Citric acid as a decalcifying agent for the excised calcified human heart valves.

OBJECTIVE: Cardiac valvular pathologies are frequently encountered as mechanical and functional disorders due to the calcification of the valves whatever the etiologies are. This pathophysiologic table usually ends up with valvular replacement. In this study, we aimed to decrease/eliminate the calcium in the excised calcified human heart valves by using citric acid in vitro hence bringing about the question for possible oral treatment of calcification of the valves by citric acid ingestion. METHODS: Fourteen pieces of mitral and/or aortic valves excised from 12 patients undergoing valve replacement were placed in a freshly prepared phosphate buffered saline solution containing 0.625% glutaraldehyde at +4 0C for 48 h. They were rinsed with 0.9% NaCl and divided into two groups; study and control. Control tissues were further treated in a freshly prepared solution with identical properties for another 5 days. Study tissues were placed into a solution containing 3.8% citric acid (pH 7.4) and kept for 48 h at +37 degrees C, then rinsed with 0.9% NaCl and transferred into a fresh solution containing 0.625% glutaraldehyde with phosphate buffer at 37 0C for 3 more days. Specimens were biochemically and histopathologically evaluated and compared using Mann Whitney U test. RESULTS: Calcium and phosphate levels in the study group were lower than in the control group (852.5+/-913.41 microg g-1 vs 413.05+/-519.53 microg g-1, p=0.001 and 207.6+/-321.86 microg g-1 vs 124.4+/-289.48 microg g-1, p=0.035, respectively). Malondialdehyde and protein level values were changed insignificantly in the control and study groups. Histopathologic evaluation showed that collagen and elastin fibers were similar in both groups. In the study group, irregular and fusiform calcific formations around the collagen fibers were significantly decreased. CONCLUSIONS: Decalcifying human heart valves in vitro conditions with citric acid without an adverse change to the morphology of the valvular tissue specimens is meaningful. We believe that forwarding and looking for the answer to the question "whether systemic application of citric acid could lead to the decalcification and/or reduction of calcification in the native human heart valves" would be expressive.

Postoperative effects of low-dose intrathecal morphine in coronary artery bypass surgery.

BACKGROUND: Intrathecal morphine has been used in hopes of providing long-lasting postoperative analgesia in patients after cardiac surgery. The aim of this study was to evaluate the effects of 7 micro/kg intrathecal morphine administration in coronary bypass surgery in the postoperative period. METHODS: We conducted a prospective, randomized, blinded, and controlled study. Twenty-three patients, who underwent primary elective coronary bypass surgery, were randomly allocated to receive morphine 7 micro/kg intrathecally, before the induction of general anesthesia (Group M, n = 12) or no intrathecal injection (Group C, n = 11). Pain scores, determined by visual analogue scale (VAS), were recorded immediately after extubation upon admission to the intensive care unit (ICU), at the 2nd, 4th, 6th, and 18th hour after extubation. Pethidine was administered if the patient's VAS > or = 4 and consumption was recorded. Extubation time and ICU length of stay were also recorded. RESULTS: VAS scores were lower in the Group M at each measured time than the control group (p = 0.016, 0.023, 0.004, 0.0001, and 0.001, respectively). According to the VAS scores, pethidine requirement was lower in the Group M than the control (p = 0.001). Extubation time (3.58 +/- 1.57 vs. 4.86 +/- 1.38 hours, p = 0.045) and ICU length of stay (16.25 +/- 2.70 vs. 19.30 +/- 2.45 hours, p = 0.014) were also significantly shorter in the Group M than the control group. No significant complications were seen in this group of patients. CONCLUSIONS: Intrathecal morphine provided effective analgesia, earlier tracheal extubation and less ICU length stay after on-pump coronary bypass surgery. The influence on ICU length of stay requires further evaluations.

Prevention of calcification with TPEN in pericardial bioprosthetic heart valve material.

OBJECTIVE: Calcification is a frequent cause of the clinical failure of bioprosthetic heart valves fabricated from glutaraldehyde pretreated bovine pericardium. The major object of the present study is to prevent calcification of pericardial bioprosthetic heart valve materials with TPEN. METHODS: Bovine pericardium was cut into 2-cm 2 pieces, rinsed in phosphate-buffered saline solution, transferred into +4 degrees C phosphate-buffered saline containing 0.625% glutaraldehyde for initial fixation for 48 h, and allocated into two groups. Control samples were treated in an identical fresh solution for five more days. Others underwent additional fixation in phosphate-buffered saline 2microM TPEN for 48 h. They were then transferred into phosphate-buffered saline + 0.625% glutaraldehyde solution at 37 degrees C (pH 7.4) for three more days. Pericardial patches were inserted into the dorsal pouches of 18 juvenile male Wistar rats as control and study groups. Rats were divided into two groups and sacrificed consecutively by the end of 9th and 12th weeks. The biomechanical properties and calcium contents of explanted tissues were tested and were also assessed histopathologically. RESULTS: The difference in the calcium contents of the control and study groups' pericardial tissues at the 9th, and 12th weeks were statistically significant (p=0.0001, p=0.0001). The comparison of calcium contents between controls of 9th and 12th weeks and study groups' of the 9th and 12th weeks pericardial tissues were also significant (p=0.0001 and p=0.0001). Histopathologic and biomechanical assessment also supported these findings. CONCLUSION: Calcific degeneration of glutaraldehyde-fixed bovine pericardium can be reduced by using TPEN without any effect on durability.

Iloprost for the attenuation of ischaemia/reperfusion injury in a distant organ.

The objective of this study was to investigate antioxidant and cytoprotective properties of iloprost in a distant organ after ischaemia reperfusion injury. Male Wistar rats were divided into two groups. After application of anesthaesia both hindlimbs were occluded. A 2-h reperfusion procedure was carried out after 60 min of ischemia. Study group (STU) rats (n=10) received 10 microg kg(-1) iloprost in 1 ml of saline from the tail vein 10 min before reperfusion. Control (CON) group rats (n=10) received an equal amount of saline. The rats were sacrificed by injection of a high dose of thiopentone sodium. Blood and tissue samples (right kidneys) were taken for analysis. Differences in malondialdehyde (MDA), myeloperoxidase (MPO), Na+-K+ ATPase and total antioxidant capacity (TAC) between the groups were analysed. MPO, MDA and TAC levels in the sera of CON and STU groups were 1.60+/-0.26 U l(-1), 11.42+/-5.23 nmol ml(-1), 8.30 x 10(-2)+/- 3.93 x 10(-2) nmol ml(-1) h(-1) and 1.07+/-0.11 U l(-1), 7.60+/-1.81 nmol ml(-1) and 0.15+/-3.23 x 10(-2) nmol ml(-1) h(-1) (p=0.0001, p=0.043 and p=0.0001 respectively). MPO, ATPase and MDA levels in kidneys for CON and STU groups were 1.24+/-0.58 U g(-1), 85.70+/-52.05 nmol mg(-1), 17.90+/-7.40 nmol ml(-1) and 0.78+/-0.31 U g(-1), 195.90+/-56.13 nmol mg(-1) and 10.10+/-0.99 nmol ml(-1) (p=0.046, p=0.0001 and p=0.009 respectively). When given prior to reperfusion, the positive effect of iloprost in the attenuation of distant organ reperfusion injury has been demonstrated.

Hearing loss in patients undergoing coronary artery bypass grafting with or without extracorporeal circulation.

BACKGROUND: Our aim was to investigate the differences in postoperative hearing thresholds in patients undergoing coronary artery bypass grafting with (Group I, n=20) or without (Group II, n=17) extracorporeal circulation. MATERIAL/METHODS: 37 patients undergoing coronary artery bypass grafting with or without extracorporeal circulation were prospectively evaluated in terms of hearing threshold changes with the intention of documenting hearing losses postoperatively. The t-test for two independent variables was used for statistical analysis. RESULTS: Hearing threshold changes were detected in 9 Group I patients (45%) and 3 Group II patients (17.65%). The difference between the two groups was statistically significant (p=0.0426). CONCLUSIONS: Postoperative hearing threshold changes, not necessarily revealed by clinical examinations, are encountered after coronary artery bypass grafting operations. Extracorporeal circulation usage seems to bring an additional risk in terms of hearing loss.

Two stage EDTA anti-calcification method for bioprosthetic heart valve materials.

BACKGROUND: The aim of this study was to investigate the effect of two-stage EDTA treatment in diminishing calcific degeneration in bovine pericardial bioprosthetic heart valve material. MATERIAL/METHODS: Conventionally preserved pericardium specimens were divided into two groups. Group I (controls, n=18) pieces were first fixed in phosphate-buffered solution (PBS)+0.6% glutaraldehyde at +4 degrees C for 24 hours, then stored in PBS+0.2% glutaraldehyde at room temperature for 6 days. Group II (study group, n=18) pieces were treated with PBS containing 100 microg/ml ethylenediaminetetraacetic acid (EDTA) at +4 degrees C for 24 hours, then fixed in PBS+0.6% glutaraldehyde as was group I at +4 degrees C for 24 hours. After a second exposure to PBS containing 100 microg/ml EDTA at room temperature for 24 hours, they were stored in PBS+0.2% glutaraldehyde at room temperature for 4 days. Pericardial patches were inserted into the dorsal pouches of 18 juvenile male Wistar rats. After 7 weeks of implantation, all the pericardium pieces were harvested from sacrificed rats. The calcium content and biomechanical properties of the explanted tissues were evaluated and also examined histopathologically. RESULTS: The difference in the calcium content of the control and study groups was statistically significant. Biomechanical and histopathologic assessment also supported these findings. CONCLUSIONS: Application of two-stage EDTA was found to be useful in the attenuation of calcification in bioprosthetic heart valve materials with mildly increased durability. As calcification was reduced by approximately 50%, it can be considered for use with other agents as an adjuvant treatment.

Comparison and evaluation of experimental mediastinitis models: precolonized foreign body implants and bacterial suspension inoculation seems promising.

BACKGROUND: Post-sternotomy mediastinitis (PSM) is a devastating surgical complication affecting 1-3% of patients that undergo cardiac surgery. Staphylococcus aureus is one of the most commonly encountered bacterial pathogen cultured from mediastinal samples obtained from patients with PSM. A component of the membrane of the gram positive bacteria, lipoteichoic acid, stimulates the blood monocytes and macrophages to secrete cytokines, radicals and nitrogen species leading to oxido-inflammatory damage. This seems to be responsible for the high mortality rate in PSM. For the evaluation of the pathogenesis of infection or for the investigation of alternative treatment models in infection, no standard model of mediastinitis seems to be available. In this study, we evaluated four mediastinitis models in rats. METHODS: The rats were divided into four groups to form different infection models. Group A: A suspension of 1 x 107 colony-forming units Staphylococcus aureus in 0,5 mL was inoculated from the right second intercostal space into the mediastinum. Group B: A hole was created in the right second intercostal space and a piece of stainless-steel implant with a length of 0.5 cm was inserted into the mediastinum and a suspension of 1 x 107 cfu bacteria in 0,5 mL was administered via the tail vein. Group C: Precolonized stainless-steel implant was inserted into the mediastinum. Group D: Precolonized stainless-steel implant was inserted into the mediastinum and the bacteria suspension was also injected into the mediastinum. On the 10th day, rats were sacrificed and the extension of infection in the mediastenae was evaluated by quantitative cultures. Myeloperoxidase activity (MPO) and malondialdehyde (MDA) levels were determined in the sera to evaluate the neutrophil activation and assess the inflammatory oxidation. RESULTS: The degree of infection in group C and D were 83.3% and 100% respectively (P < 0.001). MDA levels were significantly higher in these two groups than the others (P < 0.001). CONCLUSION: Infected implants and high bacterial concentration administration were the two important components that played a significant role in the outcome of a successful infection in mediastinum in a rat model.

Caffeic acid phenethyl ester (CAPE) supplemented St. Thomas' hospital cardioplegic solution improves the antioxidant defense system of rat myocardium during ischemia-reperfusion injury.

BACKGROUND AND AIM OF STUDY: Cardioplegic arrest remains the method of choice for myocardial protection in cardiac surgery. Caffeic acid phenethyl ester (CAPE) prevents lipid peroxidation induced by ischemia-reperfusion injury and has a potent antioxidant property. We investigated the advantages of CAPE supplemented cardioplegic solution (St. Thomas' Hospital cardioplegic solution No.: 2) on the antioxidant defense system of myocardium against ischemia-reperfusion injury. MATERIAL AND METHODS: Isolated rat hearts were mounted on a nonrecirculating type of Langendorff apparatus. The hearts were arrested for 60 min with cardioplegic solution given at 20-min intervals and then reperfused for 15 min. The hearts were divided into three groups. Cold saline (0.9%, 4 degrees C) in group 1, St. Thomas' Hospital solution in group 2 and CAPE added St. Thomas' Hospital solution in group 3 were used as the cardioplegic solution. Krebs-Henseleit buffer solution was used for reperfusion. The tissues were examined biochemically for oxidative stress. RESULTS: Significant differences among the three groups existed in tissue myeloperoxidase (MPO), catalase (CAT), Na+-K+ ATPase activity and in the concentrations of malonydealdehyde (MDA) and 3-nitrotyrosine (3-NT). Group 2 showed significant changes in MPO (P = 0.04), Na+-K+ ATPase enzyme activity (P = 0.02) and the levels of MDA (P = 0.004) and 3-NT (P = 0.01) in comparison with group 1. Group 3 efficiently reduced MDA levels (P = 0.004) and also led to significant decrease in levels of MPO (P = 0.006), 3-NT (P = 0.01) and Na+-K+ ATPase activity (P = 0.01) and increase in the level of CAT (P = 0.004) in comparison with group 1. Significant changes were also found in the levels of MDA (P = 0.03), MPO (P = 0.04) and CAT (P = 0.009) in comparison between groups 2 and 3. CONCLUSIONS: We demonstrated that the administration of CAPE into cardioplegic solutions improves the antioxidant defense system of rat heart during the ischemia-reperfusion injury.

Leaflet escape in a TRI bileaflet rotatable mitral valve.

Acute prosthetic valve dysfunction is a critical condition for any patient, and is associated with a high mortality. A 24-year-old man who had undergone mitral valve replacement with a TRI bileaflet valve four months previously at another center was admitted with acute-onset left ventricular failure. Echocardiography showed massive mitral insufficiency which was suggestive of a stuck valve. Emergency surgery was carried out, at which the cranial leaflet was found to be stuck open. There was no tissue impingement and thrombosis, the caudal leaflet was absent, and there were no signs of endocarditis or pannus formation. The TRI valve was removed and a replacement 25 mm bileaflet mechanical valve inserted. The embolized leaflet was found in the terminal aorta, but the patient died on day 66 after surgery due to sepsis which had developed from aspiration pneumonia. This is the first report of leaflet escape and terminal aortic embolization with the TRI bileaflet rotatable mitral valve. Acute deterioration of a patient with a prosthetic heart valve should suggest valve dysfunction for which appropriate treatment is rapid relief of the failing left ventricle and replacement of the defective valve with a functioning prosthesis.

Inhibition of calcification with citric acid in pericardial bioprosthetic heart valve material: a preliminary report.

BACKGROUND AND AIM OF THE STUDY: Although current bioprosthetic heart valves have low thrombogenicity and favorable hemodynamic properties, their durability remains unsatisfactory. Valve failure usually occurs from calcific degeneration. The study aim was to investigate the effect of a chelating agent, citric acid (CA), on calcification in bovine pericardium. METHODS: Freshly excised bovine pericardium was dissected free from adhering fat tissue and cut into 1- cm2 pieces; these were rinsed in phosphate-buffered saline solution (PBS), transferred into +4 degrees C PBS containing 0.625% glutaraldehyde (GA) for initial fixation, and then allocated to two groups. Control samples received the same treatment in a fresh solution for 5 days. The other samples underwent an additional fixation step in PBS (pH = 7.4, 37 degrees C) containing 3.8% CA for a period of 48 h (30 ml/g tissue) and were then transferred into freshly prepared PBS + 0.625% GA solution at 37 degrees C for a further 3 days. To investigate calcification rate, pericardial patches were inserted into the dorsal pouches of 15 juvenile male Wistar rats for 42 days. Tissue calcium levels were measured with atomic absorption spectrophotometer, and also assessed histopathologically. RESULTS: The calcium content of CA-treated pericardium was significantly lower than that of controls (66.4 +/- 33.5 and 111.4 +/- 27.2 mg/g, respectively; p = 0.000). In general, the degree of calcification in histological sections agreed well with results of the chemical analyses. Control pericardial tissues showed moderate to severe solid mineral depositions, predominantly parallel to the implant surface, whereas only minor traces of calcium were found in CA-treated tissues. CONCLUSION: These preliminary data suggest that calcific degeneration in bovine pericardium may be reduced by using CA as a chelating agent.

The effect of ethylenediaminetetraacetic acid on calcific degeneration in bovine pericardium.

Calcification is the most frequent cause of the clinical failure of bovine pericardium bioprosthetic valves, preventing their widespread application for surgical treatment. The aim of this study was to minimize calcific degeneration in bovine pericardium by using a chelating agent, ethylenediaminetetraacetic acid (EDTA). Freshly excised bovine pericardium was dissected free from adhering fat tissue and cut into 1-cm(2) pieces that were rinsed in phosphate-buffered saline solution (PBS) and transferred into 4 degrees C PBS containing 1% glutaraldehyde (GA) for initial fixation, then allocated into two groups. Group I received the same treatment in a fresh solution for 5 more days. Group II underwent an additional fixation step in PBS solution (pH 7.4, 37 degrees C) containing 11% EDTA for a period of 48 h (30 ml/g tissue) and was then transferred into freshly prepared PBS + 1% GA solution at 37 degrees C for another 3 days. To investigate the calcification rate, pericardial patches were inserted into the dorsal pouches of 25 male Wistar rats for 21 days. Calcium levels were measured with an atomic absorption spectrophotometer and examined histo-pathologically. The calcium content of EDTA-treated pericardium (Group II), 21 +/- 3.8 microg/mg, was significantly lower than that of Group I, 43.3 +/- 9.2 microg/mg. Assessment of the degree of calcification in the histological sections generally agreed well with the results of the chemical analyses. Calcium deposition in Group I samples were found to be solid mineral depositions, whereas in the Group II pericardial samples, only smaller traces of calcium were found. Calcific degeneration in bovine pericardium can be reduced by using chelates such as EDTA.

N-acetylcysteine for preventing pump-induced oxidoinflammatory response during cardiopulmonary bypass.

PURPOSE: To investigate the effect of N-acetylcysteine on preventing pump-induced oxidoinflammatory response during cardiopulmonary bypass (CPB). METHODS: Forty patients undergoing coronary artery bypass grafting (CABG) were randomly divided into a study group (n = 20), given 50 mg kg(-1) N-acetylcysteine intravenously for 3 days, and a control group (n = 20) given saline. Serum samples were collected for measurement of myeloperoxidase (MPO), malondialdehyde (MDA), interleukin-6, Alpha1-acid glycoprotein (AAGP), and C-reactive protein (CRP) during surgery and postoperatively. RESULTS: The MPO and MDA values showed a similar pattern during and after CPB in the study group, with significantly less variance than in the control group. Interleukin-6 showed similar patterns in the two groups, but the data from 30 min after the start of CPB and from 6 h post-CPB were significantly different. The AAGP and CRP values were both elevated during CPB in the two groups without a significant difference, but 6 and 24 h post-CPB, the values were significantly higher in the control group than in the study group. CONCLUSIONS: N-Acetylcysteine decreased pump-induced oxidoinflammatory response during CPB, suggesting that it could be a novel therapy for assisting in the prevention of CBP-induced oxidoinflammatory damage.

The protective effect of N-acetylcysteine on apoptotic lung injury in cecal ligation and puncture-induced sepsis model.

Apoptotic loss of parenchymal cells may lead to organ dysfunctions in critically ill patients with septic states. As an antioxidant, the protective effects of N-acetylcysteine (NAC) are documented in many experimental and clinical studies. In this experimental study, we investigated the role of chronically used NAC in septic lung injury on a cecal ligation and puncture (CLP) model. To evaluate this, 30 male Wistar rats were randomly divided into four groups as sham (n = 7), CLP (n = 8), sham + NAC (n = 7) and CLP + NAC (n = 8) groups. NAC was administered 150 mg kg(-1) day through intramuscular route beginning 6 h after the operations and lasting for a period of 1 week. One week later, histopathology and epithelial apoptosis were assessed by hematoxylin-eosin and immunohistochemically by M30 and caspase 3 staining to demonstrate septic lung injury. Additionally, lung tissue myeloperoxidase (MPO) activity, malondialdehyde (MDA), and nitrite/nitrate levels were measured. The MPO activity and MDA levels in lung homogenates were found to be increased in CLP group and the administration of NAC prevented their increase significantly (P < 0.05). However, there were no significant differences among the groups regarding nitrite/nitrate levels. The number of apoptotic cells was significantly lower in CLP+NAC group than CLP group, and this finding was supported by M30 and caspase 3 expression in lung (P < 0.05). Lung histopathology was also protected by NAC in CLP-induced sepsis. In conclusion, the chronic use of NAC inhibited MPO activity and lipid peroxidation, which resulted in reduction of apoptosis in lung in this CLP model. Because lung tissue nitrite/nitrate levels did not change significantly, organs other than the lungs may be responsible for producing the increased nitric oxide during sepsis. The chronic use of NAC needs further investigation for its possible antiapoptotic potential in septic states besides its documented antioxidant and antiinflammatory effects.

3-Nitrotyrosine in atherosclerotic blood vessels.

The effect of peroxynitrite on the development of atherosclerosis is one of the major foci of recent studies. Here, the cytotoxic effect of peroxynitrite was investigated by quantitatively measuring nitrated tyrosine, 3-nitrotyrosine (3-NT) levels in atherosclerotic blood vessels. Atherosclerotic vessels were obtained from the patients who underwent either coronary artery or peripheric artery bypass surgery. Internal thoracic arteries of the patients were treated as non-atherosclerotic control vessels. 3-NT was measured by reverse-phase HPLC and plasma nitrite-nitrate levels were measured by spectrophotometry. 3-NT levels were significantly elevated in atherosclerotic vessels (46.6 +/- 23.3 nmol/mg protein, n = 15; p < 0.001) in comparison to control vessels (15.8 +/- 2.5 nmol/mg protein, n = 10). Vessel 3-NT correlated weakly with plasma nitrate levels (r = 0.38). Thus, atherosclerotic arteries have higher 3-NT levels than non-atherosclerotic blood vessels.

Decreased serum total antioxidant status and erythrocyte-reduced glutathione levels are associated with increased serum malondialdehyde in atherosclerotic patients.

BACKGROUND: Coronary artery disease is the significant cause of morbidity and mortality today. The treatment of coronary artery disease is improving, but its prevalence is increasing. Both primary and secondary prevention measures are of vital importance. METHODS: In this study, vitamin C, total antioxidant status, malondialdehyde in serum and erythrocyte-reduced glutathione levels were investigated in patients with atherosclerosis and compared with those of controls. Levels of serum MDA, vitamin C, total antioxidant status, and erythrocyte-reduced glutathione were determined according to the methods of Yagi, Bauer et al., Miller et al., and Beutler, respectively. RESULTS: Erythrocyte-reduced glutathione, serum vitamin C, total antioxidant status, and malondialdehyde values of both patients with atherosclerosis and controls were as follows: 2.80 +/- 0.76, 5.82 +/- 0.67 micromol GSH/g Hb; 1.00 +/- 0.19, 1.62 +/- 0.30 mg/dL; 0.86 +/- 0.14, 1.43 +/- 0.16 mmol/L, and 4.26 +/- 0.9, 1.02 +/- 0.80 nmol/mL, respectively. There was a decrease in the levels of serum vitamin C, erythrocyte-reduced glutathione, and total antioxidant status (p <0.001), and increase in the levels of serum malondialdehyde (p <0.001) in patients with atherosclerosis when compared with those of controls. CONCLUSIONS: Because treatment of atherosclerosis is improving, our results suggest that antioxidant agents may have preventive roles in the formation of atherosclerosis.