Glial cell line–derived neurotrophic factor promotes invasive behaviour in testicular seminoma cells
Department of Anatomy, Histology, Forensic Medicine and Orthopedic, Section of Histology, Fondazione Pasteur Cenci Bolognetti, Sapienza University of Rome, Rome Department of Experimental Medicine, Second University of Naples, Naples Department of Radiological, Oncologic and Pathologic Sciences, Sapienza University of Rome, Rome, Italy. International Journal of Andrology
(Impact Factor: 3.7).
04/2012; 35(5):758-68. DOI: 10.1111/j.1365-2605.2012.01267.x
The glial cell line-derived neurotrophic factor (GDNF) has multiple functions that promote cell survival, proliferation and migration in different cell types. The experimental over-expression of GDNF in mouse testis leads to infertility and promotes seminomatous germ cell tumours in older animals, which suggests that deregulation of the GDNF pathway may be implicated in germ cell carcinogenesis. GDNF activates downstream pathways upon binding to its specific co-receptor GDNF family receptor-a 1 (GFRA1). This complex then interacts with Ret and other co-receptors to activate several intracellular signalling cascades. To explore the involvement of the GDNF pathway in the onset and progression of testicular germ cell tumours, we analysed GFRA1 and Ret expression patterns in seminoma samples. We demonstrated, via immunohistochemistry, that GFRA1, but not Ret, is over-expressed in in situ carcinoma (CIS) and in intratubular and invasive seminoma cells compared with normal human germ cells. Functional analysis of the GDNF biological activity was performed on TCam-2 seminoma cell line. Reverse transcription-PCR (RT-PCR) and immunohistochemical analyses demonstrate that TCam-2 cells express both GFRA1 and Ret mRNA, but only GFRA1 was detected at the protein level. In TCam-2 cells, although GDNF is not mitogenic, it is able to induce migration, as demonstrated by a Boyden chamber assay, possibly through the Src and MEK pathways. Moreover, GDNF promotes invasive behaviour, an effect dependent on pericellular protease activity, possibly through the activity of matrix metalloproteinases. GFRA1 over-expression in CIS and seminoma cells, along with the functional analyses in TCam-2 cells, suggests an involvement of the GDNF pathway in the progression of testicular germ cell cancer.
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Available from: Fabrizio D'Anselmi
- "The cells were assayed for their ability to migrate through a polycarbonate filter (pore size, 8 µm; Whatman International) using Boyden chambers (NeuroProbe) as previously described . Cells were cultured for 72 hours with or without 10% EW in the presence of 10% FBS and then maintained for 16 hours under serum-free conditions. "
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ABSTRACT: Seminoma is one of the most common Testicular Germ Cell Tumours that originates during embryonic development due to an alteration of the local niche that in turn results in a delayed or blocked differentiation of Primordial Germ Cells. The block of differentiation is actually a common way to develop cancer disease as postulated by the "embryonic rest theory of cancer". In agreement with this theory different studies have demonstrated that embryonic cues display the capacity of reprogramming aggressive cancer cells towards a less aggressive phenotype. Herein we investigate the ability of a culture medium added with 10% egg albumen (EW, Egg White) to modulate seminoma cell phenotype and behaviour, by ensuring a proper set of morphogenetic signals. We chose to use the TCam-2 seminoma cell line that has been established as the only available cell line, obtained from a primary testicular seminoma. EW is able to: 1) modify TCam-2 cell spreading rate and cell-substrate adhesion without affecting proliferation and survival indexes; 2) modulate TCam-2 actin distribution pattern increasing cortical localization of actin filaments; 3) increase TCam-2 cell-cell junction capability; 4) decrease both chemo-sensitive and collective TCam-2 migratory behaviour. According to these observations morphometric fractal analysis revealed the ability of EW to increase Circularity and Solidity parameters and, consequently, to decrease Fractal dimension. Prompted by these observations we hypothesize that EW treatment could rescue, at least in part, the neoplastic-metastatic behaviour of seminoma cells.
PLoS ONE 10/2013; 8(10):e76192. DOI:10.1371/journal.pone.0076192 · 3.23 Impact Factor
Available from: Margherita Grasso
- "Thus, the extracellular concentration of GDNF in a given area of the seminiferous tubule may vary with time . Intriguingly, GDNF has also been demonstrated to stimulate chemotaxis in both normal, transformed cells – and seminoma cells . Here, we tested the hypotheses that GDNF is a chemoattractant for undifferentiated spermatogonia, including stem/progenitor cells, and that the GDNF pathway may affect proteins involved in actin cytoskeleton rearrangement in target cells. "
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ABSTRACT: In mammals, the biological activity of the stem/progenitor compartment sustains production of mature gametes through spermatogenesis. Spermatogonial stem cells and their progeny belong to the class of undifferentiated spermatogonia, a germ cell population found on the basal membrane of the seminiferous tubules. A large body of evidence has demonstrated that glial cell line-derived neurotrophic factor (GDNF), a Sertoli-derived factor, is essential for in vivo and in vitro stem cell self-renewal. However, the mechanisms underlying this activity are not completely understood. In this study, we show that GDNF induces dose-dependent directional migration of freshly selected undifferentiated spermatogonia, as well as germline stem cells in culture, using a Boyden chamber assay. GDNF-induced migration is dependent on the expression of the GDNF co-receptor GFRA1, as shown by migration assays performed on parental and GFRA1-transduced GC-1 spermatogonial cell lines. We found that the actin regulatory protein vasodilator-stimulated phosphoprotein (VASP) is specifically expressed in undifferentiated spermatogonia. VASP belongs to the ENA/VASP family of proteins implicated in actin-dependent processes, such as fibroblast migration, axon guidance, and cell adhesion. In intact seminiferous tubules and germline stem cell cultures, GDNF treatment up-regulates VASP in a dose-dependent fashion. These data identify a novel role for the niche-derived factor GDNF, and they suggest that GDNF may impinge on the stem/progenitor compartment, affecting the actin cytoskeleton and cell migration.
PLoS ONE 04/2013; 8(4):e59431. DOI:10.1371/journal.pone.0059431 · 3.23 Impact Factor
Available from: Rossana Cecchi
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ABSTRACT: Forensic pathologists are often asked to provide evidence of asphyxia death in the trial and a histological marker of asphyxiation would be of great help. Data from the literature indicate that the reaction of lung tissue cells to asphyxia may be of more interest for forensic purposes than migrating cells. The lungs of 62 medico-legal autopsy cases, 34 acute mechanical asphyxia (AMA), and 28 control cases (CC), were immunostained with anti-P-selectin, anti-E-selectin, anti-SP-A, and anti-HIF1-α antibodies, in order to verify if some of them may be used as markers of asphyxia death. Results show that P- and E-selectins expression in lung vessels, being activated by several types of trigger stimuli not specific to hypoxia, cannot be used as indicator of asphyxia. Intra-alveolar granular deposits of SP-A seem to be related to an intense hypoxic stimulus, and when massively present, they can suggest, together with other elements, a severe hypoxia as the mechanism of death. HIF1-α was expressed in small-, medium-, and large-caliber lung vessels of the vast majority of mechanical asphyxia deaths and CO intoxications, with the number and intensity of positive-stained vessels increasing with the duration of the hypoxia. Although further confirmation studies are required, these preliminary data indicate an interesting potential utility of HIF1-α as a screening test for asphyxia deaths.
Deutsche Zeitschrift für die Gesamte Gerichtliche Medizin 05/2013; 128(1). DOI:10.1007/s00414-013-0876-x · 2.71 Impact Factor
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