Objective To study degradation of the antigen-extracted meniscus in PBS solution with no enzyme or with different enzymes. Methods Four types of enzymes (collagenase, hyaluronidase, trypsin, papain) were used to enzymolyze the antigen-extracted meniscus and the fresh meniscus for 3, 7, 15 and 30 days (37℃). The antigenextracted meniscus and the fresh meniscus were immersed in PBS solution (37℃) for 30 days. Weight loss measurement, UV spectrophotometry, and scanning electron microscopy (SEM) were used to characterize the degraded materials. Results The two types of the materials were remarkably digested under the enzymes, especially under trypsin. The degradation curves showed that the antigen-extracted meniscus was enzymolyzed less than the fresh meniscus. The degradation products were grouped as amino, peptide, and polyose by the analysis. Both of the materials could hardly behydrolyzed in PBS solution without the enzymes. The four different enzymes had different surface morphologies under the examination of SEM. Conclusion The antigen-extracted meniscus is enzymolyzed more slowly than the fresh meniscus in vitro, and the result can be used as a guideline to the further research.
Objective To investigate the effect of homograft of marrow mesenchymal stem cells (MSCs) seeded onto poly-L-lactic acid (PLLA)/gelatin on repair of articular cartilage defects. Methods The MSCs derived from36 Qingzilan rabbits, aging 4 to 6 months and weighed 2.5-3.5 kg were cultured in vitroand seeded onto PLLA/gelatin. The MSCs/ PLLA/gelatin composite was cultured and transplanted into full thickness defects on intercondylar fossa. Thirty-six healthy Qingzilan rabbits were made models of cartilage defects in the intercondylar fossa. These rabbits were divided into 3 groups according to the repair materials with 12 in each group: group A, MSCs and PLLA/gelatin complex(MSCs/ PLLA/gelatin); group B, only PLLA/gelatin; and group C, nothing. At 4,8 and 12 weeks after operation, the gross, histological and immunohistochemical observations were made, and grading scales were evaluated. Results At 12 weeks after transplantation, defect was repaired and the structures of the cartilage surface and normal cartilage was in integrity. The defects in group A were repaired by the hylinelike tissue and defects in groups B and C were repaired by the fibrous tissues. Immunohistochemical staining showed that cells in the zones of repaired tissues were larger in size, arranged columnedly, riched in collagen Ⅱ matrix and integrated satisfactorily with native adjacent cartilages and subchondral bones in group A at 12 weeks postoperatively. In gross score, group A(2.75±0.89) was significantly better than group B (4.88±1.25) and group C (7.38±1.18) 12 weeks afteroperation, showing significant differences (P＜0.05); in histological score, group A (3.88±1.36) was better than group B (8.38±1.06) and group C (13.13±1.96), and group B was better than group C, showing significant differences (P＜0.05). Conclusion Transplantation of mesenchymal stem cells seeded onto PLLA/gelatin is a promising way for the treatment of cartilage defects.
To evaluate an improved treatment of an autologous fat injection for hemifacial atrophy to increase the survival rate of the fat graft and decrease complications including colliquation, necrosis, and absorption of the graft fat. Methods From March 1999 to October 2004, 31 patients with hemifacial atrophy underwent an improved treatment by an autologous fat injection for their diseases. There were 12 males and 19 females aged 1928 years (average, 23.5 years). The patients were divided into the following 3 groups according to the atrophy extent: the mild group (n=9), the moderate group (n=19), and the severe group (n=3). Based on the previous researches on the fat transplantation techniques, the improved treatment combined the following strategies that were simply called “3L3M”: low position for the fat donation, low pressure for the fat harvesting, and lowspeed centrifugation for purification of the fat; multipoint, multitunnel, and multiplane for injections of the fat graft. The preoperative and the postoperative photos were taken and the findings were compared to make clear whether the hard and firm masses and cysts existed; then, the decision was made about whether the patients needed another operation according to whether the patients had a natural facial expression and whether the patients had comfortable feelings as well as the ray findings. Results All the patients had a satisfactory symmetrical face after 1 injection of the fat in 15 patients, 2 injections in 13 patients, and 3 injections in 3 patients. The effect of the 3rd injection was better than that of the 2nd injection; the effect of the 2nd injection was better than that of the 1st injection; the fat volume for the injection could be gradually decreased. The fat volumes for injections were as follows: 814 ml (average, 11 ml) in the submaxillary region, 1525 ml (average, 20 ml) in the buccal region, 510 ml (average, 75 ml) in the zygomatic region, and 1820 ml (average, 19 ml) in the forehead region. The followup for 35 years revealed that there wasno infection, hard and firm mass, cyst or other complications. The pigmentationin the affected face was significantly improved. Conclusion Compared with the traditional treatments, the improved treatment of an autologousfat injection for hemifacial atrophy can achieve a satisfactory symmetry of theface with no injury to the donor site or complications in the recipient site. This improved method is an ideal treatment for hemifacial atrophy.
Objective To review research progress of adipose tissuederived stromal cells (ADSCs).Methods The recent articles on ADSCs were extensively reviewed, and the culture and differentiation ability of ADSCs were investigated.Results A population of stem cells could be isolated from adult adipose tissue, they were processed to obtain a fibroblast-like population of cells and could be maintained in vitro for extended periods with stable population doubling. The majority of the isolated cells were mesenchymal origin, with a few pericytes,endothelial cells and smooth muscle cells. ADSCs could be induced to differentiate intomultiple mesenchymal cell types, including osteogenic, chondrogenic, myogenic and adipogenic cells, they could also differentiate into nerve cells.Conclusion ADSCs can substitute mesenchymal stem cells and become an alternative stem cells source for tissue engineering.
Objective To study the biocompatibility of tendon mixedextraction of bovine collagen(tMEBC) and to explore the feasibility of using the threedimensional framework as periodontal tissue engineering scaffold. Methods After being prepared, the tMEBC were cultured with the P4P6 of human periodontal ligament fibroblasts (HPDLFs) in vitro. Threedimensional framework was prepared from bovine tendon. The P4-P6 of HPDLFs (with an initial density of 5×106 cells／ml) were cultured in vitro. Cell attachment andproliferation were measured by cell counting 1 day, 3,5, and 10 days after cell seeding. Histological examination was performed with light microscope and scanning electron microscope 5 and 10 days after cell seeding. Results Porous structure, which supported the proliferation and attachment of HPDLFs, was found in tMEBC. The density of cell increased from 0.556×104 cells／ml 24 hours after cell seeding to 3.944×104 cells／ml 10 days after seeding. Light and scanning electron microscope examinationindicated that HPDLFs were attached and extended on the three-dimensional scaffolds and were well embedded in the newly formed tissue matrix. ConclusiontMEBC has good biocompatibility with the HPDLFs, and can be used as scaffold for cell transplantation in periodontal tissue engineering.
Objective To provide theoretical evidence for clinical application of the epidermal stem cells after an investigation on changes of the epidermal stem cells during the survival process after the fullthickness skin autograft. Methods On the backs of 42 Wistar rats, orthotopic transplantation models (1.5 cm×1.5 cm) of the fullthickness skin autograft were made. According to the time of the specimen taking, at 1, 3, 5, 7, 14, 21 and 30 days after operation, the rats were randomly divided in 7 groups (Groups 1-7). Specimens taken in each group before operation were used as controls. At each time point, the gross observation was made on the transplanted skin flaps, from which the skin tissues were harvested at each time point before and after operation. The routine pathological and the immunohistochemical examinations were performed on the specimens, which were stained by HE and were observed for immunohistochemical changes and the changes in the cells positive for integrinβ-1 and p63. Results All the fullthickness skin autografts survived 3 days after operation except the skin autograft in 1 rat in both Group 5 and Group 6, which was infected around the transplanted skin flap. In Groups 1-4, cell edema, inflammatory cell infiltration, and increased fibrocytes were observed. In Groups 5-7, the maturity degree of the epithelial cells became higher and higher, and the fibrocyte proportion was lowered. In each group the cell positivity rate for integrin β1 was lower than the cell positivity rate for p63. The positive cells were arranged in disorder, distributed into the layers of the epidermis and gradually concentrated in the basal layer of the epidermis and the bulge of the folliculus pili. The positive cells were also found in the other layers of the epidermis.The positive cells were gradually decreased in number, and reached the lowest level in Group 2. There was a significant difference in the above variables in Groups 1,2,3,5,6 and 7 between before and after operations (P＜0.05). Conclusion During the survival process of the fullthickness skin autograft, the proportion of theepidermal stem cells is gradually decreased at first; Then, the proportion isgradually increased, even beyond the normal level; finally, the proportion is decreased again. The distribution of the epidermal stem cells appear in disorder, almost distributed in the layers of the epidermis; finally, the almost normal distribution can be found.
Objective To explore the effective autologous bone marrow stem cell dosage for treatment of severe lower limb ischemia. Methods From December 2003 to December 2004, 22 cases of bilateral lower limb ischemia were treated with autologous bone morrow cell transplantation. All the patients were randomly divided into two groups according to ischemia degree. In group A(severe ischemia side), the amount of transplanted autologous bone marrow cells was more than 1×108, and ingroup B(mild ischemia side), the amount was less than 1×105. A series of subjective indexes, such as improvement of pain, cold sensation and numbness, and objective indexes, such as increase of ankle/brachial index (ABI) and transcutaneous oxygen pressure (TcPO2), angiography, amputation rate, and improvement of foot wound healing were used to evaluate the effect of autologous bone marrow stem cells implantation. Results The rates of pain relief were 90.0% in group A and 16.7% in group B (Plt;0.01); the rates of cold sensation relief were 90.5% in group A and 5.3% in group B（Plt;0.01）;the improvement of numbness was 62.5% in group A and 9.1% in group B（Plt;0.01）. Increase of ABI was 31.8% and 0 in groups A and B respectively（Plt;0.01） at 4 weeks after implantation. Increase of TcPO2was 94.4% and 11.1% in groups A and B respectively（Plt;0.01） at 4 weeks after implantation. Twelve cases of angiography showed rich new collateral vessels in 100% of the limbs in group A while no remarkable new collateral vessel in group B. The amputation rates were 4.5% in group A and 27.3% in group B（Plt;0.05） at 4 weeks after implantation. The rate of improvement of foot wound healing was 75% in group A and there was no changein wound healing in group B after 4 weeks of implantation. Conclusion The effectiveness of autologous bone marrow stem cell implantation depends on the number of implanted stem cells. Effectiveness is expected in most patients if the implanted stem cell is more than 1×108, whereas there would be little effect if the cell number is less than 1×105.
Objective To verify the effect of palatoplasty with or without velopharyngeal muscular reconstruction on the velar movement through the lateral radiography. Methods From October 1988 to October 2000, 62 patients with cleft palate and velopharyngeal insufficiency were treated. Of them, 32 were repaired by velopharyngeal muscular reconstruction (group A) and 30 by modified von Langenbeck’s procedure (group B). The lateral radiographs and cephalometric analysis were taken. The velopharyngeal closure, velar extensibility, the angle changes of velar elevation, the distance changes of velar levator eminence based on anatomy (LEA) to velopharyngeal closure line (VCL), the varieties in LEA, the comparison of LEA and velar levator eminence based on phonation (LEP), the distance comparison between posterior nasal spine (PNS) to LEA and LEP through soft palate line (SPL) were measured. Results During the phonation, group A was significantly greater than group B in the velopharyngeal closure rate(Plt;0.01)， the velar extensibility(Plt;0.05) and the location comparison between LEA and LEP(Plt;0.01); group B was significantly greater than group A in velar elevation angle (Plt;0.05), the varieties in LEA(Plt;0.05). In velar rest position, the distance of LEA to VCL was greater inB group than in A group(Plt;0.01). There was significant difference in thedistance comparison between LEA and LEP(Plt;0.05), difference between LEA and LEP(Plt;0.01) and the distance PNS-SPL-LEA and PNSSPL-LEP(Plt;0.05) within group B; contrary to the results within group A(P＞0.05). Conclusion The velopharyngeal muscular reconstruction in palatoplasty can result in a near normalizationof anatomic measurement of velar levator muscles and improve the velar functionand velopharyngeal competence. Repositioning of velar muscles in a more anatomic correct, transverse position is more important to improve the velar length andaccordant velar movement with velopharyngeal muscles in functional palatoplasty. The velar angle change and velar elevation during phonation are not determinative factors for velopharyngeal competence.
Objective To assess the possibility of placing the posterior pedicle screw on atlas. Methods Twenty human cadaver specimens were used to insert pedicle screws in atlas, through the posterior arch or the pedicle of C1 into the lateral mass. The screw entry point was on the posterior surface of C1 posterior arch and at the intersection of the vertical line through the center of C2 inferior articular process and the horizontal line at least 3 mm below the superior rim of the C1 lamina. The screw of 3.5 mm in diameter was placed in a direction of 10° medial angle and 5° upwardangle. After placement of C1 pedicle screw, the distance from C1 screw entry point to the mediallateral midpoint of C1 pedicle, the maximum length of screw trajectory and the actual screw trajectory angles were measured. The direction of screw penetrating through the cortical of C1 pedicle or lateral mass and the injuries to the vertebral artery and spinal cord were observed.Results Forty pedicle screws were placed on atlas, the mean distance from C1 screw entry point to the medial-lateral midpoint of C1 pedicle was (2.20±0.42)mm, the maximum length of screw trajectory averaged (30.51±1.59)mm, and the actual screw trajectory angle measured (9.7±0.67)° in a medial direction and (4.6±0.59) ° in a upward direction. Only 1 screw penetrated the upper cortical bone of the atlas pedicle because the upward angle was too large, and 8 screws were inserted so deep that the inferior cortical bone of the C1 lateral mass was penetrated. But no injuries to the vertebral artery and spinal cord wereobserved. Conclusion C1 posterior pedicle screw fixation is quite accessible and safe, but the su
Objective To study the mechanism of ectopic osteogenesis of nacre/Polylactic acid (N/P) artificial bone combined with allogenic osteoblasts, and to explore the possibility as a scaffold material of bone tissue engineering. Methods The allogenic- osteoblasts seeded onto N/P artificial bone were co-cultured in vivo 1 week.The N/P artificial bone with allogenic osteoblasts were implanted subcutaneously into the left back sites of the New Zealand white rabbits in the experimental group and the simple N/P artificial bone into the right ones in the control group. The complexes were harvested and examined by gross observation, histologic analysis and immunohistochemical investigation 2, 4 and 8 weeks after implantation respectively.Results In experimental group, the osteoid formed after 4 weeks, and the mature bone tissue withbone medullary cavities formed after 8 weeks; but in control group there was nonew bone formation instead of abundant fibrous tissue after 4 weeks, and more fibrous tissue after 8 weeks.Conclusion N/P artificial bone can be used as an optical scaffold material of bone tissue engineering.