[Lung transplantation: current situation and developments]. / Lungentransplantation: aktueller Stand und Entwicklungen.

Lung transplantation is currently the gold standard treatment for end-stage lung diseases. Advances in the preservation of donor lungs, the surgical technique and immunosuppressive therapy have led to lung transplantation now being a routine procedure. Nevertheless, the shortage of donor organs, the acute and particularly chronic lung allograft dysfunction (CLAD) still represent major challenges even in experienced centers. Research in this area is still necessary to improve the long-term survival of lung recipients.

The effects of brain death and ischemia on tolerance induction are organ-specific.

We have previously shown that 12 days of high-dose calcineurin inhibition induced tolerance in MHC inbred miniature swine receiving MHC-mismatched lung, kidney, or co-transplanted heart/kidney allografts. However, if lung grafts were procured from donation after brain death (DBD), and transplanted alone, they were rejected within 19-45 days. Here, we investigated whether donor brain death with or without allograft ischemia would also prevent tolerance induction in kidney or heart/kidney recipients. Four kidney recipients treated with 12 days of calcineurin inhibition received organs from donors rendered brain dead for 4 hours. Six heart/kidney recipients also treated with calcineurin inhibition received organs from donors rendered brain dead for 4 hours, 8 hours, or 4 hours with 4 additional hours of cold storage. In contrast to lung allograft recipients, all isolated kidney or heart/kidney recipients that received organs from DBD donors achieved long-term survival (>100 days) without histologic evidence of rejection. Proinflammatory cytokine gene expression was upregulated in lungs and hearts, but not kidney allografts, after brain death. These data suggest that the deleterious effects of brain death and ischemia on tolerance induction are organ-specific, which has implications for the application of tolerance to clinical transplantation.

Effects of Lung Cotransplantation on Cardiac Allograft Tolerance Across a Full Major Histocompatibility Complex Barrier in Miniature Swine.

A 12-day course of high-dose tacrolimus induces tolerance of major histocompatibility complex-mismatched lung allografts in miniature swine but does not induce tolerance of heart allografts unless a kidney is cotransplanted. To determine whether lungs share with kidneys the ability to induce cardiac allograft tolerance, we investigated heart-lung cotransplantation using the same induction protocol. Hearts (n = 3), heart-kidneys (n = 3), lungs (n = 6), and hearts-lungs (n = 3) were transplanted into fully major histocompatibility complex-mismatched recipients treated with high-dose tacrolimus for 12 days. Serial biopsy samples were used to evaluate rejection, and in vitro assays were used to detect donor responsiveness. All heart-kidney recipients and five of six lung recipients demonstrated long-term graft survival for longer than 272 days, while all heart recipients rejected their allografts within 35 days. Tolerant recipients remained free of alloantibody and showed persistent donor-specific unresponsiveness by cell-mediated lympholysis/mixed-lymphocyte reaction. In contrast, heart-lung recipients demonstrated rejection of both allografts (days 47, 55, and 202) and antidonor responsiveness in vitro. In contrast to kidneys, lung cotransplantation leads to rejection of both heart and lung allografts, indicating that lungs do not have the same tolerogenic capacity as kidneys. We conclude that cells or cell products present in kidney, but not heart or lung allografts, have a unique capacity to confer unresponsiveness on cotransplanted organs, most likely by amplifying host regulatory mechanisms.

A novel approach to measuring cell-mediated lympholysis using quantitative flow and imaging cytometry.

In this study, we established a novel isotope-free approach for the detection of cell-mediated lympholysis (CML) in MHC defined peripheral blood mononuclear cells (PBMCs) using multiparameter flow and imaging cytometry. CML is an established in vitro assay to detect the presence of cytotoxic effector T-lymphocytes precursors (CTLp). Current methods employed in the identification of CTLp in the context of transplantation are based upon the quantification of chromium ((51)Cr) released from target cells. In order to adapt the assay to flow cytometry, primary porcine PBMC targets were labeled with eFluor670 and incubated with major histocompatibility complex (MHC) mismatched effector cytotoxic lymphocytes (CTLs). With this method, we were able to detect target-specific lysis that was comparable to that observed with the (51)Cr-based assay. In addition, the use of quantitative cell imaging demonstrates the presence of accessory cells involved in the cytotoxic pathway. This innovative technique improves upon the standard (51)Cr release assay by eliminating the need for radioisotopes and provides enhanced characterization of the interactions between effector and target cells. This technique has wide applicability to numerous experimental and clinical models involved with effector-cell interactions.

Innovative cold storage of donor organs using the Paragonix Sherpa Pak ™ devices.

INTRODUCTION: Currently, the gold standard for donor organ preservation in clinical organ transplantation consists of 3 plastic bags and an ice box. The first plastic bag includes the organ itself immersed in preservation solution (e.g. Celsior). This bag is put in a second bag filled with saline, and then these two are put in a third bag filled with saline which is then put in the ice box.  The disadvantage of this method is that the organ usually gets too cold. It has been shown that the theoretical perfect temperature for organ preservation is 4°C - 8°C. While higher temperatures lead to hypoxic injury of the organ because the metabolism is not decreased efficiently, lower temperatures than 4°C increase the risk of cold injury with protein denaturation. In the current study, we investigated a device that keeps the organ temperature consistently in the desired range of 4°C - 8°C and can potentially decrease cold injury to donor organs. METHODS: Three different ex vivo studies were performed with the Paragonix Sherpa Pak™ devices: 1) the temperature of the fluid-filled device was measured for up to 30 hours at an outside temperature set at 22°C, 2) the temperature of the fluid-filled device was measured for up to 30 hours at extreme outside temperatures set at -8°C and 31°C, 3) the temperature of a pig heart attached to the device was measured up to 12 hours. RESULTS: All studies showed that the Paragonix Sherpa Pak™ can keep the temperature of the heart consistently between 4° and 8°C. CONCLUSIONS: The Paragonix Sherpa Pak™ device may decrease cold injury of donor organs by maintaining the temperature consistently between 4°C and 8°C and therefore may decrease primary graft failure after organ transplantation.

Kidney-induced cardiac allograft tolerance in miniature swine is dependent on MHC-matching of donor cardiac and renal parenchyma.

Kidney allografts possess the ability to enable a short course of immunosuppression to induce tolerance of themselves and of cardiac allografts across a full-MHC barrier in miniature swine. However, the renal element(s) responsible for kidney-induced cardiac allograft tolerance (KICAT) are unknown. Here we investigated whether MHC disparities between parenchyma versus hematopoietic-derived "passenger" cells of the heart and kidney allografts affected KICAT. Heart and kidney allografts were co-transplanted into MHC-mismatched recipients treated with high-dose tacrolimus for 12 days. Group 1 animals (n = 3) received kidney and heart allografts fully MHC-mismatched to each other and to the recipient. Group 2 animals (n = 3) received kidney and heart allografts MHC-matched to each other but MHC-mismatched to the recipient. Group 3 animals (n = 3) received chimeric kidney allografts whose parenchyma was MHC-mismatched to the donor heart. Group 4 animals (n = 3) received chimeric kidney allografts whose passenger leukocytes were MHC-mismatched to the donor heart. Five of six heart allografts in Groups 1 and 3 rejected <40 days. In contrast, heart allografts in Groups 2 and 4 survived >150 days without rejection (p < 0.05). These data demonstrate that KICAT requires MHC-matching between kidney allograft parenchyma and heart allografts, suggesting that cells intrinsic to the kidney enable cardiac allograft tolerance.

Induction of cardiac allograft tolerance across a full MHC barrier in miniature swine by donor kidney cotransplantation.

We have previously shown that tolerance of kidney allografts across a full major histocompatibility complex (MHC) barrier can be induced in miniature swine by a 12-day course of high-dose tacrolimus. However, that treatment did not prolong survival of heart allografts across the same barrier. We have now tested the effect of cotransplanting an allogeneic heart and kidney from the same MHC-mismatched donor using the same treatment regimen. Heart allografts (n = 3) or heart plus kidney allografts (n = 5) were transplanted into MHC-mismatched recipients treated with high-dose tacrolimus for 12 days. As expected, all isolated heart allografts rejected by postoperative day 40. In contrast, heart and kidney allografts survived for >200 days with no evidence of rejection on serial cardiac biopsies. Heart/kidney recipients lost donor-specific responsiveness in cell-mediated lympholysis and mixed-lymphocyte reaction assays, were free of alloantibody and exhibited prolonged survival of donor, but not third-party skin grafts. Late (>100 days) removal of the kidney allografts did not cause acute rejection of the heart allografts (n = 2) and did not abrogate donor-specific unresponsiveness in vitro. While kidney-induced cardiac allograft tolerance (KICAT) has previously been demonstrated across a Class I disparity, these data demonstrate that this phenomenon can also be observed across the more clinically relevant full MHC mismatch. Elucidating the renal element(s) responsible for KICAT could provide mechanistic information relevant to the induction of tolerance in recipients of isolated heart allografts as well as other tolerance-resistant organs.

[Experimental quality control of biopsy cannulas]. / Experimentelle Qualitätskontrolle von Biopsiekanülen.

To examine the quality and comparability of biopsy needles. 310 biopsy needles of 40 types were examined microscopically and their suitability for penetrating tissues were studied. For this purpose the pressure required to penetrate a phantom was evaluated. The results were correlated with the diameter, design and construction. 50 of the 310 needles (16%) showed faults on microscopic examination. Measurements of identical needle types within a single production run showed variations in penetration pressures up to 330% (measured as force required for penetration), the average variation was 88%. The results show that the quality of biopsy needles is not constant and leaves something to be desired. To what extent variations in quality affect the biopsy material cannot be estimated. Nevertheless, the present results indicate that stricter quality control of biopsy needles is desirable.