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Biomedical subjects

F Rosenfeldt

Publications and source records attributed to F Rosenfeldt.

17 recordsLinked to original sources
PubMed2001

Quantification of mitochondrial DNA in peripheral blood mononuclear cells and subcutaneous fat using real-time polymerase chain reaction.

BACKGROUND: With decreased rates of HIV mortality and disease progression attributable to treatment with nucleoside analogue reverse transcriptase inhibitors (NRTIs), attention has now become focused on the toxicities of these forms of treatment. It is believed NRTIs cause a decrease in mitochondrial DNA (mtDNA) synthesis due to their inhibition of DNA polymerase gamma. This hypothesis is supported by in vitro data from muscle biopsies and human lymphoblastic cell lines. The resulting mitochondrial toxicity is thought to manifest itself in a variety of clinical symptoms including fatigue, fat wasting and peripheral neuropathy. A non-invasive test of mitochondrial toxicity is needed to assess toxicity and optimise HIV treatment strategies. Peripheral blood mononuclear cells (PBMC) and subcutaneous fat could be ideal and accessible sources of mtDNA for examining toxicity. OBJECTIVES: The objectives of this study were (a) to develop an assay to quantify the mtDNA copy number of PBMC and obtain reproducible results and (b) to establish the utility of subcutaneous fat as a source of mtDNA for quantification. STUDY DESIGN: PBMC were isolated from blood by centrifugation over Ficoll-Paque and subcutaneous fat was obtained from two 3 mm punch skin biopsies. Following DNA extraction, the mtDNA copy number in each sample was quantified by real-time polymerase chain reaction (PCR). RESULTS: The real-time PCR assay was found to generate consistent and reproducible results with replicates of samples undertaken within the same run, and in two or more different runs, having a mean coefficient of variation of 11.3 and 17.2%, respectively. PBMC and subcutaneous fat contained 409+/-148 and 2042+/-391 copies of mtDNA per cell, respectively. CONCLUSIONS: From the work carried out it can be concluded that firstly, the real-time PCR assay generates consistent and reproducible results, and secondly that mtDNA can be extracted and quantified from PBMC and subcutaneous fat.

Adipose Tissue↗
PubMed2000

Cardioplegic strategies for calcium control: low Ca(2+), high Mg(2+), citrate, or Na(+)/H(+) exchange inhibitor HOE-642.

BACKGROUND: Ca(2+) overload plays an important role in the pathogenesis of cardioplegic ischemia-reperfusion injury. The standard technique to control Ca(2+) overload has been to reduce Ca(2+) in the cardioplegic solution (CP). Recent reports suggest that Na(+)/H(+) exchange inhibitors can also prevent Ca(2+) overload. We compared 4 crystalloid CPs that might minimize Ca(2+) overload in comparison with standard Mg(2+)-containing CP: (1) low Ca(2+) CP (0.25 mmol/L), (2) citrate CP/normal Mg(2+) (1 mmol/L Mg(2+)), (3) citrate CP/high Mg(2+) (9 mmol/L Mg(2+)), and (4) the addition of the Na(+)/H(+) exchange inhibitor HOE-642 (Cariporide). We also tested the effect of citrate titration in vitro on the level of free Ca(2+) and Mg(2+) in CPs. METHODS AND RESULTS: Isolated working rat heart preparations were perfused with oxygenated Krebs-Henseleit buffer and subjected to 60 minutes of 37 degrees C arrest and reperfusion with CPs with different Ca(2+) concentrations. Cardiac performance, including aortic flow (AF), was measured before and after ischemia. Myocardial high-energy phosphates were measured after reperfusion. The in vitro addition of citrate to CP (2%, 21 mmol/L) produced parallel reductions in Mg(2+) and Ca(2+). Because only Ca(2+) was required to be low, the further addition of Mg(2+) increased free Mg(2+), but the highest level achieved was 9 mmol/L. Citrate CP significantly impaired postischemic function (AF 58.3+/-2. 5% without citrate versus 41.6+/-3% for citrate with normal Mg(2+), P:<0.05, versus 22.4+/-6.2% for citrate with high Mg(2+), P:<0.05). Low-Ca(2+) CP (0.25 mmol/L Ca(2+)) significantly improved the recovery of postischemic function in comparison with standard CP (1.0 mmol/L Ca(2+)) (AF 47.6+/-1.7% versus 58.3+/-2.5%, P:<0.05). The addition of HOE-642 (1 micromol/L) to CP significantly improved postischemia function (47.6+/-1.7% without HOE-642 versus 62.4+/-1. 7% with HOE-642, P:<0.05). Postischemia cardiac high-energy phosphate levels were unaffected by Ca(2+) manipulation. CONCLUSIONS: (1) A lowered Ca(2+) concentration in CP is beneficial in Mg(2+)-containing cardioplegia. (2) The use of citrate to chelate Ca(2+) is detrimental in the crystalloid-perfused isolated working rat heart, especially with high Mg(2+). (3) The mechanism of citrate action is complex, and its use limits precise simultaneous control of Ca(2+) and Mg(2+). (4) HOE-642 in CP is as efficacious in preservation of the ischemic myocardium as is the direct reduction in Ca(2+).

Adenosine Diphosphate↗
PubMed2000

Tolerance to ischemia and hypoxia is reduced in aged human myocardium.

BACKGROUND: Recovery of cardiac function after cardiac surgery and other interventional cardiac procedures in elderly patients is inferior to that in younger patients, suggesting that the aged myocardium is more sensitive to ischemia and other stresses. Although convincing data from animal studies of senescence now exist, there is a dearth of controlled in vitro studies that examine the specific response of aged human myocardium to the stress of hypoxia or ischemia. OBJECTIVE: We sought to determine the effect of age on the capacity of human atrial trabeculae to recover contractile function after in vitro hypoxic or ischemic stress. METHODS: Atrial pectinate trabeculae were dissected from the tip of 58 right atrial appendages harvested during an operation in patients aged between 34 and 89 years and electrically stimulated at 1 Hz in oxygenated Ringer's solution at 37 degrees C. Tissues experienced 30 minutes of either hypoxia (N(2) and perfusate) or simulated ischemia (humidified N(2) without perfusate) and were returned to normoxia for recovery of function for 30 minutes. Developed force and other contractile variables were determined during each period. RESULTS: Under normoxic conditions, no significant age difference was observed for any contractile function variable. However, after hypoxia, the old (70-89 years) and intermediate age groups (60-69 years) showed reduced recovery of developed force (48.5% +/- 22.2% [n = 11] and 44.9% +/- 19% [n = 12], respectively) compared with that found (66.4% +/- 19.7% [n = 15]) in the younger (34-59 years) group (mean +/- SD, P =.02). Similarly, after simulated ischemia, the groups of 70- to 89-year-old and 60- to 69-year-old subjects showed reduced recovery of developed force (35.7% +/- 17% [n = 5] and 51.1% +/- 11.8% [n = 9], respectively) compared with that found (68.2% +/- 10.4% [n = 6]) in the group of 34- to 59-year-old subjects (P =.01). Multivariable analysis, comparing 20 factors of surgical patient characteristics and recovery of developed force, found that only age (P =.01) and hypertension (P =.01) were predictors of reduced recovery of developed force after either hypoxia or simulated ischemia. CONCLUSIONS: In aged human atrial myocardium, the capacity to recover contractile function after in vitro hypoxia or simulated ischemia is reduced compared with the younger myocardium of mature adults. These findings suggest that enhanced myocardial protective strategies may be indicated for elderly patients undergoing cardiac surgery.

Adult↗
PubMed1999

Mitochondrial DNA deletions in human cardiac tissue show a gross mosaic distribution.

The variability of mitochondrial DNA (mtDNA) deletional patterns has been investigated in adjacent slices of human heart atrium. Using quantitative PCR we found differential abundances of one particular mtDNA deletion, that of 4977 bp (mtDNA4977), among sets of adjacent slices of right atrial trabeculae pectinatae from 10 subjects. Some subjects had relatively constant abundance of mtDNA4977 among the tissue slices, while others covered a wide range. A qualitative PCR procedure was used to visualize the patterns of multiple deletions within an 8.64-kb segment of the mtDNA genome, in the same set of atrial trabeculae samples. Some subjects showed completely different multiple deletional patterns in each of the trabeculae slices analyzed. There was no correlation between the variation of the abundance of mtDNA4977 and that of the multiple deletions. The results are consistent with the notion that the occurrence of mtDNA deletions during aging is a random process, involving their production throughout the lifetime of an individual. In this view, the patterns of new deletions are superimposed on those already accumulated by propagation and segregation of mutations formed earlier in life.

Adult↗
PubMed1998

Tissue-specific distribution of multiple mitochondrial DNA rearrangements during human aging.

Mitochondria, according to the free radical theory of aging, are the major source of reactive oxygen species (ROS). The results, presented in this paper, question the role of reactive oxygen species in contributing significantly to the extent of mitochondrial bioenergy degradation of the tissues, which can be correlated with mtDNA rearrangements. We report here that mtDNA rearrangements, including deletions and duplications, in tissues from human aged subjects, occur in levels ranging from very low in liver, to considerable in cardiac muscle, to almost total in skeletal muscle. The extent of mtDNA rearrangements is correlated at both the individual tissue and cell level with cytochrome oxidase (COX) activity as the exemplifier of cellular bioenergy capacity. Thus, the ROS proposal in its simplest form as it affects mtDNA and mitochondrial electron transport system is not supported by the available data.

Aged↗
PubMed1992

Cardiopulmonary transplantation service establishment in the 1990s: Australia--a developing country?

1. A National Transplantation Service has been established at the Alfred Hospital performing more than 50 transplants per year. 2. Increased access to transplantation technology has facilitated an improvement in national population servicing from 2.7 to 6.2 transplants per million population per year. National funding of A$4.062 million per year has been secured. 3. Basic research into organ preservation has facilitated transcontinental organ procurement. 4. An active lung transplantation program has been established synchronous with the cardiac transplant service activities. 5. MCS program establishment has paralleled clinical transplantation activities. 6. Budget management and cost containment has been achieved through rationalisation of management protocols.

Australia↗
PubMed1991

Absence of particle-induced coronary vasoconstriction during cardioplegic infusion: is it desirable to use a microfilter in the infusion line?

Many cardiac surgical units now use a microfilter in the infusion line for delivery of crystalloid cardioplegic solution to protect against the potential hazards of particulate contamination of cardioplegic solution. The aim of this group of studies was to determine the effects of particulate contamination of cardioplegic solutions, in order to establish whether a microfilter is needed in the infusion line. Total particle counts performed on two commercial cardioplegic solutions were low, but there were sufficient particles greater than 10 microns in diameter to cause coronary vasoconstriction. In isolated rat hearts a 20-minute infusion of St. Thomas' Hospital cardioplegic solution produced a progressive reduction in coronary flow, which was not prevented by the inclusion of a 0.8 micron filter in the infusion line. Two studies were performed on canine hearts to determine the effects of unfiltered cardioplegic solution on coronary vascular resistance. In the first, cardioplegic solution at 20 degrees C was infused for 20 minutes at a constant pressure of 50 mm Hg and flow rate was measured. In the second, the same solution at 4 degrees C was infused at a constant flow rate for 2 1/2 minutes and the infusion pressure was measured. In neither study did coronary vascular resistance rise. A final clinical study involving patients undergoing coronary bypass surgery compared the effects on coronary resistance of infusion at a constant flow of filtered versus unfiltered cardioplegic solution (n = 10 in each group). There was a similar rise in coronary perfusion pressure in both groups during the infusion. We conclude that there is insufficient evidence of particle-induced coronary vasoconstriction to justify the expense of a microfilter in the cardioplegic infusion line.

Animals↗
PubMed1985

Control of refractory supraventricular arrhythmias after unsuccessful closed-chest His bundle ablation.

Six patients underwent attempted catheter ablation of the His bundle for control of refractory supraventricular tachyarrhythmias. Permanent complete heart block was achieved in only three patients. All six patients have remained asymptomatic without antiarrhythmic medications over a follow-up period of six to 17 months (mean 10 months). There were no complications of the procedure apart from mild elevation of creatine kinase levels in three patients. In this series, resumption of atrioventricular (AV) conduction following attempted His bundle ablation was not associated with recurrence of symptomatic arrhythmias. Preservation of AV conduction may also obviate the need for permanent ventricular pacing.

Adult↗