Estrogenic regulation of testicular expression of stem cell factor and c kit Implications in germ cell survival and male fertility

Capsule:
17b-estradiol unbalances the expression of SCF and c-kit in rat seminiferous tubules cultured ex vivo, which is accompanied by reduced proliferation and augmented apoptosis of germ cells.

Authors:
Sara Correia, M.S., Mário R. Alves, M.S., José E. Cavaco, Ph.D., Pedro F. Oliveira, Ph.D., Silvia Socorro, Ph.D.

Volume 102, Issue 1, Pages 299–306

Abstract:

Objective:
To study the effect of estrogens regulating the testicular expression of stem cell factor (SCF) and c-kit.

Design:
Experimental study.

Setting:
University research center.

Animal(s):
Male Wistar rats.

Intervention(s):
Rat seminiferous tubules (SeT) cultured in the presence or absence of 17β-estradiol (E2).

Main Outcome Measure(s):
Expression of SCF and c-kit as well as apoptotic factors, FasL, FasR, Bcl-2, and Bax analyzed via quantitative reverse transcription-polymerase and Western blot; enzymatic activity of apoptosis effector caspase-3 assessed by colorimetric assay; proliferation index in SeT epithelium determined via fluorescent immunohistochemistry of nuclear proliferation marker Ki67.

Result(s):
E2 (100 nM) induced a decrease in c-kit expression while increasing expression of SCF. Altered expression of the SCF/c-kit system relied on apoptosis of germ cells, as evidenced by the up-regulated expression of FasL/FasR, the increased ratio of proapoptotic/antiapoptotic proteins (Bax/Bcl-2), and the augmented activity of caspase-3. Decreased proliferation was also found in SeT in response to E2.

Conclusion(s):
A 100 nM dose of E2 unbalance the SCF/c-kit system, with a crucial impact on germ cell survival and thus male fertility. These findings contribute to our knowledge of the mechanisms underlying male idiopathic infertility associated with hyperestrogenism and open new perspectives on treatment targeting estrogen-signaling mechanisms.

  • Ruili Li

    I wonder if the authors have tested the Ki67 antibody on Western. We used the same antibody for fluorescent immunohistochemistry (IHC) and Western. We found the size of the band in Western is incorrect and fits very well with the Ki67-like protein, which is much smaller than Ki67. The IHC whti this Ki67 antibody showed the same location as Ki67 (nucleus) but incorrect time as it should.

    • Silvia Socorro

      The predicted molecular weight of the proliferation marker ki67 in WB is 359 kDa. This molecular weight is relatively high and requires a precise optimization of the WB technique procedure, namely gel percentage and time of electrotransference, in order to obtain labelling on the correct size.

      • Ruili Li

        Do you mean that you did the Western with the Ki67 antibody on the
        protein extracted from the testes/tissue and got the correct band?
        Congratulations! Will you be able to share your gel picture here and let us know the
        amount of the protein loaded on the gel, please? Thanks

        • Silvia Socorro

          No
          we haven’t did the Western with testis proteins but we have tried it in
          prostate samples. Unfortunately, we couldn’t optimize the protocol to get
          labelling of the 359 kD band.

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