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Johng S Rhim

Publications and source records attributed to Johng S Rhim.

At least 19 recordsLinked to original sources
PubMed2006

Erythropoietin stimulates growth and STAT5 phosphorylation in human prostate epithelial and prostate cancer cells.

BACKGROUND: Erythropoietin (Epo), the principal regulator of erythroid progenitor survival, growth, and differentiation, initiates its action by binding to its cognate cell surface receptor (EpoR). EpoR have been identified on a variety of non-hematopoietic cells, both normal and malignant, however, little is known about the function of EpoR on malignant cells. METHODS: RT-PCR, Western blotting, and immunohistochemistry were used to demonstrate that prostate cancer cells express EpoR at both the gene and protein level. Cell proliferation assays and STAT5 phosphorylation were used to demonstrate Epo's mitogenic action and intracellular signaling, respectively. RESULTS: We have demonstrated that transformed prostate epithelial and prostate cancer cell lines, as well as primary prostate tissue, express the EpoR. Importantly, the EpoR on prostate cells are functional, as demonstrated by the observation that each of the cell lines exhibited a dose-dependent proliferative response to Epo, and that Epo triggered STAT5b phosphorylation in the cells. CONCLUSION: Human prostatic epithelial cells and prostate cancer cells express functional EpoR, and Epo serves as a growth factor for these cells. These results have implications for our understanding of normal prostatic growth and development and of the pathobiology of human prostate cancer.

Blotting, Western↗
PubMed2006

Phenotypic characterization of telomerase-immortalized primary non-malignant and malignant tumor-derived human prostate epithelial cell lines.

In vitro human prostate cell culture models are critical for clarifying the mechanism of prostate cancer progression and for testing preventive and therapeutic agents. Cell lines ideal for the study of human primary prostate tumors would be those derived from spontaneously immortalized tumor cells; unfortunately, explanted primary prostate cells survive only short-term in culture, and rarely immortalize spontaneously. Therefore, we recently have generated five immortal human prostate epithelial cell cultures derived from both the benign and malignant tissues of prostate cancer patients with telomerase, a gene that prevents cellular senescence. Examination of these cell lines for their morphologies and proliferative capacities, their abilities to grow in low serum, to respond to androgen stimulation, to grow above the agar layer, to form tumors in SCID mice, suggests that they may serve as valid, useful tools for the elucidation of early events in prostate tumorigenesis. Furthermore, the chromosome alterations observed in these immortalized cell lines expressing aspects of the malignant phenotypes imply that these cell lines accurately recapitulate the genetic composition of primary tumors. These novel in vitro models may offer unique models for the study of prostate carcinogenesis and also provide the means for testing both chemopreventive and chemotherapeutic agents.

Adult↗
PubMed2006

Characterization of rat spiral ligament cell line immortalized by adenovirus 12-simian virus 40 hybrid virus.

OBJECTIVES: Spiral ligament fibrocytes play an important role in inner ear ion homeostasis and are classified into several subtypes according to expression of specific enzymes such as Na+, K+ -ATPase, Ca++ -ATPase, and carbonic anhydrase. Although our understanding of the cell and molecular biology of spiral ligament fibrocytes has increased over time, access to these cells still remains a significant hurdle hindering future studies. In this study, we aimed to establish a rat spiral ligament cell line with minimal disruption of the original characteristics. METHODS: The primary spiral ligament fibrocytes were exposed to adenovirus 12-simian virus 40 hybrid virus for immortalization. Karyotypic analysis was performed after stabilization of the infected cells, and the population doubling time was compared to that of the primary cell. The cell line was characterized by immunolabeling and electron microscopy. RESULTS: We describe the establishment and characterization of a line of type I spiral ligament fibrocytes immortalized with an adenovirus 12-simian virus 40 hybrid virus. CONCLUSIONS: This cell line can be a useful research tool for investigating the role of spiral ligament fibrocytes in homeostasis and inflammation of the inner ear.

Adenoviridae↗
PubMed2005

Farnesyltransferase inhibitor effects on prostate tumor micro-environment and radiation survival.

BACKGROUND: Ras activation by mutation, overexpression, or receptor signaling can increase tumor cell survival after irradiation. METHODS: We examined whether inhibiting Ras activity with farnesyltransferase inhibitors (FTI) altered the radiosensitivity and tumor micro-environment in prostate tumors. RESULTS: Treatment with FTIs L-744,832 or FTI-277 reduced clonogenic survival of prostate tumor cells expressing oncogenic H-ras after irradiation. PI3-kinase/Akt and MAPK signaling pathways were downregulated by FTIs in these cells. FTI treatment reduced tumor hypoxia and also reduced MMP-9 expression in tumors with activated mutant H-ras. FTI treatment did not, however, increase apoptosis in irradiated intestine, demonstrating that acute radiation injury of this normal tissue was not enhanced by FTIs. CONCLUSIONS: FTIs can enhance the killing of prostate tumors with activated H-Ras. Together with the absence of increased acute toxicity to normal bowel, these results imply that FTI treatment should be further studied as a possible adjuvant to radiotherapy in the treatment of abdominal cancers with activated Ras signaling.

Alkyl and Aryl Transferases↗
PubMed2005

A methylation profile of in vitro immortalized human cell lines.

Normal human diploid cells have a limited life span and undergo replicative senescence after various limited population doublings. Cells must pass the senescence barrier to become immortal. The exact mechanisms of immortalization are not clear, although inactivation of the RB pathway, and/or the p53 pathway and activation of telomerase has been shown to be necessary for immortalization of certain cell types with DNA viruses or hTERT. Methylation-associated inactivation of tumor suppressor genes plays an important role in tumor progression. To test if gene-specific methylation contributes to the immortalized and transformed phenotype, we analyzed the methylation status of 17 genes in normal cells immortalized with SV40, hTERT, Ad5, Ad12-SV40 or HPV-18. Some of these immortalized lines were progressively transformed and tumorigenic in nude mice. We observed gene-specific methylation in the in vitro immortalized and transformed cells. SV40 and HPV18 immortalization resulted in different methylation spectra. In SV40- and h-TERT-immortalized prostate epithelial cells, the most frequently methylated gene was RASSF1A, while in HPV18-immortalized cell lines, the RAR-beta2 gene was universally methylated. Immortalization with SV40 resulted in methylation of a greater number of genes than immortalization with HPV. Furthermore, in SV40-immortalized cell lines, methylation affected different genes in fibroblasts compared with epithelial cells, suggesting that different mechanisms may be used by SV40 to immortalize cell lines of different origins. In HPV18-immortalized and subsequently transformed cell lines, the most commonly methylated genes were hormone responsive genes, such as AR, ER-beta and RAR-beta2. In general, more genes were methylated in neoplastically-transformed cell lines than in only immortalized cell lines, indicating that accumulation of epigenetic abnormalities may contribute to oncogenesis.

Animals↗
PubMed2004

Selective modulation of type 1 insulin-like growth factor receptor signaling and functions by beta1 integrins.

We show here that beta1 integrins selectively modulate insulin-like growth factor type I receptor (IGF-IR) signaling in response to IGF stimulation. The beta1A integrin forms a complex with the IGF-IR and insulin receptor substrate-1 (IRS-1); this complex does not promote IGF-I mediated cell adhesion to laminin (LN), although it does support IGF-mediated cell proliferation. In contrast, beta1C, an integrin cytoplasmic variant, increases cell adhesion to LN in response to IGF-I and its down-regulation by a ribozyme prevents IGF-mediated adhesion to LN. Moreover, beta1C completely prevents IGF-mediated cell proliferation and tumor growth by inhibiting IGF-IR auto-phosphorylation in response to IGF-I stimulation. Evidence is provided that the beta1 cytodomain plays an important role in mediating beta1 integrin association with either IRS-1 or Grb2-associated binder1 (Gab1)/SH2-containing protein-tyrosine phosphate 2 (Shp2), downstream effectors of IGF-IR: specifically, beta1A associates with IRS-1 and beta1C with Gab1/Shp2. This study unravels a novel mechanism mediated by the integrin cytoplasmic domain that differentially regulates cell adhesion to LN and cell proliferation in response to IGF.

Animals↗
PubMed2004

A telomerase-immortalized primary human prostate cancer clonal cell line with neoplastic phenotypes.

Understanding of molecular genetic mechanisms underlying prostate carcinogenesis would be greatly advanced by in vitro models of prostate tumors representing primary tumors. We have successfully established a neoplastic immortalized human prostate epithelial (HPE) clonal culture derived from a primary tumor of a prostate cancer patient (RC-58T) with hTERT, the catalytic subunit of telomerase. The early passage RC-58T cells derived from a radical prostatectomy specimen of a 52-year-old white male patient was transduced through infection with a retrovirus vector expressing the hTERT for the establishment of the RC-58T/hTERT cell line. One clonal line, soft-agar derived from the RC-58T/hTERT cell line, was isolated and further characterized phenotypically and genetically. These clonal (RC-58T/hTERT SA#4) cells are currently growing well at passage 70 and exhibit transformed morphology. The RC-58T/hTERT SA#4 line expressed a high level of telomerase activity and showed anchorage-independent growth in soft agar. The clonal line like the untransduced RC-58T cells (passage 3) expressed prostate specific antigen (PSA), androgen receptor (AR), prostate stem cell antigen (PSCA), and an androgen-regulated prostate specific gene NKX3.1, P16, and cytokeratin (CK) 8. Growth is slightly stimulated by dihydrotestosterone (DHT), and lyates are immunoreactive with AR antibody by Western blot analysis. More importantly, this clonal line produced adenocarcinomas when transplanted into SCID mice. A number of chromosome alterations were observed including the loss of chromosome Y, 1q, 2p, 3p, 4q, 8p, 11p, 14p, 17p and 18q. Our results demonstrate that this primary tumor-derived HPE cell line retained its neoplastic phenotypes and its prostate specific markers and should allow elucidating molecular and genetic alterations involved in prostate cancer. This is the first documented case of an AR and PSA expressing telomerase established human prostate cancer cell line with neoplastic phenotypes from a primary tumor of a prostate cancer patient.

Animals↗
PubMed2003

Duffy antigen facilitates movement of chemokine across the endothelium in vitro and promotes neutrophil transmigration in vitro and in vivo.

The Duffy Ag expressed on RBCs, capillaries, and postcapillary venular endothelial cells binds selective CXC and CC chemokines with high affinity. Cells transfected with the Duffy Ag internalize but do not degrade chemokine ligand. It has been proposed that Duffy Ag transports chemokines across the endothelium. We hypothesized that Duffy Ag participates in the movement of chemokines across the endothelium and, by doing so, modifies neutrophil transmigration. We found that the Duffy Ag transfected into human endothelial cells facilitates movement of the radiolabeled CXC chemokine, growth related oncogene-alpha/CXC chemokine ligand 1 (GRO-alpha/CXCL1), across an endothelial monolayer. In addition, neutrophil migration toward GRO-alpha/CXCL1 and IL-8 (IL-8/CXCL8) was enhanced across an endothelial monolayer expressing the Duffy Ag. Furthermore, GRO-alpha/CXCL1 stimulation of endothelial cells expressing the Duffy Ag did not affect gene expression by oligonucleotide microarray analysis. These in vitro observations are supported by the finding that IL-8/CXCL8-driven neutrophil recruitment into the lungs was markedly attenuated in transgenic mice lacking the Duffy Ag. We conclude that Duffy Ag has a role in enhancing leukocyte recruitment to sites of inflammation by facilitating movement of chemokines across the endothelium.

Amino Acid Sequence↗
PubMed2003

Research into molecular and genetic mechanisms underlying prostate carcinogenesis would be greatly advanced by in vitro prostate cell models.

Prostate cancer is the most commonly diagnosed malignancy in the United States, as well as in the Western world, and the second leading cause of male cancer death in the United States. Despite its high incidence, the molecular and genetic events involved in prostate cancer progression remain poorly understood. A hurdle in understanding the molecular genetic changes in prostate cancer has been the difficulty in establishing premalignant lesions and primary prostate tumors as in vitro cell cultures. Primary epithelial cells grow for a finite life span and then senesce. Immortalization is defined by continuous growth of otherwise senescing cells and is believed to represent an early stage in tumor progression. In order to examine these early stages, we and others have developed in vitro models of prostate epithelial cell immortalization. Because prostate cancer is a multistep, progressive disease with a typical onset later in life and with an usually high number of latent cases that do not develop into clinically manifest cancer, the steps in the progression to malignancy are of particular interest. To understand the many factors that are suspected to contribute to the development of this malignancy, there is a need for an in vitro multistep human prostate epithelial culture system. These models have been extremely important in identifying genetic and molecular changes involved in prostate cancer progression. Recently, novel human cell culture models for the study of prostate cancer have been developed. Successful establishment of primary prostate cancer cell lines from patients' familial and sporadic prostate cancer has been accomplished using telomerase, the gene that prevents cellular senescence. The novel models will be useful for identification and characterization of prostate cancer genes and will provide the new means for testing for chemoprevention and chemotherapeutic agents.

Cell Line, Tumor↗
PubMed2003

C-Jun NH(2)-terminal kinase mediates proliferation and tumor growth of human prostate carcinoma.

PURPOSE: C-Jun NH(2)-terminal kinase (JNK) has been implicated in numerous functions including stress responses, apoptosis,and transformation. The role in transformation is based largely on studies of isolated cell types with little indication of whether JNK plays a general role in a specific human tumor type or whether this occurs in vivo. EXPERIMENTAL DESIGN: We examined 9 human prostate carcinoma cell lines in vitro and a representative line in vivo. RESULTS: For all of the cell lines proliferation is highly correlated with serum-supported JNK activity (r(Pearson) = 0.91; P = 0.004), whereas no relationship was observed for 10 human breast cancer cell lines (r(Pearson) = -0.32). Treatment with characterized antisense oligonucleotides complementary to sequences common to either the JNK1 or JNK2 family of isoforms showed that, whereas antisense JNK1 inhibited growth by a maximum of 57%, antisense JNK2 inhibited proliferation up to 80%. Sense and scrambled control oligonucleotides had little effect (average 3.7 +/- 1.5%). Moreover, systemic treatment of mice bearing established xenografts of PC3 prostate carcinoma cells with antisense JNK1 and JNK2 led to inhibition tumor growth by 57% (P < 0.002) and 80% (P < 0.001), respectively. The difference is significant (P < 0.012). Combined antisense treatment led to a significant increase in frequency of tumor regression (P = 0.022). CONCLUSION: These results indicate that JNK is required for growth of prostate carcinoma cells in vitro and in vivo, and additionally indicate that JNK2 plays a dominant role. The JNK pathway is a novel target in the treatment of prostate carcinoma.

Animals↗
PubMed2003

A novel neoplastic primary tumor-derived human prostate epithelial cell line.

Research into molecular and genetic mechanisms underlying prostate carcinogenesis would be greatly advanced by in vitro models of prostate tumors representing primary tumors. The generation of immortalized primary prostate cancer cells that will accurately reflect the in situ characteristics of malignant epithelium is greatly needed. We have successfully established a neoplastic immortalized human prostate epithelial (HPE) cell culture derived from a primary tumor. The RC-9 cells transduced through infection with a retrovirus vector expressing the E6 and E7 genes (E6E7) of human papilloma virus-16 (HPV-16) are currently growing well at passage 40, whereas RC-9 cells senesced at passage 7. RC-9/E6E7 cells exhibit epithelial morphology and high level of telomerase activity. More importantly, these immortalized cells produced tumors (SCID5038D) when inoculated into SCID mice. RC-9/E6E7 cells and SCID-5038D cells exhibit a high level of telomerase activity and androgen-responsiveness when treated with R1881. Expression of prostate specific antigen (PSA), androgen receptor (AR), prostate stem cell antigen (PSCA), an androgen-regulated prostate specific gene (NKX3.1), p16, cytokeratins 8, 15 and HPV-16 E6 gene was detected in both of these cells. RC-9/E6E7 and SCID5038D cells also showed growth inhibition when exposed to retinoic acid and transforming growth factor (TGF)-beta1, potent inhibitors of prostate epithelial cell growth. A number of chromosome alterations were observed including the loss of chromosomes 2p, 3p, 8p, 13, 14, 16, 17, 18, 21 and the gain of 7 and 20 in the tumor cell line (SCID5038D). These results demonstrate that this primary tumor-derived HPE cell line retained its neoplastic phenotypes and its prostate-specific markers and should allow studies to elucidate molecular and genetic alterations involved in prostate cancer. This is the first documented case of a malignant AR and PSA positive established human prostate cancer cell line from a primary tumor of a prostate cancer patient.

Animals↗
PubMed2002

HEPSIN inhibits cell growth/invasion in prostate cancer cells.

Expression of HEPSIN, a type II transmembrane serine protease in prostate cancer (CaP), has been highlighted by several studies analyzing CaP-specific gene expression alterations by cDNA microarray. Evaluations of the biological functions of HEPSIN in CaP cells are warranted for better assessment of its utility as a biomarker and/or therapeutic target. In stable clones of PC-3/HEPSIN transfectants, there was a dramatic reduction in the cell growth, cell invasion, and soft agar colony formation. A higher proportion of PC-3/HEPSIN cells were in the G(2)-M phase of the cell cycle, and there was also an increase in the cell population undergoing apoptosis. Preliminary analysis of HEPSIN transfections into LNCaP and DU145 cells further revealed cell growth-inhibitory effects. These results underscore that exogenous HEPSIN expression negatively regulates cell growth in metastatic CaP cell lines. Although the cause of the biological consequence of HEPSIN overexpression in primary CaP remains to be determined, the negative cell growth-regulatory effects of HEPSIN in metastatic CaP cells reported here have unraveled possible cellular and molecular mechanisms underlying observations that link decreased/loss of HEPSIN expression with poor prognosis of CaP.

3T3 Cells↗
PubMed2002

The RASSF1A tumor suppressor gene is inactivated in prostate tumors and suppresses growth of prostate carcinoma cells.

We analyzed expression status of the recently identified tumor suppressor geneRASSF1A in primary prostate carcinomas and in prostate cell lines. We found complete methylation of the RASSF1A promoter in 63% of primary microdissected prostate carcinomas (7 of 11 samples). The remaining 4 samples (37%) were partially methylated, possibly because of contamination with normal cells. No promoter methylation was observed in matching normal prostate tissues. High levels of RASSF1A transcript and no methylation of RASSF1A promoter were found in explanted primary normal prostate epithelial and stromal cells. Complete silencing and methylation of RASSF1A promoter was observed in five widely used prostate carcinoma cell lines, which acquired the ability to grow in culture spontaneously, including LNCaP, PC-3, ND-1, DU-145, 22Rv1, and one primary prostate carcinoma immortalized by overexpression of the human telomerase catalytic subunit (RC-58T/hTERT). However, no silencing of RASSF1A was found in four other prostate carcinoma cell lines, which were adapted for cell culture after transformation with human papillomaviral DNA. Suppression of cell growth in vitro was demonstrated after the reintroduction of RASSF1A-expressing construct into LNCaP prostate carcinoma cells. Our data implicate the RASSF1A gene in human prostate tumorigenesis.

Cell Division↗
PubMed2002

Retinol metabolism and lecithin:retinol acyltransferase levels are reduced in cultured human prostate cancer cells and tissue specimens.

Recent studies from our laboratory have indicated that the metabolism of vitamin A (retinol) to retinyl esters, carried out primarily by the enzyme lecithin:retinol acyltransferase (LRAT), is greatly reduced in human carcinoma cell lines of the oral cavity, skin, breast, and kidney as compared with their normal epithelial counterparts. These studies suggest that human carcinoma cells are retinoid-deficient relative to normal epithelial cells. In this study, we examined the metabolism of [(3)H]retinol and [(3)H]retinoic acid (RA) in human prostate cancer lines and in primary cultures of human prostate epithelial cells. Normal cells esterified all of the [(3)H]retinol added to the cultures. In contrast, all seven prostate cancer cell lines and four primary cultures derived from prostatic adenocarcinomas metabolized only trace amounts of [(3)H]retinol to [(3)H]retinyl esters. Correlated with this relative lack of esterification of [(3)H]retinol by the cancer cells was loss of expression of LRAT protein, whereas normal cells expressed abundant levels of LRAT protein by Western analysis. The metabolism of [(3)H]RA was also examined in these prostatic cells. Two of the prostate cancer tumor lines, DU 145 and PJ-1, exhibited rapid metabolism of [(3)H]RA; in contrast, the other tumor lines or primary cultures metabolized [(3)H]RA at a much slower rate. We also found that the immortalization of normal human prostatic epithelial cells by SV40 T antigen led to a reduction in LRAT protein expression and esterification of [(3)H]retinol. Further transformation to tumorigenicity with the ras oncogene resulted in loss of detectable LRAT expression. Finally, we analyzed LRAT protein expression in tissue sections from six prostatectomy specimens by immunohistochemistry. LRAT protein was predominantly expressed in the basal cells of normal prostatic epithelium, whereas its expression was lost in prostate cancer. Collectively, these data implicate aberrant retinoid metabolism in the process of prostatic carcinogenesis.

Acyltransferases↗
PubMed2002

Immortalization of normal adult human middle ear epithelial cells using a retrovirus containing the E6/E7 genes of human papillomavirus type 16.

A human middle ear epithelial cell line (HMEEC-1) was established using human papillomavirus E6/E7 genes. HMEEC-1 has remained morphologically and phenotypically stable, even after 50 passages. The cells are anchorage-dependent and nontumorigenic when injected into nude mice. This cell line thus provides a new tool for the study of normal cell biology and the pathological processes associated with the epithelial cells of the middle ear in otitis media. HMEEC-1 will also be useful in the search for new drugs and biological agents for the treatment of otitis media.

Adult↗
PubMed2002

Immortalization of rat eustachian tube epithelial cells by adenovirus 12-simian virus 40 hybrid virus.

The eustachian tube epithelial cells play an important role in the initial pathogenesis of otitis media. In order to study the role of the eustachian tube epithelial cells in the pathogenesis of otitis media, we have established a rat eustachian tube epithelial cell line. The cell line was derived by infecting primary cultures of eustachian tube epithelial cells with the adenovirus 12-simian virus 40 (Adl2-SV40) hybrid virus. The immortalized cells have retained the morphological characteristics of the parental cells and show positive staining with anti-cytokeratin antibodies (a marker for epithelial cells), but not with anti-vimentin antibodies (a fibroblast marker). The cells have been in continuous culture for more than 10 months and have undergone 38 passages. Western blotting and cell staining have confirmed the expression of the SV40 T antigen and p53. Chromosomal analysis indicates that the cell line is aneuploid and derived from male rat epithelial cells. Together, our results suggest that the cell line originated from eustachian tube epithelial cells from a male rat and was successfully immortalized by the Ad12-SV40 virus.

Adenoviridae↗
PubMed2002

Establishment and characterization of a schwannoma cell line from a patient with neurofibromatosis 2.

By using retroviral mediated gene transfer technique, a primary schwannoma culture from a 56-year-old Neurofibromatosis type 2 (NF2) patient was immortalized with HPV E6-E7 genes. This cell line, HEI193, has a unique splice site mutation of the NF2 gene. Both immunocytochemistry and molecular biology techniques were used to demonstrate that this cell line is of Schwann cell origin. Comparison of the primary tumor with HEI193 revealed the same NF2 mutation and an identical pattern of allele loss at multiple loci, indicating that the established cell line had maintained many of the properties of the original tumor. The immortalized cell line was non-tumorigenic in both severe combined immunodeficient (SCID) mice and nude mice, but has altered growth properties such as higher proliferation rate and independence of Schwann cell growth factors. To our knowledge, this is the first attempt to establish permanent cell lines from human NF2 patients. This Schwann tumor-derived cell line may provide a useful model system for the study of familial NF2 tumor pathogenesis, for elucidating NF2 functions and for testing new gene-based therapeutic approaches.

Alleles↗