Rationale: Gq-coupled receptors are thought to play a critical role in the induction of left vent... more Rationale: Gq-coupled receptors are thought to play a critical role in the induction of left ventricular hypertrophy (LVH) secondary to pressure overload, although mechano-sensitive channel activation by a variety of mechanisms has also been proposed, and the relative importance of calcineurin-and calmodulin kinase II (CaMKII)-dependent hypertrophic pathways remains controversial. Objective: To determine the mechanisms regulating the induction of LVH in response to mechanical pressure overload. Methods and Results: Transgenic mice with cardiac-targeted inhibition of Gq-coupled receptors (GqI mice) and their non-transgenic littermates (NTL) were subjected to neurohumoral stimulation (continuous, subcutaneous angiotensin II (AngII) infusion for 14 days) or mechanical pressure overload (transverse aortic arch constriction (TAC) for 21 days) to induce LVH. Candidate signalling pathway activation was examined. As expected, LVH observed in NTL mice with AngII infusion was attenuated in heterozygous (GqI +/-) mice and absent in homozygous (GqI-/-) mice. In contrast, LVH due to TAC was unaltered by either heterozygous or homozygous Gq inhibition. Gene expression of atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP) and α-skeletal actin (α-SA) was increased 48 hours after AngII infusion or TAC in NTL mice; in GqI mice, the increases in ANP, BNP and α-SA in response to AngII were completely absent, as expected, but all three increased after TAC. Increased nuclear translocation of nuclear factor of activated T-cells c4 (NFATc4), indicating calcineurin pathway activation, occurred in NTL mice with AngII infusion but not TAC, and was prevented in GqI mice infused with AngII. Nuclear and cytoplasmic CaMKIIδ levels increased in both NTL and GqI mice after TAC but not AngII infusion, with increased cytoplasmic phospho-and total histone deacetylase 4 (HDAC4) and increased nuclear myocyte enhancer factor 2 (MEF2) levels. Conclusion: Cardiac Gq receptors and calcineurin activation are required for neurohumorally mediated LVH but are not required for LVH induced by mechanical pressure overload (TAC); the latter is mediated by activation of the CaMKII-HDAC4-MEF2 pathway.
1. Regional differences in odorant-induced responsiveness of the rat olfactory epithelium were me... more 1. Regional differences in odorant-induced responsiveness of the rat olfactory epithelium were measured via electrophysiological recordings [negative component of electro-olfactogram (Veog(-)) made from the surface of the olfactory epithelium on the nasal septum]. The nasal septum provided a flat surface from which multiple recordings could be made. 2. Veog(-)s were recorded from a standardized grid of 16 sites. This grid of recording sites extended over most of the surface of the olfactory epithelium on the nasal septum. 3. Twenty-one animals were tested for their responses to seven odorants. The animals were divided into three groups, each of which was tested with two different odorants plus amyl acetate, which provided a comparison between the groups. 4. For each odorant in each animal, topographic maps of relative responsiveness were derived to test whether odorants elicited different patterns of responses in the same individual. Topographic maps of responsiveness were derived also for the animal groups to test for the generality of the form of the maps for different odorants. Response latencies were also measured for each odorant at each recording site. 5. All individuals showed different topographic patterns of responses to the three test odorants. For most odorants, the location of the most responsive site was similar in all animals. In different animals the topographic maps for the same odorant were remarkably similar. Topographic maps for the odorants were all different from one another. 6. These results are consistent with the hypothesis that odorant quality is encoded in the differential spatial distribution of receptor cells whose differences in responsiveness appear to be distributed as a continuum across the epithelium. The results establish for a mammalian species what was previously reported in amphibians. These differences are presumed to be due to differential expression of odorant receptor proteins. 7. The mean response latency was 32 ms. This period was similar for all odorants, all animals, and all recording sites and was independent of Veog(-) amplitude. It is concluded that diffusion through the mucus contributed approximately 6 ms to the latency of onset of the responses to these odorants.
Recombinant human platelet-derived growth factor-AB improves cardiac function and survival after ... more Recombinant human platelet-derived growth factor-AB improves cardiac function and survival after myocardial infarction in a porcine model.
Animal models of pressure overload are valuable for understanding hypertensive heart disease. We ... more Animal models of pressure overload are valuable for understanding hypertensive heart disease. We characterised a surgical model of pressure overload-induced hypertrophy in C57BL/6J mice produced by suprarenal aortic constriction (SAC). Compared to sham controls, at one week post-SAC systolic blood pressure was significantly elevated and left ventricular (LV) hypertrophy was evident by a 50% increase in the LV weight-to-tibia length ratio due to cardiomyocyte hypertrophy. As a result, LV end-diastolic wall thickness-to-chamber radius (h/R) ratio increased, consistent with the development of concentric hypertrophy. LV wall thickening was not sufficient to normalise LV wall stress, which also increased, resulting in LV systolic dysfunction with reductions in ejection fraction and fractional shortening, but no evidence of heart failure. Pathological LV remodelling was evident by the re-expression of fetal genes and coronary artery perivascular fibrosis, with ischaemia indicated by enhanced cardiomyocyte Hif1a expression. The expression of stem cell factor receptor, c-Kit, was low basally in cardiomyocytes and did not change following the development of robust hypertrophy, suggesting there is no role for cardiomyocyte c-Kit signalling in pathological LV remodelling following pressure overload. Heart failure is a common end-result of a variety of cardiovascular diseases, including ischaemic and hypertensive heart disease. Hypertension is the primary cause of pathological left ventricular (LV) hypertrophy in 75% of patients 1. While the hypertrophic response is initially compensatory to maintain heart function in the face of pressure overload (PO), pathological hypertrophy eventually becomes decompensatory, with decreased cardiac performance that precedes overt heart failure. Current treatments for heart failure merely slow the progression to end-stage heart failure, since there is no cure other than heart transplantation. Animal models of hypertensive heart disease are, thus, critical to developing potential new regenerative therapies 2. Two commonly used cardiac injury models are myocardial infarction (MI) produced by ligation of the left anterior descending coronary artery, and aortic constriction; the latter first developed and characterised by Rytand 3 and Goldblatt et al. 4 , produced by ligating the descending abdominal aorta between or immediately above the renal arteries. Constriction of the abdominal aorta induces PO leading to the rapid development of cardiac hypertrophy and heart failure 2,5. Historically, aortic constriction models were developed in larger animal species, including rabbits, dogs, guinea pigs and rats 2,5 , but over the last few decades these techniques have been adapted to mice-the preferred species for genetic studies that are also economical 6,7. Thoracic aortic constriction
B rain death causes a surge in plasma catecholamine levels, which leads to acute cardiac dysfunct... more B rain death causes a surge in plasma catecholamine levels, which leads to acute cardiac dysfunction. 1-3 It also alters the response of the myocardium to the hypothermic injury and the ischaemic reperfusion injury associated with conventional preservation and storage of the donor heart. 1-3 Minimising these combined injuries through improvements in preservation would increase the potential donor pool and improve outcome following cardiac transplantation. The objective of this brief communication is to present a large animal model of orthotopic cardiac transplantation, incorporating a brain dead donor, that we have developed to assess new strategies for cardiac allograft preservation. Final Model Methodology The experimental protocol has been approved by the St Vincent's Hospital animal experimentation ethics committee and was in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes. Inbred sibling Westran pigs (20-50 kg) were obtained in pairs. The donor pig was pre-medicated with an intramuscular injection of ketamine (10 mg/kg), midazolam (1 mg/kg) and atropine (1.2 mg). General anaesthesia was induced with intravenous thiopentone (50 mg boluses, to effect), and maintained by inhaled isoflurane (1-3% inhaled gas) and intravenous fentanyl (5 µg/kg boluses). The pig was intubated and ventilated with 100% oxygen. Normal (0.9%) saline was infused
European Journal of Cardio-Thoracic Surgery, Nov 1, 2002
Objective: Paradoxically, it has been reported that after 1.5-4 h of hypothermic ischaemic preser... more Objective: Paradoxically, it has been reported that after 1.5-4 h of hypothermic ischaemic preservation there is complete recovery of contractile function in canine cardiac allografts, as assessed by the preload recruitable stroke work (PRSW) relationship. This raises questions about the suitability of the canine heart as a model for preservation research and the PRSW relationship as an end-point. The aim of the present study was to evaluate the PRSW relationship as an index of left ventricular contractility in porcine cardiac allografts. Methods: Eighteen orthotopic heart transplants were performed in inbred Westran pigs. Brain death was induced in the donor pigs 1 h prior to explantation. The donor hearts were arrested with extracellular cardioplegia, which was stored in ice prior to administration. On explantation, the donor hearts were immersed in cardioplegia and stored in ice. The donor hearts were subjected to either 4 (IT4, n ¼ 6), 6 (IT6, n ¼ 9) or 14 (IT14, n ¼ 3) h of ischaemia. Post-transplant, all hearts were supported with dobutamine (10 mcg/kg per min). The PRSW relationship was derived from pressure-volume loops obtained by epicardial sonomicrometry and transmyocardial micromanometry. Multiple linear regression was used to describe and compare the PRSW relationship before brain death in the donor and after weaning from bypass in the recipient. Results: Eleven hearts were weaned successfully from cardiopulmonary bypass: IT4 100% (6/6), IT6 56% (5/9) and IT14 0% (0/3) (IT4 versus IT14: P ¼ 0:012). Analysis of the PRSW relationship revealed a reduction in contractility in both the IT4 and IT6 groups (both P , 0:0001), but a greater reduction in the IT6 group (P , 0:0001). Notably, the volume-axis intercept of the PRSW relationship was found to be a better discriminator of post-preservation contractile dysfunction than the slope of the PRSW relationship. Conclusions: The porcine heart's susceptibility to ischaemic injury makes it ideal for evaluating the effect of different preservation strategies on contractile recovery. The PRSW relationship can be used to evaluate the differences in contractile recovery, though the nature of the effect of ischaemic preservation necessitates analysis by multiple linear regression.
Journal of Molecular and Cellular Cardiology, Jun 1, 2021
The 'fight or flight' response to physiological stress involves sympathetic nervous syste... more The 'fight or flight' response to physiological stress involves sympathetic nervous system activation, catecholamine release and adrenergic receptor stimulation. In the heart, this induces positive inotropy, previously attributed to the β1-adrenergic receptor subtype. However, the role of the α1A-adrenergic receptor, which has been suggested to be protective in cardiac pathology, has not been investigated in the setting of physiological stress. To explore this, we developed a tamoxifen-inducible, cardiomyocyte-specific α1A-adrenergic receptor knock-down mouse model, challenged mice to four weeks of endurance swim training and assessed cardiac outcomes. With 4-OH tamoxifen treatment, expression of the α1A-adrenergic receptor was knocked down by 80-89%, without any compensatory changes in the expression of other adrenergic receptors, or changes to baseline cardiac structure and function. Swim training caused eccentric hypertrophy, regardless of genotype, demonstrated by an increase in heart weight/tibia length ratio (30% and 22% in vehicle- and tamoxifen-treated animals, respectively) and an increase in left ventricular end diastolic volume (30% and 24% in vehicle- and tamoxifen-treated animals, respectively) without any change in the wall thickness/chamber radius ratio. Consistent with physiological hypertrophy, there was no increase in fetal gene program (Myh7, Nppa, Nppb or Acta1) expression. In response to exercise-induced volume overload, stroke volume (39% and 30% in vehicle- and tamoxifen-treated animals, respectively), cardiac output/tibia length ratio (41% in vehicle-treated animals) and stroke work (61% and 33% in vehicle- and tamoxifen-treated animals, respectively) increased, regardless of genotype. These findings demonstrate that cardiomyocyte α1A-adrenergic receptors are not necessary for cardiac adaptation to endurance exercise stress and their acute ablation is not deleterious.
Pathological left ventricular hypertrophy (LVH) occurs in response to pressure overload and remai... more Pathological left ventricular hypertrophy (LVH) occurs in response to pressure overload and remains the single most important clinical predictor of cardiac mortality. The molecular pathways in the induction of pressure overload LVH are potential targets for therapeutic intervention. Current treatments aim to remove the pressure overload stimulus for LVH, but do not completely reverse adverse cardiac remodelling. Although numerous molecular signalling steps in the induction of LVH have been identified, the initial step by which mechanical stretch associated with cardiac pressure overload is converted into a chemical signal that initiates hypertrophic signalling remains unresolved. In this study, we show that selective deletion of transient receptor potential melastatin 4 (TRPM4) channels in mouse cardiomyocytes results in an approximately 50% reduction in the LVH induced by transverse aortic constriction. Our results suggest that TRPM4 channel is an important component of the mechanosensory signalling pathway that induces LVH in response to pressure overload and represents a potential novel therapeutic target for the prevention of pathological LVH.
Mutations in the giant sarcomeric protein titin (TTN) are a major cause for inherited forms of di... more Mutations in the giant sarcomeric protein titin (TTN) are a major cause for inherited forms of dilated cardiomyopathy (DCM). We have previously developed a mouse model that imitates a TTN truncation mutation we found in a large pedigree with DCM. While heterozygous Ttn knock-in mice do not display signs of heart failure under sedentary conditions, they recapitulate the human phenotype when exposed to the pharmacological stressor angiotensin II or isoproterenol. In this study we investigated the effects of pressure overload by transverse aortic constriction (TAC) in heterozygous (Het) Ttn knock-in mice. Two weeks after TAC, Het mice developed marked impairment of left ventricular ejection fraction (< 0.05), while wild-type (WT) TAC mice did not. Het mice also trended toward increased ventricular end diastolic pressure and volume compared to WT littermates. We found an increase in histologically diffuse cardiac fibrosis in Het compared to WT in TAC mice. This study shows that a pattern of DCM can be induced by TAC-mediated pressure overload in a TTN-truncated mouse model. This model enlarges our arsenal of cardiac disease models, adding a valuable tool to understand cardiac pathophysiological remodeling processes and to develop therapeutic approaches to combat heart failure.
Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA... more Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA and Western world. Polyphenolic compounds are present in our daily diet such as vegetable, fruit, onion, and processed food such as red wine, and dark chocolate. Flavonoids, a member of polyphenols and related compounds have significant antioxidant and anti-inflammatory activity, which could vary based on their structures. Recent studies indicated that quercetin; a flavonol reduces the health risks linked with obesity, heart disease, diabetes and high blood pressure. The objective: This study is designed to investigate the effects of quercetin intake and exercise in atherosclerosis. Study Design: The study compares the effect of intake of quercetin with exercise on atherosclerotic plaque and biomarkers of atherosclerosis. We conducted three separate studies as follows: Study 1: C57BL6 wild type female mice (N¼ 35). Study 2: C57BL6 wild type male mice (N¼ 40) and Study 3: C57BL6 LDLr-/-mice (N¼ 40). Mice were fed normal mouse chow or atherogenic diet and orally supplied with 100 μg/day of quercetin with or without exercising. In each study animals were divided into four groups (10 each). These groups were as follows: Control mice, left untreated; control quercetin group, exercise group without quercetin, and exercise group with quercetin supplements. The exercise groups were run on a treadmill for 30 minutes, 15-30 m/m/ 5 days/week for 30-60 days. At the end of the 1-2 month of treatment, mice were sacrificed. Livers, aortas, adipose tissue and blood were collected. Plasma lipids, and inflammatory biomarkers, and liver and aorta gene expressions and western blot were performed. Results: In wildtype males lipids modulators such as PCSK9 mRNA levels were significantly up-regulated with combination of exercise and quercetin intake (p o 0.05). ANGPLT4 mRNA levels significantly down-regulated with the combination after 8 weeks of exercise and quercetin intake compared to both
Journal of Heart and Lung Transplantation, Mar 1, 2003
Background: U74389G (16-desmethyl tirilazad), a 21-aminosteroid or "lazaroid," inhibits lipid per... more Background: U74389G (16-desmethyl tirilazad), a 21-aminosteroid or "lazaroid," inhibits lipid peroxidation, which is an important element of ischemia-reperfusion injury. The aim of this study was to determine whether the addition of U74389G to the cardioplegic preservation solution could improve early cardiac allograft function. Methods: A porcine model of donor brain death and orthotopic cardiac transplantation was used. Hearts were arrested and preserved for 6 hours in an aspartate-enriched extracellular cardioplegia that had been supplemented with either U74389G and its carrier (n ϭ 7) or the carrier alone (n ϭ 9). Epicardial sonomicrometry and transmyocardial micromanometry were used to obtain pressure-volume loops before and after transplantation. Left ventricular wall volume was measured by volume displacement. Results: A higher proportion of U74389G-treated hearts were weaned successfully from cardiopulmonary bypass, but this difference did not achieve statistical significance (86% [6 of 7] vs 56% [5 of 9]; p ϭ 0.308). In the hearts that were weaned successfully, preservation of left ventricular contractility, as judged by the pre-load recruitable stroke work relationship, was significantly better in the U74389G-treated hearts (p ϭ 0.0271). In contrast, left ventricular compliance, as judged by the end-diastolic pressure-volume relationship, was significantly better preserved in the control group (p Ͻ 0.0001). U74389G-treated hearts developed less myocardial edema, as judged by the posttransplant left ventricular wall volume/baseline steady-state epicardial end-diastolic volume ratio (64 Ϯ 9% vs 76 Ϯ 11%; p ϭ 0.045). Conclusions: The benefit obtained from U74389G-supplemented cardioplegic preservation solution was marginal for hearts stored for 6 hours. After longer ischemic times, the benefit may be clearer.
Journal of Heart and Lung Transplantation, Aug 1, 2003
Background: Acute brain death from increased intracranial pressure results in a transient increas... more Background: Acute brain death from increased intracranial pressure results in a transient increase in myocardial adenosine and lactate, which indicates that oxygen demand exceeds oxygen delivery during the sympathetic "storm". The aim of this study was to determine the functional significance of this period of ischemia. Methods: Brain death was inflicted on 40 Westran pigs (36.5-68.0 kg) by inflating a 21-ml subdural balloon over 3 minutes. In 38 animals, micromanometry and sonomicrometry were used to obtain left ventricular pressure-volume loops to determine the preload recruitable stroke work (PRSW) relationship. Data files were recorded before and at 15-minute intervals after beginning balloon inflation. Plasma troponin I was measured before and 60 minutes after beginning balloon inflation in the 38 instrumented and 2 non-instrumented animals. Results: All animals experienced the classical sympathetic storm. The slope of the PRSW relationship decreased, and the volume-axis intercept shifted to the right 15 minutes after beginning balloon inflation (p Ͻ 0.0001). Progressive incremental recovery (leftward shift) occurred between subsequent time points (p Յ 0.0018). In the instrumented animals, the mean plasma troponin I level increased from 1.4 Ϯ 1.6 g/liter to 2.8 Ϯ 2.3 g/liter (p Ͻ 0.001). However, troponin I was not detected before or after induction of brain death in the plasma of either non-instrumented animal (p ϭ 0.001). Conclusions: The sympathetic storm produced transient contractile dysfunction, consistent with ischemic injury. However, troponin I release reflected surgical instrumentation and not brain death.
of the donor's wishes, a refl ection of a generous nature and way of making sense of a person's d... more of the donor's wishes, a refl ection of a generous nature and way of making sense of a person's death. Selective consent was related to emotional attachment or religious beliefs linked to specifi c organs. Of interest, there appeared to be little concern for recipients at this time but it was suggested that they might become more relevant for the donor family at a later time.
Heart, Lung and Circulation, Volume 12, Issue 2, Pages A59, 2003, Authors:Fang Lin; Andrew W. Owe... more Heart, Lung and Circulation, Volume 12, Issue 2, Pages A59, 2003, Authors:Fang Lin; Andrew W. Owens; Scott H. Kesteven; January Michalicek; Songhai Chen; Robert M. Graham; Michael P. Feneley. Journal Home. Register or Login: Password: Auto-Login [Reminder]. ...
Background. The aim of this study was to determine the efficacy of cariporide (a sodium-hydrogen ... more Background. The aim of this study was to determine the efficacy of cariporide (a sodium-hydrogen exchanger inhibitor), BMS180448 (a pharmacologic ischemic preconditioning agent), and the combination thereof, as adjuvant therapies for extended cardiac allograft preservation. Methods. A porcine model of donor brain death and orthotopic heart transplantation was used. All hearts were arrested and stored for 14 hr in an extracellular preservation solution. Control hearts (CON; n)3؍ did not receive any additional treatment. Treated hearts received BMS180448 alone (BMS; n,)3؍ cariporide alone (CAR; n,)6؍ or both BMS180448 and cariporide (B؉C; n.)6؍ Donors of BMS180448-treated hearts received 2 mg/kg, 15 min before explantation. Donors and recipients of cariporide-treated hearts received 2 mg/kg, 15 min before explantation and reperfusion, respectively. Results. The CON and BMS arms of the study were terminated after three transplantations because initial results in these groups were poor. Significantly, none of the control hearts could be weaned successfully from bypass, whereas all of the treated hearts were weaned successfully (CAR vs. CON and B؉C vs. CON: P.)210.0؍ The rate of troponin I release during the first 3 hr after reperfusion was significantly lower in CAR (P)0810.0؍ and B؉C (P)4510.0؍ recipients than in CON recipients. Mean plasma troponin I levels (g/ mL) 3 hr after reperfusion were as follows: CON 633؎177, BMS 576؎110, CAR 346؎93, and B؉C 296؎97. Conclusion. In this porcine model of extended cardiac allograft preservation, cariporide was more effec
Journal of Molecular and Cellular Cardiology, Mar 1, 2020
Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA... more Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA and Western world. Polyphenolic compounds are present in our daily diet such as vegetable, fruit, onion, and processed food such as red wine, and dark chocolate. Flavonoids, a member of polyphenols and related compounds have significant antioxidant and anti-inflammatory activity, which could vary based on their structures. Recent studies indicated that quercetin; a flavonol reduces the health risks linked with obesity, heart disease, diabetes and high blood pressure. The objective: This study is designed to investigate the effects of quercetin intake and exercise in atherosclerosis. Study Design: The study compares the effect of intake of quercetin with exercise on atherosclerotic plaque and biomarkers of atherosclerosis. We conducted three separate studies as follows: Study 1: C57BL6 wild type female mice (N¼ 35). Study 2: C57BL6 wild type male mice (N¼ 40) and Study 3: C57BL6 LDLr-/-mice (N¼ 40). Mice were fed normal mouse chow or atherogenic diet and orally supplied with 100 μg/day of quercetin with or without exercising. In each study animals were divided into four groups (10 each). These groups were as follows: Control mice, left untreated; control quercetin group, exercise group without quercetin, and exercise group with quercetin supplements. The exercise groups were run on a treadmill for 30 minutes, 15-30 m/m/ 5 days/week for 30-60 days. At the end of the 1-2 month of treatment, mice were sacrificed. Livers, aortas, adipose tissue and blood were collected. Plasma lipids, and inflammatory biomarkers, and liver and aorta gene expressions and western blot were performed. Results: In wildtype males lipids modulators such as PCSK9 mRNA levels were significantly up-regulated with combination of exercise and quercetin intake (p o 0.05). ANGPLT4 mRNA levels significantly down-regulated with the combination after 8 weeks of exercise and quercetin intake compared to both
Journal of Heart and Lung Transplantation, Feb 1, 2005
relationship (PRSW) and stroke work index (SWI) were used to assess left ventricular (LV) functio... more relationship (PRSW) and stroke work index (SWI) were used to assess left ventricular (LV) function. Median NA use was higher in the control group at 6 h (0.6 vs 0 g/kg/min; pϽ0.01). NA was weaned off by 6 h in 5/7 HR pigs compared with 0/7 controls (pϽ0.03). These 5 pigs were also weaned off vasopressin. MAP was higher in HR pigs at 6 h (75 vs 51 mmHg; pϽ0.05) and at fixed NA (71 vs 36; pϽ0.005). PRSW analysis found a greater increase in LV contractility in the HR group at 6 h compared with 0 h (pϭ0.014), equating to an increase in SWI of 85% vs 43%. LV contractility was increased in the HR group at fixed NA compared with 0 h (pϽ0.001), equating to a SWI increase of 85% vs 3%. HR in this model reduces noradrenaline requirements and improves circulatory haemodynamics and LV contractility. These results support the use of HR in the management of the brain dead organ donor.
Rationale: Gq-coupled receptors are thought to play a critical role in the induction of left vent... more Rationale: Gq-coupled receptors are thought to play a critical role in the induction of left ventricular hypertrophy (LVH) secondary to pressure overload, although mechano-sensitive channel activation by a variety of mechanisms has also been proposed, and the relative importance of calcineurin-and calmodulin kinase II (CaMKII)-dependent hypertrophic pathways remains controversial. Objective: To determine the mechanisms regulating the induction of LVH in response to mechanical pressure overload. Methods and Results: Transgenic mice with cardiac-targeted inhibition of Gq-coupled receptors (GqI mice) and their non-transgenic littermates (NTL) were subjected to neurohumoral stimulation (continuous, subcutaneous angiotensin II (AngII) infusion for 14 days) or mechanical pressure overload (transverse aortic arch constriction (TAC) for 21 days) to induce LVH. Candidate signalling pathway activation was examined. As expected, LVH observed in NTL mice with AngII infusion was attenuated in heterozygous (GqI +/-) mice and absent in homozygous (GqI-/-) mice. In contrast, LVH due to TAC was unaltered by either heterozygous or homozygous Gq inhibition. Gene expression of atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP) and α-skeletal actin (α-SA) was increased 48 hours after AngII infusion or TAC in NTL mice; in GqI mice, the increases in ANP, BNP and α-SA in response to AngII were completely absent, as expected, but all three increased after TAC. Increased nuclear translocation of nuclear factor of activated T-cells c4 (NFATc4), indicating calcineurin pathway activation, occurred in NTL mice with AngII infusion but not TAC, and was prevented in GqI mice infused with AngII. Nuclear and cytoplasmic CaMKIIδ levels increased in both NTL and GqI mice after TAC but not AngII infusion, with increased cytoplasmic phospho-and total histone deacetylase 4 (HDAC4) and increased nuclear myocyte enhancer factor 2 (MEF2) levels. Conclusion: Cardiac Gq receptors and calcineurin activation are required for neurohumorally mediated LVH but are not required for LVH induced by mechanical pressure overload (TAC); the latter is mediated by activation of the CaMKII-HDAC4-MEF2 pathway.
1. Regional differences in odorant-induced responsiveness of the rat olfactory epithelium were me... more 1. Regional differences in odorant-induced responsiveness of the rat olfactory epithelium were measured via electrophysiological recordings [negative component of electro-olfactogram (Veog(-)) made from the surface of the olfactory epithelium on the nasal septum]. The nasal septum provided a flat surface from which multiple recordings could be made. 2. Veog(-)s were recorded from a standardized grid of 16 sites. This grid of recording sites extended over most of the surface of the olfactory epithelium on the nasal septum. 3. Twenty-one animals were tested for their responses to seven odorants. The animals were divided into three groups, each of which was tested with two different odorants plus amyl acetate, which provided a comparison between the groups. 4. For each odorant in each animal, topographic maps of relative responsiveness were derived to test whether odorants elicited different patterns of responses in the same individual. Topographic maps of responsiveness were derived also for the animal groups to test for the generality of the form of the maps for different odorants. Response latencies were also measured for each odorant at each recording site. 5. All individuals showed different topographic patterns of responses to the three test odorants. For most odorants, the location of the most responsive site was similar in all animals. In different animals the topographic maps for the same odorant were remarkably similar. Topographic maps for the odorants were all different from one another. 6. These results are consistent with the hypothesis that odorant quality is encoded in the differential spatial distribution of receptor cells whose differences in responsiveness appear to be distributed as a continuum across the epithelium. The results establish for a mammalian species what was previously reported in amphibians. These differences are presumed to be due to differential expression of odorant receptor proteins. 7. The mean response latency was 32 ms. This period was similar for all odorants, all animals, and all recording sites and was independent of Veog(-) amplitude. It is concluded that diffusion through the mucus contributed approximately 6 ms to the latency of onset of the responses to these odorants.
Recombinant human platelet-derived growth factor-AB improves cardiac function and survival after ... more Recombinant human platelet-derived growth factor-AB improves cardiac function and survival after myocardial infarction in a porcine model.
Animal models of pressure overload are valuable for understanding hypertensive heart disease. We ... more Animal models of pressure overload are valuable for understanding hypertensive heart disease. We characterised a surgical model of pressure overload-induced hypertrophy in C57BL/6J mice produced by suprarenal aortic constriction (SAC). Compared to sham controls, at one week post-SAC systolic blood pressure was significantly elevated and left ventricular (LV) hypertrophy was evident by a 50% increase in the LV weight-to-tibia length ratio due to cardiomyocyte hypertrophy. As a result, LV end-diastolic wall thickness-to-chamber radius (h/R) ratio increased, consistent with the development of concentric hypertrophy. LV wall thickening was not sufficient to normalise LV wall stress, which also increased, resulting in LV systolic dysfunction with reductions in ejection fraction and fractional shortening, but no evidence of heart failure. Pathological LV remodelling was evident by the re-expression of fetal genes and coronary artery perivascular fibrosis, with ischaemia indicated by enhanced cardiomyocyte Hif1a expression. The expression of stem cell factor receptor, c-Kit, was low basally in cardiomyocytes and did not change following the development of robust hypertrophy, suggesting there is no role for cardiomyocyte c-Kit signalling in pathological LV remodelling following pressure overload. Heart failure is a common end-result of a variety of cardiovascular diseases, including ischaemic and hypertensive heart disease. Hypertension is the primary cause of pathological left ventricular (LV) hypertrophy in 75% of patients 1. While the hypertrophic response is initially compensatory to maintain heart function in the face of pressure overload (PO), pathological hypertrophy eventually becomes decompensatory, with decreased cardiac performance that precedes overt heart failure. Current treatments for heart failure merely slow the progression to end-stage heart failure, since there is no cure other than heart transplantation. Animal models of hypertensive heart disease are, thus, critical to developing potential new regenerative therapies 2. Two commonly used cardiac injury models are myocardial infarction (MI) produced by ligation of the left anterior descending coronary artery, and aortic constriction; the latter first developed and characterised by Rytand 3 and Goldblatt et al. 4 , produced by ligating the descending abdominal aorta between or immediately above the renal arteries. Constriction of the abdominal aorta induces PO leading to the rapid development of cardiac hypertrophy and heart failure 2,5. Historically, aortic constriction models were developed in larger animal species, including rabbits, dogs, guinea pigs and rats 2,5 , but over the last few decades these techniques have been adapted to mice-the preferred species for genetic studies that are also economical 6,7. Thoracic aortic constriction
B rain death causes a surge in plasma catecholamine levels, which leads to acute cardiac dysfunct... more B rain death causes a surge in plasma catecholamine levels, which leads to acute cardiac dysfunction. 1-3 It also alters the response of the myocardium to the hypothermic injury and the ischaemic reperfusion injury associated with conventional preservation and storage of the donor heart. 1-3 Minimising these combined injuries through improvements in preservation would increase the potential donor pool and improve outcome following cardiac transplantation. The objective of this brief communication is to present a large animal model of orthotopic cardiac transplantation, incorporating a brain dead donor, that we have developed to assess new strategies for cardiac allograft preservation. Final Model Methodology The experimental protocol has been approved by the St Vincent's Hospital animal experimentation ethics committee and was in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes. Inbred sibling Westran pigs (20-50 kg) were obtained in pairs. The donor pig was pre-medicated with an intramuscular injection of ketamine (10 mg/kg), midazolam (1 mg/kg) and atropine (1.2 mg). General anaesthesia was induced with intravenous thiopentone (50 mg boluses, to effect), and maintained by inhaled isoflurane (1-3% inhaled gas) and intravenous fentanyl (5 µg/kg boluses). The pig was intubated and ventilated with 100% oxygen. Normal (0.9%) saline was infused
European Journal of Cardio-Thoracic Surgery, Nov 1, 2002
Objective: Paradoxically, it has been reported that after 1.5-4 h of hypothermic ischaemic preser... more Objective: Paradoxically, it has been reported that after 1.5-4 h of hypothermic ischaemic preservation there is complete recovery of contractile function in canine cardiac allografts, as assessed by the preload recruitable stroke work (PRSW) relationship. This raises questions about the suitability of the canine heart as a model for preservation research and the PRSW relationship as an end-point. The aim of the present study was to evaluate the PRSW relationship as an index of left ventricular contractility in porcine cardiac allografts. Methods: Eighteen orthotopic heart transplants were performed in inbred Westran pigs. Brain death was induced in the donor pigs 1 h prior to explantation. The donor hearts were arrested with extracellular cardioplegia, which was stored in ice prior to administration. On explantation, the donor hearts were immersed in cardioplegia and stored in ice. The donor hearts were subjected to either 4 (IT4, n ¼ 6), 6 (IT6, n ¼ 9) or 14 (IT14, n ¼ 3) h of ischaemia. Post-transplant, all hearts were supported with dobutamine (10 mcg/kg per min). The PRSW relationship was derived from pressure-volume loops obtained by epicardial sonomicrometry and transmyocardial micromanometry. Multiple linear regression was used to describe and compare the PRSW relationship before brain death in the donor and after weaning from bypass in the recipient. Results: Eleven hearts were weaned successfully from cardiopulmonary bypass: IT4 100% (6/6), IT6 56% (5/9) and IT14 0% (0/3) (IT4 versus IT14: P ¼ 0:012). Analysis of the PRSW relationship revealed a reduction in contractility in both the IT4 and IT6 groups (both P , 0:0001), but a greater reduction in the IT6 group (P , 0:0001). Notably, the volume-axis intercept of the PRSW relationship was found to be a better discriminator of post-preservation contractile dysfunction than the slope of the PRSW relationship. Conclusions: The porcine heart's susceptibility to ischaemic injury makes it ideal for evaluating the effect of different preservation strategies on contractile recovery. The PRSW relationship can be used to evaluate the differences in contractile recovery, though the nature of the effect of ischaemic preservation necessitates analysis by multiple linear regression.
Journal of Molecular and Cellular Cardiology, Jun 1, 2021
The 'fight or flight' response to physiological stress involves sympathetic nervous syste... more The 'fight or flight' response to physiological stress involves sympathetic nervous system activation, catecholamine release and adrenergic receptor stimulation. In the heart, this induces positive inotropy, previously attributed to the β1-adrenergic receptor subtype. However, the role of the α1A-adrenergic receptor, which has been suggested to be protective in cardiac pathology, has not been investigated in the setting of physiological stress. To explore this, we developed a tamoxifen-inducible, cardiomyocyte-specific α1A-adrenergic receptor knock-down mouse model, challenged mice to four weeks of endurance swim training and assessed cardiac outcomes. With 4-OH tamoxifen treatment, expression of the α1A-adrenergic receptor was knocked down by 80-89%, without any compensatory changes in the expression of other adrenergic receptors, or changes to baseline cardiac structure and function. Swim training caused eccentric hypertrophy, regardless of genotype, demonstrated by an increase in heart weight/tibia length ratio (30% and 22% in vehicle- and tamoxifen-treated animals, respectively) and an increase in left ventricular end diastolic volume (30% and 24% in vehicle- and tamoxifen-treated animals, respectively) without any change in the wall thickness/chamber radius ratio. Consistent with physiological hypertrophy, there was no increase in fetal gene program (Myh7, Nppa, Nppb or Acta1) expression. In response to exercise-induced volume overload, stroke volume (39% and 30% in vehicle- and tamoxifen-treated animals, respectively), cardiac output/tibia length ratio (41% in vehicle-treated animals) and stroke work (61% and 33% in vehicle- and tamoxifen-treated animals, respectively) increased, regardless of genotype. These findings demonstrate that cardiomyocyte α1A-adrenergic receptors are not necessary for cardiac adaptation to endurance exercise stress and their acute ablation is not deleterious.
Pathological left ventricular hypertrophy (LVH) occurs in response to pressure overload and remai... more Pathological left ventricular hypertrophy (LVH) occurs in response to pressure overload and remains the single most important clinical predictor of cardiac mortality. The molecular pathways in the induction of pressure overload LVH are potential targets for therapeutic intervention. Current treatments aim to remove the pressure overload stimulus for LVH, but do not completely reverse adverse cardiac remodelling. Although numerous molecular signalling steps in the induction of LVH have been identified, the initial step by which mechanical stretch associated with cardiac pressure overload is converted into a chemical signal that initiates hypertrophic signalling remains unresolved. In this study, we show that selective deletion of transient receptor potential melastatin 4 (TRPM4) channels in mouse cardiomyocytes results in an approximately 50% reduction in the LVH induced by transverse aortic constriction. Our results suggest that TRPM4 channel is an important component of the mechanosensory signalling pathway that induces LVH in response to pressure overload and represents a potential novel therapeutic target for the prevention of pathological LVH.
Mutations in the giant sarcomeric protein titin (TTN) are a major cause for inherited forms of di... more Mutations in the giant sarcomeric protein titin (TTN) are a major cause for inherited forms of dilated cardiomyopathy (DCM). We have previously developed a mouse model that imitates a TTN truncation mutation we found in a large pedigree with DCM. While heterozygous Ttn knock-in mice do not display signs of heart failure under sedentary conditions, they recapitulate the human phenotype when exposed to the pharmacological stressor angiotensin II or isoproterenol. In this study we investigated the effects of pressure overload by transverse aortic constriction (TAC) in heterozygous (Het) Ttn knock-in mice. Two weeks after TAC, Het mice developed marked impairment of left ventricular ejection fraction (< 0.05), while wild-type (WT) TAC mice did not. Het mice also trended toward increased ventricular end diastolic pressure and volume compared to WT littermates. We found an increase in histologically diffuse cardiac fibrosis in Het compared to WT in TAC mice. This study shows that a pattern of DCM can be induced by TAC-mediated pressure overload in a TTN-truncated mouse model. This model enlarges our arsenal of cardiac disease models, adding a valuable tool to understand cardiac pathophysiological remodeling processes and to develop therapeutic approaches to combat heart failure.
Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA... more Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA and Western world. Polyphenolic compounds are present in our daily diet such as vegetable, fruit, onion, and processed food such as red wine, and dark chocolate. Flavonoids, a member of polyphenols and related compounds have significant antioxidant and anti-inflammatory activity, which could vary based on their structures. Recent studies indicated that quercetin; a flavonol reduces the health risks linked with obesity, heart disease, diabetes and high blood pressure. The objective: This study is designed to investigate the effects of quercetin intake and exercise in atherosclerosis. Study Design: The study compares the effect of intake of quercetin with exercise on atherosclerotic plaque and biomarkers of atherosclerosis. We conducted three separate studies as follows: Study 1: C57BL6 wild type female mice (N¼ 35). Study 2: C57BL6 wild type male mice (N¼ 40) and Study 3: C57BL6 LDLr-/-mice (N¼ 40). Mice were fed normal mouse chow or atherogenic diet and orally supplied with 100 μg/day of quercetin with or without exercising. In each study animals were divided into four groups (10 each). These groups were as follows: Control mice, left untreated; control quercetin group, exercise group without quercetin, and exercise group with quercetin supplements. The exercise groups were run on a treadmill for 30 minutes, 15-30 m/m/ 5 days/week for 30-60 days. At the end of the 1-2 month of treatment, mice were sacrificed. Livers, aortas, adipose tissue and blood were collected. Plasma lipids, and inflammatory biomarkers, and liver and aorta gene expressions and western blot were performed. Results: In wildtype males lipids modulators such as PCSK9 mRNA levels were significantly up-regulated with combination of exercise and quercetin intake (p o 0.05). ANGPLT4 mRNA levels significantly down-regulated with the combination after 8 weeks of exercise and quercetin intake compared to both
Journal of Heart and Lung Transplantation, Mar 1, 2003
Background: U74389G (16-desmethyl tirilazad), a 21-aminosteroid or "lazaroid," inhibits lipid per... more Background: U74389G (16-desmethyl tirilazad), a 21-aminosteroid or "lazaroid," inhibits lipid peroxidation, which is an important element of ischemia-reperfusion injury. The aim of this study was to determine whether the addition of U74389G to the cardioplegic preservation solution could improve early cardiac allograft function. Methods: A porcine model of donor brain death and orthotopic cardiac transplantation was used. Hearts were arrested and preserved for 6 hours in an aspartate-enriched extracellular cardioplegia that had been supplemented with either U74389G and its carrier (n ϭ 7) or the carrier alone (n ϭ 9). Epicardial sonomicrometry and transmyocardial micromanometry were used to obtain pressure-volume loops before and after transplantation. Left ventricular wall volume was measured by volume displacement. Results: A higher proportion of U74389G-treated hearts were weaned successfully from cardiopulmonary bypass, but this difference did not achieve statistical significance (86% [6 of 7] vs 56% [5 of 9]; p ϭ 0.308). In the hearts that were weaned successfully, preservation of left ventricular contractility, as judged by the pre-load recruitable stroke work relationship, was significantly better in the U74389G-treated hearts (p ϭ 0.0271). In contrast, left ventricular compliance, as judged by the end-diastolic pressure-volume relationship, was significantly better preserved in the control group (p Ͻ 0.0001). U74389G-treated hearts developed less myocardial edema, as judged by the posttransplant left ventricular wall volume/baseline steady-state epicardial end-diastolic volume ratio (64 Ϯ 9% vs 76 Ϯ 11%; p ϭ 0.045). Conclusions: The benefit obtained from U74389G-supplemented cardioplegic preservation solution was marginal for hearts stored for 6 hours. After longer ischemic times, the benefit may be clearer.
Journal of Heart and Lung Transplantation, Aug 1, 2003
Background: Acute brain death from increased intracranial pressure results in a transient increas... more Background: Acute brain death from increased intracranial pressure results in a transient increase in myocardial adenosine and lactate, which indicates that oxygen demand exceeds oxygen delivery during the sympathetic "storm". The aim of this study was to determine the functional significance of this period of ischemia. Methods: Brain death was inflicted on 40 Westran pigs (36.5-68.0 kg) by inflating a 21-ml subdural balloon over 3 minutes. In 38 animals, micromanometry and sonomicrometry were used to obtain left ventricular pressure-volume loops to determine the preload recruitable stroke work (PRSW) relationship. Data files were recorded before and at 15-minute intervals after beginning balloon inflation. Plasma troponin I was measured before and 60 minutes after beginning balloon inflation in the 38 instrumented and 2 non-instrumented animals. Results: All animals experienced the classical sympathetic storm. The slope of the PRSW relationship decreased, and the volume-axis intercept shifted to the right 15 minutes after beginning balloon inflation (p Ͻ 0.0001). Progressive incremental recovery (leftward shift) occurred between subsequent time points (p Յ 0.0018). In the instrumented animals, the mean plasma troponin I level increased from 1.4 Ϯ 1.6 g/liter to 2.8 Ϯ 2.3 g/liter (p Ͻ 0.001). However, troponin I was not detected before or after induction of brain death in the plasma of either non-instrumented animal (p ϭ 0.001). Conclusions: The sympathetic storm produced transient contractile dysfunction, consistent with ischemic injury. However, troponin I release reflected surgical instrumentation and not brain death.
of the donor's wishes, a refl ection of a generous nature and way of making sense of a person's d... more of the donor's wishes, a refl ection of a generous nature and way of making sense of a person's death. Selective consent was related to emotional attachment or religious beliefs linked to specifi c organs. Of interest, there appeared to be little concern for recipients at this time but it was suggested that they might become more relevant for the donor family at a later time.
Heart, Lung and Circulation, Volume 12, Issue 2, Pages A59, 2003, Authors:Fang Lin; Andrew W. Owe... more Heart, Lung and Circulation, Volume 12, Issue 2, Pages A59, 2003, Authors:Fang Lin; Andrew W. Owens; Scott H. Kesteven; January Michalicek; Songhai Chen; Robert M. Graham; Michael P. Feneley. Journal Home. Register or Login: Password: Auto-Login [Reminder]. ...
Background. The aim of this study was to determine the efficacy of cariporide (a sodium-hydrogen ... more Background. The aim of this study was to determine the efficacy of cariporide (a sodium-hydrogen exchanger inhibitor), BMS180448 (a pharmacologic ischemic preconditioning agent), and the combination thereof, as adjuvant therapies for extended cardiac allograft preservation. Methods. A porcine model of donor brain death and orthotopic heart transplantation was used. All hearts were arrested and stored for 14 hr in an extracellular preservation solution. Control hearts (CON; n)3؍ did not receive any additional treatment. Treated hearts received BMS180448 alone (BMS; n,)3؍ cariporide alone (CAR; n,)6؍ or both BMS180448 and cariporide (B؉C; n.)6؍ Donors of BMS180448-treated hearts received 2 mg/kg, 15 min before explantation. Donors and recipients of cariporide-treated hearts received 2 mg/kg, 15 min before explantation and reperfusion, respectively. Results. The CON and BMS arms of the study were terminated after three transplantations because initial results in these groups were poor. Significantly, none of the control hearts could be weaned successfully from bypass, whereas all of the treated hearts were weaned successfully (CAR vs. CON and B؉C vs. CON: P.)210.0؍ The rate of troponin I release during the first 3 hr after reperfusion was significantly lower in CAR (P)0810.0؍ and B؉C (P)4510.0؍ recipients than in CON recipients. Mean plasma troponin I levels (g/ mL) 3 hr after reperfusion were as follows: CON 633؎177, BMS 576؎110, CAR 346؎93, and B؉C 296؎97. Conclusion. In this porcine model of extended cardiac allograft preservation, cariporide was more effec
Journal of Molecular and Cellular Cardiology, Mar 1, 2020
Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA... more Introduction: Cardiovascular disease (CVD) is the leading cause of mortality and morbidity in USA and Western world. Polyphenolic compounds are present in our daily diet such as vegetable, fruit, onion, and processed food such as red wine, and dark chocolate. Flavonoids, a member of polyphenols and related compounds have significant antioxidant and anti-inflammatory activity, which could vary based on their structures. Recent studies indicated that quercetin; a flavonol reduces the health risks linked with obesity, heart disease, diabetes and high blood pressure. The objective: This study is designed to investigate the effects of quercetin intake and exercise in atherosclerosis. Study Design: The study compares the effect of intake of quercetin with exercise on atherosclerotic plaque and biomarkers of atherosclerosis. We conducted three separate studies as follows: Study 1: C57BL6 wild type female mice (N¼ 35). Study 2: C57BL6 wild type male mice (N¼ 40) and Study 3: C57BL6 LDLr-/-mice (N¼ 40). Mice were fed normal mouse chow or atherogenic diet and orally supplied with 100 μg/day of quercetin with or without exercising. In each study animals were divided into four groups (10 each). These groups were as follows: Control mice, left untreated; control quercetin group, exercise group without quercetin, and exercise group with quercetin supplements. The exercise groups were run on a treadmill for 30 minutes, 15-30 m/m/ 5 days/week for 30-60 days. At the end of the 1-2 month of treatment, mice were sacrificed. Livers, aortas, adipose tissue and blood were collected. Plasma lipids, and inflammatory biomarkers, and liver and aorta gene expressions and western blot were performed. Results: In wildtype males lipids modulators such as PCSK9 mRNA levels were significantly up-regulated with combination of exercise and quercetin intake (p o 0.05). ANGPLT4 mRNA levels significantly down-regulated with the combination after 8 weeks of exercise and quercetin intake compared to both
Journal of Heart and Lung Transplantation, Feb 1, 2005
relationship (PRSW) and stroke work index (SWI) were used to assess left ventricular (LV) functio... more relationship (PRSW) and stroke work index (SWI) were used to assess left ventricular (LV) function. Median NA use was higher in the control group at 6 h (0.6 vs 0 g/kg/min; pϽ0.01). NA was weaned off by 6 h in 5/7 HR pigs compared with 0/7 controls (pϽ0.03). These 5 pigs were also weaned off vasopressin. MAP was higher in HR pigs at 6 h (75 vs 51 mmHg; pϽ0.05) and at fixed NA (71 vs 36; pϽ0.005). PRSW analysis found a greater increase in LV contractility in the HR group at 6 h compared with 0 h (pϭ0.014), equating to an increase in SWI of 85% vs 43%. LV contractility was increased in the HR group at fixed NA compared with 0 h (pϽ0.001), equating to a SWI increase of 85% vs 3%. HR in this model reduces noradrenaline requirements and improves circulatory haemodynamics and LV contractility. These results support the use of HR in the management of the brain dead organ donor.
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