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Background: Survival of surgical inpatients is a key quality metric. Patient, surgeon, and system factors all contribute to inpatient mortality, and sophisticated risk adjustment is required to assess outcomes. When the mortality of general surgery patients was determined to be high at a safety-net hospital, a comprehensive approach was undertaken to improve patient survival., Methods: General surgical service line mortality was measured in the database of the University HealthSystem Consortium from January 2013 through June 2015. Ten best practices were implemented sequentially to decrease observed and/or increase expected mortality. University HealthSystem Consortium mortality rank, observed, expected, and observed/expected index as well as early deaths were compared with control charts for 30 months., Results: University HealthSystem Consortium general surgery mortality improved from the bottom decile to the top quartile, while Case Mix Index increased from 2.48 to 2.82 (P < .05). Observed mortality decreased from 3.39 to 2.35%. Expected mortality increased from 1.40 to 2.73% (P < .05). The observed/expected mortality index decreased from 2.43 to 0.86 (P < .05). Early deaths decreased from 0.52 to 0% (P < .05)., Conclusion: Risk-adjusted mortality and early deaths decreased significantly over 30 months in general surgery patients. Systematic implementation of quality best practices was associated with improved survival of general surgery patients at a safety-net medical center., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Patient value (V) is enhanced when quality (Q) is increased and cost (C) is diminished (V = Q/C). However, calculating value has been inhibited by a lack of risk-adjusted cost data. The aim of this analysis was to measure patient value before and after implementation of quality improvement and cost reduction programs., Study Design: Multidisciplinary efforts to improve patient value were initiated at a safety-net hospital in 2012. Quality improvement focused on adoption of multiple best practices, and minimizing practice variation was the strategy to control cost. University HealthSystem Consortium (UHC) risk-adjusted quality (patient mortality + safety + satisfaction + effectiveness) and cost (length of stay + direct cost) data were used to calculate patient value over 3 fiscal years. Normalized ranks in the UHC Quality and Accountability Scorecard were used in the value equation., Results: For all hospital patients, quality scores improved from 50.3 to 66.5, with most of the change occurring in decreased mortality. Similar trends were observed for all surgery patients (42.6 to 48.4) and for general surgery patients (30.9 to 64.6). For all hospital patients, cost scores improved from 71.0 to 2.9. Similar changes were noted for all surgical (71.6 to 27.1) and general surgery (85.7 to 23.0) patients. Therefore, value increased more than 30-fold for all patients, 3-fold for all surgical patients, and almost 8-fold for general surgery patients., Conclusions: Multidisciplinary quality and cost efforts resulted in significant improvements in value for all hospitalized patients as well as general surgery patients. Mortality improved the most in general surgery patients, and satisfaction was highest among surgical patients., (Copyright © 2016 American College of Surgeons. Published by Elsevier Inc. All rights reserved.)

Academic medical centers (AMCs) and the physicians and other professionals who lead them need to recognize they are in a business that is making a transition from a system of "sickness" care to one of "health" care, accountable for the health of defined populations and for the value (quality divided by cost) of the services provided. This change has profound implications for how AMCs conceive themselves, how they function, and how they are paid for the work that they do. A failure to recognize how the disruption of the mission of AMCs is changing may impair them as irrevocably as other changes caused the demise of Kodak, once the world's leader in the manufacture and sale of photographic film and cameras. Leaders of academic medicine need to understand, respond to, and ultimately lead the transformation of our system of health. In this Commentary, the authors review the pressures driving these changes and potential responses to them-a process already under way. They summarize the issues in the question "Should the words 'health' and 'system' take the place of 'medical' and 'center' in our institutions' names and, more important, in how we conceive of what we do?" The authors propose the name "academic health system" to better identify primary objectives to measure success by the health of patients.

Throughout the last 3 years, the Society of Thoracic Surgeons (STS) has put forth a major effort towards more accurate valuation of the work performed by cardiothoracic surgeons. The culmination of these efforts was realized on November 1, 2006, when the Centers for Medicare & Medicaid Services published the Final Rule which markedly increased the physician work values for the most frequently performed cardiothoracic surgery procedures. This article recounts the innovative approach taken by the STS during these extended efforts.

Background: Accurate control of the anticoagulation level is important for safe initiation of cardiopulmonary bypass. Using the Hemochron Jr., we consistently noted a higher than customary heparin dose required to achieve an activated clotting time (ACT) that, according to the literature and our quality standards, should be more than 480 seconds. This study was designed to determine whether there existed a significant difference in ACT values measured by the newer Hemochron Jr. and the older Hemochron 801 assay system., Methods: A total of 30 patients underwent cardiovascular surgical procedures requiring heparinization (300 U/kg). Multiple samples for measurement of the ACT were obtained from all patients before heparinization, after heparinization, during cardiopulmonary bypass, and after protamine administration. Arterial samples were collected, and ACT was determined simultaneously on the same sample using both Hemochron Jr. and Hemochron 801. Activated clotting time data were analyzed with a linear mixed model using an unstructured variance-covariance matrix., Results: Descriptive statistics on all heparinized patients revealed that the Hemochron Jr. yielded ACT results that on average were 121.28 seconds lower than the determination by the standard Hemochron 801 on the same sample of blood. This difference was -139.04 in on-pump cases and -90.51 in off-pump cases, primarily a function of the fact that higher heparin doses and therefore longer ACTs were used in patients having operations using the heart-lung machine. From the linear mixed model, the estimated average paired difference between the Hemochron Jr. and Hemochron 801 was found to be -86.03, yielding a highly significant test statistic (t(28) = -6.18; p < 0.0001)., Conclusions: Lower ACT values determined by Hemochron Jr. are consistent with higher, clinically acceptable ACT values as measured by the Hemochron 801. These findings would suggest that safe levels of anticoagulation are determined in part by the specific assay used.

Background: The Society of Thoracic Surgeons National Cardiac Database data indicate that the performance of mitral repair has increased significantly (1990 to 23.2%, 1999 to 32.0%, p < 0.0001). We examined contemporary (1999 to 2000) usage of mitral repair in the United States., Methods: We analyzed National Cardiac Database data to determine the operative approach for a diagnosis of mitral regurgitation., Results: A total of 21,741 isolated and combined mitral valve procedures were identified. The overall frequency of repair was 37.7% (8206). For procedures isolated to the mitral valve, the frequency of repair was 35.7% (3027/8486) whereas repair was more common with concomitant CABG (42.9% [3088/7193], p < 0.0001). The proportion of patients having repair decreased with age (41.2% [386/936] in 20 to 39 years, 36.1% [3513/9746] in > 70 years, p = 0.0016). Repair was more common in males (43.5% [4720/10860]) than females (32.0% [3472/10842], p < 0.0001). Repair was less common as NYHA Class increased (Class I, 47.8% [949/1984] vs Class IV, 33.2% [1803/5427]) and for emergent operative status (21.2% [156/736] vs 38.5% [8000/20773] for elective/urgent, both p < 0.0001). The number of prior operations did not affect the use of repair. Simple annuloplasty was performed in the majority of reported repairs (62.8% [3837/6115]), more so with associated CABG as compared to isolated repair (70.2% [2167/3088] vs 55.2% [1670/3027]; p < 0.0001)., Conclusions: Mitral repair was performed in over one-third of the patients reported in 1999 to 2000 and has increased since the National Cardiac Database inception. Repair usage differed based on sex, age, gravity of illness, and associated procedures. This provides a base line from which to expand the application of repair.

Background: Long-term pig xenografts in monkeys demonstrated the infiltration of CD8 T cells into pig cartilage xenografts, transplanted into monkeys. The objective of the present study was to determine in an experimental animal model whether CD8 T cells in pig xenograft recipients exert any direct cytotoxic effect on pig cells., Methods: The killing of xenograft cells by CD8 T cells, obtained from xenograft recipients, was studied in alpha1,3galactosyltransferase knockout mice that were repeatedly injected intraperitoneally with pig kidney membranes. The pig kidney cell line PK15, which shares many antigens with pig kidney membranes, served as a model for xenograft target cells in cytotoxicity assays. Cell lines from other species were also studied as target cells., Results: Lymphocytes obtained freshly from spleens of mice immunized with pig kidney membranes failed to display significant cytotoxic activity against pig cells. However, incubation of these lymphocytes with irradiated PK15 cells and addition of recombinant interleukin (IL)-2 (100 U/mL), on the third day of incubation, resulted in extensive proliferation and expansion of CD8 cytotoxic T lymphocytes (CTL). These CTL, obtained after 12 days of incubation, killed nonspecifically pig, human, and mouse normal and malignant cells. These CTL were not generated in cultures in the absence of stimulatory pig cells or in the absence of IL-2. These CTL could not be generated in cultures of lymphocytes from naive mice that were incubated with PK15 cells and IL-2., Conclusions: The data obtained imply that CD8 T cells from xenograft recipients can be stimulated in vitro by xenoantigens and IL-2 to differentiate into highly reactive nonspecific CTL that are capable of killing a large variety of xenogeneic and syngeneic cells. Similar in vivo microenvironmental conditions within the xenograft may induce the local differentiation of infiltrating CD8 T cells into CTL that can destroy nonspecifically adjacent xenograft cells. Such cells may not be active outside the xenograft because of the absence of IL-2 in sufficiently high concentrations.

Background: A major barrier in pig to human organ transplantation is the binding of human anti-Gal to alpha-gal epitopes (Gal alpha 1-3Gal beta 1-4GlcNAc-R) on pig cells, resulting in hyperacute and acute vascular rejection of pig xenografts. Moreover, the immune system in xenograft recipients is activated by these epitopes to produce high affinity anti-Gal, which is also detrimental to xenografts. Production of anti-Gal can be prevented by specific elimination of anti-Gal B cells. This was achieved with the toxin ricin A, coupled to human alpha1-acid glycoprotein modified to carry alpha-gal epitopes. This complex, designated alpha-gal ricin, is targeted in vivo to anti-Gal B cells by interaction with the immunoglobulin molecules (i.e., B cell receptors) on these cells., Methods: Carbohydrate chains on alpha 1-acid glycoprotein were converted to carry alpha-gal epitopes by enzymatic treatment with recombinant alpha 1,3 galactosyltransferase (alpha 1,3GT). This molecule and ricin A were biotinylated and coupled by avidin to generate alpha-gal ricin. The efficacy of alpha-gal ricin in eliminating anti-Gal B cells was studied in the experimental model of alpha 1,3GT knockout (KO) mice. These mice produce large amounts of anti-Gal immunoglobulin G when immunized with pig kidney membranes, as measured by ELISA with alpha-gal epitopes linked to bovine serum albumin (BSA). In the absence of anti-Gal B cells, these mice lack the ability to produce anti-Gal., Results: Repeated administration of alpha-gal ricin into alpha1,3GT KO mice resulted in elimination of anti-Gal B cells, thereby preventing production of anti-Gal immunoglobulin G after immunization with pig kidney membranes. This prevention of anti-Gal production occurred with doses of alpha-gal ricin that were not toxic to the mice and did not affect production of antibodies with other specificities., Conclusions: Administration of alpha-gal ricin results in specific elimination of anti-Gal B cells in alpha 1,3GT KO mice. The elimination of these B cells may prove to be helpful in attempts to achieve immune tolerance to alpha-gal epitopes in primates.

Background: The shortage of human hearts remains a major barrier to the efficacy of heart transplantation for the treatment of end-stage heart disease. One potential solution to the supply problem would be the use of hearts from nonhuman donors (xenografts). We have established a model of mouse to rat xenogeneic bone marrow chimerism, and in this study we have hypothesized that such chimeric rats will accept both donor and recipient specific heart grafts while rejecting third-party mouse and rat grafts. We also investigated humoral responses in naive and chimeric rats with and without donor murine cardiac grafts., Methods: Recipient Lewis rats (n = 22) were given 1100 cGy lethal total body irradiation and the same day received 300 x 10(6) donor B10.BR mouse bone marrow cells intravenously. Peripheral blood of surviving rats (n = 18) was typed at 4 weeks and then monthly thereafter. Donor and recipient specific and third-party heterotopic heart transplantations were performed at 6 to 8 weeks after reconstitution with bone marrow., Results: Multilineage bone marrow chimerism was produced in all experimental animals with complete replacement of recipient marrow by donor cells. Murine donor and rat recipient strain hearts transplanted in chimeric rats survived indefinitely. Third-party rat and mouse hearts were rejected, though at a slower rate than bone marrow matched naive controls. High levels of antimouse antibodies were detected in rats with rejected hearts. These antibodies were absent in chimeric animals with long-term surviving heart grafts., Conclusions: Long-term multilineage bone marrow chimerism can be produced in a mouse --> rat bone marrow transplant model. Long-term survival of donor specific and recipient specific vascularized cardiac grafts can be produced in these chimeric animals. These animals are clinically normal but show signs of subclinical immunosuppression regimen as they reject third-party hearts later than naive animals. Our results suggest that antibodies also play a significant role in concordant xenograft rejection, and induction of bone marrow chimerism can overcome this barrier.

Aortoesophageal fistula (AEF) is an uncommon cause of upper gastrointestinal hemorrhage. Usually, but not always, patients present with a small sentinel bleed followed by a variable interval of apparent resolution, and then they experience a massive exsanguinating hemorrhage. The variable interval of time after the sentinel bleed is the period in which most AEFs resulting from thoracic aortic aneurysm have been successfully treated. Although only a few successful cases have been reported in the literature, most describe an in situ repair. We describe treatment of a late-presenting AEF due to a thoracic aneurysm with an extra-anatomic bypass graft for the aortic repair.

Background: Xenotransplantation is a potential solution to the critical shortage of transplantable organs. However, conventional immunosuppressive agents do not control the vigorous cellular and humoral rejection across species disparities. The induction of donor specific tolerance via bone marrow chimerism may be a method to avoid xenograft rejection. In xenogeneic chimeras, T cell repertoire selection plays an important role in the induction of tolerance. Until now a model of mouse-->rat multilineage chimerism has not been reported. This study reports the establishment of fully xenogeneic mouse-->rat multilineage chimeras and evaluates the role of T cell development and repertoire selection in tolerance induction in a xenogeneic environment., Methods: Recipient rats were irradiated at a dose of total body irradiation ranging between 800-1100 cGy and injected with 120-300x10(6) donor mouse bone marrow cells. Chimeras were typed for engraftment at 4 weeks and then monthly thereafter. T cell repertoire was evaluated in chimeras using two-color flow cytometry and monoclonal antibodies directed against the variable portion of the beta chain of the T cell receptor., Results: Fully xenogeneic multilineage bone marrow chimerism was produced in a mouse-->rat model by using ablative radiation and a high dose of donor cells. Mouse T cells develop in a phenotypically normal fashion in chimeric rats and the host rat is capable of deleting T cells that are reactive to the donor mouse strain., Conclusion: Long-term multilineage bone marrow chimerism can be produced in a mouse-->rat bone marrow transplant model. Mouse T cells develop in a phenotypically normal fashion and negative selection of specific T cell receptor-Vbeta occurs in a xenogeneic environment in a predictable fashion paralleling that for syngeneic or allogeneic transplantation.

Objectives: The purpose of this study was to determine risk factors for adverse events following protamine administration after cardiopulmonary bypass., Background: Intravenous protamine administration is associated with a risk of severe systemic reactions. However, risk factors for these events have not been well delineated, thus hampering development of preventive strategies., Methods: A case-control study nested within a cohort of consecutive patients undergoing surgery requiring cardiopulmonary bypass was performed. The primary case definition included those events (pulmonary hypertensive and systemic hypotensive) occurring within 10 min of protamine administration in the absence of other measurable causes of hemodynamic compromise., Results: Comparing the 53 cases to the 223 control subjects, three risk factors were independently associated with events (multivariable odds ratio [95% confidence interval]): neutral protamine Hagedorn insulin use (8.18 [2.08, 32.2]); fish allergy (24.5 [1.24, 482.3]), and a history of nonprotamine medication allergy (2.97 [1.25, 7.07]). These risk factors demonstrated an increasingly strong association with progressively more specific case definitions. An estimated 39% of cardiopulmonary bypass patients had one or more of these risk factors. Prior intravenous protamine, central venous pressure prior to protamine, preoperative ejection fraction and the need for inotropes when coming off bypass did not exhibit statistically significant associations with events (all p > 0.15). Prior protamine allergy was associated specifically with an increased risk of pulmonary hypertension (multivariable odds ratio 189; 95% confidence interval 13, 2,856)., Conclusions: Immunologic factors are important in predisposing individuals to protamine reactions, and a substantial proportion of patients are at considerably increased risk Strategies to reduce the risk of protamine-associated events are needed.

Heart failure is an important medical and public health problem. Although medical therapy is effective for many people, the only definitive therapy is heart transplantation, which is limited severely by the number of donors. Mechanical devices presently are used as "bridges" to transplantation. Their widespread use may solve the donor shortage problem, but at present, mechanical devices are limited by problems related to blood clotting, power supply, and foreign body infection. Cardiac xenotransplantation using animal donors is a potential biologic solution to the donor organ shortage. The immune response, consisting of hyperacute rejection, acute vascular rejection, and cellular rejection, currently prevents clinical xenotransplantation. Advances in the solution of these problems have been made using conventional immunosuppressive drugs and newer agents whose use is based on an understanding of important steps in xenoimmunity. The most exciting approaches use tools of molecular biology to create genetically engineered donors and to induce states of donor and recipient bone marrow chimerism and tolerance in xenogeneic organ recipients. The successful future strategy may use a combination of a genetically engineered donor and a chimeric recipient with or without nonspecific immunosuppressive drugs.

Background: Vesnarinone (VES) has been used for treatment of patients with congestive heart failure. In addition to inotropic effects, it seems to have immunosuppressive action. We tested the hypothesis that VES suppresses graft rejection, inotropic dysfunction caused by early rejection, and chronic coronary obstruction in a heterotopic rat cardiac transplantation model., Methods: (1) To study acute rejection, hearts from Lewis-Brown Norway (LBN) rats were transplanted into Lewis rats, which were treated with or without VES (50 or 100 mg/kg/day orally). (2) In a functional study, LBN hearts with or without VES (100 mg/kg/ day) were isolated and perfused on day 3 after transplantation to assess inotropic response to isoproterenol (3 x 10(-8) M). (3) To study chronic rejection, Lewis hearts were transplanted into Fisher 344 rats, which were treated with or without VES (50 mg/kg/day) for 90 days. Coronary obstructive disease was assessed by morphometric analysis. There were five to six animals in each group., Results: (1) VES (100 mg/kg/day) prolonged LBN heart survival (11.7 +/- 0.7 vs. 9.6 +/- 0.7 days in control; P < 0.05). (2) Left ventricular developed pressure was depressed in transplanted hearts regardless of VES treatment (84 +/- 12, 90 +/- 8 vs. 144 +/- 16 mmHg in untransplanted hearts; P < 0.01). The developed pressure after administration of isoproterenol in VES-treated hearts (184 +/- 20 mmHg) was higher than transplanted hearts without VES (118 +/- 16 mmHg; P < 0.05), and similar to untransplanted hearts (203 +/- 27 mmHg; P = NS). (3) Transplanted hearts treated with or without VES showed similar grades of rejection (2.0 +/- 0.3 vs. 2.6 +/- 0.2; P = NS), intimal area (6,996 +/- 3,186 vs. 13,441 +/- 5,165 microns2; NS), and coronary luminal obstruction (45 +/- 16% vs. 67 +/- 14%; NS)., Conclusions: VES produces mild prolongation in survival of rat heart grafts, but has no significant effect on chronic graft atherosclerosis. VES preserves the positive inotropic effects of isoproterenol that are otherwise deteriorated by early acute rejection.

Indefinite donor-specific tolerance to a cardiac allograft can be induced through pretransplantation intrathymic injection of donor spleen cells and a single intraperitoneal injection of antilymphocyte serum. This study was designed to determine whether this phenomenon was reproducible with grafts differing in either class I major histocompatibility complex only or class II MHC only. Donors of cells and hearts in all experiments were RP rats. Class I MHC disparate grafts were performed by placing an RP heart into a Lewis recipient, and class II disparate grafts were performed with RP donors and Wistar Furth recipients. Lewis (n = 10) and Wistar Furth (n = 10) recipients underwent intraperitoneal injection of 1 ml antilympocyte serum and intrathymic injection of 5 x 10(7) RP spleen cells. Three weeks later, heterotopic cardiac transplantation was done with a heart from an RP rat. Control rats had no pretreatment or received antilympocyte serum alone. Without pretreatment, RP hearts survived 7 to 9 days (mean 8 days) in Lewis recipients (n = 5) and 9 to 14 days (mean 12 days) in Wistar Furth recipients (n = 5). Antilymphocyte serum alone produced slight prolongation of graft survival. Lewis rats pretreated with class I disparate RP splenocytes and antilympocyte serum had graft survivals of 8 to 27 days (mean 14 days), not significantly different from the results with antilympocyte serum alone. Class II disparate RP grafts placed in pretreated Wistar Furth rats had significant prolongation of graft survival, with four of five grafts surviving longer than 60 days (p < 0.01 vs antilympocyte serum alone). These results suggest that a disparity at the class II locus of the major histocompatibility complex is critical for the induction of cardiac allograft tolerance after intrathymic inoculation of allogeneic cells.

Background: In utero fetal inoculation with allogeneic cells has produced subsequent tolerance to experimental cardiac allografts. We attempted to extend this observation to a model of xenogeneic cardiac transplantation., Methods: Lewis rat fetuses were inoculated with Golden Syrian hamster thymocytes (n = 5) or whole spleen cells (n = 5) on the tenth day of intrauterine life. Six weeks after the birth of pretreated fetuses, heterotopic cardiac transplantation using a hamster donor was performed. Three to 4 weeks after parturition, we performed heterotopic cardiac transplantation using hamster donors in the female Lewis rats whose fetuses had been treated in utero., Results: Animals treated in utero with either thymocytes or whole spleen cells had graft survival of 3 days, not different from that in untreated Lewis rats (n = 5) (p = not significant). Maternal Lewis rats whose fetuses were treated with thymocytes (n = 5) or whole spleen cells (in = 4) had markedly reduced survival of xenogeneic cardiac grafts (range, 3 to 20 hours; mean, 15 hours; p < 0.01; and range, 5 to 15 minutes; mean, 10 minutes; p < 0.01, respectively). Female Lewis rats without intrauterine inoculation (n = 5) had expected xenograft survival time (3 days) (p = not significant). Immunohistochemical staining of hyperacutely rejected grafts showed deposits of immunoglobulin M as well as immunoglobulin G and complement. In normally rejected xenografts, no immunoglobulin M was detected., Conclusions: These studies reveal the surprising observation that fetal exposure to xenogeneic cells sensitizes the maternal rat without tolerizing the fetal rat as observed in an allograft model. In addition, whole spleen cells produce a more vigorous hyperacute rejection than thymocytes, suggesting that B cells or macrophages may be the sensitizing agents. The accelerated rejection observed has the characteristics of an immunoglobulin M antibody-mediated hyperacute rejection response with deposition of complement.

Background: In a previous study we demonstrated that intrathymic exposure of neonatal rats to both alloantigens and xenoantigens produced tolerance only to subsequent cardiac allografts and not to xenografts implanted when the animals were 6 weeks old. Interestingly, graft recipients were not sensitized to the xenografts as observed in the adult model. This study was designed to investigate whether earlier grafting would produce tolerance to cardiac xenografts in animals pretreated by neonatal intrathymic inoculation with allogeneic and xenogeneic cells., Methods: All recipient animals were Lewis rats. Donors were either Lewis Brown Norway rats or Golden Syrian hamsters. Lewis Brown Norway rat and hamster splenocytes (25 x 10(6) cells in a volume of 0.01 ml) were inoculated percutaneously into the thymus of neonatal recipients (n = 22). At age 4 weeks, five pretreated recipients underwent cervical heterotopic heart transplantation with rat hearts, and 2 weeks later abdominal heterotopic transplantation was done with hamster donors. A second group ( n = 6) received hamster hearts as the first graft and then grafts from rat donors. The third group underwent rat followed by hamster heart transplantation at age 6 to 7 weeks., Results: Mean rat allograft survival time for groups 1,2, and 3, respectively was 49.8 days with 4 of 5 surviving indefinitely, 4.3 days with 2 of 3 surviving indefinitely, and 42 days with 7 of 11 surviving indefinitely (p < 0.01 versus untreated control animals, 7.8 days, n = 5). In rats undergoing transplantation at 4 weeks, cardiac xenografts were rejected in 2.5 days 2.3 days, which was significantly shorter than xenograft survival (3.3 days) in rats that underwent transplantation at an older age (p < 0.02), in naive rats (p < 0.05), and in rats treated with hamster cells alone (p < 0.05). Mixed lymphocyte reaction showed a diminished proliferative response to Lewis Brown Norway rat cells in pretreated rats, which retained the ability to respond in culture to hamster cells (p < 0.05)., Conclusions: Earlier grafting in rats pretreated as neonates produces allograft tolerance but may accelerate rejection of xenografts. Preliminary mixed lymphocyte reaction results suggest that only the alloimmune cellular proliferative response is abrogated by this pretreatment.

To study the molecular immunologic features of allograft rejection and tolerance induction by intrathymic pretreatment we developed a murine model of cardiac transplantation. In this model the transplant recipient was the C57BL/6 mouse with its major histocompatibility phenotype H-2b. Donors of cells for intrathymic pretreatment and of hearts for grafting were mice of the bm1 mutation. The bm1 mutation involves substitution of three amino acids in one of the alpha helixes of the class I H-2Kb molecule. Because of the discrete molecular configuration of the transplant antigen we hypothesized that there would be limited heterogeneity of receptor expression on C57BL/6 T cells responding to bm1 cardiac grafts and that intrathymic pretreatment would alter the T-cell repertoire of graft recipients by causing elimination of T cells responsible for graft rejection. Mice were given 0.3 ml of antilymphocyte serum intraperitoneally and had intrathymic injection of 25 x 10(6) bm1 splenocytes 12 to 21 days before transplantation with a bm1 cardiac graft. Flow cytometric analysis of lymph node and spleen cells was used to study the V beta T-cell repertoire of graft recipients. Cells were stained with monoclonal antibodies to CD3 and 13 V beta regions (n = 5, each group) of T cells in naive, sensitized, and tolerant animals. Untreated C57BL/6 mice (n = 9) rejected bm1 cardiac grafts a mean of 20.4 days after transplantation. Twelve mice pretreated with antilymphocyte serum and intrathymic bm1 cells had permanent graft survival (> 100 days, p < 0.0001). Animals treated with antilymphocyte serum alone (n = 5) or pretreated animals undergoing third-party BALB/c grafts (n = 4) rejected grafts in the normal time frame. There was significant alteration of percentage receptor expression of V beta 5.1, 7, 12, 13, and 17a in sensitized and tolerant mice. Of interest, V beta 7 expression was increased in the sensitized mice (3.8% to 8.3%,p = 0.005) and was virtually eliminated in tolerant mice (p = 0.005). In conclusion, these data suggest that V beta 7 is a critical receptor in the C57BL/6 response to subcutaneous bm1 cardiac grafts. Pretreatment of graft recipients with one dose of antilymphocyte serum and intrathymic bm1 cells appears to produce permanent tolerance to bm1 grafts with elimination of T cells expressing receptor chain V beta 7.

Permanent tolerance to an experimental heterotopic cardiac allograft can be achieved by pretreatment with antilymphocyte serum (ALS) and intrathymic inoculation of donor cells. Most successful experimental protocols have employed a time lag of 2 to 3 weeks between intrathymic pretreatment and transplantation, which makes this treatment strategy impractical for clinical heart transplantation. In these experiments we modified the standard protocol by giving ALS 24 hr prior to both intrathymic injection of donor cells and heterotopic transplantation. Seven Lewis rats had intraperitoneal injection of 1 ml of ALS and 24 hr later underwent intrathymic injection of 5 X 10(7) donor Lewis-Brown Norway (LBN) splenocytes and heterotopic cardiac transplantation using an LBN donor. Mean graft survival was 24.4 days, significantly longer than the 7.8-day graft survival observed in untreated Lewis recipients (n = 5) (P < 0.02). However, graft survival was not different from that observed in Lewis rats pretreated with ALS alone (n = 5) (25.8 days, P = NS). Permanent graft survival was produced in two rats receiving only A-LS and in one rat receiving both ALS and intrathymic inoculation. In these experiments it appears that prolongation of graft survival may have been due to the effect of A-LS alone. These results suggest that there is a critical time period between intrathymic inoculation and transplantation that is needed for permanent tolerance to be induced consistently. This may be due to the kinetics of the effects of ALS on alloreactive T-lymphocytes or to a time-dependent requirement for antigen processing in the thymus.

Permanent acceptance of an experimental cardiac allograft can be achieved in the rat by pretreating the recipient with antilymphocyte serum and intrathymic donor lymphocytes. We investigated the durability and specificity of the tolerance produced by this pretreatment in a rat model of heterotopic heart transplantation with Lewis-Brown Norway donors and Lewis recipients. Pretreated Lewis rats received 1 ml antilymphocyte serum intraperitoneally and 5 x 10(7) Lewis-Brown Norway splenocytes intrathymically, followed 21 days later by Lewis-Brown Norway cardiac transplantation. The first Lewis-Brown Norway cardiac allograft survived long term (mean 140 days) in pretreated recipients who were given no subsequent immunosuppression. After 60 days with a beating Lewis-Brown Norway allograft, tolerant Lewis recipients underwent a second cardiac allograft with either a Lewis-Brown Norway heart or a third-party Wistar-Furth heart. The second Lewis-Brown Norway cardiac allograft was not rejected (mean survival 76 days), but that from the third-party Wistar-Furth donor was rejected in a normal fashion (mean survival 10.4 days). The presence of second grafts did not affect survival of first grafts. Tolerant Lewis recipients of two Lewis-Brown Norway heart grafts underwent subsequent transplantation with Lewis-Brown Norway skin. Skin allograft survival in this group (mean 8.4 days) was not different from that in Lewis recipients without pretreatment. Rejection of skin grafts had no effect on the heart grafts. These data suggest that tolerance to cardiac allografts produced by intrathymic pretreatment is durable and extends to a second heart graft from a genetically identical donor. Tolerant rats reject third-party hearts and primary donor skin grafts normally, and tolerance to previously placed heart grafts is not abrogated by this rejection. Non-major histocompatibility complex skin antigens not present on cardiac cells may account for the tissue specificity of the tolerance produced by intrathymic treatment in this model.

Background: Donor-specific tolerance to a rat heterotopic cardiac allograft has been achieved by the pretransplantation intrathymic injection of donor splenocytes and a single intraperitoneal injection of antilymphocyte serum (ALS). Permanent tolerance is achieved without subsequent immunosuppression therapy. This study investigated the mechanisms responsible for maintenance of the tolerant state., Methods: Tolerance was produced in Lewis rats by the administration of 1 mL of ALS intraperitoneally and 5 x 10(7) Lewis Brown Norway (LBN) splenocytes intrathymically 21 days before heterotopic transplantation using an LBN donor., Results: In tolerant Lewis rats bearing LBN allografts for more than 100 days, rejection could not be produced by the intravenous injection of naive Lewis spleen cells (5 x 10(7) cells x 1 day, n = 5; 5 x 10(7) cells x 3 days, n = 5) or cells from Lewis rats sensitized to LBN tissues (5 x 10(7) cells x 3 days, n = 5). Naive Lewis recipients were pretreated with ALS and 6 days later with intravenous spleen cells (25 x 10(7), n = 5) or lymphoid cells (10 to 15 x 10(7), n = 5) from a tolerant animal bearing a viable LBN graft. When transplantation with an LBN donor was done the next day, significant prolongation of LBN allograft survival (mean survival time 32.8 days, p < 0.01; and 22.2 days, p < 0.01; respectively) was seen. Wistar-Furth allograft survival was not prolonged by treatment with ALS and intravenous spleen (n = 5) or lymph node (n = 5) cells from rats tolerant to LBN tissues (mean survival time 8.6 and 9.2 days, control 9 days; p = not significant). The administration of ALS alone (n = 5) did not prolong LBN graft survival (mean survival time 11.8 days)., Conclusion: These data suggest that transferable suppressor cells specific for the donor strain are at least in part responsible for the maintenance of long-term allograft survival after intrathymic pretreatment with allogeneic cells.