Effect of HO-1-modified BMMSCs on immune function in liver transplantation.

We examined whether haem oxygenase-1 (HO-1) could enhance the immunosuppressive effects of bone marrow mesenchymal stem cells (BMMSCs) on the rejection of transplanted liver allografts in rats. The animals were divided into three groups: the normal saline (NS) group, BMMSC group and HO-1/BMMSCs group. In vitro, the extraction, culture and HO-1 transfection of BMMSCs were performed. Mixed lymphocyte response (MLR) analysis of HO-1/BMMSCs efficacy was performed. The rejection model of orthotopic liver transplantation in rats was established when BMMSCs and HO-1/BMMSCs were transfused via the portal vein. To reduce research bias, we established an isogenic Liver transplantation model of (LEW → LEW) and (BN → BN), which can achieve tolerance. Changes in histopathology and liver function in the transplanted liver and changes in regulatory T cell (Tregs), natural killer (NK) cells and cytokines after transplantation were observed in the different groups. The severe acute rejection after liver transplantation on postoperative Day 10 was observed in the NS group. The BMMSC group showed strong protective effects against rejection within the first 10 days after transplantation, while HO-1/BMMSCs showed stronger effects on rejection than BMMSCs alone. In addition, the activity of natural killer (NK) cells decreased significantly, the levels of regulatory T cells (Tregs), interleukin-10 (IL-10) and transforming growth factor-ß (TGF-ß) increased significantly and the levels of interleukin-2 (IL-2), interleukin-6 (IL-6), interleukin-17 (IL-17), interleukin-23 (IL-23), tumour necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) decreased significantly in the HO-1/BMMSC group compared with the BMMSC group. HO-1/BMMSCs showed better immunosuppressive effects after liver transplantation than the other treatments. Our findings reveal that HO-1 can enhance the effects of BMMSCs on inhibiting acute rejection in orthotopic liver transplantation in rats.

Successful Double DIEP Syngeneic Transplantation across Monozygotic Twins for Total Back Reconstruction.

BACKGROUND: A 56-year-old woman presented with an extensive sarcoma requiring nearly total back resection. She had limited donor sites for reconstruction because of a previous laparotomy, but presented with a significantly larger, identical twin. Cancer has traditionally been considered a contraindication for vascularized composite allotransplantation; however, immunosuppression is potentially avoidable between monozygotic twins. METHODS: A preoperative genetic workup revealed 10/10 human leukocyte antigen homozygosity. Despite substantial phenotypic divergence in size and facial features, the sisters were genotypically identical. A two-stage, double deep inferior epigastric perforator transplant was planned for delayed reconstruction. At the first stage following the resection, an arteriovenous loop was performed to provide recipient vasculature to the back. At a second stage, the transplantation was performed. In addition, bilateral lumbar artery perforator flaps were created to reduce the length of the defect. Intraoperative steroid bolus and a short taper alone were used for immunosuppression. RESULTS: The resection resulted in a 22 × 29-cm specimen down to the spine. After a 4-day interval for permanent pathologic evaluation, the transplant was successfully transferred between twins. Two arteries and six veins were anastomosed to establish perfusion. Postoperatively, there have been no episodes of rejection or flap compromise at last follow-up (>36 months). CONCLUSIONS: This case represents one of the few vascularized composite allotransplantations between monozygotic twins, and the only reported successful vascularized composite allotransplantation for a recurrent cancer diagnosis. Oncologic safety depended on 100 percent histocompatibility to avoid immunosuppression. Limited patient donor sites precluded total autologous coverage, and a substantial size discrepancy between the twins favored a transplant.

Syngeneic Transplantation of Rat Olfactory Stem Cells in a Vein Conduit Improves Facial Movements and Reduces Synkinesis after Facial Nerve Injury.

BACKGROUND: Posttraumatic facial paralysis is a disabling condition. Current surgical management by faciofacial nerve suture provides limited recovery. To improve the outcome, the authors evaluated an add-on strategy based on a syngeneic transplantation of nasal olfactory stem cells in a rat model of facial nerve injury. The main readouts of the study were the recording of whisking function and buccal synkinesis. METHODS: Sixty rats were allocated to three groups. Animals with a 2-mm facial nerve loss were repaired with a femoral vein, filled or not with olfactory stem cells. These two groups were compared to similarly injured rats but with a faciofacial nerve suture. Olfactory stem cells were purified from rat olfactory mucosa. Three months after surgery, facial motor performance was evaluated using video-based motion analysis and electromyography. Synkinesis was assessed by electromyography, using measure of buccal involuntary movements during blink reflex, and double retrograde labeling of regenerating motoneurons. RESULTS: The authors' study reveals that olfactory stem cell transplantation induces functional recovery in comparison to nontransplanted and faciofacial nerve suture groups. They significantly increase (1) maximal amplitude of vibrissae protraction and retraction cycles and (2) angular velocity during protraction of vibrissae. They also reduce buccal synkinesis, according to the two techniques used. However, olfactory stem cell transplantation did not improve axonal regrowth of the facial nerve, 3 months after surgery. CONCLUSIONS: The authors show here that the adjuvant strategy of syngeneic transplantation of olfactory stem cells improves functional recovery. These promising results open the way for a phase I clinical trial based on the autologous engraftment of olfactory stem cells in patients with a facial nerve paralysis.

A Comparison of Immune Responses Exerted Following Syngeneic, Allogeneic, and Xenogeneic Transplantation of Mesenchymal Stem Cells into the Mouse Brain.

Due to their multifactorial aspects, mesenchymal stem cells (MSCs) have been widely established as an attractive and potential candidate for the treatment of a multitude of diseases. A substantial number of studies advocate that MSCs are poorly immunogenic. In several studies, however, immune responses were observed following injections of xenogeneic donor MSCs. In this study, the aim was to examine differences in immune responses exerted based on transplantations of xenogeneic, syngeneic, and allogeneic MSCs in the wild-type mouse brain. Xenogeneic, allogeneic, and syngeneic MSCs were intracerebrally injected into C57BL/6 mice. Mice were sacrificed one week following transplantation. Based on immunohistochemical (IHC) analysis, leukocytes and neutrophils were expressed at the injection sites in the following order (highest to lowest) xenogeneic, allogeneic, and syngeneic. In contrast, microglia and macrophages were expressed in the following order (highest to lowest): syngeneic, allogeneic, and xenogeneic. Residual human MSCs in the mouse brain were barely detected after seven days. Although the discrepancy between leukocytes versus macrophages/microglia infiltration should be resolved, our results overall argue against the previous notions that MSCs are poorly immunogenic and that modulation of immune responses is a prerequisite for preclinical and clinical studies in MSC therapy of central nervous system diseases.

Coberturas transitorias en quemaduras / Temporary coverage in burns

En numerosas ocasiones, los pacientes con quemaduras profundas recientemente sometidos a escarectomías no pueden recibir una cobertura inmediata con autoinjertos debido a las condiciones locales del lecho de la herida, a la no disponibilidad de áreas donantes o porque el procedimiento en sí mismo pueda resultar arriesgado para la supervivencia del paciente. En estas circunstancias es necesario cubrir temporalmente la herida para mantener su viabilidad y reducir las infecciones y el estrés metabólico secundarios a la pérdida de fluidos y al dolor. Los homoinjertos están considerados como la mejor cobertura transitoria, pero su disponibilidad y costo hacen que su uso sea limitado; en segundo lugar, y aunque su calidad es inferior a la de los homoinjertos, los heteroinjertos proveen también una cobertura cutánea adecuada en muchos de estos casos. Asistimos en la actualidad a un desarrollo constante de productos biosintéticos útiles para este mismo fin; sin embargo, hasta la fecha, no existe ningún producto que podamos considerar como patrón o modelo de elección. Este artículo revisa los productos actualmente dispo¬nibles y las situaciones clínicas en las cuales pueden ser utilizados

Sometimes excised burn wounds cannot be covered immediately with autologous skin grafts, due to local wound conditions, unavailability of donor areas, or because the procedure itself may be risky for patient sur¬vival. In these circumstances a temporary coverage is desirable to maintain wound viability, reduce infections, metabolic stress and pain. Allografts have always been considered the best temporary coverage, but availability and cost are of concern. Xenografts can provide temporary coverage, even though its quality is clearly inferior to allograft. Although there is a constant evolution in the development of skin substitutes, no single product can be considered as the gold standard. This article reviews currently available products and clinical situations in which they can be used

Intravenously administered contact allergens coupled to syngeneic erythrocytes induce in mice tolerance rather than effector immune response.

Constantly increasing prevalence of allergic diseases determines the attempts to elaborate the therapeutic strategies activating immune tolerance to particular allergen. Our current research focuses on the antigen-specific action of CD8+ suppressor T (Ts) lymphocytes induced in mice by intravenous administration of a high dose of haptenated syngeneic erythrocytes. While the regulatory activity of Ts cells mediated by exosome-delivered miRNA-150 is well de ned, the mechanism of their induction remained unclear. Therefore, the current studies investigated the immune e ects induced in mice by intravenous administration of contact allergens coupled to syngeneic erythrocytes. In mouse models of hapten-induced contact hypersensitivity (CHS) and delayed-type hypersensitivity to ovalbumin, we have shown that intravenous administration of hapten-coupled erythrocytes failed to induce CHS effector cells. Moreover, hapten-induced CHS reaction occurred to be suppressed in mice intravenously administered with syngeneic erythrocytes coupled with protein allergen. Finally, we have demonstrated that intravenously administered allergen induces immune tolerance only when bound to syngeneic erythrocytes, proving that intravenously delivered allergens are deprived of their immunizing properties when coupled with membrane of self cells. Altogether, our current studies suggest that alteration of self cell membrane by allergen binding is enough to induce Ts cell-mediated immune tolerance to nonpathogenic agents, which express a great translational potential in such conditions as allergies and hypersensitivity-related autoimmune disorders.

Heterogeneous Mixture of Amniotic Cells is Likely a Better Source of Stem Cells than Adipose Tissue.

Stem cells are increasingly being used in the course of burn treatment. As several different types of stem cells are available for the purposes, it is important to chose the most efficient and the most practicable stem cell type. The aim of this study was to compare the potential of heterogeneous amnion cell mixture with the presently used standard therapy, the adipose tissue-derived stem cells. The placenta was collected during a Cesarean section procedure. Adipose tissue tissue-derived cells were isolated using the Cytori's Celution® System. Cells were tested for fulfillment of the minimum criteria for stem cells. The efficiency of cell cultures was tested by an analysis of population doubling, cell proliferation, cell cycle and cell migration. Amniotic cells presented a higher ability for differentiation to chondrocytes and osteocytes than adipose-derived regenerative cells but a lower ability for differentiation toward adipocytes. Additionally, in vitro experiments have demonstrated a higher applicability of amniotic cells than adipose tissue-derived stem cells. Amniotic cells show several advantages: easy access to placenta, low costs and a lack of ethical dilemmas related to stem cell harvesting. The main disadvantage is, however, their availability, as isogenic treatment would only be possible for women around children-bearing age, unless personalized banks for amniotic cells would be established.

Syngeneic, in contrast to allogeneic, mesenchymal stem cells have superior therapeutic potential following spinal cord injury.

We evaluated the importance of histocompatibility of transplanted MSCs in terms of therapeutic potential. Mouse syngeneic MSCs or allogeneic MSCs were transplanted following SCI in mouse. In this study we found that syngeneic, but not allogeneic, MSCs alternatively activated macrophages resulting in a down-regulation of pro-inflammation. Syngeneic MSCs also had a general suppressive effect on the immune response as compared to allogeneic MSCs. Additionally, syngeneic, but not allogeneic, MSCs significantly enhanced the recovery of hind limb function. In this study we show that the histocompatibility of transplanted MSCs is of importance for their therapeutic potential.

Evaluating full-thickness skin grafts in intraperitoneal onlay mesh position versus onlay position in mice.

BACKGROUND: Importance: Hernia surgery requires reinforcement material with few side effects when used in the intraperitoneal position. Autologous skin grafting may meet this requirement, but animal experiments are obligatory before being applied in humans. OBJECTIVE: To compare survival and effects of isogeneic full-thickness skin grafts in the intraperitoneal onlay mesh (IPOM) position in mice, with a control group using the onlay position. Primary end point was graft survival and secondary end point adhesion formation and inflammation through NF-κB activity. METHODS: Design: Intervention study with 8-week follow-up in accordance with ARRIVE criteria, performed between 2015 and 2016. SETTING: Animal laboratory. PARTICIPANTS: Transgenic C57BL/6 mice with isogeneic background were used. Recipients were female wild-type phenotype mice >3 mo (n = 24). Donors were male or female mice >7 mo, with phenotype-positive for the luciferase gene (n = 20) or positive for NF-κB-luciferase gene (n = 4). INTERVENTION: Full-thickness skin was grafted in the IPOM position and compared with grafts in the onlay position as controls. Survival was evaluated by regular longitudinal postoperative luminescence imaging over 8 wk. Adherence formation was evaluated macroscopically after sacrifice. Inflammation of full-thickness skin grafts in IPOM position of NF-κB mice was evaluated in four additional mice. Main outcome and measure: Survival of grafts, evaluated by luminescence. RESULTS: Ten animals received grafts in the IPOM position, and 10 in the onlay position as controls. Graft survival after 8 wk was 100% (20/20). Average luminescence at the end of the 8-week period was 999,597 flux (min 162,800, max 2,521,530) in the IPOM group (n = 10) and 769,708 flux (min 76,590, max 2,164,080) in the onlay control group (n = 10). No adhesions requiring sharp dissection (Jenkins' scale >2) were seen. Four animals with grafts in the IPOM position showed peak inflammation (NF-κB activity) 5 d after surgery subsiding toward the end of follow-up. CONCLUSIONS: Full-thickness skin survives as well in the IPOM position as in the onlay control position, and few adherences develop. Further studies are required to fully characterize the tissue remodeling and repair processes associated with IPOM skin grafting. The result is relevant in the search for alternative reinforcement materials to be used in complex hernia surgery in humans.

Reparación de lesiones en nervios mediante el implante de prótesis obtenidas de segmentos acelulares de nervio isogénico / Repair of nerve injury by implanting prostheses obtained from isogenic acellular nerve segments

Introducción. Cuando se produce una sección nerviosa con separación significativa de los cabos es necesario utilizar una prótesis, a modo de puente, para suturarlos. La mejor prótesis es un segmento de nervio autógeno, pero presenta importantes inconvenientes. Nuestro objetivo es comparar la eficacia de la sutura simple con la tubulización para el implante de una prótesis de nervio isogénico descelularizado. Material y método. Se utilizan 4 grupos de ratas Wistar. Grupo 0: animales donantes de nervio ciático. Grupo 1: recibió el implante con sutura término-terminal. Grupo 2: recibió el implante dentro de un tubo de ??-caprolactona. Grupo 3: lo recibió en un tubo de poliláctico-co-glicólico. Se evaluó la función motora (índice ciático) y la extensión de la regeneración (estudio histológico) a las 3 semanas del implante. Resultados. La regeneración ha sido irregular en los 3 grupos experimentales. En todos hay implantes en los que las fibras nerviosas regeneran la longitud máxima estudiada (15mm) y otros en los que la regeneración es muy escasa. La longitud media de regeneración es mayor en el grupo de sutura directa (G1), aunque la velocidad es similar en los 3. El grupo 1 muestra el mayor porcentaje de regeneración, aunque la variabilidad de los resultados impide que esta diferencia alcance significación estadística. No hemos hallado diferencias significativas entre los dos grupos con tubos de diferentes polímeros. Conclusión. Para implantar prótesis de nervios isogénicos descelularizados es más eficaz, en nuestras condiciones experimentales, la sutura término-terminal que los tubos de polímeros biocompatibles (AU)

Introduction. When a nerve section with a significant gap occurs, it is necessary to use a prosthesis to suture it. To date an autologous nerve segment graft appears to be the best treatment; but it has several important disadvantages. Our goal is to study the effectiveness of an isogenic acellular nerve prosthesis comparing a simple suture with tubulisation. Material and method. Four groups of Wistar rats were used. The animals in Group 0 served as donors of nerve segments to graft. Group 1 received the implant with an end-to-end suture. In group 2, the implant was sutured inside an ??-caprolactone tube. Group 3 received it in a polylactic-co-glycolic acid tube. We evaluated the motor function (sciatic index and step test in motion), and the regeneration length by histological study of regeneration, after a maximum of 3 weeks. Results. Regeneration was uneven in the three groups. In all groups, there were implants with regenerated nerve fibres at the maximum studied length (15mm) and others where regeneration was scarce. The mean regeneration length was greater in the direct end-to-end suture group (G1), although the regeneration speed was similar in the three groups. Group 1 showed the highest percentage of regeneration, but the variability of results prevents this difference reaching statistical significance. We found no significant differences between the two groups with polymer tubes. Conclusion. For the implantation of isogenic acellular nerve prosthesis, under our experimental conditions, the direct end-to-end suture was more effective than when it isprotected with biopolymer tubes (AU)

Repair of nerve injury by implanting prostheses obtained from isogenic acellular nerve segments. / Reparación de lesiones en nervios mediante el implante de prótesis obtenidas de segmentos acelulares de nervio isogénico.

INTRODUCTION: When a nerve section with a significant gap occurs, it is necessary to use a prosthesis to suture it. To date an autologous nerve segment graft appears to be the best treatment; but it has several important disadvantages. Our goal is to study the effectiveness of an isogenic acellular nerve prosthesis comparing a simple suture with tubulisation. MATERIAL AND METHOD: Four groups of Wistar rats were used. The animals in Group 0 served as donors of nerve segments to graft. Group 1 received the implant with an end-to-end suture. In group 2, the implant was sutured inside an ɛ-caprolactone tube. Group 3 received it in a polylactic-co-glycolic acid tube. We evaluated the motor function (sciatic index and step test in motion), and the regeneration length by histological study of regeneration, after a maximum of 3 weeks. RESULTS: Regeneration was uneven in the three groups. In all groups, there were implants with regenerated nerve fibres at the maximum studied length (15mm) and others where regeneration was scarce. The mean regeneration length was greater in the direct end-to-end suture group (G1), although the regeneration speed was similar in the three groups. Group 1 showed the highest percentage of regeneration, but the variability of results prevents this difference reaching statistical significance. We found no significant differences between the two groups with polymer tubes. CONCLUSION: For the implantation of isogenic acellular nerve prosthesis, under our experimental conditions, the direct end-to-end suture was more effective than when it isprotected with biopolymer tubes.

Fibrosis in small syngeneic rat liver grafts because of damaged bone marrow stem cells from chronic alcohol consumption.

A patient with liver failure due to chronic and acute alcohol abuse under consideration for an urgent liver transplant shortly after stopping alcohol may have residual abnormalities that threaten transplant success, particularly for a small graft. To address this, we studied a model in which reduced-size (50%) Lewis rat livers are transplanted into green fluorescence protein transgenic Lewis recipients after they are fed alcohol or a control diet for 5 weeks. Here we show that normal small Lewis grafts transplanted to alcohol-fed Lewis hosts developed fibrosis, whereas no fibrosis was observed in control-fed recipients. Host-derived CD133 + 8-hydroxy-2'-deoxyguanosine (8-OHdG) cells were significantly increased in livers recovered from both alcohol-fed and control recipients, but only alcohol-fed recipients demonstrated co-staining (a marker of oxidative DNA damage). α smooth muscle actin (α-SMA) staining, a marker for myofibroblasts, also co-localized with CD133 + cells only in the livers of alcohol-fed recipients. Immunostaining and polymerase chain reaction analysis confirmed that chronic alcohol consumption decreased the proportion of bone marrow stem cells (BMSCs) expressing CD133, c-Kit, and chemokine (C-X-C motif) receptor 4 markers and caused oxidative mitochondria DNA (mtDNA) damage. Culture of CD133 + cells from normal rats with medium containing 3% ethanol for 48 hours resulted in elevated mitochondrial 8-OHdG and mtDNA deletion, and ethanol exposure diminished CD133 expression but dramatically increased α-SMA expression. In conclusion, oxidative mtDNA damage and deletions occur in BMSCs of chronic alcohol-fed recipients, and these damaged cells mobilize to the small liver grafts and become myofibroblasts where they play a key role in the subsequent development of fibrosis. Liver Transplantation 23 1564-1576 2017 AASLD.

A bilaminated decellularized scaffold for islet transplantation: Structure, properties and functions in diabetic mice.

Ectopic transplantation of islets provides a beta cell-replacement approach that may allow the recovery of physiological regulation of the blood sugar level in patients with Type I diabetes (T1D). In development of new extrahepatic islet transplantation protocols in support of the islet engraftment, it is pivotal to develop scaffold materials with multifaceted functions to provide beneficial microenvironment, mediate host response in favor of vascularization/islet integration and maintain long-term islet function at the transplantation site. In this study, a new composite bilaminar decellularized scaffold (CDS) was fabricated with differential structural, degradation and mechanical properties by the combination of a fast-degrading porous collagen matrix and a mechanically supportive porcine pericardium. When investigated in the epididymal fat pad in syngeneic mouse models, it was shown that CDS could serve as superior scaffolds to promote islet adhesion and viability, and islet-CDS constructs also allowed rapid reversal of the hyperglycemic condition in the host. The engraftment and effects of islets were achieved at low islet numbers, accompanied by minimal adverse tissue reactions and optimal islet integration with the surrounding fat tissue. The bioactive surface, mechanical/chemical durability and biocompatibility of the CDS may all have played important roles in facilitating the engraftment of islets. Our study provided new insights into scaffold's function in the interplay of cells, materials and host tissue and the extracellular matrix-based scaffolds have potential for clinical translation in the beta cell-replacement therapy to treat T1D.

Tumor growth limited to subcutaneous site vs tumor growth in pulmonary site exhibit differential effects on systemic immunities.

To evaluate systemic immunity associated with tumor growth limited to a subcutaneous site versus growth proceeding at multiple tumor sites, we established syngeneic mouse subcutaneous and pulmonary tumor models by local subcutaneous and intravenous injection of colon carcinoma CT26 cells. We found that splenic myeloid-derived suppressor cell (MDSC) levels were significantly increased in the subcutaneous tumor model but not in the pulmonary tumor model. Furthermore, both CD4+ and CD8+ T cells as well as CD4+ Foxp3+ T cells were significantly decreased in the subcutaneous tumor model and were largely unchanged in the pulmonary tumor model. In addition, the subcutaneous model, but not the pulmonary model, displayed a Th1 polarization bias. This bias was characterized by decreased IL-4, IL-9, and IL-10 production, whereas the pulmonary model displayed increased production of IL-10. These results suggest that the mode of tumor development has differential effects on systemic immunity that may, in turn, influence approaches to treatment of cancer patients.

A simplified cuff technique for abdominal aortic transplantation in mice.

BACKGROUND: Allograft arteriopathy is still a leading cause of late organ failure. The aortic allograft model in mice has been used to study chronic rejection and has given useful information in the development of graft arteriosclerosis. However, the technical difficulties of small vessel anastomoses still continue to limit its widespread use. We introduce a new simple method for aortic transplantation in mice. METHODS: The descending aorta or infrarenal aorta from the donor mouse was anastomosed to the infrarenal aorta using a cuff technique. Aortic transplantation was performed in 30 mice, 10 isografts and 20 allografts. No immunosuppression was administered, and the recipients were sacrificed at day 28. The grafts were histologically analyzed. RESULTS: Implantation of grafts could be completed in an average of 23 min. There was no technical failure in all 60 anastomoses. The overall survival rate was 93.3%. Histology of aortas revealed typical aspects of chronic rejection in the allografts at day 28. No significant lesion was observed in isografts. CONCLUSIONS: We have developed an innovative, stable, and simple aortic transplantation model in mice, which is useful for vascular research in transplantation and beyond.

Isogenic enteric neural progenitor cells can replace missing neurons and glia in mice with Hirschsprung disease.

BACKGROUND: Transplanting autologous patient-derived enteric neuronal stem/progenitor cells (ENSCs) is an innovative approach to replacing missing enteric neurons in patients with Hirschsprung disease (HSCR). Using autologous cells eliminates immunologic and ethical concerns raised by other cell sources. However, whether postnatal aganglionic bowel is permissive for transplanted ENSCs and whether ENSCs from HSCR patients can be successfully isolated, cultured, and transplanted in vivo remains unknown. METHODS: ENSCs isolated from the ganglionic intestine of Ednrb(-/-) mice (HSCR-ENSCs) were characterized immunohistochemically and evaluated for their capacity to proliferate and differentiate in vitro. Fluorescently labeled ENSCs were co-cultured ex vivo with aganglionic Ednrb(-/-) colon. For in vivo transplantation, HSCR-ENSCs were labeled with lentivirus expressing green fluorescent protein (GFP) and implanted into aganglionic embryonic chick gut in ovo and postnatal aganglionic Ednrb(-/-) rectum in vivo. KEY RESULTS: HSCR-ENSCs maintain normal capacity self-renewal and neuronal differentiation. Moreover, the Ednrb(-/-) aganglionic environment is permissive to engraftment by wild-type ENSCs ex vivo and supports migratrion and neuroglial differentiation of these cells following transplantation in vivo. Lentiviral GFP-labeled HSCR-ENSCs populated embryonic chick hindgut and postnatal colon of Ednrb(-/-) HSCR, with cells populating the intermuscular layer and forming enteric neurons and glia. CONCLUSIONS & INFERENCES: ENSCs can be isolated and cultured from mice with HSCR, and transplanted into the aganglionic bowel of HSCR littermates to generate enteric neuronal networks. These results in an isogenic model establish the potential of using autologous-derived stem cells to treat HSCR and other intestinal neuropathies.

Syngeneic Transplants with Modified Chimeric Hematopoietic Tumors.

This protocol describes strategies to rapidly transduce tumor cells ex vivo and then transplant modified cells into immunocompetent-recipient mice. Inherent in the definition of a bona fide murine hematopoietic malignancy, unlike a myelo- or lympho-proliferative disease, is the ability to transplant tumors and give rise to a malignancy in recipient animals. This characteristic of hematopoietic disease makes these tumors a tractable model for examining the role of specific genes in tumor growth, dissemination, or therapeutic response. Additionally, because of the systemic nature of hematopoietic malignancies, transplanted tumors are frequently pathologically indistinguishable from donor malignancies-allowing one to perform decisive therapy studies on large cohorts of transplant recipients. Finally, following ex vivo manipulation, transplanted tumors can be made chimeric for the presence of defined retrovirally induced alterations. Thus, these malignancies can be made to resemble genetically heterogeneous human tumors that are in the process of acquiring new capabilities. In these experiments, fluorescent markers serve as a surrogate marker for the expression of a defined alteration, and the change in the percentage of fluorescent cells in a tumor population over time or in response to therapy can be used to gauge the impact of specific alterations on tumor behavior.

Izogenic cartilage transfer in rhinoplasty procedure.

Cartilage is commonly grafted during primary and secondary rhinoplasties as a means of addressing both functional and esthetic issues. Generally, such grafts are taken from the nasal septum, but auricular conchae or ribs may serve as donor sites if needed. However, the latter often entail considerable morbidity and graft mismatch. To circumvent these drawbacks, use of implants or processed cartilage (allogenic or xenogenic in origin) has been proposed. Herein, the isogenic transfer of nasal septal cartilage between identical twins is reported.

Hydrogen preconditioning during ex vivo lung perfusion improves the quality of lung grafts in rats.

BACKGROUND: Although the benefits of ex vivo lung perfusion (EVLP) have been globally advocated, the potentially deleterious effects of applying EVLP, in particular activation of proinflammatory cascades and alteration of metabolic profiles, are rarely discussed. This study examined proinflammatory events and metabolic profiles in lung grafts on EVLP and tested whether preconditioning lung grafts with inhaled hydrogen, a potent, cytoprotective gaseous signaling molecule, would alter the lungs' response to EVLP. METHODS: Rat heart-lung blocks were mounted on an acellular normothermic EVLP system for 4 hr and ventilated with air or air supplemented with 2% hydrogen. Arterial and airway pressures were monitored continuously; perfusate was sampled hourly to examine oxygenation. After EVLP, the lung grafts were transplanted orthotopically into syngeneic rats, and lung function was examined. RESULTS: Placing lung grafts on EVLP resulted in significant upregulation of the messenger RNAs for several proinflammatory cytokines, higher glucose consumption, and increased lactate production. Hydrogen administration attenuated proinflammatory changes during EVLP through upregulation of the heme oxygenase-1. Hydrogen administration also promoted mitochondrial biogenesis and significantly decreased lactate production. Additionally, in the hydrogen-treated lungs, the expression of hypoxia-inducible factor-1 was significantly attenuated during EVLP. These effects were maintained throughout EVLP and led to better posttransplant lung graft function in the recipients of hydrogen-treated lungs. CONCLUSIONS: Lung grafts on EVLP exhibited prominent proinflammatory changes and compromised metabolic profiles. Preconditioning lung grafts using inhaled hydrogen attenuated these proinflammatory changes, promoted mitochondrial biogenesis in the lungs throughout the procedure, and resulted in better posttransplant graft function.