ABSTRACT
Objective To explore the expressions of Smad4 and estrogen receptor (ER) and their interrelation,and the relationship with the clinicopathological features of breast cancer.Methods The immunohistochemical SP method was used to detect the expressions of Smad4 and ER in 50 case of invasive cancer,12 cases of carcinoma in situ and 15 cases of normal breast tissues.The differences in different clinical stages,differentiation degrees and nodal metastases were analyzed.The correlation between Smad4 and ER was explored.Results The positive expression rate of Smad4 in invasive cancer was 52.00％,which lower than that in normal breast tissue (93.33％),with a significant difference (x2 =8.329,P =0.004),positive expression rates of ER were 60.00％ and 40.00％ respectively,with no significant difference (x2 =1.868,P =0.172).The positive expression rates of Smad4 in carcinoma in situ and invasive cancer were 75.00％ and 52.00％ respectively,with no significant difference (x2 =2.082,P =0.149).The positive expression rates of ER were 58.33％ and 60.00％ respectively,with no significant difference (x2 =0.011,P =0.916).The positive expression of Smad4 was related to the TNM stage (x2 =6.392,P =0.011) and the lymph node metastasis (x2 =6.738,P =0.009),but it was not associated with the histologic grade (x2 =0.542,P =0.462).The positive expression of ER was related to the lymph node metastasis (x2 =4.133,P =0.042) and histologic grade (x2 =5.357,P =0.021),but it was not associated with the TNM stage (x2 =1.159,P =0.282).There was positive correlation between Smad4 and ER in breast cancer tissue (r =0.263,P =0.032).Conclusion Smad4 is expressed at lower level in breast cancer than in normal breast tissue.The expressions of Smad4 and ER are related to the different clinicopathological features of breast cancer with positive correlation.
ABSTRACT
The transforming growth factor beta (TGF-beta) is a cytokine that plays crucial roles in the regulation of angiogenesis, immune response, proliferation, migration and apoptosis of cells. In addition, it can inhibit cell progression and stimulate apoptosis in early stages of cancer. TGF-beta is a multifunctional homodimeric protein secreted by various cell lines, which have three different isoforms: TGF-beta1, TGF-beta2 and TGF-beta3. In normal conditions, TGF-beta1 activates some tumor suppressor cell signaling pathways that inhibit proliferation and are involved in cell migration, differentiation and apoptosis. However, in more advanced stages of cancer, when TGF-beta1 is altered, it acts as a promoter of tumorigenesis and may cause: 1) increased TGF-beta1, 2) overexpression of TGF-beta1 receptors (TbetaR), 3) TbetaR mutations, and 4) downregulation of TGF-beta receptor. In oral squamous cell carcinoma, the path is altered especially at the level of transmembrane receptors, with the TbetaR-II and TbetaR-III subtypes being the most affected. However, there is little information on the prognostic role it plays in the various types of cancers. It is important to study the signaling pathways of TGF-beta in order to develop techniques that may help detect their alterations and restore their normal operation. The objective of this review is to describe the alterations of TGF-beta in oral squamous cell carcinoma.
El factor de crecimiento transformante beta (TGF-beta) es una citocina que cumple funciones fundamentales en la regulación de la angiogénesis, respuesta inmune, proliferación, migración y apoptosis celular. Además, puede inhibir la progresión celular y estimular la apoptosis en etapas tempranas del cáncer. El TGF-beta es una proteína homodimérica multifuncional secretada por diversas líneas celulares, que presentan 3 isoformas: TGF-beta1, TGF-beta2 y TGF-beta3. En condiciones normales TGF-beta1 activa a algunas vías de señalización celular supresoras de tumores que inhiben la proliferación, y participan en la migración, diferenciación y apoptosis. Sin embargo, cuando esta se ve alterada, en etapas más avanzadas del cáncer actúa como promotor de la tumorogénesis, pudiendo producir: 1) aumento del TGF-beta1, 2) sobre expresión de los receptores del TGF-beta1 (TbetaR), 3) mutaciones de TbetaR, y 4) falla en la regulación negativa de TbetaR. En el carcinoma oral de células escamosas, la vía se ve alterada especialmente a nivel de sus receptores transmembranales, siendo los subtipos TbetaR-II y TbetaR-III los más afectados. Sin embargo, es escasa la información sobre el rol pronóstico que juega en los diversos tipos de cánceres. Es importante estudiar las vías de señalización de TGF-beta para desarrollar técnicas que detecten sus alteraciones y restauren el funcionamiento del sistema. El objetivo de esta revisión es describir las alteraciones de TGF-beta en carcinoma oral de células escamosas.
Subject(s)
Humans , Carcinoma, Squamous Cell/metabolism , Mouth Neoplasms/metabolism , Transforming Growth Factor beta1/metabolism , /metabolismABSTRACT
Objective To investigate miR-146a-Smad4 expression during ultraviolet A(UVA)-induced photoaging of human skin fibroblasts (HSFs), and to evaluate effects of up-regulation of miR-146a expression on its target gene Smad4 and cell photoaging. Methods HSFs were isolated from the prepuce, and subjected to primary culture and maintained up to 10th passage. Then, the HSFs were classified into 4 groups: blank control group receiving no treatment, UVA group irradiated with 10 J/cm2 UVA, miR-146a group transfected with a lentiviral vector expressing miR-146a, UVA+ miR-146a group transfected with the lentiviral vector expressing miR-146a followed by UVA radiation. Real time PCR was performed to measure miR-146a expression in HSFs in the UVA group on day 0, 3, 7 and 14 after UVA radiation.Fluorescence microscopy was carried out to estimate transfection efficiency on day 7 and 14 in the miR-146a group after transfection, and real time PCR was performed to quantify miR-146a expression in these cells. Methyl thiazolyl tetrazolium (MTT)assay was conducted to evaluate proliferative activity of HSFs, real time PCR to quantify mRNA expressions of photoaging-related genes p53, p21 and p16, and Western blot analysis to measure Smad4 protein expression in these cells. Statistical analysis was carried out by using repeated-measures analysis of variance and factorial design analysis of variance. Results Repeated-measures analysis of variance showed that the expression of miR-146a decreased over time in both the UVA group and blank control group(F = 213.840, P 0.05). Factorial design analysis of variance showed that UVA radiation had an inhibitory effect on the proliferative activity of HSFs (P 0.05). Real time PCR and Western blot analysis both revealed that UVA radiation could increase the expressions of p53, p21 and p16 mRNAs as well as Smad4 protein(all P 0.05). Conclusion The expression of miR-146a is inhibited in UVA-induced photoaged HSFs, and its up-regulation may counteract cell photoaging by suppressing Smad4 expression in, and promoting proliferation of, photoaged HSFs.
ABSTRACT
BACKGROUND:Chinese nourishing kidney herbs can prevent osteoporosis and improve bone metabolism, which has been proved in animal and cel experiments. But there are few reports on the compatibility of Chinese nourishing kidney herbs, and it is difficult to screen the optimal compatibility, as the interaction of active ingredients and drug substance basis are uncertain. OBJECTIVE: To determine the proliferation, differentiation and Smad4 mRNA expression of neonatal Sprague-Dawley rat osteoblasts cultured by Chinese nourishing kidney herbs with different compatibility so as to find out the optimal compatibility of Chinese nourishing kidney herbs. METHODS: Passage 5 osteoblasts were divided into five groups: group A, 1×10-5mol/L icarin; group B, 1×10-5mol/L icarin+1×10-5 mol/L naringin; group C, 1×10-5mol/L icarin+1×10-5 mol/L diosgenin; group D, 1×10-5mol/L icarin+ 1×10-5mol/L catalpol; group E, 10 μL normal saline (control group). There were six wels in each group. RESULTS AND CONCLUSION: Compared with the group E, the proliferative ability of osteoblasts and expression of Smad4 mRNA were increased in the groups B and C; until the 72nd hour, the proliferative ability of osteoblasts in the group B reached the peak. At 48 hours of culture, the activity of alkaline phosphatase in groups B and C was higher than that in group E; at 72 hours of culture, the activity of alkaline phosphatase in groups B and D was higher than that in group E. These findings indicate that the compatibility of Chinese nourishing kidney herbs can influence the activity of osteoblasts, and icarin+naringin has the strongest effect.
ABSTRACT
Objective To study the effect of NLK on TGFβsignaling pathway and explore the molecular mechanism .Methods Protein stability assay was used to determine the influence of NLK on the degradation of Smad 4 .In vivo ubiquitination assay was applied to detect the effect of NLK on the ubiquitination of Smad4 .Luciferase reporter gene assay was used to detect the effect of NLK on CAGA‐luc and 3TP‐luc ,the responsive reporter of TGFβ signaling pathway .Real time PCR was applied to examine the effect of NLK on the expression of p21 and PAI‐1 ,the target genes of TGFβsignaling pathway .Results In HEK293T cell ,over ex‐pression of NLK promotes the degradation and ubiquitination of Smad4 .In HEK293T cells ,Ectopic expression of NLK inhibits the activity of CAGA luc and 3TP luc stimulated by TGFβ.In HepG2 cells ,over expression of NLK inhibits the expression of p21 and PAI 1 ,the target genes of TGFβsignaling pathway .Conclusion NLK promotes the ubiquitination and degradation of Smad4 ,conse‐quently inhibits TGFβsignaling pathway .
ABSTRACT
Objective To investigate the anti-tumor effect and mechanism of curcumin in pancreatic cancer (PC). Methods Smad4 and Jab1 expressions were detected by immunohistochemistry in tumor tissues and pericarcinomatous tis?sue from 35 PC cases, and the correlation of Smad4 and Jab1 were analyzed based on the percentage of positive staining in?tissues from 21 random selected PC cases. The effect of curcumin on expressions of tumor suppressors p53, Smad4 and cell cycle inhibitor p27 were examined by Western Blotting after human pancreatic cancer cell line PANC-1 were divided into PANC-1 control group (no treatments were given) and PANC-1 curcumin group (treated with cell culture medium containing 10μmol/L curcumin). The effect of curcumin on expressions of combination of β-TrCP1 and Smad4 was examined by Co-Immunoprecipitation after human embryonic kidney cell line 293T were divided into 293T control group (no treatments were given), 293T curcumin group (treated with cell culture medium containing 10μmol/L curcumin) and 293T Jab1 group (trans?fected by HA-Jab1 plasmid). Results Compared with expressions in pericarcinomatous tissues, Smad4 was down regulated while the expression of Jab1 was upregulated in PC tissues (P<0.01), and the expression of Smad4 was negatively correlated with the expression of Jab1 (n=21, r=-0.71, P=0.007). After treated with curcumin, the protein expression of p53, Smad4 and p27 was increased in PANC1 cell, and the protein expression of the combination ofβ-TrCP1 and Smad4 was decreased in 293T cell (P<0.05). After transfected by HA-Jab1 plasmid, the protein expression of the combination ofβ-TrCP1 and Smad4 was increased in 293T cell (P<0.05). Conclusion Curcumin may have suppression effect of PC through increasing the protein expression of p53, Smad4 and p27, and the mechanism of Smad4 upregulation may be related with the inhibition of Smad4 ubiquitination process, while Jab1 may be also involved in Smad4 degradation through ubiquitination.
ABSTRACT
Objective To investigate the expression and significance of Smad4 and Smad7 in newborn rats with hyperoxia-induced chronic lung disease(CLD).Methods Sixty-four newborn Wistar rats 12 h after birth were divided into high-oxygen group (n =32) and air group (n =32,control group) by random number table method.The high-oxygen group was placed in the oxygen glass tank with continuous infusion of oxygen.And 1,3,7,14 d after experiment,tracheal separated,the chest opened to expose heart and lung,slices were Masson staining,undergo dynamic observation of the pulmonary pathological changes under light microscope.Lung fibrosis score was carried out to determine the degree of pulmonary fibrosis,and immunohistochemical technique was used to detect Smad4 and Smad7 protein expression in lung tissue.The expression levels of Smad4 and Smad7 protein in lung tissue were detected with Western blot.Results Compared with the air group,there was statistically significant difference in pulmonary fibrosis score on day 7 (2.67 ± 0.21 vs 0.58 ± 0.17) and day 14 (4.48 ± 0.24 vs 0.63 ± 0.13) in high-oxygen group (P ＜ 0.05) ; Smad4 and Smad7 was main in visible lung epithelial cells and interstitial fibroblasts.Smad4 expression in the high-oxygen group gradually enhanced,compared with the air group (P ＜ 0.05) on day 7 (122.35 ± 10.3 vs 140.08 ±7.77) and day 14(129.7 ± 7.33 vs 144.99 ± 6.49).Smad7 expression in the high-oxygen group first increased and then decreased,expression in the high-oxygen group increased on day 7 (122.35 ± 10.29 vs 130.56 ±9.8),and compared decreased with the air group(P ＜0.05) on day 14(132.16 ±4.38 vs 126.22 ±6.49).Conclusion The newborn rat exposed hyperoxia,the up-regulation of Smad4 protein expression and the down-regulation of Smad7 protein expression are imposible closely related to the happen and development of CLD pulmonary fibrosis.
ABSTRACT
INTRODUCTION: A cleft palate is a common birth defect in humans with an incidence of 1/500 to 1/1,000 births. It appears to be caused by multiple genetic and environmental factors during palatogenesis. Many molecules are involved in palate formation but the biological mechanisms underlying the normal palate formation and cleft palate are unclear. Accumulating evidence suggests that transforming growth factor beta/bone morphogenetic proteins (TGF-beta/BMP) family members mediate the epithelial-mesenchymal interactions during palate formation. However, their roles in palatal morphogenesis are not completely understood. MATERIALS AND METHODS: To understand the roles of TGF-beta/BMP signaling in vivo during palatogenesis, mice with a palatal mesenchyme-specific deletion of Smad4, a key intracellular mediator of TGF-beta/BMP signaling, were generated and analyzed using the Osr2Ires-Cre mice. RESULTS: The mutant mice were alive at the time of birth with open eyelids and complete cleft palate but died within 24 hours after birth. In skeletal preparation, the horizontal processes of the palatine bones in mutants were not formed and resulted in a complete cleft palate. At E13.5, the palatal shelves of the mutants were growing as normally as those of theirwild type littermates. However, the palatal shelves of the mutants were not elevated at E14.5 in contrast to the elevated palatal shelves of the wild type mice. At E15.5, the palatal shelves of the mutants were elevated over the tongue but did not come in contact with each other, resulting in a cleft palate. CONCLUSION: These results suggest that mesenchymal Smad4 mediated signaling is essential for the growth of palatal processes and suggests that TGF-beta/BMP family members are essential regulators during palate development.
Subject(s)
Animals , Humans , Mice , Cleft Palate , Congenital Abnormalities , Eyelids , Incidence , Morphogenesis , Palate , Parturition , Proteins , Smad4 Protein , Tongue , Transforming Growth FactorsABSTRACT
BACKGROUND: Minichromosome maintenance protein 7 (MCM 7) performs a direct role in the initiation of DNA replication, which suggests that it may prove useful as a marker of cell proliferation. Smad 4 is a tumor suppressor gene that mediates the transforming growth factor beta pathway. The principal objective of this study was to characterize the expression of MCM 7 and Smad 4 and to analyze their relationship to clinicopathological parameters in patients with esophageal squamous cell carcinoma. METHODS: Expression levels of MCM 7 and Smad 4 were evaluated via immunohistochemistry on formalin-fixed and paraffin-embedded tissues from 67 cases of esophageal squamous cell carcinoma. RESULTS: High levels of MCM 7 expression were detected in 53 cases (74.6%), and were associated with higher T stages (p = 0.030). Kaplan-Meier survival curves demonstrated that patients with higher levels of MCM 7 expression had poorer prognoses, although this association was not significant (p = 0.086). Loss of Smad 4 expression was noted in 18 cases (23.4%), and was not associated with clinicopathological characteristics, including MCM 7 expression, or prognosis. CONCLUSIONS: MCM 7 expression is associated with the invasiveness of esophageal squamous cell carcinoma. Altered expression of Smad 4 does not appear to have pathobiological significance in esophageal carcinoma.
Subject(s)
Humans , Carcinoma, Squamous Cell , Cell Proliferation , DNA Replication , Esophageal Neoplasms , Esophagus , Genes, Tumor Suppressor , Immunohistochemistry , Kaplan-Meier Estimate , Prognosis , Transforming Growth Factor betaABSTRACT
Objective To investigate the expression and significance of Smad4 in peripheral hepatocytes of lesions in mice with hepatic alveolar echinococcosis.Methods Eight mice in the test group were inoculated with alveolar echinococcosis and 8 mice in the control group were injected with normal saline.The expression of Smad4 protein in the hepatic tissue of the mice was detected by immunohistochemistry method,and the data were analyzed by chi-square test.The effect of alveolar echinocoeeosis on the expression of Smad4 protein was investigated.Results Smad4 was detected in cell nuclei and partly in the cytoplasm.Six months after the establishment of the mice model for alveolar echinococeosis,the expression of Smad4 in the hepatic tissue in the test group was significantly higher than in the control group(x2=19.869,P＜0.05).The number of Smad4 with positive expression in the hepatocytes in the test group was slightly higher than in the control group,and the expression intensity of Smad4 in the test group was greater than in the control group(x2=58.3 10,P＜0.05).Conclusions The increase of the expression of Smad4 protein in the periphery hepatocytes and tissues of hepatic alveolar echinococcosis lesions may play a role in hepatic cirrhosis and immune evasion.
ABSTRACT
Objective To investigate the expression of transforming growth factor β1,transforming growth factor beta receptor(TBR)Ⅰ,TβR Ⅱ,Smad4 and C-Jun in rats with nonalcoholic fatty liver disease(NAFLD)and to find out the mechanisms of liver fibrosis in patients with NAFLD.Methods A total of 18 male SD rats were randomly divided into normal control group(n=9)and model group(n=9).The rats in control group were fed with normal diet,and those in model group were fed with fat-rich diet(consisted of 10%lard oil+2%cholesterol).An rats were sacrificed at the 20th week.The levels of TGFβ1,TβR Ⅰ and TβR Ⅱ mRNA were examined by RT-PCR.The expressions of TGFβ1 and Smad4 in liver tissue were detected by immunohistochemistry.The expression of C-Jun protein was detected by Western blotting.Results The NAFLD model was successfully established.The immunohistochemistry examination revealed that TGFβ1 and Smad4 were expressed weekly in control group,but strongly expressed in model group.RT-PCR showed that A values of TGFβ1,TβR Ⅰ and TβR Ⅱ mRNA were 0.46±0.12,5.z4±2.70 and 3.35±1.95,respectively,in model group,which were higher than those in control group(0.21±0.09,1.36±0.77 and 0.52±0.19,all P values＜0.01).The Western blotting results demonstrated that the expression of C-Jun protein in model group(0.93±0.41)was higher than that in control group (0.32±0.25,P=0.001).Conclusion TGFβ1/Smad4 signal pathway might be involved in the development of hepatic fibrosis in NAFLD.Blocking TGFβ1/Smad4 signal pathway will be helpful in treatment of NAFLD.
ABSTRACT
Objective To determine the correlation between Smad4 mRNA,protein expression,abnormal methylation of CpG islands of Smad4 and colorectal cancer.Methods The expression of Smad4 in 43 cases of colorectal cancer and its adjacent tissue.30 cases of colorectal adenoma and 12 normal colon mucosa were analyzed by RT-PCR,sequencing,semi-quantitative RT-PCR,methylation-specific PCR(MSP)and immunohistochemistry.Results The mRNA expression of Smad4 was detected in 25 cases of colorectal cancer(58.14%),with expression levels at 0.73±0.25,which was significantly lower than those in corresponding adjacent tissues(88.37%,0.95±0.29),eolorectal adenoma(90.63%,1.01±0.37)and normal mucosa(100.00%,1.18 ±0.33)(P＜0.05).The positive rates of Smad4 gone promoter methylation in colorectal caner(60.53%)was significantly higher than that in other tissues(27.03%,25.00%and 16.67%,respectively,P＜0.05).The expression of Smad4 protein was significantly lower in colorectal cancer(44.19%)than that in other tissues(81.40%,87.50%,91.67%,respectively,P＜0.05),which was correlated with tumor invasion and lymph node metastasis.Conclusion Down-regulation of Smad4 expression may be associated with the development,the biological behavior and prognosis of colorectal cancer,and it can be an important biological marker in evaluation of disease progression.
ABSTRACT
Objective: To investigate the relationship between the expressions of the transforming growth factor beta 1 (TGF-β1), Smad4, and TβR II in the different stages of gallbladder carcinoma and discuss their roles in the development of gallbladder carcinoma. Methods: Immunohistochemical SP method was used to examine the expression of TGF-β1, Smad4, and TβR II in 30 cases of gallbladder carcinoma, 11 cases of gallbladder adenoma, and 30 cases of cholecystitis. The relationship between the expressions of TGF-β1, Smad4 and TβR II and the clinicopathological features of gallbladder carcinoma were analyzed. Results: The positive rates of TGF-β1, Smad4 and TβR II was 73.3%, 20%, and 16.7% in 30 cases of gallbladder carcinoma; 90.9%, 63.7%, and 54.5% in 11 cases of gallbladder adenoma; 96.7%, 93.3%, and 90% in 30 cases of cholecystitis. The expression of TGF-β1 was significantly lower in gallbladder carcinoma than that in cholecystitis (P<0.05). The expressions of Smad4 and TPR H were significantly lower in gallbladder carcinoma than that in cholecystitis (P<0.05). Gallbladder carcinoma at stages I and II had lower TGF-β1 expression but higher Smad4 and TβR II expression than those at stage III and V (P<0.05). TGF-β1 expression was significantly higher (94.1%) in gallbladder carcinoma with metastasis than that without metastasis (P<0.05). Conclusion: The decreased expression of TGF-β1 in gallbladder carcinoma may be related with the cell malignant transformation and uncontrolled growth. Over-expression of TGF-β could not inhibit the proliferation of gallbladder carcinoma which may be due to the decreased expression of Smad4 and TβR II. The high expression of TGF-β1 is related with the progression, invasion, and metastasis of gallbladder carcinoma.