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PubMed2005

Expression of Smad4 during rat ovarian development.

OBJECTIVE: To clarify the signal transduction pathway of transforming growth factor-betasuperfamily (TGF-betas) in the regulation of follicle growth by investigating the expressions of Smad4 protein and mRNA in rat ovaries in different developmental stages. METHODS: Rat ovaries of different developmental stages were obtained to determine the expression of Smad4 protein by immunohistochemistry and image analysis system, with Smad4 mRNA measured by semi-quantitative RT-PCR. Specific primers of Smad4 and GAPDH (internal control) were used for amplification by RT-PCR, and the ratios of their integrated optical densities were calculated to estimate the relative quantity of Smad4 mRNA expression. RESULTS: Smad4 protein was widely expressed in the ovary, mainly in the follicles, and the location and intensity of Smad4 expression varied with the degree of maturation of the ovary. In the early developmental stages, Smad4 protein expressed mainly in the primordial and preantral follicles, but little in the stromal cells, and its expression intensity in the stroma increased gradually in the course of ovarian maturation. After sexual maturity, Smad4 expression intensity varied only insignificantly among the granulosa cells, theca cells and stromal cells of the antral and mature follicles (P>0.05). The staining intensity of Smad4 in the follicles also underwent changes in relation with their development, being less intense in the oocytes of the antral and matured follicles as compared to the preantral follicles (P<0.05 and P<0.01, respectively) but markedly greater in the theca cells of the antral and matured follicles than in the preantral follicles (P<0.01). No significant difference in Smad4 expression was found in the granulosa cells of different developmental stages (P>0.05). RT-PCR demonstrated that Smad4 mRNA was expressed in all the developmental stages of the rat ovary; and from the 3rd week on, the integrated optical density of Smad4 and GAPDH was significantly higher than that in 1-day-old neonatal rats. CONCLUSION: The expression patterns of Smad4 protein and mRNA in rat ovary in the course of its development indicate that Smad signal transduction may play a role in the folliculogenesis.

Animals↗
PubMed2002

Proteolytic degradation of Smad4 in extracts of AML blasts.

Loss of transforming growth factor (TGF) beta signaling has been implicated in malignant transformation of various tissues. To investigate a potential role of Smad4 in acute myeloid leukemia (AML), the expression of Smad4 was determined in blast cells from AML patients. Western analysis of nuclear extracts of nine AML samples indicated the absence of Smad4 protein in two cases. Smad4 RT-PCR analysis of these cases indicated normal Smad4 mRNA expression, and sequencing of one of these cases revealed no mutations as compared to wild type Smad4. Next, it was investigated whether Smad4 protein from these AML cases was subject to proteolytic degradation by incubating cell extracts of these Smad4-negative AML cells with extracts from COS-7 cells in which a tagged Smad4 was overexpressed. Inhibitor studies indicated that the extracts of AML blasts lacking Smad4 possessed a serine-dependent proteolytic activity, capable of degrading Smad4. Transfection studies using an SBE containing reporter construct as well as RT-PCR analysis of endogenous TGFbeta1 responsive genes indicated that the AML blasts were still able to respond to TGFbeta1, despite the observed degradation of Smad4. It was, therefore, concluded that the degradation of Smad4 was possibly AML subtype-dependent, in vitro phenomenon, occurring during the preparation of nuclear and cellular extracts despite the addition of a protease inhibitor cocktail. The results indicate that care should be taken when interpreting data obtained from protein expression studies using AML blast cells.

Acute Disease↗
PubMed2006

[Expression of Smad4 in leukemia cells].

Loss of transforming growth factor (TGF)-beta signaling has been implicated in malignant transformation of various tissues. Smad4 plays a central role in the signal transduction of TGF-beta. Deletion or mutation of Smad4 has been described in a number of cancers. This study was aimed to investigate a potential role of Smad4 in leukemia including its expression and location in blast cells. The mononuclear cells were separated from bone marrow of leukemia patients. The samples, blast cells of which were more than 90% in mononuclear cells, were selected. The expression and location of Smad4 protein were analyzed by immunohistochemistry methods. The results showed that the Smad4 protein located mainly in nucleus, part of this protein located in cytoplasma, the expressions of Smad4 were not detected in 6 out of 9 ALL patients, in 7 out of 24 AML patients and in 1 out of 2 CML patients; these leukemia patients, in whose cells the expression of Smad4 was not detected, included one L1 and one L3, four L2, one M0, one M1, two M2a, one M3a, one M4b, one M6 and one CML. In conclusion, the Smad4 protein was mainly in nucleus, the deletion or functional change of Smad4 may related with the pathogenesis of human AML.

Humans↗
PubMed2007

Inactivation of SMAD4 tumor suppressor gene during gastric carcinoma progression.

PURPOSE: Mothers against decapentaplegic homologue 4 (SMAD4) is a tumor suppressor gene associated with gastrointestinal carcinogenesis. The aim of the present study is to more precisely characterize its role in the development and progression of human gastric carcinoma. EXPERIMENTAL DESIGN: The expression of SMAD4 was investigated in 283 gastric adenocarcinomas and related lesions, as well as in 9 gastric carcinoma cell lines. We also analyzed the methylation status of SMAD4 gene by using methylation-specific PCR, examined loss of heterozygosity (LOH) of this gene locus by using a vicinal marker, and detected exon mutation of SMAD4 through exon-by-exon amplification. Moreover, we assessed whether MG132, a proteasome inhibitor, affected the SMAD4 protein level. RESULTS: We found loss of SMAD4 protein expression in the cytoplasm (36 of 114, 32%) and in the nucleus (46 of 114, 40%) of gastric cancer cells. The loss of nuclear SMAD4 expression in primary tumors correlated significantly with poor survival, and was an independent prognostic marker in multivariate analysis. We also found a substantial decrease in SMAD4 expression at both the RNA and protein level in several human gastric carcinoma cell lines. In addition, we found that LOH (20 of 70, 29%) and promoter hypermethylation (4 of 73, 5%) were associated with the loss of SMAD4 expression. SMAD4 protein levels were also affected in certain gastric carcinoma cell lines following incubation with MG132. CONCLUSION: Taken together, our results indicate that the loss of SMAD4, especially loss of nuclear SMAD4 expression, is involved in gastric cancer progression. The loss of SMAD4 in gastric carcinomas was due to several mechanisms, including LOH, hypermethylation, and proteasome degradation.

Aged↗
PubMed2002

Decreased Smad4 expression in the transforming growth factor-beta signaling pathway during progression of esophageal squamous cell carcinoma.

BACKGROUND: Transforming growth factor-beta (TGF-beta) has antiproliferative effects in various cells, and inactivation of the TGF-beta signaling pathway contributes to tumor progression or development. Smad4, a tumor suppressor gene, is a central mediator in the signaling pathways of the TGF-beta superfamily. This study was undertaken to clarify the correlation between Smad4 expression and the clinicopathologic characteristics of patients with esophageal squamous cell carcinoma (SCC). The authors also investigated the expression of components of the TGF-beta signaling pathway in seven established cell lines derived from esophageal SCC. METHODS: Immunohistochemistry for Smad4 using monoclonal anti-Smad4 antibody was performed on surgical specimens obtained from 80 patients with esophageal SCC. In seven cell lines, the authors examined the expression of components of the TGF-beta signaling pathway using Western and Northern blot analyses. RESULTS: There was a significant inverse correlation between Smad4 expression and both depth of invasion (P = 0.0008) and pathologic stage (P = 0.0079). The expression of Smad4 proteins could be detected in five of seven cell lines. The expression of TGF-beta type II receptor protein was decreased in two of seven cell lines, and the expression of both Smad2 and Smad3 proteins was decreased in only one cell line. The level of expression of Smad4 mRNA did not differ dramatically between cell lines and was not correlated with the quantity of Smad4 protein. CONCLUSIONS: In this study, the expression of Smad4 protein appeared to be correlated with the depth of invasion of esophageal SCC. The loss of Smad4 expression was not regulated at the level of transcription.

DNA-Binding Proteins↗
PubMed2002

Transforming growth factor beta 1 induces proliferation in colon carcinoma cells by Ras-dependent, smad-independent down-regulation of p21cip1.

Transforming growth factor beta1 (TGFbeta1) can act as a tumor suppressor or a tumor promoter depending on the characteristics of the malignant cell. We recently demonstrated that colon carcinoma cells transfected with oncogenic cellular K-rasV12, but not oncogenic cellular H-rasV12, switched from TGFbeta1-insensitive to TGFbeta1-growth-stimulated and also became more invasive (Yan, Z., Deng, X., and Friedman, E. (2001) J. Biol. Chem. 276, 1555-1563). We now demonstrate that TGFbeta1 growth stimulation of colon carcinoma cells is Ras-dependent and smad-independent. In U9 colon carcinoma cells, which are responsive to TGFbeta1 by growth stimulation, a truncating mutation at Gln-311 was found in the smad4 gene. Very little smad4 protein was detected in these cells. Loss of smad4 protein was confirmed by functional studies. In U9 cells co-transfected wild-type smad4, but not mutant smad4, mediated response of the 3TP-lux and pSBE promoter reporter constructs to TGFbeta1. Proliferation initiated by TGFbeta1 in U9 cells required Ras-mediated down-regulation of p21cip1 protein. Less p21cip1 was associated with cdk2 small middle dotcyclin complexes in TGFbeta1-treated U9 cells, and the cdk2 complexes had increased kinase activity. Elevation of p21cip1 levels diminished proliferative response to TGFbeta1. U9 cells expressing DN-N17ras neither proliferated in response to TGFbeta1 nor down-regulated the cdk inhibitor p21cip1, and TGFbeta1 activation of 3TP-lux in U9 cells was inhibited by DN-N17ras in a dose-dependent manner. TGFbeta1 also decreased p21cip1 levels and stimulated proliferation in SW480 cells, which express mutant K-Ras but no smad4 protein. TGFbeta1 did not activate or inhibit the p21cip1 promoter construct in U9 cells even in the presence of co-transfected smad4, or alter p21cip1 mRNA levels. Thus the decrease in p21cip1 levels was mediated by a TGFbeta-initiated Ras-dependent, but smad-independent post-transcriptional mechanism.

Blotting, Northern↗
PubMed2005

Effect of deleted pancreatic cancer locus 4 gene transfection on biological behaviors of human colorectal carcinoma cells.

AIM: To investigate the effect of deleted pancreatic cancer locus 4 (DPC4) gene transfection on biological behaviors of human colorectal carcinoma cells and the role of DPC4 gene in colorectal carcinogenesis. METHODS: PcDNA3.1-DPC4 plasmid was re-constructed by gene-recombination technology. SW620 cells, a human colorectal carcinoma cell line, were transfected with PcDNA3.1-DPC4 plasmid using lipofectamine transfecting technique. Transfected cells were selected with G418. Expression of Smad4 protein was detected in cells transfected with DPC4 gene by immunohistochemistry and Western blot. Biological characterristics of transfected cells were evaluated by population-doubling time and cloning efficiency. Alterations of percentage of S phage cells (S%) and apoptosis rate were determined by flow- cytometry. RESULTS: PcDNA3.1-DPC4 plasmid was constructed successfully. SW620 cells transfected with PcDNA3.1-DPC4 plasmid (DPC4+-SW620 cells) showed a strong intracellular expression of Smad4 protein, and the positive signal was localized in cytoplasm and nuclei, mainly in cytoplasm, where the expressions of Smad4 protein in SW620 cells transfected with PcDNA3.1 plasmid (PcDNA3.1-SW620 cells) and non-transfected SW620 cells (SW620 cells) were weaker than those in DPC4+-SW620 cells. The population- doubling time in DPC4+-SW620 cells (116 h) was significantly longer than that in SW620 cells (31 h) and PcDNA3.1-Sw620 cells (29 h) (P<0.01). The cloning efficiencies of DPC4+-SW620 cells (12%) were markedly lower than those of SW620 cells (69%) and PcDNA3.1-Sw620 cells (67%) (P<0.01). Compared with SW620 cells and PcDNA3.1-Sw620 cells, the G0-G1% of DPC4+-SW620 cells was obviously higher and the S% was markedly lower (P<0.05). Apoptosis rate of DPC4+-SW620 cells was significantly higher than that of SW620 cells and PcDNA3.1-SW620 cells. CONCLUSION: PcDNA3.1-DPC4 plasmid can be successfully re-constructed and stably transfected into human SW620 cells, thereby the cells can steadily express Smad4. DPC4 protein may regulate proliferation of colorectal carcinoma cells by inhibiting cell growth and inducing cell apoptosis.

Apoptosis↗
PubMed2006

Potentiation of Smad-mediated transcriptional activation by the RNA-binding protein RBPMS.

Smad2, Smad3 and Smad4 proteins are considered to be key mediators of transforming growth factor-beta (TGF-beta) signaling. However, the identities of the Smad partners mediating TGF-beta signaling are not fully understood. Here, we show that RNA-binding protein with multiple splicing (RBPMS), a member of the RNA-binding protein family, physically interacts with Smad2, Smad3 and Smad4 both in vitro and in vivo. The presence of TGF-beta increases the binding of RBPMS with these Smad proteins. Consistent with the binding results, overexpression of RBPMS enhances Smad-dependent transcriptional activity in a TGF-beta-dependent manner, whereas knockdown of RBPMS decreases this activity. RBPMS interacts with TGF-beta receptor type I (TbetaR-I), increases phosphorylation of C-terminal SSXS regions in Smad2 and Smad3, and promotes the nuclear accumulation of the Smad proteins. Moreover, RBPMS fails to enhance the transcriptional activity of Smad2 and Smad3 that lack the C-terminal phosphorylation sites. Our data provide the first evidence for an RNA-binding protein playing a role in regulation of Smad-mediated transcriptional activity and suggest that RBPMS stimulates Smad-mediated transactivation possibly through enhanced phosphorylation of Smad2 and Smad3 at the C-terminus and promotion of the nuclear accumulation of the Smad proteins.

Animals↗
PubMed2005

SMAD4 levels and response to 5-fluorouracil in colorectal cancer.

We have recently reported that low tumor levels of SMAD4, a key mediator of transforming growth factor-beta superfamily signaling, can predict the probability of recurrence in patients with Dukes C colorectal cancer who had surgery as the only form of treatment. However, standard treatment for Dukes C colorectal cancer patients currently involves the administration of 5-fluorouracil (5-FU)-based adjuvant chemotherapy after surgery. Approximately 30% to 40% of these patients present with recurrence and die within 5 years, and there is great need for markers capable of predicting poor prognosis after the combined surgery/adjuvant treatment. In this study, we evaluate the prognostic value of SMAD4 in patients treated with surgery and 5-FU-based adjuvant therapy. We used immunohistochemistry and quantitative real-time reverse transcription-PCR to measure the levels of SMAD4 protein and mRNA expression in the primary tumors and a number of lymph node metastases from a series of 75 Dukes C colorectal cancer patients with at least 6 years of follow-up. Patients with tumors expressing low levels of SMAD4 protein or mRNA showed significantly shorted disease-free and overall survival than patients with high tumor levels of SMAD4. The median survival of patients with low SMAD4 protein or mRNA tumor levels was 1.4 and 1.2 years, respectively, whereas patients with high SMAD4 tumor level had a median survival of >9.3 years. In addition, the protein and mRNA levels of SMAD4 in lymph node metastases was significantly lower than in primary tumors (P = 0.006). In contrast, allelic imbalance in chromosome 18q21 was of no prognostic significance in these patients. In conclusion, low SMAD4 tumor levels identified a subset of patients with poor prognosis following surgery and 5-FU-based adjuvant therapy; therefore, these patients could be good candidates to receive combined treatment with additional chemotherapeutic agents such as CPT-11 and/or oxaliplatin.

Allelic Imbalance↗
PubMed2004

[Effects of TGF beta 1 on the cell cycle of irradiated human lung fibroblasts and its signal transduction mechanism].

AIM: To investigate the effects of TGF beta 1 on the proliferative activity and cell cycle of normal and gamma ray-irradiated human lung fibroblasts (HLFs). METHODS: HLFs were cultured and irradiated with 3Gy gamma rays and then stimulated by TGF beta 1 at 0.01, 0.1, 1 and 10 microg/L. The proliferative activity and cell cycle were detected by MTT colorimetry and FACS. Furthermore, the expressions of signal proteins Smad3/4 in HLFs were observed by immunohistochemical staining and FACS. RESULTS: TGF beta 1 of 10 microg/L could promote the proliferation of HLFs, with the number of cells in S phase increased. It also could enhance the expression of Smad3 protein and promote the nuclear translocation of Smad4 protein. CONCLUSION: A given dose of TGF beta 1 can promote the proliferation of gamma rays-irradiated HLFs by regulating their cell cycle in which the activation of Smad pathway may play a role.

Cell Cycle↗
PubMed2001

FLRG, an activin-binding protein, is a new target of TGFbeta transcription activation through Smad proteins.

The FLRG gene encodes a secreted glycoprotein that binds to activin and is highly homologous to follistatin, an activin ligand. We cloned the promoter region of the human FLRG gene, and defined the minimal region necessary for transcription activation in a reporter-system assay. We showed that the fragment between positions -130 and +6, which consists of multiple consensus Sp1-binding sites, is required for the constitutive expression of the FLRG gene. We demonstrate here that FLRG mRNA expression is rapidly induced by TGFbeta or by transfection with Smad protein expression vectors in human HepG2 cells. We investigated the transcription-regulation mechanism of FLRG expression in HepG2 cells following treatment with TGFbeta. By deletion and point-mutation analysis of the FLRG promoter, we identified a Smad-binding element involved in the TGFbeta-inducible expression of the FLRG gene. Moreover, transactivation of the FLRG promoter by TGFbeta was compromised by dominant-negative mutants of Smad3 and Smad4 proteins. In addition, gel electrophoresis mobility-shift assays demonstrated the specific interaction of Smad3 and Smad4 proteins with the Smad-binding element consensus motif found in the FLRG promoter. Taken together, our data imply that Smad proteins participate in the regulation of expression of FLRG, a new target of TGFbeta transcription activation.

Activins↗
PubMed2002

SMIF, a Smad4-interacting protein that functions as a co-activator in TGFbeta signalling.

Proteins of the transforming growth factor beta(TGFbeta) superfamily regulate diverse cellular responses, including cell growth and differentiation. After TGFbeta stimulation, receptor-associated Smads are phosphorylated and form a complex with the common mediator Smad4. Here, we report the cloning of SMIF, a ubiquitously expressed, Smad4-interacting transcriptional co-activator. SMIF forms a TGFbeta/bone morphogenetic protein 4 (BMP4)-inducible complex with Smad4, but not with others Smads, and translocates to the nucleus in a TGFbeta/BMP4-inducible and Smad4-dependent manner. SMIF possesses strong intrinsic TGFbeta-inducible transcriptional activity, which is dependent on Smad4 in mammalian cells and requires p300/CBP. A point mutation in Smad4 abolished binding to SMIF and impaired its activity in transcriptional assays. Overexpression of wild-type SMIF enhanced expression of TGFbeta/BMP regulated genes, whereas a dominant-negative SMIF mutant suppressed expression. Furthermore, dominant-negative SMIF is able to block TGFbeta-induced growth inhibition. In a knockdown approach with morpholino-antisense oligonucleotides targeting zebrafish SMIF, severe but distinct phenotypic defects were observed in zebrafish embryos. Thus, we propose that SMIF is a crucial activator of TGFbeta signalling.

Active Transport, Cell Nucleus↗
PubMed1999

G1 cell cycle arrest and apoptosis induction by nuclear Smad4/Dpc4: phenotypes reversed by a tumorigenic mutation.

The tumor suppressor Smad4/Dpc4 is a transcription activator that binds specific DNA sequences and whose nuclear localization is induced after exposure to type beta transforming growth factor-like cytokines. We explored an inducible system in which Smad4 protein is activated by translocation to the nucleus when cell lines that stably express wild-type or mutant Smad4 proteins fused to a murine estrogen receptor domain are treated with 4-hydroxytamoxifen. This induced Smad4-mediated transcriptional activation and a decrease in growth rate, attributable to a cell cycle arrest at the G1 phase and an induction of apoptosis. A tumor-derived mutation (Arg-100 --> Thr) affecting a residue critical for DNA-binding demonstrated an "oncogenic" phenotype, having decreases in both the G1 fraction and apoptosis and, consequently, an augmentation of population growth. This model should be useful in the exploration and control of components that lie further downstream in the Smad4 tumor-suppressor pathway.

Amino Acid Substitution↗
PubMed2002

Smad4 overexpression in hepatocellular carcinoma is strongly associated with transforming growth factor beta II receptor immunolabeling.

In the normal liver, the transforming growth factor beta (TGF-beta) signaling pathway plays an important role in inhibiting hepatocyte growth. This effect is mediated through Smad4 (or Dpc4), a tumor-suppressor gene that affects gene transcription and controls cell growth. A loss of Smad4 is associated with carcinoma in a number of other organs, including the pancreas and colon. Despite these facts, several recent studies using cDNA microarrays have surprisingly shown overexpression of Smad4 in hepatocellular carcinoma (HCC). Because Smad4 plays a central role in the TGF-beta signaling pathway, we hypothesized that activation of the TGF-beta signaling pathway may explain Smad4 overexpression. To investigate this, 21 surgically resected HCCs were immunostained with antibodies to Smad4 and TGF-beta receptor II. Tumor and normal liver tissues were stained in all cases, and expression in the tumor was scored in comparison to the nonneoplastic liver. Thirteen hepatic adenomas were also immunostained as a control group. The average age at resection was 58 +/- 16 years for the 17 men and 4 women with HCC. TGF-beta receptor II was weakly expressed in the hepatocyte cytoplasm of all normal livers and was overexpressed in 10 of 21 HCCs. Of these 10 HCCs increased Smad4 immunolabeling was also present in 10 of 10 cases. In contrast, of the 11 of HCCs that did not show TGF-beta overexpression, only 1 showed increased Smad4 immunolabeling. Increased TGF-beta receptor II and Smad4 labeling was associated with a worse nuclear grade and increased mitotic activity. For the hepatic adenomas, the 13 women had an average age at resection of 36 +/- 10 years. Whereas 2 adenomas showed over expression of TGF-beta receptor II, there was no Smad4 overexpression in any case. In conclusion, increased Smad4 protein expression in HCC is tightly linked to overexpression of TGF-beta II receptors and is associated with increased mitoses and a worse nuclear grade. Hepatic adenomas only rarely show overexpression of TGF-beta II receptors and did not show increased Smad4 labeling. The results from this study indicate that Smad4 protein overexpression is present in a subset of HCCs and is strongly correlated with immunostaining for TGF-beta II receptor, findings that may represent activation or dysregulation of the TGF-beta signaling pathway.

Adenoma↗
PubMed2003

Activin A induces phosphorylation of Smad2 but not complex formation of Smad2 with Smad4 in human colon cancer cell line HT-29.

Western blotting coupled with immunoprecipitation showed that activin A treatment induced phosphorylation of Smad2 but not complex formation of Smad2/4 in human colon cancer-derived HT-29 cells. Because HT-29 cells expressed neither Smad4 mRNA nor Smad4 protein, it is suggested that deletion of Smad4 leads to a defect of formation of Smad2/4 complex upon activin A stimulation in HT-29 cells.

Activins↗
PubMed2005

Two somatic biallelic lesions within and near SMAD4 in a human breast cancer cell line.

Loss of chromosome arm 18q is a common event in human pancreatic, colon, and breast cancers and is often interpreted as representing loss of one or more tumor-suppressor genes. In this article, we describe two novel biallelic deletions at chromosome band 18q21.1 in a recently characterized human breast cancer cell line, HCC-1428. One lesion deletes a fragment of approximately 300 kb between SMAD4 and DCC that encodes no known genes. The second lesion is an in-frame SMAD4 deletion (amino acids 49-51) that affects the level of SMAD4 protein but not the SMAD4 message. This change accelerates 26S proteasome-mediated degradation of both endogenous and exogenous mutant SMAD4. Examination of normal DNA from the same patient demonstrated that both lesions are somatic and associated with loss of both normal alleles. These data support the concept that two independent tumor-suppressor loci exist at chromosome segment 18q21.1, one at SMAD4 and the other potentially at an enhancer of DCC or an unrelated novel gene.

Alleles↗
PubMed2001

Apoptotic cell death and Smad4 expression in transitional cell carcinoma of the renal pelvis and ureter.

PURPOSE: To investigate the frequency of apoptosis and the expression of Smad4 protein as well as their roles in transitional cell carcinoma (TCC) of the renal pelvis and ureter. METHODS: Apoptosis was detected by using terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) technique in 34 formalin-fixed and paraffin-embedded specimens of renal pelvic and ureteral TCC. The expression of Smad4 was immunohistochemically studied. RESULTS: The incidence of apoptosis ranged from 1.10 to 3.75% with a median of 2.50% in TCC of the renal pelvis and ureter. The incidence of apoptosis was noted to be closely related to histologic grade but not to pathologic stage of the cancer. The expression of Smad4 was detected in six of 34 cases (17.6%). Regarding subcellular distribution, Smad4 protein was localized both in cytoplasm and nucleus of the cancer cells. In comparing the incidence of apoptosis with the expression of Smad4, no significant associations were seen between them. The expression of Smad4 was not related to the tumor grade nor stage of the cancer. CONCLUSIONS: The present study demonstrated close association of the incidence of apoptosis with the tumor grade of TCC of the renal pelvis and ureter. Significance of Smad4 expression was not noted in the study. It suggests that apoptotic cell death may play an important role in the tumor progression of renal pelvic and ureteral TCC.

Adult↗
PubMed2004

Coexpression of matrix metalloproteinase-7 (MMP-7) and epidermal growth factor (EGF) receptor in colorectal cancer: an EGF receptor tyrosine kinase inhibitor is effective against MMP-7-expressing cancer cells.

PURPOSE: Matrix metalloproteinase-7 (MMP-7) plays an important role in carcinoma invasion and metastasis of cancer. Recent studies focus on diverse roles of MMP-7, other than as a protease, during cancer progression. MMP-7 activates the epidermal growth factor (EGF) receptor by releasing an EGF ligand, tumor growth factor (TGF)-alpha. EXPERIMENTAL DESIGN: We examined expression of MMP-7 and EGF receptor in an immunohistochemical study of 40 colorectal cancer (CRC) cases. To determine the relationship between the EGF receptor and MMP-7, with a potential curative application, we compared the antitumor activity of the EGF receptor tyrosine kinase inhibitor (gefitinib) between MMP-7 transfectant, KYSE150 and HT29, and control cells. RESULTS: We found a statistically significant correlation (P = 0.04) between MMP-7 and activated (phosphorylated) EGF receptor expression, both being positive in six (15%) cases. Gefitinib reduced the cell number ratio more for MMP-7 transfectant than mock cells, and the proportion of apoptotic cells was 1.5 times higher in MMP-7 transfectant than mock cells by annexin/propidium iodide staining. This was mediated by activation of a TGF-beta signal as confirmed by the abundant expression of TGF-beta protein, the cytoplasmic to nuclear translocation of Smad4 protein by the administration of gefitinib, and the quantitative assay of the plasminogen activator inhibitor-1 promoter/luciferase construction. CONCLUSIONS: We propose that there are some cancers with up-regulated MMP-7 expression that leads to the activation of apoptotic activity of TGF-beta, which is susceptible to treatment with EGF receptor tyrosine kinase inhibitor.

Adult↗