Objective To investigate the effect of machine-enzyme digestion method on the residual quantity of small intestinal submucosa (SIS) cell and the content of growth factors. Methods Fresh jejunum of pig within 4 hours after harvesting was prepared into SIS after machine digestion (removing placenta percreta, mucosa, and muscular layer), degrease,trypsinization, abstergent processing, and freeze drying. Samples were kept after every preparation step serving as groups A, B, C, D, and E, respectively (n=4 per group). And the fresh jejunum served as control group (group F, n=4). The histological alteration in each preparation process was reviewed with HE staining and scanning electron microscope (SEM). Nest-polymerase chain reaction (PCR) was used to determine the content of death associated protein 12 (DAP12), and enzyme-linked immunosorbent assay (ELISA) was appl ied to detect the content of vascular endothel ial growth factor (VEGF), basic fibroblast growth factor (bFGF), transforming growth factor β (TGF-β), tumor necrosis factor α (TNF-α). Results HE staining and SEM observation showed that there were residual cells in groups A and B, and there were no residual cells in groups C, D, and E. Nest-PCR test revealed the occurrence of DAP12 in each group. The contents of DAP12 in groups A, B, C, D, E, and F were (18.01 ± 9.53), (11.87 ± 2.35), (0.59 ± 0.27), (0.29 ± 0.05), (0.19 ± 0.04), and (183.50 ± 120.13) copy × 106/cm2. The content of DAP12 in group F was significant higher than that of other groups (P lt; 0.05), groups A and B was higher than groups C, D, and E (P lt; 0.05), there were significantdifferences among groups C, D, and E (P lt; 0.05), and there was no significant difference between groups A and B (P gt; 0.05). The ELISA test showed the content of VEGF, bFGF, TGF-β, and TNF-α in group A was significantly higher than that of groups B, C, D, and E (P lt; 0.05), and there was no significant difference among groups B, C, D, and E (P gt; 0.05). Conclusion SIS prepared by simple mechanical method has more residual cells, while the machine-enzyme digestion method can effectively remove the cells and significantly reduce the DAP12 content. This approach can not obviously reduce the growth factor content in SIS.
Objective To review the development of researches on the stem cells and the tissue engineering technique used in the intestines. Methods We comprehensively reviewed the literature related to the stem cells and the tissue engineering technique used in the intestines, and summarized the conclusions made by the researches concerned. Results The researches on the stem cells and the tissue engineering technique used in the intestines were attractive topics in the recent years and obtained some developments, especially in the field dealing with the characteristics, proliferation and differentiation of the intestinal stem cells as well as the tissue engineering framework of the small intestinal submucosa in vivo. However, the markers for the differentiation of the intestinal stem cells were still a critical problem, which had not been solved yet, and besides, the researches on the intestinal tissue engineering were still in the initial stage. Conclusion There is a broad prospective application of the intestinal stem cells and the tissue engineering technique to the intestinal problem solution. Substantial achievements can be obtained in the treatment of the inflammatory bowel disease, inan exploration on the oncogenesis mechanism, and in the clinical application ofthe intestinal tissue engineering.
Objective
To review the recent progress of the small intestinal submucosa (SIS) in application research of tissue repair and reconstruction.
Methods
The domestic and international articles on the SIS were reviewed and summarized.
Results
As a natural extracellular matrix, SIS has outstanding biological advantages, such as good mechanical property, tissue compatibility, and lower immunogenicity. SIS has been used to repair and reconstruct various types of tissue defects in animal models and clinical application, especially in the treatment of hernia, urinary system disease, and refractory skin trauma. The development of the tissue engineering technology expands the field of SIS repair and reconstruction and promotes the intensive study of SIS. However, the long-term effect of SIS in tissue repair and reconstruction still remains to be further observation, while the cell/SIS material construction by tissue engineering technology also needs more studies.
Conclusion
SIS has a widely promising application future in the tissue repair and reconstruction.
Objective To study an optimal ratio of small intestinal submucosa (SIS) and (hydroxyapatite-tricalcium phosphate,HA-TCP,SIS/HA-TCP) compositions according to the effect of SIS/HA-TCP compositions with different ratios on repairing rabbit femoral condyle defect. Methods Thirty-six rabbits were made into bone defect models of 6 mm in diameter and 10 mm in depth in both sides of femoral condyles. Three different ratios of SIS/HA-TCP compositions (w/w: 1, 0.5, 0.25) were implanted into rabbit femoral condyle defect. After 2, 4, 8 and 12 weeks of operation, the repair effect wasobserved grossly. The histological evaluations were performed by histological scoring system and computer imaging analysis system. Results The amount of new bone formation in SIS/HA-TCP(0.5) group was more than that in SIS/HA-TCP(1) and SIS/HA-TCP(0.25) groups. Histological observation: In SIS/HA-TCP(1) group, few new bone formation was seen and bone defect was repaired in the 12th week. In SIS/HA-TCP(0.5) group, immature woven bone was found in the defect in the 2nd week; more immature woven bone appeared and formed trabeculae in the 4th week; the regenerated bone was vigorously growing into the interspaces of the implanted materials in the 8th week; the implanted materials was basically replaced by bony structure and the lamellar bone appeared in the 12thweek. The results of SIS/HA-TCP (0.25) group were similar to that of SIS/HA-TCP(0.5) group. The histological scoring was higher in SIS/HA-TCP(0.5) and SIS／HA-TCP(0.25) groups than that in SIS/HA-TCP(1) group (Plt;0.05) in the 2nd, 4th, 8th, and 12th weeks. The scoring was higher in SIS/HA-TCP(0.5) roup than that in SIS/HA-TCP(0.25) group in the 2nd and 12th weeks(P＜0.05). In new bone formation and the degradation of HA-TCP, SIS/HA-TCP(0.5) and SIS／HA-TCPC(0.25) groups were superior to SIS/HA-TCP(1) group(Plt;0.05), SIS/HA-TCP(0.5) group was superior to SIS/HA-TCP(0.25) group (Plt;0.05). Conclusion SIS/HA-TCP(0.5) has better effects of repairing bone defect and it can be used as a reference ratio in constructing bone scaffolds.
ObjectiveTo study the feasibility of human adipose-derived stem cells (hADSCs) combined with small intestinal submucosa powder (SISP)/chitosan chloride (CSCl)-β-glycerol phosphate disodium (GP)-hydroxyethyl cellulose (HEC) for adipose tissue engineering.
MethodshADSCs were isolated from human breast fat with collagenase type I digestion, and the third passage hADSCs were mixed with SISP/CSCl-GP-HEC at a density of 1×106 cells/mL. Twenty-four healthy female nude mice of 5 weeks old were randomly divided into experimental group (n=12) and control group (n=12), and the mice were subcutaneously injected with 1 mL hADSCs+SISP/CSCl-GP-HEC or SISP/CSCl-GP-HEC respectively at the neck. The degradation rate was evaluated by implant volume measurement at 0, 1, 2, 4, and 8 weeks. Three mice were euthanized at 1, 2, 4, and 8 weeks respectively for general, histological, and immunohistochemical observations. The ability of adipogenesis (Oil O staining), angiopoiesis (CD31), and localized the hADSCs (immunostaining for human Vimentin) were identified.
ResultsThe volume of implants of both groups decreased with time, but it was greater in experimental group than the control group, showing significant difference at 8 weeks (t=3.348, P=0.029). The general observation showed that the border of implants was clear with no adhesion at each time point;fat-liked new tissues were observed with capillaries on the surface at 8 weeks in 2 groups. The histological examinations showed that the structure of implants got compact gradually after injection, and SISP gradually degraded with slower degradation speed in experimental group;adipose tissue began to form, and some mature adipose tissue was observed at 8 weeks in the experimental group. The Oil O staining positive area of experimental group was greater than that of the control group at each time point, showing significant difference at 8 weeks (t=3.411, P=0.027). Immunohistochemical staining for Vemintin showed that hADSCs could survive at each time point in the experimental group;angiogenesis was most remarkable at 2 weeks, showing no significant differences in CD31 possitive area between 2 groups (P>0.05), but angiogenesis was more homogeneous in experimental group.
ConclusionSISP/CSCl-GP-HEC can use as scaffolds for hADSCs to reconstruct tissue engineered adipose.
Objective To investigate effects of the autologous bone mesenchymal stem cells (MSCs) enriched by the small intestinal submucosa (SIS) film implantation on the myocardial structure, cardiac function, and compensator y circulation after myocardial infarction in the goats. Methods Sixteen black goats were selected and divided randomly into the control group (n=8)and the experimental group (n=8). The chronic myocardial infarction models were made by the ligation of the far end of the left anterior desc ending coronary artery. At the same time, MSCs were aspired from the thigh bone of the goats in the experimental group. MSCs were isolated by the centrifu gation through a percoll step gradient and purified by the plating culture and depletion of the non-adherent cells. Primary MSCs were cultured in the DMEM me dium supplemented with the fetal bovine serum in vitro. After that, the cultures were labeled by 5- BrdU. The active cells were transplanted into the SIS film. Six weeks after the ligation, the MSCs-SIS film was implanted by its being sutured onto the infarction area; whereas, the control group underwent a shamoperation. In both groups, echocardiographic measurements were performed before infarction, 6 weeks after infarction and 6 weeks after the MSC-collagen mplantion, respectively, to assess the myocardial structure and ca rdiac function. The left coronary artery angiography was performed with the digi tal subtraction angiography. Results In an assessment of the left ventricular function, at 6 weeks after operation, t he stroke volume and the ejection fraction of the control group and the experim ental group were 42.81±4.91, 37.06±4.75 ml and 59.20%±5.41%, 44.56%±4.23%, respectively (Plt;0.05). The enddisatolic volume and the endsystolic volume of the control group and the experimental group were 72.55±8.13, 83.31±8.61 ml and 29.75±5.98, 46.25±6.68 ml, respectively (Plt;0.05). The maximal velocity of peak E of contral group and experimental group were 54.8 5±6.35 cm/s and 43.14±4.81cm/s (Plt;0.01); and the maximal velocity of peak A o f control group and experimental grouop were 52.33±6.65 cm/s and 56.91±6.34 cm/s (Pgt;0.05). Echocowdiogr aphy sho wing a distinctly dilatation of left ventricle with the ventricular dyskinesia i n contral group, but without the ventricular dyskinesia in experimental group. T he selective-coronary evngiography revealed that the obvious compensatory circu l ation established between the anterior descending branch and the left circumflex branch in the experimental group. Conclusion Implantation of the autologus MSCs enriched by the SIS film can prevent dilatation of the left ventricular chamber and can improve the contractile ability of the myocardium, cardiac function, and collateral perfusion.
Objective to determine the modulus of elasticity (E) of small intestinal submucosa (SIS), a new biological graft material. Methods The longitudinal tensile testing was performed on 21 specimens of canine jejunum with the electronic material test machine. Results Stress (σ)strain (ε) data were obtained. It was found that the stress (σ)strain (ε) data fitted the expressionσ＝Kεα very well, the mean correlation coefficients R2 was0.991 6.Then the expression of the modulus of elasticity (E) of SIS was E=K1/ασ1-1/α. The mean values of α and K were 3.966 9 and 374.55，so E=4.3992σ0.75. Conclusion The modulus of elasticity was found to increase with increasing stress. The variations law is similar to that of the vessels. Furthermore when σ is 001333 MPa(100 mmHg),E is about 0.16 MPa, which is similar to that of the vessels.
Objective To summarize the basic research and the cl inical use of small intestinal submucosa (SIS), which is used as a degradable material for tissue repair. Methods Recent l iterature concerning SIS at home and abroad was extensively reviewed, and current developments of the basic research and the cl inical use of SIS were investigated. Results SIShad many biological advantages in tissue repair, and was used to repair various tissue defects in animal trials. It had successful outcomes in many cl inical trials to repair hernia, anal fistula and Peyronie diseases. And it also had good results at the early stage to treat dilation of the anastomosis, urethroplasty, hypospadias, and other diseases, however, the long-term follow-up was needed. Conclusion SIS is one kind of good material for tissue repair, and has promising future in the cl inical use.
OBJECTIVE: To review the research advance of the preparation and characteristics of small intestinal submucosa(SIS). METHODS: Recent original articles related to such aspects of small intestinal submucosa were reviewed extensively. RESULTS: Small intestinal submucosa was an easily obtained biomaterial. SIS was a bio-absorbable and degradable material. SIS had tissue specific regeneration properties. CONCLUSION: SIS is a suitable bio-derived material for tissue engineering of blood vessel, muscle tendon, urinary bladder and abdomen.
ObjectiveTo investigate the feasibility of tissue engineered periosteum (TEP) constructed by porcine small intestinal submucosa (SIS) and bone marrow mesenchymal stem cells (BMSCs) of rabbit to repair the large irregular bone defects in allogenic rabbits.
MethodsThe BMSCs were cultivated from the bone marrow of New Zealand white rabbits (aged, 2 weeks-1 month). SIS was fabricated by porcine proximal jejunum. The TEP constructed by SIS scaffold and BMSCs was prepared in vitro. Eighteen 6-month-old New Zealand white rabbits whose scapula was incompletely resected to establish one side large irregular bone defects (3 cm×3 cm) model. The bone defects were repaired with TEP (experimental group,n=9) and SIS (control group,n=9), respectively. At 8 weeks after operation, the rabbits were sacrificed, and the implants were harvested. The general condition of the rabbits was observed; X-ray radiography and score according to Lane-Sandhu criteria, and histological examination (HE staining and Masson staining) were performed.
ResultsAfter operation, all animals had normal behavior and diet; the incision healed normally. The X-ray results showed new bone formation with normal bone density in the defect area of experimental group; but no bone formation was observed in control group. The X-ray score was 6.67±0.32 in experimental group and was 0.32±0.04 in control group, showing significant difference (t=19.871,P=0.001). The general observation of the specimens showed bone healing at both ends of the defect, and the defect was filled by new bone in experimental group; no new bone formed in the control group. The histological staining showed new bone tissue where there were a lot of new vessels and medullary cavity, and no macrophages or lymphocytes infiltration was observed in the defect area of experimental group; only some connective tissue was found in the control group.
ConclusionTEP constructed by porcine SIS and BMSCs of rabbit can form new bone in allogenic rabbit and has the feasibility to repair the large irregular bone defects.
