Lymphodepletive effects of rabbit anti-pig thymocyte globulin in neonatal swines.

Lymphodepletive agents play important role in different clinical applications or experimental transplant studies. In order to facilitate preclinical pediatric transplant studies, we have developed the rabbit anti-pig thymocyte globulin (pATG) and studied its effects in neonatal swines. In vitro assays showed that pATG can bind to lymphocytes and neutrophils in a dose-dependent manner and lyse peripheral blood mononuclear cells by apoptosis and complement-dependent cytotoxicity. In vivo, pATG as a monotherapy was administered at different doses (2.5, 5, 20, 40 and 80mg/kg) in newborn pigs. Results showed that pATG induced a dose-dependent but transient T-cell depletion in peripheral blood. Lymphodepletion was also observed in lymph nodes, spleen and thymus. Pharmacokinetic studies showed dose-related cell-bound pATG on lymphocytes, as well as the presence of free pATG in the serum. Both cell-bound and free pATG levels decreased gradually after administration. Interestingly, adjuvant mycophenolate mofetil (MMF) given at 1g/m2/day for 1week successfully maintained pATG-induced T-cell depletion. In conclusion, pATG administration can cause transient T-cell depletion in neonatal pigs and this effect can be maintained by MMF. Therefore, we have developed an original immunosuppressive regimen that can be used for transplantation studies in swine model.

Face graft? Extrapolation of facial allotransplantation to children.

The possibility to imagine a vascularized composite allotransplantation for disfigured children is felt more critical than for adults non on technical point of view but in terms of indications and justifications. The question is not about surgery. It is related to the pathologies themselves for which transplant could be suitable. Moreover the procurement of face transplant will be more difficult because of immunologic criteria but also age and phototype. Specificity of the newborn malformative face is usually not only a question of tissue defect. It is reasonably not an indication for VCA. It should be added that nothing is known about the future of transplantation in terms of duration but also morbidities due to immunosuppression. Indications are rather negative. To rise the question of VCA for children has a double benefit. The first is to point out that surgical innovation often arise from a non imaginable or non imagined clinical situation. The second is the question of VCA in newborn regarding the tolerance.

Chronic Rejection in Human Vascularized Composite Allotransplantation (Hand and Face Recipients): An Update.

Vascularized composite tissue allografts (VCA) have become a viable option to restore severely damaged parts of the body that cannot be repaired with conventional surgical techniques. Acute rejection develops frequently in the early postgraft period both in human and experimental VCA, but the possibility of human VCA to undergo chronic rejection (CR) remained initially unknown. The experience gained over the years shows that, similar to solid organ transplants (SOT), human VCA can also develop CR. Chronic rejection is clinically mostly apparent on the skin and targets preferentially skin and deep vessels, leading, as in SOT, to graft vasculopathy and often to graft loss. Dermal sclerosis and adnexal atrophy are additional features of CR. The pathogenetic immune mechanisms involved (cell-mediated versus humoral) remain incompletely known. The changes of CR can be detected with skin and deep tissue biopsies. Modern in vivo imaging tools can detect vascular narrowing and have the advantage of being noninvasive. However, the diagnosis and treatment of CR remain challenging, as several important questions remain to be answered: a more accurate definition of CR in VCA is needed to establish criteria allowing an accurate and early diagnosis. The pathogenetic mechanisms of CR need to be better understood to allow more efficacious treatment. Favoring/triggering factors of CR need to be better known so that they can be avoided. As in SOT, there is a need for efficient tolerance-inducing protocols that will favor graft acceptance and (ideally) circumvent the necessity of lifelong immunosuppression.

Transplant Tolerance Induction in Newborn Infants: Mechanisms, Advantages, and Potential Strategies.

Although several tolerance induction protocols have been successfully implemented in adult renal transplantation, no tolerance induction approach has, as yet, been defined for solid organ transplantations in young infants. Pediatric transplant recipients have a pressing demand for the elaboration of tolerance induction regimens. Indeed, since they display a longer survival time, they are exposed to a higher level of risks linked to long-term immunosuppression (IS) and to chronic rejection. Interestingly, central tolerance induction may be of great interest in newborns, because of their immunological immaturity and the important role of the thymus at this early stage in life. The present review aims to clarify mechanisms and strategies of tolerance induction in these immunologically premature recipients. We first introduce the discovery and mechanisms of neonatal tolerance in murine experimental models and subsequently analyze tolerance induction in human newborn infants. Hematopoietic mixed chimerism in neonates is also discussed based on in utero hematopoietic stem cell (HSC) transplant studies. Then, we review the recent advances in tolerance induction approaches in adults, including the infusion of HSCs associated with less toxic conditioning regimens, regulatory T cells/facilitating cells/mesenchymal stem cells transplantation, costimulatory blockade, and thymus manipulation. Finally, IS withdrawal in pediatric solid organ transplant is discussed. In conclusion, the establishment of transplant tolerance induction in infants is promising and deserves further investigations. Future studies could focus on the selection of patients, on less toxic conditioning regimens, and on biomarkers for IS minimization or withdrawal.

Premalignant and Malignant Skin Lesions in Two Recipients of Vascularized Composite Tissue Allografts (Face, Hands).

Recipients of solid organ transplants (RSOT) have a highly increased risk for developing cutaneous premalignant and malignant lesions, favored by the lifelong immunosuppression. Vascularized composite tissue allografts (VCA) have been introduced recently, and relevant data are sparse. Two patients with skin cancers (one with basal cell carcinoma and one with squamous cell carcinomas) have been so far reported in this patient group. Since 2000 we have been following 9 recipients of VCA (3 face, 6 bilateral hands) for the development of rejection and complications of the immunosuppressive treatment. Among the 9 patients, one face-grafted recipient was diagnosed with nodular-pigmented basal cell carcinoma of her own facial skin 6 years after graft, and one patient with double hand allografts developed disseminated superficial actinic porokeratosis, a potentially premalignant dermatosis, on her skin of the arm and legs. Similar to RSOT, recipients of VCA are prone to develop cutaneous premalignant and malignant lesions. Prevention should be applied through sun-protective measures, regular skin examination, and early treatment of premalignant lesions.

Bilateral hand transplantation: Functional benefits assessment in five patients with a mean follow-up of 7.6 years (range 4-13 years).

Between January 2000 and July 2009, five adults who had suffered bilateral traumatic below-elbow amputations, received bilateral hand-forearm allografts performed by the Lyon team. We report the functional benefits achieved over a mean follow-up period of 7.6 years (range 4-13 years), up to December 31st, 2013. Clinical measurement is hampered by the lack of specific validated assessment tools, obliging us to use non-specific standardized evaluation means. Our assessment shows that the restoration of motion, strength, and sensibility are fair. Functional results (Carroll upper extremity function test, 400-point test, Activities of daily living) are good, as well as quality of life evaluation (RAND-36). Subjective and overall results explored with questionnaires - Disabilities of the Arm Shoulder and Hand (DASH), Hand Transplantation Score System (HTSS), are very good. Improvement was seen to continue during the first three years, and then tend to become stable. Continued efforts should be directed at designing comprehensive, condition-specific, reliable outcome measurement tools. Continuous monitoring and evaluation of patients is required to assess the long-term risk-benefit balance.

Oral cyclosporine A in neonatal swines for transplantation studies.

The purpose of this study is to define the optimal dose of oral cyclosporine A (CsA) microemulsion in newborn swine for transplantation studies and to describe its pharmacokinetics and acute renal effects in short-term administration. Thirteen neonatal pigs were randomized into four groups: one control and three groups with CsA administration at 4, 8 and 12 mg/kg/d for 15 days (D). Blood samples were collected on D 0, 2, 4, 9 and 14 to determine the changes of the CsA trough concentrations, the creatinine (Cr) and blood urea nitrogen (BUN) serum concentrations. On D 14, blood samples were collected every hour from 1 h to 10 h after CsA administration to determine the area under the curve (AUC). On D 15, kidneys were removed for histological analysis. We observed a stabilization of CsA trough concentrations from D 4 to D 14. On D 14, in the three treated groups, CsA trough concentrations were 687 ± 7, 1200 ± 77 and 2211 ± 1030 ng/ml, respectively; AUC (0-10 h) were 6721 ± 51 ng·h/ml in group 4 mg/kg/d, 13431 ± 988 ng·h/ml in group 8 mg/kg/d and 28264 ± 9430 ng·h/ml in group 12 mg/kg/d. Cr concentrations were not significantly different among the four groups; but compared to control group, BUN concentrations of the three treated groups increased significantly. CsA was well tolerated; neither acute, severe adverse event nor renal histological abnormality was observed. In conclusion, a 15-d course of oral CsA treatment ranged from 4 to 12 mg/kg/d is safe for newborn pigs, which need much lower CsA dose than adult pigs to reach comparable trough level and AUC. As immunosuppressive therapy in newborn pigs, we recommend a CsA dose of 4 mg/kg/d to achieve a trough blood concentration between 400 and 800 ng/ml.

Outcomes after bilateral hand allotransplantation: a risk/benefit ratio analysis.

BACKGROUND: The clinic era of composite tissue allotransplantation was inaugurated by hand allotransplantation in 1998, giving rise to many controversies and scepticism because of the lifelong immunosuppression, the unclear risk-benefit ratio, and the uncertain long-term functional results of the procedure. The aim of this study was to evaluate the outcomes and the risk/benefit balance in bilateral hand allotransplantation. METHODS: The study included 5 cases of bilateral hand allotransplantation performed in a single center, with a follow-up ranging from 3 to 13 years. The recipients (4 men, 1 woman) were young. The level of amputation was distal in all cases except for 2 patients amputated at the midforearm level. All the recipients initially received the same immunosuppressive treatment that included tacrolimus, mycophenolate mofetil, prednisone, and, for induction, antithymocyte globulins. RESULTS: Patient and graft survival was 100%. All recipients showed adequate sensorimotor recovery (protective and tactile sensitivity and partial recovery of intrinsic muscles), they were able to perform the majority of activities of daily living, and had a normal social life. Most complications occurred in the first posttransplant year and were successfully managed. All recipients experienced at least 1 episode of acute rejection, which was easily reversed by increasing oral steroid dose or by intravenous steroids, except for patient 3, who presented 6 episodes of acute rejection, the latest 2 treated with Campath-1H. CONCLUSIONS: Although bilateral hand transplantation may be a satisfactory treatment option for amputees, a careful selection of candidates and a rigorous evaluation of recipients after transplantation are imperative.

Methodological choices for the clinical development of medical devices.

Clinical evidence available for the assessment of medical devices (MDs) is frequently insufficient. New MDs should be subjected to high quality clinical studies to demonstrate their benefit to patients. The randomized controlled trial (RCT) is the study design reaching the highest level of evidence in order to demonstrate the efficacy of a new MD. However, the clinical context of some MDs makes it difficult to carry out a conventional RCT. The objectives of this review are to present problems related to conducting conventional RCTs and to identify other experimental designs, their limitations, and their applications. A systematic literature search was conducted for the period January 2000 to July 2012 by searching medical bibliographic databases. Problems related to conducting conventional RCTs of MDs were identified: timing the assessment, eligible population and recruitment, acceptability, blinding, choice of comparator group, and learning curve. Other types of experimental designs have been described. Zelen's design trials and randomized consent design trials facilitate the recruitment of patients, but can cause ethical problems to arise. Expertise-based RCTs involve randomization to a team that specializes in a given intervention. Sometimes, the feasibility of an expertise-based randomized trial may be greater than that of a conventional trial. Cross-over trials reduce the number of patients, but are not applicable when a learning curve is required. Sequential trials have the advantage of allowing a trial to be stopped early depending on the results of first inclusions, but they require an independent committee. Bayesian methods combine existing information with information from the ongoing trial. These methods are particularly useful in situations where the number of subjects is small. The disadvantage is the risk of including erroneous prior information. Other types of experimental designs exist when conventional trials cannot always be applied to the clinical development of MDs.

Composite tissue allotransplantation in newborns: a swine model.

BACKGROUND: Management of congenital limb aplasia or facial malformations could be improved by composite tissue allotransplantation (CTA), a technique that has never been performed in newborns. For this, however, the induction of donor-specific tolerance would be mandatory, as long-term immunosuppression is not acceptable in this non-lifesaving procedure. Induction of tolerance has been shown to be possible in a newborn CTA rat model but has never been tested in large-animal models. Our goals were to establish a model of CTA in newborn swine to see if tolerance could be obtained without immunosuppression and to assess rejection or tolerance properties via clinical and histologic examinations. MATERIALS AND METHODS: We applied a CTA heterotopic knee swine model. We performed two series of surgical procedures: Series 1 was 20 autografts in 6-day-old (1-10) 2,544 kg (1,140-4,060 kg) piglets; Series 2 was 10 allografts without immunosuppression between outbred animals aged 7.8 d (6-10) and weighing 2,770 kg (2,200-3,550 kg). RESULTS: In Series 1, six early deaths and two cases of vascular failure were observed. In Series 2, no spontaneous deaths were observed and all piglets presented clinical and histologic rejection. CONCLUSIONS: Our findings strongly suggest that newborn immunologic status is not sufficient for the development of tolerance in large animals without immunologic intervention. Complications and animal death after transplantation correlate with age and weight. Low rates for both vascular failure and postoperative death permit the use of this model in piglets weighing over 2 kg and aged more than 6 d for research on newborn CTA.

Fifteen years later: main lessons from composite tissue allografts.

Composite tissue allografts (CTA) are also called "reconstructive transplantation" as they are a valid alternative approach to repairing complex tissue defects. These procedures are still considered "experimental" and their therapeutic value remains to be validated. An immunosuppressive treatment similar to that used in solid organ transplantation allows CTA survival and function despite a high rate of acute rejection (AR) episodes. Clinical experience seems to confirm that skin is the most antigenic tissue and the first target of AR episodes, which are easy to reverse and do not seem to adversely influence graft survival and function when promptly treated. Chronic rejection can also occur in CTA, although its features are still unclear. Upper-extremity or face-transplanted patients show a relevant sensorimotor recovery. Patients are able to perform the majority of daily activities and to lead normal social lives. Global cortical remodeling occurs in the months following transplantation, reversing the functional reorganization induced by the amputation. Appropriation of the graft occurs in parallel with functional recovery. The patients' compliance is essential for the success of CTAs as well as careful recipient selection and patient follow-up to prevent complications of long-term immunosuppression.

First human face transplantation: 5 years outcomes.

BACKGROUND: The first human facial allotransplantation, a 38-year-old woman, was performed on November 27, 2005. The aesthetic aspect and functional recovery and the risk-to-benefit ratio are evaluated 5 years later. MATERIALS AND METHODS: The facial transplantation included nose, chin, part of cheeks, and lips. The immunosuppressive protocol included tacrolimus, mycophenolate mofetil, prednisone, and antithymocyte globulins. In addition, donor bone marrow cells were infused on days 4 and 11 after transplantation. RESULTS: The aesthetic aspect is satisfying. The patient has normal protective and discriminative sensibility. She showed a rapid motion recovery, which has remained stable for 3 years posttransplantation. She can smile, chew, swallow, and blow normally whereas pouting and kissing is still difficult. Phonation recovery was impressive therefore the patient can talk normally. Two episodes of acute rejection developed during the first year. Donor-specific anti-human leukocyte antigen antibodies were never detected. Five-year mucosal biopsy showed a slight perivascular inflammatory infiltrate while skin biopsy was normal. The main side effect of the immunosuppressive treatment was a progressive decrease in renal function, which improved after switching from tacrolimus to sirolimus. Moreover, she developed arterial hypertension, an increase in lipid levels, and in situ cervix carcinoma treated by conization. Since 2008, she showed mild cholangitis possibly caused by sirolimus. In September 2010, bilateral pneumopathy occurred and was successfully treated with antibiotics. CONCLUSION: Despite some long-term complications, which are similar to those reported after solid organ transplantation, the patient is satisfied of her new face and has normal social interaction.

World experience after more than a decade of clinical hand transplantation: update on the French program.

The first hand transplantation was realized in Lyon and the results achieved in this case showed the feasibility of the surgical technique, the efficacy of the immunosuppressive protocol, the limited adverse effects and the importance of a patient's compliance and rehabilitation to ensure graft viability and functional recovery. Based on these findings and the positive results achieved in other single hand transplants realized around the world the authors performed also the first double hand transplantation, then followed by other four cases. The recipients received the same immunosuppressive treatment including tacrolimus, prednisone, mycophenolate mofetil and antithymocyte globulins for induction, nevertheless they showed some episodes of acute rejection episodes which reversed after a prompt treatment. All the bilateral hand grafted patients showed a relevant sensorimotor recovery particularly of sensibility and activity of intrinsic muscles. They were able to perform the majority of daily activities and to lead a normal social life. On the basis of the authors' experience the results achieved in hand allotransplantation are very encouraging as major adverse effects due to surgery and immunosuppressive regimen did not occur and patients' quality of life improved considerably.

Self-renewal capacity of human epidermal Langerhans cells: observations made on a composite tissue allograft.

Epidermal Langerhans cells (LC) are dendritic, antigen-presenting cells residing within mammalian epidermis and mucosal epithelia. When massively depleted, they are replaced by cells of bone-marrow origin. However, their renewal within normal skin under steady-state conditions is not precisely known. We observed that epidermal LC within a human hand allograft remain stable in the long term (10 years) and are not replaced by cells of recipient's origin; furthermore, we observed a Langerhans cell in mitosis within the epidermis 8 years postgraft. These results show that under almost physiological conditions, human LC renew in the epidermis by local mitoses of preexisting cells.

The International Registry on Hand and Composite Tissue allotransplantation.

The International Registry on Hand and Composite Tissue Transplantation includes hand and face allotransplantations: 39 patients who received 57 upper extremity transplantations (18 bilateral and 21 single transplantations--17 right and 4 left allografted hands); and 15 cases of partial or total face allotransplantation. The recipients of upper extremity allotransplantations are: 32 males and 7 females, median age 32 years. In 57.9% of cases, the level of amputation was at the wrist. The followup periods range from 6 months to 13 years. The recipients of face allotransplantations are: 12 males and 3 females, median age 34 years. In the majority of cases, the deficit included cheek, nose, chin, lips, and perioral area. The patients presented impairment of swallowing, eating, and speaking. The follow-up periods range from 8 months to 6 years. In hand and face transplantation, the imunosuppressive therapy included: tacrolimus, mycophenolate mofetil, and steroids. Polyclonal or monoclonal antibodies were used for induction. Within the first post-transplant year, eighty-five percent of hand and face recipients experienced at least one episode of acute rejection, which was reversible when promptly treated. Side-effects included: opportunistic infections, metabolic complications, and malignancies. Hand-grafted patients developed protective sensibility: 90% of them tactile sensibility and 82.3% also a discriminative sensibility. Motor recovery enabled patients to perform most daily activities. Face-grafted patients improved their aesthetic aspects and enhanced some activities such as eating, drinking, and speaking, living a normal social life. Five upper allotransplantation losses occurred. One of these patients who underwent simultaneous face and bilateral hand transplantation died on day 65. Hand and face transplantations are successful procedures, however, careful evaluation of patients before and after transplantation, and their compliance are indispensable.

[The kidney]. / Le rein.

Whole-organ engineering of the kidney is particularly difficult because of its structural complexity and of the morphological and functional heterogeneity of renal cell types. As for other organs, research is currently focused on:--the matrix to support recellularization: synthetic, biodegradable or biological. Use of the extracellular matrix as a biological scaffold is the most promising approach. Rodent, porcine and rhesus monkey kidneys have been decellularized to obtain scaffolds with a preserved extracellular matrix and vasculature.--The source of cells for recellularization: embryonic stem cells, fetal cells, adult-derived stem cells, progenitor cells and adult-derived inducible, pluripotent stem cells have all been used. Nephron development results from mutual inducive interactions between the urethral bud and the metanephric mesenchyme, a process that can be reproduced in vitro. Ex-vivo "fabrication" of a kidney that could be implanted with no risk of rejection in patients with chronic renal failure appears ultimately feasible. Another area of research is the use of renal assistance devices--the bioartificial kidney--based on a bioreactor containing renal epithelial cells derived from tubules that maintain their reabsorptive, metabolic and endocrine functions. Phase II clinical trials have given encouraging results.