Lung transplant acceptance is facilitated by early events in the graft and is associated with lymphoid neogenesis.

Early immune responses are important in shaping long-term outcomes of human lung transplants. To examine the role of early immune responses in lung rejection and acceptance, we developed a method to retransplant mouse lungs. Retransplantation into T-cell-deficient hosts showed that for lungs and hearts alloimmune responses occurring within 72 h of transplantation are reversible. In contrast to hearts, a 72-h period of immunosuppression with costimulation blockade in primary allogeneic recipients suffices to prevent rejection of lungs upon retransplantation into untreated allogeneic hosts. Long-term lung acceptance is associated with induction of bronchus-associated lymphoid tissue, where Foxp3(+) cells accumulate and recipient T cells interact with CD11c(+) dendritic cells. Acceptance of retransplanted lung allografts is abrogated by treatment of immunosuppressed primary recipients with anti-CD25 antibodies. Thus, events contributing to lung transplant acceptance are established early in the graft and induction of bronchus-associated lymphoid tissue can be associated with an immune quiescent state.

Costimulatory blockade-mediated lung allograft acceptance is abrogated by overexpression of Bcl-2 in the recipient.

Lung allografts are considered to be more immunogenic than other solid organs. Little is known about the effectiveness of immunosuppressive regimens after lung transplantation. Herein, we describe a novel model of murine vascularized orthotopic lung transplantation we used to study the effects of costimulatory blockade on lung rejection. Transplants were performed in the Balb --> B6 strain combination. Recipients were either not immunosuppressed or received perioperative CD40/CD40L and CD28/B7 costimulatory blockade. Nonimmunosupressed Balb/c --> B6 lung transplants had severe acute rejection 7 days after transplantation and CD8(+) T cells outnumbered CD4(+) T cells within the allografts. Alternatively, B6 recipients that received perioperative costimulatory blockade had minimal inflammation and there were nearly equal numbers of CD8(+) and CD4(+) T cells in these grafts. Approximately one third of graft-infiltrating CD4(+) T cells expressed Foxp3. CD4(+) T cells isolated from these grafts induced apoptosis of alloreactive CD8(+) T cells that were stimulated with donor splenocytes in vitro. In contrast with wild-type B6 recipient mice, we observed severe rejection of Balb/c lungs 7 days after transplantation into Bcl-2 transgenic B6 recipients that had received costimulatory blockade. CD8(+) T cells outnumbered CD4(+) T cells in these immunosuppressed Bcl-2 transgenic recipients and, compared with immunosuppressed wild-type B6 recipients, a lower percentage of graft-infiltrating CD4(+) T cells expressed Foxp3, and a higher percentage of graft-infiltrating CD8(+) T cells expressed intereferon-gamma. Thus, our results show that perioperative blockade of the CD40/CD40L and CD28/B7 costimulatory pathways markedly ameliorates acute rejection of lung allografts in wild type but not Bcl-2 transgenic recipients.

Monocyte differentiation is controlled by MyD88 after mouse orthotopic lung transplantation.

In lung grafts, ischemia-reperfusion signals rapidly induce the recruitment and differentiation of host monocytes into macrophages and dendritic cells. The nature of ischemia-reperfusion signals are antigen independent, but have been hypothesized to initiate Toll-like receptor (TLR) and interleukin (IL)-1R-mediated signaling pathways that are thought to potentiate alloimmune responses. We wondered whether MyD88, an adaptor molecule critical for both TLR and IL-1R-mediated inflammatory responses, regulated monocyte differentiation in a mouse model of vascularized orthotopic lung transplantation. Orthotopic left lung transplants were performed in the following syngeneic combinations: CD45.1(+) B6 --> CD45.2(+) MyD88(-/-) and CD45.1(+) B6 --> CD45.2(+) B6. One day later, recipient-derived dendritic cells and macrophage numbers were assessed in the bronchiolar lavage by FACS analysis. Compared with the bronchiolar lavage of wildtype recipients, MyD88(-/-) recipients had lower numbers of dendritic cells in lung graft airways that were of recipient origin. Lower numbers of newly differentiated lung graft dendritic cells was coincident with the appearance of higher numbers of undifferentiated monocytes in the lung airways of MyD88(-/-) recipients as compared with wild-type recipients. Moreover, adoptive transfer experiments demonstrated that MyD88(-/-) monocytes were poorer at differentiating into lung dendritic cells as compared with wild-type monocytes. Taken together, these data show that MyD88 regulates graft-infiltrating monocyte differentiation and suggests a mechanism by which TLR/IL-1R-signaling pathways control adaptive responses in lung allografts through controlling monocyte fate.

Sphingosine 1-phosphate inhibits ischemia reperfusion injury following experimental lung transplantation.

Ischemia reperfusion (I/R) injury following lung transplantation is exacerbated by the destruction of the endothelial cell barrier leading to pulmonary edema and dysregulated activated lymphocyte migration. Sphingosine 1-phosphate (S1P), a G-coupled protein receptor (GPCR) agonist, has been previously shown to promote endothelial cell tight junction formation and prevent monocyte chemotaxis. We asked if S1P treatment could improve pulmonary function and attenuate I/R injury following syngeneic rat lung transplantation. In comparison to vehicle-treated recipients, S1P administered before reperfusion significantly improved recipient oxygenation following transplantation. Improved graft function was associated with reduced inflammatory signaling pathway activation along with attenuated intragraft levels of MIP-2, TNF-alpha and IL-1beta. Moreover, S1P-treated recipients had significantly less apoptotic endothelial cells, pulmonary edema and graft accumulation of neutrophils than did vehicle-treated recipients. Thus our data show that S1P improves lung tissue homeostasis following reperfusion by enhancing endothelial barrier function and blunting monocytic graft infiltration and inflammation.

Myocardial tissue engineering and regeneration as a therapeutic alternative to transplantation.

Ischemic cardiomyopathy leading to congestive heart failure remains the leading source of morbidity and mortality in Western society and medical management of this condition offers only palliative treatment. While allogeneic heart transplantation can both extend and improve the quality of life for patients with end-stage heart failure, this therapeutic option is limited by donor organ shortage. Even after successful transplantation, chronic cardiac rejection in the form of cardiac allograft vasculopathy can severely limit the lifespan of the transplanted organ. Current experimental efforts focus on cellular cardiomyoplasty, myocardial tissue engineering, and myocardial regeneration as alternative approaches to whole organ transplantation. Such strategies may offer novel forms of therapy to patients with end-stage heart failure within the near future.

Emergent lung retransplantation after discovery of two primary malignancies in the donor.

A 50-year-old woman underwent single lung transplantation for advanced chronic obstructive pulmonary disease. Shortly after the procedure, it was discovered that the donor suffered from both a renal cell carcinoma and a spindle-cell sarcoma of the ascending aorta, which had metastasized to the spleen. The patient was emergently listed for a retransplantation and underwent bilateral lung transplantation after a new donor became available 4 days after the initial transplantation procedure. After 24 months, the patient is without evidence of malignancy. This case illustrates the role of immediate retransplantation for patients who have inadvertently received thoracic organs from donors harboring occult malignancies.

Hand-assisted laparoscopic vertical banded gastroplasty: early results.

BACKGROUND: Minimally invasive hand-port-assisted laparoscopic vertical banded gastroplasty has the potential to reduce postoperative complications after bariatric surgery. METHODS: We analyzed the postoperative course of 46 hand-port-assisted laparoscopic vertical banded gastroplasties (LVBG) completed between January 1998 and April 1999. RESULTS: The operating time for the LVBG was shorter (140.8 +/- 6.0 vs 180.2 +/- 6.3 min; p < 0.05). Individuals were able to ambulate sooner (1.36 +/- 0.09 vs 2.44 +/- 0.16 days; p < 0.05), and start oral intake earlier (2.7 +/- 0.27 vs 3.7 +/- 0.17 days; p < 0.05) than the open vertical banded gastroplasty (VBG) controls. Three staple line leaks were detected in this group. Two leaks resolved without clinical sequelae, but one patient developed intraabdominal sepsis. This complication extended the average hospital stay to 6.8 +/- 2.00 days, as compared to 7.71 +/- 0.18 days for historical controls. By discounting this patient from the analysis, we arrive at a more representative length of hospitalization of 4.82 +/- 0.34 days (p < 0.05). CONCLUSIONS: LVBG offers a good alternative to the standard open VBG. Although this procedure has a relatively short learning curve, it should be done at centers with an interest in bariatric surgery.

Donor antigen-presenting cells are important in the development of obliterative airway disease.

OBJECTIVE: Obliterative airway disease, which resembles obliterative bronchiolitis histologically, develops in murine heterotopic tracheal allografts. Chimeric tracheas were used to examine whether donor-type antigen-presenting cells are important in the development of obliterative airway disease. To separate the contributions of CD4(+) and CD8(+) direct pathways, we transplanted tracheas from knockout mice lacking major histocompatibility complex (MHC) class I or II antigens. METHODS: Chimeric tracheas were created via bone marrow transplantation in fully MHC-mismatched combinations. Tracheas from naive B6, autologously reconstituted B6, chimeric B6 bearing recipient-type C3H antigen-presenting cells, MHC class I knockout B6 (B6(I-)), MHC class II knockout B6 (B6(II-)), or C3H mice were transplanted into C3H recipients. The tracheas were harvested at days 14 and 28. RESULTS: At day 28, isografts showed no occlusion, normal respiratory epithelium, and minimal infiltrates. Naive or autologously reconstituted B6, B6(I-), and B6(II-) tracheas showed minimal occlusion at day 14 but contained intraepithelial infiltrates. By day 28, the naive or autologously reconstituted B6 tracheas had occlusion of 69.5% +/- 11.6% (mean +/- standard error of the mean), and in comparison, B6(I-) and B6(II-) tracheas had occlusions of 53.0% +/- 16.3% and 52.2% +/- 15.9%, respectively (P =. 20,.19). In chimeric B6 tracheas, minimal occlusion was seen at day 14 and remained 33.6% +/- 16.2% (P =.039) at day 28. Subtle epithelial changes and minimal infiltrates were seen. CONCLUSIONS: Obliterative airway disease appears to involve donor-type antigen-presenting cells and develops in the absence of either MHC class I or II antigens. These findings suggest that either CD8(+) or CD4(+) direct allorecognition is important in the development of obliterative airway disease.

Calcium ionophore activation of chronic myelogenous leukemia progenitor cells into dendritic cells is mediated by calcineurin phosphatase.

We have previously demonstrated that Ph+ myeloid progenitor cells of patients with chronic myeloid leukemia (CML) can acquire characteristics of mature dendritic cells (DC) following calcium mobilization with calcium ionophore (A23187, CI). In this study we characterize the intracellular signaling pathway by which CI induces the acquisition of DC features in these leukemic cells. CI-induced activation of CML cells is attenuated by the calcineurin phosphatase inhibitor cyclosporin A (CsA) as well as the calmodulin (CaM) antagonist W-7. These cause ablation of both the CI-induced immunophenotypic expression of DC markers and immunostimulatory properties in mixed leukocyte responses (MLR). Minimal blocking effect was observed when Ca(2+)/CaM kinase II (281-301) inhibitor was added to the cultures. These findings suggest a Ca(2+)-dependent mechanism for the CI-induced activation of CML cells into antigen-presenting cells (APC), which is primarily mediated through the CaM/calcineurin pathway.

Replacement of graft-resident donor-type antigen presenting cells alters the tempo and pathogenesis of murine cardiac allograft rejection.

BACKGROUND: Graft-resident antigen presenting cells (APCs) are potent stimulators of the alloresponse. To test whether replacement of graft-resident donor-type APCs with those of recipient-type alters allorecognition and the pathogenesis of both acute and chronic rejection, we created chimeric hearts for transplantation into naive recipients. METHODS: To replace donor-type APCs with those of recipient-type, chimeric animals were created by bone marrow transplantation (BMT) in fully allogeneic mouse and rat strain combinations. The degree of APC replacement in chimeric organs was assessed phenotypically and functionally. Chimeric hearts were transplanted heterotopically into untreated recipients. RESULTS: Flow cytometric and immunohistochemical analysis did not detect residual bone marrow recipient-type APCs in mouse BMT chimeras. Although semi-quantitative reverse transcription polymerase chain reaction detected 0.001-0.01% residual cells, APCs isolated from chimeric organs were functionally unable to stimulate donor-type cells. When transplanted into naive recipients, chimeric mouse hearts had significantly prolonged survival but were nevertheless rejected acutely. Similar results were obtained in the ACI --> LEW rat strain combination. However, in the PVG --> DA rat model, the majority of chimeric hearts survived >100 days and all long-surviving hearts developed cardiac allograft vasculopathy. CONCLUSIONS: BMT leads to near complete replacement of organ-resident APCs. The virtual absence of donor-type APCs in chimeric hearts delays or prevents acute rejection in a strain-dependent manner. In contrast, this type of graft modification does not prevent cardiac allograft vasculopathy. This suggests that, although the CD4+ direct pathway may play a role in acute rejection, it is not essential for the development of chronic rejection in rodent cardiac allografts.

Changing presentation and management of neutropenic enterocolitis.

OBJECTIVE: To characterize the current clinical presentation and management of neutropenic enterocolitis. DESIGN: Retrospective review of records of oncology unit patients requiring general surgical consultation for abdominal complaints in a 1-year period. SETTING: Oncology unit of a tertiary care, university teaching hospital. PATIENTS AND INTERVENTIONS: Fourteen patients diagnosed as having neutropenic enterocolitis were managed conservatively with operation reserved for failure of conservative therapy. MAIN OUTCOME MEASURES: Clinical data from patients at the time of presentation and during treatment for neutropenic enterocolitis. RESULTS: All 14 patients diagnosed as having neutropenic enterocolitis were receiving chemotherapy for solid tumors or leukemias. Seven patients were undergoing stem cell or autologous bone marrow transplantation. Presenting symptoms and physical examination findings were nonspecific. All patients except one had neutropenia at the time of diagnosis. Computed tomographic scans of the abdomen were the most useful confirmatory study for the diagnosis of neutropenic enterocolitis. All patients except one had resolution of neutropenic enterocolitis with conservative therapy. One patient whose course of conservative management failed had protracted neutropenia and required operation for resection of bowel with full-thickness necrosis. CONCLUSIONS: Neutropenic enterocolitis has evolved from a complication of patients with leukemia to a disease of patients receiving high-dose chemotherapy for many malignancies, solid as well as hematologic. Diagnosis of neutropenic enterocolitis continues to be a challenge, as patients typically present with nonspecific gastrointestinal tract symptoms. Neutropenia and computed tomographic scan findings are useful adjuncts in diagnosing neutropenic enterocolitis. Timely conservative treatment frequently allows resolution of neutropenic enterocolitis without operation.

Surgical antibiotic prophylaxis and Clostridium difficile toxin positivity.

OBJECTIVE: To examine a possible relationship between prophylactic antibiotic therapy (PAT) and the development of Clostridium difficile toxin (CDT) positivity. DESIGN: Retrospective case-control study. SETTING: Tertiary care medical center in New York, NY. PATIENTS: A total of 357 patients, admitted from November 1992 to April 1994, with positive. CDT assays. Of these, 23 patients (6%) received only PAT for elective surgical procedures. Thirty-nine patients were matched as controls for age, sex, and surgical procedure. MAIN OUTCOME MEASURES: Both CDT positivity and inappropriate use of PAT. RESULTS: Appropriate PAT was used in 26 (42%) of 62 patients (17% cases, 56% controls). The Mantel-Haenszel estimator for the summary odds ratio for the development of CDT positivity from inappropriate use of PAT was 5.1 (95% confidence interval, 1.10 to 23.64). Main duration between the operation and the final antibiotic dose was significantly longer in the CDT-positive group compared with the control group (3.1 vs 1.7 days, P < .05). The length of hospital stay was significantly longer in the CDT-positive group compared with the control group (16.5 vs 10.2 days, P < .05). CONCLUSIONS: The prolonged use of PAT in elective surgical cases increases the risk of developing CDT positivity. The appropriate use of PAT could significantly reduce health costs by eliminating unnecessary doses of antibiotics, by decreasing the rate of CDT positivity, and by shortening the hospital stay. Restrictive policies may need to be implemented to prevent further antibiotic misuse.

An improved model for rat liver transplantation including arterial reconstruction and simplified microvascular suture techniques.

For experimental liver transplantation in the rat, the models that have been used most frequently do not include reconstruction of the arterial blood supply to the liver. In these procedures, specially developed cuff anastomoses rather than the conventional microvascular suture technique are used almost exclusively in the recipient operation, so that the anhepatic time is minimized. In this study the technical details of an improved rat model for orthotopic liver transplantation are described. During the donor operation in this experimental method, the liver is prepared with an arterial pedicle that includes the abdominal segment of the aorta, permitting perfusion in situ of the portal vein as well as the hepatic artery. The transplantation of the excised donor organ into the recipient site is carried out with simplified microvascular suture techniques and includes reconstruction of the arterial supply to the liver. Anastomosis of the bile duct is accomplished by choledocho-choledochostomy with a splint technique and supplemental suturing. For the entire procedure, magnifying glasses with 2- to 2.5-fold magnification are sufficient. When this technique has been mastered, the average duration of the anhepatic phase is about 20 min, well below the critical 30-min limit for survival of the experimental animals. As proficiency increased, the perioperative mortality was reduced to 9.2% (n = 130). With the combination of portal and arterial in situ flushing during the donor operation and the rearterialization of the transplant during the recipient operation, the clinical conditions can be approximated more closely than is possible when the transplanted rat liver is supplied only by the portal vein. Use of microvascular suture techniques, without cuff anastomoses, reduces the need for ex situ handling of the donor organ.