Publications by authors named "Pirkko Härkönen"

76 Publications

Critical evaluation of the subcutaneous engraftments of hormone naïve primary prostate cancer.

Transl Androl Urol 2020 Jun;9(3):1120-1134

Institute of Biomedicine, University of Turku, Turku, Finland.

Background: Patient-derived xenografts (PDXs) are considered to better recapitulate the histopathological and molecular heterogeneity of human cancer than other preclinical models. Despite technological advances, PDX models from hormone naïve primary prostate cancer are scarce. We performed a detailed analysis of PDX methodology using a robust subcutaneous model and fresh tissues from patients with primary hormone naïve prostate cancer.

Methods: Clinical prostate tumor specimens (n=26, Gleason score 6-10) were collected from robotic-assisted laparoscopic radical prostatectomies at Turku University Hospital (Turku, Finland), cut into pieces, and implanted subcutaneously into 84 immunodeficient mice. Engraftments and the adjacent material from prostatic surgical specimens were compared using histology, immunohistochemistry and DNA sequencing.

Results: The probability of a successful engraftment correlated with the presence of carcinoma in the implanted tissue. Tumor take rate was 41%. Surprisingly, mouse hormone supplementation inhibited tumor take rate, whereas the degree of mouse immunodeficiency did not have an effect. Histologically, the engrafted tumors closely mimicked their parental tumors, and the Gleason grades and copy number variants of the engraftments were similar to those of their primary tumors. Expression levels of androgen receptor, prostate-specific antigen, and keratins were retained in engraftments, and a detailed genomic analysis revealed high fidelity of the engraftments with their corresponding primary tumors. However, in the second or third passage of tumors, the carcinoma areas were almost completely replaced by benign tissue with frequent degenerative or metaplastic changes.

Conclusions: Subcutaneous primary prostate engraftments preserve the phenotypic and genotypic landscape. Thus, they serve a potential model for personalized medicine and preclinical research but their use may be limited to the first passage.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.21037/tau.2020.03.38DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7354344PMC
June 2020

Dovitinib dilactic acid reduces tumor growth and tumor-induced bone changes in an experimental breast cancer bone growth model.

J Bone Oncol 2019 Jun 19;16:100232. Epub 2019 Mar 19.

University of Turku, Kiinamyllynkatu 10, 20520 Turku, Finland.

Advanced breast cancer has a high incidence of bone metastases. In bone, breast cancer cells induce osteolytic or mixed bone lesions by inducing an imbalance in bone formation and resorption. Activated fibroblast growth factor receptors (FGFRs) are important in regulation of tumor growth and bone remodeling. In this study we used FGFR1 and FGFR2 gene amplifications containing human MFM223 breast cancer cells in an experimental xenograft model of breast cancer bone growth using intratibial inoculation technique. This model mimics bone metastases in breast cancer patients. The effects of an FGFR inhibitor, dovitinib dilactic acid (TKI258) on tumor growth and tumor-induced bone changes were evaluated. Cancer-induced bone lesions were smaller in dovitinib-treated mice as evaluated by X-ray imaging. Peripheral quantitative computed tomography imaging showed higher total and cortical bone mineral content and cortical bone mineral density in dovitinib-treated mice, suggesting better preserved bone mass. CatWalk gait analysis indicated that dovitinib-treated mice experienced less cancer-induced bone pain in the tumor-bearing leg. A trend towards decreased tumor growth and metabolic activity was observed in dovitinib-treated mice quantified by positron emission tomography imaging with 2-[F]fluoro-2-deoxy-D-glucose at the endpoint. We conclude that dovitinib treatment decreased tumor burden, cancer-induced changes in bone, and bone pain. The results suggest that targeting FGFRs could be beneficial in breast cancer patients with bone metastases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jbo.2019.100232DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6434100PMC
June 2019

Alendronate-induced disruption of actin cytoskeleton and inhibition of migration/invasion are associated with cofilin downregulation in PC-3 prostate cancer cells.

Oncotarget 2018 Aug 24;9(66):32593-32608. Epub 2018 Aug 24.

University of Turku, Institute of Biomedicine, FI-20520 Turku, Finland.

Bisphosphonates are used for prevention of osteoporosis and metastatic bone diseases. Anti-invasive effects on various cancer cells have also been reported, but the mechanisms involved are not well-understood. We investigated the effects of the nitrogen-containing bisphosphonate alendronate (ALN) on the regulation of actin cytoskeleton in PC-3 cells. We analyzed the ALN effect on the organization and the dynamics of actin, and on the cytoskeleton-related regulatory proteins cofilin, p21-associated kinase 2 (PAK2), paxillin and focal adhesion kinase. Immunostainings of cofilin in ALN-treated PC-3 cells and xenografts were performed, and the role of cofilin in ALN-regulated F-actin organization and migration/invasion in PC-3 cells was analyzed using cofilin knockdown and transfection. We demonstrate that disrupted F-actin organization and decreased cell motility in ALN-treated PC-3 cells were associated with decreased levels of total and phosphorylated cofilin. PAK2 levels were also lowered but adhesion-related proteins were not altered. The knockdown of cofilin similarly impaired F-actin organization and decreased invasion of PC-3 cells, whereas in the cells transfected with a cofilin expressing vector, ALN treatment did not decrease cellular cofilin levels and migration as in mock transfected cells. ALN also reduced immunohistochemical staining of cofilin in PC-3 xenografts. Our results suggest that reduction of cofilin has an important role in ALN-induced disruption of the actin cytoskeleton and inhibition of the PC-3 cell motility and invasion. These data also support the idea that the nitrogen-containing bisphosphonates could be efficacious in inhibition of prostate cancer invasion and metastasis, if delivered in a pharmacological formulation accessible to the tumors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.25961DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6135693PMC
August 2018

SERMs Promote Anti-Inflammatory Signaling and Phenotype of CD14+ Cells.

Inflammation 2018 Aug;41(4):1157-1171

Institute of Biomedicine, University of Turku, Turku, Finland.

Signaling via estrogen receptors (ER) is recognized as an essential part of the immune regulation, and ER-mediated signaling is involved in autoimmune reactions. Especially ERα activation in immune cells has been suggested to skew cytokine production toward Th2/M2-type mediators, which can have protective effect on inflammatory diseases and reduce Th1 and Th17 responses. These effects are caused by increased alternative activation of macrophages and changes in the activation of different T cell populations. In humans, hormonal status has been shown to have a major impact on several inflammatory diseases. Selective estrogen receptor modulators (SERMs) are ER ligands that regulate ER actions in a tissue-specific manner mostly lacking the adverse effects of steroid hormones. The impact of SERMs on the immune system is less studied, but it is suggested that certain SERMs may also produce immunoprotective effects. Here, we show that two novel SERMs and raloxifene affect immune cells by promoting M2 macrophage phenotype, alleviating NFκB activity, inhibiting T cell proliferation, and stimulating the production of anti-inflammatory compounds such as IL10 and IL1 receptor antagonist. Thus, these compounds have high potency as drug candidates against autoimmune diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10753-018-0763-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6061028PMC
August 2018

Oncolytic alphavirus SFV-VA7 efficiently eradicates subcutaneous and orthotopic human prostate tumours in mice.

Br J Cancer 2017 Jun 30;117(1):51-55. Epub 2017 May 30.

Department of Biotechnology and Molecular Medicine, A. I. Virtanen Institute for Molecular Sciences, University of Eastern Finland, Kuopio 70211, Finland.

Background: Despite recent therapeutic and diagnostic advances, prostate cancer remains the second leading cause of cancer-related deaths among men in the Western world. Oncolytic viruses that replicate selectively in tumour cells represent a novel treatment candidate for these malignancies.

Methods: We analysed infectivity of avirulent Semliki Firest virus SFV-VA7 in human prostate cancer cell lines VCaP, LNCaP and 22Rv1 and in nonmalignant prostate epithelial cell line RWPE-1. Therapeutic potency of SFV-VA7 was evaluated in subcutaneous and orthotopic mouse LNCaP xenograft models.

Results: SFV-VA7 infected and killed the tested human prostate cancer cell lines irrespective of their hormone response status, while the nonmalignant prostate epithelial cell line RWPE-1 proved highly virus resistant. Notably, a single peritoneal dose of SFV-VA7 was sufficient to eradicate all subcutaneous and orthotopic LNCaP tumours.

Conclusions: Our results indicate that SFV-VA7 is a novel, promising therapeutic virus against prostate cancer warranting further testing in early clinical trials.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/bjc.2017.151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5520213PMC
June 2017

Breast cancer in neurofibromatosis type 1: overrepresentation of unfavourable prognostic factors.

Br J Cancer 2017 Jan 8;116(2):211-217. Epub 2016 Dec 8.

Department of Cell Biology and Anatomy, University of Turku, Kiinamyllynkatu 10, 20520 Turku, Finland.

Background: An increased breast cancer incidence and poor survival have been reported for women with neurofibromatosis 1 (NF1). To explain the poor survival, we aimed to link the histopathology and clinical characteristics of NF1-associated breast cancers.

Methods: The Finnish Cancer Registry and the Finnish NF Registry were cross-referenced to identify the NF1 patients with breast cancer. Archival NF1 breast cancer specimens were retrieved for histopathological typing and compared with matched controls.

Results: A total of 32 breast cancers were diagnosed in 1404 NF1 patients during the follow-up. Women with NF1 had an estimated lifetime risk of 18.0% for breast cancer, and this is nearly two-fold compared with that of the general Finnish female population (9.74%). The 26 successfully retrieved archival NF1 breast tumours were more often associated with unfavourable prognostic factors, such as oestrogen and progesterone receptor negativity and HER2 amplification. However, survival was worse in the NF1 group (P=0.053) even when compared with the control group matched for age, diagnosis year, gender and oestrogen receptor status. Scrutiny of The Cancer Genome Atlas data set showed that NF1 mutations and deletions were associated with similar characteristics in the breast cancers of the general population.

Conclusions: These results emphasise the role of the NF1 gene in the pathogenesis of breast cancer and a need for active follow-up for breast cancer in women with NF1.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/bjc.2016.403DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5243991PMC
January 2017

Toll-like receptor 9 expression is associated with breast cancer sensitivity to the growth inhibitory effects of bisphosphonates in vitro and in vivo.

Oncotarget 2016 Dec;7(52):87373-87389

Department of Chemistry, University of Alabama at Birmingham, Birmingham, AL, U.S.A.

Bisphosphonates are standard treatments for bone metastases. When given in the adjuvant setting, they reduce breast cancer mortality and recurrence in bone but only among post-menopausal patients. Optimal drug use would require biomarker-based patient selection. Such biomarkers are not yet in clinical use. Based on the similarities in inflammatory responses to bisphosphonates and Toll-like receptor (TLR) agonists, we hypothesized that TLR9 expression may affect bisphosphonate responses in cells. We compared bisphosphonate effects in breast cancer cell lines with low or high TLR9 expression. We discovered that cells with decreased TLR9 expression are significantly more sensitive to the growth-inhibitory effects of bisphosphonates in vitro and in vivo. Furthermore, cancer growth-promoting effects seen with some bisphosphonates in some control shRNA cells were not detected in TLR9 shRNA cells. These differences were not associated with inhibition of Rap1A prenylation or p38 phosphorylation, which are known markers for bisphosphonate activity. However, TLR9 shRNA cells exhibited increased sensitivity to ApppI, a metabolite that accumulates in cells after bisphosphonate treatment. We conclude that decreased TLR9-expression sensitizes breast cancer cells to the growth inhibitory effects of bisphosphonates. Our results suggest that TLR9 should be studied as a potential biomarker for adjuvant bisphosphonate sensitivity among breast cancer patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.13570DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5349995PMC
December 2016

Effects of ospemifene, a novel selective estrogen-receptor modulator, on human breast tissue ex vivo.

Menopause 2016 07;23(7):719-30

1Department of Cell Biology and Anatomy, Institute of Biomedicine 2Department of Biostatistics, University of Turku, Finland 3Hormos Ltd, Turku, Finland 4IMC FH Krems, Austria 5Department of Obstetrics and Gynaecology, Turku University Central Hospital, Finland.

Objective: Ospemifene (Osp) is a novel selective estrogen-receptor modulator (SERM) accepted for the treatment of dyspareunia, a symptom of postmenopausal vulvovaginal atrophy. We aimed to analyze the effects of Osp on human breast tissue (HBT), in comparison with the clinically established SERMs raloxifene (Ral) and tamoxifen (Tam), using ex vivo explant cultures.

Methods: HBT samples were obtained from postmenopausal women undergoing mammoplasty and cultured with or without Osp, Ral, Tam, or 17β-estradiol (E2) for 7 and 14 days, and studied for morphology, proliferation, and apoptosis. The expression of epithelial markers, the estrogen-receptor alpha (ERα), the androgen receptor (AR), TFF1, and apolipoprotein D was evaluated using immunohistochemistry and quantitative reverse transcription-polymerase chain reaction. The PvuII polymorphism of ERS1 was determined.

Results: Osp, similar to Ral and Tam, decreased the number of proliferating cells in a concentration-dependent manner (at 100 nM, P < 0.01) and strongly opposed 10 nM E2-stimulated proliferation (P < 0.001). Corresponding effects were observed in the proportions of cells expressing ERα and TFF1 (P < 0.001). At 14 days apoptosis was increased by 100 nM SERMs (P < 0.01), but, notably, decreased by 1 nM Osp and Ral at day 7 (P < 0.05). The SERMs exerted ER-agonist effects on AR-positive cell populations at 1 nM (P < 0.05), but not at 100 nM concentrations. The effects on proliferation and ERα expressing cell numbers were associated with the ERS1 PvuII genotype.

Conclusions: In summary, Osp inhibited proliferation and opposed E2 stimulation in normal HBT in an efficacious, but less potent way than Ral and Tam. The ESR1 PvuII polymorphisms may influence the responsiveness of HBT to E2 and SERMs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/GME.0000000000000624DOI Listing
July 2016

Fam3c modulates osteogenic cell differentiation and affects bone volume and cortical bone mineral density.

Bonekey Rep 2016 6;5:787. Epub 2016 Apr 6.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Turku, Finland.

Fam3c, a cytokine-like growth factor, has been suggested to have a role in epithelial-to-mesenchymal transition (EMT), tumor growth and metastasis. A single-nucleotide polymorphism affecting bone mineral density has been found in the first intron of the Fam3c gene in a study analyzing an Asian population cohort. Other independent studies on different population cohorts have found the fam3c locus to be associated with bone mineral density and fractures. In order to investigate the role of Fam3c in bone biology, we have generated a Fam3c knock-out (KO) mouse strain. The Fam3c KO mice were found to have normal appearance, behavior and fertility, but small changes in bone morphology and content were also observed. Micro-CT analysis of tibiae of the female mice revealed decreased number of trabeculae. In male mice the changes in the bone phenotype were smaller, but hematological changes were observed. Furthermore, there was a negative correlation between body weight and tibial trabecular and cortical bone volume in the male KO mice. There was a small increase in cortical bone mineral density, but in the lateral direction of tibiae the breaking strength was reduced. Fam3c KO bone marrow cells showed accelerated osteogenic differentiation and mineralization in vitro. The reduced number of bone trabeculae in Fam3c KO mice and the stimulated osteogenic differentiation indicate a role for Fam3c in osteoblast differentiation and bone homeostasis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/bonekey.2016.14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4822344PMC
April 2016

Increased expression of fibroblast growth factor 13 in prostate cancer is associated with shortened time to biochemical recurrence after radical prostatectomy.

Int J Cancer 2016 Jul 12;139(1):140-52. Epub 2016 Mar 12.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku, Turku, Finland.

Fibroblast growth factor homologous factors (FHFs) belong to the fibroblast growth factor (FGF) superfamily, which plays an important role in prostate cancer (PCa). Mining of public database suggests that FGF13 (FHF2) mRNA expression is altered in over 30% of PCa cases. This study examined the FGF13 expression pattern in human PCa specimens and evaluated its potential as a biomarker for patient outcome after radical prostatectomy (RP). Immunohistochemistry (IHC) showed that FGF13 was detectable in the majority of human PCa samples, and FGF13 IHC scores were higher in high-grade prostatic intraepithelial neoplasia, in primary PCa and in metastatic PCa than in benign prostatic tissue. There was a significant association between high cytoplasmic FGF13 staining and a risk of biochemical recurrence (BCR) after RP. This was also evident in the intermediate to high-risk patient groups. In contrast, positive nuclear FGF13 staining along with low cytoplasmic FGF13 group showed a decreased BCR risk. Multivariate regression analysis indicated that high cytoplasmic FGF13 staining was associated with BCR and that this could serve as an independent prognostic marker in PCa. Several PCa cell lines showed increased FGF13 expression at the mRNA and protein levels compared to the immortalized prostate epithelial cell line PNT1a. Analysis of co-labeled cells suggested a possible interaction of FGF13 with α-tubulin and the voltage-gated sodium channel proteins (Na(V)s/VGSCs). Our data indicate that, for PCa patients after RP, FGF13 serves as a potential novel prognostic marker that improves prediction of BCR-free survival, in particular if combined with other clinical parameters.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/ijc.30048DOI Listing
July 2016

Spheroid culture of LuCaP 136 patient-derived xenograft enables versatile preclinical models of prostate cancer.

Clin Exp Metastasis 2016 Apr 12;33(4):325-37. Epub 2016 Feb 12.

Department of Urology, Stanford University School of Medicine, Stanford, CA, 94305, USA.

LuCaP serially transplantable patient-derived xenografts (PDXs) are valuable preclinical models of locally advanced or metastatic prostate cancer. Using spheroid culture methodology, we recently established cell lines from several LuCaP PDXs. Here, we characterized in depth the features of xenografts derived from LuCaP 136 spheroid cultures and found faithful retention of the phenotype of the original PDX. In vitro culture enabled luciferase transfection into LuCaP 136 spheroids, facilitating in vivo imaging. We showed that LuCaP 136 spheroids formed intratibial, orthotopic, and subcutaneous tumors when re-introduced into mice. Intratibial tumors responded to castration and were highly osteosclerotic. LuCaP 136 is a realistic in vitro-in vivo preclinical model of a subtype of bone metastatic prostate cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10585-016-9781-2DOI Listing
April 2016

Comparison of the Effects of the Selective Estrogen Receptor Modulators Ospemifene, Raloxifene, and Tamoxifen on Breast Tissue in Ex Vivo Culture.

Methods Mol Biol 2016 ;1366:327-336

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku, Kiinamyllynkatu 10, Turku, 20520, Finland.

Explant tissue culture provides a model for studying the direct effects of steroid hormones, their analogs, and novel hormonally active compounds on normal freshly isolated human breast tissues (HBTs). For this purpose, pre- and postmenopausal HBTs can be maintained in this culture system. The results demonstrate that the morphological integrity of HBT explants can be maintained in tissue culture up to 2 weeks and expression of differentiation markers, steroid hormone receptors, proliferation and apoptosis ratios can be evaluated as a response to hormonal stimulation. This chapter describes an ex vivo culture model that we have applied to study the effects of various hormonally active substances, including 17β-estradiol and selective estrogen receptor modulators (SERMs), on normal human breast tissues.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-4939-3127-9_25DOI Listing
August 2016

Pim Kinases Promote Migration and Metastatic Growth of Prostate Cancer Xenografts.

PLoS One 2015 15;10(6):e0130340. Epub 2015 Jun 15.

Section of Genetics and Physiology, Department of Biology, University of Turku, 20500 Turku, Finland.

Background And Methods: Pim family proteins are oncogenic kinases implicated in several types of cancer and involved in regulation of cell proliferation, survival as well as motility. Here we have investigated the ability of Pim kinases to promote metastatic growth of prostate cancer cells in two xenograft models for human prostate cancer. We have also evaluated the efficacy of Pim-selective inhibitors to antagonize these effects.

Results: We show here that tumorigenic growth of both subcutaneously and orthotopically inoculated prostate cancer xenografts is enhanced by stable overexpression of either Pim-1 or Pim-3. Moreover, Pim-overexpressing orthotopic prostate tumors are highly invasive and able to migrate not only to the nearby prostate-draining lymph nodes, but also into the lungs to form metastases. When the xenografted mice are daily treated with the Pim-selective inhibitor DHPCC-9, both the volumes as well as the metastatic capacity of the tumors are drastically decreased. Interestingly, the Pim-promoted metastatic growth of the orthotopic xenografts is associated with enhanced angiogenesis and lymphangiogenesis. Furthermore, forced Pim expression also increases phosphorylation of the CXCR4 chemokine receptor, which may enable the tumor cells to migrate towards tissues such as the lungs that express the CXCL12 chemokine ligand.

Conclusions: Our results indicate that Pim overexpression enhances the invasive properties of prostate cancer cells in vivo. These effects can be reduced by the Pim-selective inhibitor DHPCC-9, which can reach tumor tissues without serious side effects. Thus, Pim-targeting therapies with DHPCC-9-like compounds may help to prevent progression of local prostate carcinomas to fatally metastatic malignancies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0130340PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4467846PMC
May 2016

Induction of morphological changes in death-induced cancer cells monitored by holographic microscopy.

J Struct Biol 2015 Mar 28;189(3):207-12. Epub 2015 Jan 28.

Department of Biomedical Science, Health and Society, Malmö University, Malmö, Sweden. Electronic address:

We are using the label-free technique of holographic microscopy to analyze cellular parameters including cell number, confluence, cellular volume and area directly in the cell culture environment. We show that death-induced cells can be distinguished from untreated counterparts by the use of holographic microscopy, and we demonstrate its capability for cell death assessment. Morphological analysis of two representative cell lines (L929 and DU145) was performed in the culture flasks without any prior cell detachment. The two cell lines were treated with the anti-tumour agent etoposide for 1-3days. Measurements by holographic microscopy showed significant differences in average cell number, confluence, volume and area when comparing etoposide-treated with untreated cells. The cell volume of the treated cell lines was initially increased at early time-points. By time, cells decreased in volume, especially when treated with high doses of etoposide. In conclusion, we have shown that holographic microscopy allows label-free and completely non-invasive morphological measurements of cell growth, viability and death. Future applications could include real-time monitoring of these holographic microscopy parameters in cells in response to clinically relevant compounds.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jsb.2015.01.010DOI Listing
March 2015

Adenosine inhibits tumor cell invasion via receptor-independent mechanisms.

Mol Cancer Res 2014 Dec 30;12(12):1863-74. Epub 2014 Jul 30.

Department of Medical Microbiology, University of Turku, Turku, Finland.

Unlabelled: Extracellular adenosine mediates diverse anti-inflammatory, angiogenic, and other signaling effects via binding to adenosine receptors, and it also regulates cell proliferation and death via activation of the intrinsic signaling pathways. Given the emerging role of adenosine and other purines in tumor growth and metastasis, this study evaluated the effects of adenosine on the invasion of metastatic prostate and breast cancer cells. Treatment with low micromolar concentrations of adenosine, but not other nucleosides or adenosine receptor agonists, inhibited subsequent cell invasion and migration through Matrigel- and laminin-coated inserts. These inhibitory effects occurred via intrinsic receptor-independent mechanisms, despite the abundant expression of A2B adenosine receptors (ADORA2B). Extracellular nucleotides and adenosine were shown to be rapidly metabolized on tumor cell surfaces via sequential ecto-5'-nucleotidase (CD73/NT5E) and adenosine deaminase reactions with subsequent cellular uptake of nucleoside metabolites and their intracellular interconversion into ADP/ATP. This was accompanied by concurrent inhibition of AMP-activated protein kinase and other signaling pathways. No differences in the proliferation rates, cytoskeleton assembly, expression of major adhesion molecules [integrin-1β (ITGB1), CD44, focal adhesion kinase], and secretion of matrix metalloproteinases were detected between the control and treated cells, thus excluding the contribution of these components of invasion cascade to the inhibitory effects of adenosine. These data provide a novel insight into the ability of adenosine to dampen immune responses and prevent tumor invasion via two different, adenosine receptor-dependent and -independent mechanisms.

Implications: This study suggests that the combined targeting of adenosine receptors and modulation of intracellular purine levels can affect tumor growth and metastasis phenotypes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1158/1541-7786.MCR-14-0302-TDOI Listing
December 2014

The role of PI3K/AKT-related PIP5K1α and the discovery of its selective inhibitor for treatment of advanced prostate cancer.

Proc Natl Acad Sci U S A 2014 Sep 28;111(35):E3689-98. Epub 2014 Jul 28.

Division of Experimental Cancer Research, Department of Laboratory Medicine, Clinical Research Center,

Nitrogen-containing heterocyclic compounds are an important class of molecules that are commonly used for the synthesis of candidate drugs. Phosphatidylinositol-4-phosphate 5-kinase-α (PIP5Kα) is a lipid kinase, similar to PI3K. However, the role of PIP5K1α in oncogenic processes and the development of inhibitors that selectively target PIP5K1α have not been reported. In the present study we report that overexpression of PIP5K1α is associated with poor prognosis in prostate cancer and correlates with an elevated level of the androgen receptor. Overexpression of PIP5K1α in PNT1A nonmalignant cells results in an increased AKT activity and an increased survival, as well as invasive malignant phenotype, whereas siRNA-mediated knockdown of PIP5K1α in aggressive PC-3 cells leads to a reduced AKT activity and an inhibition in tumor growth in xenograft mice. We further report a previously unidentified role for PIP5K1α as a druggable target for our newly developed compound ISA-2011B using a high-throughput KINOMEscan platform. ISA-2011B was discovered during our synthetic studies of C-1 indol-3-yl substituted 1,2,3,4-tetrahydroisoquinolines via a Pictet-Spengler approach. ISA-2011B significantly inhibits growth of tumor cells in xenograft mice, and we show that this is mediated by targeting PIP5K1α-associated PI3K/AKT and the downstream survival, proliferation, and invasion pathways. Further, siRNA-mediated knockdown of PIP5K1α exerts similar effects on PC3 cells as ISA-2011B treatment, significantly inhibiting AKT activity, increasing apoptosis and reducing invasion. Thus, PIP5K1α has high potential as a drug target, and compound ISA-2011B is interesting for further development of targeted cancer therapy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1405801111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4156761PMC
September 2014

Deficiency of ERβ and prostate tumorigenesis in FGF8b transgenic mice.

Endocr Relat Cancer 2014 Aug 17;21(4):677-90. Epub 2014 Jun 17.

Departments of Cell Biology and AnatomyPharmacologyDrug Development and TherapeuticsTurku Center for Disease ModelingInstitute of Biomedicine, University of Turku, Kiinamyllynkatu 10, 20520 Turku, FinlandFunctional Foods ForumUniversity of Turku, Turku, FinlandDepartment of Laboratory MedicineMAS University Hospital, Lund University, Malmö, SwedenDepartments of Cell Biology and AnatomyPharmacologyDrug Development and TherapeuticsTurku Center for Disease ModelingInstitute of Biomedicine, University of Turku, Kiinamyllynkatu 10, 20520 Turku, FinlandFunctional Foods ForumUniversity of Turku, Turku, FinlandDepartment of Laboratory MedicineMAS University Hospital, Lund University, Malmö, Sweden.

Estrogens contribute to the development and growth of the prostate and are implicated in prostate tumorigenesis. In their target tissues, estrogens mediate their effects via estrogen receptor α (ERα (ESR1)) and β (ERβ (ESR2)). Hyperplasia and decreased differentiation of epithelial cells in the prostate have been reported in ERβ knockout (BERKO) mice. Herein, we studied the effect of ERβ deficiency on prostate tumorigenesis by crossing BERKOFVB mice with prostate-targeted human fibroblast growth factor 8b transgenic (FGF8b-Tg) mice. Consistent with results described in our previous report, the prostates of 1-year-old FGF8b-Tg mice displayed stromal aberrations, prostatic intraepithelial neoplasia (mPIN) lesions, inflammation, and occasionally cancer. The prostates of BERKOFVB mice exhibited mild epithelial hypercellularity and inflammation. The prostate phenotypes of FGF8b-Tg-BERKOFVB mice closely resembled those of FGF8b-Tg mice. However, mucinous metaplasia, indicated by Goblet-like cells in the epithelium, was significantly more frequent in the prostates of FGF8b-Tg-BERKOFVB mice when compared with FGF8b-Tg mice. Furthermore, compared with FGF8b-Tg mice, there was a tendency for increased frequency of inflammation but milder hyperplasias in the prostate stroma of FGF8b-Tg-BERKOFVB mice. The expression levels of mRNAs for FGF8b-regulated genes including osteopontin (Spp1), connective tissue growth factor (Ctgf), fibroblast growth factor receptors (Fgfrs), and steroid hormone receptors and cytokines were similar in the prostates of FGF8b-Tg and FGF8b-Tg-BERKOFVB mice. Our results indicate that ERβ plays a role in the differentiation of the prostatic epithelium and, potentially, in the defensive mechanism required for protection against inflammation but do not support a direct tumor-suppressive function of ERβ in the prostate of FGF8b-Tg mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1530/ERC-13-0480DOI Listing
August 2014

Tumor models for prostate cancer exemplified by fibroblast growth factor 8-induced tumorigenesis and tumor progression.

Reprod Biol 2014 Mar 21;14(1):16-24. Epub 2014 Jan 21.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku, Turku, Finland. Electronic address:

Prostate cancer is a very common malignancy among Western males. Although most tumors are indolent and grow slowly, some grow and metastasize aggressively. Because prostate cancer growth is usually androgen-dependent, androgen ablation offers a therapeutic option to treat post-resection tumor recurrence or primarily metastasized prostate cancer. However, patients often relapse after the primary response to androgen ablation therapy, and there is no effective cure for cases of castration-resistant prostate cancer (CRPC). The mechanisms of tumor growth in CRPC are poorly understood. Although the androgen receptors (ARs) remain functional in CRPC, other mechanisms are clearly activated (e.g., disturbed growth factor signaling). Results from our laboratory and others have shown that dysregulation of fibroblast growth factor (FGF) signaling, including FGF receptor 1 (FGFR1) activation and FGF8b overexpression, has an important role in prostate cancer growth and progression. Several experimental models have been developed for prostate tumorigenesis and various stages of tumor progression. These models include genetically engineered mice and rats, as well as induced tumors and xenografts in immunodeficient mice. The latter was created using parental and genetically modified cell lines. All of these models greatly helped to elucidate the roles of different genes in prostate carcinogenesis and tumor progression. Recently, patient-derived xenografts have been studied for possible use in testing individual, specific responses of tumor tissue to different treatment options. Feasible and functional CRPC models for drug responsiveness analysis and the development of effective therapies targeting the FGF signaling pathway and other pathways in prostate cancer are being actively investigated.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.repbio.2014.01.002DOI Listing
March 2014

Inactivation of the androgen receptor in bone-forming cells leads to trabecular bone loss in adult female mice.

Bonekey Rep 2013 6;2:440. Epub 2013 Nov 6.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Turku, Finland.

Removal of the androgen receptor (AR) from bone-forming cells has been shown to reduce trabecular bone volume in male mice. In female mice, the role of AR in the regulation of bone homeostasis has been poorly understood. We generated a mouse strain in which the AR is completely inactivated only in mineralizing osteoblasts and osteocytes by breeding mice carrying osteocalcin promoter-regulated Cre-recombinase with mice possessing loxP recombination sites flanking exon 2 of the AR gene (AR(ΔOB/ΔOB) mice). In female AR(ΔOB/ΔOB) mice, the trabecular bone volume was reduced owing to a smaller number of trabeculae at 6 months of age compared with the control AR(fl/fl) animals. In male AR(ΔOB/ΔOB) mice, an increase in trabecular bone separation could already be detected at 3.5 months of age, and at 6 months, the trabecular bone volume was significantly reduced compared with that of male AR(fl/fl) mice. No AR-dependent changes were observed in the cortical bone of either sex. On the basis of micro-computed tomography and histomorphometry, we conclude that in male mice, the AR is involved in the regulation of osteoclast number by osteoblasts, whereas in female mice, the lack of the AR in the bone-forming cells leads to a decreased number of trabeculae upon aging.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/bonekey.2013.174DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3844973PMC
January 2014

Cyclin A1 modulates the expression of vascular endothelial growth factor and promotes hormone-dependent growth and angiogenesis of breast cancer.

PLoS One 2013 8;8(8):e72210. Epub 2013 Aug 8.

Department of Clinical Sciences, Lund University, Malmö, Sweden.

Alterations in cellular pathways related to both endocrine and vascular endothelial growth factors (VEGF) may contribute to breast cancer progression. Inhibition of the elevated levels of these pathways is associated with clinical benefits. However, molecular mechanisms by which endocrine-related pathways and VEGF signalling cooperatively promote breast cancer progression remain poorly understood. In the present study, we show that the A-type cyclin, cyclin A1, known for its important role in the initiation of leukemia and prostate cancer metastasis, is highly expressed in primary breast cancer specimens and metastatic lesions, in contrasting to its barely detectable expression in normal human breast tissues. There is a statistically significant correlation between cyclin A1 and VEGF expression in breast cancer specimens from two patient cohorts (p<0.01). Induction of cyclin A1 overexpression in breast cancer cell line MCF-7 results in an enhanced invasiveness and a concomitant increase in VEGF expression. In addition, there is a formation of protein-protein complexes between cyclin A1 and estrogen receptor ER-α cyclin A1 overexpression increases ER-α expression in MCF-7 and T47D cells. In mouse tumor xenograft models in which mice were implanted with MCF-7 cells that overexpressed cyclin A1 or control vector, cyclin A1 overexpression results in an increase in tumor growth and angiogenesis, which is coincident with an enhanced expression of VEGF, VEGFR1 and ER-α Our findings unravel a novel role for cyclin A1 in growth and progression of breast cancer, and suggest that multiple cellular pathways, including cell cycle regulators, angiogenesis and estrogen receptor signalling, may cooperatively contribute to breast cancer progression.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0072210PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3744130PMC
April 2014

Novel interaction of Rab13 and Rab8 with endospanins.

FEBS Open Bio 2013 22;3:83-8. Epub 2013 Jan 22.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku, Turku, Finland.

Rab GTPases regulate vesicular traffic in eukaryotic cells by cycling between the active GTP-bound and inactive GDP-bound states. Their functions are modulated by the diverse selection of effector proteins that bind to specific Rabs in their activated state. We previously described the expression of Rab13 in bone cells. To search for novel Rab13 interaction partners, we screened a newborn rat bone marrow cDNA library for Rab13 effectors with a bacterial two-hybrid system. We found that Rab13 binds to the C-terminus of Endospanin-2, a small transmembrane protein. In addition to Rab13 also Rab8 bound to Endospanin-2, while no binding of Rab7, Rab10, Rab11 or Rab32 was observed. Rab13 and Rab8 also interacted with Endospanin-1, a close homolog of Endospanin-2. Rab13 and Endospanin-2 colocalised in perinuclear vesicular structures in Cos1 cells suggesting direct binding also in vivo. Endospanin-2 is implicated in the regulation of the cell surface growth hormone receptor (GHR), but the inhibition of Rab13 expression did not affect GHR cell surface expression. This suggests that the Rab13-Endospanin-2 interaction may have functions other than GHR regulation. In conclusion, we have identified a novel interaction for Rab13 and Rab8 with Endospanin-2 and Endospanin-1. The role of this interaction in cell physiology, however, remains to be elucidated.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.fob.2013.01.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3668521PMC
June 2013

Low expression of SHP-2 is associated with less favorable prostate cancer outcomes.

Tumour Biol 2013 Apr 29;34(2):637-42. Epub 2012 Nov 29.

Department of Laboratory Medicine, Division of Tumor Biology, Lund University, Lund, Sweden.

Src homology 2 domain-containing tyrosine phosphatase-2 (SHP-2) is an important regulator of cell signaling because of its ability to dephosphorylate receptors of growth factors as well as the cytokines and tyrosine-phosphorylated proteins associated with these receptors. In the current study, we used four different prostate cancer cell lines: PC3, DU145, LNCaP and LNCaP-IL6+. Tumor specimens from 122 patients with prostate cancer were analyzed using a tissue microarray. Our data demonstrate that all four prostate cancer cell lines express the SHP-2 protein. Additionally, low staining intensity and SHP-2 expression in the cytoplasm of cancer cells in prostate tumor specimens was inversely correlated with prostate volume (p = 0.041 and p = 0.042, respectively) whereas nuclear staining was positively correlated with extracapsular extension (p = 0.039). In our post-prostatectomy specimens, we found that patients with low SHP-2 expression had less favorable outcomes with respect to biochemical recurrence and clinical progression (p = 0.005 and p = 0.018, respectively). The loss of cytoplasmic SHP-2 expression is associated with increased growth and prostatic cancer progression.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s13277-012-0590-1DOI Listing
April 2013

Differential roles of fibroblast growth factor receptors (FGFR) 1, 2 and 3 in the regulation of S115 breast cancer cell growth.

PLoS One 2012 21;7(11):e49970. Epub 2012 Nov 21.

Institute of Biomedicine, Department of Cell Biology and Anatomy, University of Turku, Turku, Finland.

Fibroblast growth factors (FGFs) regulate the growth and progression of breast cancer. FGF signaling is transduced through FGF receptors 1-4, which have oncogenic or anti-oncogenic roles depending on the ligand and the cellular context. Our aim was to clarify the roles of FGFR1-3 in breast cancer cell growth in vitro and in vivo. Pools of S115 mouse breast cancer cells expressing shRNA against FGFR1, 2 and 3 were created by lentiviral gene transfer, resulting in cells with downregulated expression of FGFR1, FGFR2 or FGFR3 (shR1, shR2 and shR3 cells, respectively) and shLacZ controls. FGFR1-silenced shR1 cells formed small, poorly vascularized tumors in nude mice. Silencing of FGFR2 in shR2 cells was associated with strong upregulation of FGFR1 expression and the formation of large, highly vascularized tumors compared to the control tumors. Silencing FGFR3 did not affect cell survival or tumor growth. Overexpressing FGFR2 in control cells did not affect FGFR1 expression, suggesting that high FGFR1 expression in shR2 cells and tumors was associated with FGFR2 silencing by indirect mechanisms. The expression of FGFR1 was, however, increased by the addition of FGF-8 to starved shLacZ or MCF-7 cells and decreased by the FGFR inhibitor PD173074 in shR2 cells with an elevated FGFR1 level. In conclusion, our results demonstrate that FGFR1 is crucial for S115 breast cancer cell proliferation and tumor growth and angiogenesis, whereas FGFR2 and FGFR3 are less critical for the growth of these cells. The results also suggest that the expression of FGFR1 itself is regulated by FGF-8 and FGF signaling, which may be of importance in breast tumors expressing FGFs at a high level.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0049970PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3503871PMC
May 2013

Inactivation of estrogen receptor α in bone-forming cells induces bone loss in female mice.

FASEB J 2013 Feb 16;27(2):478-88. Epub 2012 Oct 16.

Institute of Biomedicine, Department of Cell Biology and Anatomy, University of Turku, FI20520 Turku, Finland.

The role of the estrogen receptor α (ERα) in bone-forming cells is incompletely understood at present. To examine the in vivo effects of ERα in these cells, we generated a mouse strain in which the ERα gene is inactivated in osteoblasts via osteocalcin promoter-regulated cyclic recombinase (Cre) activity (ERα(ΔOB/ΔOB)). This enabled micro-computed tomography- and histomorphometry-based analysis of ERα-mediated effects in bone-forming cells in mice, in which the systemic ERα-mediated effects are intact. In female ERα(ΔOB/ΔOB) mice, trabecular and cortical bone volumes were significantly reduced (31.5 and 11.4%, respectively) at 3.5 mo of age compared with control ERα(fl/fl) animals, and their response to ovariectomy was small compared with that of controls. In contrast with females, no differences could be detected in the bone phenotype of young males, whereas in 6-mo-old ERα(ΔOB/ΔOB) males, trabecular bone volume (Tb.BV) was decreased (27.5%). The ERα inactivation-related effects were compared with those of controls having a similar genetic background. Parental osteocalcin-Cre mice did not show Cre-related changes. Our results suggest that in female mice, Tb.BV and cortical bone volume are critically dependent on the ERα regulation of osteoblasts, whereas in male mice, osteoblastic ERα is not required for the regulation of bone formation during rapid skeletal growth, but it is involved in the maintenance of Tb.BV.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.12-213587DOI Listing
February 2013

Low TLR9 expression defines an aggressive subtype of triple-negative breast cancer.

Breast Cancer Res Treat 2012 Sep 31;135(2):481-93. Epub 2012 Jul 31.

Department of Medicine, Division of Hematology-Oncology, University of Alabama at Birmingham, SHEL 514, 1825 University Blvd, Birmingham, AL 35294-3300, USA.

Toll-like receptor-9 (TLR9) is a DNA receptor widely expressed in cancers. Although synthetic TLR9 ligands induce cancer cell invasion in vitro, the role of TLR9 in cancer pathophysiology is unclear. We discovered that low tumor TLR9 expression is associated with significantly shortened disease-specific survival in patients with triple negative but not with ER+ breast cancers. A likely mechanism of this clinical finding involves differential responses to hypoxia. Our pre-clinical studies indicate that while TLR9 expression is hypoxia-regulated, low TLR9 expression has different effects on triple negative and ER+ breast cancer invasion in hypoxia. Hypoxia-induced invasion is augmented by TLR9 siRNA in triple negative, but not in ER+ breast cancer cells. This is possibly due to differential TLR9-regulated TIMP-3 expression, which remains detectable in ER+ cells but disappears from triple-negative TLR9 siRNA cells in hypoxia. Our results demonstrate a novel role for this innate immunity receptor in cancer biology and suggest that TLR9 expression may be a novel marker for triple-negative breast cancer patients who are at a high risk of relapse. Furthermore, these results suggest that interventions or events, which induce hypoxia or down-regulate TLR9 expression in triple-negative breast cancer cells may actually induce their spread.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10549-012-2181-7DOI Listing
September 2012

Rab13 is upregulated during osteoclast differentiation and associates with small vesicles revealing polarized distribution in resorbing cells.

J Histochem Cytochem 2012 Jul 4;60(7):537-49. Epub 2012 May 4.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku, Turku, Finland.

Osteoclasts are bone-resorbing multinucleated cells that undergo drastic changes in their polarization due to heavy vesicular trafficking during the resorption cycle. These events require the precise orchestration of membrane traffic in order to maintain the unique characteristics of the different membrane domains in osteoclasts. Rab proteins are small GTPases involved in regulation of most, if not all, steps of vesicle trafficking. The investigators studied RAB genes in human osteoclasts and found that at least 26 RABs were expressed in osteoclasts. Out of these, RAB13 gene expression was highly upregulated during differentiation of human peripheral blood monocytic cells into osteoclasts. To study its possible function in osteoclasts, the investigators performed immunolocalization studies for Rab13 and various known markers of osteoclast vesicular trafficking. Rab13 localized to small vesicular structures at the superior parts of the osteoclast between the trans-Golgi network and basolateral membrane domain. Rab13 localization suggests that it is not involved in endocytosis or transcytosis of bone degradation products. In addition, Rab13 did not associate with early endosomes or recycling endosomes labeled with EEA1 or TRITC-conjugated transferrin, respectively. Its involvement in glucose transporter traffic was excluded as well. It is suggested that Rab13 is associated with a putative secretory function in osteoclasts.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1369/0022155412448069DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3460351PMC
July 2012

Androgens inhibit the stimulatory action of 17β-estradiol on normal human breast tissue in explant cultures.

J Clin Endocrinol Metab 2012 Jul 24;97(7):E1116-27. Epub 2012 Apr 24.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku, Kiinamyllynkatu 10, 20520 Turku, Finland.

Background: The data concerning the effects and safety of androgen in human breast tissue are conflicting.

Objective: Our aim was to analyze the effects of androgens on normal human breast tissue (HBT).

Approach: We cultured explants of HBT (obtained from reduction mammoplasty operations of postmenopausal women) with or without testosterone (T) and 5α-dihydrotestosterone (DHT) or in combination with 17β-estradiol (E(2)) for 7 and 14 d to study the effects of androgens on proliferation, apoptosis, target gene expression, and steroid receptors. The androgen receptor (AR) and estrogen receptor (ER) dependences of the effects were studied with the antihormones bicalutamide and fulvestrant, respectively.

Results: The hormone responsiveness of cultured breast tissue was assessed by assaying apolipoprotein-D and prostate-specific antigen expression increased by androgens and amphiregulin and trefoil factor-1 expression induced by E(2) treatment. T and DHT reduced proliferation and increased apoptosis in breast epithelium, the effects of which were reversed by bicalutamide. In combination with E(2), they suppressed E(2)-stimulated proliferation and cell survival. DHT also inhibited basal (P < 0.05) and E(2)-induced expression of cyclin-D1 mRNA (P < 0.05). Immunohistochemistry showed that T (P < 0.05) and DHT (P < 0.05) increased the relative number of AR-positive cells, whereas ERα-positive (P < 0.001) cell numbers were strongly decreased. The percentage of ERβ-positive cells remained unchanged. E(2) treatment increased ERα-positive (P < 0.01) cells, whereas AR- (P < 0.05) and ERβ-expressing (P < 0.001) cells diminished. These effects were repressed in combination cultures of E(2) with T and DHT.

Conclusion: T and DHT inhibited proliferation and increased apoptosis in the epithelium of cultured normal HBT and opposed E(2)-stimulated proliferation and cell survival in an AR-dependent manner. These effects were associated with changes in the proportions of ERα- and AR-positive epithelial cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1210/jc.2011-3228DOI Listing
July 2012

Fibroblast growth factor 8b causes progressive stromal and epithelial changes in the epididymis and degeneration of the seminiferous epithelium in the testis of transgenic mice.

Biol Reprod 2012 May 17;86(5):157, 1-12. Epub 2012 May 17.

Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku, Turku, Finland.

Transgenic (Tg) mice expressing human fibroblast growth factor 8b (FGF8-b) under the probasin promoter (Tg [Pbsn-FGF8] L2-L5Elo; hereafter referred to as FGF8-b-Tg) were shown to produce FGF8-b at high levels in the prostate and epididymis and at lower levels in the testis. The present study examined the effects of FGF8-b expression on the epididymis and testis. In old (age, >6 mo) FGF8-b-Tg mice, epididymides were frequently enlarged, with epithelial and stromal hypercellularity progressing upon aging to epithelial dysplasia and malignant transformation of stroma. In addition, oligospermia, dilatation of the duct, and inflammation were frequently observed in the epididymides. In association with the epididymal changes, some FGF8-b-Tg mice presented a degenerative seminiferous epithelium of the testis. Consistent with this observation, infertile males were found in two FGF8-b-Tg mouse lines. Masson trichrome staining and immunohistochemical analysis of smooth muscle actin, laminin, and androgen receptor revealed that changes in the epididymal stroma closely resembled those previously found in the prostates of the FGF8-b-Tg mice. Genes previously found to be upregulated in the prostate of FGF8-b-Tg mice, such as osteopontin (Spp1) connective tissue growth factor (Ctgf), apolipoprotein D (Apod), and FGF receptor 1c (Fgfr1-c), were also upregulated in the epididymides, suggesting that similar molecular mechanisms were active in both tissues. However, unlike in the prostate, the changes in the epididymal epithelium of the FGF8-b-Tg mice did not progress into invasive carcinoma. The results suggest that prolonged and enhanced FGF signaling induces dramatic changes in the epididymis and testis that lead to infertility in a portion of the FGF8-b-Tg males.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1095/biolreprod.111.097352DOI Listing
May 2012

Tracer level electrophilic synthesis and pharmacokinetics of the hypoxia tracer [(18)F]EF5.

Mol Imaging Biol 2012 Apr;14(2):205-12

Turku PET Centre, Radiopharmaceutical Chemistry Laboratory, University of Turku, Turku, Finland.

Purpose: 2-(2-nitro-1H-imidazol-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)-acetamide labeled with [(18)F]-fluorine ([(18)F]EF5), a promising tracer for tumor hypoxia, has previously been synthesized in low yields and low specific radioactivity. In pharmacokinetic evaluations, in the presence of non-radioactive EF5, a uniform and low background uptake and high in vivo stability of [(18)F]EF5 have been demonstrated. Our purpose was to increase the specific radioactivity of [(18)F]EF5 to enable to study the pharmacokinetics at trace level.

Procedures: [(18)F]EF5 was synthesized using high specific radioactivity electrophilic [(18)F]F(2) as labelling reagent. Biodistribution of [(18)F]EF5 was determined in a prostate tumor mouse model, and formation of radiolabelled metabolites was studied in mouse, rat and human plasma.

Results: On average, 595 ± 153 MBq of [(18)F]EF5 was produced. Specific radioactivity was 6.6 ± 1.9 GBq/μmol and the radiochemical purity exceeded 99.0%. [(18)F]EF5 was distributed uniformly in tissues, with highest uptake in liver, kidney, and intestine. Several radiolabelled metabolites were detected in mouse plasma and tissues, whereas low amounts of metabolites were detected in human and rat plasma.

Conclusions: [(18)F]EF5 was synthesized by electrophilic labelling with high quality and high yields. Pharmacokinetics of [(18)F]EF5 was determined at trace level in several species. Our results suggest that the trace-level approach does not affect the biodistribution of [(18)F]EF5. Extensive metabolism was seen in mouse.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s11307-011-0484-4DOI Listing
April 2012

Multiparametric luminescence method for quantitative cell surface protein expression analysis and imaging.

J Immunol Methods 2011 Mar 28;367(1-2):40-6. Epub 2011 Jan 28.

Laboratory of Biophysics, Department of Cell Biology and Anatomy and Medicity Research Laboratories, Institute of Biomedicine, University of Turku, Turku, Finland.

A luminometric method for quantitative cell surface protein expression analysis has been developed in a microtiter plate format. The method is based on immunocytochemistry, the use of long-lived europium(III) and terbium(III) chelates and platinum(II) porphyrin luminescence labels in addition to short-lived syto13 DNA stain, and detection of photoluminescence emission from adhered cells by both time-resolved luminescence and conventional fluorescence. After the immunoreactions, the wells were evaporated to dryness, allowing repeated and postponed luminescence analysis even after months and cellular protein localization studies by microscopy imaging. The multiparametric method assayed the cell surface expression of ß1-integrin, E-selectin and intercellular adhesion molecule 1 (ICAM-1) in HUVE cells (human umbilical vein endothelial cells). The expression of E-selectin and ICAM-1 was enhanced by treating HUVECs with tumor necrosis factor α (TNF-α), while the expression level of ß1-integrin remained unchanged. The sensitivity limit of TNF-α detection by the method was ca. 1 pg/ml and the Z'-factors for the quantification of E-selectin and ICAM-1 were >0.7 suggesting a highly robust method. The novel approach proposed in this paper can be potentially applied to cell surface protein expression analysis in screening applications combined with localization studies of the target proteins by fluorescence microscopy imaging.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jim.2011.01.008DOI Listing
March 2011