Features of an alternative hemodialysis method using a hemoconcentrator during cardiopulmonary bypass surgeries.

PURPOSE: This study clarified the features of a hemoconcentrator-based, alternative hemodialysis (ALTHD) method that improves the speed of serum potassium (K(+)) concentration adjustments, compared with dilutional ultrafiltration (DUF), during cardiopulmonary bypasses. METHODS: Standardized bovine blood was recirculated (300 ml/min) through an in vitro hemoconcentrator circuit; hematocrit, K(+) and glucose levels were measured at 5-20 min after DUF or ALTHD. We evaluated DUF at dialysis speeds of 50-250 ml/min and ALTHD at speeds of 50-1000 ml/min. RESULTS: ALTHD rapidly corrected K(+) and glucose concentrations at speeds up to 800 ml/min. ALTHD took 8.9 min to reach a K(+) level of 4.5 mmol/L, faster than DUF (12.8 min). The ALTHD efficiency curves plateaued at 600 ml/min. CONCLUSION: ALTHD allowed faster adjustment of electrolyte levels, with peak efficiency at 600 ml/min. ALTHD has possible clinical application if available for potential use during all cardiopulmonary bypass surgeries involving extracorporeal circulation.

Prospects for using a hemoconcentrator as an alternative hemodialysis method in cardiopulmonary bypass surgeries.

OBJECTIVE: Cardioplegic solutions often cause high blood concentrations of potassium. The conventional hemoconcentration circuit was improved to correct electrolyte imbalances through a method involving dilutional ultrafiltration (DUF) and an alternative hemodialysis (ALTHD) method. This study aimed to determine the effectiveness of this ALTHD method. METHODS: Bovine blood was used, in conjunction with a hemoconcentrator, in an experimental hemodialysis (HD) circuit to evaluate an ALTHD method. The effectiveness of the method was determined by electrolyte and hematocrit measurements following the procedure. RESULTS: The ALTHD method corrected electrolyte levels as effectively as DUF and was less affected by dilution than DUF. CONCLUSION: The ALTHD method may provide faster electrolyte adjustments than DUF because its efficiency depends on both the blood and dialysate flow rates. In addition, the ALTHD method is expected to provide increased efficiency. Thus, our DUF/ALTHD circuit-switching method may be clinically useful when rapid electrolyte correction is required.

Evidence that graft coronary arteriosclerosis begins in the early phase after transplantation and progresses without chronic immunoreaction. Histopathological analysis using a retransplantation model.

The histopathological features of chronic rejection and its initiation were assessed using rat heterotopic heart transplantation and retransplantation models. Fully allogeneic or minor, non-MHC antigen-mismatch heart grafts transplanted into recipient rats treated with a short course of FK506 showed long-term survival but developed graft atherosclerosis after 40 days posttransplantation. Retransplantation of allografts back into the original donor strain did not prevent graft atherosclerosis if the grafts had resided in the primary recipient for up to 5 days; residence in the primary allogeneic recipient for less than 4 days did not result in graft atherosclerosis in the secondary recipient. Short-course administration of FK506 did not affect the production of these changes. Graft coronary arteriosclerosis begins between 3 and 5 days posttransplantation and progresses without continuous allogeneic immunological drive. The present findings will provide a new means by which to approach the analysis of development of chronic allograft rejection.

Analysis of rejection mechanism in the rat to mouse cardiac xenotransplantation. Role and characteristics of anti-endothelial cell antibodies.

Recent reports have demonstrated that humoral factors, especially antibodies elicited by xenoantigens, play an important role in the rejection of concordant cardiac xenografts. These induced antibodies, however, have not been well characterized. Therefore, we investigated the rejection mechanism, especially the role of humoral immunological responses in the concordant rat to mouse cardiac xenograft model. Lewis rat hearts transplanted into C3H/HeN mice were rejected in 5-6 days. The essential role of humoral factors in the rejection was demonstrated by histological analysis of the rejected hearts showing interstitial hemorrhage, scant cellular infiltration, and the dense deposition of mouse IgG, IgM, and C3 on the graft endothelial cells. In addition, mice that received hyperimmune serum (serum at the 10th day after transplantation) rejected rat hearts hyperacutely. Flow cytometrical analysis using cultured donor rat coronary endothelial cells demonstrated the xenoreactive antibodies of all subclasses, but especially strong reactivity of IgM and IgG2a in the serum at rejection. These xenoreactive antibodies were produced against not only MHC, but also non-MHC antigens on graft endothelial cells. In vivo depletion of L3T4+ T cells led to the suppression of xenoreactive antibody production and the prolongation of graft survival, indicating that antibody production in this model needs L3T4+ T cell help.

Effects of transfected complement regulatory proteins, MCP, DAF, and MCP/DAE hybrid, on complement-mediated swine endothelial cell lysis.

We established several swine endothelial cell (SEC) lines, expressing human MCP (CD46), DAF (CD55), and MCP/DAF hybrid by transfection of cDNA, and assessed the function of these transfectant molecules on complement-mediated cell lysis as an in vitro hyperacute rejection model of swine to human discordant xenograft. Discordant organ xenografts are hyperacutely rejected by complement activation. Amelioration of complement-mediated lysis by these transfectant molecules was tested in each SEC line by lactate dehydrogenase assay. Naive swine endothelial cells were markedly damaged by human complement mainly via the classical pathway, activating only minimally the alternative pathway of human complement. Both MCP and DAF protected SEC from human complement attack in parallel with the expression density, with DAF being more effective than MCP. The MCP/DAF hybrid was more effective than MCP alone, and as effective as DAF in this system. The results suggest that the transfection of DAF or the MCP/DAF hybrid cDNA into organs to be transplanted could protect against hyperacute rejection.

Initial T-cell activation required for transplant vasculopathy in retransplanted rat cardiac allografts.

BACKGROUND: A precise understanding of immunological mechanisms is needed to prevent transplant vasculopathy. METHODS: The developing process of transplant vasculopathy was investigated by retransplanting rat cardiac allografts and measuring the expressions of 21 different genes inside the retransplanted allografts under nonimmunosuppressive conditions. RESULTS: Significant transplant vasculopathy developed if WKY hearts were grafted to LEW and retransplanted to WKY 5 days after the initial grafting, but it did not in allografts retransplanted 3 days after the initial grafting. The disease did not progress in retransplanted isografts or if nude rats were used as the initial recipients. However, the development of transplant vasculopathy was not affected by changing the second recipients to the F1 progeny of donor x recipient or to nude animals. Among the expressions of 21 different genes observed in allografts at 1, 14, 30, or 60 days after retransplantation, those of T-cell activation-related genes, such as interferon-y and Fas ligand, showed the earliest and the most dramatic difference between 3- and 5-day-retransplanted allografts whereas macrophage/monocyte activation-related genes showed little difference. Furthermore, reverse transcription-polymerase chain reaction analyses of allografts retransplanted to nude animal indicated that T cells of the initial recipient origin survive and remain activated even 60 days after retransplantation. CONCLUSIONS: The T-cell response occurring between 3 and 5 days after grafting was identified as the critical parameter to the disease progression. Once alloreactive T cells enter a graft, they may be able to survive a long period and promote chronic rejection.

Sutureless coronary artery bypass with biologic glued anastomoses: preliminary in vivo and in vitro results.

OBJECTIVE: As heart surgery becomes increasingly focused on minimally invasive techniques, it has become apparent that conventional techniques of anastomosis will need to be severely altered or abandoned. Toward that end, we developed and tested in vitro and in vivo coronary artery bypass graft anastomoses using a biologic glue formulated from bovine albumin and glutaraldehyde. We used a double-balloon catheter as a temporary internal stent to create and seal the anastomosis during gluing. METHODS: Initially, anastomoses were made between cryopreserved human saphenous vein segments and coronary arteries in vitro on 12 intact bovine hearts. A total of 42 anastomoses were created with the catheter system introduced into the distal end of the graft, exiting the back wall, and entering the anterior wall of the coronary artery. Two balloons (one in the graft and one in the coronary artery) held the anastomosis stable while the biologic glue was applied externally and allowed to set for 2 minutes. The balloon catheter was then removed from the end of the graft simulating a side-to-side internal thoracic artery anastomosis. After the graft had been flushed to assure distal end patency, the open end of the graft was clipped, turning the anastomosis into an end-to-side graft. A pressure transducer was then attached to the graft and saline solution forcefully infused. RESULTS: All grafts easily held a pressure of 300 mm Hg; 10 grafts were tested up to 560 mm Hg without leaks. Distal and proximal coronary artery patency was checked by examining flow out of the coronary ostia and by cutting arteries distal to the grafts. All anastomoses were patent on being opened and no glue was seen intraluminally. Subsequently, 3 anastomoses of the left internal thoracic artery to the left anterior descending artery have been constructed in goats, with autopsies at 24 hours, 10 months, and 1 year revealing patent anastomoses. CONCLUSION: A biologic glue and catheter system has been developed that allows a coronary anastomosis with a high bursting strength to be performed. When the system has been further developed and tested, truly minimally invasive heart surgery may be possible.

Active cell migration in retransplanted rat cardiac allografts during the course of chronic rejection.

BACKGROUND: Chronic allograft vasculopathy (CAV) is caused by the infiltration of host immune cells to a graft, but it has been technically difficult to monitor the movements of the cells in graft rejection. METHODS: We used a male-specific gene, SRY, as a marker to investigate the dynamics of host cells in a model of CAV in which immunosuppression was unnecessary and anti-male responses were practically negligible. Fluorescent-based real-time quantitative polymerase chain reaction (PCR) was adapted to estimate the fraction of host cells in a graft by the ratio of SRY to IL-2 gene. Using this technique, we studied the turnover and migration of host cells during the course of CAV progression by retransplanting female allografts from male to female or from female to male rats. RESULTS: We detected histologic CAV 60 days after retransplantation in allografts retransplanted to the F(1) progeny of donor x recipient on the 5th day, but not in those retransplanted on the 3rd day, regardless of the mismatches in the genders. Most of the initial infiltrating cells disappeared rapidly in both cases. The fraction of migrating cells from the second recipient, however, continuously increased in allografts developing CAV, and 60 days after retransplantation exceeded 50%, whereas it stayed at 5% to 15% in those not developing CAV. ED-1-positive macrophages/monocytes were likely candidates for the migrated cells. CONCLUSION: We have developed a simple method to measure the migration of host cells into a graft. This technique was useful, at least in certain rat strains, to investigate the cellular mechanisms of chronic cardiac allograft rejection.

Essential initial immunostimulation in graft coronary arteriosclerosis induction detected by retransplantation technique in rats: the participation of T cell subsets.

Graft coronary arteriosclerosis occurs as chronic rejection after heart transplantation. In the previous studies, we have examined the minimum period of allogeneic stimulation to induce this change, using heterotopic rat heart transplantation and a retransplantation model. Retransplantation of allografts back into the donor strain did not prevent graft arteriosclerosis if the grafts had resided in the primary recipient for up to five days. In this study, the participation of the T cell subset causing graft coronary arteriosclerotic change was assessed using the same model. The transplanted rats in fully allogeneic or non-MHC antigen mismatch combinations were treated with a short-course administration of FK506. The graft was removed and retransplanted into the donor strain rats to escape from further immunological stimulation. CD4+ T cells and/or CD8+ T cells of first recipient rats in both combinations were eliminated by monoclonal antibodies. The grade of arteriosclerosis in the retransplanted hearts was evaluated on a basis of a scale from 0-4 according to the histological appearance of the vessel injury on day 40 after initial engraftment. While neither anti-CD4 nor anti-CD8 monoclonal antibody alone had little effect, the administration of both mAbs reduced this arteriosclerotic change and development. In conclusion, the T cell subsets, CD4+ T cell and CD8+ T cell play a certain role in the induction of the graft coronary arteriosclerotic change.

Multichannel myocardial temperature probe.

We present a new system to measure myocardial temperature at multiple points in myocardium. This system consists of a probe with multiple thermocouples, a data collector, and a personal computer. We have demonstrated a change in distribution of temperature in myocardium during hypothermic cardioplegic arrest.

Right ventricular outflow tract reconstruction using a Goretex membrane monocusp valve in infant animals.

We examined the feasibility of using a polytetrafluoroethylene membrane (goretex) valve and transannular patch (TAP) for right ventricular outflow tract (RVOT) reconstruction in growing animal models. Eleven infant goats (Group A) and 12 infant sheep (Group B) underwent RVOT reconstruction under cardiopulmonary bypass. In Group A, a monocusp valve was constructed of goretex, and the RVOT was roofed over utilizing a TAP of bovine pericardium. In Group B, both a monocusp valve and a TAP were constructed of goretex. Animals were sacrificed at 6 or 12 postoperative months. Two goats in Group A died at 9 days and 4 months postoperatively due to RVOT obstruction caused by thrombus formation. Seven goats in Group A and 11 in Group B showed no pressure gradient across the valve. All animals in Group B revealed mild to moderate valvular insufficiency, but no ventricular deterioration. In contrast to the bovine pericardium, which showed a prominent fibroinflammatory reaction with calcified areas causing TAP shrinkage and RVOT stenosis, there was minimal calcification and inflammatory reaction directed against the goretex valve and TAP. We conclude that goretex can be used as a material from which to create both a monocusp valve and TAP for long-term RVOT reconstruction.

Successful experimental multiorgan transplant from non-heart-beating donors using percutaneous cardiopulmonary support.

Organ procurement from non-heart-beating donors (NHBDs) may expand donor pools. In this study, the method of reanimation of heart, lung, and kidney in NHBDs by percutaneous cardiopulmonary support (PCPS) was evaluated. Thirteen beagles were asphyxiated after being given prostacyclin analogue, verapamil, propranolol, and nafmostat mesilate intravenously. Thirty minutes after cardiac arrest, the body was reperfused by PCPS for 1 hr. PCPS priming fluid contained the four drugs above and KCl. Eight hearts immersed in University of Wisconsin (Madison, WI) solution for 24 hr were transplanted orthotopically using leukocyte depleted blood cardioplegia, five left lungs immersed in modified Collins solution were transplanted orthotopically, and five kidneys immersed in University of Wisconsin solution were transplanted heterotopically. All donor hearts arrested without ventricular fibrillation. All transplanted hearts beat spontaneously, and all animals were weaned from cardiopulmonary bypass without inotropic support. The oxygen and carbon dioxide pressure of pulmonary vein blood in the donor lung were no different from those in the recipient lung. All transplanted kidneys made urine soon after reperfusion. These data suggested that hearts, lungs, and kidneys from NHBDs pretreated with four drugs and reanimated with PCPS can be transplanted successfully and that this method may expand the donor pool.