Altered endothelium-derived hyperpolarizing factor-mediated endothelial function in coronary microarteries by St Thomas' Hospital solution.

OBJECTIVES: We examined the effect of St Thomas' Hospital solution on endothelium-derived hyperpolarizing factor-mediated function in the porcine coronary microarteries with emphasis on the effect of temperature and washout time. METHODS: Microartery rings (diameter, 200-450 micrometers) were studied in myograph. The arteries were incubated in St Thomas' Hospital or Krebs solution (control) at 4 degrees C for 4 hours followed by 45 minutes (group Ia) or 90 minutes washout (group Ib) or at 22 degrees C for 1 hour followed by 45 minutes (group IIa) or 90 minutes washout (group IIb) and precontracted with -8.5 log M U 46619. The endothelium-derived hyperpolarizing factor-mediated relaxation to bradykinin was studied when endothelium-derived nitric oxide and prostaglandin I2 were inhibited with the presence of 7 micromol/L indomethacin and 300 micromol/L NG-nitro-L -arginine. RESULTS: After exposure to St Thomas' Hospital solution, the maximal endothelium-derived hyperpolarizing factor-mediated relaxation (percentage of the precontraction) was significantly reduced at either temperature after washout for 45 minutes (group Ia, 42.7% +/- 3.5% vs 69.0% +/- 5.3%; n = 9; P =.000; and group IIa, 12.3% +/- 1.6% vs 56.1% +/- 4. 4%; n = 8; P =.000) but fully recovered after washout for 90 minutes. The U46619-induced contraction force was also significantly reduced after washout for 45 minutes (P <.001) but fully recovered at 90 minutes. CONCLUSIONS: Under profound and moderate hypothermia, St Thomas' Hospital solution impairs endothelium-derived hyperpolarizing factor-mediated relaxation and smooth muscle contraction in the coronary microarteries. These effects exist during the reperfusion period for at least 45 minutes after exposure to St Thomas' Hospital solution and may account for the possible myocardial dysfunction during reperfusion.     

Electrophysiologic and mechanical evidence of superiority of hyperpolarizing versus depolarizing cardioplegia in protection of endothelium-derived hyperpolarizing factor-mediated endothelial function: a study in coronary resistance arteries

OBJECTIVE: The advantages of hyperpolarizing cardioplegia with potassium-channel openers versus depolarizing cardioplegia have been suggested but not demonstrated in coronary microarteries. This study examined the simultaneous electric and tonic alteration of coronary microarteries at the cellular level during and after exposure to depolarizing cardioplegia or hyperpolarizing cardioplegia, with emphasis on endothelium-derived hyperpolarizing factor-mediated relaxation and hyperpolarization. METHODS: Porcine coronary microarteries (diameter, approximately 200-400 microm) were incubated with depolarizing cardioplegia (20 mmol/L KCl) or hyperpolarizing cardioplegia (10 micromol/L aprikalim) for 1 hour. Cellular membrane potential with a glass microelectrode in a coronary smooth muscle cell and isometric force of the muscle were simultaneously measured in a myograph. RESULTS: Depolarizing cardioplegia incubation produced a stable contraction (from 4.9 +/- 0.3 mN to 7.3 +/- 0.4 mN) and depolarization (from -51 +/- 1 mV to -41 +/- 2 mV). In contrast, hyperpolarizing cardioplegia relaxed (from 4.8 +/- 0.3 mN to 3.5 +/- 0.3 mN) and hyperpolarized (from -51 +/- 2 mV to -56 +/- 1 mV) the smooth muscle. After exposure to depolarizing cardioplegia, the bradykinin-induced, endothelium-derived hyperpolarizing factor-mediated relaxation reduced from 66.2% +/- 5.0% to 18.4% +/- 3.7% (P <.001), and the membrane hyperpolarization reduced from 18 +/- 1 mV to 7 +/- 1 mV (P <.001) in the presence of indomethacin and N(G)-nitro-L-arginine. In contrast, hyperpolarizing cardioplegia did not affect the bradykinin-induced responses. CONCLUSIONS: In the coronary microarteries, exposure to hyperpolarizing cardioplegia preserves whereas depolarizing cardioplegia reduces the endothelium-derived hyperpolarizing factor-mediated electric (hyperpolarization) and mechanical (relaxation) responses. Thus hyperpolarizing cardioplegia is superior to depolarizing cardioplegia in protecting the endothelial function in the coronary microcirculation.     

Alteration of cellular electrophysiologic properties in porcine pulmonary microcirculation after preservation with University of Wisconsin and Euro-Collins solutions

BACKGROUND: The effect of cold storage of porcine pulmonary microvessels in University of Wisconsin (UW) and Euro-Collins (EC) solutions on the cellular electrophysiologic properties remains unknown. METHODS: The pulmonary microarteries (PA, 381.6 +/- 62.8 microm; n = 60) and microveins (PV, 360.8 +/- 54.5 microm; n = 60) were incubated with Krebs (control), UW, or EC solution at 4 degrees C for 4 hours in a myograph. The resting membrane potential and the endothelium-derived hyperpolarizing factor-mediated hyperpolarization to bradykinin (0.1 micromol/L) in the presence of inhibitors of nitric oxide and prostacyclin, N(omega)-nitro-l-arginine, hemoglobin, and indomethacin, in a single smooth muscle cell were directly measured. RESULTS: The resting membrane potential (-60.8 +/- 1.3 mV in PA and -48.1 +/- 0.7 mV in PV, n = 6) was depolarized after exposure to UW solution (to -18.4 +/- 0.7 mV in PA and -13.6 +/- 0.8 mV in PV; n = 8; p < 0.001). The amplitude of endothelium-derived hyperpolarizing factor-mediated hyperpolarization to bradykinin was also decreased (from 7.4 +/- 0.7 mV to 2.6 +/- 0.7 mV in PA and from 4.6 +/- 0.5 mV to 0.9 +/- 0.4 mV in PV; p < 0.001). In comparison, EC depolarized the membrane potential to a lesser extent (to -28.3 +/- 0.9 mV in PA and to -21.3 +/- 0.8 mV in PV; n = 8; p < 0.001) and almost abolished the hyperpolarization to bradykinin. After washout, hyperpolarization was partially restored (UW, 4.9 +/- 0.7 mV in PA and 2.0 +/- 0.3 mV in PV. p < 0.01; EC, 2.3 +/- 0.5 mV in PA and 1.0 +/- 0.3 mV in PV. p < 0.01). CONCLUSIONS: Cold storage of porcine PA and PV with UW or EC solution impairs the electrophysiologic properties (hyperpolarization) related to endothelium-smooth muscle interaction. The alteration is more profound with EC than UW solution and in veins than in arteries. The findings urge further studies on lung preservation solutions.     

Protective effect of magnesium on the endothelial function mediated by endothelium-derived hyperpolarizing factor in coronary arteries during cardioplegic arrest in a porcine model

OBJECTIVES: Hyperkalemia in cardioplegia impairs the function mediated by endothelium-derived hyperpolarizing factor. This study examined the effect and mechanism of magnesium ion on the relaxation mediated by endothelium-derived hyperpolarizing factor. METHODS: In the isometric force study, porcine coronary microarteries in a myograph (diameter 200-450 microm) were incubated in Krebs solution (subgroups Ia, IIa, and IIIa), potassium ion (20 mmol/L, subgroups Ib, IIb, and IIIb), magnesium ion (16 mmol/L, subgroups Ic, IIc, and IIIc), or potassium ion plus magnesium ion (subgroups Id, IId, and IIId) for 1 hour at 37 degrees C in group I or II, followed by washout for 45 minutes in group III (n = 8). Relaxation to bradykinin (groups I and III) or sodium nitroprusside (group II) in U(46619)-stimulated contraction was established. In the electrophysiologic study, the membrane potentials of single smooth muscle cells of arteries were measured by microelectrode after superfusion with the previously described solutions (subgroups IVa-IVc). RESULTS: In group I, 20-mmol/L potassium ion greatly reduced the bradykinin-induced relaxation (35.0% +/- 4.9% vs 86.0% +/- 5.3%, P <.001), which was significantly restored by magnesium ion (51.9% +/- 4.0%, P =.017). In groups II and III, the bradykinin- or nitroprusside-induced relaxation had no significant differences. In group IV, potassium ion depolarized the smooth muscle and decreased the bradykinin-induced hyperpolarization (-72.0 +/- 1.5 vs -61.7 +/- 0.7 mV, n = 7, P <.001), which was significantly restored by magnesium ion (-68.0 +/- 2.5 mV vs -72.5 +/- 1.5 mV, n = 6, P =.029). CONCLUSIONS: Magnesium ion, either alone or added to hyperkalemic solutions, preserves or helps to restore the endothelial function mediated by endothelium-derived hyperpolarizing factor. The mechanism is related to preservation of the membrane hyperpolarization and reversal of the potassium-induced membrane depolarization of the smooth muscle cell.     

Effect of 11,12-epoxyeicosatrienoic acid as an additive to St. Thomas' cardioplegia and University of Wisconsin solutions on endothelium-derived hyperpolarizing factor-mediated function in coronary microarteries: influence of temperature and time

BACKGROUND: We examined the effect of 11,12-epoxyeicosatrienoic acid (EET(11,12)) added to St. Thomas' Hospital (ST) solution or University of Wisconsin (UW) solution on endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation under clinically relevant temperature and exposure time. METHODS: Porcine coronary microarteries (200 to 450 microm) were incubated with Krebs' solution (control), ST with or without EET(11,12) (300 nmol/L) at 22 degrees C for 1 hour as well as at 4 degrees C for 1 or 4 hours, and UW with or without EET(11,12) at 4 degrees C for 4 hours. The EDHF-mediated relaxation was induced by bradykinin (-10 to approximately -6.5 log M) in the precontraction evoked by U(46619) (10 nmol/L) or U(46619) (1 nmol/L) plus endothelin-1 (6 nmol/L). RESULTS: The EDHF-mediated relaxation was reduced after exposure to UW (79.7% +/- 4.6% versus 93.6% +/- 2.8%, p = 0.01) at 4 degrees C for 4 hours. One-hour exposure to ST under 22 degrees C or 4 degrees C decreased the relaxation (75.2% +/- 7.6% versus 96.7% +/- 1.6%, p < 0.05) or the sensitivity to bradykinin (-8.04 +/- 0.15 versus -8.50 +/- 0.20 log M, p < 0.05). The relaxation increased to 86.8% +/- 5.3% by addition of EET(11,12) to ST (1 hour at 22 degrees C, p < 0.05) but was unchanged when added to either ST or UW at 4 degrees C for 1 or 4 hours. CONCLUSIONS: As an additive to ST solution, EET(11,12) may partially restore EDHF-mediated endothelial function under moderate hypothermia but had no significant effect under profound hypothermia when added to either ST or UW solution. Further investigation is necessary to improve the effect.     

Histidine-tryptophan-ketoglutarate solution maximally preserves endothelium-derived hyperpolarizing factor-mediated function during heart preservation: comparison with University of Wisconsin solution.

BACKGROUND: The University of Wisconsin (UW) solution is used widely in heart preservation but has been demonstrated to be detrimental to the endothelial function. The present study compares the effect of histidine-tryptophan-ketoglutarate (HTK) and UW solutions on endothelium-derived hyperpolarizing factor (EDHF)-mediated function in porcine small coronary arteries. METHODS: An isometric force study was performed in a myograph and the membrane potential of a single smooth muscle cell was measured electrophysiologically. Small coronary arteries (diameter 457 +/- 15 microm) were incubated with UW (n = 8), HTK (n = 7) or Krebs solution (n = 15) at 4 degrees C for 4 hours. After washout, in the presence of indomethacin (Indo; 7 micromol/liter), N(G)-nitro-l-arginine (l-NNA; 300 micromol/liter) and oxyhemoglobin (HbO; 20 micromol/liter), bradykinin (BK; -10 to -6.5 log M)-induced relaxation was compared in U46619 (-8 log M) pre-contraction. EDHF-mediated hyperpolarization was elicited by BK (-6.5 log M) in the presence of Indo, l-NNA and HbO. RESULTS: BK-induced, EDHF-mediated relaxation was reduced from 93.6 +/- 2.8% to 79.7 +/- 4.6% after UW preservation (p = 0.01 by unpaired t-test and p = 0.005 by 2-way analysis of variance [ANOVA]), whereas HTK incubation did not decrease EDHF-mediated relaxation (87.0 +/- 6.5%, p = 0.3 by unpaired t-test and p = 0.6 by 2-way ANOVA, compared with control, and p = 0.001 by 2-way ANOVA, compared with UW). EDHF-mediated hyperpolarization (10.3 +/- 1.6 mV) was attenuated by UW exposure (3.4 +/- 0.6 mV, [p = 0.002] vs control), but not by HTK exposure (8.3 +/- 1.1 mV, [p = 0.3] vs control). CONCLUSIONS: HTK is superior to UW solution in protecting EDHF-mediated endothelial function in porcine small coronary arteries. The present findings supports the use of HTK solution in heart preservation.     

Effect of potassium-channel openers on the release of endothelium-derived hyperpolarizing factor in porcine coronary arteries stored in cold hyperkalemic solution

Hyperkalemic solution is widely used to protect the myocardium during open-heart surgery or to preserve donor hearts during heart or heart/lung transplants. The inhibitory effects of hvperkalemic solution on the release of endothelium-derived hyperpolarizing factor (EDHF) of coronary arteries following deep hypothermic storage (4 degrees C) has been well studied. However, it has not been established whether potassium channel openers have protective effects on the coronary endothelial function after cold storage. This study was designed to examine this. Porcine coronary artery rings were studied in organ baths. Relaxation in response to the EDHF stimulus A23187 (nonreceptor-mediated stimulus calcium ionophore) in thromboxane A2 mimetic U46619 (30 nmol/L)-induced precontraction after incubation with hyperkalemic solution (20 mmol/L) with nicorandil (10 micromol/L) (either at 37 degrees C in the oxygenated organ chamber or at 4 degrees C in a refrigerator for 6 h) was compared with the control. There was significant difference between hyperkalemia group and hyperkalemia with nicorandil group under normothermia (p = .04). The difference was significant in the same solution between normothermia and hypothermia. After incubation in hyperkalemic solution without or with nicorandil, the A23187-induced relaxation was 32.8% +/- 9.1% and 72.6% +/- 16.9%, respectively (N = 8, p < .01). Potassium channel opener can attenuate the inhibitory effect of hyperkalemic solution on the release of EDHF after cold storage.     

Comparison of University of Wisconsin and St Thomas' Hospital solutions on endothelium-derived hyperpolarizing factor-mediated function in coronary micro-arteries

BACKGROUND: It is controversial whether coronary endothelial function is impaired after cold exposure to University of Wisconsin (UW) or St. Thomas' Hospital (ST) solution during heart transplantation. We therefore examined the effects of cold storage of coronary micro-arteries with UW or ST solution on endothelium-derived hyperpolarizing factor (EDHF)-mediated function. METHODS: Porcine and human coronary arteries were immersed in either UW or ST solution at 4 degrees C for 4 hr and then normalized in a wire myograph. RESULTS: In the rings (normalized diameter: 200-500 microM) precontracted by U46619, EDHF-mediated relaxation and hyperpolarization were initiated by bradykinin (BK) or A23187 in the presence of indomethacin and NG-nitro-L-arginine. In the human coronary arteries, the EDHF-mediated relaxation to BK was reduced by UW solution from 53.2+/-5.6% to 24.0+/-2.7% (P=0.006). The reduced EDHF-mediated relaxation occurred concurrently with the decreased hyperpolarization to BK (17.0+/-1.5 vs. 10.5+/-1.1 mV, n=10, P=0.004) or A23187 in porcine coronary arteries. In the control arteries, K+ channel blockers, either glybenclamide or tetraethylammonium reduced the EDHF-mediated relaxation. After exposure to UW solution, the EDHF-mediated relaxation was further significantly inhibited. In contrast, ST solution did not affect these responses. CONCLUSIONS: These results show that in coronary micro-arteries, UW, but not ST, solution impairs the EDHF-mediated function and inhibits the Ca2+-activated and ATP-sensitive K+ channels. Our comparative study suggests that ST solution may be superior to UW solution in preserving the EDHF-related endothelial function of coronary micro-arteries.     

Hypoxic preconditioning in coronary microarteries: role of EDHF and K+ channel openers

BACKGROUND: Hypoxic preconditioning may provide a useful method of myocardial protection in cardiac operations. The present study was designed to investigate the possible mechanisms of preconditioning regarding endothelium-derived hyperpolarizing factor (EDHF) and the effect of a potassium channel opener KRN4884 on the porcine coronary microartery in mimicking hypoxic preconditioning. METHODS: Porcine coronary microartery rings (diameter 200 to 500 microm) studied in a myograph were divided into seven groups: (1) control group; (2) hypoxia-reoxygenation group (hypoxia for 60 minutes followed by reoxygenation for 30 minutes); (3) preconditioning group (hypoxia for 5 minutes followed by reoxygenation for 10 minutes before hypoxia reoxygenation); (4) KRN4884 pretreatment group (KRN4884 was added into the myograph chamber 20 minutes before hypoxia reoxygenation); (5) 5-hydroxydecanoate + KRN group (5-hydroxydecanoate was given 20 minutes before KRN4884 pretreatetment); (6) glibenclamide (GBC) + KRN group (GBC was added 20 minutes before KRN4884 pretreatment); and (7) endothelium denuded group (the endothelium was removed). The endothelium-derived hyperpolarizing factor-mediated relaxation to bradykinin was studied in the rings precontracted with U46619 in the presence of N(omega)-nitro-L-arginine and indomethacin. RESULTS: The maximal relaxation induced by bradykinin was reduced in hypoxia reoxygenation (40.7% +/- 2.8% vs 66.9% +/- 2.5% in control, p = 0.000). This reduced relaxation was recovered in either preconditioning (64.6% +/- 4.6%, p = 0.002), or KRN4884 pretreatment (67.1% +/- 3.6%, p = 0.000). The 5-hydroxydecanoate, but not GBC pretreatment abolished the effect of KRN44884 pretreatment (67.1% +/- 3.6% vs 42.9% +/- 3%, p = 0.001). CONCLUSIONS: Hypoxia reoxygenation reduces the relaxation mediated by endothelium-derived hyperpolarizing factor in the coronary microartery. This function can be restored by either hypoxic preconditioning or the K(ATP) channel opener KRN4884, and therefore K(ATP) channel openers may provide similar effect as preconditioning. The mechanism is mainly related to the mitochondrial ATP-sensitive K+ channels.     

Epoxyeicosatrienoic acids (EET(11,12)) may partially restore endothelium-derived hyperpolarizing factor-mediated function in coronary microarteries.

Background. Endothelial cells derive nitric oxide, prostacyclin, and endothelium-derived hyperpolarizing factor (EDHF). The cytochrome P-450–monooxygenase metabolites of arachidonic acid (epoxyeicosatrienoic acids [EETs]) have been suggested to be EDHF. This study was designed to examine the effect of EET_11,12 with regard to the possibility of restoring EDHF function when added into hyperkalemic cardioplegic solution.

Methods. Porcine coronary microartery rings were studied in a myograph. In groups 1 and 2, paired arteries were incubated in either hyperkalemic solution (K⁺ 20 mmol/L) or Krebs’ solution (control). In group 3, the paired arteries were incubated in hyperkalemia plus EET_11,12 (1 × 10^−6.5 mol/L) or hyperkalemia alone (control) at 37°C for 1 hour, followed by Krebs’ washout and then precontracted with 1 × 10^−8.5 mol/L U46619. The EDHF-mediated relaxation to EET_11,12 (group 1) or bradykinin (groups 2 and 3) was studied in the presence of N^G-nitro-l-arginine, indomethacin, and oxyhemoglobin.

Results. After exposure to hyperkalemia, the EDHF-mediated maximal relaxation by bradykinin (72.5% ± 7.8% versus 41.6% ± 10.6%; p < 0.05), but not by EET_11,12 (18.4% ± 3.3% versus 25.1% ± 4.9%; p > 0.05) was significantly reduced. Incubation with EET_11,12 partially restored EDHF function (33.3% ± 9.5% versus 62.0% ± 8.5%; p < 0.05).

Conclusions. In coronary microarteries, hyperkalemia impairs EDHF-mediated relaxation, and EET_11,12 may partially mimic the EDHF function. Addition of EET_11,12 into cardioplegic solution may partially restore EDHF-mediated function reduced by exposure to hyperkalemia.     

University of Wisconsin solution for pulmonary preservation in a rat transplant model.

University of Wisconsin and modified Euro-Collins solutions for pulmonary preservation were compared in a rat orthotopic left lung isotransplant model. Heart-lung blocks of donor rats were flushed with and preserved in one of the preservation solutions at 0 degrees C. After 6 or 12 hours of cold ischemia, the left lungs were transplanted into recipient rats and reperfused for 1 hour. Pulmonary function was assessed by measuring oxygen and carbon dioxide tensions in arterial blood after removal of the right lung. Lipid peroxide concentrations were measured as thiobarbiturate acid-reactive substances. The ratios of wet to dry weight of grafts after ischemia and after reperfusion were calculated. Histologic changes of ischemia-reperfusion injury of the lung tissue were evaluated using a graded scale. Oxygen tension after 6 hours of preservation followed by reperfusion was significantly higher with University of Wisconsin solution (308.8 +/- 81.1 mm Hg) than with Euro-Collins solution (50.8 +/- 17.8 mm Hg; p less than 0.001). Carbon dioxide tension in the University of Wisconsin solution group was also significantly lower than in the Euro-Collins solution group (28.2 +/- 2.3 versus 46.0 +/- 4.5 mm Hg; p less than 0.05). Lipid peroxide concentration after 6 hours' preservation in University of Wisconsin solution was significantly lower (0.88 +/- 0.07 mumol/g) than that in Euro-Collins solution (1.26 +/- 0.12 mumol/g; p less than 0.05). After 12 hours of preservation only lipid peroxide concentration with University of Wisconsin solution was significantly lower (1.30 +/- 0.09 mumol/g) than with Euro-Collins solution (1.71 +/- 0.15 mumol/g; p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Raffinose improves the function of rat pulmonary grafts stored for twenty-four hours in low-potassium dextran solution

OBJECTIVES: The perfect strategy for pulmonary graft preservation remains elusive. Experimental work supports the use of perfusates, such as Euro-Collins, University of Wisconsin, and low-potassium dextran solutions. We use low-potassium dextran solution in our clinical program, but we aim for continued improvement. The trisaccharide raffinose has been shown to be responsible for the efficacy of University of Wisconsin perfusate in lung preservation. Raffinose is superior to a variety of other saccharides for this purpose. We tested the hypothesis that the addition of raffinose to low-potassium dextran solution might further improve graft function. METHODS: In a randomized blinded study with a rat left lung transplant model, donor lungs were flushed with either standard low-potassium dextran solution or low-potassium dextran solution modified by the addition of 30 mmol/L raffinose (n = 5 for each group). Alprostadil (prostaglandin E(1), 500 microg/L) was added to the perfusates in accordance with our clinical practice. Grafts were stored inflated at 4 degrees C for 24 hours. After transplantation, recipients were ventilated with a fraction of inspired oxygen of 1 and a positive end-expiratory pressure of 2 cm H(2)O. Graft function was evaluated by measuring oxygenation at 2 hours after graft reperfusion, peak airway pressure throughout the reperfusion period, and the wet/dry lung weight ratio. RESULTS: The group receiving low-potassium dextran solution with raffinose demonstrated significantly higher oxygenation (oxygen tension, 370 +/- 45 mm Hg vs 150 +/- 64 mm Hg; P =.0025), lower peak airway pressures at 2 hours after lung reperfusion (11 +/- 2.7 mm Hg vs 16 +/- 2.4 mm Hg; P <.001), and a lower wet/dry weight ratio (4.7 +/- 1.26 vs 11 +/- 5. 0; P =.017). CONCLUSION: Modification of low-potassium dextran solution with the trisaccharide raffinose resulted in a significant improvement in graft function in this model and merits further evaluation with respect to the mechanisms involved.     

Procaine in cardioplegia: the effect on EDHF-mediated function in porcine coronary arteries

OBJECTIVES: Hyperkalemia in cardioplegia impairs the endothelium-derived hyperpolarizing factor (EDHF)-mediated function. This study examined the effect of procaine in cardioplegia on the EDHF-mediated response in porcine coronary arteries. METHODS: An isometric force study was performed in a myograph. Two rings taken from the same artery (diameter 200-450 microm) were incubated with Krebs solution (group I) or 20 mM K+ (group II) with/without procaine (1 mM) at 37 degrees C for 1 hour. The EDHF-mediated relaxation was induced by bradykinin (BK, -10 approximately -6.5 log M) after U46619 (-8 log M, in group I) or K+-precontraction (in group II) in the presence of indomethacin (7 microM), NG-nitro-L-arginine (300 microM), and hemoglobin (20 microM). The membrane potential of a single smooth muscle cell was measured by a microelectrode after superfusion with Krebs solution with/without procaine for 1 hour. RESULTS: The EDHF-mediated relaxation was increased by the treatment with procaine with the EC50 shifted leftward (97.3 +/- 0.6% vs. 83.0 +/- 5.1% at -7 log M and 99.4 +/- 0.6% vs. 96.7 +/- 1.6% at -6.5 log M, p < 0.05; EC50: -8.57 +/- 0.24 vs. -7.92 +/- 0.23 log M, p < 0.05). Procaine decreased the BK-induced hyperpolarization from -72.3 +/- 0.7 mV to -68.8 +/- 0.8 mV (-6.5 log M, p < 0.01). The EDHF-mediated relaxation in arteries exposed to 20 mM K+ was not altered by procaine (49.9 +/- 7.4% vs. 55.8 +/- 7.6%, p > 0.05). CONCLUSIONS: In the coronary arteries, procaine has a depolarizing effect but it enhances EDHF-mediated relaxation. Addition of procaine in cardioplegia did not change the EDHF-mediated endothelial function.     

Beneficial effect of polyethylene glycol in lung preservation: early evaluation by proton nuclear magnetic resonance spectroscopy

BACKGROUND: Proton nuclear magnetic resonance spectroscopy can be used to measure organic molecules in biological fluids. In this study, proton nuclear magnetic resonance spectroscopy of bronchoalveolar lavage was assessed to detect cellular damage in lung transplants. Also we evaluated a polyethylene glycol solution in lung preservation. METHODS: An isolated perfused and working pig lung was used to assess initial pulmonary function after in situ cold flush and cold storage for 6 hours in three preservation solutions: (1) Euro-Collins solution, (2) University of Wisconsin solution, and (3) low potassium solution with polyethylene glycol (PEG). Pulmonary vascular resistance and partial pressure of arterial oxygen were measured during reperfusion. Bronchoalveolar lavage was studied by proton nuclear magnetic resonance spectroscopy and a histologic study of the lungs was done at the harvest after ischemia and after reperfusion. RESULTS: Partial pressure of arterial oxygen and pulmonary vascular resistance were significantly better in PEG compared with Euro-Collins solution (p = 0.011). Interstitial edema was significantly higher in Euro-Collins solution (2.4 +/- 0.24; p = 0.02) and University of Wisconsin solution (2.7 +/- 0.20; p = 0.0003) than PEG (2 +/- 0.16). Mitochondria scale was better in PEG (8.1 +/- 0.46) than in Euro-Collins solution (6.2 +/- 0.37; p = 0.0001) or University of Wisconsin solution (5.6 +/- 1.36; p = 0.0046). In bronchoalveolar lavage proton nuclear magnetic resonance spectroscopy spectra, lactate, pyruvate, citrate, and acetate were only detected after reperfusion, with a significantly reduced production of acetate in PEG. Pyruvate was reduced at the limit of significance in PEG versus University of Wisconsin solution. CONCLUSIONS: Proton nuclear magnetic resonance spectroscopy seems to be a simple and suitable method for assessment of early injury to the lung transplant. In this experimental study, PEG preserved the lung better than University of Wisconsin solution and Euro-Collins solution in both the proton nuclear magnetic resonance spectroscopy study as well as the physiologic study.     

Superior protection in orthotopic rat lung transplantation with cyclic adenosine monophosphate and nitroglycerin-containing preservation solution.

BACKGROUND: Primary lung graft failure is common, and current lung preservation strategies are suboptimal. Because the decline in lung levels of cyclic adenosine monophosphate and cyclic guanosine monophosphate during preservation could enhance adhesiveness of endothelial cells for leukocytes as well as increase vascular permeability and vasoconstriction, we hypothesized that buttressing these levels by means of a preservation solution would significantly improve lung preservation. METHODS: An orthotopic rat left lung transplantation model was used. Lungs were harvested from male Lewis rats and preserved for 6 hours at 4 degrees C with (1) Euro-Collins solution (n = 8); (2) University of Wisconsin solution (n = 8); (3) low-potassium dextran glucose solution (n = 8); (4) Columbia University solution (n = 8), which contains a cyclic adenosine monophosphate analog (dibutyryl cyclic adenosine monophosphate) and a nitric oxide donor (nitroglycerin) to buttress cyclic guanosine monophosphate levels; or (5) Columbia University solution without cyclic adenosine monophosphate or nitroglycerin (n = 8). PaO2, pulmonary vascular resistance, and recipient survival were evaluated 30 minutes after left lung transplantation and removal of the nontransplanted right lung from the pulmonary circulation. RESULTS: Among all groups studied, grafts stored with Columbia University solution demonstrated the highest Pa O2 (355 +/- 25 mm Hg for Columbia University solution versus 95 +/- 22 mm Hg for Euro-Collins solution, P <.01, 172 +/- 55 mm Hg for University of Wisconsin solution, P <.05, 76 +/- 15 mm Hg for low-potassium dextran glucose solution, P <.01, and 82 +/- 25 mm Hg for Columbia University solution without cyclic adenosine monophosphate or nitroglycerin, P <.01) and the lowest pulmonary vascular resistances (1 +/- 0.2 mm Hg * mL-1 * min-1 for Columbia University solution versus 12 +/- 4 mm Hg * mL-1 * min-1 for Euro-Collins solution, P <.01, 9 +/- 2 mm Hg * mL-1 * min-1 for University of Wisconsin solution, 14 +/- 6 mm Hg * mL-1 * min-1 for low-potassium dextran glucose solution, P <.01, and 8 +/- 2 mm Hg * mL-1 * min-1 for Columbia University solution without cyclic adenosine monophosphate and nitroglycerin). These functional and hemodynamic improvements provided by Columbia University solution were accompanied by decreased graft leukostasis and decreased recipient tumor necrosis factor alpha and interleukin 1alpha levels compared with the other groups. In toto, these improvements translated into superior survival among recipients of Columbia University solution-preserved grafts (100% for Columbia University solution, 37% for Euro-Collins solution, P <.01, 50% for University of Wisconsin solution, P <.05, 50% for low-potassium dextran glucose solution, P <.05, and 13% for Columbia University solution without cyclic adenosine monophosphate and nitroglycerin, P <.01). CONCLUSION: Nitroglycerin and cyclic adenosine monophosphate confer beneficial vascular effects that make Columbia University solution a superior lung preservation solution in a stringent rat lung transplantation model.     

Donor heart preservation with a novel hyperpolarizing solution: superior protection compared with University of Wisconsin solution

OBJECTIVES: A donor heart preservation solution was designed to use hyperpolarized arrest with the adenosine triphosphate-sensitive potassium-channel opener pinacidil. This solution contained concentrations of potassium, sodium, calcium, magnesium, lactobionate, and the buffer histidine specifically chosen to minimize intracellular calcium accumulation associated with prolonged ischemia. METHODS: Twenty-four rabbit hearts were randomly assigned to receive 1 of 3 preservation solutions in a crystalloid-perfused Langendorff model: (1) prototype solution containing a 0.5 mmol/L concentration of pinacidil, (2) prototype solution without pinacidil as control, and (3) University of Wisconsin solution. Thirty minutes of initial perfusion preceded baseline data acquisition. Data comprised left ventricle pressure-volume curves generated by inflating an intraventricular latex balloon. After cardioplegic administration, hearts underwent 4 hours of hypothermic storage, followed by 60 minutes of reperfusion and postischemic data acquisition. RESULTS: Postischemic developed pressure was better preserved by pinacidil solution (92.4% +/- 4.5%) than by the control (74.9% +/- 3.4%, P =.01) and University of Wisconsin solutions (66.7% +/- 5.1%, P =.001). Diastolic negative dP/dT was better preserved by pinacidil solution (104.4% +/- 10.2%) than by the control (80.2% +/- 4.2%, P =.034) and University of Wisconsin solutions (71.7% +/- 7.0%, P =.015). Diastolic compliance, expressed as baseline/postischemic diastolic slope ratios, was more poorly preserved by University of Wisconsin solution (0.67 +/- 0.07) than by the pinacidil (0.88 +/- 0.05, P =.041) and control solutions (0.87 +/- 0.05, P =.021). Postischemic coronary flow was higher in hearts exposed to pinacidil solution (77.8% +/- 3.0%) than in those exposed to the control (66.8% +/- 2.4%) and University of Wisconsin solutions (70.9% +/- 4.0%, P =.07). CONCLUSIONS: The superiority of the pinacidil solution in this experiment demonstrated that hyperpolarized arrest with potassium-channel openers improves donor heart preservation when administered in a novel histidine-buffered lactobionate-enriched vehicle.     

Epoxyeicosatrienoic acids (EET11,12) may partially restore endothelium-derived hyperpolarizing factor–mediated function in coronary microarteries

Background. Endothelial cells derive nitric oxide, prostacyclin, and endothelium-derived hyperpolarizing factor (EDHF). The cytochrome P-450–monooxygenase metabolites of arachidonic acid (epoxyeicosatrienoic acids [EETs]) have been suggested to be EDHF. This study was designed to examine the effect of EET_11,12 with regard to the possibility of restoring EDHF function when added into hyperkalemic cardioplegic solution.Methods. Porcine coronary microartery rings were studied in a myograph. In groups 1 and 2, paired arteries were incubated in either hyperkalemic solution (K⁺ 20 mmol/L) or Krebs’ solution (control). In group 3, the paired arteries were incubated in hyperkalemia plus EET_11,12 (1 × 10^−6.5 mol/L) or hyperkalemia alone (control) at 37°C for 1 hour, followed by Krebs’ washout and then precontracted with 1 × 10^−8.5 mol/L U46619. The EDHF-mediated relaxation to EET_11,12 (group 1) or bradykinin (groups 2 and 3) was studied in the presence of N^G-nitro-l-arginine, indomethacin, and oxyhemoglobin.Results. After exposure to hyperkalemia, the EDHF-mediated maximal relaxation by bradykinin (72.5% ± 7.8% versus 41.6% ± 10.6%; p < 0.05), but not by EET_11,12 (18.4% ± 3.3% versus 25.1% ± 4.9%; p > 0.05) was significantly reduced. Incubation with EET_11,12 partially restored EDHF function (33.3% ± 9.5% versus 62.0% ± 8.5%; p < 0.05).Conclusions. In coronary microarteries, hyperkalemia impairs EDHF-mediated relaxation, and EET_11,12 may partially mimic the EDHF function. Addition of EET_11,12 into cardioplegic solution may partially restore EDHF-mediated function reduced by exposure to hyperkalemia.     

Impaired EDHF-Mediated Relaxation in Porcine Pulmonary Micro-Arteries by Cold Storage with University of Wisconsin and Euro-Collins Solutions

Background: Vascular endothelium plays a key role in regulation of vascular tone. Hyperkalemia has been demonstrated to impair the EDHF‐mediated endothelial function in coronary circulation. University of Wisconsin (UW) and Eruo‐collins (EC) solutions are used for organ preservation in transplantation surgery. The potassium concentration in UW or EC solutions is as high as 125 mmol/L or 115 mmol/L, respectively. This study was designed to examine whether hyperkalemia or storage with UW and EC solutions affects the relaxation mediated by EDHF in the porcine pulmonary micro‐arteries. Methods: Porcine pulmonary micro‐artery rings (diameter 200–450 μm) were studied in myograph (n = 8 in each group). After incubation with hyperkalemia (K⁺ 125 mmol/L, at 37° C), UW or EC solutions (at 4° C for 4 hours), EDHF‐mediated relaxation induced by bradykinin (BK, ?10 to ?6.5 log M) in the presence of inhibitors for cyclooxygenase (Indomethacin, 7 μM), nitric oxide synthase (N^G‐nitro‐_L‐arginine, 300 μM), and oxyhemoglobin (20 μM) was compared with control (Krebs' solution) in precontraction with U₄₆₆₁₉ (?7.5 log M). Results: The EDHF‐mediated relaxation to BK was 69.6 ± 6.3% compared with 97.1 ± 1.7% (p= 0.003) in control (no inhibitors). After incubation with hyperkalemia, the relaxation significantly decreased (38.6 ± 3.0% vs. 59.1 ± 7.4%, p= 0.03 ). Storage with UW or EC solutions also significantly decreased the relaxation (49.3 ± 7.3% vs. 65.2 ± 3.5%, p= 0.04 and 51.9 ± 8.4% vs. 60.3 ± 6.1%, p= 0.02 , respectively). Conclusions: In porcine pulmonary micro‐arteries, exposure to hyperkalemia or storage with UW or EC solutions at 4°C for 4 hours impairs the EDHF‐mediated endothelial function. The clinical significance of this effect should be further studied.     

Cellular electrophysiological and mechanical effects of celsior solution on endothelial function in resistance coronary arteries

BACKGROUND: We investigated a relatively new organ preservation (Celsior) solution regarding its effect on the endothelium-derived hyperpolarizing factor (EDHF)-mediated function with comparison to St. Thomas Hospital (ST) solution. METHODS: The EDHF-mediated relaxation was induced by bradykinin (BK, -10 to -6.5 logM) in the presence of inhibitors of nitric oxide and prostacyclin in porcine small resistance coronary arteries, before and after incubation in ST (Group Ia, n=11), Celsior (Group Ib, n=13), or Krebs (Group Ic, control, n=12) at 4 degrees C for 4 hr. The EDHF-mediated hyperpolarization of the membrane potential of smooth muscle cells was measured by microelectrode with simultaneous relaxation after cold storage in ST (IIa, n=7), Celsior (IIb, n=6), or Krebs (IIc, control, n=6), or followed by washout with Krebs (ST: IIIa, n=6, Celsior: IIIb, n=6). RESULTS: The EDHF-mediated relaxation was significantly decreased in Group Ia (56.4+/-7.2% vs. 71.2+/-5.3%, P<0.05) and Ib (44.8+/-4.9% vs. 74.7+/-3.3%, P<0.05) but not in Ic. The sensitivity to BK was also significantly decreased (Ia: -7.51+/-0.14 vs. -7.76+/-0.12 log M, P<0.05; Ib: -7.36+/-0.09 vs. -7.60+/-0.09 logM, P<0.05). The resting membrane potential was depolarized in IIa (-44.3+/-1.9 mV, n=7, P<0.05) and IIb (-33.0+/-2.2 mV, n=6, P<0.05) compared with IIc (-57.1+/-1.5 mV, n=6). The EDHF-mediated hyperpolarization decreased significantly in IIa and IIb (3.4+/-0.3 and 3.0+/-0.2 vs. 6.3+/-0.5 mV, P<0.05) and partially restored in IIIa (5.0+/-0.2 vs. 3.4+/-0.3 mV, P<0.05) and IIIb (4.1+/-0.3 vs. 3.0+/-0.2 mV, P<0.05). CONCLUSIONS: Storage with Celsior and ST solutions reduces the EDHF-mediated endothelial function (hyperpolarization and associated relaxation) in porcine small resistance coronary arteries.     

Low-potassium dextran solution ameliorates reperfusion injury of the lung and protects surfactant function

OBJECTIVE: This study was designed to compare the effect of lung preservation with low-potassium dextran solution and Euro-Collins solution on reperfusion injury and surfactant function by using an in situ model of warm ischemia. METHODS: The left lungs of 6 minipigs were selectively perfused with Euro-Collins solution. In an additional 6 animals low-potassium dextran solution was used for flush perfusion. After 90 minutes of warm ischemia, the lungs were reperfused, and the contralateral pulmonary artery and bronchus were clamped. Hemodynamic and respiratory measurements were obtained for 7 hours of reperfusion. Surface tension of bronchoalveolar lavage and surfactant small and large aggregates were determined before perfusion (right lung) and after 2 hours of reperfusion (left lung). RESULTS: In the group receiving Euro-Collins solution, right heart failure developed within 215 +/- 39 minutes of reperfusion. An increase in minimal surface tension (P =.03), surfactant small aggregates/large aggregates ratio (P =.003), and bronchoalveolar lavage protein content (P =.012) were found after 2 hours of reperfusion. In the group receiving low-potassium dextran solution, all minipigs survived (P =.0001). Dynamic lung compliance (P =.034) and oxygen tension/inspired oxygen fraction ratios were higher (P =. 0001). Lung water content was lower (P =.049). The increase of minimal surface tension (P =.02) and bronchoalveolar lavage protein concentration (P =.015) were significantly less. CONCLUSION: Preservation of the lung with Euro-Collins solution leads to a reduction of physical surfactant function during reperfusion. Low-potassium dextran solution protects surfactant function and metabolism, thereby reducing reperfusion injury of the lung.