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. 2001 Mar 13;98(6):3316-21.
doi: 10.1073/pnas.051566098. Epub 2001 Feb 27.

Testicular FasL is expressed by sperm cells

A D'Alessio  1 , A Riccioli, P Lauretti, F Padula, B Muciaccia, P De Cesaris, A Filippini, S Nagata, E Ziparo
Affiliations

Testicular FasL is expressed by sperm cells

A D'Alessio et al. Proc Natl Acad Sci U S A. .

Abstract

The testis is the main source of Fas ligand (FasL) mRNA in rodents; it is generally believed that this molecule, expressed on bordering somatic Sertoli cells, bestows an immune-privileged status in the testis by eliminating infiltrating inflammatory Fas-bearing leukocytes. Our results demonstrate that the attribution of testicular expression of FasL to Sertoli cells is erroneous and that FasL transcription instead occurs in meiotic and postmeiotic germ cells, whereas the protein is only displayed on mature spermatozoa. These findings point to a significant role of the Fas system in the biology of mammalian reproduction.

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Figures

Figure 1
Figure 1
(a) Age-dependent expression of FasL in the rat testis. Total RNA (20 μg/lane) extracted from total testis from animals of various ages were analyzed by Northern blot. Nylon filter was hybridized with 32P-labeled rat FasL cDNA probe (b) Northern blot analysis of FasL expression in Sertoli cells at different ages. Total RNA (20 μg/lane) from cultured and crude (*) Sertoli cells were subjected to Northern blot analysis for detection of rat FasL. N2a-FasL+ and N2a-Neo cell lines were used as positive and negative controls for FasL, respectively. The integrity and equal loading of RNA was ascertained by ethidium bromide staining of the gel before transfer (Lower). Data are representative of three independent experiments.
Figure 2
Figure 2
Morphological study of seminiferous epithelium from SCE adult rats. The semithin sections show: (a) Aspermatogenic Sertoli-cell-only testis from 41-day-old rat irradiated in utero; (b) the recovery of spermatogenesis 63 days after birth (arrow heads indicate spermatocytes); (c) the abdominal testis, showing the absence of spermatogenesis; and (d) controlateral scrotal testis in unilaterally cryptorchid SCE rat (210× magnification for all). (e) Northern blot analysis of FasL expression in SCE rat testes. Total RNA from unilaterally cryptorchid SCE rat testis and Sertoli-cell-only testis from irradiated in utero rats of different ages were analyzed by Northern blot analysis. Each band was normalized by ethidium bromide staining of the gel before transfer (Lower). Results are representative of three independent experiments.
Figure 3
Figure 3
(a) Northern blot analysis of FasL expression in purified populations of rat and mouse germ cells. Results are representative of three experiments. (b) Expression of mRNA for FasL in adult rat testis by ISH analysis with antisense probe under dark-field illumination. The high density of silver grain indicates that the site of FasL expression is in the elongated spermatids. (c) ISH analysis with sense probe under dark-field illumination shows the probe specificity. (50× magnification)
Figure 4
Figure 4
Evaluation of DNA content of different testicular cell types by propidium iodide incorporation. (a–c) The percentage of gated cells with indicated ploidy is shown in brackets. Flow cytometric analysis of FasL expression in the seminiferous epithelium and in activated T lymphocytes. The expression of FasL was determined by biotin-conjugated anti-FasL monoclonal antibody (MFL4). (d–o) The white histograms indicate isotype control and gray histograms indicate the staining with specific antibody. l and m show negative and positive controls for FasL, respectively. Results are representative of three independent experiments.
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