Autocrine effect of DHT on FGF signaling and cell proliferation in LNCaP cells: role of heparin/heparan-degrading enzymes.

BACKGROUND: LNCaP cells are androgen-sensitive human prostate cancer cells. They are characterized by a bell-shaped growth curve in response to increasing doses of dihydrotestosterone (DHT) in culture. At a low concentration of DHT (0.1 nM), these cells show an increase in proliferation, but their growth is arrested at a high concentration (100 nM) of DHT. Results of our previous study demonstrated that the inhibitory effect of DHT at a high concentration was mediated through the action of TGF-beta1. The objective of the present study was to elucidate the mechanism of the proliferative effect of DHT in LNCaP cells. METHODS AND RESULTS DHT stimulated LNCaP proliferation only when cells were cultured in the presence of serum. In serum-free cultures, the characteristic DHT-induced proliferation was not observed. The addition of neutralizing antibody against FGF-2 (basic fibroblast growth factor) was able to inhibit this DHT-induced proliferation. These results suggest that the proliferative effect of DHT was mediated through the action of FGF-2. However, results of the reverse transcriptase polymerase chain reaction indicated that LNCaP cells did not express FGF-2 message. As a result, the source of FGF-2 in these cultures must be the serum supplemented in the culture media. FGF-2 can bind to heparin sulfate chains within the extracellular matrix (ECM). In cultures treated with exogenous heparin, the proliferative effect of DHT was abolished. These results led to the development of the hypothesis that DHT treatment mediates the release of FGF-2 entrapped in the ECM through increased heparinase activity. The addition of heparinase to cultures of LNCaP cells, in the absence of DHT, was able to stimulate cell proliferation. Moreover, 0.1 nM DHT caused a significant increase in heparinase activity. CONCLUSIONS: These results provide a possible mechanism for DHT action in LNCaP cells. In the absence of DHT, FGF-2 in culture was trapped in the extracellular matrix and was not available to interact with LNCaP cells. However, in the presence of 0.1 nM DHT, heparinase activity in the culture was elevated and, as a result, it liberated the trapped FGF-2 which, in turn, stimulated proliferation in LNCaP cells.

Transforming growth factor-beta1-induced proliferation of the prostate cancer cell line, TSU-Pr1: the role of platelet-derived growth factor.

The results of our previous study revealed that transforming growth factor-beta1 (TGFbeta1) stimulated proliferation of the prostate cancer cell line, TSU-Pr1. This observation is unexpected, for TGFbeta usually inhibits proliferation in prostate cancer cells. The present study examines possible mechanisms through which TGFbeta1 induces this proliferation. We postulate that TGFbeta1 action is mediated through an indirect mechanism by inducing the expression of platelet-derived growth factor (PDGF), which, in turn, stimulates proliferation. The TGFbeta1-induced proliferation can be abrogated by treatment with a PDGF-neutralizing antibody. Treatment with exogenous PDGF significantly increased TSU-Pr1 proliferation. Finally, treatment of TSU-Pr1 cells with TGFbeta1 resulted in an increase in PDGF secretion. These results indicate that TGFbeta1-induced proliferation in TSU-Pr1 cells is at least mediated through an increased secretion of PDGF.

Transforming growth factor-beta in benign and malignant prostate.

BACKGROUND: The present review summarizes the cellular action of TGF-beta in benign and malignant growth of the prostate. METHODS: TGF-beta is a pleiotropic growth factor. It plays an important role in the regulation of growth and differentiation in many cells. In benign prostatic epithelia, its action is mediated through a paracrine mechanism. It inhibits proliferation and induces apoptosis in prostatic epithelia. It provides a mechanism to maintain epithelial homeostasis in the prostate. In prostatic stroma, its continual action leads to smooth muscle differentiation. This effect of TGF-beta may regulate the development of prostatic smooth muscle nodules in benign prostatic hyperplasia. RESULTS: As prostatic epithelial cells undergo malignant transformation, two major events occur regarding TGF-beta action. These include the loss of expression of functional TGF-beta receptors and overproduction of TGF-beta in malignant cells. The loss of expression of functional TGF-beta receptors provides a growth advantage to cancer cells over their benign counterparts. The overproduction of TGF-beta by cancer cells has a multitude of adverse consequences. TGF-beta can promote extracellular matrix production, induce angiogenesis, and inhibit host immune function. The biological consequence of these activities is an enhanced tumorigenicity in prostate cancer. Results of our recent studies with a rat prostate cancer model suggest that the immunosuppressive effect of TGF-beta seems to be the primary cause of tumor progression. This is because, if these cancer cells were engineered to reduce the production of TGF-beta, tumor growth was inhibited in syngeneic hosts but not in immune compromised hosts. CONCLUSIONS: Our future research should take advantage of this knowledge to devise therapeutic strategies aimed at eradicating prostate cancer.

Cytotoxic sensitivity to tumor necrosis factor-alpha in PC3 and LNCaP prostatic cancer cells is regulated by extracellular levels of SGP-2 (clusterin).

BACKGROUND: SGP-2 is a ubiquitous secreted glycoprotein that prevents cellular apoptosis. This study was carried out to determine the extracellular action of SGP-2 in a model of tumor necrosis factor-alpha (TNF)-induced cytotoxicity using two human prostatic cancer lines, LNCaP and PC3. These two lines were selected because LNCaP cells are highly sensitive to the cytotoxic effect of TNF, while PC3 cells are resistant to TNF at 24 hr. METHODS: Cells were cultured in the presence or absence of TNF (10 ng/ml). LNCaP cells were treated with varying concentrations of exogenous SGP-2, while PC3 cells were treated with antisera to SGP-2 with and without exogenous SGP-2. Following a 24-hr treatment, cultures were assessed by counting of cell number and by the trypan blue exclusion assay. RESULTS: Western blot analysis of conditioned media revealed that PC3 secreted more SGP-2 than did LNCaP. The sensitivity to TNF in LNCaP cells was reduced by the addition of exogenous SGP-2. PC3 cells became sensitive to TNF when SGP-2 antibody was added to the culture. The effect of SGP-2 antibody on PC3 cells was reversed by the addition of exogenous SGP-2 to the culture. CONCLUSIONS: These results suggest that SGP-2 can act as an extracellular mediator of anti-TNF-induced cytotoxicity.

A proliferative effect of transforming growth factor-beta1 on a human prostate cancer cell line, TSU-Pr1.

Transforming growth factor -beta (TGF-beta) is growth inhibitory to many malignant cells, including prostate cancer cells. The present study reports an unusual observation in that TGF-beta is growth stimulatory to a human prostate cancer cell line, TSU-Pr1. The TSU-Pr1 line is highly aggressive and exhibits a rapid rate of proliferation in culture. These cells underwent further proliferation in response to TGF-beta1. Both type I and II receptors to TGF-beta (TPR-I, TPR-II) are expressed in TSU-Pr1 cells. Activation of a luciferase reporter gene, which contains a TGF-beta response element, confirmed that the TGF-beta receptors in TSU-Pr1 cells were functional. RT-PCR analysis and an ELISA assay determined that TSU-Pr1 cells secreted TGF-beta. In conclusion, TSU-Pr1 cells contain functional TGF-beta receptors but instead of the usual growth inhibition by TGF-beta1, these cells undergo proliferation. The present observation provides a proliferative role of TGF-beta in TSU-Pr1 cells, which may play a part in the aggressive phenotype of these cells and, perhaps other prostate cancer cells.

Intracellular levels of SGP-2 (Clusterin) correlate with tumor grade in prostate cancer.

Our previous observations in LNCaP cells in vitro demonstrated an association between apoptotic cell death resistance and SGP-2 (Clusterin) overexpression. Accordingly, we hypothesized that high levels of cellular SGP-2 would aid in identifying biologically aggressive prostate cancer cells with unique survival advantages. To test this hypothesis, 40 archival radical prostatectomy and/or biopsy specimens of varying grades of prostate cancer were subjected to immunohistochemical SGP-2 staining. The resulting epithelial stains were quantified subjectively on a scale of 1-3 by four independent observers. Benign prostatic epithelial cells from young donors served as controls and showed a consistently weak staining intensity. In contrast, prostate cancer specimens showed varying degrees of staining intensity that correlated with a Gleason pattern (P = 0.006). This correlation supports the hypothesis that protection from apoptotic death may account, in part, for biologically aggressive tumor behavior.