-senescence p53 protein expression, confirming a causal regulation of these fibrotic and senescence mediators by miR-15a/b. Moreover, conditioned medium (CM) collected from cardiomyocytes treated with miR-15a/b markedly diminished the differentiation of diabetic human cardiac fibroblasts.Conclusion: Our results provide first evidence that early down-regulation of miR-15a/b activates fibrotic signalling in diabetic heart, and hence could be a potential target for the treatment/prevention of diabetes-induced fibrotic remodelling of the heart.

Integrated microRNA and messenger RNA analysis in aortic stenosis.

Aortic valve stenosis (AS) is a major cause of morbidity and mortality, with no effective medical therapies. Investigation into the underlying biology of AS in humans is limited by difficulties in obtaining healthy valvular tissue for use as a control group. However, micro-ribonucleic acids (miRNAs) are stable in post-mortem tissue. We compared valve specimens from patients undergoing aortic valve replacement for AS to non-diseased cadaveric valves. We found 106 differentially expressed miRNAs (p < 0.05, adjusted for multiple comparisons) on microarray analysis, with highly correlated expression among up- and down-regulated miRNAs. Integrated miRNA/gene expression analysis validated the microarray results as a whole, while quantitative polymerase chain reaction confirmed downregulation of miR-122-5p, miR-625-5p, miR-30e-5p and upregulation of miR-21-5p and miR-221-3p. Pathway analysis of the integrated miRNA/mRNA network identified pathways predominantly involved in extracellular matrix function. A number of currently available therapies target products of upregulated genes in the integrated miRNA/mRNA network, with these genes being predominantly more peripheral members of the network. The identification of a group of tissue miRNA associated with AS may contribute to the development of new therapeutic approaches to AS. This study highlights the importance of systems biology-based approaches to complex diseases.

Type-2 diabetes increases autophagy in the human heart through promotion of Beclin-1 mediated pathway.

BACKGROUND: Diabetes promotes progressive loss of cardiac cells, which are replaced by a fibrotic matrix, resulting in the loss of cardiac function. In the current study we sought to identify if excessive autophagy plays a major role in inducing this progressive loss. METHODS AND RESULTS: Immunofluorescence and western blotting analysis of the right atrial appendages collected from diabetic and non-diabetic patients undergoing coronary artery bypass graft surgery showed a marked increase in the level of autophagy in the diabetic heart, as evidenced by increased expression of autophagy marker LC3B-II and its mediator Beclin-1 and decreased expression of p62, which incorporates into autophagosomes to be efficiently degraded. Moreover, a marked activation of pro-apoptotic caspase-3 was observed. Electron microscopy showed increased autophagosomes in the diabetic heart. In vivo measurement of autophagic flux by choloroquine injection resulted in further enhancement of LC3B-II in the diabetic myocardium, confirming increased autophagic activity in the type-2 diabetic heart. Importantly, in-vitro genetic depletion of beclin-1 in high glucose treated adult rat cardiomyocytes markedly inhibited the level of autophagy and subsequent apoptotic cell death. CONCLUSIONS: These findings demonstrate the pathological role of autophagy in the type-2 diabetic heart, opening up a potentially novel therapeutic avenue for the treatment of diabetic heart disease.

Data supporting the activation of autophagy genes in the diabetic heart.

This data article contains full list of autophagy related genes that are altered in diabetic heart. This article also shows data from in vitro cultured cardiomyocytes that are exposed the high glucose treatment to simulate hyperglycemic state in vitro. The interpretation of these data and further extensive insights into the regulation of SG biogenesis by AMPK can be found in "Type-2 diabetes increases autophagy in the human heart through promotion of Beclin-1 mediated pathway" (Munasinghe et al., in press) [1].

Impaired relaxation despite upregulated calcium-handling protein atrial myocardium from type 2 diabetic patients with preserved ejection fraction.

BACKGROUND: Diastolic dysfunction is a key factor in the development and pathology of cardiac dysfunction in diabetes, however the exact underlying mechanism remains unknown, especially in humans. We aimed to measure contraction, relaxation, expression of calcium-handling proteins and fibrosis in myocardium of diabetic patients with preserved systolic function. METHODS: Right atrial appendages from patients with type 2 diabetes mellitus (DM, n = 20) and non-diabetic patients (non-DM, n = 36), all with preserved ejection fraction and undergoing coronary artery bypass grafting (CABG), were collected. From appendages, small cardiac muscles, trabeculae, were isolated to measure basal and ß-adrenergic stimulated myocardial function. Expression levels of calcium-handling proteins, sarcoplasmic reticulum Ca2+ ATPase (SERCA2a) and phospholamban (PLB), and of ß1-adrenoreceptors were determined in tissue samples by Western blot. Collagen deposition was determined by picro-sirius red staining. RESULTS: In trabeculae from diabetic samples, contractile function was preserved, but relaxation was prolonged (Tau: 74 ± 13 ms vs. 93 ± 16 ms, non-DM vs. DM, p = 0.03). The expression of SERCA2a was increased in diabetic myocardial tissue (0.75 ± 0.09 vs. 1.23 ± 0.15, non-DM vs. DM, p = 0.007), whereas its endogenous inhibitor PLB was reduced (2.21 ± 0.45 vs. 0.42 ± 0.11, non-DM vs. DM, p = 0.01). Collagen deposition was increased in diabetic samples. Moreover, trabeculae from diabetic patients were unresponsive to ß-adrenergic stimulation, despite no change in ß1-adrenoreceptor expression levels. CONCLUSIONS: Human type 2 diabetic atrial myocardium showed increased fibrosis without systolic dysfunction but with impaired relaxation, especially during ß-adrenergic challenge. Interestingly, changes in calcium-handling protein expression suggests accelerated active calcium re-uptake, thus improved relaxation, indicating a compensatory calcium-handling mechanism in diabetes in an attempt to maintain diastolic function at rest despite impaired relaxation in the diabetic fibrotic atrial myocardium. Our study addresses important aspects of the underlying mechanisms of diabetes-associated diastolic dysfunction, which is crucial to developing new therapeutic treatments.

Rapid onset of cardiomyopathy in STZ-induced female diabetic mice involves the downregulation of pro-survival Pim-1.

BACKGROUND: Diabetic women are five times more likely to develop congestive heart failure compared with two fold for men. The underlying mechanism for this gender difference is not known. Here we investigate the molecular mechanisms responsible for this female disadvantage and attempt safeguarding cardiomyocytes viability and function through restoration of pro-survival Pim-1. METHODS AND RESULTS: Diabetes was induced by injection of streptozotocin in CD1 mice of both genders. Functional and dimensional parameters measurement using echocardiography revealed diastolic dysfunction in female diabetic mice within 8 weeks after STZ-induced diabetes. This was associated with significant downregulation of pro-survival Pim-1 and upregulation of pro-apoptotic Caspase-3, microRNA-1 and microRNA-208a. Male diabetic mice did not show any significant changes at this time point (P < 0.05 vs. female diabetic). Further, the onset of ventricular remodelling was quicker in female diabetic mice showing marked left ventricular dilation, reduced ejection fraction and poor contractility (P < 0.05 vs. male diabetic at 12 and 16 weeks of STZ-induced diabetes). Molecular analysis of samples from human diabetic hearts confirmed the results of pre-clinical studies, showing marked downregulation of Pim-1 in the female diabetic heart (P < 0.05 vs. male diabetic). Finally, in vitro restoration of Pim-1 reversed the female disadvantage in diabetic cardiomyocytes. CONCLUSIONS: We provide novel insights into the molecular mechanisms behind the rapid onset of cardiomyopathy in female diabetics. These results suggest the requirement for the development of gender-specific treatments for diabetic cardiomyopathy.

Kinky circumflex grafts: the left appendage flip maneuver.

The "too-long," or redundant, circumflex graft is notorious for its tendency to form a kink. The acute angle (the kink) typically occurs over the front of the pulmonary trunk. Described herein are (1) the left appendage flip maneuver, a simple solution to correct graft kink, and (2) analysis and explanation of vein graft kink.

Top-end aortocoronary anastomosis made easy: the visor technique.

The importance of the proximal aortocoronary anastomosis is overshadowed by its lack of literary attention. This aortic connection, sometimes difficult to perform, is simplified by the Visor technique.

Rupture of aortic pseudoaneursym arising from a transverse aortotomy line 12 years following mechanical aortic valve replacement.

Pseudoaneurysm formation at the site of aortotomy is a rare complication following aortic valve replacement; it appears to be more common in those whose valve is replaced for endocarditis or in those who develop an early postoperative endocarditis or sepsis. We discuss the case of a 77-year-old male, who presented 12 years following mechanical aortic valve replacement, with rupture of an ascending aortic pseudoaneurysm arising from a transverse aortotomy line.