Objective To investigate the rat model of cardiac allograft vasculopathy after heart transplantation in rat abdominal cavity. Methods Forty Wistar rats and 40SDrats were divided into control group and experiment group randomly pair-matching. Rat model ofheterotopic heart transplantation was developed. Low doseCyclosporine A were injected into the abdominal cavity in experiment group, while the control group had not received the Cyclosporine A. Transplant hearts were harvested at two weeks and four weeks post-operatively and changes of coronary artery were observed by light microscope. Results There were no alteration of tunica intima of coronary artery in control group at two weeks and four weeks post-transplantation. Tunica intima of coronary artery increased in thickness at two weeks post-transplantation in experiment group and concentric circular change occurred at four weeks post-transplantation. Lumen of coronary artery constricted transparent and cardiac allograft vasculopathy occurred. Conclusion This animal model is reliable of cardiac allograft vasculopathy.
Objective
To investigate how to establish stable mice cervical heart transplantation model.
Methods
Totally, 40 male C57 mice with the age of 6-8 weeks and weight of 19-24 g were randomly divided into recipients and donors (n=20 in each group). Mice cervical heart transplantation model was established by connecting the ascending aorta of donors to the right cervical common artery of recipients through end to side anastmosis and the pulmonary artery of donors to the right external jugular vein of recipients through end to end anastmosis.
Results
More than 95% recipients survived after surgery. Cold ischemia time was 15±5 min, warm ischemia time 23±6 min, and the whole operation took about 55±15 min. The recipients survived more than 30 d with functional heart grafts. Histologically, there was no difference between the heart graft one month after the transplantion and the normal heart.
Conclusion
Cervical heart transplantation of mice model is reliable and feasible, which is easy to monitor the survival condition of heart graft by visual examination and palpation, which will benefit the basic research in transplantation field.
Objective To insure early detection and hence efficient prevention of allograft rejection in transplanted heart, investigate possible applications of NAD(P)H fluorescence components analysis at the level of living cardiac cells to propose new approaches for diagnosis of rejection. Methods NAD(P)H was studied for noninvasive fluorescent probing of the mitochondrial function. Human cardiomyocyte were isolated from one additional endomyocardial biopsy (EMB) of 14 pediatric patients with heart ransplantation. Rat cardiomyocyte (n=5, 13-14 week old) were also isolated by the same approach for human myocytes. Autofluorescence(AF) was recorded in living cardiomyocytes following excitation with 375 nm UVlight and detection by spectrallyresolved time correlated single photon counting (TCSPC), based on the simultaneous measurement of the fluorescence spectra and lifetimes. Rat cardiac cells were divided into four groups: normoxic condition, normoxia with Rotenone, ischemic condition and ischemia with Rotenone. Comparison of cardiomyocyte AF between human and rat; compared kinetics of rat cardiomyocytes AF in normoxic conditions to ischemiamimicking ones, induced at physiological temperatures by reducing cell pH and oxygen content; comparison of cardiomyocyte AF dynamic changes in transplanted pediatric patients presenting either no rejection (R0) or mild rejection (R1). Results We have achieved appropriate isolation of living cardiomyocytes from human biopsies, as well as from rat cardiac tissues and determined their AF. At least a 3-exponential decay with 0.5-0.7ns, 1.9-2.4 ns and 9.0-15.0 ns lifetime pools is necessary to describe human cardiomyocyte AF within 420560 nm spectral range. Rat cardiomyocyte steadystate AF in ischemiamimicking condition was significantly increased when compared normoxic ones (Plt;0.05); application of Rotenone induced a significant increase in AF intensity in ischemic and normoxic condition, however no significant difference between the two groups (Plt;0.05).Human cardiomyocyte AF was found significantly lower in comparison to experimental rat model in the same condition(Plt;0.05). A correlation between changes in steadystate NAD(P)H fluorescence and rejection grades was found when comparison of R1 to R0. R1 showed significantly increased fluorescence intensity (Plt;0.05), without change in the spectra shape, results can be comparable to the effect of ischemiamimic conditions. Conclusion Our studies clearly demonstrated that spectrallyresolved fluorescence spectral analysis coupled to fluorescence lifetime are high sensitive approaches to examine mitochondrial metabolic oxidative state directly in living human cardiomyocytes with good reproducibility. Human cardiomyocytes are more metabolically active than the rat ones, while this activity (and thus ATP production) seems lowered during rejection process. In perspective, the advantage of this method is the possibility of its combination to multiphoton confocal microscopy, which can result in the adaptation of this approach directly to tissue biopsy, as well as in vivo directly via cardiac catheterization without the necessity of cell isolation. This approach provides promising new tool for clinical diagnosis and treatment of allograft rejection, and will enhance our knowledge about cardiomyocyte oxidative metabolism and/or its dysfunction at a cellular level.
Objective To investigate the effect of alltrans retinoic acid (atRA) on proliferative artery disease after heart transplantation. Methods Heterotopic heart transplantation model was established by Ono model with 16 inbred healthy male Wistar rats as donors and 16 SD rats as recipients. The rats were divided into chronic rejection group and atRAtreated group by complete random design, and there were 8 rats in each group. Rats in chronic rejection group were given Cyclosporine A 10 mg/(kg·d) by subcutaneous injection after operation, and those in atRAtreated group were given Cyclosporine A 10 mg/(kg·d) in the same way and atRA 10mg/(kg·d) by gavage. The transplanted hearts of rats were taken out 60 days after the transplantation. HE stain, masson stain and Van Gieson were done to analyze the rejection of transplanted hearts, the degree of vascular stenosis and myocardial fibrosis respectively.Immunohistochemistry was used to test proliferating cell nuclear antigen (PCNA). Results The area of myocardial fibrosis in chronic rejection group was obviously larger than that in atRAtreated group（63.99%±11.91% vs.34.68%±6.34%）, and there was significant difference between two groups（t=8.377，P=0.000）. The index of vascular stenosis in chronic rejection group was higher than that in atRAtreated group（62.86±17.18 vs. 40.10±8.20）. Vascular stenosis in atRAtreated group alleviated significantly, and there was significant difference between two groups（t=3.913, P=0.006）. The PCNA positive cells in chronic rejection group were obviously more than that in atRAtreated group（60.17±17.74 vs. 33.96±8.65）, and there was significant difference between two groups（t=5.387, P≤0.001）. There was a positive correlation between the PCNA positive cell ratio and the index of vascular stenosis（r=0.854, P=0.007）. Conclusion Alltrans retinoic acid can inhibit vascular disease after heart transplantation by cell proliferative pathway.
Abstract: Human leukocyte antigen (HLA) is the key antigen mediating rejection and panel reactive antibody (PRA) represent anti-HLA antibodiesin circulation. HLA typing and PRA testing are carried out generally before organ transplantation. With research on the relationship among HLA, PRA and heart transplantation developing, the value of HLA typing and PRA testing in heart transplantation has received more attention and their clinical using strategy has been improved. This article will review the strategy of HLA typing, the clinical value of HLA typing, time-selection in HLA typing, reason and mechanism of rising PRA, clinical sense of PRA testing and treatment of sensitized patients.
Abstract:Objective To investigate the expression and significance of Voltage-gated Cl channel-3 （ClC-3） in acute cardiac allograft rejection in rats. Methods The model of heterotopic cardiac allograft of SD to Wistar rats was established. The rats were divided into two groups： control group and cyclosporin A（CsA） treated group （CsA group）. Living span of the transplants in eight rats of each group were observed. Allograft samples were harvested separately on the day 1, 3, 5, 7 after operation （n = 6）. The rejection was evaluated by routine pathological examinations. The myocardial apoptosis by terminal deoxylnucleotidyl transferase mediated-dUTP nick end labeling （TUNEL） method and the local expression of ClC-3 were detected by reverse transcriptase polymerase chain reaction （RT-PCR）. Results The allografts survival time was significantly longer in CsA group compared with that in control group （15.4±5.1dvs. 7.6±1.5d, P〈0.05）. There was lesser pathological changes in CsA group than that in control group. The apoptosis index were significantly higher in control group and the expression of ClC-3 was significantly lower（P〈0.05）. CsA could inhibit the rise of apoptosis index and the decrease of the ClC-3 expression. Conclusion The ClC-3 expression is closely related with the severity of myocardial necrosis and apoptosis index, which indicates that ClC-3 plays a very important role in the necrosis and apoptosis during acute cardiac allograft rejection of rat.
Objective
To investigate the effect of N-acetylcysteine (NAC) on the apoptosis during myocardial ischemia reperfusion injury in rats’ heart transplantation, and to explore the possible role of NAC in myocardial apoptosis.
Methods
Sixty healthy male Lewis rats (weighing, 200-220 g) were randomly divided into 3 groups, 20 rats each group (10 donors and 10 recipients). In control group, 1 mL normal saline was infused via inferior vena cava at 30 minutes before donor harvesting; in donor preconditioning group, NAC (300 mg/kg) was infused via inferior vena cava at 30 minutes before donor harvesting, but no treatment in recipients; and in recipient preconditioning group, NAC (300 mg/kg) was infused via inferior vena cava at 30 minutes before recipient transplantation, but no treatment in donors. Heart transplantation was established in each group. Blood was drawn at 6 and 24 hours after reperfusion for analysis of aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH) as markers of graft injury; myocardial tissue was harvested to determine the superoxide dismutase (SOD) and lipid hydroperoxide (LPO) activity at 24 hours after reperfusion and to observe the histology and ultrastructural changes. Graft active Caspase-3 protein expression was measured by immunohistochemistry staining, and apoptosis index (AI) was calculated by TUNEL.
Results
The heart transplantation operation was successfully completed in all groups, and the rats survived to the end of the experiment. The serum levels of AST, ALT, and LDH in donor and recipient preconditioning groups were significantly lower than those in control group at 6 hours after reperfusion (P lt; 0.05); the levels of AST and ALT in donor preconditioning group and the levels of AST and LDH in recipient preconditioning group were significantly lower than those in control group at 24 hours (P lt; 0.05); and no significant difference was found between donor and recipient perconditioning groups (P gt; 0.05). The levels of AST, ALT, and LDH at 24 hours were significantly lower than those at 6 hours in each group (P lt; 0.05) except the level of ALT in recipient preconditioning group (P gt; 0.05). SOD activity and SOD/LPO in donor and recipient preconditioning groups were significantly higher than those in control group (P lt; 0.05), but no significant difference between donor and recipient preconditioning groups (P gt; 0.05); there was no significant difference in LPO activity among 3 groups (P gt; 0.05). Histological staining and transmission electron microscope showed that myocardial injury in recipient preconditioning group was obviously lighter than that in donor preconditioning group and control group. Active Caspase-3 in recipient pretreatment group was significantly higher than that in donor preconditioning group and control group (P lt; 0.05). AI of donor and recipient preconditioning groups was significantly lower than that of control group (P lt; 0.05), but no significant difference was found between donor and recipient preconditioning groups (P gt; 0.05).
Conclusion
NAC can relieve ischemia reperfusion injury in rats’ heart transplantation by improving myocardial SOD content, and reducing active Caspase-3 activity and AI, which has a protective effect on myocardial cell of donor heart.
Objective To analyze the relation between preoperative pulmonary artery pressure(PAP) and postoperative complications in heart transplant patients, and summarize the experience of perioperative management of pulmonary hypertension (PH), to facilitate the early period heart function recovery of postoperative heart transplant patients. Methods A total of 125 orthotopic heart transplant patients were divided into two groups according to preoperative pulmonary arterial systolic pressure(PASP) and pulmonary vascular resistance(PVR), pulmonary [CM(1583mm]hypertension group (n=56): preoperativePASPgt;50 mm Hg or PVRgt;5 Wood·U; control group (n=69): preoperative PASP≤50 mmHg and PVR≤5 Wood·U. Hemodynamics index including preoperative cardiac index (CI)，preoperative and postoperative PVR and PAP were collected by SwanGanz catheter and compared. The extent of postoperative tricuspid regurgitation was evaluated by echocardiography. Postoperative pulmonary hypertension was treated by diuresis，nitrogen oxide inhaling，nitroglycerin and prostacyclin infusion, continuous renal replacement therapy(CRRT)and extracorporeal membrane oxygenation(ECMO). Results All patients survived except one patient in pulmonary hypertension group died of multiorgan failure and severe infection postoperatively in hospital. Acute right ventricular failure occurred postoperatively in 23 patients, 10 patients used ECMO support, 10 patients with acute renal insufficiency were treated with CRRT. 124 patients were followed up for 2.59 months，7 patients died of multiple organ failure, infection and acute rejection in follow-up period, the survivals in both groups have normal PAP, no significant tricuspid regurgitation. No significant difference in cold ischemia time of donor heart, cardiopulmonary bypass(CPB) and circulation support time between both groups; but the patients of pulmonary hypertension group had longer tracheal intubation time in comparison with the patients of control group (65±119 h vs. 32±38 h, t=2.17，P=0.028). Preoperative PASP，mean pulmonary artery pressure(MPAP) and PVR in pulmonary hypertension group were significantly higher than those in control group, CI was lower in pulmonary hypertension group ［PASP 64.30±11.50 mm Hg vs. 35.60±10.20 mm Hg; MPAP 43.20±8.50 mm Hg vs. 24.20±7.20 mm Hg; PVR 4.72±2.26 Wood·U vs. 2.27±1.24 Wood·U; CI 1.93±0.62 L/（min·m2） vs. 2.33±0.56 L/（min·m2）; Plt;0.05］. Postoperative early PASP, MPAP and PVR in pulmonary hypertension group were significantly higher than those in control group (PASP 35.40±5.60 mm Hg vs. 31.10±5.70 mm Hg, MPAP 23.10±3.60 mm Hg vs. 21.00±4.00 mm Hg, PVR 2.46±0.78 Wood·U vs. 1.79±0.62 Wood·U; Plt;0.05). Conclusion Postoperative right heart insuficiency is related to preoperative pulmonary hypertension in heart transplant patients. Donor heart can quickly rehabilitate postoperatively by effectively controlling perioperative pulmonary hypertension with good follow-up results.
Objective To study efficiency and security of the recombinant adenoviralmediated gene transfer to the donor heart during the heart transplantation. Methods A total of 140 healthy male Wistar rats,aged 10 weeks, weighing 200250 g, were equally divided into the donor group and the recipient group, and then 70 rats in the recipient group were randomly andequally divided into 2 subgroups: the gene transfer group and the control group. The rat model of heterotopic heart transplantation(Abdomen)was developed, the donor hearts were removed and their coronary arteries were perfused with 800 μlof the recombinant adenoviral vectors encoding the β-galactosidase gene（Ad-LacZ）. The grafts were stored in the 4℃ cold saline solution for 30 minutes, and then the syngeneic transplant was performed. In the control group, saline of tales doses was perfused. The donor hearts were harvested at 3, 5, 7, 14, and 28days (n=7)after transplantation, and the β-galactosidase activity was assessed by the X-gal staining. At 28 days the major organs of the recipients were tested by the histopathological analysis and the polymerase chain reaction of the adenoviral E1A sequences. Results The successful gene transfer of the βgalactosidase gene was demonstrated in the adenovirus-perfused hearts, with no staining in the control group. The gene expression reached a peak level at 3, 5 and 7 days, and the averaged numbers of the total βgalactosidase positive staining cells per slice were 66.4±23.1, 91.3±32.4 and 68.7±22.7, respectively, with no significant difference between the groups (Pgt;0.05). At 14 days the gene expression gradually declined (32.1±13.9), and the significant difference was found when compared with that at 3, 5 and 7 days (Plt;0.05). At 28 days the cells positive for β-galactosidase were sparse (3.9±3.4), and the gene transfer was significantly less efficient compared with that at 3, 5, 7 and 14 days (Plt;0.05). The major organs of the recipients were not affected seriously at 28 days. No virus spread to other organs in this experimental protocol. Conclusion The ex vivo adenoviralmediated gene transfer intracoronarily to the donor heart during the heart transplantation is feasible and safe.
Objective To report the experiences using orthotopic heart transplantation （HTX） to treat a patient with primary cardiac pheochromocytoma （PCT）. Methods On June 2, 2005, a 48-year-old woman received orthotopic HTX because she suffered from cardiac PCT which can not be resected. The procedure for the recipient was uneventful. The aortic crossclamp time was 95 min, assist circulation time 64 min. Results Twenty days after the operation, the patient was discharged without any events, and serum norepinepherine dropped to 1. 339 ng/ml, and urinary norepinepherine 108μg/24h. Conclusion Orthotopic HTX is an effective treatment for cardiac PCT when it cannot be resected.
