高生物活性复合人工骨修复骨缺损的实验研究

目的 :探讨载有转化生长因子 β1(TGF- β1) ,具有高生物活性复合人工骨修复骨缺损的价值。方法 :建立SD大鼠下颌骨缺损模型。采用组织学和分子生物学 Slot Blot杂交法 ,检测羟基磷灰石复合 TGF- β1实验组 ,单纯羟基磷灰石对照组 ,无材料修复的空白对照组在骨缺损愈合过程中的组织学及 型胶原 m RNA的表达状况。结果 :实验组的新骨形成及 型胶原 m RNA的表达水平最高。结论 :提示载有 TGF-

Termination of TGF-β Superfamily Signaling Through SMAD Dephosphorylation——A Functional Genomic View

The transforming growth factor-β （TGF-β） and related growth factors activate a broad range of cellular responses in metazoan organisms via autocrine, paracrine, and endocrine modes. They play key roles in the pathogenesis of many diseases especially cancer, fibrotic diseases, autoimmune diseases and cardiovascular diseases. TGF-β receptor-mediated phosphorylation of R-SMADs represents the most critical step in the TGF-β signaling pathways that triggers a cascade of intracellular events from SMAD complex assembly in the cytoplasm to transcriptional control in the nucleus. Conversely, dephosphorylafion of R-SMADs is a key mechanism for terminating TGF-β signaling. Our labs have recently taken an integrated approach combining functional genomics, biochemistry and development biology to describe the isolation and functional characterization of protein phosphatase PPM1A in controlling TGF-β signaling. This article briefly reviews how dynamic phosphorylation and dephosphorylation of SMADs control or fine-tune the signaling strength and duration and ultimately the physiological consequences in TGF-β signaling.

The involvement of p38 MAPK in transforming growth factor β1-induced apoptosis in murine hepatocytes

We reported in this manuscript that TGF-beta1 induces apoptosis in AML12 murine hepatocytes, which is associated with the activation of p38 MAPK signaling pathway. SB202190, a specific inhibitor of p38 MAPK, strongly inhibited the TGF-beta1-induced apoptosis and PAI-1 promoter activity. Treatment of cells with TGF-beta1 activates p38. Furthermore, over-expression of dominant negative mutant p38 also reduced the TGF-beta1-induced apoptosis. The data indicate that the activation of p38 is involved in TGF-beta1-mediated gene expression and apoptosis.

Transforming growth factor beta can be a parameter of aggressiveness of pT1 colorectal cancer

AIM: To evaluate the significance of transforming growth factor beta (TGF β) expression, in correlation with histopathological parameters, at the front of invasion in T1 colorectal cancer (CRC) and presence of metastases. METHODS: TGF p immunohistochemical expression was studied in 34 specimens of colorectal adenocarcinomas (pT1). A three-step avidin-biotinylated immuno-peroxidase (ABCu-NCL) staining technique was performed on 4-μm paraffin-embedded tissue sections with a monoclonal antibody to TGF β (Novocastra, NCL-TGFB, clone TGFB 17, dilution 1:40). RESULTS: Seventeen (50%) out of 34 lesions were positive for TGF p expression. The TGF β-positive rate in patients with vascular invasion was significantly higher than in those without vascular invasion (11/14 cases, P<0.01, P= 0.005). The TGF p-positive rate was observed in 91.7% of patients with presence of tumor budding at the front of invasion (11/12 cases, P<0.01, P= 0.0003). A statistically significant correlation was found between the presence of lymph node metastases and positive expression of TGF β (14/16 cases, P<0.01, P= 0.0001). We also observed that the TGF β-positive rates in groups with distant and non-distant metastases were 92.8% and 20% respectively, and a significant correlation between TGF β expression and distant metastasis was shown (P<0.01, P= 0.00003). CONCLUSION: The evaluation of TGF β expression of protein in association with histological parameters can be used as a parameter of the aggressiveness of pT1 CRC.

Tetrandrine inhibits activation of rat hepatic stellate cells in vitro via transforming growth factor-β signaling

AIM: To investigate the effect of various concentrations of tetrandrine on activation of quiescent rat hepatic stellate cells (HSCs) and transforming growth factor-β (TGF-β) signaling in vitro.METHODS: HSCs were isolated from rats by in situperfusion of liver and 18% Nycodenz gradient centrifugation, and primarily cultured on uncoated plastic plates for 24 hwith DMEM containing 20% fetal bovine serum (FBS/DMEM) before the culture medium was substituted with 2% FBS/DMEM for another 24 h. Then, the HSCs were cultured in 2% FBS/DMEM with tetrandrine (0.25, 0.5, 1,2 mg/L, respectively). Cell morphological features were observed under an inverted microscope, smooth muscleα-actin (α-SMA) was detected by immunocytochemistry and image analysis system, laminin (LN) and type Ⅲprocollagen (PCⅢ) in supernatants were determined byradioimmunoassay. TGF-β1 mRNA, Smad 7 mRNA and Smad 7 protein were analyzed with RT-PCR and Western blotting, respectively.RESULTS: Tetrandrine at the concentrations of 0.25-2 mg/L prevented morphological transformation of HSC from the quiescent state to the activated one, while α-SMA, LN and PCⅢ expressions were inhibited. As estimated by gray values, the expression of α-SMA in tetrandrine groups (0.25, 0.5, 1, 2 mg/L) was reduced from 21.3% to 42.2%(control: 0.67, tetrandrine groups: 0.82, 0.85, 0.96, or 0.96, respectively, which were statistically different from the control, P＜0.01), and the difference was more significant in tetrandrine at 1 and 2 mg/L. The content of LN in supernatants was significantly decreased in tetrandrine groups to 58.5%, 69.1%, 65.8% or 60.0% that of the control respectively, and that of PCⅢ to 84.6%, 81.5%,75.7% or 80.7% respectively (P＜0.05 vs control), with no significant difference among tetrandrine groups. RTPCR showed that TGF-β1 mRNA expression was reduced by tetrandrine treatments from 56.56% to 87.90% in comparison with the control, while Smad 7 mRNA was increased 1.4-4.8 times. The TGF-β1 mRNA and Smad 7 mRNA expression was in a significant negative correlation (r= -0.755, P＜0.01), and both were significantly correlated with α-SMA protein expression (r = -0.938, P＜0.01;r = 0.938, P＜0.01, respectively). The up-regulation of Smad 7 protein by tetrandrine (1 mg/L)was confirmed by Western blotting as well.CONCLUSION: Tetrandrine has a direct inhibiting effect on the activation of rat HSCs in culture. It up-regulates the expression of Smad 7 which in turn blocks TGF-β1 expression and signaling.

TGF-β and cancer： Is Smad3 a repressor of hTERT gene？

Transforming growth factor β （TGF-β） carries out tumor suppressor activity in epithelial and lymphoid cells, whereas telomerase is required for most cancers. Although the molecular mechanisms by which TGF-β acts as a tumor suppressor are yet to be fully established, a link between TGFb and its tumor suppressor activity by telomerase has been suggested. Recently, we have noted a novel mode of action for TGF-β through which human telomerase reverse transcriptase （hTERT） gene is repressed in immortal and neoplastic cells, confirming that one of the mechanisms underlying TGF-β suppression of tumor growth may be through inhibiting hTERT gene transcription. Moreover, the inhibition of hTERT gene by TGF-β suggests a cis action of the TGF-β signaling molecule Smad3 on hTERT promoter directly. This article examines our current understanding and investigation of TGF-β regulation of telomerase activity, and presents a model in which Smad3 participates in regulating hTERT gene transcription by acting as a repressor directly. Engineering the interface between Smad3 and hTERT gene may lead to a new strategy to inhibit telomerase activity in cancer.

Effect of transforming growth factor beta and bone morphogenetic proteins on rat hepatic stellate cell proliferation and transdifferentiation

AIM: To explore different roles of TGF-β (transforming growth factor beta) and bone morphogenetic proteins (BMPs)in hepatic stellate cell proliferation and trans-differentiation.METHODS: Hepatic stellate cells were isolated from male Sprague-Dawley rats. Sub-cultured hepatic stellate cells were employed for cell proliferation assay with WST-1 reagent and Western blot analysis with antibody against smooth muscle alpha actin (SMA).RESULTS: The results indicated that TGF-β1 significantly inhibited cell proliferation at concentration as low as 0.1 ng/ml, but both BMP-2 and BMP-4 did not affect cell proliferation at concentration as high as 10 ng/ml. The effect on hepatic stellate cell trans-differentiation was similar between TGFβ1 and BMPs. However, BMPs was more potent at transdifferentiation of hepatic stellate cells than TGF-β1. In addition, we observed that TGF-β1 transient reduced the abundance of SMA in hepatic stellate cells.CONCLUSION: TGF-β may be more important in regulation of hepatic stellate cell proliferation while BMPs may be the major cytokines regulating hepatic stellate cell transdifferentiation.

Expression patterns of transforming growth factor-beta and its receptors in gastric mucosa of patients with refractory gastric ulcer

AIM: Transforming growth factor-β (TGF-β) plays a regulatory role in tissue repair. In a previous study, we found that TGF-β and its receptors were expressed in gastric mucosa of patients with well-healed gastric ulcers, as demonstrated by immunohistochemistry. To further characterize the role of TGF-β and its receptors in repairing gastric ulcers, we investigated the expression patterns of TGF-β and its receptors in gastric mucosa by in situ hybridization and reverse transcriptase-polymerase chain reaction (RT-PCR).METHODS: Seventy-four patients with endoscopically proven gastric ulcers were eligible for participation in this study. All patients had routine biopsies on initial endoscopy and were then treated for 12 wk with an H2 blocker. Repeat endoscopy was then performed. There were 8 patients with poorly healed ulcers, and biopsies were taken from the margin of the residual ulcers. These tissue samples, along with biopsy of gastric mucosa near the original ulcers from 8 randomly selected patients with well-healed ulcers were examined for TGF-β and TGF-β receptor Ⅱ mRNA by RT-PCR and in situ hybridization, as well as immunohistochemistry.RESULTS: TGF-β and TGF-β receptor Ⅱ were strongly expressed in tissues from patients with well-healed ulcers.Four of the 8 patients with poor healing had low or absent expression of TGF-β or TGF-β receptor Ⅱ mRNA. All cases positive by RT-PCR assay were confirmed by in situ hybridization as well as immunohistochemistry.CONCLUSION: It is suggested that TGF-β and its receptors are important for gastric ulcer healing. These results may have implications for further investigation of the healing process and in predicting response to therapy.

ROLE OF TRANSFORMING GROWTH FACTOR β(TGF-β)IN REPAIRING OF BONE DEFECTS

TGF-β is a multifunctional cytokine that regulates many aspects of cellular function, including periosteal mesenchymal cell proliferation, differentiation. This experiment is to study its effects on bone defect repair. A rabbit radial bone defect model was used to evaluate the effect of TGF-β, which was extracted and purified from bovine blood platelets, on the healing of a large segmental osteoperiosteal defect. A 1. 5-centimeter segmental defect was created in the mid-upper part of the radial shaft of adult rabbits. The defect was filled with implant containing TGF-β that consisted of carrier and bovine TGF-β. Limbs served as controls received carrier alone. The defectswere examined radiographically and histologically at 4, 8,12 , 16 and 20 weeks after implantation. The results showed that in TGF-β implant group . the defect areas at 12 weeks post operation were bridged by uniform new bone and the cut ends of cortex could not be seen;while in control group, the defects remained clear. Only a small amount of new bone formed as a cap on the cut bone ends. In the experimental group, new lamellar and woven bone formed in continuity with the cut ends of the cortex. An early medullar canal appears to be forming and contained normal-appearancing marrow elements; while the control group displayed entirely fibrous tissue within the defect site. Remnants of the cancellous bone carrier were observed in the control specimen. These data demonstrate that exogenous TGF-β initiate osteogenesis and stimulate the bone defects repair in animal model.

EXPRESSION AND LOCALIZATION OF SMAD4 PROTEIN IN RAT TESTIS DURING THE POSTNATAL DEVELOPMENT

Objective.To study expression and localiza tion of Smad4protein,the common-mediator Smad,which is one of intracellular signaling molecules of transforming growth factor-afamily,in rat t estis during postnatal development.Methods.In this study,whole testes were co llected from S.D rats aged3days,7days,14days and28days,and adult respe ctively.We examined the cellular localization and developmental change of Smad 4in rat testis by immunohistochemical ABC method with glucose oxidase-DAB-nic kel enhancement technique;the quantitative analysis of the immunostaining by t he image analysis system;the Smads pro-teins coexistence in the adult rat test is by the double immune staining for CD14-Smad4and Smad2-Smad4;the prote in expression of Smad during rat testicular development by means of Western blo ts.Results.The protein of Smad4was present in rats from3days of age to adul thood,and the im-munolocalization was exclusively localized to the cytoplasm o f Leydig cells with negative nuclei in the in-terstitial tissue at any time po int.No expression was detected in germ cells.The result of image and sta-tis tical analysis showed that generally,there was a tendency that the expression o f Smad4in the testes increased gradually with the rats developing maturation.C onclusion.Our data provide direct evidence for the molecular mechanism of TGF-aaction in rat testes during postnatal development and spermatogenesis of ra ts.

Association of overexpression of TIF1γ with colorectal carcinogenesis and advanced colorectal adenocarcinoma

AIM:To determine the expression and clinical significance of transcriptional intermediary factor 1 gamma (TIF1γ),Smad4 and transforming growth factor-beta (TGFβR) across a spectrum representing colorectal cancer (CRC) development.METHODS:Tissue microarrays were prepared from archival paraffin embedded tissue,including 51 colorectal carcinomas,25 tubular adenomas (TA) and 26 HPs,each with matched normal colonic epithelium.Immunohistochemistry was performed using antibodies against TIF1γ,Smad4 and TGFβ RⅡ.The levels of expression were scored semi-quantitatively (score 0-3 or loss and retention for Smad4).RESULTS:Overexpression of TIF1γ was detected in 5/26 (19%) HP;however,it was seen in a significantly higher proportion of neoplasms,15/25 (60%) TAs and 24/51 (47%) CRCs (P<0.05).Normal colonic mucosa,HP,and TAs showed strong Smad4 expression,while its expression was absent in 22/51 (43%) CRCs.Over-expression of TGFβ RⅡ was more commonly seen in neoplasms,13/25 (52%) TAs and 29/51 (57%) CRCs compared to 9/26 (35%) HP (P<0.05).Furthermore,there was a correlation between TIF1γ overexpression and Smad4 loss in CRC (Kendall tau rank correlation value=0.35,P<0.05).The levels of TIF1γ overexpression were significantly higher in stage Ⅲ than in stage Ⅰ and Ⅱ CRC (P<0.05).CONCLUSION:The findings suggest that over-expression of TIF1γ occurs in early stages of colorectal carcinogenesis,is inversely related with Smad4 loss,and may be a prognostic indicator for poor outcome.

Mechanistic basis and clinical relevance of the role of transforming growth factor-β in cancer

Transforming growth factor-β(TGF-β) is a key factor in cancer development and progression. TGF-β can suppress tumorigenesis by inhibiting cell cycle progression and stimulating apoptosis in early stages of cancer progression. However, TGF-β can modulate cancer-related processes, such as cell invasion, distant metastasis, and microenvironment modification that may be used by cancer cells to their advantage in late stages. Corresponding mechanisms include angiogenesis promotion, anti-tumor immunity suppression, and epithelial-to-mesenchymal transition(EMT) induction. The correlation between TGF-β expression and cancer prognosis has also been extensively investigated. Results suggest that TGF-β pathway can be targeted to treat cancer; as such, the feasibility of this treatment is investigated in clinical trials.

Role of TGF-β1 and its Receptors in Breast Carcinogenesis:Evaluation of Gene Expression Patterns and Clinical Implications

OBJECTIVE Transforming growth factor β1 （TGF-β1） is a multifunc- tional cytokine that may play an important role in tumor development and progression. METHODS We evaluated gene expression patterns of TGF-β1 and its receptors [transforming growth factor β type Ⅰ receptor （TβR- Ⅰ ） and transforming growth factor β type Ⅱ receptor （TβR- Ⅱ ）] in tumor tissue from patients with breast cancer or with benign breast diseases （BBD） and adjacent normal tissue from the patients with breast cancer. Included in the study were 527 breast cancer patients and 213 BBD patients who participated in the Shanghai Breast Cancer Study. RESULTS The expression levels of the TGF-β1, TβR- Ⅰ and TβR-Ⅱ genes in breast tissue were quantified using real-time PCR. TIER- Ⅱ expression in cancer tissue was decreased by over 50% as compared to either adjacent normal tissue from the same patients or benign tumor tissue from BBD patients （p〈0.001）. TGF-β1 expression was lower by approximately 20% in cancer tissue compared to adjacent normal tissue （p=0.14） or to benign tumor tissue （p=0.002）. Although TβR-Ⅰ expression was also reduced in cancer tissue compared to adjacent normal tissue, or benign tumor tissue, the magnitude of the reduction was less apparent than that for TβR- Ⅱ. Compared to patients with the lowest tertile value for TβR- Ⅱ, patients with median tertile value for TβR- Ⅱ had more favorable overall survival （HR 0.47, 95% CI 0.27-0.85） and disease-free survival （HR 0.65, 95% CI 0.39-1.06）. No apparent associations, however, were observed between TGF-β1 or TβR- Ⅰ expression and overall or disease-free survival. CONCLUSION The results from this study support the hypothesis that a decreased level of TβR-Ⅱ gene expression, and thus reduced TGF-β1 sensitivity, is related to breast tumor progression.

TRANSFORMING GROWTH FACTOR-β AND FIBROBLAST GROWTH FACTOR INDUCE LENS EPITHELIAL EXPLANT METAPLASIA:IMPLICATIONS FOR THE FORMATION OF SUBCAPSULAR OPACIFICATION

Objective. This study was to investigate the effects of transforming growth factor-β(TGFβ) and fi- broblast growth factor (FGF) in the subcapsular opacification formation of the lens. Methods. Lens epithelial explants from 10-day-old rats were cultured with TGFβ1 or TGFβ2 in the presence of FGF for 5 days, then were examined by light and electron microscopy, and by immunolocal- ization of smooth muscle(α-sm) actin and type I collagen. Results. In TGFβ/FGF-treated explants,extensive proliferation occured, with formation of spindle and star-shaped cells. These cells showed ultrastructure and biochemical features of fibroblast or myofibroblast. Prominent Golgi apparatus and rough endoplaic reticulum were observed in some cells. Intracellular micro- filaments with cytoplasmic dense babies and membrane associated dense bodies, features of smooth muscle cells, were also observed. Some cells showed reactivity to -sin actin antibody. TGFβ/FGF-treated ex- plants were strongly stained with type I collagen antibody. Condusion. In the presence of FGF, TGFβ1 and TGFβ2 induced lens epithelial cell (LEC ) proliferation and transformation into fibroblast or myofibroblast-like cells, with producing of abundant collagen matrix in the explants. The changes are similar to the metaplasia that occurrs in subcapsular opacification of the lens. The findings suggest that TGFβ and FGF plays a role in the pathogenesis of subcapsular opacification of the lens.

TGF-β receptors in mouse ES-5 cells and their differentiated derivatives

By radioreceptor binding studies with iodinated TGF-β1, it has been shown that an undifferentiated ES-5 cell expresses approximately 3270 receptors with a dissociation constant Kd=130pM, but after the induction of differenti-ation by retinoic acid and dBcAMP, the receptor number of a differentiated RA-ES-5 cell was increased about 80% and the Kd was also increased to 370 pM. Furthermore,more direct evidence supporting the expression of TGF-βtype Ⅰand type Ⅱ receptors in both ES-5 and RA-ES-5 cells has come from dot blot hybridization of cellular mRNA with cDNA probes for type Ⅰ and type Ⅱ recep-tors. Meanwhile, mRNA expression level of types Ⅰ and Ⅱreceptors in RA-ES-5 cells were higher than that in ES-5 cells. Down regulation of TGF-β receptors with a signifi-cant decrease in the rate of cell proliferation in both cells, was found by employing a pretreatment with neutralizing antibody to TGF-β1. The possible role of receptors for TGF-β in cen differentiation is discussed here.

Activation of phospholipase D activity in transforming growth factor-beta-induced cell growth inhibition

Cells regulate phospholipase D (PLD) activity in response to numerous extracellular signals. Here, we investigated the involvement of PLD activity in transforming growth factor-β(TGF-β1)-mediated growth inhibition of epithelial cells. TGFβ1 inhibits the growth of MDCK, Mv1Lu, and A-549 cells. In the presence of 0.4 % butanol, TGF-β1 induces an increase in the formation of phosp hat idylbutanol, a unique product catalyzed by PLD. TGF-β1 also induces an increase in phosphatidic acid (PA) level in A-549 and MDCK cells. TGF-β1 induces an increase in the levels of DAG labeled with [~3H]-myristic acid in A-549 and MDCK cells but not in Mv1Lu cells- No increase of DAG was observed in cells prelabeled with [~3H]-arachidonic acid.The data presented suggest that PLD activation is involved in the TGF-β1-induced cell growth inhibition.

Clinical efficacy of Salvia miltiorrhiza in pathological scar treatment:A systematic review and meta-analysis

To systematically assess the clinical efficacy of Salvia miltiorrhiza(SM)in treating pathological scars and provide a reference basis for scar pharmacotherapy,we conducted a comprehensive literature search in both English and Chinese databases from database inception to December 2022.Key search terms included Salvia miltiorhiza,cryptotanshinone,tanshinone IIA,sodium-tanshinol,compound Salvia miltiorrhiza dripping pills,cicatrix,cicatrices,and scar.The inclusion criteria encompassed all clinical randomized controlled studies on the treatment of pathological scars with SM,without regard to blinding or allocation concealment,as well as irrespective of patient nationality,race,or age.Data from the selected literature were subjected to analysis using Rev Man 5.4 software,employing the standard mean difference or weighted mean difference for numerical variables and odds ratios(ORs)for dichotomous variables.Statistical significance was set at P<0.05.Six eligible studies involving a total of 778 patients met the inclusion criteria.The analysis revealed a significant therapeutic efficacy with an OR of 3.83(95%CI:2.65–5.54),indicating a substantially higher therapeutic efficacy rate in SM group compared to the control group.Furthermore,the total effective rate of treatment exhibited an OR of 6.94(95%CI:2.53–19.06),signifying a significantly superior treatment outcome in SM group.Regarding scar scores,no significant improvement was observed in SM group compared to the control group after 3 months of treatment(mean difference[MD]=–0.96,95%CI:–2.29–0.36).However,after 6 months of treatment,the scar score demonstrated a noteworthy improvement in SM group(MD=–1.37,95%CI:–2.44 to–0.30)compared to the control group.In summary,this study affirmed that SM treatment markedly enhanced the therapeutic efficacy and overall treatment efficiency for clinical scar patients,underscoring its positive clinical therapeutic impact on scar patients.

T helper cells in patients with chronic hepatitis B virus infection

OBJECTIVE: To investigate the compositions of Th1/Th2/Th3 cells in chronic hepatitis B virus (HBV)-infected individuals by determining the expression of interleukin-4 (IL-4), inetrferon-gamma (IFN-gamma), and transform growth factor-beta (TGF-beta) in single CD4(+) T cells isolated from peripheral blood mononuclear cells (PBMCs) and the role of polarized Th cell populations in chronic HBV-infection was discussed. METHODS: PBMCs from chronically infected HBV individuals were isolated, stimulated by PMA/Ionomycin/Monensin, and IL-4, IFN-gamma and TGF-beta production by CD4(+) T cells was determined by using fluorescence activated cell sorter (FACS) analysis. RESULTS: The percentage of IFN-gamma-producing T cells, IL-4-producing T cells and TGF-beta-producing T cells ranged from 2.3% - 18.6%, 1.1% - 8.7% and 0.7% - 7.1% respectively in CD4(+) T cells from non-infected individuals. Most of CD4(+) T cells from PBMCs in chronically infected HBV individuals were Th0 cells. The proportion of Th1 cells increased significantly with hepatic inflammatory activity, and in the active period of chronic hepatitis B infection were higher than those in the non-active period (P 0.05), but were higher than that from controls (P

Effect of substance P on gene expression of transforming growth factor β-1 and its receptors in rat's fibroblasts

Objective: To investigate the effect of substance P (SP) on gene expression of transforming growth factor β-1 (TGFβ-1), transforming growth factor receptor-1 (TGFR-1) and transforming growth factor receptor-2 (TGFR-2) in fibroblasts cultured in vitro from rat’s granulation tissues. Methods: The fibroblasts from the granulation tissues in the skeletal muscle of rat’s hind limbs injured by formaldehyde were cultured in vitro. When different concentrations (10 -9-10 -5 mol/L) of SP were added into the culture medium, the changes of gene expression of TGFβ-1, TGFR-1 and TGFR-2 in the cultured fibroblasts were observed with reverse transcription polymerase chain reaction at different intervals (0, 3, 6, 12 and 24 hours after incubation). Results: The gene expression of TGFβ-1, TGFR-1 and TGFR-2 in the fibroblasts cultured from rat’s granulation tissues was up-regulated by SP. The peak level of the mRNA expression was found at 10 -8 mol/L SP and the up-regulation effect was not found at 10 -5 mol/L and 10 -6 mol/L. The peak levels of gene expression of TGFβ-1, TGFR-1 and TGFR-2 in the fibroblasts treated with SP were achieved at 6 and 12 hours, respectively. Conclusions: SP has up-regulation effect on the gene expression of TGFβ-1, TGFR-1 and TGFR-2 in fibroblasts from rat’s granulation tissues in vitro, and the effect is related to different stimulating concentrations of SP. It may be concerned with proliferation and differentiation of fibroblasts and formation of scar tissues during wound healing.

Effect of Moxibustion on Expressions of TGF-β and Smad4 in Colonic Mucosa of Rats with Intestinal Fibrosis in Crohn Disease

Objective： To observe the effect of moxibustion on the expressions of transforming growth factor-β （TGF-β） protein, and Smad4 protein and mRNA in rats with intestinal fibrosis in Crohn disease （CD）, and to discuss the action mechanism of acupuncture-moxibustion in treating intestinal fibrosis in CD from the perspective of TGF-β/Smads signal pathway. Methods： A CD intestinal fibrosis model was developed by using male SD rats of clean grade. The rats were randomized into a normal group, a model group, a mild moxibustion group, an electroacupuncture （EA） group, and a herb-partitioned moxibustion group. The rats in the normal and the model groups did not receive any interventions. Tianshu （ST 25） and Qihai （CV 6） were selected for the rats in the mild moxibustion group, the EA group, and the herb-partitioned moxibustion group. Mild moxibustion, EA, and herb-partitioned moxibustion were given respectively. After treatment, the expressions of TGF-β and Smad4 proteins in rat’s colon were detected by using immunohistochemistry （IHC）; the expression of Smad4 mRNA was determined by using fluorescence quantitative polymerase chain reaction （FQ-PCR）. Results： Compared with the normal group, the expressions of TGF-β protein, Smad4 protein and Smad4 mRNA significantly increased in the model group （P0.01）. Compared with the model group, the expressions of TGF-β protein, Smad4 protein, and Smad4 mRNA dropped markedly （P0.01 or P0.05） after treatments of mild moxibustion, EA, and herb-partitioned moxibustion. Conclusion： The expressions of TGF-β protein, Smad4 protein and Smad4 mRNA significantly increase in CD rats withintestinal fibrosis; while mild moxibustion, EA and herb-partitioned moxibustion at Tianshu （ST 25） and Qihai （CV 6） all can down-regulate the abnormal expressions.