Heterotopic Transplantation Model Research Articles

Mouse Model for Pancreas Transplantation Using a Modified Cuff Technique.

Mouse models have several advantages in transplantation research, including easy handling, a variety of genetically well-defined strains, and the availability of the widest range of molecular probes and reagents to perform in vivo as well as in vitro studies. Based on our experience with various murine transplantation models, we developed a heterotopic pancreas transplantation model in mice with the intent to analyze mechanisms underlying severe ischemia reperfusion injury-associated early graft damage. In contrast to previously described techniques using suture techniques, herein we describe a new procedure using a non-suture cuff technique. In recent years, we have performed more than 300 pancreas transplantations in mice with an overall success rate of >90%, a success rate never described before in mouse pancreas transplantation. The backbone of this non-suture cuff technique for graft revascularization consists of two major steps: (I) pulling the recipient vessel over a polyethylene/polyamide cuff and fixing it with a circumferential ligature, and (II) placing the donor vessel over the everted recipient vessel and fixing it with a second circumferential ligature. The resultant continuity of the endothelial layer results in less thrombogenic lesions with high patency rates and, finally, high success rates.In this model, arterial anastomosis is achieved by pulling the abdominal aorta of the donor graft over the everted common carotid artery of the recipient animal. Venous drainage of the graft is achieved by pulling the portal vein of the graft over the everted external jugular vein of the recipient. This manuscript provides details and crucial steps of the organ recovery and organ implantation procedures, which will allow researchers with microsurgical skills to perform the transplantation successfully in their laboratories.

Effect of cobra venom factor on the immune function of the pig to monkey liver xeno-transplantation

Objective To explore the effect of high purity venom factor (CVF) and other factors on immune function after pig to monkey liver xeno-transplantation. Methods To establish the pig to monkey heterotopic auxiliary liver transplantation model, the occurrence of super acute rejection (HAR) and delayed rejection (DXR) was observed after liver transplantation while interfered by high purity CVF. The expression of IgG, IgM and C3 protein in liver tissues was detected by Western blotting at different time points. Results The donor liver used for the adsorption all occured HAR, respectively of 10, 12, 11, 9, 13 minutes, the average time was (11.00±1.41) minutes. The pathological findings were ischemic necrosis and thrombosis. After CVF and other multiple factors, all the recipient monkey liver did not observe HAR and DXR until death, the IgG, IgM and C3 protein deposition in liver tissue after the death of the receptor were lower than the donor liver used for the adsorption, which is statistically significant difference (P=0.026, 0.047, 0.032). Conclusion CVF and other multiple factors can effectively overcome the occurrence of HAR in pig to monkey liver xeno-transplantation, and has the potential to prevent or delay the DXR. Key words: Xenograft; Snake venom factor; Rejection

An Anatomical Murine Model of Heterotopic Periorbital Subunit Transplantation.

Conventional reconstructive methods fail to achieve satisfactory results in total eyelid defect cases. Vascularized composite tissue allotransplantation might provide both good appearance and function for these patients. The structure of the eyelid is exceptional because it simultaneously consists of skin, connective tissue, the striated muscle, fiber structure, aponeuroses, and mucosa. Thus, before clinical application of eyelid allotransplantation, more experiments are needed to clarify the impact of ischemia, immunal suppressive agents, and deinnervation effects on these sophisticated structures. We developed an heterotopic periorbital transplantation model in rats to facilitate further experiment in this field. Twenty-five inbred male Lewis rats were used for anatomy study (n=10), and as donors or recipients of the operations (n=10). In the anatomy study, the vascular distribution and innervation to the periorbital unit was identified and recorded. Then, according to the anatomy study, 10 heterotopic transplantations and 2 transplantations with pedicle ligated were performed. The posterior facial vein and the external carotid artery are selected as the graft pedicle. All transplanted eyelids were assessed daily. Micro-CT scanning and hematoxylin and eosin staining of the grafts were performed 60 days after the operation. All recipients tolerated the operation well. All grafts without pedicles ligated survived and new hair growth was observed. All of the transplanted eyelids were pink and pliable during the entire observation period, and we did not observe any signs of arterial or venous occlusion. In the recipients with graft pedicle ligated, the grafts were necrosed and mummified within 4 to 5 days. MicroCT of the survived grafts showed good blood supply and histologic staining revealed normal histologic morphologies. Our study proved the anatomical feasibility of periorbital transplantation by establishing a heterotopic transplantation model, which might facilitate future eyelid allotransplantation-related experiments.

IL-17 contributes to the pathogenesis of obliterative bronchiolitis via regulation of M1 macrophages polarization in murine heterotopic trachea transplantation models

Acute allograft rejection is a principal conundrum in lung obliterative bronchiolitis (OB). Monocytes/macrophages infiltration has been proved to be the main reason for acute rejection. IL-17 contributes to the recruitment and function of macrophages. However, the mechanism of IL-17 underlying OB progression remains elusive. In the present study, we showed that the deficiency of IL-17 attenuated the pathology of murine heterotopic trachea allografts. Compared to WT recipients, IL-17−/− mice displayed higher frequency of CD206+ cells and lower ratio of CD86+ cells among F4/80+ macrophages in allografts and spleens on day 7 post heterotopic trachea transplantation. Moreover, mRNA levels of pro-inflammatory cytokines including IL-6, TNF-α, and IL-1β decreased in allografts of IL-17−/− recipients, but these of MRC1 and Arg-1 increased in comparison with WT. IL-17 deficiency can inhibit LPS induced M1 while promote IL-4 induced M2 polarization of bone marrow-derived macrophages. Further data demonstrated that the deficiency of IL-17 suppressed the lipopolysaccharide-induced M1 polarization and function through prevention of phosphorylation of both STAT3 and STAT5. Therefore, IL-17 contributes to OB pathogenesis through regulating macrophages function, thereby it may unravel part of the complexity of IL-17 in OB and enhance future therapeutic development.

Metastasized murine oral squamous cell carcinoma cells induce intratumoral polymorphonuclear myeloid derived suppressor cells.

Myeloid derived suppressor cells (MDSCs) localize to hematopoietic organs and peripheral blood during inflammation or tumor tissues and lymph nodes in the presence of a tumor. However, whether there are differences in MDSCs found in the primary tumor and metastases is unknown. In the present study, we established a cell line of metastasized tumor cells to a lymph node, L5-11, which were derived from the Sq-1979 mouse buccal mucosa squamous cell carcinoma cell line. We then analyzed tumor immunogenicity, especially with regard to MDSCs, to clarify the differences between the primary tumor and metastases, using an isogenic heterotopic tumor transplantation model. Our data showed that the population of intratumoral MDSCs, especially polymorphonuclear MDSCs in the lymph node metastasis model were significantly increased compared with syngeneic grafts from the primary cell line Sq-1979 after 21 days. Furthermore, we identified that the lymph node metastasis cell line had increased expression of genes that promote the expansion of MDSCs, tumor growth and metastasis. Hence, these data suggest that tumor immunosuppression can occur via activation of MDSCs. However, further examination is required to clarify whether all or a subset of these factors from the lymph node metastasis tumor cells are required to induce intratumoral MDSCs.

Tu1875 - Ovarian Cancer G-Protein-Coupled Receptor 1 Deficiency Protects from Fibrogenesis in the Heterotopic Transplantation Model of Intestinal Fibrosis

Tu1875 - Ovarian Cancer G-Protein-Coupled Receptor 1 Deficiency Protects from Fibrogenesis in the Heterotopic Transplantation Model of Intestinal Fibrosis

Intralipid® attenuates acute cardiac allograft rejection in relation to promoting CD4+CD25+Foxp3+ regulatory T-cells and inhibiting toll-like receptor 4 expression

Clinically available agents that promote the suppression of inflammatory processes with minimal adverse effects are highly desired. An FDA-approved nutrition supplement, Intralipid®, has shown its immunomodulatory potential. However, its influence on organ transplant rejection has not been tested. Here, we have investigated whether Intralipid® affect allograft outcome in a fully major histocompatibility complex (MHC)-mismatched rat model of heterotopic working-heart transplantation. Recipients were treated intravenously with Intralipid® (1.5g/kg, clinical dosage) or saline (untreated control) on post-transplantation days (PODs) 1, 3, and 6. Treatment with Intralipid® results in a significant prolongation of median graft survival from 7 days to 12 days. On POD 7, while the allografts from our untreated (i.e., control) group exhibit severe rejection, the allografts from Intralipid®-treated group show milder rejection demonstrating preserved myocardium integrity and decreasing infiltration of inflammatory cells (ED1+ and CD3+ cells). Flow cytometry analysis reveals a marked increase of CD4+CD25+Foxp3+ regulatory T-cells (Tregs) in the peripheral blood of Intralipid®-treated recipients beginning on POD 3 (7.0% vs. 14.4%, untreated vs. Intralipid®-treated). Intralipid® treatment also promotes Tregs in the spleens and allograft hearts. Toll-like receptor 4 expression is markedly suppressed in the Intralipid®-treated allografts. Our results suggest a new use of Intralipid® to modulate immune responses in the setting of heart transplantation. This study could stimulate further mechanistic as well as treatment studies leading to the clinical application of Intralipid® as an adjunctive therapy with current immunosuppression agents to manage organ rejection.

Triptolide Reduces the Required Dose of Tacrolimus by Attenuating Inflammation, Enhancing Immunosuppression, and Increasing Donor Chimerism in a Heterotopic Hindlimb Transplantation Model.

Induction of tolerance and minimizing the toxicity of immunosuppression are two fundamental goals in vascularized composite allotransplantation. Accumulating data indicate that triptolide is an agent that may have the capacity to suppress inflammation and immunologic rejection. A heterotopic hindlimb allotransplantation model from Brown Norway to Lewis rats was established and treated with different doses of tacrolimus combined with or without triptolide. Mean survival time of the transplants was monitored, and histopathologic examination of the skin was performed. The level of inflammatory cytokine interleukin-1β, interleukin-6, and tumor necrosis factor-á in peripheral blood was assayed. The percentage of T lymphocytes and its subsets was measured using flow cytometry. The level of recipient peripheral chimerism and the apoptosis of donor bone marrow cells were evaluated. The apoptotic related genes bcl-2 and Bax were detected by real-time polymerase chain reaction. The authors' results showed that triptolide not only reduces the dose of tacrolimus required for immunosuppression, but also decreased drug side effects in terms of weight gain and diarrhea. Triptolide had an obvious effect on proinflammatory cytokine expression and T-lymphocyte proliferation in the peripheral blood. Interestingly, triptolide could increase the mixed chimerism level of recipients, possibly by inhibiting the apoptosis of transplanted bone marrow cells by means of regulation of the apoptotic genes bcl-2 and Bax. Triptolide reduces the dose of tacrolimus required for immunosuppression by attenuating inflammation and by T-cell suppression. Furthermore, triptolide increases the chimerism level, which might contribute to acceptance of the allografts.

Prolonged survival effects induced by immature dendritic cells and regulatory T cells in a rat liver transplantation model.

Dendritic cells (DCs) and regulatory T (Treg) cells are crucial for inducing immune tolerance. However, the suppressive function of infused Treg cells and immature DCs (imDCs) following solid organ transplantation remains unclear. ImDCs derived from DA-donor rats and Treg cells isolated from spleens of Lewis rats were prepared. A heterotopic liver transplantation model was established to examine the immune tolerance effects of infusion of Treg-imDCs, imDCs and Treg cells individually. Th1/Th2 cytokines and TRAL were detected by ELISA. The overall rejection grade was assessed and the rejection activity index (RAI) was calculated. TUNEL-positive lymphocytes were detected in the portal area in liver sections. The infusion of Treg-imDCs was more effective than imDCs or Treg cells individually. Moreover, the expression of IL-10 and TGF-β1 was significantly up-regulated, and IL-12 expression was significantly down-regulated, especially in the Treg-imDCs group. The percentage of TUNEL-positive cells was significantly higher in the Treg cells and imDCs groups. The RAI values in Treg-imDCs group on days 3 and 7 were lower than control, imDCs and Treg cells groups individually (p<0.05). Both TBIL and ALT levels in the Treg-imDCs and imDCs groups were significantly lower than those of the control and Treg cells groups, and serum TRAL levels increased significantly 10days after transplantation in the imDC and Treg-imDC groups compared with the control and Treg cells groups (P<0.001). These data demonstrated that infusion of Treg cells and/or imDCs induces alloantigen tolerance and prolongs liver allograft survival. The infusion of Treg-imDCs was more effective than imDCs or Treg cells individually. ImDCs synergize with Treg cells in inducing and maintaining the feedback loop between imDCs and Treg cells in vivo.

Establishment and comparison of stoma and stoma-free heterotopic small intestine transplantation models in mice

To establish stoma and stoma-free murine models of heterotopic small intestine transplantation in order to choose a more effective and reliable model. A total of 140 male 8-10 weeks age C57BL/6(B6) mice weighted 25-30 g were enrolled in the experiment. Syngeneic heterotopic small intestine transplantation was performed between C57BL/6 mice, and recipient mice were divided into either stoma or stoma-free group. Heterotopic small intestine transplantation was performed in 70 mice, with 35 mice in each group. After closing the proximal end of the graft by ligation, the distal end of graft was exteriorized as a stoma then secured to the skin of the abdominal wall in stoma group. In stoma-free group, the distal end of graft was anastomosed end-to-side to the recipient ileum. Successful rate of operation, two-week survival rate, operation time, associated complications, postoperative care time and body weight change were recorded and compared between two groups. The successful rate of stoma group was 65.7%, while it was 80.0% of stoma-free group (χ(2)=1.806, P=0.179). The operation time of donor in stoma group was (48.1±6.6) minutes, while it was (47.2±5.9) minutes in stoma-free group (t=0.598, P=0.552). The operation time of recipient in stoma group was (77.9±9.1) minutes, while it was (76.4±8.3) minutes in stoma-free group (t=0.683, P=0.497). The cold ischemic time of graft in stoma group was (34.7±4.0) minutes, while it was (33.9±4.6) minutes in stoma-free group(t=0.667, P=0.507). The two-week survival rate of stoma group was 45.7%, and it was 77.1% of stoma-free group(χ(2)=7.295, P=0.007). The stoma group had more complications[54.3%(19/35) vs. 22.9%(8/35), χ(2)=7.295, P=0.007], which needed more postoperative care time(191 min vs. 35 min). The weight loss in stoma group in the third day after operation was more significant [(81.52±5.20)% vs. (85.46±4.65)%, t=2.856, P=0.006]. By 2 weeks after operation, the weight of mice in both groups retruned to 95% of the postoperative wight. The murine heteropotic small intestine transplantation model with stoma-free appears to be more reasonable and reliable.

Fast-track surgery and exclusive enteral nutrition applied to a rat model of heterotopic intestinal transplantation.

The present study applied fast-track surgery (FTS) concepts and exclusive enteral nutrition (EEN) to a rat model of heterotopic intestinal transplantation (HIT). A total of 96 pairs of Sprague-Dawley rats were randomly distributed into three groups, as follows: i) The conventional group (group 1); ii) the FTS group (group 2); and iii) the FTS with EEN group (EEN group). FTS alterations to the HIT protocol were as follows: i) The use of sevoflurane as an anesthetic; ii) alterations to the order of the procedure and iii) a modified suturing technique. In addition, the EEN group rats underwent an early EEN gavage. The operation time, success rate, recovery state and morphological characteristics of the grafts were compared among the groups. The average operative time was significantly decreased in the group 2 and EEN group rats (137.44±16.03 and 139.67±15.25 min, respectively), as compared with the group 1 rats (169.36±13.72 min; P<0.05). In addition, the percentage of rats surviving >14 days was significantly increased in the group 2 (87.5%) and EEN group (90.6%) rats, as compared with the group 1 rats (68.7%; P<0.05). Furthermore, the villi of graft in EEN group appeared longer, and exhibited narrower interspaces. The ischemia-reperfusion injury and mononuclear cell infiltration were attenuated at postoperative day 7. The results of the present study suggested that the application of FTS concepts and EEN gavage to HIT may accelerate recovery and ameliorate graft damage following surgery.

Genetic and Pharmacological Modulation of Akt1 for Improving Ovarian Graft Revascularization in a Mouse Model.

Ovarian tissue cryopreservation and transplantation is one of a few available treatments for fertility preservation in women diagnosed with cancer. Rapid revascularization is essential for reducing hypoxic damage after grafting and protecting the primordial follicles reserve. Using a mouse model of heterotopic ovarian graft transplantation, we have delineated the role of endothelial Akt1 expression using longitudinal magnetic resonance imaging follow-up to quantify angiogenic response. Endothelial Akt1 activation in ovarian grafts promoted angiogenesis to support the graft during posttransplantation hypoxic period. Similarly, simvastatin therapy activated Akt1 at the transplantation site and improved the revascularization and vascular support of ovarian grafts. These results serve as an important first step toward pharmacological intervention to improve revascularization of ovarian grafts and restoration of fertility in cancer survivors. The pro-angiogenic effects reported here may extend beyond improving ovarian graft reception in fertility preservation and could potentially be used for different organ or tissue transplantation.

Increased expression of Notch 1 in a rat liver transplantation model.

Liver transplantation is the standard treatment for end‑stage liver failure; however, rejection can result in allograft failure. In order to investigate the role of Notch 1 during rejection, the present study evaluated Notch 1 expression, as well as the levels of immune reactivity, in rat liver allografts. A heterotopic liver transplantation model was established using Dark Agouti (DA) rats as donors and Lewis rats as recipients (DA/Lewis), with DA recipient rats serving as controls (DA/DA). The concentration levels of immune reactivity markers and serum Notch 1 were measured on days 3, 5, and 7. The overall survival was significantly shorter (<10 days) in the DA/Lewis group, as compared with the DA/DA group (P<0.0001). The concentration levels of serum alanine aminotransferase and total bilirubin were significantly higher 5 and 7 days following transplantation in the DA/Lewis group, as compared with the DA/DA group (P<0.001). The concentration levels of serum Notch 1 were significantly higher in the DA/Lewis group, as compared with the DA/DA group on days 3, 5, and 7 following transplantation (P<0.0001). These results indicate that the expression levels of serum Notch 1 significantly increase during liver allograft rejection, suggesting that Notch 1 is involved in the mechanism underlying liver allograft rejection. Notch 1 may serve as a marker of acute rejection in a rat liver transplantation model.

Mesenchymal stromal cells improve cardiac function and left ventricular remodeling in a heart transplantation model

Ischemia/reperfusion (I/R) injury is an inevitable consequence of organ transplantation and a major determinant of patient and graft survival in heart transplantation. Bone marrow-mesenchymal stromal cell (BM-MSC) treatment is a potentially effective cell therapy for cardiac disease. We investigated the effects of intravenous delivery of BM-MSCs in the acute phase post-transplant in a heterotopic heart transplantation (HHT) model associated with I/R injury. Hearts of wild-type Lewis (WT LEW) rats were harvested and transplanted heterotopically into the necks of recipient WT LEW rats. Forty-eight hours after HHT, BM-MSCs were injected intravenously into animals in the experimental group, whereas controls received normal saline (NS). Eight days after BM-MSC injection, fractional shortening of transplanted hearts was significantly higher and left ventricular systolic diameter was lower in the BM-MSC group compared with controls, whereas no differences were found 28 days after infusion. A reduction in ventricular remodeling and cardiac fibrosis was observed by histochemical analysis and confirmed by cardiac magnetic resonance imaging in the BM-MSC group. The perivascular stromal cells' density and the number of capillaries were increased whereas the number of apoptotic cells decreased significantly in transplanted hearts in the BM-MSC group compared with the NS group. We showed early improvement in cardiac function and subsequent enhanced ventricular remodeling, reduced cardiac fibrosis, augmented neo-vascularization and decreased cardiomyocyte apoptosis of the transplanted heart in a heterotopic transplantation model after intravenous infusion of BM-derived MSCs. Our data suggest that clinical studies with BM-MSCs are warranted to understand their effects on cardiac graft and transplant recipient survival.

Endocardial fibroelastosis is caused by aberrant endothelial to mesenchymal transition.

Endocardial fibroelastosis (EFE) is a unique form of fibrosis, which forms a de novo subendocardial tissue layer encapsulating the myocardium and stunting its growth, and which is typically associated with congenital heart diseases of heterogeneous origin, such as hypoplastic left heart syndrome. Relevance of EFE was only recently highlighted through the establishment of staged biventricular repair surgery in infant patients with hypoplastic left heart syndrome, where surgical removal of EFE tissue has resulted in improvement in the restrictive physiology leading to the growth of the left ventricle in parallel with somatic growth. However, pathomechanisms underlying EFE formation are still scarce, and specific therapeutic targets are not yet known. Here, we aimed to investigate the cellular origins of EFE tissue and to gain insights into the underlying molecular mechanisms to ultimately develop novel therapeutic strategies. By utilizing a novel EFE model of heterotopic transplantation of hearts from newborn reporter mice and by analyzing human EFE tissue, we demonstrate for the first time that fibrogenic cells within EFE tissue originate from endocardial endothelial cells via aberrant endothelial to mesenchymal transition. We further demonstrate that such aberrant endothelial to mesenchymal transition involving endocardial endothelial cells is caused by dysregulated transforming growth factor beta/bone morphogenetic proteins signaling and that this imbalance is at least in part caused by aberrant promoter methylation and subsequent transcriptional suppression of bone morphogenetic proteins 5 and 7. Finally, we provide evidence that supplementation of exogenous recombinant bone morphogenetic proteins 7 effectively ameliorates endothelial to mesenchymal transition and experimental EFE in rats. In summary, our data point to aberrant endothelial to mesenchymal transition as a common denominator of infant EFE development in heterogeneous, congenital heart diseases, and to bone morphogenetic proteins 7 as an effective treatment for EFE and its restriction of heart growth.

Pig BMSCs Transfected with Human TFPI Combat Species Incompatibility and Regulate the Human TF Pathway in Vitro and in a Rodent Model

Background: The activation of tissue factor (TF) is one of the major reasons for coagulation dysregulation after pig-to-primate xenotransplantation. Tissue factor pathway inhibitor (TFPI) is the most important inhibitor of TF. Studies have demonstrated species incompatibility between pig TFPI and human TF. Methods: A pig-to-macaque heterotopic auxiliary liver transplantation model was established to determine the origin of activated TF. Chimeric proteins of human and pig TFPI were constructed to assess the role of Kunitz domains in species incompatibility. Immortalised pig bone marrow mesenchymal stem cells transfected with human TFPI were tested for their ability to inhibit clotting in vitro. Results: TF from recipient was activated early after liver xenotransplantation. Pig TFPI Kunitz domain 2 bound human FXa, but Kunitz domain 1 did not effectively inhibit human TF/FVIIa. Immortalised pig bone marrow mesenchymal cells (BMSCs) transfected with human TFPI showed a prolonged recalcification time in vitro and in a rodent model. Conclusion: Recipient TF is relevant to dysregulated coagulation after xenotransplantation. Kunitz domain 1 plays the most important role in species incompatibility between pig TFPI and human TF, and clotting can be inhibited by human TFPI-transfected pig BMSCs. Our study shows a possible way to resolve the incompatibility of pig TFPI.

A new rat model of auxiliary partial heterotopic liver transplantation with liver dual arterial blood supply.

Auxiliary partial heterotopic liver transplantation (APHLT) with portal vein arterialization is a valuable procedure to be considered in the treatment of patients with acute liver failure and metabolic liver diseases. The aim of this study was to develop a new rat model of APHLT with liver dual arterial blood supply (LDABS). A total of 20 rats were used. The donor liver was resected, and the celiac trunk was reserved. Left and medial hepatic lobes accounting for 70% of the liver mass were removed en bloc and the suprahepatic caval vein was ligated simultaneously. Thus, 30% of the donor liver was obtained as the graft. Sleeve anastomosis of the graft portal vein and splenic artery were performed after narrowing the portal vein lumen through suturing. The right kidney of the recipient was removed, and sleeve anastomosis was performed between the celiac trunk of the graft and the right renal artery of the recipient. In addition, end-to-end anastomosis was performed between the infrahepatic caval vein of the graft and the right renal vein of the recipient. Following the reperfusion of the graft, the blood flow of the arterialized portal vein was controlled within the physiological range through suturing and narrowing under monitoring with an ultrasonic flowmeter. The bile duct of the graft was implanted into the duodenum of the recipient through an internal stent catheter. A 70% section of the native liver (left and medial hepatic lobes) was resected using bloodless hepatectomy. The mean operative duration was 154.5±16.4 min, and the warm and cold ischemia times of the graft were 8.1±1.1 min and 64.5±6.6 min, respectively. The blood flow of the arterialized portal vein to the graft was 1.8±0.3 ml/min/g liver weight. The success rate of model establishment (waking with post-surgical survival of >24 h) was 70% (7/10). Following successful model establishment, all rats survived 7 days post-surgery (100%; 7/7). The graft was found to be soft in texture and bright red in color following exploratory laparotomy. In conclusion, a new rat model of APHLT with LDABS without stent for vascular reconstruction was developed. This is a feasible and reliable rat model for liver transplantation study.

Establishment of mice heterotopic cardiac transplantation and observation between different strains

Objective To establish the mice heterotopic cardiac transplantation as well as to observe the different graft reaction between different strains.Methods According to the surgical procedures of Ono's pattern in mice cardiac heterotopic transplantation model,we did allograft cardiac transplantation using same (BALB/c→BALB/c ; CBA→CBA; C57BL/6j→C57BL/6j) or different (BALB/c→C57BL/6j ; C57BL/6j→BALB/c; C57BL/6j→CBA; CBA→C57BL/6j)strains male mice.Survival time and pathological examination were observed after transplantation Results The operative survival was 88.6％ (62/70).Mean donor graft harvest time of was 4.8 min,mean anatomic time was (28.70 ± 3.56) min (BALB/c→BALB/c),(20.70±4.22) min (CBA→CBA) and (27.90±2.51) min (C57BL/6j→C57BL/6j) in same strains while (28.00 ± 2.71) min (BALB/c→C57BL/6j),(22.10 ± 4.33) min (C57BL/6j→CBA),(29.90±3.14) min (C57BL/6j→BALB/c) and (30.33 ±23.00) min (CBA→C57BL/6j) in different strains.The survival time was over 100 days in same strains and (7.10 ±0.74) d (C57BL/6j→BALB/c) ; (7.40 ± 0.82) d (BALB/c→ C57 BL/6j) ; (8.90 ± 1.10) d (C57 BL/6j→ CBA) ; (8.50 ± 0.85) d (CBA→C57BL/6j) d in different strains.It was confirmed with pathological examination.Conclusion On the base of mastering the microsurgical techniques,we can establish the mice cardiac heterotopic transplantation model.The immune response in C57BL/6j to BALB/c model was very strong,while weaker in C57BL/6j to CBA models.The different transplant immune response can be used in the transplantation research. Key words: Mice; Cardiac transplantation; Models ; Strains

Transplantation of pulmonary valve using a mouse model of heterotopic heart transplantation.

Tissue engineered heart valves, especially decellularized valves, are starting to gain momentum in clinical use of reconstructive surgery with mixed results. However, the cellular and molecular mechanisms of the neotissue development, valve thickening, and stenosis development are not researched extensively. To answer the above questions, we developed a murine heterotopic heart valve transplantation model. A heart valve was harvested from a valve donor mouse and transplanted to a heart donor mouse. The heart with a new valve was transplanted heterotopically to a recipient mouse. The transplanted heart showed its own heartbeat, independent of the recipient's heartbeat. The blood flow was quantified using a high frequency ultrasound system with a pulsed wave Doppler. The flow through the implanted pulmonary valve showed forward flow with minimal regurgitation and the peak flow was close to 100 mm/sec. This murine model of heart valve transplantation is highly versatile, so it can be modified and adapted to provide different hemodynamic environments and/or can be used with various transgenic mice to study neotissue development in a tissue engineered heart valve.

Carbon monoxide-releasing molecule attenuates allograft airway rejection

Acute rejection after lung transplantation is the main risk factor for the development of bronchiolitis obliterans (BO). Carbon monoxide (CO) can provide anti-inflammatory effects and may serve to limit tissue injury in airway transplant. Here, we tested the ability of carbon monoxide releasing molecule-2 (CORM-2) to prevent airway rejection. Tracheal grafts from BALB/c or C57BL/6 were transplanted to C57BL/6 recipients. Experimental groups were treated with multiple doses of CORM-2. Histopathological evaluation of luminal obliteration was blindly reviewed. Immunohistochemistry and real-time RT-PCR analyses were performed. Allografts treated with CORM-2 revealed a striking reduction of thickening in epithelial and subepithelial airway layers (P < 0.01) at day 7 in orthotopic trachea transplantation model compared with allografts treated with vehicle. In heterotopic trachea transplantation model, CORM-2 treated allografts showed a reduction of luminal obliteration (P < 0.01) at days 14 and 21. There was also a concordant decrease in CD3(+) lymphocytes and macrophages in CORM-2 treated allografts. IFN-γ, IL-2 and IL17A mRNA expressions were reduced dramatically by systemic administration of CORM-2. These data implicate CORM-2-derived CO has an important protective function in experimental BO, and may represent a target for the therapeutic intervention of chronic lung allograft rejection.