Viability effects on cell cycle synchronization of different prostate cancer cell lines: A brief report

Aim: Serum deprivation is often used to synchronize cells in G0/G1 phase for comparative in vitro studies. Here we aimed to investigate the effect on viability of the three commonly used prostate cancer-cell lines PC3, LNCaP and DU145 after serum deprivation. Methods: The cell lines were cultured in standard culture medium (controls) and under serum deprivation for 48 and 72 h. Then the proportion of cells in G0/G1 phase was analysed by flow cytometry and cell morphology was microscopically investigated. The cells were allowed to recover for three days in standard culture condition before cell viability (using MTT assay) was analysed. Results: In comparison to control cells, significant effects on cell cycle arrest in G0/G1 phase were noted for DU145 and PC3 cells and the cell morphology was negatively affected in a time-dependent manner. These parameters were unaffected in LNCaP cells. After three days of recovery, the viability of DU145 and PC3 cells was significantly reduced compared to LNCaP. Conclusions: Serum deprivation showed different effects on the prostate cancer cells, probably due to differences in growth rate. Such effects on viability should be considered as an obstacle for comparative studies

Expression and function of the protein tyrosine phosphatases SHP-1 and SHP-2 in prostate cancer

Prostate cancer is one of the most frequent cancers in males in Western countries. Radical prostatectomy serves as the first line of treatment for patients with localised prostate cancer. However, in many cases the cancer will recur. It is currently difficult to identify those patients at a high risk of tumour recurrence. Therefore, molecular markers associated with and predicting disease recurrence are needed to help identify patients and to better understand the signalling pathways in prostate cancer cells. The protein tyrosine kinase (PTK) receptors and cytosolic signalling proteins as well as the protein tyrosine phosphatases (PTPs) have important roles in the regulation of growth of the benign and malignant prostate gland. Here, we have studied the expression and function of the PTP SHP-1 in prostate cancer cell lines and in clinical material from patients that underwent radical prostatectomy. We have also analysed the expression of SHP-2 in prostate cancer cell lines and human prostate cancer tissue. We performed several different in vitro and in vivo experiments that were evaluated with methods such as immunohistochemical staining of tissue micro arrays (TMA), polymerase chain reaction (PCR), Western blot, eukaryotic cell transfections, proliferation assays, PTP enzymatic activity assay, caspase assay, flow cytometric assays, cell growth in vivo in nude mice, migration and invasion assays. Our results show that a low or absent SHP-1 expression might serve as a marker to identify prostate cancer patients at an increased risk of tumour recurrence. We also demonstrate that proliferation and apoptosis can be regulated by modulations of the SHP-1 expression in prostate cancer cells. SHP-1 expression and activity is reduced in the LNCaP cell line that was long-term stimulated with IL-6, showing that the cytokine might regulate SHP-1 activity. SHP-1 is also able to reduce growth of PC3 cells growing in vivo in nude mice. These results suggest that SHP-1 is a negative modulator by inhibiting prostate cancer cell proliferation through the growth factor and cytokine-induced pathways in prostate cancer cells. Our results also suggest that SHP-2 may have similar activity in prostate cancer

In vitro cytotoxicity evaluation of HDAC inhibitor Apicidin in pancreatic carcinoma cells subsequent time and dose dependent treatment.

Apicidin is a potent histone deacetylase inhibitor (HDACI) that selectively binds to histone deacetylases (HDACs) class I and interferes with the deacetylation process, which results in modification of acetylation level of cellular proteins. The aim of the study was to investigate the potential time and dose dependent cytotoxicity of the test compound, Apicidin, in pancreatic cancer cells Capan-1 and Panc-1 as well as estimate maximal tolerable dose (MTD) of the test agent and determine EC50 using four complementary colorimetric cytotoxicity or viability assays. The cells were treated with increasing concentrations of Apicidin (0-5000nM) for 2, 4 and 6h (short term exposure) or 24, 48 and 72h (long term exposure) before conducting cytotoxic analyses with lactate dehydrogenase assay or viability analyses with sulforhodamine B (SRB), methyl tetrazolium (MTT) and crystal violet (CV) assays. In order to investigate whether Apicidin irreversibly affects the cells already during the short term exposure, the medium containing Apicidin was removed and replaced with fresh culturing medium after 6h of treatment. The cells were then incubated for additional 24, 48 or 72h before carrying out the analysis. The results obtained from cytotoxicity and viability assays indicated, that Apicidin was well tolerated by both cell lines at concentrations below 100nM at any given time point and at all applied concentrations during the short term (6h or less) treatment. Continuous prolonged term exposures (48h or greater) of the cells to Apicidin with concentration exceeding 100nM resulted in significantly increasing cytotoxicity and sustained significant loss of cell viability. Moreover, long term exposure of pancreatic cancer cells Capan-1 and Panc-1 to Apicidin concentrations exceeding 100nM showed an initial anti-proliferative effect before cytotoxicity onset. In summary, MTD was exposure time dependent and estimated to 100nM for long term treatment and to at least 5000nM for treatment not greater than 6h. EC50 concentration of Apicidin was established after long term treatment, however with some variation when comparing the different assays and cell lines. Results from this study may encourage reinvestigating the capacity of potent HDACI Apicidin as an attractive agent for interfering with the deacetylation process catalyzed by HDACs for potential pancreatic cancer intervention

Miljö, medicin och undervisning, hur hänger det ihop? : exempel från projektet mangan och prostatacancer

Forskningsmiljön ”Man and Biosphere Health” är engruppering där forskare från helt olika biologiskakunskapsområden träffas och knyter kontakter. Forskning inom området ”Life Science” (Livsvetenskap), som framförallt innefattar biologi, medicin och biokemi, är världens största tvärdisciplinära forskningsområde med studier av biologisktliv samt de förutsättningar som utgör grunden för fortsatt liv. Unikt för samarbetet inom MABH är kombinationen avekologisk och biomedicinsk kompetens, vilket i vårt fall har inneburit att cellbiologisk forskning har knutits ihop med miljöforskning på ett nyskapande sätt

Miljö, medicin och undervisning, hur hänger det ihop? : exempel från projektet mangan och prostatacancer

Forskningsmiljön ”Man and Biosphere Health” är engruppering där forskare från helt olika biologiskakunskapsområden träffas och knyter kontakter. Forskning inom området ”Life Science” (Livsvetenskap), som framförallt innefattar biologi, medicin och biokemi, är världens största tvärdisciplinära forskningsområde med studier av biologisktliv samt de förutsättningar som utgör grunden för fortsatt liv. Unikt för samarbetet inom MABH är kombinationen avekologisk och biomedicinsk kompetens, vilket i vårt fall har inneburit att cellbiologisk forskning har knutits ihop med miljöforskning på ett nyskapande sätt

Role of protein tyrosine phosphatase SHP-1 in interleukin-6 regulation of prostate cancer

Background Interleukin-6 (IL-6) is a multifunctional cytokine that has been implicated in the modulation of growth and progression of prostate cancer. Decreased expression of the tyrosine phosphatase SHP-1, involved in regulation of cytokine and tyrosine kinase receptor signaling, has been shown to be associated with less favourable outcome for prostate cancer patients. Methods Parental LNCaP and LNCaP-IL6+ subline, derived from parental LNCaP cells by continuous culture of the cells in the presence of recombinant IL-6 were used in the study. Expression of SHP-I, PTEN, STAT3, pSTAT3, ERK, pERK, AKT, pAKT, PTEN and SHP-1 was analysed by immunohistochemistry, Western blot, cDNA microarray quantitative PCR and reverse transcriptase PCR. Proliferation and apoptosis of transfected cells was analysed with caspase3/7 and flow cytometry. Results Phosphorylation of ERK and STAT3 was increased in the LNCaP-IL6+ subline compared to LNCaP cells, whereas pAkt was decresaed. This could be due tore-expression of PTEN in LNCaP-IL6+ cells. Overexpression and inhibition experiments with SHP-1 siRNA showed that SHP-1 reduced proliferation and increased apoptosis in both cell lines. A cDNA analysis revealed 80 up-regulated and 87 down-regulated SHP-1 related genes in the LNCaP-IL6+ cell line compared to LNCaP cells. Conclusions SHP-1 suppresses growth and increases apoptosis in both LNCaP and LNCaP-IL6+ cells which suggests that SHP-1 could provide a therapeutic target for the prostate cancer cells even in the case that have gained an IL-6 related growth advantage

Recent Advances in Molecularly Imprinted Polymers and Their Disease-Related Applications

Molecularly imprinted polymers (MIPs) and the imprinting technique provide polymeric material with recognition elements similar to natural antibodies. The template of choice (i.e., the antigen) can be almost any type of smaller or larger molecule, protein, or even tissue. There are various formats of MIPs developed for different medical purposes, such as targeting, imaging, assay diagnostics, and biomarker detection. Biologically applied MIPs are widely used and currently developed for medical applications, and targeting the antigen with MIPs can also help in personalized medicine. The synthetic recognition sites of the MIPs can be tailor-made to function as analytics, diagnostics, and drug delivery systems. This review will cover the promising clinical applications of different MIP systems recently developed for disease diagnosis and treatment

Role of the ProteinTyrosine Phosphatase SHP-I in Interleukin-6 Regulation of Prostate Cancer Cells

BACKGROUND. Interleukin-6 (IL-6) is a multifunctional cytokine that has been implicated in the modulation of growth and progression of prostate cancer. Decreased expression of the tyrosine phosphatase SHP-1, involved in regulation of cytokine and tyrosine kinase receptor signaling, has been shown to be associated with less favorable outcome among prostate cancer patients. METHODS. Parental LNCaP cells and an LNCaP-IL6+subline, derived from parental LNCaP cells by continuous culture of the cells in the presence of recombinant IL-6 were used in the study. Expression of STAT3, pSTAT3, ERK, pERK, AKT, pAKT, PTEN, and SHP-1 was analyzed by immunohistochemistry, Western blots, cDNA microarray, quantitative PCRs, and reverse transcriptase PCRs. Proliferation and apoptosis of transfected cells were analyzed by caspase3/7 assay and flow cytometry. RESULTS. Phosphorylation of ERK and STAT3 was increased in the LNCaP-IL6+ subline compared with LNCaP cells, whereas pAKT was decreased. Overexpression and inhibition experiments with SHP-1 siRNA showed that SHP-1 reduced proliferation and increased apoptosis in both cell lines. Microarray analysis revealed 80 up-regulated and 87 down-regulated SHP-1-related genes in the LNCaP-IL6+ cell line compared with LNCaP cells. CONCLUSIONS. SHP-1 suppresses growth and increases apoptosis in both LNCaP and LNCaP-IL6+ cells, which suggests that SHP-1 could be a therapeutic target in prostate cancer, even when there is an IL-6-related growth advantage. Prostate 70: 1491-1500, 2010. (C) 2010 Wiley-Liss, Inc