Differentiation Of Retinoblastoma Cells Research Articles

MicroRNA-106b promotes the proliferation, migration and invasion of retinoblastoma cells by inhibiting the expression of ZBTB4 protein.

The present study investigated the function of microRNA (miR)-106b in the proliferation, migration and invasion of retinoblastoma (RB) cells, and aimed to elucidate the underlying mechanism. A total of 56 patients with RB were enrolled in the present study. The expression of miR-106b in RB tissues was measured by reverse transcription quantitative polymerase chain reaction. After transfection with miR-106b mimics or miR-106b inhibitor, a Cell-Counting kit-8 assay was used to determine the proliferation of WERI-Rb-1 cells and a Transwell assay was employed to measure the migration and invasion of the cells. Western blot analysis was performed to determine the expression of zinc finger and BTB domain containing 4 (ZBTB4) protein. By silencing or overexpression of ZBTB4 protein, the biological functions of ZBTB4 in WERI-Rb-1 cells were studied. A dual luciferase reporter assay was performed to test whether ZBTB4 was a target gene of miR-106b. The expression of miR-106b in RB tissues was elevated and closely associated with the severity of the disease. Overexpression of miR-106b increased but inhibition of miR-106b expression decreased the proliferation, migration and invasion abilities of WERI-Rb-1 cells. In addition, overexpression of miR-106b decreased but inhibition of miR-106b expression increased ZBTB4 protein expression in WERI-Rb-1 cells. Similarly, overexpression of ZBTB4 reduced but inhibition of ZBTB4 expression promoted the proliferation, migration and invasion of WERI-Rb-1 cells. Finally, miR-106b regulated the expression of ZBTB4 by binding to the 3′-untranslated region of the ZBTB4 gene. The present study demonstrated that increased expression of miR-106b in RB tissues is positively associated with the metastasis and differentiation of RB cells. As an oncogene, miR-106b promotes the proliferation, migration and invasion of WERI-Rb-1 cells by inhibiting the expression of ZBTB4 protein.

Correlations between claudin-1 and PIGF expressions in retinoblastoma.

To investigate the expressions of claudin-1 and placental growth factor (PlGF) proteins in retinoblastoma (RB) and their relationships with the differentiation of RB, the infiltration of optic nerve and choroid and clinical stages. Immunohistochemical (IHC) method was used to detect the expressions of claudin-1 and PlGF proteins in 56 cases of RB paraffin-embedded tissue samples. The x2-test and Fisher exact test were used to compare the qualitative variables. The Pearson correlation coefficient was used to detect the correlation of the expression of claudin-1 with that of PlGF in RB tissues. 1) Among RB tissues, the positive expression rates of claudin-1 in clinical stage I tumors and clinical stage III tumors were 69.2% and 38.9%, respectively, and claudin-1 was not expressed in all clinical stage II tumors (p=0.002). In case of optic nerve invasion, the lowly positive expression of claudin-1 was detected, and the difference was significant (p=0.001). 2) The positive expression rate of PlGF proteins in RB was 73.8%, which was higher in tumors with optic nerve invasion than in tumors without the invasion; the expression was significantly different (p=0.001). In addition, the positive expression rate of PlGF in tumors with choroidal invasion was 74.1%. 3) The expression of claudin-1 in RB was negatively correlated with the presence of choroidal invasion (r=0.52, p≤0.0001) and optic nerve infiltration (r=0.49, p=0.0003). There was a significant positive correlation between the expression of PlGF and the presence of optic nerve invasion (r=0.30, p=0.009). In addition, there was a significant positive correlation between the expression of claudin-1 and that of PlGF (r=0.41, p=0.006). The expression level of claudin-1 is negatively correlated with the differentiation of RB cells, optic nerve infiltration and clinical stages, while the expression of PlGF was positively correlated with the optic nerve infiltration and clinical stages of RB. The role of claudin-1 may be opposite to that of matrix metalloproteinase-2 (MMP-2) in the development of RB.

Anti-tumor effects of pigment epithelium-derived factor (PEDF): implication for cancer therapy. A mini-review.

Pigment epithelium-derived factor (PEDF) is a secreted glycoprotein and a non-inhibitory member of the serine protease inhibitor (serpin) family. It is widely expressed in human fetal and adult tissues but its expression decreases with age and in malignant tissues. The main anti-cancer activities of PEDF derive from its dual effects, either indirectly on the tumor microenvironment (indirect antitumor action) or directly on the tumor itself (direct antitumor influence). The indirect antitumor activities of PEDF were uncovered from the early findings that it stimulates retinoblastoma cell differentiation and that additionally it possesses anti-angiogenic, anti-tumorigenic and anti-metastatic properties. The mechanisms of its direct antitumor effect, however, have not been fully elucidated. This review highlights recent progress in our understanding of the multifunctional activities of PEDF and, in particular, its anti-cancer signaling mechanisms. Additionally, we discuss the possibility of using novel phosphaplatin compounds that can upregulate PEDF expression as a chemotherapy for cancer treatment.

Lithium release from β-tricalcium phosphate inducing cementogenic and osteogenic differentiation of both hPDLCs and hBMSCs.

It is accepted that the accelerated differentiation of tissue cells on bioactive materials is of great importance to regenerate the lost tissues. It was previously reported that lithium (Li) ions could enhance the in vitro proliferation and differentiation of retinoblastoma cells and endometrium epithelia by activating the Wnt canonical signalling pathway. It is interesting to incorporate Li ions into bioactive ceramics, such as β-tricalcium phosphate (Li-β-TCP), in order to stimulate both osteogenic and cementogenic differentiation of different stem cells for the regeneration of bone/periodontal tissues. Therefore, the aim of this study was to investigate the interactions of human periodontal ligament cells (hPDLCs) and human bone marrow stromal cells (hBMSCs) with Li-β-TCP bioceramic bulks and their ionic extracts, and further explore the osteogenic and cementogenic stimulation of Li-β-TCP bioceramics and the possible molecular mechanisms. The results showed that Li-β-TCP bioceramic disks supported the cell attachment and proliferation, and significantly enhanced bone/cementum-related gene expression, Wnt canonical signalling pathway activation for both hPDLCs and hBMSCs, compared to conventional β-TCP bioceramic disks without Li. The release of Li from Li-β-TCP powders could significantly promote the bone/cementum-related gene expression for both hPDLCs and hBMSCs compared to pure β-TCP extracts without Li release. Our results suggest that the combination of Li with β-TCP bioceramics may be a promising method to enhance bone/cementum regeneration as Li-β-TCP possesses excellent in vitro osteogenic and cementogenic stimulation properties by inducing bone/cementum-related gene expression in both hPDLCs and hBMSCs.

The effects of PEDF on cancer biology: mechanisms of action and therapeutic potential

The potent actions of pigment epithelium-derived factor (PEDF) on tumour-associated cells, and its extracellular localization and secretion, stimulated research on this multifunctional serpin. Such studies have identified several PEDF receptors and downstream signalling pathways. Known cellular PEDF responses have expanded from the initial discovery that PEDF induces retinoblastoma cell differentiation to its anti-angiogenic, antitumorigenic and antimetastatic properties. Although the diversity of PEDF activities seems to be complex, they are consistent with the varied mechanisms that regulate this multimodal factor. If PEDF is to be used for cancer management, a deeper appreciation of its many functions and mechanisms of action is needed.

Correction: Inhibitory Activity of Bevacizumab to Differentiation of Retinoblastoma Cells

The fifth author's affiliations are incomplete. Dong Hun Kim is affiliated with both 3 Department of Radiology, College of Medicine, Sooncheonhyang University, Bucheon, Korea, and 5 Department of Radiology, College of Medicine, Chosun University, Gwangju, Korea

Inhibitory Activity of Bevacizumab to Differentiation of Retinoblastoma Cells

Vascular endothelial growth factor (VEGF) is a major regulator in retinal and choroidal angiogenesis, which are common causes of blindness in all age groups. Recently anti-VEGF treatment using anti-VEGF antibody has revolutionarily improved the visual outcome in patients with vaso-proliferative retinopathies. Herein, we demonstrated that bevacizumab as an anti-VEGF antibody could inhibit differentiation of retinoblastoma cells without affection to cellular viability, which would be mediated via blockade of extracellular signal-regulated kinase (ERK) 1/2 activation. The retinoblastoma cells expressed VEGFR-2 as well as TrkA which is a neurotrophin receptor associated with differentiation of retinoblastoma cells. TrkA in retinoblastoma cells was activated with VEGF treatment. Interestingly even in the concentration of no cellular death, bevascizumab significantly attenuated the neurite formation of differentiated retinoblastoma cells, which was accompanied by inhibition of neurofilament and shank2 expression. Furthermore, bevacizumab inhibited differentiation of retinoblastoma cells by blockade of ERK 1/2 activation. Therefore, based on that the differentiated retinoblastoma cells are mostly photoreceptors, our results suggest that anti-VEGF therapies would affect to the maintenance or function of photoreceptors in mature retina.

Differentiation Potential of Human Retinoblastoma Cells

Retinoblastoma (RB), an intraocular tumor of childhood, contains small subpopulation(s) of stem-like cells expressing the ABCG2 drug transporter that can efflux standard chemotherapies. Since chemo-resistant stem-like cells appear to be a driving force in tumor progression and metastasis for a variety of cancers, innovative treatment strategies are necessary to eradicate these rare cell populations. Terminal differentiation, as a means to deplete the pool of stem-like cells in RB, is an intriguing approach to cancer therapeutics. However, the full extent of RB differentiation remains unknown. Differentiation of RB cells has been examined in response to a variety of different agents, including retinoic acid/sodium butyrate, Pigment Epithelial-Derived Factor, as well as Succinylated Concanavalin A. RB cells exhibit morphologic and phenotypic responses to these differentiating agents, although the permanence of these effects is questionable due to reversibility. Further study of differentiation programs may lead to new approaches in the design of strategies to combat the initiation and progression of RB in vivo.

PEDF - A Noninhibitory Serpin with Neurotrophic Activity

The pigment epithelium-derived factor (PEDF) is a glycoprotein with a molecular weight of 50 kDa belonging to the noninhibitory serpin family. It regulates several physiological processes, such as stimulation of retinoblastoma cell differentiation into neuron cells, and facilitation of the growth and viability of photoreceptor cells and neurons of the central nervous system. Moreover, this factor protects neuronal cells against apoptosis. PEDF is not only a neurotrophic factor, but also a natural angiogenesis inhibitor. This protein, as well as its biologically active fragments, possesses significant neuroprotective, neurotrophic, and antiangiogenic capabilities. The precise molecular mechanisms underpinning the effects of PEDF are still not quite clear. However, this protein generates great interest as a promising drug for the therapy of a wide range of neurodegenerative, ophthalmological, and oncological diseases. This review is a summary of what is known today about the structural features, biochemical properties, and multimodal functions of PEDF.

Differential Roles of Matrix Metalloproteinase-9 and -2, Depending on Proliferation or Differentiation of Retinoblastoma Cells

To investigate the differential roles of matrix metalloproteinase (MMP)-9 and MMP-2 in the proliferation or differentiation of retinoblastoma cells. Cell proliferation assay with an MMP-9 inhibitor and cell viability assay with an MMP-2 inhibitor were performed in retinoblastoma cells with 5 ng/mL fibroblast growth factor 2 for proliferation, 0.1% bovine serum albumin for differentiation, or reverse transcriptase-polymerase chain reaction (RT-PCR) for MMP-9, MMP-2, and their tissue inhibitors TIMP-1 and TIMP-2. Immunohistochemistry for MMP-2 and nm23 was performed using an experimental model of retinoblastoma. With the use of an MMP-2 inhibitor, Western blot analysis was performed for neurofilament, extracellular signal-regulated kinases 1 and 2 (ERK 1/2), and phospho-ERK 1/2, and neurite length was measured in differentiated retinoblastoma cells. With the proliferation of retinoblastoma cells, MMP-9 expression was upregulated without alteration of MMP-2, TIMP-1, or TIMP-2. However, proliferation was not affected by the inhibition of MMP-9 activity. Interestingly, only MMP-2 expression, colocalized with differentiated cells in retinoblastoma tissue, was significantly increased in the differentiation of retinoblastoma cells. Inhibition of MMP-2 activity did not affect cellular viability but attenuated neurite outgrowth and neurofilament expression of differentiated retinoblastoma cells, which was mediated through the suppression of ERK 1/2 activation. The authors suggest that differential expression of MMP-9 and -2 could reflect biological features, such as proliferation and differentiation, of retinoblastoma cells. In particular, MMP-2 could be directly involved in the regulation of differentiation of retinoblastoma cells. Therefore, therapeutic targeting to MMP-2 may prove useful for reducing malignancy through the differentiation of retinoblastoma cells.

PEDF inhibits growth of retinoblastoma by anti‐angiogenic activity

Pigment epithelium-derived factor (PEDF), an angiogenesis inhibitor with multiple other functions, balances angiogenesis in the eye and blocks tumor progression. Retinoblastoma, an angiogenesis-dependent tumor, is the most common ocular cancer in children without effective treatment. It has been reported that PEDF can induce neuronal differentiation of retinoblastoma cells; however, its anti-angiogenic potential for inhibition of retinoblastoma growth in vivo has not been elucidated. The present study was designed to investigate the effect of PEDF on growth of retinoblastoma and the possible molecular mechanism. Soluble and non-fusion recombinant PEDF were generated in E. coli. Recombinant PEDF dose-dependently inhibited proliferation and induced apoptosis of endothelial cells. PEDF had no effects on the proliferation and apoptosis of retinoblastoma cell line SO-Rb50. Intraperitoneal injection of PEDF resulted in growth inhibition of heterotopic retinoblastoma xenografts at 68.78%. MVD in tumor tissues treated with PEDF was significantly decreased. These results suggested that PEDF suppressed tumor growth by blocking angiogenesis instead of a direct cytotoxic effect on tumor cells. Vascular endothelial growth factor (VEGF), a major angiogenic stimulator, was down-regulated by PEDF in both SO-Rb50 cells and retinoblastoma xenografts. Hypoxia-inducible factor (HIF)-1alpha, a crucial transcriptional factor for VEGF expression, was also down-regulated by PEDF both in vitro and in vivo. PEDF reduced HIF-1alpha nuclear translocation, which may be responsible for the down-regulation of VEGF. Down-regulation of VEGF expression in tumor cells through inhibiting HIF-1alpha, thus attenuating the paracrine effect of VEGF on endothelial cell proliferation and vascular permeability in tumor tissues, may represent a mechanism for the anti-angiogenic activity of PEDF.

Neurotrophin Receptors TrkA and TrkB in Retinoblastoma Are Differentially Expressed Depending on Cellular Differentiation

Retinoblastoma is the most common primary intraocular malignancy in children. With the progression of retinoblastoma, retinoblastoma cells lose their ability to differentiate. Regardless of many attempts to identify prognostic factors in retinoblastoma, further investigation for prognostic factors of retinoblastoma progression is still required because of the lack of sensitivity and specificity of these prognostic factors in predicting disease progression. We demonstrated that the differential expression of the neurotrophin receptors TrkA and TrkB is closely related to the differentiation of retinoblastoma cells. While retinoblastoma cells expressed TrkA as well as TrkB, their growth rates were not influenced by the addition of nerve growth factor to the culture medium. In experimental animal models of retinoblastoma, TrkA expression was primarily detected in more differentiated areas with high nm23 immunoreactivity whereas TrkB expression was apparent in more proliferative areas with high Ki67 immunoreactivity. With retinoic-acid-induced differentiation of retinoblastoma cells, TrkA expression significantly increased whereas TrkB significantly decreased. The differential expression of TrkA and TrkB with differentiation of retinoblastoma cells was mediated by extracellular-signal-regulated kinase 1/2 activation, which was confirmed by immunocytochemistry of TrkA. Therefore, our results suggest that the differential expression of TrkA and TrkB could be valuable as a therapeutic target, for instance using specific inhibitors.

Activin Inhibits Cell Growth and Induces Differentiation in Human Retinoblastoma Y79 Cells

Purpose: Activin is a member of the transforming growth factor-beta (TGF-β) superfamily and exerts certain effects on differentiation and apoptosis. We investigated the effects of activin on retinoblastoma cell line. Materials and Methods: We used retinoblastoma cell line Y79. Intracellular signal transduction of activin was investigated with RT-PCR, immunofluorescence study, and luciferase reporter assay. The effect of activin on cell growth was examined with fluorescence cell viability assays. To determine the effect of activin on apoptosis, a TUNEL assay and an immunofluorescence study of cleaved PARP were performed. The effect of activin on cell differentiation was examined with RT-PCR and Western blotting. Results: Intracellular signal transduction of activin was confirmed in Y79 cells. Activin inhibited Y79 cell growth. Activin induced the expression of neural retina leucine zipper (Nrl) at the mRNA and protein levels. Conclusions: Nrl is a specific gene in rod photoreceptor development and is a gene indispensable to differentiation into rod photoreceptors, so the present results suggest that activin affects the differentiation of retinoblastoma cells into rod photoreceptor cells.

Antitumor Activity of Arsenic Trioxide on Retinoblastoma: Cell Differentiation and Apoptosis Depending on Arsenic Trioxide Concentration

Arsenic trioxide (ATO) targets multiple pathways in malignant cells, resulting in the promotion of differentiation or in the induction of apoptosis. The antitumor activity of ATO on retinoblastoma was investigated. Human retinoblastoma cells were incubated with various ATO concentrations. The antiproliferative effect of ATO was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and the effect of ATO on cell-cycle progression was validated by flow cytometry. At a low concentration, the ATO-induced differentiation of retinoblastoma cells was evaluated by neurofilament expression and extracellular signal-regulated kinase (ERK)1/2 activation, which was confirmed by the inhibition of ERK1/2. At a high concentration, ATO-induced H(2)O(2) production was investigated with the cell-permeable fluorescent dye 2'7'-dichlorofluorescein-diacetate, and the relationship of ATO-induced H(2)O(2) production with caspase-3-dependent apoptosis was validated by Western blot and 4'6-diamidino-2-phenolindole staining, which was confirmed by reactive oxygen species (ROS) inhibition. The effect of ATO on tumor formation was assessed with an orthotopic animal model of retinoblastoma. The antitumor activity of ATO in retinoblastoma was related to two main mechanisms, differentiation and apoptosis, which were determined by the level of ATO. At a low dose (<or= 1 microM), ATO induced the differentiation of retinoblastoma cells through ERK1/2 activation, whereas ROS generation by a high dose (>or= 2 microM) of ATO induced apoptosis in retinoblastoma cells. Moreover, ATO at low and high doses effectively inhibited tumor formation. These results suggest that ATO can be used as an effective alternative therapeutic for the treatment of retinoblastoma.

Inhibition of Nuclear Translocation of Apoptosis-Inducing Factor Is an Essential Mechanism of the Neuroprotective Activity of Pigment Epithelium-Derived Factor in a Rat Model of Retinal Degeneration

Photoreceptor apoptosis is a critical process of retinal degeneration in retinitis pigmentosa (RP), a group of retinal degenerative diseases that result from rod and cone photoreceptor cell death and represent a major cause of adult blindness. We previously demonstrated the efficient prevention of photoreceptor apoptosis by intraocular gene transfer of pigment epithelium-derived factor (PEDF) in animal models of RP; however, the underlying mechanism of the neuroprotective activity of PEDF remains elusive. In this study, we show that an apoptosis-inducing factor (AIF)-related pathway is an essential target of PEDF-mediated neuroprotection. PEDF rescued serum starvation-induced apoptosis, which is mediated by AIF but not by caspases, of R28 cells derived from the rat retina by preventing translocation of AIF into the nucleus. Nuclear translocation of AIF was also observed in the apoptotic photoreceptors of Royal College of Surgeons rats, a well-known animal model of RP that carries a mutation of the Mertk gene. Lentivirus-mediated retinal gene transfer of PEDF prevented the nuclear translocation of AIF in vivo, resulting in the inhibition of the apoptotic loss of their photoreceptors in association with up-regulated Bcl-2 expression, which mediates the mitochondrial release of AIF. These findings clearly demonstrate that AIF is an essential executioner of photoreceptor apoptosis in inherited retinal degeneration and provide a therapeutic rationale for PEDF-mediated neuroprotective gene therapy for individuals with RP.

Growth inhibition and differentiation of retinoblastoma cells induced by all trans retinoic acid

To investigate the inhibition of proliferation of retinoblastoma cells (SO-Rb50) and the differentiation of tumor cells into normal cell types by all trans retinoic acid (ATRA). This was an experimental study. SO-Rb 50 cells were treated with ATRA at different concentrations and growth curves were plotted. IC50 was analyzed by MTI method. Cell cycles before and after drug treatment were analyzed using flow cytometry. Morphologic characters of living cells and hematoxylin and eosin (HE) stained cells were observed microscopically before and 20 days after drug treatment The cell markers of NSE, vimentin, GFAP were detected with immunohistochemical method before and 20 days after drug treatment. Cell growth was inhibited by ATRA treatment dose-dependently from 0 to 1 x 10(-5) mol/L and the cell viability was decreased. The IC50 of ATRA measured by MTT was approximately 14.05 microm/L. Treated with 1 x 10(-5) mol/L ATRA, the G0/G1 stage cells increased from 56.5% before treatment to 66.6%-81.0% after treatment, whilst S stage cells decreased from 33.1% to 22.3%-15.9%. Cells could attach to poly-lysine coated cover slides and formed small colonies. Some cells changed from round in shape to oval or fusiform shapes. The ratio of nuclear to cytoplasm decreased after the treatment of ATRA. Some cells extended long or short processes 20 days after treatment. Immunohistochemical studies showed that cells were stained moderately positive, strongly positive and weakly positive by NSE, vimentin and GFAP antibodies, respectively before treatment. After the ATRA treatment, cells were stained strongly positive, strongly positive and moderately positive by NSE, vimentin and GFAP antibodies, respectively. ATRA significantly inhibits the proliferation of SO-Rb 50 cells and cells arrested at G0/G1 stage of cell cycle. ATRA induces differentiation of retinoblastoma cells towards the neurons and gliocytes.

Heparin Binding Induces a Conformational Change in Pigment Epithelium-derived Factor

Pigment epithelium-derived factor (PEDF) is a noninhibitory serpin found in plasma and in the extracellular space. The protein is involved in different biological processes including cell differentiation and survival. In addition, it is a potent inhibitor of angiogenesis. The function is likely associated with binding to cell surface receptors in a heparin-dependent way (Alberdi, E. M., Weldon, J. E., and Becerra, S. P. (2003) BMC Biochem. 4, 1). We have investigated the structural basis for this observation and show that heparin induces a conformational change in the vicinity of Lys(178). This structural change was evident both when binding to intact heparin and specific heparin-derived oligosaccharides at physiological conditions or simply when exposing PEDF to low ionic strength. Binding to other glycosaminoglycans, heparin-derived oligosaccharides smaller than hexadecasaccharides (dp16), or type I collagen did not affect the structure of PEDF. The conformational change is likely to expose the epitope involved in binding to the receptor and thus regulates the interactions with cell surface receptors.

Pigment Epithelium-Derived Factor Delays the Death of Photoreceptors in Mouse Models of Inherited Retinal Degenerations

Pigment epithelium-derived factor (PEDF) is a member of the serine protease inhibitor superfamily produced by retinal pigment epithelial cells in the developing and adult retina. In vitro, it induces neuronal differentiation of retinoblastoma cells and promotes survival of cerebellar granule neurons. The pedf gene is closely linked to an autosomal-dominant locus for retinitis pigmentosa, suggesting that PEDF could be a survival factor for photoreceptors. We have investigated this possibility by injecting PEDF into the eyes of homozygous retinal degeneration (rd) and retinal degeneration slow (rds) mice, two mutants displaying apoptotic photoreceptor loss. This procedure resulted in a transient delay of photoreceptor loss in the rd mouse and a reduction in apoptotic photoreceptor profiles in the rds mouse. We conclude that PEDF can act as a survival-promoting factor for photoreceptors in vivo and could potentially be useful for the treatment of photoreceptor diseases.

Effects of medium conditioned by retinal pigmented epithelial cells on neurotransmitter phenotype in retinoblastoma cells

We previously reported that medium conditioned by retinal pigmented epithelial cells can induced cellular differentiation in human retinoblastoma cells. Extensive neurite outgrowth, increased expression of neuronal marker molecules and decreased expression of glial marker molecules are characteristic of the differentiated phenotype. In the studies described here, we examine whether modulations in the expression of potential neurotransmitter molecules, catecholamines and indolealkyl amines, might be associated with the differentiation of retinoblastoma cells. Concentrations of serotonin, 5-hydroxyindoleacetic acid, 3-methoxytyrosine, homovanillic acid, and 3-methoxy-4-hydroxyphenylacetic acid in extracts of differentiated and undifferentiated retinoblastoma cells were assessed by HPLC. The results show that serotonin and its metabolite, 5-hydroxyindoleacetic acid, are characteristically present in undifferentiated cells. Dopa metabolites, 3-methoxytyrosine, homovanillic acid and 3-methoxy-4-hydroxyphenylacetic acid, are uniquely present in differentiated cells. It appears that differentiation of retinoblastoma cells induced by factors secreted by retinal pigmented epithelial cells involves a switch from a serotonergic phenotype to one dominated by metabolites of dopa. These findings may provide clues about the factors that control retinoblastoma cells and metastasis.

Laminin-induced retinoblastoma cell differentiation: possible involvement of a 100-kDa cell-surface laminin-binding protein.

Gene and protein expression of Y-79 retinoblastoma cells growing on poly(D-lysine) is switched from a photoreceptor-like to a conventional neuron-like pathway by the basement membrane glycoprotein laminin. Unlike other cell systems where laminin influences differentiation, Y-79 cells can neither attach to nor chemotactically respond to laminin. However, laminin increases attachment to poly(D-lysine). The laminin effects therefore seem to occur via an adhesion- and chemotaxis-independent mechanism. Moreover, these tumor cells do not exhibit high-affinity laminin binding, having only a single binding site of intermediate affinity. Laminin-Sepharose affinity chromatography of Y-79 cell surface proteins labeled with 125I revealed a single major radiolabeled 100-kDa protein eluted by 20 mM EDTA, with an electrophoretic behavior different from that of integrins. No other proteins were eluted under more stringent conditions. This material, which we call LBM-100 (100-kDa laminin-binding molecule), may be a "differentiative" laminin-binding protein through which laminin influences gene expression and development independently of attachment.