Regulation of stem cells by transforming growth factor Β3/polylactic acid-glycolic acid microspheres / 中国组织工程研究

BACKGROUND: Transforming growth factor Β3/polylactic acid-glycolic acid (TGF-Β3/PLGA) sustained-release microspheres can maintain the effective drug concentration at the site of action and provide the feasibility for efficient utilization of growth factors. OBJECTIVE: To optimize the manufacturing process of TGF-Β3/PLGA sustained-release microspheres, and investigate their effects on the proliferation and migration of rabbit adipose-derived mesenchymal stem cells (ADSCs). METHODS: TGF-Β3/PLGA sustained-release microspheres were prepared by emulsification-solvent evaporation method. The morphology, particle size, drug spatial distribution, encapsulation efficiency, drug loading, and sustained release properties of the microspheres were characterized. The TGF-Β3/PLGA sustained-release microspheres were dissolved in phosphate buffered saline. The concentration of TGF-Β3 in the supernatant was detected at the corresponding time points. The microsphere morphology was observed by scanning electron microscopy at the corresponding time point. Adipose-derived mesenchymal stem cells were divided into six groups and then cultured with single culture medium (negative control) or culture medium containing TGF-Β3 or blank PLGA, or culture medium containing 10,100,1 000 g/L TGF-Β3/PLGA microspheres. Cell proliferation was detected by CCK-8 assay at the corresponding time point. Cells in each group were cultured for 24 hours with corresponding medium in a non-contact manner. The number of migratory cells was counted. RESULTS AND CONCLUSION: (1) TGF-Β3/PLGA sustained-release microspheres were spherical with smooth surface, no adhesion, and evenly distributed particle size. The microspheres had a diameter of 2-50 µm, and the protein drugs in the microspheres were evenly distributed, with high encapsulation efficacy and encapsulation dose. (2) The TGF-Β3/PLGA sustained-release microspheres had good degradation properties and were completely degraded after 6 months in vitro. At the same time, these microspheres had good sustained-release performance and released TGF-Β3 slowly for 45 days in vitro. (3) Blank microspheres and the sustained-release microspheres containing TGF-Β3 had no effect on the proliferation of adipose-derived mesenchymal stem cells. (4) Blank microspheres had no effect on the migration of adipose-derived mesenchymal stem cells, and the transforming growth factor 3 and the sustained-release microspheres containing TGF-Β3 promoted the migration of adipose-derived mesenchymal stem cells. There was no significant difference in the migration promotion between different concentrations of TGF-Β3. (5) These findings suggest that the TGF-Β3/PLGA sustained-release microspheres can promote the migration of adipose-derived mesenchymal stem cells without affecting their proliferation.

Experimental study on the transforming growth factor β3 combined with dental pulp stem cells in early bone integration of implant / 中华口腔医学杂志

Objective@#To establish the experimental model of rabbit mandibular anterior implant repair and evaluate the effects of transforming growth factor (TGF)-β3 and dental pulp stem cells (DPSC) in promoting the bone integration of implant.@*Methods@#The New Zealand rabbits were randomly divided into experimental group, control group and blank group (6 rabbits for each group) . In the experimental group, the implant area was filled with the mixture of TGF-β3, DPSC and Bio-oss powder. In the control group, the implant area was filled with the mixture of DPSC and Bio-oss powder. In the blank group, the implant area was filled with the mixture of phosphate buffer solution and Bio-oss powder. Eighteen New Zealand rabbits were sacrificed in 2 weeks after procedure. The treated alveolar bone tissue was observed. The bone tissue around the implant were estimated by HE staining, immunocytochemical staining and real-time quantitative PCR.@*Results@#The implants were no shedding nor loose. HE staining shows the blank group had a sparse trabecular bone and a small amount of blood vessel around the implant and no obvious new bone formation. The control group showed that the bone trabecula around the implant was sparse and slender, the osteoblasts were arranged linearly around the trabecular bone, a small amount of new bone formation was found around the implant. In the experimental group, there were more thick and dense trabecular bone around the implant, the surrounding osteoblasts were arranged in clusters. The osteoblasts were active and many new bone formed. Typical bone lacunae, bone cells and a large number of new blood vessels can be observed. Immunohistochemistry showed that the proportion of average positive area in the experimental group, control group, blank group were (24.6±5.3) %, (11.3±2.8) % and (7.6±3.8) % respectively. The expression of bone sialoprotein in experimental group were significantly higher than the other 2 groups(P=0.000). Real-time quantitative PCR results showed that the expression level of Runt-related transcription factor 2 (RUNX2), type Ⅰcollagen (COL-Ⅰ), alkaline phosphatase in the experimental group was higher than in the blank group. The expression level of RUNX2 and COL-Ⅰ in the experimental group was higher than that of the control group (P=0.023).@*Conclusions@#TGF-β3 has potential to promote the transformation of DPSC into osteoblasts, which can promote the integration of bone around the implant.

The experimental research of calcium alginate gel loaded transforming growth factor beta 3 and compounded with adipose mesenchymal stem cells to repair rabbit articular cartilage defect / 中国医师杂志

Objective To investigate the effect of tissue engineered cartilage on the repair of artic ular cartilage defects in rabbits with calcium alginate gel (CAG) loaded transforming growth factor beta 3 (TGF-β3) and compounded with adipose mesenchymal stem cells (ADSCs).Methods The ADSCs were separated and cultured in subcutaneous fat of New Zealand white rabbits.The full-thickness articular cartilage defect model was made at the patellar groove by exposure of the femoral ankle joint.ADSCs were implanted into calcium alginate scaffolds loaded with TGF-β3 to construct tissue-engineered cartilage and transplanted into rabbit articular cartilage defects.The animals were divided into four groups:control group (injected with sterile isotonic saline),CAG group (injected with CAG),ADSCs + CAG group (injected with CAG loaded with ADSCs),TGF-β3 + CAG group (injected with CAG loaded with TGF-β3) and TGF-β3 + ADSCs + CAG group (injected with CAG loaded with TGF-β3 and ADSCs).At the end of 12 weeks,the repair of articular cartilage defects was observed by gross observation,hematoxylin-eosin (HE) staining,immunohistochemical staining of safranin-O and type Ⅱ] collagen,and the scoring method developed by In ternational Association of Cartilage Repair (ICRS) Histological score.Results The cartilage repair effect of TGF-β3 + ADSCs + CAG group was better than that of other groups,not only the neonatal tissue and the surrounding normal tissue closely,but also the secretion of extracellular matrix and normal tissue similar.The ICRS scores of each group were (7.06 ±+ 0.18) score,(7.15 + 0.23) score,(7.45 + 0.25) score,(7.47 + 0.24) score and (15.78 ±+ 0.24) score.That of TGF-β3 + ADSCs + CAG group was better than that of other groups,the difference was significant (P ＜ 0.05).Conclusions CAG loaded with TGF-β3 and combined with ADSCs has a good effect in repairing articular cartilage defects in rabbits,and is a structural repair.

Experimental study on the transforming growth factor β3 combined with dental pulp stem cells in early bone integration of implant / 中华口腔医学杂志

To establish the experimental model of rabbit mandibular anterior implant repair and evaluate the effects of transforming growth factor (TGF)-β3 and dental pulp stem cells (DPSC) in promoting the bone integration of implant. The New Zealand rabbits were randomly divided into experimental group, control group and blank group (6 rabbits for each group) . In the experimental group, the implant area was filled with the mixture of TGF-β3, DPSC and Bio-oss powder. In the control group, the implant area was filled with the mixture of DPSC and Bio-oss powder. In the blank group, the implant area was filled with the mixture of phosphate buffer solution and Bio-oss powder. Eighteen New Zealand rabbits were sacrificed in 2 weeks after procedure. The treated alveolar bone tissue was observed. The bone tissue around the implant were estimated by HE staining, immunocytochemical staining and real-time quantitative PCR. The implants were no shedding nor loose. HE staining shows the blank group had a sparse trabecular bone and a small amount of blood vessel around the implant and no obvious new bone formation. The control group showed that the bone trabecula around the implant was sparse and slender, the osteoblasts were arranged linearly around the trabecular bone, a small amount of new bone formation was found around the implant. In the experimental group, there were more thick and dense trabecular bone around the implant, the surrounding osteoblasts were arranged in clusters. The osteoblasts were active and many new bone formed. Typical bone lacunae, bone cells and a large number of new blood vessels can be observed. Immunohistochemistry showed that the proportion of average positive area in the experimental group, control group, blank group were (24.6±5.3) %, (11.3±2.8) % and (7.6±3.8) % respectively. The expression of bone sialoprotein in experimental group were significantly higher than the other 2 groups(0.000). Real-time quantitative PCR results showed that the expression level of Runt-related transcription factor 2 (RUNX2), type Ⅰcollagen (COL-Ⅰ), alkaline phosphatase in the experimental group was higher than in the blank group. The expression level of RUNX2 and COL-Ⅰ in the experimental group was higher than that of the control group (0.023). TGF-β3 has potential to promote the transformation of DPSC into osteoblasts, which can promote the integration of bone around the implant.

Experimental study of transforming growth factor-β3 combined with dental pulp stem cells in promoting the implant's osseointegration / 中华口腔医学杂志

Objective@#To investigate the effect of transforming growth factor-β3 (TGF-β3) and dental pulp stem cells (DPSC) in promoting the implant's osteointegration.@*Methods@#Thirty-three New Zealand white rabbits were randomly divided into phosphate buffer saline (PBS) group, DPSC group and TGF-β3 + DPSC group (12 rabbits/group). Two teeth from the rabbits's mandibular incisors or molars were pulled out randomly, then implant were placed in the tooth extraction site immediately. In PBS group, the implant area was filled with Bio-Oss powder 0.30 g mixed by PBS 20 μl only; while the implant area was filled with Bio-oss powder 0.30 g and 1×108/L DPSC 20 μl in DPSC group; in the the TGF-β3+DPSC group the implant area was filled with Bio-Oss powder 0.30 g mixed with 1×108/L DPSC 20 μl and 80 μg/L TGF-β3 20 μl. Eighteen New Zealand rabbits were executed in the 4 weeks and 8 weeks respectively. The treated alveolar bone tissue and implant were collected for plastic section. Alizarin red staining (ARS), immunohistochemical detection (IHC) of bone sialoprotein (BSP), osteocalcin (OC) and type Ⅰ collagen (COL-Ⅰ) were performed after 4 weeks and 8 weeks. Combined bone lamelta width (CBLW) and implant bone contact rate (IBCR), trabecular width (TW) and trabecular area percentage (TA) were observed by histomorphometric measurement.@*Results@#ARS staining: 4 weeks after the operation, the TGF-β3+ DPSC group showed more red calcified nodules than the other two groups; 8 weeks after operation, the red calcified nodule was further increased. 4 weeks after the operation, the expression of BSP, OC and COL-Ⅰ was (0.35± 0.04), (0.36 ± 0.03) and (0.39 ± 0.01) respectively in TGF-β3+ DPSC group, (0.27 ± 0.02), (0.24 ± 0.01) and (0.28±0.03) respectively in DPSC group, and (0.13±0.03), (0.15±0.02) and (0.16±0.02) respectively in PBS group. Eight weeks after operation, the expression of BSP, OC and COL-Ⅰ was (0.51±0.02), (0.49±0.03) and (0.53±0.02) respectively in TGF-β3+DPSC group, (0.35±0.02), (0.37±0.01) and (0.38±0.01) respectively in DPSC group, and (0.21±0.03), (0.19±0.01) and (0.22±0.02) respectively in PBS group. After 4 weeks and 8 weeks, the expression of BSP, OC and COL-Ⅰ in TGF-β3+DPSC group were significantly higher than the other groups (P<0.05), there was no significant difference between DPSC group and PBS group (P>0.05). Eight weeks after operation, the CBLW, IBCR, TW and TA around implant in TGF-β3+ DPSC group were significantly higher than that in the other groups (P<0.05), there was no significant difference between DPSC group and PBS group (P>0.05).@*Conclusions@#The DPSC has the potential osteogenic differentiation ability; TGF-β3 can accelerate the osteogenic differentiation of DPSC to some extent; TGF-β3 combined with DPSC can effectively promote the implant's osseointegration.

Experimental research on methylation status of CpG islands in promoter region of TGF-β3 and Dnmts during TCDD-induced fetal palatogenisis / 中华整形外科杂志

Objective@#To investigate the correlation between CpG islands methylation statuses of TGF-β3, Dnmts and their expression during TCDD-induced mouse embryonic palatal development.@*Methods@#Eithtteen pregnant C57BL/6J mice were randomly divided into 2 groups: the control group(n=9) and TCDD-exposure group(n=9). At gestation day 10.5(GD10.5), the mice in TCDD-group were orally administrated with TCDD 28 μg/kg, while the mice in the control group received equivalent corn oil. The pregnant mice were sacrificed at GD13.5, GD14.5, GD15.5, fetal palates were collected. CpG island methylation statuses were analysed by methylation specific polymerase chain reaction(MSP). IBM SPSS 20.0 software was applied for statistical analysis. Kolmogorov-Smirnov test was used for normal distribution check, and the distributions were normal. Independent t-test was carried out between two groups. P<0.05 was considered statistically significant.@*Results@#CpG island in promoter region of gene TGF-β3 were all at low methylation level at all GDs of both groups, there were no differences at same GD between two groups [GD13.5: (8.6±0.8)% vs (8.7±0.8)%, P>0.05; GD14.5: (11.5±1.4)%vs (11.7±1.0)%, P>0.05; GD15.5: (12.0±0.7)% vs (12.1±0.5)%, P>0.05]. CpG island in promoter region of gene Dnmt1 were all highly methylated with no differences showed at same GD between two groups [GD13.5: (73.9±1.1)%vs (72.6±0.8)%, P>0.05; GD14.5: (70.8±1.7)% vs (70.7±1.0)%, P>0.05; GD15.5: (69.4±2.2)% vs (69.7±0.5)%, P>0.05]. The methylation level of CpG island 1 in promoter region of gene Dnmt3a in TCDD group was higher than that in control group at GD13.5 and GD15.5 [(21.9±1.1)% vs (8.1±0.6)%, P<0.01, (43.4±0.4)% vs(32.9±0.7)%, P<0.01], while lower at GD14.5[(33.2±0.5)% vs (42.9±0.3)%, P<0.01]. The methylation level of CpG island 2 in promoter region of gene Dnmt3a in control group was higher than that in TCDD group at all GDs [GD13.5: (82.0±0.7)% vs (32.3±0.6)%, P<0.01; GD14.5: (62.7±1.0)%vs (25.5±1.4)%, P<0.01; GD15.5: (47.2±0.4)% vs (30.3±1.4)%, P<0.01].@*Conclusions@#Low methylation level of CpG island 2 which is close to the first exon in promoter region of gene Dnm3a may be the cause of highly expressed Dnmt3a mRNA at GD13.5 during mice palatogenesis induced by TCDD, thus the global DNA methylation is extremely high to induce cleft palate. TCDD-treatment doesn′t influence the CpG methylation statuses in promoter region of TGF-β3 and Dnmt1.

Expression of Tenocyte Lineage-Related Factors from Tonsil-Derived Mesenchymal Stem Cells

Human palatine tonsil-derived mesenchymal stem cells (TMSCs) are known to be a new source of progenitor cells. Using waste tissue after tonsillectomy as a cell provider can be the biggest benefit of TMSCs, compared with other stem cells. The purpose of this study was to investigate tenogenic differentiation of TMSCs and to access the differential effects of transforming growth factor beta 3 (TGF-β3) on the tenogenesis of TMSCs. Human tonsil was obtained after tonsillectomy. Using a cytometric analysis, we were able to find that the TMSCs had typical mesenchymal stem cell markers: positive for CD73, CD90, and CD105, and negative for CD14, CD34, and CD45. Using TGF-β3, the expressions of tenocyte-specific genes and proteins, such as collagen type 1 (COL1), tenomodulin (TNMD), and scleraxis (SCX), were measured by a quantitative polymerase chain reaction (PCR), immunofluorescence staining, immunohistochemistry and Western blot analyses. Quantitative PCR assay showed that TGF-β3 significantly increased the expressions of tenocyte lineage marker genes, including COL1, TNMD, and SCX, at a 3-day treatment, compared with control. However, these increases were not found at long-term exposures (7 or 10 days), except that TNMD expression was maintained at 50 ng/mL at a 7-day exposure to TGF-β3. Like genes, the protein expression levels of COL1, TNMD, and SCX were also induced in TGF-β3-treated TMSCs in a 3-day treatment, which were maintained for 10 days, as evidenced by immunofluorescence staining, immunohistochemistry and Western blot analyses. This study demonstrated that TMSCs in tenogenic stimulation with TGF-β3 have a high tenogenic differentiation potential.

Expression of transforming growth factor beta3 in meniscus development / 中国组织工程研究

BACKGROUND:Since the meniscal blood supply is only in the red zone, the synovial fluid becomes the main nutrition source of the meniscus. As its poor self-healing ability, the meniscus is almost impossible to heal in non-blood supply regions. OBJECTIVE:To investigate the expression and characteristics of transforming growth factor-β3 in the rat meniscus during postnatal development. METHODS:Meniscus tissues of Wistar rats with different ages (1, 7, 14, 21, 28, 35 days) were adopted, and immunohistochemistry and RT-PCR methods were performed to detect the location and expression pattern of transforming growth factor-β3 within the meniscus. RESULTS AND CONCLUSION: Immunohistochemistry results revealed that the expression of transforming growth factor-β3 started from the joint capsule, and gradualy extended to the red zone, gray zone, and white area;RT-PCR results showed the mRNA expression of transforming growth factor-β3 in the rat meniscus at different ages, and during development process gradualy increased until adult stage (rat weight > 100 g), which remained relatively stable.

Effects of transforming growth factor beta 3 on the proliferation and mineralization of dental pulp stem cells from human deciduous teeth / 中国组织工程研究

BACKGROUND:The role of transforming growth factorβsuperfamily has been reported in bone mineralization of various types of stem cells, but the effects of transforming growth factorβ3 (TGF-β3) combined with heparin on proliferation and mineralization of dental pulp stem cells from human deciduous teeth remains to be studied. OBJECTIVE:To evaluate the effects of TGF-β3 on the proliferation and mineralization of dental pulp stem cells from human deciduous teeth. METHODS:The enzyme digestion method was utilized to separately culture dental pulp stem cells from human deciduous teeth. The cellcolony forming efficiency was determined. Flow cytometry was utilized to identify cellsurface marker CD146. Immunochemistry for Vimentin and STRO1 was performed to measure dental pulp stem cells from human deciduous teeth. The third passage dental pulp stem cells from human deciduous teeth cultured in vitro were intervened with heparin and TGF-β3 of 1, 5, 25μg/L mass concentration. The MTS method was applied to measure cellgrowth curves. Alizarin red staining was carried out. The changes in alkaline phosphatase activity were determined with alkaline phosphatase kit. RESULTS AND CONCLUSION:The cellcolony forming efficiency was high. cells were positive for CD146, and strongly positive for Vimentin and STRO1. Dental pulp stem cells from human deciduous teeth were identified. MTS assay indicated that there was no obvious effect on promoting proliferation of dental pulp stem cells from human deciduous teeth after stimulation of TGF-β3. Detection results of alkaline phosphatase activity demonstrated that the combination of TGF-β3 and heparin could strengthen the alkaline phosphatase activity of dental pulp stem cells from human deciduous teeth with increased concentration. Alkaline phosphatase activities were significantly higher in the TGF-β3+heparin group, TGF-β3 group and heparin group than in the control group (P<0.01). Alizarin red staining was positive in the TGF-β3+heparin group, and the staining was strongest in the 5μg/L TGF-β3+heparin group. Results indicated that TGF-β3 combined with heparin promoted mineralization of dental pulp stem cells from human deciduous teeth.

Transforming growth factor beta 3 induced odontoblast-like differentiation of stem cells from human exfoliated deciduous teeth / 中国组织工程研究

BACKGROUND:Studies have reported that the superfamily of transforming growth factors exert a role in the mineralization of various stem cells, but the combination effects of transforming growth factorβ3 and heparin on proliferation and mineralization ability of stem cells from human exfoliated deciduous teeth remains to be studied. OBJECTIVE:To explore the effect of transforming growth factorβ3 on odontoblast-like differentiation of stem cells from human exfoliated deciduous teeth. METHODS:Human deciduous teeth were col ected using enzyme digestion. The 3rd cells were incubated with 25μg/L recombinant human transforming growth factorβ3, 10 U/mL heparin or their combination. The dentin sialophosphoprotein mRNA and dentinsialoprotein expressions were detected by Q-PCR and western blot assay. Alkaline phosphatase activity was determined using alkaline phosphatase kit. RESULTS AND CONCLUSION:Stem cells from human exfoliated deciduous teeth grew wel after induction. The activity of alkaline phosphatase in the combination group was significantly higher than that in the transforming growth factorβ3, heparin and control groups (P<0.01). After combination induction, the cells were strongly positive for alizarin red staining. Results fromα-PCR and western blot assay showed that the expressions of dentin sialophosphoprotein were both remarkably increased at mRNA and protein levels. In summary, stem cells from human exfoliated deciduous teeth can differentiate into odontoblast-like cells under the induction of transforming growth factorβ3 plus heparin.

Effects of exogenous transforming growth factor-β3 on the expression of endogenous transforming growth factor-β3 and proliferation of hepatic stellate cells / 中华消化杂志

Objective To investigate the effects of exogenous transforming growth factor-β3 (TGF-β3) on the expression of endogenous TGF-β3 and proliferation of rat hepatic stellate cells (HSC).MethodsRat HSC cells were seeded in 24-well plates and were divided into 2 groups.One group of cells were exposed to TGF-β3 of different concentrations (0.08,0.4,2,10 and 50 ng/ml) for 2 hours and then the cell culture supernatant was collected; the other group of cells were exposed to 10 ng/ml TGF-β3 and the cell culture supernatant was collected at different time point (0.25,0.5,1,2,4,8 and 13 h).The content TGF-β3 was determined by ELISA method.Some other HSC cells were seeded in 96-well plates and divided into 2 groups.One group of cells were cultured in the presence of exogenous TGF-β3 of different concentrations (0.001,0.005,0.02,0.08,0.32,1.25,5,20,100,500 ng/ml) for 24 hours and then the cell proliferation was detected; the other group of cells were treated with 5 ng/ml TGF-β3 for 24 and 48 hours.The cell proliferation was measured by MTT method.The HSC cell morphology of control group and TGF-β3 treated group was observed under inverted microscope. Results The endogenous TGF-β3 expression of HSC cells obviously increased after exogenous TGF β3 treated at 2 ng/ml and reached the peak at 3 hour [(0.845±0.028) ng/ml vs (0.026±0.021) ng/ml,F=210.168,P=0.00].Low concentrations of exogenous TGF-β3 did not affect the proliferation of HSC cells.Above 0.32 ng/ml,exogenous TGF-β3 could promote HSC proliferation.There was no dose-dependent relationship between HSC cell proliferation and the concentration of exogenous TGF-β3 (F=0.68,P=0.57).Under microscope,lhere was no significant difference in HSC cell morphology between control group and TGF-β3 treated group.Conclusions Exogenous TGF-β3 can promote the expression of endogenous TGF-β3 in HSC cells,and can promote HSC cell proliferation.There is no obvious effect of exogenous TGF-β3 on HSC morphology.

Identification of a mutation in the human raloxifene response element of the transforming growth factor-3 gene

The human transforming growth factor-3 (TGF-3) is an important cytokine to maintain bone mass by inhibiting osteoclast differentiation. Recently raloxifene response element (RRE), a new enhancer with a polypurine sequence for estrogen receptor (ER)-mediated gene activation, was identified on the TGF-3 gene. Functional analysis of the RRE-mediated pathway has shown that this would be an important pathway for bone preserving effect. We found a novel mutation in the RRE sequence by single-strand conformational polymorphism analysis in one of 200 Korean women. Cloning and sequencing revealed a heterozygote in which one allele had an insertion of 20 nucleotides (AGAGAGGGAGAGGGAGA GGG) between nucleotide +71 and +72 and a point mutation at nucleotide +75 (G-A transition), and the other allele had normal sequence. The insertion was a nearly perfect tandem duplication of the wild type DNA sequence. The bone mineral density of the affected woman was not much lower than that of age-matched controls. Transient transfection of the mutant allele showed no significantly different activity compared with that of the wild type allele. These observations suggest that the heterozygote variation of the RRE sequence seems not to be operative in determination of bone mass.

Maxillary growth following cultured epidermal tissue graft and the administration of TGF-beta3 on surgically created palatal defects in rat

This study was designed to evaluate the influence of cultured epidermal tissue graft and the administration of transforming growth factor(TGF)-beta3 on maxillary growth in surgically created palatal defects. A total of 155 rats were divided into 2 groups according to surgical timing : postnatal 2 weeks(n=95), 4 weeks(n=40) and control(unoperated) group(n=20). The postnatal 2-week surgical group was subdivided into 3 groups according to repair methods: conventional surgery(Von Langenbeck technique)group(n=23); cultured tissue graft group(n=25); and full thickness skin graft group(n=25). Additionally, recombinant human TGF-beta3 was administered(30ng or 150ng) on collagen matrix in surgically created palatal defects during surgery(9 conventional surgeries, 9 cultured tissue grafts) in 2-week-old rats. The results showed that all types of surgical treatment decreased maxillary growth compared with the control(unoperated) group(p<0.0001). On the other hand, the tissue graft group, whether cultured tissue or grafted skin, contributed to increased maxillary growth(p<0.0001).And exogenous TGF-beta3 might play a role in connective tissue proliferation and new bone generation during wound healing on palatal defects. Our results suggest that grafting cultured epidermis with collagen matrix decreases the scar tension on maxillary growth more than conventional palatal surgery does. Therefore, exogenous TGF-beta3 may contribute to accelerate wound healing on palatal defects.