Distribution of Transforming Growth Factor-β and Its Receptors in Gastric Carcinoma Tissue

The distribution of the three mammalian isoforms of transforming growth factor (TGF)-β (TGF-β1,-β2, and -β3) as well as their signaling receptors, TGF-β type I and type II receptors (TβR-I and TβR-II, respectively), in gastric carcinoma tissue was examined by immunohistochemistry using specific antibodies. Tissue specimens were obtained from 25 cases of gastric carcinoma, which were classified into two groups according to Lauren's classification, i.e. 15 cases of diffuse carcinoma and 10 cases of intestinal carcinoma. In normal gastric mucosa apart from carcinoma nests, all of TGF-β1, -β2, -β3, TβR-I and TβR-II were clearly demonstrated in fundic glands. In sharp contrast, none of them was detectable in surface mucous cells. In carcinoma cells, strong staining for TGF-β1, -β2 and β3 was obtained only in diffuse-type carcinoma. In particular, carcinoma cells scattered as single cells or small nests had a tendency to show strong staining for TGF-βs. The receptors tended to be distributed concomitantly with the ligands, and diffuse-type carcinoma showed stronger receptor staining than intestinal-type carcinoma. In cancer stroma, TGF-βs and receptors were detected in both diffuse and intestinal types, but the area with positive staining was wider and more dispersed in diffuse-type carcinoma than in intestinal carcinoma. These results suggest that TGF-β may contribute in part to the variety of histogenesis and mode of progression of gastric carcinoma.     

Close Correlation between the Dephosphorylation of p53 and Growth Suppression by Transforming Growth Factor-β1 in Nasopharyngeal Carcinoma Cells Transduced with Adenovirus Early Region Genes

The mechanism of growth inhibition by transforming growth factor (TGF)-β1 was investigated. We examined the growth inhibitory effects of TGF-β1 on human nasopharyngeal carcinoma (KB) cells which constitutively expressed p53. TGF-β1 suppressed the DNA synthesis of KB cells in a dose-dependent manner. It had minimal effect on adenovirus-2-transduced KB cells expressing either adenovirus early region 1B (E1B) or 1A (E1A) product, which respectively binds to p53 or Rb product and inhibits its function, and no growth inhibition at all was observed with KB cells expressing both E1B and E1A products. Dephosphorylation of the p53 was promoted by TGF-β1 stimulation in KB cells, but not in E1B-producing KB cells, which sequestrate the function of p53. The growth inhibition of KB cells by TGF-β1 was significantly reduced by treatment with okadaic acid. These results suggest that p53 transduces the antiproliferative signal of TGF-β1 possibly through its dephosphorylation.     

Elevated Levels of Plasma Transforming Growth Factor-β in Patients with Hepatocellular Carcinoma

We measured the plasma transforming growth factor-β (TGF-β) concentration in 14 patients with human hepatocellular carcinoma (HCC) and 9 age-matched normal subjects using growth inhibition assay of mink lung epithelial cells. The calculated plasma TGF-β concentration in the patients with HCC was 28.6 ± 27.9 ng/ml (mean± SE), showing significant elevation compared with that in 9 normal subjects (5.3 ± 3.3 ng/ml, P<0.01). In three cases, we could measure plasma TGF-β levels before and after their treatment for HCC. The plasma TGF-β levels decreased from 59.0 to 18.2 ng/ml after hepatic resection in one case, and from 24.0 to 10.7 ng/ml and from 12.4 to 3.4 ng/ml after transhepatic arterial embolization in the other two cases. These data indicate that plasma TGF-β level is elevated in patients with HCC, probably due to release from HCC tissues.     

Detection of Active Form of Transforming Growth Factor-β in Cerebrospinal Fluid of Patients with Glioma

We examined transforming growth factor-β (TGF-β) activity in cerebrospinal fluid of 39 patients with various brain tumors, and found it in 10 glioma cases that had lesions related to subarachnoid space or ventricle. In one glioma case, TGF-β detected on admission disappeared after radiation and chemotherapy. We confirmed that five glioma cell lines produced TGF-β, and that four of them produced active form of TGF-β directly. The active form of TGF-β was also identified from cerebrospinal fluid before the acidification treatment in two cases. The calculated contents were 110 ng/ml and 18 ng/ml. These results indicate that active form of TGF-β is directly produced by tumor cells in patients with glioma, and may contribute to immunodeficiency of the host.     

Potentiation of Metastatic Capacity by Transforming Growth Factor-β1 Gene Transfection

This study was designed to assess whether the excessive secretion of transforming growth factor-β1 (TGF-β1) by Chinese hamster ovary (CHO) cells transfected with TGF-β1 gene may be linked to the development of a metastatic phenotype. We observed large numbers of metastatic colonies in the lungs of nude mice inoculated with the transfected CHO cells. The tumors derived from these transfected cells demonstrated marked angiogenesis. We postulate that the overproduction of TGF-β1 by these tumors may participate in the metastatic progression following establishment of angiogenesis at the primary tumor site.     

Mutations and Reduced Expression of the Transforming Growth Factor-β Receptor II Gene in Rat Lung Adenocarcinomas Induced by N-Nitrosobis-(2-hydroxypropyl)amine

Mutations and expression of the transforming growth factor-β receptor type II (TGF-βRII) gene were investigated in lung adenocarcinomas induced by N -nitrosobis(2-hydroxypropyl)amine (BHP) in rats. Males of the Wistar strain, 6 weeks old, were given 2000 ppm of BHP in their drinking water for 12 weeks and then maintained without further treatment until killed at week 25. Total RNA was extracted from 12 adenocarcinomas and mutations in TGF-βRII were investigated by RT-PCR-restriction-SSCP analysis followed by sequencing analysis. Two out of 12 adenocarcinomas showed band shifts, indicative of mutations (16.7%). One was a CTG-to-TTG (Leu to Leu) transition at codon 308 without amino acid alteration and the other a frameshift deletion of one of two guanines at nucleotides 1434 to 1435 (codon 477 to 478). Semi-quantitative RT-PCR analysis demonstrated significantly reduced TGF-βRII expression in adenocarcinomas, as compared with normal lung tissue. These results suggest that TGF-βRII alterations may play a role in the acquisition of growth advantage by lung adenocarcinomas induced by BHP in rats.     

Improvement of Macrophage Dysfunction by Administration of Anti-transforming Growth Factor-β Antibody in EL4-bearing Hosts

An experimental therapy for improvement of macrophage dysfunction caused by transforming growth factor-β (TGF-β) was tried in EL4 tumor-bearing mice. TGF-β was detected in cell-free ascitic fluid from EL4-bearers, but not in tbat from normal mice, by western blot analysis. The ascites also showed growth-suppressive activity against MvlLn cells, and the suppressive activity was potentiated by transient acidification. To investigate whether the functions of peritoneal macrophagcs were suppressed in EL4-bearers, the abilities to produce nitric oxide and tumor necrosis factor-α (TNF-α) upon lipopolysaccharide (LPS) stimulation were measured. Both abilities of macrophages in EL4-bearing mice were suppressed remarkably on day 9, and decreased further by day 14, compared with non-tumor-bearing controls. TGF-β activity was abrogated by administration of anti-TGF-α antibody to EL4-bearing mice. While a large amount of TGF-β was detected in ascitic fluid from control EL4-bearers, little TGF-β was detectable in ascites from EL4-bearers given anti-TGF-β antibody. Furthermore, while control macrophages exhibited little or no production of nitric oxide and TNF-α on LPS stimulation in vitro , macrophages from EL4-bearers administered with anti-TGF-β antibody showed the same ability as normal macrophages. These results clearly indicate that TGF-β contributes to macrophage dysfunction and that the administration of specific antibody for TGF-β reverses macrophage dysfunction in EL4-bearing hosts.     

Inhibition of Tumor Necrosis Factor-α and β Secretion by Lymphokine Activated Killer Cells by Transforming Growth Factor-β

Transforming growth factor- β (TGF- β ) has a variety of immunosuppressive properties. We investigated the effect of TGF- β secreted by glioblastoma (T98G) cells on the secretion of tumor necrosis factor- α and - β (TNFs) by lymphokine activated killer (LAK) cells stimulated with tumor cells. The supernatant from T98G cells was preincubated with anti-TGF- β l and - β 2 neutralizing antibodies or untreated, and added to a coculture of LAK and Daudi cells. The neutralizing antibodies were added to LAK/Daudi and LAK culture, and natural human TGF- β and recombinant human TGF- β were also added to the LAK/Daudi culture. LAK cells were also cultured with T98G cells, of which the supernatant contained both active and latent forms of TGF-/ β 1 and TGF- β 2, and the neutralizing antibodies were added to the coculture. TNFs activity in the supernatants from LAK/ Daudi cultures was examined by a specific bioassay. Addition of the supernatant from T98G cells to LAK/Daudi culture resulted in the inhibition of TNFs secretion by LAK cells. The inhibition was abrogated by the pretreatment of the supernatants with the anti-TGF- β antibodies. Addition of TGF- β and TGF- β to LAK/Daudi culture inhibited TNFs secretion by LAK cells in a dose-dependent manner. Addition of anti-TGF β - antibodies to LAK culture resulted in an increase of TNFs secretion. These results suggest that, if tumor cells have the capacity to convert TGF- β from a latent to an active form, the active TGF- β suppresses TNFs secretion by LAK cells stimulated with the tumor cells, and that TGF- β secreted and activated by glioblastoma cells suppresses the propagation of immune reaction by inhibiting TNFs secretion by activated lymphocytes adjacent to tumor cells.     

Reduced Levels of Transforming Growth Factor-beta Type I Receptor in Human Gastric Carcinomas

The expressions of transforming growth factor beta (TGF-β) and its receptor and TGF-β inhibitory element (TIE)-binding protein were examined on human gastric carcinomas by Northern blot hybridization, immunohistochemistry, affinity labeling and gel retardation analysis. TGF-β mRNA was expressed in tumor and normal tissues at various levels. Immunohistochemically, TGF-β expression was confirmed to be present within tumor cells. Out of the 17 human gastric carcinoma tissues, 14 (82%) showed a reduction in the level of type I receptor (65 kDa) for TGF-β when compared to corresponding normal mucosas. Interestingly, in seven of the 14 tumors the level of TIE-binding protein in the tumor tissue was lower than that in normal mucosa. Human gastric carcinoma cell line TMK-1, whose growth was inhibited by TGF-β, had only type I receptor for TGF-β and showed a high level of TIE-binding protein. Conversely, MKN-1, a TGF-β -resistant cell line, exhibited an extremely low level of TGF-β receptor and had no TIE-binding protein. These results overall indicate that although human gastric carcinoma cells produced TGF-β, they showed a reduction in TGF-β type I receptor and a low level of TIE-binding protein, resulting in escape from growth inhibition by TGF-β.     

Transforming Growth Factor-α Expression of Renal Proximal Tubules in Wistar Rats Treated with Ferric and Aluminum Nitrilotriacetate

A high incidence of renal adenocarcinoma has been observed in rats treated with ferric nitrilotriacetate (Fe-NTA) but not in rats treated with aluminum nitrilotriacetate (Al-NTA). Transforming growth factor (TGF)-α is one of the several cytokines that is known to be expressed in human and rat renal adenocarcinomas. However, its role in neoplastic transformation is still questionable. Therefore, we investigated the effect of repeated Fe-NTA and Al-NTA administration on renal TGF-α expression. Male Wistar rats were given Fe-NTA (n = 16, 5–10 mg FeAg) and Al-NTA (n = 19, 1–2 mg Al/kg) i.p., three times a week for 3 or 12 weeks. Another group of rats (n=4) was given Fe-NTA (5–10 mg FeAg) three times a week for 12 weeks and then left untreated for one year. Immunoreactivity for TGF-α was positive in the collecting ducts and on the apical surface of proximal tubules in the outer stripe of the outer medulla in all the animals including NTA-injected control animals. However, TGF-α immunoreactivity in the regenerative proximal tubular epithelium was observed only in the animals treated with Fe-NTA for 12 weeks. Northern blot analysis also showed expression of TGF-α mRNA only in animals treated with Fe-NTA for 12 weeks. The expression of TGF-α mRNA in the kidney was stronger than that in the liver or brain, TGF-α was also positive in renal cell carcinoma found in animals treated with Fe-NTA for 12 weeks and left untreated for one year. These results suggest that TGF-α expression may play an important role in renal carcinogenesis and that it may be a sensitive marker during the induction stage of renal cell carcinoma.     

Transforming Growth Factor-β and Hepatocyte Growth Factor Produced by Gastric Fibroblasts Stimulate the Invasiveness of Scirrhous Gastric Cancer Cells

Scirrhous gastric carcinoma is characterized by cancer cells that infiltrate rapidly in the stroma with extensive growth of fibroblasts. In the present study, we examined the effect of gastric fibroblasts on the invasiveness of a Scirrhous gastric cancer cell line, OCUM-2D, using an invasion assay. Gastric fibroblast-derived conditioned medium (CM) significantly stimulated the invasiveness of OCUM-2D cells, as did transforming growth factor- β (TGF- β ) and hepatocyte growth factor (HGF). The stimulating activity of gastric fibroblast-derived CM was inhibited significantly by anti-TGF- β neutralizing antibody or anti-HGF neutralizing antibody. TGF- β and HGF were detected in the gastric fibroblast-derived CM, and TGF- β receptor and C-met (HGF receptor) were expressed on OCUM-2D cells. Thus, TGF- β and HGF produced by gastric fibroblasts appear to affect the invasiveness of scirrhous gastric cancer cells. TGF- β was also detected in the conditioned medium derived from OCUM-2D cells, though HGF was not. TGF- β appears to affect the invasiveness of OCUM-2D cells in both paracrine and autocrine fashions.     

Potentiation of Invasive Capacity of Rat Ascites Hepatoma Cells by Transforming Growth Factor-β

The in vitro invasive capacity of poorly invasive cells (W₁), which were cloned from rat ascites hepatoma cells (AH 130), was potentiated dose- and time-dependently by pretreating the cells with transforming growth factor-β (TGF-β). This potentiation of invasive capacity was completely abolished by anti-TGF-β antibody. When the treated cells were ip inoculated into rats, the cells extensively invaded the peritoneum, and formed penetrating tumor nodules. The effect of TGF-β was reversed by subculturing the treated cells without TGF-β. The potentiation of invasive capacity by TGF-β might participate in platelet-associated enhancement of tumor cell metastasis.     

Inhibitory Action of Transforming Growth Factor-β on Induction of Differentiation of Myeloid Leukemia Cells

The effect of transforming growth factor-β (TGF-β) on induction of differentiation of mouse myeloid leukemia M1 cells was investigated. TGF-β1 induced adherence of M1 cells to plastic dishes and inhibited their proliferation. However, it did not induce differentiation-associated properties, such as phagocytic activity, lysozyme activity or morphological maturation. TGF-β1 also caused dosedependent inhibition of dexamethasone-induced differentiation of M1 cells. The inhibitory activity of TGF-β1 was 20 times that of TGF-β2 on M1 cells. These results suggest that TGF-β1 inhibits proliferation and dexamethasone-induced differentiation of M1 cells by interacting with receptors that can distinguish between TGF-β1 and TGF-β2. TGF-β1 had a much lower inhibitory effect on the growth of a variant M1 cell clone, which was resistant to differentiation inducers, and it did not induce adherence of the resistant M1 cells.     

Transforming Growth Factor-β CD4+ T cells Tumor-bearing state Immunosuppression Enhanced Production of TGF-β and a Progressive Increase in TGF-β Susceptibility of Anti-tumor CD4+ T Cell Function

The present study deals with the effect of transforming growth factor-β (TGF-β) on anti-tumor immune responsiveness at various stages of the tumor-bearing state. Spleen cells from BALB/c mice bearing a syngeneic tumor (CSA1M) 1–3 wk after inoculation with CSA1M cells produced interleukin-2 (IL-2) and macrophage-activating factor (MAF)/interferon-γ (IFN-γ) upon in vitro culture without addition of exogenous tumor antigens. This lymphokine production was achieved through collaboration between anti-CSA1M CD4⁺ T cells and antigen-presenting cells that had been pulsed with CSA1M tumor antigens in vivo in the tumor-bearing state. The IL-2-producing capacity of CD4⁺ T cells reached the maximal level as early as one week after tumor implantation but decreased with the progress of tumor-bearing stages. In contrast, the capacity of CD4⁺ T cells to produce MAF/IFN-γ was not affected but was maintained at high levels even late in the tumor-bearing state. The addition of recombinant TGF-β (rTGF-β) to cultures of spleen cells from various tumor-bearing stages resulted in the suppression of lymphokine production. However, the magnitude of the TGF-β-induced suppression varied depending on which tumor-bearing stages of splenic cells were tested as a responding cell population; it was slight in cells from early (1–3 wk) tumor-bearing stages but increased in cells from donor mice at later tumor-bearing stages. Thus, spleen cells from late tumor-bearing stages with weak but significant IL-2-producing and considerable MAF/IFN-γ producing capacities failed to produce these lymphokines when rTGF-β was present in cultures. A progressive increase in the TGF-β susceptibility was also observed for IL-4-producing Th2 as well as IL-2/MAF-producing Th1 cells. In addition, increased levels of TGF-β were detected in plasma from tumor-bearing mice at late stages. Taken together, these results indicate that tumor-bearing mice exhibit enhanced production of TGF-β as well as a progressive increase in the susceptibility of anti-tumor CD4⁺ T cells to TGF-β-induced suppressive mechanisms.     

Growth Inhibition, Enhancement of Intercellular Adhesion, and Increased Expression of Carcinoembryonic Antigen by Overexpression of Phosphoinositides-specific Phospholipase C β1 in LS174T Human Colon Adenocarcinoma Cell Line

By using a retrovirus-derived system we generated derivatives of the human colon adenocarcinoma cell line LS174T (ATCC CL 188) that stably overexpress a full-length cDNA encoding the β1 isoform of bovine phosphoinositides-specific phospholipase C (PI-PLC). This was confirmed by the elevated levels of catalytic activity to release phosphoinositides from phosphatidylinositol (PI-PLC) or phosphatidylinositol-bis-phosphate (PIP₂-PLC), and the enhanced expressions of messenger RNA and protein. PI-PLC β1 overexpresser clones grew to form cell clumps floating in liquid medium, whereas the pMV7-introduced control clones displayed morphologic characteristics that were very similar to those of the parent LS174T cell line. Three individual PI-PLC β1 overexpresser cell lines displayed increased doubling time (18.0 h, 21.5 h, and 23.8 h) when compared with 4 individual pMV7-introduced control cell lines (13.1 h, 10.7 h, 12.9 h, and 9.3 h). Anchorage-independent growth ability in soft agar medium was dramatically suppressed by overexpression of PLC β1, and the ability of PLC-overproducer clones to form aggregates when cultured in liquid medium was dramatically enhanced when compared with that of pMV7-introduced control clones. Tumorigenicity of PLC β1-overproducers was much weaker than that of vector-transduced control clones. The spontaneous release of carcinoembryonic antigen from PLC β1-overproducer clones was much higher than that from pMV7 control clones. The ability of PLC β1-overproducer clones to form aggregates during suspension culture was much stronger than that of the control clones. These results provide the first evidence that elevated levels of endogenous PI-PLC β1 suppress tumor cell growth, but enhance the ability to form cell aggregates and to release carcinoembryonic antigen, an intercellular adhesion molecule.     

MMP-9在上皮性卵巢癌组织中的表达及临床意义

目的 探讨MMP-9在上皮性卵巢癌组织中的表达及临床意义.方法 采用免疫组化方法检测15例正常卵巢组织、15例良性上皮性卵巢肿瘤组织和89例上皮性卵巢癌组织中MMP-9蛋白的表达.结果 正常卵巢组织及良性卵巢肿瘤组织中,MMP-9蛋白不表达或低表达.上皮性卵巢癌中,MMP-9表达水平明显升高,高表达率为56.18%.MMP-9高表达与肿瘤晚期、瘤细胞的低分化、肿瘤转移有显著的相关性(P均＜0.05).结论 上皮性卵巢癌组织中,MMP-9蛋白表达上调.MMP-9蛋白在上皮性卵巢癌的生长和转移过程中发挥着重要的作用,可能作为判断卵巢癌预后的指标.     

Modulation of c-myc Expression by Transforming Growth Factor β1 in Human Hepatoma Cell Lines

The effects of transforming growth factor β1 (TGF-β1) on cell proliferation of human hepatoma cell lines, PLC/PRF/5 and Mahlavu, were investigated under serum-free conditions. DNA synthesis was strongly inhibited in the PLC/PRF/5 cells by addition of TGF-β1 (0.5 to 4.0 ng/ml), but remained unchanged in the Mahlavu cells. Also the expression of c- myc mRNA was suppressed by the addition of TGF-β1 in the PLC/PRF/5 cells but not in the Mahlavu cells. These results indicate that TGF-β1 might regulate cell growth, in part, by modulating c- myc expression, although there is no direct proof that c- myc expression is really relevant to DNA synthesis mediated by TGF-β1.     

Meta-analysis of Excision Repair Cross-complementation Group 1 (ERCC1) Association with Response to Platinumbased Chemotherapy in Ovarian Cancer

Recent studies suggested that the ovarian cancers with negative excision repair cross-complementation group1 enzyme (ERCC1) expression have a better response to platinum-based chemotherapy than those with positiveERCC1 expression. The objective of this study was to evaluate whether ERCC1 expression is associated withresponse to platinum-based chemotherapy in ovarian cancers. MEDLINE, PubMed, Web of Science and CNKIdatabases were used for searching studies relating to ERCC1 protein expression and response to platinum-basedchemotherapy in ovarian cancers. Statistical analysis was based on the method for a fixed effects meta-analysis.Pooled odds ratios (ORs) with 95% confidence intervals for ERCC1 protein expression and response to platinumbasedchemotherapy were generated. Publication bias was investigated with Begg’s test. Five studies involving306 patients with ovarian cancer were included. Compared to patients with positive ERCC1 expression, thosewith negative ERCC1 expression had a better response to platinum-based chemotherapy. The pooled OR was5.264 (95% CI: 2.928 – 9.464, P < 0.001) and publication bias was not found (P = 0.904). The result was similarin both in Asians and Caucasians (P < 0.001 and P = 0.028, respectively). ERCC1 protein e