Christina L. Kaufman

Mutations in proto-oncogene GFI1 cause human neutropenia and target ELA2

Mice lacking the transcriptional repressor oncoprotein Gfi1 are unexpectedly neutropenic. We therefore screened GFI1 as a candidate for association with neutropenia in affected individuals without mutations in ELA2 (encoding neutrophil elastase), the most common cause of severe congenital neutropenia (SCN; ref. 3). We found dominant negative zinc finger mutations that disable transcriptional repressor activity. The phenotype also includes immunodeficient lymphocytes and production of a circulating population of myeloid cells that appear immature. We show by chromatin immunoprecipitation, gel shift, reporter assays and elevated expression of ELA2 in vivo in neutropenic individuals that GFI1 represses ELA2, linking these two genes in a common pathway involved in myeloid differentiation.

Graft vasculopathy in clinical hand transplantation.

Allogeneic hand transplantation is now a clinical reality. While results have been encouraging, acute rejection rates are higher than in their solid‐organ counterparts. In contrast, chronic rejections, as defined by vasculopathy and/or fibrosis and atrophy of skin and other tissues, as well as antibody mediated rejection, have not been reported in a compliant hand transplant recipient. Monitoring vascularized composite allograft (VCA) hand recipients for rejection has routinely involved punch skin biopsies, vascular imaging and graft appearance. Our program, which has transplanted a total of 6 hand recipients, has experience which challenges these precepts. We present evidence that the vessels, both arteries and veins may also be a primary target of rejection in the hand. Two of our recipients developed severe intimal hyperplasia and vasculopathy early post‐transplant. An analysis of events and our four other patients has shown that the standard techniques used for surveillance of rejection (i.e. punch skin biopsies, DSA and conventional vascular imaging studies) are inadequate for detecting the early stages of vasculopathy. In response, we have initiated studies using ultrasound biomicroscopy (UBM) to evaluate the vessel wall thickness. These findings suggest that vasculopathy should be a focus of frequent monitoring in VCA of the hand.

Outcomes of the First 2 American Hand Transplants at 8 and 6 Years Posttransplant

PURPOSE The feasibility of hand allotransplantation has been demonstrated. The purpose of the article is to report the (1) functional return, (2) psychosocial outcomes, (3) clinical and histological assessment for rejection, (4) complications, and (5) graft survival in the 2 American hand transplant recipients. METHODS We present 2 patients 106 and 81 months, respectively, after unilateral transplantation of an allogeneic hand and forearm. We analyzed clinical course, number of rejection episodes, adverse events, function of the allograft, and quality of life. Clinical laboratory results, biopsy histology, and patient clinical examinations were used to compare the clinical course. Standard hand function tests were used to evaluate function. Psychological interviews were used to assess acceptance and quality of life. RESULTS Our patients have allograft survival with improvements in intrinsic muscle activity, total active motion and return of functional grip, pinch strength, and sensibility. Rejection episodes were restricted primarily to the first 6 months after transplantation, and all responded to treatment. The major posttransplantation complications were a cytomegalovirus infection in patient 1 and osteonecrosis of the hip requiring both hips to be replaced, 1 at year 4 and the other at year 6, as well as transient immunosuppression-related diabetes in patient 2. Recently we have weaned both patients off maintenance steroids. Current Carroll scores are fair for patient 1 (72/99) and fair for patient 2 (55/99), although patient 2 has not had good recovery of intrinsic function. Both patients are back at work and report an excellent quality of life at nearly 9 and 7 years, respectively, after transplantation. CONCLUSIONS Our intermediate long-term results of hand transplants have demonstrated functional return similar to that of replants. Graft survival and quality of life after hand transplantation has far exceeded initial expectations. We conclude that allogeneic hand transplant is feasible and holds promise as a treatment modality for catastrophic upper extremity loss. TYPE OF STUDY/LEVEL OF EVIDENCE Therapeutic IV.

Hand transplantation in the United States: experience with 3 patients.

BACKGROUND Composite tissue allotransplantation (CTA) is a newly emerging field of transplantation that involves the simultaneous transfer of multiple tissues with differing antigenicity. Hand transplantation, the most widely recognized form of CTA, aims to improve function and the quality of life of upper limb amputees. METHODS In 1999, an institutional review board-approved hand transplantation protocol was implemented at the Jewish Hospital, University of Louisville. Suitable patients were evaluated and underwent hand transplantation. The surgical technique was akin to that used in limb reimplantation, and the immunosuppression protocol used was similar to renal transplantation. RESULTS Between 1999 and 2006, 3 patients underwent hand transplantation at our center. Although episodes of acute rejection were seen in all patients during the early postoperative period, only 1 immunologic event occurred after the first year. Graft function improved with time period. Carroll test scores were superior to those recorded with a prosthesis at the end of 1 year. Additionally, recovery of protective sensation was seen in all 3 patients and limited discriminatory sensation in 2. Complications related to immunosuppression have included cytomegalovirus infection in 2 patients, diabetes in 1, hyperlipidemia in 2, and osteonecrosis in 1. At a follow-up of 8, 6, and 1 year(s), all the recipients are healthy and have returned to a productive life. CONCLUSIONS The long-term success reported here should encourage wider application of the CTA in general and hand transplantation in particular. Methods of minimizing long-term immunosuppression need to be pursued.

Superparamagnetic iron oxide particles transactivator protein-fluorescein isothiocyanate particle labeling for in vivo magnetic resonance imaging detection of cell migration: Uptake and durability

Conjugation of dextran-coated superparamagnetic iron oxide (SPIO) particles with transactivator protein (Tat)-peptide and fluorescein isothiocyanate (FITC) allows cells to readily uptake SPIO particles. This makes possible high-resolution, real-time imaging of these cells by magnetic resonance imaging (MRI). First, we need to understand how various subpopulations take up and maintain SPIO particles. In this report, we have focused on differences in T cells, B cells, and macrophages with respect to cross-linked (CL)-SPIO Tat-FITC particle uptake over 72 hours. We have found that cells quickly take up the particles and that the bead loss that does occur is not related to cell death or apoptosis. In contrast with reports in the literature, we have observed migration of the Tat-peptide conjugates primarily to the cytoplasm rather than the nucleus.

MRI detection of macrophages labeled using micrometer-sized iron oxide particles.

To evaluate cellular labeling of immune cells using micron‐sized iron oxide particles (MPIOs) and evaluate the MR relaxivity and MRI detection of the labeled cells.

Mixed xenogeneic chimerism (mouse + rat → mouse) to induce donor-specific tolerance to sequential or simultaneous islet xenografts

We previously reported that donor-specific rat islet xenografts were accepted by fully xenogeneic (rat-->mouse) chimeras when the islets were transplanted at least six weeks following reconstitution. The purpose of the present study was to examine whether a similar outcome would occur for mixed xenogeneic (mouse+rat-->mouse) chimeras if the islets were placed coincident with the time of bone marrow infusion. As with fully xenogeneic chimeras (rat-->mouse), synchronous donor-specific F344 rat (Rt1A1) islet xenografts were significantly prolonged (MST > 139 days) in mixed xenogeneic (mouse+rat-->mouse) chimeras, while MHC-disparate third-party WF rat (Rt1Au) grafts were rejected (MST = 21.2 days). The transplanted donor-specific islets were functional to maintain euglycemia and they were regulated in function to respond to a glucose challenge. For potential clinical application, it would be of obvious benefit if the islet xenografts could be placed at the time of bone marrow transplantation. We therefore performed similar studies using donor islets administered simultaneously with bone marrow. Donor-specific islet xenografts were permanently accepted by all mouse recipients (n = 5). When MHC-disparate third-party rat islets were transplanted, only 3 of 8 islet xenografts were rejected; the other 5 remained functional from 77 to 90 days posttransplantation. Although prolonged and functional, the MHC-disparate islets that were accepted exhibited histologic evidence of fibrosis and rejection, while those matched to the donor did not. These data therefore suggest that donor-specific islet xenografts are permanently accepted if placed simultaneously or sequentially following mixed xenogeneic bone marrow reconstitution.

A New Option for Amputees: Transplantation of the Hand

The permanent loss of a hand or limb results in significant challenges. A number of options are available to individuals who must deal with this loss, including reconstructive surgery using a persons own tissue to repair the damage or the use of prosthetic devices. We present an update on the most recent addition to the list of options, namely, composite tissue allotransplantation (CTA). In this procedure, tissue to repair the loss is taken from deceased donors who are giving hearts, kidneys, and tissue for transplantation. We report on the worlds longest follow-up of CTA of the hand, as well as four other American hand transplant recipients. In very select patient populations, we propose that transplantation is now a clinical option for amputees.

Allogeneic chimerism induces donor-specific tolerance to simultaneous islet allografts in nonobese diabetic mice

BACKGROUND Nonobese diabetic (NOD) mice develop a systemic autoimmune disease that resembles type I diabetes in human beings. Previous studies in NOD mice have shown that transplantation of bone marrow from normal donors to lethally conditioned recipients prevented the development of diabetes (B10.BR-->NOD, BALB/c-->NOD). The focus of the present study was to examine whether donor-specific transplantation tolerance for islet allografts could be achieved if islet transplantations were performed coincident with the time of bone marrow infusion in diabetic NOD mice. Moreover, the transplanted islets were evaluated for evidence of recurrent autoimmunity. METHODS Female NOD mice were followed until autoimmune diabetes occurred (urine glucose, to +; blood glucose level, 300 mg/dl or greater). Diabetic NOD mice were irradiated with 950 cGy total body irradiation and received 30 x 10(6) untreated B10.BR bone marrow cells (B10.BR-->NOD). A simultaneous islet allograft was placed under the renal capsule within 24 hours after infusion of the bone marrow cells. Mice were monitored by means of blood glucose levels, and histologic analyses were performed on the transplanted islet. RESULTS Islet allografts genetically matched to the bone marrow donor were significantly prolonged (n = 5; mean survival time, 206 days or more) and showed no evidence for chronic rejection or recurrent insulitis, whereas major histocompatibility complex-disparate third-party allografts were rejected (n = 3; mean survival time, 37 days) and exhibited lymphocytic infiltration compatible with rejection on histologic evaluation. CONCLUSIONS These data suggest that permanent donor-specific tolerance to islet allografts placed simultaneously with bone marrow transplantation can be achieved in diabetic NOD mice. Moreover, recurrent autoimmune destruction of the pancreatic tissue is prevented by the bone marrow chimerism.

Monitoring and long-term outcomes in vascularized composite allotransplantation.

Purpose of reviewThe field of vascularized composite allotransplantation (VCA) is young, with less than 150 transplants worldwide. However, we now possess as much as 14 years of clinical follow-up. There are similarities and distinct differences between solid-organ transplantation (SOT) and VCA. This review will summarize how VCA recipients are monitored, outcomes observed, and what aspects are unique to VCA. Recent findingsOf about 90 documented cases, 10% of VCA recipients are out more than 10 years and 14% are out 5 or more years. There have been both graft losses and patient mortality. In most cases, these losses have been acute, most within the first year, and all within 3 years. Unlike SOT, VCA grafts function well during severe rejection. Chronic rejection-like sequelae are less frequent than in SOT, but do appear. Immunosuppression ranges from standard protocols to novel trials aimed at immunosuppression minimization. Patient selection greatly affects the outcome. Graft loss after year 1 is associated with compliance issues. SummaryFunctional outcomes have exceeded expectations. VCA recipients enjoy a quality of life not achievable with conventional reconstruction. Outstanding long-term results of more than a decade have been achieved. Monitoring of VCA patients will require new strategies to incorporate external visualization and effects of environment on rejection. Graft loss has occurred early, suggesting we focus improvement on this time period. More follow-up is needed to determine the rates and targets of chronic rejection, and the characteristics of VCA unique to face vs. hand transplantation.