Prolonging myocardial preservation with a modified University of Wisconsin solution containing 2,3-butanedione monoxime and calcium

Stringham, James C.; Paulsen, Karen L.; Southard, James H.; Mentzer, Robert M.; Belzer, Folkert O.

doi:10.1016/s0022-5223(94)70332-9

Cited by 25 publications

(5 citation statements)

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“…BDM, an uncoupling agent, is used in many slicing protocols (e.g., Watson et al, 2017 ), and preparations of cardiac muscle strips ( Mulieri et al, 1989 ) to reduce tissue damage. BDM is also used to improve cardiac and skeletal muscle function after long cold-storage periods ( Stringham et al, 1994 ; van der Heijden et al, 2000 ) and to reduce reperfusion injury ( Voigtlander et al, 1999 ). All processing steps were performed on a cooled plate on a laminar flow workbench.…”

Section: Methodsmentioning

confidence: 99%

Long-Term Cultivation of Human Atrial Myocardium

et al. 2022

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Organotypic culture of human ventricular myocardium is emerging in basic and translational cardiac research. However, few institutions have access to human ventricular tissue, whereas atrial tissue is more commonly available and important for studying atrial physiology. This study presents a method for long-term cultivation of beating human atrial myocardium. After written informed consent, tissues from the right-atrial appendage were obtained from patients with sinus rhythm undergoing open heart surgery with cardiopulmonary bypass. Trabeculae (pectinate muscles) prepared from the samples were installed into cultivation chambers at 37°C with a diastolic preload of 500 μN. After 2 days with 0.5 Hz pacing, stimulation frequency was set to 1 Hz. Contractile force was monitored continuously. Beta-adrenergic response, refractory period (RP) and maximum captured frequency (fmax) were assessed periodically. After cultivation, viability and electromechanical function were investigated, as well as the expression of several genes important for intracellular Ca2+ cycling and electrophysiology. Tissue microstructure was analyzed by confocal microscopy. We cultivated 19 constantly beating trabeculae from 8 patient samples for 12 days and 4 trabeculae from 3 specimen for 21 days. Functional parameters were compared directly after installation (0 d) with those after 12 d in culture. Contraction force was 384 ± 69 μN at 0 d and 255 ± 90 μN at 12 d (p = 0.8, n = 22), RP 480 ± 97 ms and 408 ± 78 ms (p = 0.3, n = 9), fmax 3.0 ± 0.5 Hz and 3.8 ± 0.5 Hz (p = 0.18, n = 9), respectively. Application of 100 nM isoprenaline to 11 trabeculae at 7 d increased contraction force from 168 ± 35 μN to 361 ± 60 μN (p < 0.01), fmax from 6.4 ± 0.6 Hz to 8.5 ± 0.4 Hz (p < 0.01) and lowered RP from 319 ± 22 ms to 223 ± 15 ms. CACNA1c (L-type Ca2+ channel subunit) and GJA1 (connexin-43) mRNA expressions were not significantly altered at 12 d vs 0 d, while ATP2A (SERCA) and KCNJ4 (Kir2.3) were downregulated, and KCNJ2 (Kir2.1) was upregulated. Simultaneous Ca2+ imaging and force recording showed preserved excitation-contraction coupling in cultivated trabeculae. Confocal microscopy indicated preserved cardiomyocyte structure, unaltered amounts of extracellular matrix and gap junctions. MTT assays confirmed viability at 12 d. We established a workflow that allows for stable cultivation and functional analysis of beating human atrial myocardium for up to 3 weeks. This method may lead to novel insights into the physiology and pathophysiology of human atrial myocardium.

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Section: Methodsmentioning

confidence: 99%

Long-Term Cultivation of Human Atrial Myocardium

et al. 2022

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“…While our respiratory trials show no significant difference between the two chloride substitutes, there may be certain advantages of using one over the other. For example, lactobionate may possess several beneficial properties for mitochondria [ 18 , 70 ]. Lactobionate also has calcium-chelating properties, which helped decrease calcium contamination levels in buffer B2 without EGTA [ 20 , 22 ].…”

Section: Discussionmentioning

confidence: 99%

“…Lactobionate also has calcium-chelating properties, which helped decrease calcium contamination levels in buffer B2 without EGTA [ 20 , 22 ]. And prior research has found that lactobionate acts to decrease cellular injury in preserved tissue for organ transplantation, but the mechanism has yet to be demonstrated [ 18 , 70 ]. In addition to lower chloride concentrations, the lactobionate- and gluconate-based buffers could also improve mitochondrial function by favoring a more condensed, energy-transducing, mitochondrial state associated with ADP stimulated respiration [ 71 , 72 ].…”

Section: Discussionmentioning

confidence: 99%

The effect of respiration buffer composition on mitochondrial metabolism and function

et al. 2017

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Functional studies on isolated mitochondria critically rely on the right choice of respiration buffer. Differences in buffer composition can lead to dramatically different respiration rates leading to difficulties in comparing prior studies. The ideal buffer facilities high ADP-stimulated respiratory rates and minimizes substrate transport effects so that the ability to distinguish between various treatments and conditions is maximal. In this study, we analyzed a variety of respiration buffers and substrate combinations to determine the optimal conditions to support mitochondrial function through ADP-stimulated respiration and uncoupled respiration using FCCP. The buffers consisted of a standard KCl based buffer (B1) and three modified buffers with chloride replaced by the K-lactobionate, sucrose, and the antioxidant taurine (B2) or K-gluconate (B3). The fourth buffer (B4) was identical to B2 except that K-lactobionate was replaced with K-gluconate. The substrate combinations consisted of metabolites that utilize different pathways of mitochondrial metabolism. To test mitochondrial function, we used isolated cardiac guinea pig mitochondria and measured oxygen consumption for three respiratory states using an Oroboros Oxygraph-2k. These states were the leak state (energized mitochondria in the absence of adenylates), ADP-stimulated state (energized mitochondria in the presence of saturating ADP concentrations), and uncoupled state (energized mitochondria in the presence of FCCP). On average across all substrate combinations, buffers B2, B3, and B4 had an increase of 16%, 26%, and 35% for the leak state, ADP-simulated state, and uncoupled state, respectively, relative to rates using B1. The common feature distinguishing these buffers from B1 is the notable lack of high chloride concentrations. Based on the respiratory rate metrics obtained with the substrate combinations, we conclude that the adenine nucleotide translocase, the dicarboxylate carrier, and the alpha-ketoglutarate exchanger are partially inhibited by chloride. Therefore, when the goal is to maximize ADP-stimulated respiration, buffers containing K-lactobionate or K-gluconate are superior choices compared to the standard KCl-based buffers.

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“…In contrast to these theoretical considerations, there is strong evidence from various experimental settings that adjunctive application of BDM to cardioplegic solutions is beneficial. 4,10,11 Contraction uncoupling during initial postischemic reperfusion by BDM has been equally effective when applied over a 20-minute period in a dosage of 20 mmol/L. 9 The drug is fast acting and its effects are fully reversible within minutes.…”

Section: Discussionmentioning

confidence: 99%