OBJECTIVETo establish stoma and stoma-free murine models of heterotopic small intestine transplantation in order to choose a more effective and reliable model.

METHODSA 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.

RESULTSThe 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.

CONCLUSIONThe murine heteropotic small intestine transplantation model with stoma-free appears to be more reasonable and reliable.

Animals ; Digestive System Surgical Procedures ; Ileum ; surgery ; Intestine, Small ; transplantation ; Male ; Mice ; Mice, Inbred C57BL ; Surgical Stomas ; Transplantation, Heterotopic ; methods ; Transplantation, Isogeneic

BACKGROUNDA practical problem impeding clinical translation is the limited bone formation seen in artificial bone grafts. Low-pressure/vacuum seeding and dynamic culturing in bioreactors have led to a greater penetration into the scaffolds, enhanced production of bone marrow cells, and improved tissue-engineered bone formation. The goal of this study was to promote more extensive bone formation in the composites of porous ceramics and bone marrow stromal cells (BMSCs).

METHODSBMSCs/β-tricalcium phosphate (β-TCP) composites were subcultured for 2 weeks and then subcutaneously implanted into syngeneic rats that were split into a low-intensity pulsed ultrasound (LIPUS) treatment group and a control group. These implants were harvested at 5, 10, 25, and 50 days after implantation. The samples were then biomechanically tested and analyzed for alkaline phosphate (ALP) activity and osteocalcin (OCN) content and were also observed by light microscopy.

RESULTSThe levels of ALP activity and OCN content in the composites were significantly higher in the LIPUS group than in the control group. Histomorphometric analysis revealed a greater degree of soft tissue repair, increased blood flow, better angiogenesis, and more extensive bone formation in the LIPUS groups than in the controls. No significant difference in the compressive strength was found between the two groups.

CONCLUSIONLIPUS treatment appears to enhance bone formation and angiogenesis in the BMSCs/β-TCP composites.

Animals ; Bone Marrow Cells ; physiology ; Bone Transplantation ; Calcium Phosphates ; pharmacology ; Male ; Osteogenesis ; physiology ; Rats ; Stromal Cells ; Tissue Engineering ; methods ; Transplantation, Isogeneic ; Ultrasonics ; methods

Syngeneic bone marrow transplantation (syn-BMT), as a novel therapy for type 1 diabetes (T1D), has been used more and more widely. This study was aimed to detect the changes of peripheral CD4(+) T lymphocytes, CD8(+) T lymphocytes, CD4(+)/CD8(+) T lymphocytes and NK cells before and after T1D mice were treated with syn-BMT, and to investigate the effects of these cells in T1D and the effects of syn-BMT-inducing immunotolerance. T1D mouse model was established by multiple low dose streptozotocin injection, the syn-BMT was performed on 10 day after the onset of diabetes. The T1D model mice were divided into group of diabetic mice treated with syn-BMT and group of diabetic control mice (DC), 6 normal C57BL/6J mice were regarded as normal control group (NC). On 30 day after syn-BMT, peripheral proportion of CD4(+) T lymphocytes, CD8(+) T lymphocytes, CD4(+)/CD8(+) T lymphocytes and NK cells were detected by flow cytometry. These cells of normal control mice (NC), diabetes control mice (DC) and diabetes mice treated by syn-BMT were also detected. Blood glucose level in three groups was assayed during the whole observation period. The results showed that syn-BMT could reduce blood glucose level of T1D mice to near normal (p > 0.05). Hematopoietic reconstitution happened in a month. The proportion of peripheral CD4(+) T lymphocytes, CD4(+)/CD8(+) T lymphocytes, NK cells all increased in new-onset diabetic mice (p < 0.01), while the proportion of peripheral CD8(+) T lymphocytes decreased (p < 0.01). On 30 day after T1D mice were treated with syn-BMT, the proportion of peripheral CD4(+) T lymphocytes was significantly lower than that in DC mice (p < 0.01), but still higher than NC (p < 0.05). The proportion of CD8(+) T lymphocytes was higher than that in DC and NC mice (p < 0.01). The ratio of CD4(+)/CD8(+) T lymphocytes and proportion of NK cells were both obviously lower than that in DC and NC mice (p < 0.01). It is concluded that the syn-BMT can reverse hyperglycemia and immune disorder in diabetic mice. On early period of diabetes onset, the proportions of CD4(+) T lymphocytes and NK cells, the ratio of CD4(+)/CD8(+) T lymphocytes increase, while proportion of CD8(+) T lymphocytes decreases in peripheral blood which mye be associated with onset of diabetes.
Animals ; B-Lymphocytes ; immunology ; Bone Marrow Transplantation ; CD4-CD8 Ratio ; Diabetes Mellitus, Type 1 ; immunology ; surgery ; Flow Cytometry ; Killer Cells, Natural ; immunology ; Lymphocyte Count ; Male ; Mice ; Mice, Inbred C57BL ; T-Lymphocytes ; immunology ; Transplantation, Isogeneic

The objective of this study was to investigate the effect of ligustrazine on the expression of stem cell factor mRNA (SCF) in bone marrow tissue and explore the mechanism of hematopoietic reconstitution after bone marrow transplantation (BMT). The colony forming unit of spleen (CFU-S) were counted, the survival rate at days 7, 14 and 21 after BMT were measured, as well as the expression level of SCF mRNA was detected by RT-PCR. The results showed that in ligustrazine group CFU-S counts on day 10 and survival rate, expression level of SCF mRNA on day 7, 14 and 21 after BMT were higher than that in the control group (p < 0.01 or p < 0.05). In conclusion, ligustrazine promotes the recovery of hematopoietic cells in bone marrow, enhances the repair of bone marrow microvessels, and then improves bone marrow microenvironment and promotes hematopoietic reconstitution.
Animals ; Bone Marrow Transplantation ; Hematopoiesis ; drug effects ; Male ; Mice ; Mice, Inbred BALB C ; Pyrazines ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Stem Cell Factor ; genetics ; metabolism ; Transplantation, Isogeneic

OBJECTIVETo screen specific functional proteins from lymphocyte involved in acute rejection using differential proteomics research.

METHODSTwo groups of rat liver transplantation models were established (isograft as control and allograft as acute rejection groups) by transplantation within Wistar rats, and between Wistar and SD. Morphology study were performed by histochemistry tech, followed by serum cytokine detection with ELISA. With 2-dimensional electrophoresis, proteomes of lymphocyte from the rats of different groups were separated and 2 proteome profiles were established. Comparing with the 2 profiles, 25 spots were selected and picked for in gel digestion, followed for analysis by matrix assisted laser desorption ionization (MALDI)-time of fly (TOF)/TOF MS. Two of the proteins were detected with Western blot to verify the changing profiles.

RESULTSThe results of morphology analysis and detection of cytokines (IL-2 and IFN-gamma) indicate that the animal models were established successfully and acute rejection happened after transplantation for 3 days. Twenty-five differential proteins were found out to be associated with acute rejection, among which 13 proteins were upregulated and 12 downregulated. The expression alterations of 2 proteins (beta-actin and carbonic anhydrase) are consistent with proteomics analysis results showing in Western blot.

CONCLUSIONSTwenty-five specific proteins exploiting mechanism of acute rejection are screened out, including IL-2 and carbonic anhydrase, which maybe benefit for the further works.

Acute Disease ; Animals ; Disease Models, Animal ; Graft Rejection ; metabolism ; pathology ; Interferon-gamma ; blood ; Interleukin-2 ; blood ; Liver ; pathology ; Liver Transplantation ; methods ; Lymphocytes ; metabolism ; Peptide Mapping ; Proteomics ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Transplantation, Homologous ; Transplantation, Isogeneic

A novel combined treatment of conventional chemotherapy with an intratumoral injection of syngeneic dendritic cells (DCs) has emerged as a potent cancer treatment strategy. In this study, we evaluated the synergistic effect of an intraperitoneal (i.p.) injection of a chemotherapeutic drug, paclitaxel, and an intratumoral (i.t.) injection of syngeneic bone marrow- derived DCs for the treatment of pre-existing fibrosarcoma. Subcutaneous tumors were established using MCA102 fibrosarcoma cells in syngeneic C57BL/6 mice. The results demonstrated that the combined treatment of paclitaxel chemotherapy and the injection of DCs led to complete tumor regression, in contrast to only partial eradication of the tumors with chemotherapy or DCs alone. Furthermore, the tumor-free mice were able to resist a repeat challenge with the same type of tumor. These findings suggest that a combination therapy of systemic chemotherapy along with the intratumoral administration of DCs is a potent treatment strategy for fibrosarcoma.
Treatment Outcome ; Transplantation, Isogeneic ; Phenotype ; Paclitaxel/administration & dosage/*therapeutic use ; Mice ; Injections, Intraperitoneal ; Immunologic Memory ; Fibrosarcoma/drug therapy/pathology/*therapy ; Dendritic Cells/cytology/*transplantation ; Combined Modality Therapy ; Cells, Cultured ; Cell Line, Tumor ; Bone Marrow Cells/cytology ; Antineoplastic Agents, Phytogenic/administration & dosage/*therapeutic use ; Animals

To explore the effect of ligustrazine on the expression of adherent molecule VCAM-1/VLA-4 of bone marrow cells in syngenic bone marrow transplantation (BMT) mice, the mice were divided into 3 groups: normal group (which received no treatment), BMT control group and ligustrazine-treated groups. BMT mouse models were established. The BMT control group and the ligustrazine-treated group were orally administered 0.2 ml saline per mouse and 2 mg ligustrazine per mouse, respectively, twice a day. On the day 7, 14, 21, 28 after BMT, mice were respectively killed. Bone marrow nucleated cells were detected, and then the expression of VCAM-1/VLA-4 was assayed by immunohistochemistry, RT-PCR and flow cytometry analysis, respectively. The results showed that in ligustrazine-treated group, the accounts of bone marrow nucleated cells on the day 7, 14, 21, 28 after BMT were all higher than that in BMT control group. The expression level in the ligustrazine-treated group was significantly higher than that in the BMT control group (P < 0.05 or P < 0.01). It is concluded that ligustrazine can enhance VCAM-1/VLA-4 expression in bone marrow after syngenic bone marrow transplantation in mice, which may be related to the mechanisms underlying the ligustrazine accelerating hematopoietic reconstitution in allogenic bone marrow transplantation.
Animals ; Bone Marrow Cells ; drug effects ; metabolism ; Bone Marrow Transplantation ; methods ; Flow Cytometry ; Immunohistochemistry ; Integrin alpha4beta1 ; biosynthesis ; genetics ; Male ; Mice ; Mice, Inbred BALB C ; Pyrazines ; pharmacology ; Reverse Transcriptase Polymerase Chain Reaction ; Transplantation, Isogeneic ; Vascular Cell Adhesion Molecule-1 ; biosynthesis ; genetics

The p53 gene has a significant role in controlling genomic stability of cancer. The purpose of this study was to evaluate the tumor response of allograft colorectal tumor treated with Ad5CMV-p53 in a syngeneic rat model. Two weeks after the inoculation of WB-2054-M5 tumor cells in the flank of rats, rats were randomly assigned by tumor size to one of three groups (n=18 in each): phosphate buffered saline (PBS), Ad5CMV, and Ad5CMV-p53. Recombinant adenovirus or PBS was administered through intratumoral injection at three divided doses every other day for 4 weeks. Apoptosis of the tumors was evaluated using TUNEL assay. After 2 and 4 weeks of treatment, the volume (cm3) of tumors in PBS, Ad5CMV, and Ad5CMV-p53 was as follows: 2 week: 1.66 +/-0.43, 1.40 +/-0.47, 0.75 +/-0.26 (p<0.001), 4 week: 4.41 +/-0.88, 3.93 +/-1.86, 2.33 +/-0.51 (p<0.001). Tumor growth showed no statistically significant difference between the PBS and Ad5CMV groups (6-week vol. p=0.32). The TUNEL assay results revealed more apparent apoptotic cells in Ad5CMV-p53-treated tumors than in other groups. Growth of allograft colorectal cancer in the syngeneic rat model was significantly suppressed by intratumoral Ad5CMV-p53 gene therapy. These results demonstrate that gene replacement therapy with p53 may provide a novel modality of treatment in conjunction with other present treatments for metastatic colorectal cancer.
Adenocarcinoma/genetics/pathology/therapy ; Adenoviridae/*genetics ; Animals ; Cell Line, Tumor ; Cell Proliferation ; Cell Survival/genetics ; Colorectal Neoplasms/*genetics/pathology/*therapy ; Disease Models, Animal ; Female ; Gene Therapy/*methods ; Gene Transfer Techniques ; Men ; Protein p53/*genetics/*therapeutic use ; Rats ; Rats, Inbred WF ; Recombinant Proteins/therapeutic use ; Research Support, Non-U.S. Gov't ; Transplantation, Isogeneic ; Treatment Outcome

Comparison of pancreaticoduodenal transplants (PDT) and duct-ligated pancreas transplant (DLPT) were performed using syngeneic and allogeneic studies in rats. Both DLPT and PDT allogeneic grafts showed mild rejection. DLPT groups showed disorganized pathology and acini replaced by fat. Eventually, massive fibrosis was seen in the Islets of Langerhans, as well as rejection cellular infiltrates. In both PDT groups, normal histology was observed in the same period. Thus the effect of duct occlusion is highly detrimental for the grafts.
Animals ; Graft Rejection/pathology ; Ligation/adverse effects ; Pancreas/*pathology ; Pancreas Transplantation/*adverse effects ; Pancreatic Ducts/surgery ; Postoperative Period ; Rats ; Rats, Inbred Lew ; Rats, Sprague-Dawley ; Research Support, Non-U.S. Gov't ; Transplantation, Homologous ; Transplantation, Isogeneic

To estimate the effect of ligustrazine on the expression of PECAM-1/CD31 and hematopoietic reconstitution in syngenic bone marrow transplanted mice, 56 BALB/c mice were divided into 3 groups: normal control group, BMT control group, ligustrazine treated group (BMT + Ligustrazine) and syngenic BMT mouse models were established according to the literatures. The BMT control group and the ligustrazine treated group were given orally 0.2 ml saline and 2 mg ligustrazine twice a day respectively. On days 7, 14, 21 after BMT, mice were killed and peripheral blood cells, bone marrow nucleated cells (BMNC) were detected. Histological observation of bone marrow was made and the CD31 expression was assayed by flow cytometry. The results showed that in ligustrazine treated group the peripheral blood cell, BMNC counts on days 7, 14, 21 after BMT were higher than in the BMT control group (P < 0.01 or P < 0.05). The expression of CD31 in ligustrazine treated group on days 7, 14, 21 after BMT was also higher than in the BMT control group (P < 0.01 or P < 0.05). In conclusion, ligustrazine enhances CD31 expression in bone marrow cells after syngenic bone marrow transplantation of mice, which may be related to the mechanisms underlying the ligustrazine accelerating hematopoietic reconstitution in syngeneic bone marrow transplantation.
Animals ; Blood Cell Count ; Bone Marrow Examination ; Bone Marrow Transplantation ; Female ; Hematopoiesis ; Male ; Mice ; Mice, Inbred BALB C ; Platelet Endothelial Cell Adhesion Molecule-1 ; analysis ; Pyrazines ; pharmacology ; Transplantation, Isogeneic