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Intratubular germ cell neoplasia of the human testis: heterogeneous protein expression and relation to invasive potential.

Mitchell RT, E Camacho-Moll M, Macdonald J, Anderson RA, Kelnar CJ, O'Donnell M, Sharpe RM, Smith LB, Grigor KM, Wallace WH, Stoop H, Wolffenbuttel KP, Donat R, Saunders PT, Looijenga LH - Mod. Pathol. (2014)

Bottom Line: We compared these subpopulations with normal human fetal testis and with seminoma cells.OCT4(+)/MAGEA4(-) cells showed a significantly increased rate of proliferation compared with the OCT4(+)/MAGEA4(+) population (12.8 versus 3.4%, P<0.0001) irrespective of histological tumor type, reflected in the predominance of OCT4(+)/MAGEA4(-) cells in the invasive tumor component.Surprisingly, OCT4(+)/MAGEA4(-) cells in patients with preinvasive disease showed significantly higher proliferation compared to those with seminoma or non-seminoma (18.1 versus 10.2 versus 7.2%, P<0.05, respectively).

View Article: PubMed Central - PubMed

Affiliation: 1] MRC Centre for Reproductive Health, The Queen's Medical Research Institute, The University of Edinburgh, Edinburgh, UK [2] Edinburgh Royal Hospital for Sick Children, Edinburgh, UK.

ABSTRACT
Testicular germ cell cancer develops from premalignant intratubular germ cell neoplasia, unclassified cells that are believed to arise from failure of normal maturation of fetal germ cells from gonocytes (OCT4(+)/MAGEA4(-)) into pre-spermatogonia (OCT4(-)/MAGEA4(+)). Intratubular germ cell neoplasia cell subpopulations based on stage of germ cell differentiation have been described, however the importance of these subpopulations in terms of invasive potential has not been reported. We hypothesized that cells expressing an immature (OCT4(+)/MAGEA4(-)) germ cell profile would exhibit an increased proliferation rate compared with those with a mature profile (OCT4(+)/MAGEA4(+)). Therefore, we performed triple immunofluorescence and stereology to quantify the different intratubular germ cell neoplasia cell subpopulations, based on expression of germ cell (OCT4, PLAP, AP2γ, MAGEA4, VASA) and proliferation (Ki67) markers, in testis sections from patients with preinvasive disease, seminoma, and non-seminoma. We compared these subpopulations with normal human fetal testis and with seminoma cells. Heterogeneity of protein expression was demonstrated in intratubular germ cell neoplasia cells with respect to gonocyte and spermatogonial markers. It included an embryonic/fetal germ cell subpopulation lacking expression of the definitive intratubular germ cell neoplasia marker OCT4, that did not correspond to a physiological (fetal) germ cell subpopulation. OCT4(+)/MAGEA4(-) cells showed a significantly increased rate of proliferation compared with the OCT4(+)/MAGEA4(+) population (12.8 versus 3.4%, P<0.0001) irrespective of histological tumor type, reflected in the predominance of OCT4(+)/MAGEA4(-) cells in the invasive tumor component. Surprisingly, OCT4(+)/MAGEA4(-) cells in patients with preinvasive disease showed significantly higher proliferation compared to those with seminoma or non-seminoma (18.1 versus 10.2 versus 7.2%, P<0.05, respectively). In conclusion, this study has demonstrated that OCT4(+)/MAGEA4(-) cells are the most frequent and most proliferative cell population in tubules containing intratubular germ cell neoplasia, which appears to be an important factor in determining invasive potential of intratubular germ cell neoplasia to seminomas.

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Representative image for expression of VASA (green) and AP2γ (red) in tubules from adult patients with testicular germ cell cancer. A) Tubule with apparently normal spermatogenesis: VASA is expressed in the germ cells with no expression of the intratubular germ cell neoplasia/gonocyte protein AP2γ. B) Tubule with abnormal spermatogenesis: VASA expression is seen in the presumptive spermatocytes towards the lumen (white arrowhead), however germ cells along the basement membrane express VASA (yellow arrowhead) or AP2γ (yellow arrow). A small proportion of cells co-express VASA and AP2γ (white arrow). C) Intratubular germ cell neoplasia tubule: The majority of cells express the intratubular germ cell neoplasia protein AP2γ (yellow arrow) with a small number of cells adjacent to the basement membrane expressing VASA (presumptive spermatogonia; pink arrow). A small number of germ cells expressing VASA are located towards the lumen (presumptive spermatocytes; white arrowhead).
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Figure 2: Representative image for expression of VASA (green) and AP2γ (red) in tubules from adult patients with testicular germ cell cancer. A) Tubule with apparently normal spermatogenesis: VASA is expressed in the germ cells with no expression of the intratubular germ cell neoplasia/gonocyte protein AP2γ. B) Tubule with abnormal spermatogenesis: VASA expression is seen in the presumptive spermatocytes towards the lumen (white arrowhead), however germ cells along the basement membrane express VASA (yellow arrowhead) or AP2γ (yellow arrow). A small proportion of cells co-express VASA and AP2γ (white arrow). C) Intratubular germ cell neoplasia tubule: The majority of cells express the intratubular germ cell neoplasia protein AP2γ (yellow arrow) with a small number of cells adjacent to the basement membrane expressing VASA (presumptive spermatogonia; pink arrow). A small number of germ cells expressing VASA are located towards the lumen (presumptive spermatocytes; white arrowhead).

Mentions: For the identification of intratubular germ cell neoplasia cells and to attempt to distinguish these cells from normal germ cells, AP2γ and VASA co-expression was investigated (Fig 2). In tubules with normal-appearing spermatogenesis there was expression of VASA in the cytoplasm of germ cells, but no expression of AP2γ (Fig 2A). In tubules containing a mixture of germ cells characteristic of either intratubular germ cell neoplasia or normal spermatogonia the putative intratubular germ cell neoplasia cells (located on the basement membrane) were identified as AP2γ+/VASA−, whilst a small proportion of these cells were AP2γ+/VASA+ (Fig 2B). AP2γ−/VASA+cells, located nearer the lumen were also identified in intratubular germ cell neoplasia tubules. These putative spermatocytes were also identified in tubules in which the majority of the cells were intratubular germ cell neoplasia cells (AP2γ+/VASA−; Fig 2C). Similar populations of germ cells were also identified within the human fetal testis, based on co-staining for OCT4 and VASA (Supp. Fig. 1; [7]).


Intratubular germ cell neoplasia of the human testis: heterogeneous protein expression and relation to invasive potential.

Mitchell RT, E Camacho-Moll M, Macdonald J, Anderson RA, Kelnar CJ, O'Donnell M, Sharpe RM, Smith LB, Grigor KM, Wallace WH, Stoop H, Wolffenbuttel KP, Donat R, Saunders PT, Looijenga LH - Mod. Pathol. (2014)

Representative image for expression of VASA (green) and AP2γ (red) in tubules from adult patients with testicular germ cell cancer. A) Tubule with apparently normal spermatogenesis: VASA is expressed in the germ cells with no expression of the intratubular germ cell neoplasia/gonocyte protein AP2γ. B) Tubule with abnormal spermatogenesis: VASA expression is seen in the presumptive spermatocytes towards the lumen (white arrowhead), however germ cells along the basement membrane express VASA (yellow arrowhead) or AP2γ (yellow arrow). A small proportion of cells co-express VASA and AP2γ (white arrow). C) Intratubular germ cell neoplasia tubule: The majority of cells express the intratubular germ cell neoplasia protein AP2γ (yellow arrow) with a small number of cells adjacent to the basement membrane expressing VASA (presumptive spermatogonia; pink arrow). A small number of germ cells expressing VASA are located towards the lumen (presumptive spermatocytes; white arrowhead).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4012991&req=5

Figure 2: Representative image for expression of VASA (green) and AP2γ (red) in tubules from adult patients with testicular germ cell cancer. A) Tubule with apparently normal spermatogenesis: VASA is expressed in the germ cells with no expression of the intratubular germ cell neoplasia/gonocyte protein AP2γ. B) Tubule with abnormal spermatogenesis: VASA expression is seen in the presumptive spermatocytes towards the lumen (white arrowhead), however germ cells along the basement membrane express VASA (yellow arrowhead) or AP2γ (yellow arrow). A small proportion of cells co-express VASA and AP2γ (white arrow). C) Intratubular germ cell neoplasia tubule: The majority of cells express the intratubular germ cell neoplasia protein AP2γ (yellow arrow) with a small number of cells adjacent to the basement membrane expressing VASA (presumptive spermatogonia; pink arrow). A small number of germ cells expressing VASA are located towards the lumen (presumptive spermatocytes; white arrowhead).
Mentions: For the identification of intratubular germ cell neoplasia cells and to attempt to distinguish these cells from normal germ cells, AP2γ and VASA co-expression was investigated (Fig 2). In tubules with normal-appearing spermatogenesis there was expression of VASA in the cytoplasm of germ cells, but no expression of AP2γ (Fig 2A). In tubules containing a mixture of germ cells characteristic of either intratubular germ cell neoplasia or normal spermatogonia the putative intratubular germ cell neoplasia cells (located on the basement membrane) were identified as AP2γ+/VASA−, whilst a small proportion of these cells were AP2γ+/VASA+ (Fig 2B). AP2γ−/VASA+cells, located nearer the lumen were also identified in intratubular germ cell neoplasia tubules. These putative spermatocytes were also identified in tubules in which the majority of the cells were intratubular germ cell neoplasia cells (AP2γ+/VASA−; Fig 2C). Similar populations of germ cells were also identified within the human fetal testis, based on co-staining for OCT4 and VASA (Supp. Fig. 1; [7]).

Bottom Line: We compared these subpopulations with normal human fetal testis and with seminoma cells.OCT4(+)/MAGEA4(-) cells showed a significantly increased rate of proliferation compared with the OCT4(+)/MAGEA4(+) population (12.8 versus 3.4%, P<0.0001) irrespective of histological tumor type, reflected in the predominance of OCT4(+)/MAGEA4(-) cells in the invasive tumor component.Surprisingly, OCT4(+)/MAGEA4(-) cells in patients with preinvasive disease showed significantly higher proliferation compared to those with seminoma or non-seminoma (18.1 versus 10.2 versus 7.2%, P<0.05, respectively).

View Article: PubMed Central - PubMed

Affiliation: 1] MRC Centre for Reproductive Health, The Queen's Medical Research Institute, The University of Edinburgh, Edinburgh, UK [2] Edinburgh Royal Hospital for Sick Children, Edinburgh, UK.

ABSTRACT
Testicular germ cell cancer develops from premalignant intratubular germ cell neoplasia, unclassified cells that are believed to arise from failure of normal maturation of fetal germ cells from gonocytes (OCT4(+)/MAGEA4(-)) into pre-spermatogonia (OCT4(-)/MAGEA4(+)). Intratubular germ cell neoplasia cell subpopulations based on stage of germ cell differentiation have been described, however the importance of these subpopulations in terms of invasive potential has not been reported. We hypothesized that cells expressing an immature (OCT4(+)/MAGEA4(-)) germ cell profile would exhibit an increased proliferation rate compared with those with a mature profile (OCT4(+)/MAGEA4(+)). Therefore, we performed triple immunofluorescence and stereology to quantify the different intratubular germ cell neoplasia cell subpopulations, based on expression of germ cell (OCT4, PLAP, AP2γ, MAGEA4, VASA) and proliferation (Ki67) markers, in testis sections from patients with preinvasive disease, seminoma, and non-seminoma. We compared these subpopulations with normal human fetal testis and with seminoma cells. Heterogeneity of protein expression was demonstrated in intratubular germ cell neoplasia cells with respect to gonocyte and spermatogonial markers. It included an embryonic/fetal germ cell subpopulation lacking expression of the definitive intratubular germ cell neoplasia marker OCT4, that did not correspond to a physiological (fetal) germ cell subpopulation. OCT4(+)/MAGEA4(-) cells showed a significantly increased rate of proliferation compared with the OCT4(+)/MAGEA4(+) population (12.8 versus 3.4%, P<0.0001) irrespective of histological tumor type, reflected in the predominance of OCT4(+)/MAGEA4(-) cells in the invasive tumor component. Surprisingly, OCT4(+)/MAGEA4(-) cells in patients with preinvasive disease showed significantly higher proliferation compared to those with seminoma or non-seminoma (18.1 versus 10.2 versus 7.2%, P<0.05, respectively). In conclusion, this study has demonstrated that OCT4(+)/MAGEA4(-) cells are the most frequent and most proliferative cell population in tubules containing intratubular germ cell neoplasia, which appears to be an important factor in determining invasive potential of intratubular germ cell neoplasia to seminomas.

Show MeSH
Related in: MedlinePlus