Altered TGF-β signaling in a subpopulation of human stromal cells promotes prostatic carcinogenesis

Carcinoma-associated fibroblasts (CAF) play a critical role in malignant progression. Loss of TGF-β receptor II (TGFβR2) in the prostate stroma is correlated with prostatic tumorigenesis. To determine the mechanisms by which stromal heterogeneity because of loss of TGFβR2 might contribute to cancer progression, we attenuated transforming growth factor beta (TGF-β) signaling in a subpopulation of immortalized human prostate fibroblasts in a model of tumor progression. In a tissue recombination model, loss of TGFβR2 function in 50% of the stromal cell population resulted in malignant transformation of the nontumorigenic human prostate epithelial cell line BPH1. Mixing fibroblasts expressing the empty vector and dominant negative TGFβR2 increased the expression of markers of myofibroblast differentiation [coexpression of vimentin and alpha smooth muscle actin (αSMA)] through elevation of TGF-β1 and activation of the Akt pathway. In combination, these two populations of stromal cells recapitulated the tumor inductive activity of CAFs. TGFβR2 activity in mixed stromal cell populations cultured in vitro caused secretion of factors that are known to promote tumor progression, including TGF-β1, SDF1/CXCL12, and members of the fibroblast growth factor (FGF) and bone morphogenetic protein (BMP) families. In vivo, tissue recombination of fibroblasts overexpressing TGF-β1 and SDF1/CXCL12 not only induced transformation of BPH1 cells, but also promoted a robust growth of highly invasive cells, similar to effects produced by CAFs. While the precise nature and/or origin of the particular stromal cell populations in vivo remain unknown, these findings strongly link heterogeneity in TGF-β signaling to tumor promotion by tumor stromal cells.     

A short-hairpin RNA targeting osteopontin downregulates MMP-2 and MMP-9 expressions in prostate cancer PC-3 cells

Osteopontin (OPN), a secreted phosphoglycoprotein, is frequently associated with cell proliferation and tumor metastatic spread in a variety of cancers. It has been reported that OPN induce matrix metalloproteinase (MMP)-2 and MMP-9 activations through nuclear factor kappaB (NF-κB)-mediated signaling pathways. In this study, we investigated the roles of OPN in human prostate cancer cells and provided clues about the possible functions of IkappaB kinase (IKK) in NF-κB-mediated OPN-induced activations of MMP-2 and MMP-9. Short-hairpin RNA (shRNA) expression vectors were used to inhibit OPN expression in PC-3 cells, human prostate cancer cell line, and IKK inhibitor VII were applied to inhibit the activities of IKK-1 and IKK-2. The results showed that OPN shRNA-mediated RNA interference can downregulate OPN, MMP-2 and MMP-9 expressions, thereby resulting in suppression of the proliferation, migration and invasion of PC-3 cells in vitro and tumor growth in vivo. Moreover, the inhibition of IKK-2 can suppress MMP-2 and MMP-9 expressions, in contrast, the inhibition of IKK-1 has no effects on the OPN, MMP-2 and MMP-9 expression levels. Thus, this study demonstrated that OPN knockdown could downregulate MMP-2 and MMP-9 expressions result in inhibiting the malignant physiological behaviors of PC-3 cell and that IKK-2 may play a crucial role in OPN-induced MMP-2 and MMP-9 expressions via NF-κB-mediated signaling pathways.     

Interleukin 8 expression regulates tumorigenicity and metastases in androgen-independent prostate cancer.

Interleukin 8 (IL-8) is mitogenic and chemotactic for endothelial cells. Within a neoplasm, IL-8 is secreted by inflammatory and neoplastic cells. The highly metastatic PC-3M-LN4 cell line overexpresses IL-8 relative to the poorly metastatic PC-3P cell line. We evaluated whether IL-8 expression by human prostate cancer growing within the prostate of athymic nude mice regulates tumor angiogenesis, growth, and metastasis. PC-3P cells were transfected with the full-length sense IL-8 cDNA, whereas PC-3M-LN4 cells were transfected with the full-sequence antisense IL-8 cDNA. Control cells were transfected with the neomycin resistance gene (Neo). In vitro, sense-transfected PC-3P cells overexpressed IL-8-specific mRNA and protein, which resulted in up-regulation of matrix metalloproteinase 9 (MMP-9) mRNA, and collagenase activity, resulting in increased invasion through Matrigel. After antisense transfection of the PC-3M-LN4 cells, IL-8 and MMP-9 expression, collagenase activity, and invasion were markedly reduced relative to controls. After orthotopic implantation, the sense-transfected PC-3P cells were highly tumorigenic and metastatic, with significantly increased neovascularity and IL-8 expression compared with either PC-3P cells or controls. Antisense transfection significantly reduced the expression of IL-8 and MMP-9 and tumor-induced neovascularity, resulting in inhibition of tumorigenicity and metastasis. These results demonstrate that IL-8 expression regulates angiogenesis in prostate cancer, in part by induction of MMP-9 expression, and subsequently regulates the growth and metastasis of human prostate cancer.     

Human mesenchymal stem cells inhibit metastasis of a hepatocellular carcinoma model using the MHCC97-H cell line

The effects of mesenchymal stem cells (MSC) on the growth and metastasis of human malignancies including hepatocellular carcinoma (HCC) are controversial, and the underlying mechanisms are not yet understood. The aim of this study was to explore the role of MSC in the progression of HCC. We investigated the effect of MSC on in vitro proliferation and invasion and in vivo tumor growth and pulmonary metastasis of MHCC97-H HCC cells with a high metastatic potential. The mRNA and protein levels of transforming growth factor-beta 1 (TGFβ1) and MMP, and their association with the effects of MSC on HCC cells were also evaluated. Co-culture of MHCC97-H cells with MSC conditioned medium significantly enhanced in vitro proliferation but inhibited invasiveness. Following MSC treatment of a nude mouse model bearing human HCC, the MSC were predominantly located in the HCC tissues. Compared with controls, MSC-treated mice exhibited significantly larger tumors (3080.51 ± 1234.78 mm(3) vs 2223.75 ± 1000.60 mm(3), P = 0.045), but decreased cellular numbers of lung metastases (49.75 ± 18.86 vs 227.22 ± 74.67, P = 0.046). Expression of TGFβ1 and MMP-2 was significantly downregulated in the MSC-treated HCC cells. TGFβ siRNA concurrently downregulated expression of TGFβ and MMP-2 in HCC cells and blocked the MSC-induced proliferation and invasiveness of MHCC97-H cells. The MSC enhanced tumor growth but significantly inhibited the invasiveness and metastasis of HCC, possibly through downregulation of TGFβ1. These findings suggest that MSC could be useful in controlling metastatic recurrence of HCC.     

Gene expression of angiogenic factors correlates with metastatic potential of prostate cancer cells

We hypothesize that expression of proangiogenic genes correlates with the metastatic potential of prostate cancer cells. LNCaP, DU-145, and PC-3 are prostate cancer cell lines with low, moderate, and high metastatic potential, respectively, as we demonstrated by their capacity to invade an extracellular matrix, an established tumor invasion assay. The constitutive gene expression of the proangiogenic factors, vascular endothelial growth factor, intercellular adhesion molecule-1, interleukin-8, and transforming growth factor-beta2, was significantly greater in the more metastatic DU-145 and PC-3 cells as compared with LNCaP cells. Matrix metalloproteinase (MMP)-9 is thought to contribute to the invasive phenotype of tumor cells. PC-3 cells showed increased expression of MMP-9 and membrane type 4-MMP as compared with LNCaP and DU-145. Tissue inhibitors of metalloproteinase 1 and 4 gene expression were elevated in DU-145 and PC-3 cells, but paradoxically, LNCaP cells had undetectable levels of these genes. We transfected and overexpressed MMP-9 in poorly metastatic LNCaP cells and measured their invasive activity. Transient expression of human MMP-9 in LNCaP cells produced a 3-5-fold increase in MMP-9 activity with a comparable increase in invasiveness. Antisense ablation of the expression of MMP-9 in DU-145 and PC-3 cells produced concomitant inhibition of the gene expression of the proangiogenic factors, vascular endothelial growth factor, and intercellular adhesion molecule-1 (ICAM-1). Treatment of DU-145 and PC-3 cells with a selective chemical inhibitor of MMP-9 proteinase activity also inhibited their invasive activity. These results support our hypothesis that metastatic potential of prostate cancer cells correlates with expression of proangiogenic factors.     

Gallic acid suppresses the migration and invasion of PC-3 human prostate cancer cells via inhibition of matrix metalloproteinase-2 and -9 signaling pathways

Epidemiological studies have demonstrated that a natural diet or consumption of fruits or vegetables can decrease the risk of cancer development. Cancer cells can migrate to and invade other organs or tissues that cause more difficulty to treat them and this also results in the need for treatments targeting multiple cellular pathways. Gallic acid (GA) has been demonstrated to possess multiple biological activities including anticancer function. However, no report exist on GA inhibited invasion and migration of human prostate cancer cells. We investigated the effects of migration and invasion in GA-treated PC-3 human prostate cancer cells with a series of in?vitro experiments. Boyden chamber transwell assay was used to examine the migration and invasion of PC-3 cells. Western blotting, real-time PCR and gelatin zymography were used for determining the protein levels, gene expression and enzyme activities of matrix metalloproteinase-2 (MMP-2) and -9 in?vitro. Results indicated that GA inhibited the invasion and migration of PC-3 cells and these effects are dose-dependent. GA inhibited the protein levels of MMP-2 and -9, son of sevenless homolog?1 (SOS1), growth factor receptor-bound protein 2 (GRB2), protein kinase C (PKC) and nuclear factor-κ B (NF-κB) p65, c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase 1/2 (ERK1/2), p38, p-AKT (Thr308) and p-AKT (Ser473), but it promoted the levels of phosphatidylinositol 3-kinase (PI3K) and AKT in PC-3 cells. GA also reduced the enzyme activities of MMP-2 and -9 in the examined cells. Moreover, the down-regulation of focal adhesion kinase (FAK) and Ras homolog gene family, member A (Rho A) mRNA expression levels, and up-regulation of the tissue inhibitor of metalloproteinase-1 (TIMP1) gene levels occurred in GA-treated PC-3 cells after 24?h treatment. Based on these observations, we suggest that GA might modulate through blocking the p38, JNK, PKC and PI3K/AKT signaling pathways and reducing the NF-κB protein level, resulting in the inhibition of MMP-2 and -9 of PC-3 human prostate cancer cells.     

Expression of collagenase-1 (MMP-1) promotes melanoma growth through the generation of active transforming growth factor-beta

Tumor cell invasion through basement membranes and into stromal tissue are key steps for promoting growth and metastasis. Tumor cells express various extracellular-matrix-degrading enzymes such as matrix metalloproteinases (MMPs) to degrade extracellular matrix components to facilitate tumor migration and invasion. Histological and clinical studies suggest a role for MMP-1 (collagenase-1) in malignant melanoma invasion. In this study, we evaluated MMP-1 in regulating malignant phenotypes of human melanoma cells by generating human melanoma cells stably transfected with pro-MMP-1 cDNA. The transfectants expressed the active form of MMP-1 associated with cells and showed enhanced invasive and growth abilities in type I collagen gel. Furthermore, MMP-1 expression promoted anchorage-independent growth, which was inhibited in the presence of type II transforming growth factor (TGF)-beta receptor:Fc fusion protein that scavenges TGF-beta receptors. Finally, we demonstrated that MMP-1 directly generated active TGF-beta from its latent form. Thus, these results suggest that MMP-1 produced from melanoma cells would play a role in tumor progression by both degrading matrix proteins and generating active growth factors such as TGF-beta in vivo.     

Nuclear receptor 4A1 (NR4A1) antagonists target paraspeckle component 1 (PSPC1) in cancer cells

Paraspeckles compound 1 (PSPC1) is a multifunctional protein that plays an important role in cancer cells, where PSPC1 is a master regulator of pro-oncogenic responses that includes activation of TGFβ (TGFβ1), TGFβ-dependent EMT, and metastasis. The pro-oncogenic activities of PSPC1 closely resembled those observed for the orphan nuclear receptor 4A1 (NR4A1, Nur77) and knockdown of NR4A1 decreased expression of PSPC1 in MDA-MB-231 breast, H1299 lung, and SNU449 liver cancer cells. Similar results were observed in these same cell lines after treatment with bisindole–derived (CDIMs) NR4A1 antagonists. Moreover, PSPC1-dependent regulation of TGFβ, genes associated with cancer stem cells and epithelial to mesenchymal transition (EMT) were also downregulated after NR4A1 silencing or treatment of breast, lung, and liver cancer cells with CDIM/NR4A1 antagonists. Results of chromatin immunoprecipitation (ChIP) assays suggest that NR4A1 regulates PSPC1 through interaction with an NBRE sequence in the PSPC1 gene promoter. These results coupled with in vivo studies showing that NR4A1 antagonists inhibit breast tumor growth and downregulate PSPC1 in tumors indicate that the pro-oncogenic nuclear PSPC1 factor can be targeted by CDIM/NR4A1 antagonists.     

RRR-alpha-tocopheryl succinate inhibits human prostate cancer cell invasiveness

RRR-alpha-tocopheryl succinate (alpha-vitamin E succinate, VES), one of the vitamin E derivatives, can effectively inhibit the proliferation of human prostate cancer cells. However, little is known about its effect on prostate cancer cell invasive ability. Tumor metastasis is a complex process and the extracellular matrix (ECM) is the first barrier that tumor cells encounter. Therefore, we tested the effect of VES on the invasion of different prostate tumor cells, PC-3, DU-145, and LNCaP, through Matrigel, a reconstituted ECM, using an in vitro cell invasion assay. The invasion of PC-3 and DU-145 cells through Matrigel was inhibited by 20 microM VES after treating for 24 h. The condition did not alter cell survival, cell cycle, cell adhesion or cell motility. We further investigated whether the ability of VES to inhibit prostate cancer cell invasiveness was associated with its ability to inhibit the activity of matrix metalloproteinases (MMPs), the key enzymes in the proteolysis of basement membrane during invasion. PC-3 and DU-145 cells that were treated with VES showed a significant reduction in the levels of MMP-9 in the culture medium. In contrast, LNCaP cells, which did not secrete MMP-9, were poorly invasive in Matrigel and were hardly affected by treatment with VES. This is the first report suggesting that VES inhibits human prostate cancer cell invasiveness and the reduction of secreted MMP-9 activity could be one of the contributory factors, which points to the potential use of VES in the prevention and therapy of prostate cancer invasion.     

The influence of CD44v3-v10 on adhesion, invasion and MMP-14 expression in prostate cancer cells

The expression of certain CD44 variants has been linked with metastasis and tumour progression. In particular, high molecular weight forms of CD44 show restricted expression in tumours and may correlate with tumour development and metastasis. In this study, we examined the expression of CD44 variants in prostate cancer cell lines: the invasive PC-3 and DU-145, low invasive LNCaP, and two non-invasive prostate epithelial cell lines. PC-3 prostate cancer cells were transfected with a high molecular weight CD44 variant isoform, CD44v3-v10, isolated from non-invasive prostate epithelial cell lines. These transfected cells (PC-NIVO) were assessed using in vitro invasion, tumour-endothelial, growth, and migration assays. The expression of MMP-14 was examined using SDS-PAGE and Western blot analysis. Transfected PC-3 cells (PC-NIVO) were found to be less adherent to endothelial cells and had significantly reduced invasiveness compared to wild-type PC-3 or control cells. In addition, tumour cell adhesion to endothelial cells and invasiveness was increased after exposure to HGF/SF, and can be blocked by the presence of anti-CD44 antibodies. Further investigation revealed a reduction in the expression of MMP-14 in PC-NIVO cells, but not in PC-3 or control cells. In conclusion, non-invasive prostate epithelial cells express a high molecular weight CD44 isoform, CD44v3-v10, which may counteract the standard isoform function of CD44 by reducing adhesion and invasion of endothelium by prostate tumour cells through negation of the MMP-14 function.     

The Chemokine CCL2 Increases Prostate Tumor Growth and Bone Metastasis through Macrophage and Osteoclast Recruitment

CC chemokine ligand 2 (CCL2, also known as monocyte chemoattractant protein-1) has been demonstrated to recruit monocytes to tumor sites. Monocytes are capable of being differentiated into tumor-associated macrophages (TAMs) and osteoclasts (OCs). TAMs have been shown to promote tumor growth in several cancer types. Osteoclasts have also been known to play an important role in cancer bone metastasis. To investigate the effects of CCL2 on tumorigenesis and its potential effects on bone metastasis of human prostate cancer, CCL2 was overexpressed into a luciferase-tagged human prostate cancer cell line PC-3. In vitro, the conditioned medium of CCL2 overexpressing PC-3luc cells (PC-3^lucCCL2) was a potent chemoattractant for mouse monocytes in comparison to a conditioned medium from PC-3^lucMock. In addition, CCL2 overexpression increased the growth of transplanted xenografts and increased the accumulation of macrophages in vivo. In a tumor dissemination model, PC-3^lucCCL2 enhanced the growth of bone metastasis, which was associated with more functional OCs. Neutralizing antibodies targeting both human and mouse CCL2 inhibited the growth of PC-3^luc, which was accompanied by a decrease in macrophage recruitment to the tumor. These findings suggest that CCL2 increases tumor growth and bone metastasis through recruitment of macrophages and OCs to the tumor site.     

Overexpression of Fn14 promotes androgen-independent prostate cancer progression through MMP-9 and correlates with poor treatment outcome

Fibroblast growth factor-inducible 14 (Fn14), a transmembrane receptor binding to the multifunctional cytokine tumor necrosis factor-like weak inducer of apoptosis (TWEAK), is known to modulate many cellular activities including cancer progression. Here, we demonstrated the significant role of Fn14 in invasion, migration and proliferation of androgen-independent prostate cancer (AIPC) cells. Fn14 and its ligand TWEAK were highly expressed in two AIPC cell lines, DU 145 and PC-3, whereas expression was weak in androgen-sensitive LNCaP cells. Fn14 knockdown using small-interfering RNAs attenuated migration, invasion and proliferation and enhanced apoptosis in the AIPC cell lines. Both forced overexpression of Fn14 by stable Fn14 complementary DNA transfection to PC-3 cells (PC-3/Fn14) and ligand activation by recombinant TWEAK in PC-3 cells enhanced invasion. Fn14 was shown to modulate expression of matrix metalloproteinase (MMP)-9, and MMP-9 mediated the invasive potential influenced by Fn14 in PC-3 cells. In vivo, subcutaneous xenografts of PC-3/Fn14 grew significantly faster than xenograft of PC-3/Mock, and the invasive capacity in PC-3/Fn14 was found to be higher than that of PC-3/Mock as evaluated in an invasion model of the diaphragm. Furthermore, the messenger RNA expressions of MMP-9 in PC-3/Fn14 xenografts were significantly higher than those in PC-3/Mock xenografts. Clinically, high expression of Fn14 was significantly associated with higher prostate-specific antigen recurrence rate in patients who underwent radical prostatectomy. In conclusion, the overexpression of Fn14 may contribute to multiple malignant cellular phenotypes associated with prostate cancer (PCa) progression, in part via MMP-9. TWEAK-Fn14 signaling may be a novel therapeutic target of PCa.     

Transforming growth factor ß1 induces apoptosis by suppressing FLICE‐like inhibitory protein in DU145 prostate epithelial cells

Transforming growth factor ß (TGFß) is a paracrine mediator of prostate epithelial cell apoptosis. In rodents, castration induces production of TGFβ by stromal cells, which leads to apoptosis of epithelial cells. To identify potential mediators of this cell death pathway, we developed a model using DU145 cells, a tumorigenic human prostate epithelial cell line. We discovered that at low density, in low mitogen media, DU145 cells apoptose when treated with TGFβ1. Prior to the onset of death, TGFβ1 treatment downregulated the expression of the caspase inhibitor FLICE‐like inhibitory protein (FLIP), at both the mRNA and protein level, suggesting a causal role between FLIP downregulation and cell death. To confirm the importance of FLIP in TGFβ1‐induced apoptosis, we employed small interfering RNA (siRNA) to silence FLIP expression. Doing so led to apoptosis, which is consistent with the hypothesis that FLIP prevents death in these cells. Furthermore, inhibition of caspase‐8 by siRNA knockdown partially rescued the apoptotic effects of TGFβ1, suggesting a role for death receptor signaling components in TGFß‐mediated death of prostate epithelial cells. © 2008 Wiley‐Liss, Inc.     

Bone marrow macrophages support prostate cancer growth in bone

Resident macrophages in bone play important roles in bone remodeling, repair, and hematopoietic stem cell maintenance, yet their role in skeletal metastasis remains under investigated. The purpose of this study was to determine the role of macrophages in prostate cancer skeletal metastasis, using two in vivo mouse models of conditional macrophage depletion. RM-1 syngeneic tumor growth was analyzed in an inducible macrophage (CSF-1 receptor positive cells) ablation model (MAFIA mice). There was a significant reduction in tumor growth in the tibiae of macrophage-ablated mice, compared with control non-ablated mice. Similar results were observed when macrophage ablation was performed using liposome-encapsulated clodronate and human PC-3 prostate cancer cells where tumor-bearing long bones had increased numbers of tumor associated-macrophages. Although tumors were consistently smaller in macrophage-depleted mice, paradoxical results of macrophage depletion on bone were observed. Histomorphometric and micro-CT analyses demonstrated that clodronate-treated mice had increased bone volume, while MAFIA mice had reduced bone volume. These results suggest that the effect of macrophage depletion on tumor growth was independent of its effect on bone responses and that macrophages in bone may be more important to tumor growth than the bone itself. In conclusion, resident macrophages play a pivotal role in prostate cancer growth in bone.     

Farnesyltransferase inhibitor SCH-66336 downregulates secretion of matrix proteinases and inhibits carcinoma cell migration

The ras oncogenes are among those most frequently found in human cancers. Blocking Ras farnesylation is a promising strategy for arresting cancer growth. Ras activates several signaling pathways with key roles in cellular proliferation, invasion, metastasis and angiogenesis. Furthermore, proteolytic activities of matrix proteinases such as urokinase-type plasminogen activator (uPA) and matrix metalloproteinases (MMPs) are regulated by Ras isoforms. Thus, we investigated the effects of SCH-66336, a farnesyltransferase inhibitor, on secretion of components of the plasminogen activation system as well as on the gelatinases MMP-2 and MMP-9, which play pivotal roles in matrix remodeling. SCH-66336 up to 5 microM did not significantly alter the viability of prostate (PC-3) and renal (Caki-1) cancer cells incubated in serum-depleted medium. SCH-66336 partly inhibited the processing of H-Ras, while levels of mature N-Ras and K-Ras remained unaffected. Under these noncytotoxic conditions, uPA and tPA levels were lowered in culture medium but raised in cell lysates, suggesting inhibition of trafficking pathways. In contrast, SCH-66336 had no effect on uPAR expression or on secreted PAI-1 levels. As expected, the reduction of uPA and tPA activities by SCH-66336 inhibited the conversion of plasminogen to plasmin by about 25% in PC-3 cells. SCH-66336 also inhibited the levels of secreted pro-MMP-2 and pro-MMP-9 as well as the release of their inhibitors TIMP-1 and TIMP-2. SCH-66336 decreased both the adhesion and even more so the migration of PC-3 cells on gelatin. Thus, SCH-66336 inhibited farnesylation in both cancer cell types, and H-Ras functions should be reduced by the drug. In addition, the lower levels of secreted proteinases in the presence of SCH-66336 suggest that reduced matrix remodeling and cell migration should occur in treated tumors.     

Manganese superoxide dismutase overexpression inhibits the growth of androgen-independent prostate cancer cells

This study investigates the role of the antioxidant enzyme manganese superoxide dismutase (MnSOD) in androgen-independent human prostate cancer (PC-3) cells' growth rate in vitro and in vivo. MnSOD levels were found to be lower in parental PC-3 cells compared to nonmalignant, immortalized human prostate epithelial cells (P69SV40T). To unravel the role of MnSOD in the prostate cancer phenotype, PC-3 cells were stably transfected with MnSOD cDNA plasmid. The MnSOD protein and activity levels in clones overexpressing MnSOD were increased seven- to eightfold. These cell lines showed elongated cell doubling time, reduced anchorage-independent growth in soft agar compared to parental PC-3 (Wt) cells, and reduced growth rate of PC-3 tumor xenografts in athymic nude mice. Flow cytometric studies showed an increase in membrane potential in the MnSOD-overexpressing clone (Mn32) compared to Wt and Neo cells. Also, production of extracellular H(2)O(2) was increased in the MnSOD-overexpressing clones. As determined by DNA cell cycle analysis, the proportion of cells in G(1) phase was enhanced by MnSOD overexpression. Therefore, MnSOD not only regulates cell survival but also affects PC-3 cell proliferation by retarding G(1) to S transition. Our results are consistent with MnSOD being a tumor suppressor gene in human prostate cancer.     

Human mesenchymal stem cells promote growth of osteosarcoma: Involvement of interleukin‐6 in the interaction between human mesenchymal stem cells and Saos‐2

Our previous study showed that exogenous human mesenchymal stem cells (hMSCs) targeted established osteosarcoma and promoted its growth and pulmonary metastasis in vivo. As a follow‐up, the present study aimed to investigate how hMSCs would interact with Saos‐2 through autocrine/paracrine communication. The results showed that co‐injection of hMSCs with Saos‐2 into the proximal tibia of nude mice could promote tumor growth and progression. In vitro, the proliferation of Saos‐2 and hMSCs was promoted by each other’s conditioned medium, in which interleukin‐6 (IL‐6) played an important role. Osteogenic differentiation of hMSCs could be inhibited by conditioned medium of Saos‐2, in which IL‐6 was also involved. Furthermore, decreased IL‐6 secretion by hMSCs during its osteogenesis and increased IL‐6 secretion in response to conditioned medium of Saos‐2 were observed. Based on these data, we suggest that there was a positive feedback loop of IL‐6 in the interaction between hMSCs and Saos‐2. (Cancer Sci 2010; 101: 2554–2560)     

A comparison of the expression of the malignant phenotype in two androgen-independent human prostate cancer cell lines after orthotopic implantation in nude mice

Expression of some features of the malignant phenotype were compared in the DU145 and PC-3M human prostate cancer cell lines after their intraprostatic growth in nude mice. At necropsy, 27/74 (36%) mice injected with DU145 cells and 41/75 (55%) injected with PC-3M cells had either invasive macroscopic tumors, or microscopic intraprostatic tumor cell nests (p = 0.02). Para-aortic lymph node metastases had occurred in 19% of the DU145 cell, and 44% of the PC-3M cell tumor-bearing animals (p = 0.033). Immunohistochemical staining showed high mutant p53 and vascular endothelial growth factor (VEGF) expression by DU145 cells; PC-3M cells did not express detectable p53, and had relatively low VEGF immunohistochemical reactivity. Assays by ELISA established a statistically significant difference in VEGF levels between the cell lines (p < 0.001). Urokinase-like plasminogen activator (uPA) levels, determined by ELISA, were ten-fold higher in the PC-3M cell tumors, as were matrix metalloproteinase-9 (MMP-9) activities assessed by zymography. These findings of high expression of uPA and MMP-9, two key proteolytic enzymes in the invasive/metastatic process, by PC-3M cell prostatic tumors are consistent with their aggressive behavior; the low VEGF levels compared with those in the poorly metastatic DU145 cell tumors suggest that other angiogenic factors may be critical for prostate cancer cell progression in this model.