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Mouse cells expressing the human complement regulatory proteins decay accelerating factor (DAF) or membrane cofactor protein (MCP) were produced both by hybridoma technology and by transfection with the appropriate cDNAs. The expression of either or both of these products protected the mouse cell from lysis by human (though not rabbit) complement in the presence of naturally occurring human anti-mouse antibody. This effect could be abrogated by the addition of monoclonal antibody against DAF or MCP. These data suggested that the production of animals transgenic for human complement regulatory proteins should in principle be similarly protected from hyperacute xenograft rejection.

It is clear that it is the events which occur within an operating theatre themselves which are the primary determinants of success following a cardiac surgical procedure performed with the use of cardiopulmonary bypass. It is also clear, however, that with greater understanding of the pathophysiological changes occurring after intracardiac procedures are performed the optimal management of such patients has now become a complex and often a multidisciplinary endeavour. Special problems are created after such operations from a combination of the basic cardiac disease, the trauma of the operation, the repair itself, the body’s response to extracorporeal circulatory support, profound hypothermia and from circulatory arrest.

Hyperacute discordant xenograft rejection can be simulated by blood perfused working isolated heart. The survival of the heart is dependent on its functional integrity, and the preparation is thus sensitive to early myocardial damage. Perfusion of rabbit hearts with human blood results in immediate graft destruction by a thrombotic process. Prevention of this process results in rapid rejection at about 20 min by the alternative pathway of complement.

Hyperacute discordant xenograft rejection can be simulated by a blood-perfused working isolated heart. The survival of the heart is dependent on its functional integrity, and the preparation is thus sensitive to early myocardial damage. Perfusion of rabbit hearts with human blood produces immediate graft destruction by a thrombotic process which is a distinct component of hyperacute rejection.

Acute cardiac rejection remains an important cause of death during the first year following cardiac transplantation. Right ventricular biopsies at regular intervals are the main method of detecting rejection, although it is invasive. Unfortunately, non-invasive methods of detecting rejection have not proved reliable enough to replace cardiac biopsies. We assessed the usefulness of technetium-99m 2-methoxy-isobutyl-isonitrile (99mTc-Sestamibi) perfusion scanning in detecting acute rejection in 12 human orthotopic cardiac transplant recipients. Rest and exercise studies and right ventricular biopsies were performed on two occasions. Isotopic evidence of rejection was defined as a perfusion abnormality on either the resting or exercise studies. 99mTc-Sestamibi studies successfully identified acute rejection in 8 of 11 rejection episodes (p less than 0.04). The calculated sensitivity and specificity of 99mTc-Sestamibi scanning were both 72%. Perfusion scanning with 99mTc-Sestamibi may be useful in the diagnosis of acute rejection in cardiac transplant recipients.

Investigation of hyperacute rejection of discordant xenografts has been hampered by the lack of a model for the study of rapid time course events. In vivo models are unsuitable for observation of early rejection processes and, along with most ex-vivo perfusion preparations, are insensitive since no functional demand is placed on the organ which may have undergone extensive damage whilst still appearing viable. For this reason a blood perfused isolated working heart preparation has been developed. With left atrial and left ventricular loading the heart performs measurable work as it ejects into a mock circulation with both afterload and compliance components. When cardiac function is compromised the heart is no longer able to eject against the fixed afterload and both cardiac output and coronary circulation cease with resultant organ failure. The model is thus highly sensitive to minimal organ damage and has an easily identifiable endpoint. In the present study, we used this preparation to study the discordant species combination of rabbit hearts perfused with pig blood.

The functional effects of coronary occlusive disease (COD) in cardiac transplant patients on small-resistance coronary vessels are unclear. We investigated the changes in coronary flow reserve (CFR) in response to the non-specific smooth muscle vasodilator papaverine. A 3F Doppler probe was inserted into the left anterior descending (LAD) coronary artery in 61 patients following orthotopic heart transplantation. Studies were performed in 57 males and 4 females with a mean age of 46 years (range 20-61 years). The median time from operation was 4 years (range 3 months to 10 years). Coronary blood velocity was measured at rest (RFV) and maximum hyperaemia (PFV) produced by intracoronary papaverine. Coronary flow reserve (CFR) was defined as the ratio of PFV to RFV. Minor lesions in epicardial vessels were found in 23 transplant patients. The mean percentage diameter of the most severe lesion in the coronary tree was 23% SD 3% including 12 lesions in the LAD coronary artery itself (mean 24% SD 4%). Patients with COD had an impaired CFR (2.6 SEM 0.2) compared with normals (3.9 SEM 0.2, P = 0.0003), adjusting for year after operation. Mean resting flow velocity was similar in both groups (minor COD, 6.8 cm/s SEM 1.2; normals, 7.1 cm/s SEM 0.6), but mean peak flow velocity response to papverine was reduced (16.5 cm/s SEM 2.5 versus 27.3 cm/s SEM 2.6; P = 0.007). In the presence of minor epicardial disease, coronary flow reserve in resistance vessels was reduced due to impairment of peak flow. This demonstrates that non-endothelial-dependent coronary resistance vessel vasodilatation is abnormal and may be caused by a defect in vascular smooth muscle function.

Prolonged survival of concordant organ xenografts as typified by hamster-to-rat heart transplants is difficult to produce. Studies have revealed that T cells are not primarily involved in rejecting such xenografts and that the rat recipients produce high titres of lytic anti-hamster antibodies. In this study, 200 hamster-to-rat cardiac xenografts performed in 30 different experiments revealed that cyclophosphamide (CyP) and cyclosporin A (CyA) could inhibit this antibody production. CyP alone was relatively ineffective in prolonging graft survival (the median survival time was 14 days versus 3 days in untreated controls). Combining CyP and CyA virtually abolished rejection in this model. Four critically timed doses of CyP combined with continuous CyA resulted in recipients not producing anti-hamster antibodies, despite cessation of CyP therapy, and prolonged graft survival time (median survival time was more than 100 days). Cessation of CyA at 60 and 100 days resulted in the rejection of the xenografts and the appearance of the rat anti-hamster antibodies. Xenografts in recipients given only one or two doses of CyP (and continuous CyA) had a median survival time of 7 and 12 days respectively. However xenograft rejection in rats given only 1 or 2 doses of CyP could be averted by complement depletion using a 3-week course of cobra venom factor (CoF) starting on day 4 or day 7 post-transplantation respectively. Discontinuation of CoF after 3 weeks did not result in graft rejection. These results showed that immunosuppressive therapies directed at inhibiting antibody production may be of value in preventing rejection of concordant xenografts. Short-term complement depletion could rescue xenografts from rejection such that rescued grafts appear to be accommodated.

A prospective study of 219 bronchoscopies in 54 heart-lung and in 2 single lung transplant recipients was undertaken over a 12-month period by a single operator. For histologic study, an average of 17.3 transbronchial biopsy specimens (range, 6 to 56) were taken from three lobes (or from two lobes and lingula of one lung). A further two specimens were taken for culture. The average procedure time was 14.4 minutes (SE 0.31). An estimate of the probability of rejection being missed, depending on the number of specimens taken and based on the method of Gilman and Wang, suggests 18 biopsy specimens are required to have 95% confidence of diagnosing rejection. Sensitivity for diagnosing rejection by histologic study of transbronchial biopsy specimens was 94%, and specificity was 90%. The simple grading of severity of rejection that was used was related both to the number of specimens demonstrating rejection and to the severity of graft airway mucosal inflammation seen at bronchoscopy. The major complication encountered, on 27 occasions, was bleeding of more than 100 ml. On no occasion did bleeding result in any long-term complication. Extensive transbronchial biopsy is a simple, relatively safe, and quick procedure, with a high sensitivity and specificity for diagnosing rejection and lung infection.

The study of endomyocardial biopsy specimens taken in the first 130 days after transplantation has yielded no histologic features predictive of later development of transplant-related coronary artery disease. This study, however, indicated that a combination of the following factors might be predictive in cyclosporine-treated patients: untreated histologically proven episodes of rejection, infection with cytomegalovirus or reactivation of infection, ischemic heart disease in the recipient as the reason for heart transplantation, and possibly HLA-B5 or -B8 mismatch.

Sixty patients underwent heart-lung transplantation at our institution between April 1984 and March 1989. The first five donor organs were removed in an adjacent operating room. Subsequently, organs were removed from distant centers. The method of preservation consisted of cold crystalloid cardioplegic arrest of the heart followed by a cold colloid pulmonary artery flush of a perfusate developed at Papworth Hospital. The lung perfusate is preceded by an infusion of prostacyclin into the pulmonary artery, during preliminary dissection of the donor organs. The total ischemic time ranged from 48 to 51 minutes (mean, 49.6 minutes) for the near-procurement group and from 70 to 249 minutes (mean, 157.6 minutes) for the distant-procurement group. Function of the lungs was assessed by gas exchange, pulmonary function tests, extubation, and survival data. Serial x-ray films were used to monitor graft performance in the postoperative period. We record our clinical experience of early graft function after heart-lung transplantation.

Three hundred-thirteen heart transplant procedures were performed in 303 patients in the 10 years between January 1979 and December 1988. The incidence, histologic type, and site of malignant disease have been analyzed in the 275 patients (91%) who survived more than 1 month after heart transplantation. Four immunosuppressive regimens have been used during this period. Patients 1 through 29 received azathioprine and steroids with 28 days of intravenous antithymocyte globulin. Patients 30 through 89 received cyclosporine and low-dose steroids; the next 61 patients were randomized between cyclosporine and low-dose steroids and cyclosporine and azathioprine; and after patient 150 we have used triple therapy. All cyclosporine-based regimens have included a short course of prophylactic antithymocyte globulin. Eleven malignant tumors have been diagnosed, between 2 months and 6 years after the transplant procedure, and these have been treated by a variety of means. These tumors were two squamous cell carcinomas of the skin; a malignant anal wart that recurred after excision and radiotherapy and necessitated abdominoperineal resection: two small-cell carcinomas of the lung: one squamous cell carcinoma of the esophagus and one of the larynx; two carcinomas of the kidney were found at postmortem examinations; and there were two malignant lymphomas, one of which disseminated rapidly, whereas the other has had prolonged remissions with reduction in immunotherapy with acyclovir. These 11 tumors were responsible for four deaths. No relationship has been demonstrated between type of immunosuppression and tumor development. All patients who are immunosuppressed remain at increased risk for malignant changes. Close surveillance is needed to detect tumors at an early stage.

Chronic rejection is the major cause of death of long-term survivors of heart-lung transplantation. Of our 61 patients who have received heart-lung transplants, 29 have been followed up for a year or longer. Seven patients had clinical evidence of chronic rejection within 15 months of transplantations of these seven, four died with postmortem confirmation of extensive obliterative bronchiolitis, interstitial and pleural fibrosis, and vascular sclerosis in the heart and lungs. All seven patients had evidence on transbronchial biopsy specimens of submucosal fibrosis and vascular sclerosis. Twelve of our remaining patients have shown similar areas of lung fibrosis on transbronchial biopsy specimens, and the other 10 are well and without fibrosis seen on transbronchial biopsy specimens. Studies of the 201 biopsy specimens obtained from 29 patients confirmed rejection on 130 occasions, with more frequent, more persistent, and more severe rejection in the chronic-rejection group than in the without-fibrosis or lung-fibrosis group. Opportunistic infections resulted in pneumonia on 19 occasions, and these were most commonly found in patients with lung fibrosis. We conclude that chronic lung rejection is the likely outcome in patients with early, poorly controlled, severe rejection.

Cross-correlation of cyclosporine concentrations with results of biochemical tests of renal and liver function, measured during the first three months post-operatively, was carried out retrospectively in 24 heart and eight heart-lung transplant recipients to assess the temporal relationship between cyclosporine treatment and the development of possible toxic side-effects. We found a statistically significant negative correlation (95% confidence interval of the mean correlation coefficient did not overlap zero) between the five-day mean concentration of cyclosporine in whole blood (but not plasma) as measured with nonselective (NSRIA) and selective radioimmunoassays (SRIA) and the mean reciprocal creatinine concentration measured in the subsequent five days. In 15 of 32 (47%) patients the negative correlation coefficient exceeded 0.7 (high susceptibility), whereas in 11 of 32 (34%) it was between 0.5 and 0.7 (medium susceptibility), and in only six of 32 (19%) was it less than 0.3 (low susceptibility). We found no consistent correlations between cyclosporine measurements and results of other renal-function tests or liver-function tests. This suggests that therapeutic doses of the drug are not hepatotoxic in most patients. There was, however, a significantly correlated decrease in the NSRIA/SRIA ratio and in serum bilirubin concentration with time post-operatively, reflecting improvement in hepatic function and clearance of the cyclosporine metabolites that are detected by NSRIA. Assays of cyclosporine in whole blood, but not in plasma, are of value in anticipating changes in renal function after heart and heart-lung transplantation.

Endothelium-dependent vasorelaxation mediated by endothelium-derived relaxing factors (EDRF) has been extensively studied in animals but only limited studies in man are available. Demonstration of EDRF-mediated dilatation of human vessels is fundamental for understanding the mechanisms of vascular diseases in man. We have investigated endothelium-dependent relaxation of isolated human pulmonary arteries. Vascular segments, taken from uninvolved regions of resected lung from eight patients undergoing lobectomy for lung carcinoma, were cut into rings. In rings precontracted with phenylephrine, both acetylcholine (ACh) and adenosine diphosphate (ADP) induced dose-dependent relaxation in the presence of endothelium but not when the endothelium had been carefully removed. The rings without endothelium relaxed completely with sodium nitroprusside, a vasodilator agent acting directly on vascular smooth muscle. Pre-incubation with indomethacin, a cyclo-oxygenase inhibitor which blocks production of prostacyclin, did not alter the vasorelaxant responses to ACh and ADP, suggesting that one (or several) non-prostanoid EDRF(s) are responsible for the endothelium-dependent relaxation of isolated human pulmonary arteries.

Twelve patients with severe primary pulmonary hypertension and pronounced disability unresponsive to oral vasolidators were assessed as to their suitability for heart-lung transplantation. During routine right heart catheterisation, intravenous infusions of both prostacyclin (PG1(2] and its synthetic analogue, iloprost (IL), were given in random order. The dose of each drug was increased whilst the improvements in cardiac output (Q) and pulmonary vascular resistance (PVR) were recorded. The maximum rate of each infusion was determined by the onset of side effects or a fall in mean systemic blood pressure of more than 20 per cent. The mean maximum dose of PG1(2) was 6 +/- 2 ng/kg/min and that of IL 3.4 +/- 1.8 ng/kg/min, and both infusions achieved similar haemodynamic effects. In this open, randomised study, intravenous IL was as effective as prostacyclin in lowering PVR, but with a lower infused dose. Iloprost may have an important role in the acute assessment of patients with primary pulmonary hypertension, and when available for prescription should be assessed as an alterative long-term intravenous vasodilator to prostacyclin.

The histologic changes in transbronchial lung biopsy specimens of heart-lung transplant patients were graded during episodes of acute rejection and when patients were well. Infection was strictly excluded from all episodes studied. Grade of severity of rejection was determined by the magnitude and extent of the inflammatory infiltrate. Biopsy specimens, obtained 1 year after the initial biopsies, were examined for histologic evidence of airway submucosal fibrosis, and each patient's clinical status at this time was recorded. The biopsy material from 22 long-term survivors was studied. On 16 occasions the specimens showed no evidence of rejection (grade 0). Twelve of these sets of specimens were from clinically well patients, and four were from the patients who had clinical evidence of rejection. The other six sets of specimens, from clinically well patients, showed evidence of rejection: three grade 1 and three grade 2. One year later, the clinically well patients had normal biopsy histology without fibrosis and normal lung function. Bronchiolitis obliterans had not developed in any patient. There were 27 episodes of rejection in the 22 patients, of which 23 were confirmed histologically. Eleven sets of specimens had grade 1 acute rejection, eight grade 2, and four grade 3. Three of these patients died, and bronchiolitis obliterans was confirmed at necropsy. Lung fibrosis was more common in specimens taken after 1 year, and lung function was depressed in these patients. The histologic grading of transbronchial lung biopsy material, although still in the early stages of development, provides some predictive value to the long-term outcome of the lung transplant patient, in development of both bronchiolitis obliterans and lesser fibrotic changes.

Triple-drug immunosuppression with cyclosporine, azathioprine, and steroids was introduced at Papworth Hospital in 1986. The management of rejection episodes in the first 40 patients receiving this regimen was reviewed for a minimal follow-up period of 18 months. Routine endomyocardial biopsy specimens were taken and classified into nil, minimal, mild, moderate, and severe rejection categories. Management was dependent on the biopsy result, in addition to the clinical state of the patient. In the early postoperative period (day 0 to 30) 2% of biopsy results showed severe rejection and 27% showed moderate rejection; all but one of the latter patients received augmentation of immunosuppression. In the intermediate period (days 31 to 90) severe rejection was present in 1% and moderate rejection in 36 (31%) of biopsies. Immunosuppression was augmented in 13 symptomatic patients, but in 23 asymptomatic patients additional therapy was withheld and the biopsy was repeated in 5 to 7 days. If moderate rejection was still present (11 cases), immunosuppression was augmented. Later (3 to 12 months) augmentation of immunosuppression was given on only four occasions. When the histologic criteria for diagnosis of moderate rejection are present during the early period after transplantation, we recommend augmented immunosuppression. In cases of later rejection we suggest that the decision be based on the biopsy result and the clinical condition of the patient; however, a prospective controlled trial will be required to confirm that this policy is correct.

1. Using cultured epithelia from sweat glands, derived from both cystic fibrosis and normal subjects, the relationship between amiloride concentration and the inhibition of electrogenic sodium transport was measured, under short circuit conditions. 2. The Kd for amiloride in cultures from normal subjects was 0.64 microM (n = 6) while in cultures derived from CF patients the value was 1.07 microM (n = 4). The values were significantly different (P less than 0.02, ANOVA). 3. In cultures from normal sweat glands, bathed in solutions free of permeable anions (chloride/bicarbonate), the Kd for amiloride rose to a value greater than found in CF tissues (2.3 microM). In CF epithelia subject to the same conditions the abnormally high value increased further, so that in solutions without permeable anions normal and CF cultures behaved similarly. 4. In cultures derived from normal and CF tissues lowering the sodium concentration to 10 mM also lowered the Kd for amiloride, however the shift was greater for CF cultures. 5. Several possible explanations for the results are discussed. The most probable is that the relatively more positive apical membrane potential in CF epithelia opposes the interaction of amiloride with the sodium channel, implying that complex formation is potential sensitive.

Combined heart—lung transplantation has advanced from an innovative surgical technique to a clinical treatment of severe pulmonary vascular disease1, chronic end-stage lung disease2, and cystic fibrosis3. Developments in organ preservation4 (Chapters 7 and 35) and careful selection of recipients and donors, together with active investigation of lung rejection using transbronchial biopsy (TBB), have led to effective use of limited donor resources.

A 39 year old female presented with bilateral pneumothoraces and interstitial shadowing on chest x ray. A diagnosis of lymphangioleiomyomatosis was made following an open lung biopsy. Over the next eight years she developed respiratory failure leading to single lung transplantation but she died in the immediate postoperative period. Necropsy examination and review of the previous open lung biopsy revealed multiple pulmonary metastases from a low grade endometrial stromal sarcoma of the uterus. This case high-lights the importance of an accurate diagnosis before transplantation.

The case history is presented of a patient with common variable immunodeficiency in whom heart lung transplantation has been carried out with success. Transplantation was the only long term therapeutic option in this patient due to the progressive respiratory failure resulting from bronchiectasis, emphysema, and granulomatous lung disease.