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Case Presentation A 26-year-old man presented with a 2-week history of productive cough and a 1-year history of effort-related dyspnea. His medical history was significant for hay fever and exertion-triggered asthma. He was not taking medicines regularly but was using inhaled salbutamol as needed. He was an ex-smoker, with a previous history of 2-pack years.

Objectives Our objective was to examine intermediate-term survival and reinterventions in unselected patients, stratified by indication, who received a Freestyle® bioprosthesis as a full aortic root replacement. Methods Data from medical records were retrospectively collected for patients who had aortic root replacement using Freestyle® bioprostheses between 1999 and 2018 at six North-Atlantic centers. Survival status was extracted from national registries and results stratified for indication for surgery. Results We included 1030 implantations in 1008 patients with elective indications for surgery: aneurysm (39.8%), small root (8.3%) and other (13.8%), and urgent/emergent indications: endocarditis (26.7%) and Stanford type A aortic dissection (11.4%). Across indications, 46.3% were non-elective cases and 34.0% were reoperations. Median age was 66.0 years (IQR 58.0-71.8) and median follow-up was 5.0 years (IQR 2.6-7.9). Thirty-day mortality varied from 2.9% to 27.4% depending on indication. Intermediate survival for 90-day survivors with elective indications were not different from the general population standardized by age and sex (p-values 0.95, 0.83 and 0.16 for aneurysms, small roots and other, respectively). In contrast, patients with endocarditis and type A dissection had excess mortality (p-values Objectives: Our objective was to examine intermediate-term survival and reinterventions in unselected patients, stratified according to indication, who received a Freestyle (Medtronic Inc, Minneapolis, Minn) bioprosthesis as a full aortic root replacement. Methods: Data from medical records were retrospectively collected for patients who had aortic root replacement using Freestyle bioprostheses between 1999 and 2018 at 6 North-Atlantic centers. Survival status was extracted from national registries and results stratified according to indication for surgery. Results: We included 1030 implantations in 1008 patients with elective indications for surgery: aneurysm (39.8%), small root (8.3%), and other (13.8%), and urgent/emergent indications: endocarditis (26.7%) and Stanford type A aortic dissection (11.4%). Across indications, 46.3% were nonelective cases and 34.0% were reoperations. Median age was 66.0 (interquartile range, 58.0-71.8) years and median follow-up was 5.0 (interquartile range, 2.6-7.9) years. Thirty-day mortality varied from 2.9% to 27.4% depending on indication. Intermediate survival for 90-day survivors with elective indications were not different from the general population standardized for age and sex (P = .95, 83, and .16 for aneurysms, small roots, and other, respectively). In contrast, patients with endocarditis and type A dissection had excess mortality (P < .001). Freedom from valve reinterventions was 95.0% and 94.4% at 5 and 8 years, respectively. In all, 52 patients (5.2%) underwent reinterventions, most because of endocarditis. Conclusions: At intermediate term follow-up this retrospective study provides further support for the use of the Freestyle bioprosthesis in the real-world setting of diverse, complex, and often high-risk aortic root replacement and suggests that outcome is determined by patient and disease, rather than by prosthesis, characteristics.

Background Postoperative heart failure (PHF) is the main cause for mortality after cardiac surgery but unbiased evaluation of PHF is difficult. We investigated the utility of postoperative NT-proBNP as an objective marker of PHF after coronary artery bypass surgery (CABG). Methods Prospective study on 382 patients undergoing isolated CABG for acute coronary syndrome. NT-proBNP was measured preoperatively, the first (POD1) and third postoperative morning (POD3). A blinded Endpoints Committee used prespecified criteria for PHF. Use of circulatory support was scrutinized. Results After adjusting for confounders PHF was associated with 1.46 times higher NT-proBNP on POD1 (p = 0.002), 1.54 times higher on POD3 (p < 0.0001). In severe PHF, NT-proBNP was 2.18 times higher on POD1 (p = 0.001) and 1.81 times higher on POD3 (p = 0.019). Postoperative change of NT-proBNP was independently associated with PHF (OR 5.12, 95% CI 1.86–14.10, p = 0.002). The use of inotropes and ICU resources increased with incremental quartiles of postoperative NT-proBNP. Conclusions Postoperative NT-proBNP can serve as an objective marker of the severity of postoperative myocardial dysfunction. Due to overlap in individuals, NT-proBNP is useful mainly for comparisons at cohort level. As such, it provides a tool for study purposes when an unbiased assessment of prevention or treatment of PHF is desirable. Trial registration ClinicalTrials.gov Identifier: NCT00489827https://clinicaltrials.gov/ct2/show/NCT00489827?term=glutamics&draw=2&rank=1.

Background Surgical cryothermia and radiofrequency (RF) ablations for atrial fibrillation (AF) seem to result in similar sinus rhythm restoration, but the biochemical consequences of the two methods are unclear. We aimed to compare the biochemical responses to the two ablative methods in concomitant mitral valve surgery (MVS). Methods Sixty mitral valve surgery patients with AF were prospectively included. Forty-one patients planned for ablation were randomized to cryothermia (n = 20) or radiofrequency (n = 21) ablation and 19 served as controls. Markers for myocardial injury, inflammation, cell stress, apoptosis, and heart failure were analyzed pre- and postoperatively at different time points. Results Troponin T and creatine kinase isoenzyme MB (CK-MB) peak levels were significantly higher in the cryothermia group compared with the RF group (12,805 [6140–15,700] vs. 2790 [1880–4180] ng/L; P = 0.002 and 271 [217–357] vs. 79 [66–93] μg/L; P rs = 0.41, P = 0.001) and NT-proBNP (rs = 0.48, P Conclusions Cryoablation results in a larger myocardial injury and possibly more elevated apoptotic activity and cell stress compared with the RF technique. The type of ablation device did not have any significant influence on the postoperative inflammatory response nor on the early postoperative levels of NT-proBNP. Trial registration DOI:10.1186/ISRCTN14454361. Registered 16 November 2014 – Retrospectively registered, http://www.isrctn.com/ISRCTN14454361

Objectives. The predictive value of preoperative N-terminal pro-B-type natriuretic peptide (NT-proBNP) was evaluated in patients with acute coronary syndrome undergoing coronary artery bypass grafting (CABG). Design. As a substudy to a clinical trial 383 patients with acute coronary syndrome undergoing CABG were studied. 17 patients had a concomitant procedure. NT-proBNP was measured immediately preoperatively and evaluated with regard to in-hospital mortality, and severe circulatory failure postoperatively according to prespecified criteria. Follow-up was 3.2 ± 0.9 years. Results. In patients with isolated CABG, receiver operating characteristics (ROC) analysis showed an area under the curve (AUC) of 0.82 for in-hospital mortality and 0.87 for severe circulatory failure respectively with a best cut-off for preoperative NT-proBNP of 1028 ng/L. This cut-off level independently predicted severe circulatory failure. Patients with NT-proBNP < 1028 ng/L had significantly better long-term survival (p = 0.004). Preoperative NT-proBNP was higher in patients with concomitant procedure than isolated CABG (2146 ± 1858 v 887 ± 1635 ng/L; p = 0.0005). In patients with concomitant procedure ROC analysis showed an AUC of 0.93 for severe circulatory failure with a best cut-off for preoperative NT-proBNP of 3145 ng/L. Conclusions. Preoperative NT-proBNP predicted in-hospital mortality, severe circulatory failure postoperatively and long-term survival in patients undergoing surgery for acute coronary syndrome but a higher threshold was found in patients having concomitant procedures. Trial registration:ClinicalTrials.gov identifier: NCT00489827.

Objective The primary aim was to investigate the role of underlying heart disease on preoperative NT-proBNP levels in patients admitted for adult cardiac surgery, after adjusting for the known confounders age, gender, obesity and renal function. The second aim was to investigate the predictive value of preoperative NT-proBNP with regard to severe postoperative heart failure (SPHF) and postoperative mortality. Methods A retrospective cohort study based on preoperative NT-proBNP measurements in an unselected cohort including all patients undergoing first time surgery for coronary artery disease (CAD; n = 2226), aortic stenosis (AS; n = 406) or mitral regurgitation (MR; n = 346) from April 2010 to August 2016 in the southeast region of Sweden (n = 2978). Concomitant procedures were not included, with the exception of Maze or tricuspid valve procedures. Results Preoperative NT-proBNP was 1.67 times (pamp;lt;0.0001) and 1.41 times (pamp;lt;0.0001) higher in patients with AS or MR respectively, than in patients with CAD after adjusting for confounders. NT-proBNP demonstrated significant discrimination with regard to SPHF in CAD (AUC = 0.79, 95% CI 0.73 +/- 0.85, pamp;lt;0.0001), MR (AUC = 0.80, 95% CI 0.72 +/- 0.87, pamp;lt;0.0001) and AS (AUC = 0.66, 95% CI 0.51 +/- 0.81, p = 0.047). In CAD patients NT-proBNP demonstrated significant discrimination with regard to postoperative 30-day or in-hospital mortality (AUC = 0.78; 95% CI 0.71 +/- 0.85, pamp;lt;0.0001). The number of deaths was too few in the AS and MR group to permit analysis. Elevated NT-proBNP emerged as an independent risk factor for SPHF, and postoperative mortality in CAD. Conclusions Patients with AS or MR have higher preoperative NT-proBNP than CAD patients even after adjusting for confounders. The predictive value of NT-proBNP with regard to SPHF was confirmed in CAD and MR patients but was less convincing in AS patients. Funding Agencies|Region Ostergotland [LIO-443891]

Postoperative heart failure remains the major cause of death after cardiac surgery. As N-terminal pro-B-type natriuretic peptide (NT-proBNP) is a predictor for postoperative heart failure, the aim was to evaluate if preoperative NT-proBNP could provide additional prognostic information to the recently launched EuroSCORE II.A total of 365 patients with acute coronary syndrome (ACS) undergoing isolated coronary artery bypass graft (CABG) surgery were studied prospectively. Preoperative NT-proBNP and EuroSCORE II were evaluated with regard to severe circulatory failure after operation according to prespecified criteria. To assess what clinical outcomes are indicated by NT-proBNP levels in different risk categories, the patients were stratified according to EuroSCORE II. Based on receiver operating characteristics analysis, these cohorts were assessed with regard to preoperative NT-proBNP below or above 1028 ng litre(-1). The follow-up time averaged 4.4 (0.7) yr.Preoperative NT-proBNP≥1028 ng litre(-1) [odds ratio (OR) 9.9, 95% confidence interval (CI) 1.01-98.9; P=0.049] and EuroSCORE II (OR 1.24, 95% CI 1.06-1.46; P=0.008) independently predicted severe circulatory failure after operation. In intermediate-risk patients (EuroSCORE II 2.0-10.0), NT-proBNP≥1028 ng litre(-1) was associated with a higher incidence of severe circulatory failure (6.6% vs 0%; P=0.007), renal failure (14.8% vs 5.4%; P=0.03), stroke (6.6% vs 0.7%; P=0.03), longer intensive care unit stay [37 (35) vs 27 (38) h; P=0.002], and worse long-term survival.Combining EuroSCORE II and preoperative NT-proBNP appears to improve risk prediction with regard to severe circulatory failure after isolated CABG for ACS. NT-proBNP may be particularly useful in patients at intermediate risk according to EuroSCORE II.NCT00489827.

In patients with severe aortic valve disease, aortic valve intervention is performed when the risk for mortality or morbidity with conservative management is significantly increased. Left ventricular (LV) longitudinal motion decreases before conventional echocardiographic measures such as ejection fraction, are influenced in patients with severe aortic valve disease. This thesis is devoted to the assessment of cardiac function, including ventricular longitudinal motion, by echocardiography before and after aortic valve intervention in patients with chronic severe aortic regurgitation (AR) or severe aortic stenosis (AS).Patients with chronic AR (n=29) were studied preoperatively, and 6 months and 4 years postoperatively by echocardiography, including tissue Doppler imaging, at rest and during exercise. LV longitudinal function (atrioventricular plane displacement, AVPD, and peak systolic velocity, PSV) decreased postoperatively, and patients with low PSV continued to show reduced longitudinal function 6 months after surgery. Preoperative exercise echocardiographic variables showed a strong correlation to late LV function variables, while preoperative variables obtained at rest were not useful for prediction. Exercise echocardiography and longitudinal LV function could therefore be useful complements in the timing of aortic valve surgery for AR.Patients with chronic AR (n=29) were studied preoperatively, and 6 months and 4 years postoperatively by echocardiography, including tissue Doppler imaging, at rest and during exercise. LV longitudinal function (atrioventricular plane displacement, AVPD, and peak systolic velocity, PSV) decreased postoperatively, and patients with low PSV continued to show reduced longitudinal function 6 months after surgery. Preoperative exercise echocardiographic variables showed a strong correlation to late LV function variables, while preoperative variables obtained at rest were not useful for prediction. Exercise echocardiography and longitudinal LV function could therefore be useful complements in the timing of aortic valve surgery for AR.

OBJECTIVE: The indications for and the risk and benefit of concomitant surgical ablation for atrial fibrillation (AF) have not been fully delineated. Our aim was to survey whether the Cox-maze IV procedure is associated with postoperative heart failure (PHF) or other adverse short-term outcomes after mitral valve surgery (MVS). DESIGN: Consecutive patients with AF undergoing MVS with (n = 50) or without (n = 66) concomitant Cox-maze IV cryoablation were analysed regarding perioperative data and one-year mortality. RESULTS: The patients in the Maze group were younger, were in lower NYHA classes, had better right ventricular function and had lower pulmonary artery pressure. The Maze group had 30 min longer median cross-clamp time (CCT) and 50% had PHF compared with 33% in the No-maze group, p = 0.09. Two patients in the No-maze group died within one year of surgery. Congestive heart failure (OR 4.3 [CI 95%: 1.8-10], p Funding agencies: ALF funding, County Council of Ostergotland, Sweden

Aims Knowledge of longitudinal left and right ventricular (LV and RV) function after transcatheter aortic valve implantation (TAVI) is scarce. We hypothesized that the longitudinal systolic biventricular function in aortic stenosis (AS) patients is affected differently by TAVI and surgical aortic valve replacement (SAVR). less thanbrgreater than less thanbrgreater thanMethods and results Thirty-three AS patients (all-TAVI group, age 81 +/- 9 years, 18 female), with EuroSCORE 18 +/- 9%, were accepted for TAVI. Seventeen of these patients were matched (by gender, age, and LV function) to 17 patients undergoing SAVR. Conventional echocardiographic parameters, systolic atrioventricular plane displacement (AVPD) at standard sites and peak systolic velocity (PSV) by pulsed tissue Doppler at basal RV free wall, LV lateral wall, and septum were studied before and 8 weeks after the procedure. Procedural success was 100%, and 30-day mortality 9%. In all TAVI patients, AVPD(lateral), PSV(lateral), AVPD(septal), and PSV(septal) increased (P andlt; 0.001, 0.003, 0.006 and 0.002). When studying the matched patients postoperatively, both the SAVR and TAVI patients had increased PSV(lateral) and AVPD(lateral) (SAVR: P = 0.03 and P = 0.04, TAVI: P = 0.04 and P = 0.01). The PSV(RV) increased in the all-TAVI group (P = 0.007), while the AVPD(RV) was unchanged. SAVR patients had decreased AVPD(RV) (P = 0.001) and PSV(RV) (P = 0.004), while the matched TAVI patients had unchanged RV function parameters. less thanbrgreater than less thanbrgreater thanConclusion An improvement in regional longitudinal LV function in the septal and lateral wall could be seen after TAVI. Among the matched patients, both the TAVI and SAVR patients seemed to improve LV function in the lateral wall. RV systolic function increased in TAVI patients, but was impaired in the matched SAVR group at the 8-week follow-up. Funding Agencies|Swedish Heart Lung Foundation||Medical Research Council of Southeast Sweden||County Council of Ostergotland, Sweden

Background: Intravenous glutamate reduced the risk of developing severe circulatory failure after isolated coronary artery bypass graft surgery (CABG) for acute coronary syndrome (ACS) in a double-blind randomised clinical trial (GLUTAMICS-ClinicalTrials. gov Identifier:NCT00489827). Here our aim was to study if glutamate was associated with reduced the use of inotropes. Methods: Post-hoc analysis of 824 patients undergoing isolated CABG for ACS in the GLUTAMICS-trial. ICU-records were retrospectively scrutinised including hourly registration of inotropic drug infusion, dosage and total duration during the operation and postoperatively. Results: ICU-records were found for 171 out of 177 patients who received inotropes perioperatively. Only one fourth of the patients treated with inotropes fulfilled study criteria for postoperative heart failure at weaning from cardiopulmonary bypass (CPB) or later in the ICU. Inotropes were mainly given preemptively to facilitate weaning from CPB or to treat postoperative circulatory instability (bleeding, hypovolaemia). Except for a significantly lower use of epinephrine there were only trends towards lower need of other inotropes overall in the glutamate group. In patients treated with inotropes (glutamate n = 17; placebo n = 13) who fulfilled study criteria for left ventricular failure at weaning from CPB the average duration of inotropic treatment (34 +/- 20 v 80 +/- 77 h; p = 0.014) and the number of inotropes used (1.35 +/- 0.6 v 1.85 +/- 0.7; p = 0.047) were lower in the glutamate group. Conclusions: Intravenous glutamate was associated with a minor influence on inotrope use overall in patients undergoing CABG for ACS whereas a considerable and significant reduction was observed in patients with heart failure at weaning from CPB. Funding Agencies|Swedish Heart-Lung Foundation [20030595]; Capio Research Foundation [2005-1021, 2006-1203]; Linkoping University; Ostergotlands Lans Landsting

Objective To evaluate the relevance of the individual components of the Valve Academic Research Consortium (VARC)-2 criteria for periprocedural myocardial infarction (MI) in transcatheter aortic valve implantation (TAVI). The association between biomarkers and adverse procedural outcome has been established. However, the additive prognostic importance of signs and symptoms are more uncertain. Methods A total of 125 consecutive TAVI patients were prospectively included in this study. Biomarkers for MI were analyzed and signs and symptoms according to VARC-2 criteria were collected from clinical records. Results The criteria of elevated biomarkers and of signs or symptoms were found in 27 (22%) and 32 (26%) of the patients, respectively. According to VARC-2 definition, 12 (10%) had MI. VARC-2 definition of MI, Troponin T (TnT) greater than 600 ng/L, and presence of signs or symptoms correlated with 6 months mortality, prolonged ICU stay, elevation of N-terminal prohormone brain natriuretic peptide, and renal impairment. No signs or symptoms were found in 7 (44%) of the patients who fulfilled the criterion of elevated TnT greater than 600 ng/L. In the group with positive TnT criterion, there were no significant differences between those with and without signs or symptoms in respect to levels of TnT (1014 [585-1720] ng/L versus 704 [515-905] ng/L, p = 0.17) or creatine kinase-MB (36 [25-52] mu g/L versus 29 [25-39] mu g/L, p = 0.32). In the multivariate Cox regression analysis, TnT greater than 600 ng/L was the only significant independent variable associated with 6-months postprocedural mortality. Conclusions Myocardial injury in TAVI, measured with biomarkers, correlates well with adverse procedural outcome. In this study it is also the strongest predictor for early postprocedural mortality. The additional requirement of signs or symptoms for the diagnosis of MI results in omission of a considerable number of clinically significant MI. Funding Agencies|Medical Research Council of Southeast Sweden [FORSS-12657, FORSS-23891, FORSS-159851, FORSS-311341]; ALF founding, County Council of Ostergotland, Sweden

Glutamate plays an important role for myocardial metabolism in association with ischaemia. Patients with coronary artery disease characteristically demonstrate increased uptake of glutamate. Improved recovery of myocardial metabolism and haemodynamic state after coronary surgery has been reported in patients treated with glutamate infusion. However, the effect of glutamate has not been studied after other cardiac surgical procedures. In addition, the effects of glutamate on peripheral tissue metabolism remain to be described.Twenty patients undergoing surgery for aortic stenosis were studied after randomisation to blinded infusion of glutamate or saline during 1h immediately after skin closure. Myocardial and leg tissue metabolism were assessed with organ balance techniques.Postoperative glutamate infusion induced a marked increase in myocardial and leg tissue uptake of glutamate. This was associated with a significant uptake of lactate in the heart. The negative arterial-venous differences of amino acids and free fatty acids across the leg were significantly smaller in the glutamate group. Haemodynamic state remained stable and did not differ between groups.The heart and peripheral tissues consumed the exogenously administered glutamate after surgery for aortic stenosis. Potentially favourable effects of glutamate on myocardial and peripheral tissue metabolism are suggested.

Neurological complication remains a feared and increasing problem in association with cardiac surgery. The aim of this study was to analyze risk factors for neurological complications in a cohort of patients in whom inotropes for weaning from cardiopulmonary bypass was gradually replaced by metabolic treatment.The records of 775 consecutive patients undergoing coronary artery bypass grafting (CABG) or combined CABG+valve procedures were examined. Forward stepwise multiple logistic regression analysis was used for statistical evaluation of independent risk factors.The incidence of neurological injury was 1.8% in patients undergoing isolated CABG and 5.4% after combined CABG+valve procedures. After cross-validation multivariate analysis identified history of cerebrovascular disease, advanced age and aortic cross-clamp time as independent risk factors for postoperative cerebral complications. Chronic obstructive pulmonary disease and number of bypasses also emerged as risk factors in the primary analysis.In general, markers for advanced atherosclerosis, with history of cerebrovascular disease as the most important, emerged as predictors for neurological injury. Although it did not enter the final risk model, the results also suggest that postoperative heart failure deserves further surveillance as a potential risk factor for neurological complications. However, no evidence for untoward neurological effects associated with glutamate or glucose-insulin-potassium treatment was found.

We present the results obtained with an in vitro model system that resembles the in vivo tumour micro-environment, where malignant cells are in close contact with the infiltrating lymphocytes. Unmanipulated blood lymphocytes were cytotoxic against the autologous ex vivo tumour cells in 3/19 patients and this function was generated in 6-day mixed cultures in five additional cases. Production of transforming growth factor beta (TGFbeta) by the freshly separated tumour cells was determined in parallel. Cytotoxicity was generated by a small number of tumour cells (2-5/100 lymphocytes), while a large number (10-20/100 lymphocytes) inhibited not only the generation but also the existing lytic activity. The presence of a neutralising TGFbeta-specific mAb (2G7) potentiated the activation of lymphocytes and suspended the suppression inflicted by the tumour cells. In those tumours, which expressed relatively high levels of MHC class I and ICAM-1 molecules, the quantity of secreted TGFbeta interfered with the ability of tumour cells to generate cytotoxic lymphocytes. In the tumours with low expression of class I, such a correlation was not detected, indicating the primordial role of MHC class I expression in the regulation of autologous tumour recognition. Our results demonstrate the involvement of TGFbeta in the impaired lymphocyte-mediated reactivity against immunogenic tumours and support a mechanism that contrasts the tolerance or anergy. Since presence of TGFbeta in the microenvironment of tumours counteracts the function of cytotoxic T lymphocytes, production of this cytokine can contribute to the failure of immunotherapy.

The frequent use of diuretic drugs in cardiac surgical practice contrasts with the lack of documentation regarding diuretic treatment in this setting. The aims of this study were to delineate the need for diuretic drugs in adult cardiac surgical practice and to evaluate the impact of adding a combination of 50 mg hydrochlorothiazide and 5 mg amiloride orally to patients responding poorly to furosemide.Two hundred ten consecutive patients, 159 undergoing coronary artery bypass grafting procedures and 51 having valve operations, were studied.Seventy-seven patients received large doses of furosemide (or = 80 mg/24 h) at some time during the postoperative course, and of these 20 responded poorly to furosemide (weight loss 0.3 +/- 0.2 kg) despite considerable fluid retention. The addition of hydrochlorothiazide and amiloride provided a prompt and effective remedy to relative furosemide resistance. Average weight loss was 2.3 +/- 0.2 kg (p0.01 compared with response to furosemide) and average diuresis was 2,949 +/- 156 mL in the following 24 hours.Relative furosemide resistance is common after cardiac operations. Thiazides, although they are mild diuretic agents, may serve as useful adjuncts in this setting.

Human carcinoma cells vary appreciably in the expression of MHC class I, class II, ICAM-1 (CD54) and B7 (CD80) molecules. Short-term in vitro exposure of ex vivo carcinoma cells to interferon gamma and tumour necrosis factor alpha elevated/induced the surface expression of MHC class I, class II and ICAM-1, but only rarely of B7. We found that cytokine treatment elevated the cytotoxic susceptibility and the stimulatory potential of ex vivo tumour cells. This was demonstrated (a) by the increased frequency and elevated level of auto-tumour lysis and (b) by induction of DNA synthesis and generation of cytotoxic lymphocytes in autologous mixed lymphocyte/tumour cell culture (MLTC). The MHC class I and ICAM-1 molecules on the tumour cells were required for interaction with the lymphocytes as indicated by the inhibitory effect of specific mAb both in the stimulation and in the cytotoxic tests. While the cytokine-induced increases in MHC and ICAM-1 on the low-expression tumours were probably important for the modification of functional interaction with the autologous lymphocytes, it is likely that alterations in other properties of tumour cells were also induced which contributed to the phenomenon. This was indicated by the results obtained with several tumours, which expressed indigenously high levels of these molecules but activated the autologous lymphocytes only after cytokine treatment. In several experiments the untreated targets that did not activate the lymphocytes were sensitive to the cytotoxicity of the effectors activated in MLTC. The results show that the afferent and efferent arms of the immune response have different requirements for functional interactions between lymphocytes and tumour cells.

Although the EuroSCORE was developed for predicting operative mortality after cardiac surgery, it has also been shown to predict long-term mortality. It has been reported that postoperative heart failure (PHF) in association with surgery, albeit comparatively benign in the short term, has a profound impact on five-year survival after surgery for aortic stenosis (AS). The study aim was to determine the combined impact of EuroSCORE and PHF on long-term survival after isolated aortic valve replacement (AVR) for AS.A total of 397 patients (48% females; average age 70 +/- 10 years) who underwent AVR for AS at the authors' institution between 1995 and 2000 was studied. The cohort was subdivided according to the additive EuroSCORE into a high-risk group (EuroSCORE7) and a low-risk group (EuroSCOREor = 7), and further analyzed in relation to PHF.The average follow up was 8.1 years (range: 5.2-11.2 years). Forty-five patients (11%) were treated for procedure-associated PHF. Patients with or without PHF and a high-risk EuroSCORE had crude five-year survivals of 57% and 64%, respectively (p = 0.6), whereas those with or without PHF but with a low-risk EuroSCORE had crude five-year survivals of 58% and 89%, respectively (p = 0.0003).Both PHF and a high EuroSCORE were associated with poor long-term survival. The role of PHF per se for the long-term prognosis was illustrated by the fact that the negative impact on long-term survival was almost as profound in patients of the low-risk group as of the high-risk group.

Objectives. To describe short-term clinical and echocardiography outcomes in patients undergoing transcatheter aortic valve implantation (TAVI) and surgical aortic valve replacement (SAVR). To explore patient selection criteria for treatment with TAVI. Design. TAVI patients (n = 45) were matched to SAVR patients (n = 45) with respect to age within +/- 10 years, sex and systolic left ventricular function. Results. TAVI patients were older, 82 +/- 8 versus 78 +/- 5 years (p = 0.005) and they had higher logEuroSCORE, 16 +/- 11% versus 8 +/- 4% (p andlt; 0.001). There were no significant differences in 30 days mortality, stroke and myocardial infarction. TAVI patients received less erythrocyte (53% vs. 78%, p = 0.03) and thrombocyte (7% vs. 27%, p = 0.02) transfusions. Postoperative atrial fibrillation was less common (18% vs. 60%, p andlt; 0.001) in the TAVI group. Paravalvular regurgitation was more common in TAVI patients (87% vs. 0%, p andlt; 0.001) and 27% had access site complications. Aortic transvalvular velocity was 2.3 +/- 0.4 m/s versus 2.6 +/- 0.5 m/s (p = 0.002) and mean valve pressure gradient was 12 +/- 4 mmHg versus 15 +/- 5 mmHg (p = 0.01) in the TAVI and SAVR groups, respectively. Twenty-nine (64%) of the TAVI patients had logEuroSCORE andlt; 15%. Conclusions. Both TAVI and SAVR have good short term clinical outcome with excellent hemodynamic result. In clinical practice, factors other than high logEuroSCORE play an important role in patient selection for TAVI. Funding Agencies|Swedish Heart and Lung Foundation||Medical Research Council of Southeast Sweden (FORSS)||ALF||County Council of Ostergotland

ObjectiveGlutamate has been claimed to protect the heart from ischemia and to facilitate metabolic and hemodynamic recovery after ischemia. The GLUTAmate for Metabolic Intervention in Coronary Surgery trial investigated whether an intravenous glutamate infusion given in association with surgery for acute coronary syndrome could reduce mortality and prevent or mitigate myocardial injury and postoperative heart failure.MethodsIn the present prospective, triple-center, double-blind study, 861 patients undergoing surgery for acute coronary syndrome were randomly assigned to an intravenous infusion of glutamate (n = 428) or saline (n = 433) perioperatively.ResultsThe incidence of the primary endpoint—a composite of 30-day mortality, perioperative myocardial infarction, and left ventricular heart failure at weaning from cardiopulmonary bypass—was 7.3% versus 5.8% (P = .41) in the glutamate and control groups, respectively. Patients with left ventricular failure at weaning from cardiopulmonary bypass had a shorter median intensive care unit stay (25 vs 92 hours; P = .02) if they were treated with glutamate. In patients with unstable angina (Canadian Cardiovascular Society class IV) undergoing isolated coronary artery bypass grafting (n = 458), the incidence of severe circulatory failure according to the prespecified criteria was significantly lower in the glutamate group (1.3% vs 6.9%; P = .004). On multivariate analysis, glutamate infusion was associated with a reduced risk of developing severe circulatory failure (odds ratio, 0.17; 95% confidence interval, 0.04-0.72; P = .02). A relative risk reduction exceeding 50% for developing severe circulatory failure was seen in most risk groups undergoing isolated coronary artery bypass grafting, with those with diabetes a notable exception.ConclusionsThe primary endpoint did not differ significantly between the groups. The secondary outcomes and post hoc analyses warrant additional studies with regard to the potential beneficial effect of glutamate on postischemic myocardial recovery.

Myocardial ischemia is a major cause of postoperative heart failure and adverse outcome in coronary artery bypass graft surgery (CABG). Conventional treatment of postoperative heart failure with inotropic drugs may aggravate underlying ischemic injury. Glutamate has been claimed to increase myocardial tolerance to ischemia and promote metabolic and hemodynamic recovery after ischemia. The aim of this work was to investigate if intravenous glutamate infusion given in association with CABG for acute coronary syndrome can reduce mortality and prevent or mitigate myocardial injury and postoperative heart failure. We also wanted to assess neurological safety issues, as a concern with the use of glutamate is that it may act as an excitotoxin under certain conditions.A metabolic strategy for perioperative care was assessed in an observational study on 104 consecutive patients with severe left ventricular dysfunction undergoing CABG. Based on encouraging clinical results, unsurpassed in the literature, the GLUTAMICS-trial was initiated. 861 patients undergoing CABG for acute coronary syndrome were randomly allocated to blinded intravenous infusion of L-glutamicacid solution or saline. The primary endpoint was a composite of postoperative mortality (≤30 days), perioperative myocardial infarction and left ventric ular heart failure in association with weaning from cardiopulmonary bypass. Secondary endpoints included neurological safety issues, degree of myocardial injury,postoperative hemodynamic state, use of circulatory support and cardiac mortality.The event rate was lower than anticipated and the primary endpoint did not differ significantly between the groups. Regarding secondary endpoints there were significant differences compatible with a beneficial effect of glutamate on post-ischemic myocardial recovery. The putative effect of glutamate infusion was seen in more ischemic patients (CCS class IV) and in patients with evident or anticipated LV-failure on weaning from CPB. No evidence for increased incidence of clinical or subclinical neurological injury was found. In conclusion, intravenous glutamate infusion is safe in the dosages employed and could provide a novel and important way of promoting myocardial recovery after ischemic injury.

Background. Complications of an inadequate haemodynamic state are a leading cause of morbidity and mortality after cardiac surgery. Unfortunately, commonly used methods to assess haemodynamic status are not well documented with respect to outcome. The aim of this study was to investigate SV(O2) as a prognostic marker for short-and long-term outcome in a large unselected coronary artery bypass grafting (CABG) cohort and in subgroups with or without treatment for intraoperative heart failure. less thanbrgreater than less thanbrgreater thanMethods. Two thousand seven hundred and fifty-five consecutive CABG patients and subgroups comprising 344 patients with and 2411 patients without intraoperative heart failure, respectively, were investigated. SV(O2) was routinely measured on admission to the intensive care unit (ICU). The mean (SD) follow-up was 10.2 (1.5) yr. less thanbrgreater than less thanbrgreater thanResults. The best cut-off for 30 day mortality related to heart failure based on receiver-operating characteristic analysis was SV(O2) 60.1%. Patients with SV(O2) andlt;60% had higher 30 day mortality (5.4% vs 1.0%; P andlt; 0.0001) and lower 5 yr survival (81.4% vs 90.5%; P andlt; 0.0001). The incidences of perioperative myocardial infarction, renal failure, and stroke were also significantly higher, leading to a longer ICU stay. Similar prognostic information was obtained in the subgroups that were admitted to ICU with or without treatment for intraoperative heart failure. In patients admitted to ICU without treatment for intraoperative heart failure and SV(O2) andgt;= 60%, 30 day mortality was 0.5% and 5 yr survival 92.1%. less thanbrgreater than less thanbrgreater thanConclusions. SV(O2) andlt;60% on admission to ICU was related to worse short- and long-term outcome after CABG, regardless of whether the patients were admitted to ICU with or without treatment for intraoperative heart failure. Funding Agencies|Östergotlands Läns Landsting||Linköping University Hospital

Autologous mixed lymphocyte-tumor cell cultures (MLTC) were initiated with cytokine (IFN gamma and TNF alpha)-treated ex-vivo tumor cells of lung, ovarian, breast and stomach carcinomas. The cytokine-treated tumors expressed class-I but not class-II molecules. Although the proportion of CD8+ lymphocytes increased in the bulk culture of MLTCs, in 5/7 experiments the majority of the established T-cell clones were CD4+. Among the CD8+ clones a high proportion (77%) was cytotoxic, while the proliferative response was more frequent among the CD4+ clones (70%). In 4/26 cytotoxic T-lymphocyte (CTL) clones (3/17 CD4+ and 1/9 CD8+), derived from a patient with class I+ class II- stomach carcinoma, lysis was restricted to the autologous tumor cells. These auto-tumor-specific clones did not lyse the autologous ConA blasts, the 5 allogeneic ex-vivo tumors, the NK-sensitive K562 or the relatively sensitive Daudi cells. The cytotoxicity of these clones was inhibited by pre-incubation of the tumor cells with W6/32 (alpha-class I) MAb, or by preincubation of the lymphocytes with OKT3 (alpha-CD3) MAb. The alpha-CD4 (OKT4) MAb had only a marginal effect on the CD4+ clones, while the lytic function of the CD8+ clone was inhibited by the alpha-CD8 (OKT8) MAb. The 3 CD4+ CTL clones also responded with proliferation to the autologous tumor cells. This proliferative response was inhibited by the presence of W6/32 MAb. Our results indicate that the auto-tumor lysis exerted by CD4+ CTL clones was restricted by the class-I antigens, and that the CD4 molecules of the clones were not essential for the lytic interaction.

It has been reported that the assembly of MHC class I molecules in mutagenized cell lines could be induced by specific binding peptides. We have now demonstrated that the defect in assembly between heavy and light chains of class I molecules naturally occurred in tumor cells of one spontaneous ovarian carcinoma detected by one-dimensional isoelectric focusing of immunoprecipitates with anti-monomorphic class I MAb (W6/32) and by immunostaining with free heavy chain and beta 2m-specific MAbs. In vitro treatment of the tumor cells with IFN-gamma induced the assembly and surface expression of majority class I molecules (A2.1, B7, B15, Cw6, Cw7 out of A2.1, A2*, B7, B15, Cw6, Cw7). Moreover, assembly of A2 and Cw6 was induced by exposure of the tumor cells to a HLA A2-binding peptide K62 derived from influenza A matrix protein. Autologous blood T lymphocytes were activated in mixed lymphocyte-tumor cell culture (MLTC) by the IFN-gamma-treated but not by the unmanipulated tumor cells. Although activated lymphocytes damaged both IFN-gamma-treated and untreated tumor cells, the alpha class I MAb (W6/32) efficiently inhibited the lysis of IFN-gamma-treated targets, but not the untreated targets. These results indicate that the defect of MHC class I assembly may result in the escape of tumor cells from immune response.

Concerns have been raised about potential neurological injury related to exogenous glutamate. In cardiac surgery glutamate has been administered as a putative cardioprotective agent by cardioplegia or intravenous infusion. In the GLUTAMICS trial, in addition to surveillance of clinical neurological injuries, a prespecified subgroup was analyzed with regard to postoperative S-100B levels to detect potential subclinical neurological injury related to glutamate infusion.Sixty-nine patients operated on for unstable coronary syndrome were randomized to intravenous infusion of glutamate (n=35) or saline (n=34) perioperatively. Plasma levels of S-100B were obtained on the third postoperative day.S-100B in the glutamate group and the control group were 0.079+/-0.034microg/L and 0.090+/-0.042microg/L respectively (p=0.245). There were no patients with stroke or mortality. Three patients in the control group and two in the glutamate group had postoperative confusion. These patients had significantly elevated S-100B compared with those without confusion (0.132+/-0.047vs 0.081+/-0.036microg/L; p=0.003). Overall, 21 patients had S-100B above reference level (or =0.10microg/L) and these patients had significantly more calcifications in the ascending aorta on epiaortic scanning.Intravenous glutamate infusion during surgery for unstable coronary artery disease did not initiate a sustained elevation of plasma S-100B. Thus, no evidence for subclinical neurological injury related to glutamate infusion was found. In contrast, postoperative elevation of plasma S-100B was linked to calcification of the ascending aorta and postoperative confusion.

We analyzed the expression of class-I antigens in ex vivo human tumor cells by isoelectric focusing (IEF) the anti-class-I mAb W6/32 immunoprecipitates prepared from cell lysates. Out of 42 experiments, 27 were technically successful. The patient's blood lymphocytes were used as controls. In vitro exposure of the tumor cells to IFN gamma and TNF alpha elevated class-I antigen expression. In 11 cases, defects in MHC-class-I-antigen expression were observed. In 2 cases the antigens were detected only in the cytokine-treated tumor samples, probably due to a defect in the association between beta 2m and class-I heavy chains. Selective changes in the expression of alleles were seen in 10 cases and might involve HLA A, B and C antigens. Alterations in class-I expression as compared with the lymphocytes were observed in 9 of 13 cases in which the tumor cells were collected from metastases, and only in 2 of 14 primary tumors.

By one-dimension isoelectric focusing we analysed the major histocompatibility complex class I antigen expression on human tumours. Blood lymphocytes of the patients, processed in parallel, served as a basis for comparison. The prerequisite for the analysis is the preparation of metabolically active tumour cell suspensions devoid of significant leucocyte contamination. The method was found to be suitable for study of the expression of HLA alleles on ex vivo tumour cells and allowed the detection of changes imposed by in vitro treatment with interferon gamma and tumour necrosis factor alpha.

In a group of 30 human tumors, comprising 12 lung, 14 ovarian, 2 breast carcinomas, 1 hypernephroma and 1 mid-gut carcinoid, the expression of major histocompatibility complex (MHC) class I molecules and the intercellular adhesion molecule 1 (CAM-1, CD54) was found to vary independently. Some tumors expressed both or neither of these molecules. Among 9/13 ICAM-1+ tumors, in which greater than 50% cells reacted with the anti-ICAM-1 monoclonal antibody (mAb) (LB-2), the class I antigen was also detected on greater than 50% of the cells. Only 2 ICAM-1+ tumors were class-I-. In 5/17 cases the tumors were MHC-class-I+ and ICAM-1-. Lymphocytes collected from the blood or from the tumor site were assayed for recognition on the tumor cells in the auto-tumor cytotoxicity test and in mixed lymphocyte tumor cell culture (MLTC). Positive results were obtained only with the MHC-class-I+/ICAM-1+ tumors. In vitro treatment of the tumor cell suspensions with interferon gamma and tumor necrosis factor alpha (TNF alpha) induced or enhanced the ICAM-1 and/or class I antigen expression in 8/12 cases. Of the tumor samples treatged, 8/9 acquired stimulatory capacity and 3/10 became susceptible to lysis by the lymphocytes. In 6/6 MLTC performed with the cytokine-treated tumor cells, cytotoxicity against the autologous tumor was generated. Three of these MLTC lymphocytes also lysed the untreated targets. mAb directed to class I antigens or to ICAM-1 inhibited both the stimulation by and the lysis of tumor cells when confronted with fresh lymphocytes. The cytotoxicity generated in the MLTC was also inhibited. If, however, the cytotoxic function was induced in MTLC containing interleukin-2 (5 U/ml), inhibition was obtained only by pretreatment of the targets with mAb against ICAM-1. The results show thus (a) that the lymphocytes react in vitro with tumor cells only if these express both MHC class I molecules and ICAM-1; (b) that expression of these molecules can be induced by interferon alpha and TNF alpha; (c) that cytotoxic effectors generated in the MLTC with cytokine-treated tumors can also act on the untreated tumor cells. The requirement of the two surface moieties for the interaction with lymphocytes was also substantiated by blockade with relevant mAb.

Although postoperative heart failure is a major determinant of operative mortality in cardiac surgery it has received little attention in the literature, and long-term consequences remain to be addressed. Therefore, the impact of postoperative heart failure on long-term survival in relation to other risk factors was studied.All patients undergoing aortic valve replacement (AVR) for aortic stenosis from 1995 through 2000 in the southeast region of Sweden (n = 398) were compared with a cohort, matched for age and sex, undergoing coronary artery bypass grafting (CABG [n = 398]). Risk factors for 5-year mortality were analyzed.Forty-five AVR and 47 CABG patients required treatment for postoperative heart failure. Thirty-day, 1-year, and 5-year mortality in patients with and without postoperative heart failure after AVR were 6.7% versus 1.4% (p = 0.05), 8.9% versus 4.0% (p = 0.13), and 42.2% versus 14.2% (p0.0001) respectively. Corresponding results in the CABG group were 21.3% versus 1.1% (p0.0001), 25.5% versus 3.1% (p0.0001), and 36.2% versus 11.1% (p = 0.0015). Postoperative heart failure, preoperative renal dysfunction, procedure-associated stroke, body mass index less than 19 kg/m2, older age, preoperative atrial fibrillation, and preoperative anemia turned out as independent risk factors for 5-year mortality after AVR. In the CABG group, postoperative heart failure, diabetes mellitus, older age, and procedure-associated stroke emerged as independent risk factor for 5-year mortality.Postoperative heart failure was associated with high early mortality after CABG whereas the grave consequences of postoperative heart failure after AVR for aortic stenosis became evident only with time.

We have analyzed 30 human tumor specimens (two breast, six lung, and 21 ovarian carcinomas and one malignant melanoma) for the expression of the transporter associated with antigen processing, TAP-1. Cell suspensions judged to contain negligible contamination with non-tumor cells were tested for reactivity with the antibody directed to TAP-1 in Western blot. According to these results all lysates prepared from the tumor samples contained the protein, but with considerable quantitative variations. Tumors exposed in vitro for a short time to IFN-gamma and TNF-alpha had elevated TAP-1 levels in 12 out of 30 experiments. In accordance with previous results, the tumor cell populations were heterogeneous with regard to MHC class I expression, as analyzed by indirect immunofluorescence on viable cell suspensions. Short term in vitro treatment with IFN-gamma and TNF-alpha elevated MHC class I expression in several tumors. In several mixed cultures, cytokine treated tumor cells induced cytotoxic activity in autologous or allogeneic lymphocyte populations. In vitro treatment with IFN-gamma and TNF-alpha upregulated thus the expression of MHC class I and TAP-1 in a fraction of the tumor samples. However the potentiation of the capacity to interact with T cells induced by the cytokines was not always parallel with the changes in these two parameters. It is therefore likely that the cytokine treatment induced additional changes in the tumors which contributed to their capacity to elicit the T cell response.

We have developed an in vitro model to study mechanisms by which ovarian tumor cells that over-express the HER-2/neu proto-oncogene escape recognition by TCD8+. Nine tumor-specific, HLA A2-restricted TCD8+ clones were isolated from 2 ovarian tumor-specific TCD8+ lines derived from tumor-infiltrating or -associated lymphocytes. Of these, 2 clones recognized the previously defined HER-2/neu epitope E75 (a.a. 369-377) and one recognized the C85 epitope (a.a. 971-979), whereas the specificity of the remaining 6 clones was unknown. Three different tumor escape variants (EVC8, EVC22 and EVC36) were produced by co-culturing an ovarian tumor line over-expressing HER-2/neu with these autologous TCD8+ clones. Cell surface expression of HLA A2 was markedly decreased on all 3 escape variants, relative to the parental tumor line, while no significant decrease in their expression of the HER-2/neu, ICAM-1 or LFA-3 molecules was found. There was a correlation between the level of tumor-specific recognition and HLA A2 expression among the tumor clones isolated from 2 of the escape variants (EVC8 and EVC36). In contrast, high HLA A2-expressing tumor clones isolated from the EVC22 variant, or EVC22 which had regained high HLA A2 expression through IFN-gamma treatment, were not recognized by the HER-2/neu-specific TCD8+ clone C-22. No mutations were found in the cDNA or the genomic DNA derived from the PCR product corresponding to a 496 bp fragment including the region coding for the E75 epitope of the HER2/neu gene in the EVC22 variant. Collectively, this in vitro model underlines the importance of decreased expression of the HLA restriction element for escape from tumor-specific TCD8+ but also demonstrates that additional mechanisms exist.

We tested 20 human carcinoma samples for the production of transforming growth factor beta (TGFbeta) in vitro. Tumour cell suspensions without obvious contamination with non-malignant cells were kept in culture conditions for 16 h and their supernatants were added to CCL-64 cells. The proliferation of these cells is inhibited by TGFbeta. According to this assay, the supernatants contained both active and latent TGFbeta. In addition, the supernatants were found to suppress the spontaneous cytotoxic function and activation of T-cell-enriched lymphocyte populations. A specific monoclonal antibody (mAb) counteracted these effects and therefore we concluded that they were mediated to a large extent by TGFbeta. In line with the results obtained with the supernatants, activation of lymphocytes could also be inhibited by tumour cells and their inhibitory effect was weaker in the presence of the TGFbeta-specific mAb. It is important to note that, when TGFbeta-specific mAb was added to autologous mixed lymphocyte/tumour cell cultures, lymphocyte activation occurred more often. These results thus substantiate the assumption that production of TGFbeta may help the survival of potentially immunogenic tumour cells in immunocompetent patients.

Short-term exposure of ex vivo carcinoma and sarcoma cells to IFN-gamma and TNF-alpha induced or elevated to detectable levels the surface expression of MHC class I, class II, and ICAM-1 (CD54), but only rarely the B7 (CD80) molecules. The cytokine-treated tumor cells interacted more efficiently with allogeneic blood lymphocytes collected from healthy donors compared with untreated cells. This was demonstrated (1) by the induction of DNA synthesis and generation of cytotoxic activity in mixed cultures and (2) by the elevated susceptibility to the cytotoxic effectors. Although the cytokine-induced increase in MHC and ICAM-1 on the low-expressor tumors were probably important to the interaction with lymphocytes, it is likely that other properties were also induced that contributed to the phenomenon. This was indicated by the results obtained with several tumors that expressed indigenously high levels of these molecules but reacted with the allogeneic lymphocytes only or more efficiently after treatment with IFN-gamma and TNF-alpha. In these experiments B7 expression did not influence the efficiency of interactions between lymphocyte and tumor cells. The results also showed that, under the conditions used, the untreated tumor cells that did not activate allogeneic lymphocytes were sensitive to appropriately activated effectors. Thus the afferent and efferent arms of lymphocyte-tumor cell interactions appeared to have different requirements.

Twenty-nine samples of ex vivo ovarian and lung carcinomas were investigated for the relationship between the presence of mutated protein 53 (mp53) and cytotoxic susceptibility. Unaltered expression of MHC class I alleles was required for the cytotoxic susceptibility of tumour cells to the autologous ex vivo blood lymphocytes, i.e. all 4 sensitive tumours belonged to the group of 11 tumours without defect in MHC class I expression. In contrast, the susceptibility did not correlate with the presence of mp53, i.e. cases with mp53 were randomly distributed between the sensitive and resistant tumours (2/4 and 10/17 respectively). There was no correlation either between the p53 mutation and down-regulation of MHC class I alleles. The results suggest that in these tumours the mutated p53 is not the source of immunogenic peptides and that the lack of recognition of the tumours with mp53 is not caused by a defect in the expression of MHC class I molecules.

Human cell lines and blood lymphocytes were treated for short time periods with IFN-gamma. This treatment increased the amount of the assembled MHC class I molecules on the plasma membrane after 30 min. This early increase of the membrane expression subsided in the next few hours. A second wave of elevation occurred after 8-24 hr. Analysis of cytoplasmic and membrane molecules in pulse chase experiments showed that the cytokine enhanced both the assembly of available heavy and light chains and the transport of the complex to the plasma membrane. The membrane level of the HLA-A2 molecules showed similar kinetics. Addition of an A2 specific binding peptide stabilized the IFN-gamma induced molecules on the cell surface. It seems that IFN-gamma alone or together with a binding peptide can influence MHC class I expression solely through post-transcriptional events utilizing an available pool of free heavy and light chains already after a short time, before the enhancement of the synthesis starts.

T-cell-enriched lymphocyte populations derived from the malignant exudate of a patient with ovarian carcinoma were exposed to autologous tumor cells in the mixed lymphocyte-tumor-cell culture (MLTC) and propagated for 42 days. Proliferation of lymphocytes depended on exposures to autologous tumor cells and on the presence of IL-2. After 7 days, the MLTC-lymphocytes lysed K562 and the autologous tumor cells. The latter effect was not inhibited by monoclonal antibodies (MAbs) reactive with MHC class-I antigens or with CD3. After 7 restimulations, the culture was enriched in CD8+ cells (92%) and showed selective lytic activity against the autologous tumor. This function was inhibited by the alpha-class I or alpha-CD3 MAbs, and also by antibodies reactive with the HLA B locus or B5 allele products. The antibodies reactive with HLA A molecules had no such effect. It seems therefore that the function of the CTLs was restricted by HLA B5. Analysis of the TCR beta genes indicated clonal T-cell expansion in this culture. This MLTC was 1 of 21 initiated with 11 blood- and 10 tumor-derived lymphocyte (TIL) populations prepared from the malignant effusions of ovarian carcinoma patients. None of these ex-vivo lymphocytes lysed autologous tumor cells. In 17 MLTCs the lymphocytes did not proliferate, and in 3 cultures the proliferation was maintained only for 2-3 weeks. In 3 of 4 cultures auto-tumor cytotoxicity was induced.

We have studied the effect of polysaccharide K (PSK) in the in vitro recognition of ex vivo carcinoma, sarcoma and lymphoma cells by the autologous blood lymphocytes. In 4/25 experiments PSK treatment activated the lymphocytes for auto-tumour lysis. Tumour cells alone generated lytic activity both in short- (16 h) and in long-term (6 days) mixed lymphocyte/tumour cell cultures (MLTC), in 2/12 and 3/13 cases respectively. The tumours that activated the lymphocytes expressed high levels of major histocompatibility complex (MHC) class I molecules. In vitro cytokine (interferon gamma and tumour necrosis factor alpha) treatment of the tumour cells elevated the amounts of class I antigens and the treated cells acquired stimulatory potential. When PSK was added to the MLTC, in which untreated tumour cells were used, lytic potential was induced in 9/13 short-term and in 11/12 long-term cultures. It is noteworthy that in the presence of PSK the untreated, negative or low-class-I-expressor tumours also activated the cytotoxic function of the lymphocytes in 4/5 long-term and in 6/7 short-term cultures. Even in the case of those lymphocytes that could be activated by PSK or tumour cells alone, the simultaneous exposure was more efficient. The effect of PSK was dose-dependent, being optimal at 1 micrograms/ml and 10 micrograms/ml. The presence of EDTA and/or cytochalasin B in the cytotoxic test performed with the activated effectors abrogated the lysis, indicating the requirement of contacts with the effector cells.

T cell responses against autologous tumors with samples from patients with a variety of tumors were examined. The abilities of T lymphocytes to lyse the autologous tumor cells were analyzed after short-term mixed lymphocyte/tumor cell cultures (MLTC). Southern blot analysis was used to evaluate whether particular rearrangements of the TCR β-chain gene predominate in these cultures. Tumor specific lysis could be induced in a proportion of the mixed cultures. In two cases enrichment of T lymphocytes with similar TCR β-chain gene rearrangements was detected after repeated stimulations with autologous tumor cells.

The kinetics of interleukin-2 receptor (IL-2R) expression and the [3H]dT incorporation of blood lymphocytes after the first and the second stimulation with allogeneic leukocytes (primary and secondary MLC) or with the autologous tumor cells (primary and secondary MLTC) were compared. The expression of IL-2R paralleled the induction of DNA synthesis. The proportion of IL-2R+ cells of the unprimed donors peaked earlier in the secondary MLC as compared to the primary MLC (on days 3 and 5 respectively). In MLC of alloimmunized healthy individuals and in the MLTC of cancer patients the highest proportions of IL-2R+ cells were detected between days 2 and 3 after both the first and second stimulations. Thus the first in vitro stimulation in the MLTC showed similar kinetics to those of the secondary MLC of unprimed individuals and to the primary MLC response of the allo-immunized individuals. The findings in the MLTC substantiate the hypothesis that cancer patients can be sensitized to their own tumors. The kinetics of the appearance of the IL-2R together with the characteristics of the IL-2-propagated cultures provide useful information for the strategy of expansion of auto-tumor reactive lymphocyte populations.