Clinicopathological significance of CEACAM1 gene expression in breast cancer.

Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a cell adhesion molecule expressed in a variety of cell types. The role of CEACAM1 in breast cancer development and progression is largely unknown. Immunohistochemical analysis was used to examine CEACAM1 expression in breast cancer with long-term follow-up. CEACAM1 expression level in primary breast cancer was low or undetectable. In 65% of the cases, CEACAM1 expression within tumor tissue was lower than that in adjacent tissues. In 20% of the cases, CEACAM1 was negative. In 28.3% of cases, equivalent CEACAM1 expression level was detected in tumor and adjacent tissues. The expression level of CEACAM1 in tumor tissue was negatively correlated with patient mortality, while positively correlated with the expression level of ER+/PR+. CEACAM1 expression was not related with patients' age, pathological classification, lymphatic involvement and the size of tumor. The down-regulation of CEACAM1 was correlated with negative ER-/PR- and might be attributed to the malignant process of breast cancer. The prognosis of the patients with low CEACAM1 expression and high tumor pathological grade were poorer than those patients with high expression and low pathological grade, P < 0.05. Clinically, it is possible to predict the prognosis among the patients of breast cancer by measuring CEACAM1 gene expression in the tumor tissues.

The possible roles of OPN-regulated CEACAM1 expression in promoting the survival of activated T cells and the apoptosis of oral keratinocytes in oral lichen planus patients.

Oral lichen planus is a chronic inflammatory disorder of the oral mucosa that represents T cell-mediated autoimmune diseases. The regulation and roles of carcinoembryonic antigen-related cellular adhesion molecule 1 (CEACAM1), a novel immune molecule, in the immunopathogenesis of T cell-mediated autoimmune diseases remain unclear. In the current paper, CEACAM1 was found to be overexpressed in peripheral T cells and epithelial cells in oral lichen planus patients. A fraction of infiltrating inflammatory mononuclear cells in the lamina propria of the oral lichen planus mucosa also expressed CEACAM1. Importantly, for the first time, CEACAM1 expression in T cells and in normal human oral keratinocytes was demonstrated to be regulated differently by osteopontin in vitro. Furthermore, the apoptosis of oral keratinocytes and activated T cells can be markedly suppressed by CEACAM1-specific monoclonal antibodies. In conclusion, OPN-regulated CEACAM1 expression may play a critical role in the immunopathogenesis of oral lichen planus.

[Effects of silencing inhibitor of DNA binding-1 gene on the growth and invasiveness of adenoid cystic carcinoma cells].

OBJECTIVE: To investigate the role of inhibitor of DNA binding-1 (Id-1) gene in adenoid cystic carcinoma cell growth and invasion behavior. METHODS: With salivary adenoid cystic carcinoma cell lines ACC-M and ACC-2, dedected Id-1 gene expression was screened with immunofluorescence assay. After Id-1 mRNA knocking-down using small interfering RNA, RT-PCR and Western blot were used to detect the different expressions before and after interference, and the growth of cells before and after interference was deceted using the MTT assay, and the cell invasion ability was checked with the use of Transwell chamber assay. RESULTS: Id-1 were both expressed in the ACC-M and ACC-2, and the expression in ACC-M was higher than that in ACC-2. After Id-1 RNA interference, the growth and invasiveness of ACC-M and ACC-2 were inhibited with the restrained degree in ACC-M much stronger than that in the ACC-2. CONCLUSION: In view of the important role of Id-1 in the behavior of growth and invasion in ACC cell, interfering the expression of Id-1 gene is expected to be a novel and effective means for the treatment of adenoid cystic carcinoma.

Over-expression of Gadd45a enhances radiotherapy efficacy in human Tca8113 cell line.

AIM: To investigate the effect of the growth arrest- and DNA damage-inducible Gadd45a gene on the radiosensitivity of human tongue squamous cell carcinoma cell line to ionizing radiation (IR). METHODS: Short interfering ribonucleic acid (si-RNA) targeting Gadd45a or an irrelevant mRNA (nonsense si-RNA) was chemically synthesized. The constructed si-RNAs were transfected into Tca8113 cells and Gadd45a expression was determined using quantitative real-time PCR and Western-blot. After 24-h exposure to IR at a dose rate of 4 Gy/min, apoptosis of Tca8113 cells was detected using flow cytometry, and radiosensitivity was measured using MTT assays. RESULTS: IR apparently increased the expression of Gadd45a at mRNA and protein levels in Tca8113 cells. The effect was efficiently inhibited by transfection with Gadd45a si-RNA (P<0.01). Furthermore, silencing Gadd45a gene significantly increased cell viability and decreased the percentage of apoptotic cells during irradiation, which indicated that IR-induced Gadd45a over-expression could increase the radiosensitivity of Tca8113 cells. CONCLUSION: These results indicated that targeting Gadd45a may have important therapeutic implications in sensitizing Tca8113 cells to IR.

OPN promotes survival of activated T cells by up-regulating CD44 in patients with oral lichen planus.

Oral lichen planus (OLP) is a chronic inflammatory disorder of oral mucosa, which represents cell-mediated autoimmune diseases. Pathological study demonstrated that abundant T lymphocytes infiltrated the oral mucosa, in which the activated T cells that trigger apoptosis of oral epithelial cells is an important mechanism for OLP. However, to date the molecular mechanisms underlying the T lymphocytes infiltration and accumulation in OLP remain unclear. In this paper, we found that the levels of plasma OPN were elevated and were associated with the up-regulated expressions of CD44 in OLP patients. In vitro, the addition of exogenous OPN can suppress the apoptosis of activated CD8(+) T cells via CD44, and this T cell resistance to apoptosis may be attributed to the reduction of endogenous mature granzyme B. Our results suggested that the abnormally elevated levels of OPN may contribute to the abnormal infiltration and accumulation of the activated T cells by up-regulating CD44 in OLP.

[The expression of interleukin-10 mRNA in gingival lesion of different clinical states in patients with adult periodontitis].

OBJECTIVE: To investigate the expression of interleukin-10 (IL-10) mRNA in gingival tissue of active and stable stage in patients with adult periodontitis. METHODS: 12 patients with acute abscesses of the periodontium, 12 patients after periodontal initial treatment and 6 periodontal healthy patients having extraction of impacted wisdom tooth were randomly divided into group A (active stage group), group B (stable stage group) and the control group. Biopsies of gingival tissues were collected from every subject of three groups. Technique of in situ hybridization was applied to observe the expression of IL-10 mRNA in the biopsies from three groups semi-quantitatively. RESULTS: IL-10 mRNA was positively expressed in lymphocytes, macrophages and fibroblasts. The quantity of IL-10 mRNA of group A was the lowest in the three groups and was significantly lower than that of control group and group B respectively (P < 0.01). The quantity of IL-10 mRNA of group B was the highest in the three groups and was significantly higher compared with the control group and group A (P < 0.01). CONCLUSION: The quantities of IL-10 mRNA expression are closely related with various clinical states of periodontitis.

Overexpression of Kif2a promotes the progression and metastasis of squamous cell carcinoma of the oral tongue.

The aim of this study was to examine the Kif2a expression and its role in tumor progression, invasion and metastasis in squamous cell carcinoma of the oral tongue (SCCOT). The study included 44 cases of primary tumor and the corresponding adjacent tissues, 20 cases of primary tumor with lymph node metastasis. Immunohistochemistry was used to observe the Kif2a expression and its correlation with clinicopathologic factors in oral tongue cancer. The immunohistochemistry showed that Kif2a expression was stronger in oral tongue cancer tissues than in paired adjacent tissues (P<0.01), and the higher expression of Kif2a was also significantly associated with lymph node metastasis (P<0.01), tumor clinical stage (P<0.01). In addition, in vitro results from transwell chamber assay showed that Tca8113 cells transfected with Kif2a-siRNA had a decreased migratory ability (P<0.01) compared to nonsense-siRNA-transfected cells. Therefore we speculate the overexpression of Kif2a might be involved in the progression, invasion and metastasis of SCCOT and Kif2a should be as a predictor for prognosis.

[Morphologic observation of oral cancer cells cocultured with mesenchymal cells in vitro].

OBJECTIVE: To study the morphologic and growing alterations of oral cancer cell line Tca8113 before and after cocultured with tumor stromal fibroblasts (TSF) and normal stromal fibroblasts (NSF) respectively, and evaluate the influence of mesenchymal cells on tumor cells. METHODS: TSF and NSF were isolated and cultured. To observe the morphologic change of Tca8113 cells after cocultured with TSF and NSF respectively. RESULTS: When cocultured with NSF, the Tca8113 cells proliferated as rapidly as monocultured to form colonies, while the NSF proliferated slowly to form pieces and then joined each other to form network. The NSF network segmented and surrounded the colonies of cancer cells so that the cancer cells shrank, turn round, broke away from the bottom and floated into the medium. The cancer cells proliferated actively but they were elbow out entirely in the end. TSF proliferated slowly when cocultured with cancer cells, projected several branched protrusions. The cancer cells proliferated along the two sides of protrusions of TSF, or projected short protrusions to connect the body or protrusions of TSF, and overlaid the protrusions gradually, finally, cover the body. In the end, TSF melt away, and the cancer cells took on the figure of TSF. CONCLUSION: The results do suggest that, oral cancer cell line Tca8113 are restrained when coculture with NSF, but are promoted when with TSF.

CEACAM1 distribution and it's effects on angiogenesis and lymphangiogenesis in oral carcinoma.

To investigate the expression of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) and its effects on angiogenesis and lymphangiogenesis in oral carcinoma. Immunohistochemistry was used to study the expression of CEACAM1, LYVE1 and CD31, double-labelling immunofluorescence was used to detect the co-expression of CEACAM1 and LYVE1, and double-labelling immunohistochemistry was performed to observe the co-expression of LYVE1 and CD31 in vessels. Membranous CEACAM1 was expressed in well-differentiated squamous cell carcinoma and cytoplastic CEACAM1 in poorly and moderately differentiated carcinoma (P<0.05). More CEACAM1-positive vessels were observed in CEACAM1-positive tumors with cytoplasmic expression than with membranous expression (P<0.001). Co-expression of CEACAM1 and LYVE1, LYVE1 and CD31 in vessels was more common in CEACAM1-positive tumors with cytoplasmic expression than with membranous expression (P<0.001). CEACAM1 has different distribution in oral carcinoma. Membranous CEACAM1 inhibits angiogenesis and lymphangiogenesis, but cytoplasmic CEACAM1 promotes angiogenesis, and even promotes lymphangiogenesis by mediating the transformation of vascular endothelial cells (VECs) into lymphatic endothelial cells (LECs).

The different expression of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) and possible roles in gastric carcinomas.

The purpose of this study was to investigate the expression of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) and its effects on promoting angiogenesis in gastric adenocarcinomas. Paraffin wax sections of 222 patients with gastric adenocarcinomas having undergone surgery between 2001 and 2006 were classified into three histotypes: intestinal, diffuse, and mixed carcinomas following the Laurén classification. Immunohistochemistry (IHC) was used to study the distribution of CEACAM1, and double-labeling immunohistochemistry was used to observe the relationship between CEACAM1 expression and neovascularization in carcinoma areas. No CEACAM1 expression was found in normal non-metaplastic mucosa adjacent to the tumors; but in metaplastic mucosa, CEACAM1 was expressed on the apical surface. However, all of the collected gastric carcinomas expressed CEACAM1 with cytoplasmic or membranous staining. CEACAM1 was expressed mainly with a membranous pattern in the intestinal carcinomas, and with a cytoplasmic pattern in the diffuse carcinomas. There was a significant difference between the expression patterns and the histotypes (P<0.0001). CEACAM1 expression was classified as high (> or =66% positive cells) and low (<66% positive cells), and high CEACAM1 expression was associated with lymph nodes metastasis (P<0.05). High microvessel density (MVD) was observed more frequently in the tumors with membranous expression, and low MVD in the tumors with cytoplasmic staining (P<0.0001). The transformation of CEACAM1 distribution from membrane to cytoplasm is an important incident for the reverse effects on the tumorous angiogenesis, and high expression of CEACAM1 facilitates the metastasis of carcinoma cells to lymph nodes. Moreover, the different distribution of CEACAM1 in the intestinal and diffuse carcinomas indicates a different tumorigenic pathway.

Expression of ER, Ki-67 and cylinD1 in the pre-cancerous breast of Chinese patients.

To investigate the expression and association of ER, Ki-67 and cyclinD1 in usual ductal hyperplasia(UDH), atypical ductal hyperplasia (ADH) and ductal carcinoma in situ(DCIS) in the breast. The study included 56 cases of pre-cancerous lesions which were surgically excised at Qi Lu Hospital of Shangdong University. Immunohistochemistry was used to determine the expression of ER, Ki-67 and cyclinD1 and double-labelling immunofluorescence technique was used to observe the coexpression of ER and Ki-67. The expression and distribution of ER-positive cells were significantly different in UDH, ADH and DCIS. The ER-positive cells were much more in UDH than in normal TDLUs (terminal duct lobular units). The distribution of ER-positive cells interspersed amid ER-negative cells within UDH. However , the ER positive cells showed marked increases in ADH and low grade nuclear DCIS (P < 0.05), distributing in almost all constituent cells. The expression of ki-67 and cyclinD1 were significantly different between UDH and DCIS (P < 0.05) , and a positive correlation was found between expression of Ki-67 and morphological classification of pre-cancerous lesions (r = 0.3522, P < 0.05) as well as cyclinD1 (r = 0.3901, P < 0.05). Double-labelling immunofluorescence showed that there was no coexpression of ER and Ki-67 in normal breast tissue. The coexpression of the two markers was found in ADH and increased in DCIS. Overexpression of ER, Ki-67 and cyclinD1 significantly accompanies the transition of normal cells and UDH to ADH and DCIS. The coexpression of ER and ki-67 may present the early change in carcinogenesis of breast cancer.

[Effect of curcumin on invasion and migration of tongue squamous cell carcinoma cell line Tca8113].

OBJECTIVE: To study the effect of curcumin on invasion and migration of the tongue squamous cell line Tca8113. METHODS: Tca8113 cells were treated with curcumin (0 - 100 micromol/L) for 24 h and the conditional medium was collected. The gelatinases - matrix metalloproteinase -2 and -9 (MMP-2, -9) in the conditional medium were detected by gelatin zymography. The cell invasion and migration model in vitro was conducted using transwell chamber with or without matrigel. Using this model, the effects of 50 micromol/L curcumin on invasion and migration of Tca8113 were detected. RESULTS: Curcumin reduced the activities of MMP-2 and MMP-9 on a dose-dependent manner. Curcumin can suppress cell invasion and migration significantly (P <0.01). CONCLUSIONS: Curcumin can suppress Tca8113 invasion and migration by reducing the activities of MMP-2 and MMP-9.

[Selection and construction of cell line stably expressing survivin gene in lower level through eukaryotic plasmid vector of shRNA].

PURPOSE: To construct a short hairpin RNA(shRNA) interference expression plasmid vector of survivin gene, transfect tongue squamous cell carcinoma line Tca8113 which expressed survivin gene in a high level, and choose the cells whose survivin gene were suppressed significantly. METHODS: Two pairs of oligonucleotide sequences specific for survivin gene were designed and synthesized, and cloned into pSilencer-2.1U6-neo plasmid. The recombinant plasmids (named PS1 and PS2) were amplified in Ecoli. DH5alpha was identified by restriction digestion, PCR and sequencing. The vectors were transfected into Tca8113 cells with lipofectamine 2000. After selection with G418, the stable cell clones were attained. Survivn expression was assayed with real-time quantitative PCR and Western blotting. SAS8.0 software package was used for Student t test. RESULTS: Two vectors were constructed successfully and stable cell clones with PS1 or PS2 plasmid were obtained. As compared with those of control, survivin expression of transfected cell with PS1 or PS2 in mRNA level was significantly suppressed (P<0.05). In protein level, only those of transfected cell with PS2 was significantly suppressed (P<0.01). CONCLUSIONS: The shRNA interference expression plasmid vectors of survivin gene are successfully constructed, and Tca8113 cells which express survivin gene in a stable lower level are attained, which enable us to carry out further research on gene therapy of oral squamous cell carcinoma. Supported by National Natural Science Foundation of China (Grant No.30572056).

[Effect of bone morphogenetic proteins-2 on mandibular distraction osteogenesis in rabbits].

OBJECTIVE: To investigate the Effect of recombinant adenovirus vectors containing human Bone morphogenetic proteins-2 (Ad-hBMP-2) on the for mandibular distraction osteogenesis (DO) in rabbits. METHODS: Twenty-four New Zealand white rabbits were randomly divided into experimental group, and control group and underwent mandibular distraction osteogenesis. After 5 days latency, the distracters were activated at a speed of 0.5 mm every 12 hours for 7 days, then on the first day in the consolidation period, the distraction gaps of experimental group were injected with 0.2 ml Ad-hBMP2 10(12) pfu/L, while the animals of control group were injected with 0.2 ml Ad-EGFP 10(12) pfu/L. At the 7 th and 28 th day of consolidation period, specimens were obtained, X-ray and histomorphology were performed. The bone density and the quantity of new bone formation in the distraction gaps were observed and compared between the two groups at different consolidation period. RESULTS: Ad-hBMP-2 treated specimens demonstrated an increased amount of new bone formation CONCLUSIONS: Adenovirally-mediated delivery of BMP-2 can locally increase bone deposition during DO, which may potentially shorten the consolidation period.

[Effect of hypoxia inducible factor-1 alpha on vascular endothelial growth factor expression in human tongue squamous carcinoma cells (Tca8113) under hypoxia].

OBJECTIVE: To investigate the effect of hypoxia inducible factor-1 alpha (HIF-1 alpha) on vascular endothelial growth factor (VEGF) expression in Tca8113 cells under hypoxia. METHODS: The expression of the mRNA of HIF-1 alpha and VEGF in Tca8113 cells was examined by RT-PCR technique at different culture times (1/2 h, 1 h, 3 h, 6 h, 12 h, 24 h) under normoxic and hypoxic conditions. RESULTS: The expression of HIF-1 alpha under hypoxia showed the trend of increasing first and then decreasing, and was higher than that of the control (normoxic group) at 6h and 12 h (P < 0.05). The expression of VEGF under hypoxia was higher than that of the control group at 1/2 h, 1 h, 3 h, 12 h, 24 h (P < 0.05). The expression of hypoxia-induced VEGF mRNA increased with the increased expression of HIF-1 alpha mRNA in the cell lines tested at the initial stage of hypoxia. But no statistical significant association was observed between HIF-1 alpha and VEGF expression within 24 h under hypoxia (rs = 0.5750, P > .005). CONCLUSIONS: The increased expression of VEGF in Tca8113 cells might be mediated by multiple factors, including HIF-1 alpha.

[Experimental study of mandibular periosteal distraction in rabbits].

OBJECTIVE: To investigate a novel technique for new bone formation--periosteal distraction osteogenesis. METHODS: A custom made periosteal distraction device was fixed to bilateral surface of the mandible in three rabbits. Periosteal distraction was performed on the left side of the mandible, the right side of the mandible served as the control. The animals were sacrificed at the end of distraction process. All the specimens were X-rayed and histologically examinated. RESULTS: All three animals survived with no obvious complications. Both in mass specimens and X-rays, there showed new bone formation on the distracted side of the mandible. In histological examinations, there was osteoblast-like cell infiltration and bone tissue formation in the distracted area. CONCLUSION: Periosteal distraction osteogenesis can provide a novel technique for the repair of bone defects.

[Study of induction of tumor specific cytotoxic T lymphocyte by using tumor-derived exosome].

OBJECTIVE: To investigate whether the exosomes derived from Tca8113 could induce production of tumor-specific T cells when pulsed onto dendritic cells. METHODS: Tca8113 cell was cultured with RPIM1640, isolated and purified the tumor-derived exosomes from the culture supernatants by ultrafiltration with Millipore centrifual filter devices; frozen and thawed Tca8113 cells to get frozen tumor antigens (FTA). The exosomes and FTA was pulsed onto DC generated from normal human peripheral blood mononuclear cell (PBMC) in vitro. The DC pulsed with FAP or exosomes cocultured with the peripheral blood lymphocytes to transform T cell into specific CTL. To observe the killing and wounding activity of CTL, the CTL and Tca8113 cells were mixed at a ratio of 20 to 1, SPCA-1 cells and 95-D cells was evaluated as control group. RESULTS: The CTL induced by DC pulsed with FAP or exosomes had significant activity killing Tca8113 (P < 0.01); Moreover the CTL induced by DC pulsed with exosomes could also kill SPCA-1 cells (P < 0.05), but the CTL induced by DC pulsed with FTA had not this function. CONCLUSION: Exosomes derived from tumour accumulate in culture supernatants. Exosomes are a natural and new source of tumour-rejection antigens, opening up new avenues for immunotherapy against oral cancers. The exosome-specific CTL could kill another kind of tumor, so tumor-derived exosomes may contain shared tumor-rejection antigens.

[Expression of stromal CD34 and alpha-smooth muscle actin in oral invasive cancer].

OBJECTIVE: To observe the expression of stromal CD34 and alpha-smooth muscle actin (alpha-SMA) in oral invasive cancer. METHODS: The distribution and expression of stromal CD34 and alpha-SMA in 60 patients with invasive oral squamous cell cancer and 20 resections with tumor-free margins from 60 patients with invasive oral squamous cell cancer were examined by immunohistochemistry SP method. RESULTS: Twenty cases of resections of mucosa with tumor-free margins exhibited CD34 expression in fibrocytes cytoplasm in the vicinity of vessels, mucosa and submucosa but were free of alpha-SMA expression, only the walls of muscularized vessels stained positive for alpha-SMA in the stroma. Sixty invasive oral squamous cell carcinomas were free of stromal CD34 expression, only the endothelia of small vessels stained positive for CD34, of which 53 invasive oral squamous cell carcinomas expressed stromal alpha-SMA in myofibroblasts cytoplasm. CONCLUSIONS: The detection of CD34 and alpha-SMA is an adjunctive tool in judging oral cancer invasion in small oral mucosa biopsy specimens based on the absence of CD34 expression paralleled by the gain of alpha-SMA in the stroma of oral invasive cancer.

[Clinical study of oral and maxillofacial malignancies associated with multiple primary malignant neoplasms].

OBJECTIVE: To study the clinical characteristics, diagnosis and treatment strategy of oral and maxillofacial malignancies in multiple primary malignant neoplasms (MPMNs). METHODS: 21 cases of oral and maxillofacial malignancies associated with MPMNs admitted to our hospital between 1985 and 2000 were studied respectively. RESULTS: There were 44 malignant cases in the 21 MPMNs patients. Among the 44 cases, there were 24 cases in alimentary and respiratory tract such as oral, pharynx, esophagus, stomach and lung, and 10 cases in salivary gland, breast and female reproductive system. There were 25 cases malignant neoplasms in oral and maxillofacial region where squamous cell carcinoma was the most common pathologic type, secondly adenoid cystic carcinoma. In oral and maxillofacial region, MPMNs were often found in tongue, parotid and submandibular gland, buccal mucosa and gingival. CONCLUSION: Tongue and salivary gland were the common locations with MPMNs, and they were closely associated with alimentary and respiratory tract. Patients with malignancies of oral and maxillofacial region associated with MPMNs must be submitted to a long-term and careful follow-up. For female patients, breast and female reproductive system should be examined specially. Regular follow-up, early detection, early diagnosis, active and effective treatment can help to improve the survival quality of MPMNs patients.

[Repair of mandibular central fissures in rabbits with hBMP-2 gene modified tissue engineered bone].

PURPOSE: To evaluate the effectiveness of the tissue engineered bone substitute loaded with periosteal-derived osteoblasts (POBs) transfected by adenovirus mediated human bone morphogenetic protein-2 gene (Ad-hBMP-2) in the repair of rabbit mandibular central fissures. METHODS: 45 rabbits with the soft tissue in the mandibular central fissures removed were randomly divided into 5 groups, group I: Ad-hBMP-2 transfected POBs/bioglass group (n=10); group II: adenovirus mediated enhanced green fluorescent protein (Ad-EGFP) gene transfected POBs/bioglass group (n=10); group III: untransfected POBs/bioglass group (n=10); group IV: single bioglass group (n=10); group V: control group (n=5). The above bone substitutes were implanted in the rabbit mandibular central fissures respectively except group V. The samples were studied by gross, X-ray, histomorphology, histomorphometrical analysis and biomechanics after 2, 4, 8, 12 weeks respectively. One-way ANOVA was used for statistical analysis. RESULTS: In gross view, the rabbit mandibular central fissures in group I were replaced by new bone including cortical bone from the 4th week. X-ray examination showed that the higher bone density was found in the rabbit mandibular central fissures of group I 4 to 8 weeks after implantation. Histomorphometrical analysis showed much more new bony callus in group I than in other groups (P<0.01). The maximal anti-bending load and bending rigidity of the implanted bone substitute of group I were significantly higher than those of group II, III and IV (P<0.01). CONCLUSION: The tissue engineered bone substitute loaded with POBs transfected by human BMP-2 gene could get the best result in the repair of rabbit mandibular central fissures, therefore, it is likely to be used in the repair of alveolar clefts.