Robert M. Lust

Carnitine insufficiency caused by aging and overnutrition compromises mitochondrial performance and metabolic control.

In addition to its essential role in permitting mitochondrial import and oxidation of long chain fatty acids, carnitine also functions as an acyl group acceptor that facilitates mitochondrial export of excess carbons in the form of acylcarnitines. Recent evidence suggests carnitine requirements increase under conditions of sustained metabolic stress. Accordingly, we hypothesized that carnitine insufficiency might contribute to mitochondrial dysfunction and obesity-related impairments in glucose tolerance. Consistent with this prediction whole body carnitine dimunition was identified as a common feature of insulin-resistant states such as advanced age, genetic diabetes, and diet-induced obesity. In rodents fed a lifelong (12 month) high fat diet, compromised carnitine status corresponded with increased skeletal muscle accumulation of acylcarnitine esters and diminished hepatic expression of carnitine biosynthetic genes. Diminished carnitine reserves in muscle of obese rats was accompanied by marked perturbations in mitochondrial fuel metabolism, including low rates of complete fatty acid oxidation, elevated incomplete β-oxidation, and impaired substrate switching from fatty acid to pyruvate. These mitochondrial abnormalities were reversed by 8 weeks of oral carnitine supplementation, in concert with increased tissue efflux and urinary excretion of acetylcarnitine and improvement of whole body glucose tolerance. Acetylcarnitine is produced by the mitochondrial matrix enzyme, carnitine acetyltransferase (CrAT). A role for this enzyme in combating glucose intolerance was further supported by the finding that CrAT overexpression in primary human skeletal myocytes increased glucose uptake and attenuated lipid-induced suppression of glucose oxidation. These results implicate carnitine insufficiency and reduced CrAT activity as reversible components of the metabolic syndrome.

Video-assisted minimally invasive mitral valve surgery

OBJECTIVE This study was done to determine the potential benefits of minimally invasive mitral surgery performed with intraoperative video assistance. METHODS From May 1996 until March 1997, a minithoracotomy and video assistance were used in 31 consecutive patients undergoing mitral repair (n = 20) and replacement (n = 11). Their ages ranged from 18 to 77 years (59 +/- 2.6 years; mean +/- standard error of the mean). Ejection fractions were 35% to 62% (55% +/- 1.5%). Operations were done with either antegrade/retrograde (n = 10) or antegrade (n = 19) cold blood cardioplegia and a new transthoracic crossclamp or with ventricular fibrillation (n = 2). Peripheral arterial cannulation (n = 28) and pump-assisted right atrial drainage (n = 26) were used most often. RESULTS No hospital deaths occurred, but the 30-day mortality was 3.2%. Complications included deep venous thrombosis and a phrenic nerve palsy in one patient each. No patient had a stroke or required reoperation for bleeding. Postoperative echocardiography showed excellent valve function in all but one patient. Cardiopulmonary bypass and arrest times averaged 183 +/- 7.2 and 136 +/- 5.5 minutes, respectively. Compared with 100 patients having conventional mitral valve operations, these patients had significantly shorter hospitalization times (8.6 +/- 0.5 vs 5.1 +/- 0.9 days, p = 0.05). Moreover, 81% of the later cohort were discharged between day 3 and 5 (3.6 +/- 0.2 days). Hospital charges (decreases 27%, p = 0.05) and costs (decreases 34%, p < 0.05) were less than in conventional operations. Patient follow-up suggested minimal perioperative pain and rapid recovery. CONCLUSIONS Early results suggest that video-assisted minimally invasive mitral operations can be done safely. These methods may benefit patients through less morbidity, earlier discharge, and lower cost.

Mitochondrial permeability transition in the diabetic heart: Contributions of thiol redox state and mitochondrial calcium to augmented reperfusion injury

Mitochondria from diabetic hearts are sensitized to mitochondrial permeability transition pore (PTP) opening, which may be responsible for the increased propensity for cardiac injury in diabetic hearts. The purpose of this study was to determine if redox-dependent PTP opening contributes to augmented injury in diabetic hearts, and if compounds targeted at mitochondrial PTP, ROS, and calcium influx protected diabetic hearts from injury. Hearts from control or streptozotocin-induced diabetic rats were excised for either whole-heart or isolated mitochondria experiments. Myocardial glutathione content was oxidized in diabetic hearts when compared to control, and this translated to increased oxidation of the adenine nucleotide translocase in diabetic hearts. Diabetic mitochondria displayed significantly greater sensitivity to PTP opening than non-diabetic counterparts, which was reversed with the thiol-reducing agent dithiothreitol. The thiol-oxidant diamide increased calcium sensitivity in control, but not diabetic mitochondria. Diabetic animals treated with the mitochondria-targeted ROS suppressing peptide MTP-131 also showed improved resistance to PTP opening. In separate experiments hearts underwent ex vivo ischemia/reperfusion (IR). Diabetic hearts were more susceptible to IR injury, with infarct sizes of 60 ± 4% of the area-at-risk (vs. 46 ± 2% in non-diabetics; P<0.05). Administration of the PTP blocker NIM811 (5 μM), MTP-131 (1 nM) or the mitochondrial calcium uniporter blocker minocycline (1 μM) at the onset of reperfusion reduced infarct sizes in both control and diabetic hearts. These findings suggest that augmented susceptibility to injury in the diabetic heart is mediated by redox-dependent shifts in PTP opening, and that three novel mitochondria-targeted agents administered at reperfusion may be suitable adjuvant reperfusion therapies to attenuate injury in diabetic patients.

Transformation of nonvascular acellular tissue matrices into durable vascular conduits.

BACKGROUND Prosthetic grafts commonly used for vascular reconstruction are limited to synthetics and cross-linked tissue grafts. Within these devices, graft infections are common, compliance mismatch is significant, and handling qualities are poor. Natural biological tissues that are unfixed have been shown to resist infections and be durable and compliant. A natural biological matrix that could be remodeled appropriately after implantation would be a desirable graft for vascular reconstruction. METHODS SynerGraft tissue engineering strategies have been used to minimize antigenicity and produce stable unfixed vascular grafts from nonvascular bovine tissues. These grafts have replaced the abdominal aortas of 8 dogs that have been followed for up to 10 months. RESULTS Early evaluation indicates rapid recellularization by recipient smooth muscle actin positive cells, which become arranged circumferentially, into the media. Arterioles were present in the adventitial areas and endothelial cells were seen to cover lumenal surfaces. After 10 months, grafts were patent and not aneurysmal. CONCLUSIONS These data indicate that SynerGraft processing of animal tissues is capable of producing stable vascular conduits that exhibit long-term functionality in other species.

Mast cells contribute to altered vascular reactivity and ischemia-reperfusion injury following cerium oxide nanoparticle instillation

Abstract Cerium oxide (CeO2) represents an important nanomaterial with wide ranging applications. However, little is known regarding how CeO2 exposure may influence pulmonary or systemic inflammation. Furthermore, how mast cells would influence inflammatory responses to a nanoparticle exposure is unknown. We thus compared pulmonary and cardiovascular responses between C57BL/6 and B6.Cg-KitW-sh mast cell deficient mice following CeO2 nanoparticle instillation. C57BL/6 mice instilled with CeO2 exhibited mild pulmonary inflammation. However, B6.Cg-KitW-sh mice did not display a similar degree of inflammation following CeO2 instillation. Moreover, C57BL/6 mice instilled with CeO2 exhibited altered aortic vascular responses to adenosine and an increase in myocardial ischemia/reperfusion injury which was absent in B6.Cg-KitW-sh mice. In vitro CeO2 exposure resulted in increased production of PGD2, TNF-α, IL-6 and osteopontin by cultured mast cells. These findings demonstrate that CeO2 nanoparticles activate mast cells contributing to pulmonary inflammation, impairment of vascular relaxation and exacerbation of myocardial ischemia/reperfusion injury.

Effect of chronic native flow competition on internal thoracic artery grafts

Residual competitive flow from the native coronary artery has been proposed as a mechanism that reduces flow in an internal thoracic artery graft (ITA), resulting in narrowing and ultimately failure of the graft. Results from acute experiments have indicated that competitive flow from a fully patent native artery did not abolish ITA graft flow. The present study was designed to examine the consequences of dynamic flow competition between the native vessel and the ITA graft in a chronic model. Fifteen mongrel dogs underwent coronary artery bypass grafting using the pedicled left ITA anastomosed to the normal, fully patent circumflex (CFX) coronary artery. The procedure was performed through a sterile thoracotomy, without systemic cardiopulmonary bypass, using a brief local occlusion to construct the anastomosis. Intraoperatively, ITA flow was measured in situ on the chest wall, before the pedicle was mobilized. Internal thoracic artery graft and distal CFX flow were measured after the anastomosis was completed, with and without brief occlusion of the proximal CFX. Angiography was performed 72 hours, 4 weeks, and 8 weeks later; graft patency and diameter were evaluated. After 8 weeks, open-chest direct flow measurements comparable with the intraoperative assessment were obtained. Two grafts (13%) occluded early, the technical result of poor anastomotic construction. In the 13 remaining animals, all grafts were widely patent at all time points. Internal thoracic artery flow in situ averaged 10.9 +/- 7.8 mL/min (mean +/- standard deviation), and was maintained after grafting (11.5 +/- 4.4 mL/min; p = not significant).(ABSTRACT TRUNCATED AT 250 WORDS)

Competitive flow from a fully patent coronary artery does not limit acute mammary graft flow

The shriveled, stenotic mammary graft sometimes observed after internal mammary artery (IMA) to coronary artery bypass grafting has been attributed to competitive flow from the insufficiently stenosed native coronary vessel. To study further the effects of native coronary artery competing flow on IMA graft flow, 10 dogs (mean weight, 23.5 +/- 3.69 kg) underwent coronary artery bypass grafting using the pedicled left IMA anastomosed to a normal, fully patent proximal circumflex (CFX) coronary artery. The procedure was performed through a left thoracotomy, off pump, using a brief local occlusion to perform the anastomosis. Native in situ IMA flow, CFX flow distal to the anastomosis, and IMA graft flow were measured using calibrated electromagnetic flow probes. When the CFX proximal to the anastomosis was occluded transiently, IMA flow increased to supply 100% of the previously measured distal CFX flow (60.2 +/- 7.9 mL/min). When both the IMA graft and CFX proximal to the anastomosis were patent, total distal perfusion was maintained (58.9 +/- 7.8 mL/min) and relative IMA graft flow (26.5 +/- 3.3 mL/min) was proportional to the relative diameter of the IMA graft to the native coronary artery (r = 0.96). The mean flow in the IMA in situ on the chest wall before its division was 23.8 +/- 8.1 mL/min. These results suggest that, at least acutely in a canine model, IMA graft flow is maintained above in situ levels even when grafted to a completely patent coronary artery and that acute competitive flow probably does not cause mammary artery shriveling.

Transfemoral balloon aortic occlusion during open cardiopulmonary resuscitation improves myocardial and cerebral blood flow

These experiments were designed to determine whether the limited cardiac output during open cardiac massage could be preferentially directed to the coronary and cerebral vessels by balloon occlusion of the descending thoracic aorta. Sixteen dogs were instrumented to monitor cardiac output and left atrial, right atrial, right ventricular, left ventricular, and arterial blood pressures. Measurements of myocardial and cerebral blood flow distribution during massage were made using the radioactive microsphere technique. Each animal underwent two episodes of fibrillation and resuscitation. In one episode the arrest was managed by open massage alone, and in the other, open massage was accompanied by balloon occlusion, with the order randomized. When compared to control, open cardiac massage was associated with a significant decrease in mean arterial pressure; however, the addition of balloon occlusion produced a 130% increase in the mean arterial pressure that was obtained during open CPR (control, 93 +/- 5 mm Hg; massage alone, 35 +/- 2 mm Hg; massage + balloon, 76 +/- 2 mm Hg, P less than 0.01). In a similar fashion, although the absolute blood flow was reduced by 50% when compared to control, the blood flow (ml/min/g) to the brain and heart during massage was 100% better when balloon occlusion was employed (brain: control, 0.41 +/- 0.03; massage only, 0.05 +/- 0.01; massage + balloon, 0.25 +/- 0.02, P less than 0.01; heart: control, 1.46 +/- 0.11; massage alone, 0.35 +/- 0.05; massage + balloon, 0.71 +/- 0.05, P less than 0.01). These results suggest that aortic occlusion significantly increased myocardial and cerebral perfusion patterns during ventricular fibrillation and open cardiac massage.(ABSTRACT TRUNCATED AT 250 WORDS)

Cardiac and Vasular Changes in Mice After Exposure to Ultrafine Particulate Matter

Increased ambient air particulate matter (PM) concentrations are associated with risk for myocardial infarction, stroke, and arrhythmia, and ultrafine PM (UFPM) might be particularly toxic to the cardiovascular system. Recent epidemiological studies are beginning to offer mechanistic insights, yet the rodent model remains a valuable tool to explore potential mechanisms. This article reviews a series of studies from our laboratory demonstrating the promise of mouse models to link health effects to biological mechanisms. Specifically, data from 6- to 10-wk-old male ICR mice exposed to intratracheal instillation of 100 μ g of UFPM collected from the Chapel Hill, NC airshed are described. Studies of ischemia/reperfusion, vascular function, and hemostasis are described. In summary, UFPM exposure doubles the size of myocardial infarction attendant to an episode of ischemia and reperfusion while increasing postischemic oxidant stress. UFPM alters endothelial-dependent and -independent regulation of systemic vascular tone; increases platelet number, plasma fibrinogen, and soluble P-selectin levels; and reduces bleeding time, implying enhanced thrombogenic potential. Taking these findings together, this model of acute UFPM exposure in the mouse indicates that UFPM induces a prothrombotic state and decreases vasomotor responsiveness, thereby offering insight into how UFPM could contribute to vascular events associated with thrombosis and ischemia and increasing the extent of infarction.

Attenuation of myocardial injury in mice with functional deletion of the circadian rhythm gene mPer2

Variations in circadian rhythms are evident in the incidence of cardiovascular disease, and the risk of cardiovascular events increases when rhythms are disrupted. The suprachiasmatic nucleus is the central circadian pacemaker that regulates the daily rhythm of peripheral organs. Diurnal rhythms have more recently been shown to exist in myocardial tissue and are involved in metabolism and contractile function. Thus we sought to determine whether the functional deletion of the circadian rhythm mouse periodic gene 2 (mPer2) would protect the heart against ischemic injury. Nonreperfused myocardial infarction was induced in anesthetized, ventilated C57 (n = 17) and mPer2 mutant (mPer2-M; n = 15) mice via permanent ligation of the left anterior descending coronary artery. At 4 days post-myocardial infarction, we observed a 43% reduction of infarct area in mPer2-M mice compared with wild-type mice. This is coincident with 25% less macrophage infiltration, 43% higher capillary density, 17% increase in hypertrophy, and 15% less cardiomyocyte apoptosis in the infarct zone. Also, matrix metalloproteinase-9 was expressed in inflammatory cells in both groups, but total protein was 40% higher in wild-type mice, whereas it was not elevated in mPer2-M mice in response to injury. The functional deletion of the mPer2 gene reduces the severity of myocardial infarct injury by limiting the inflammatory response, reducing apoptosis, and inducing cardiomyocyte hypertrophy, thus preserving cardiac function. These findings collectively imply that the disruption of the circadian clock gene mPer2 is protective. Understanding the interactions between circadian rhythm genes and cardiovascular disease may provide insights into potential preventative and therapeutic strategies for susceptible populations.