Using RT-PCR, western blot and enzyme and fluorescence immunocytochemical t
echniques, the three isoforms of neurofilament proteins (NFPs), namely NF-L
(NFP-68 kDa), NF-M (NFP-160 kDa) and NF-H (NFP-200 kDa) were found in Sert
oli and Leydig cells of human testes. RT-PCR showed specific for the three
NFP fragments in testicular tissue, in isolated seminiferous tubules and in
isolated Leydig cells. In protein preparations from the same testicular co
mponents, western blot analysis detected bands with molecular weights chara
cteristic for NF-H, NF-M and NF-L. Application of immunofluorescence and im
munoenzyme methods on cryostat and paraffin sections resulted in difference
s in the staining pattern in Sertoli cells and Leydig cells. In these cells
, the NFPs showed predominantly a perinuclear location from which bundles e
merge that were directed towards the basal, apical and lateral extensions o
f the Sertoli cells as well as the periphery of Leydig cells. NF-H coexists
with vimentin-type filaments as seen by dual staining and staining of cons
eccutive serial sections of material embedded in paraffin. In Sertoli cells
, vimentin and NF-H showed distinct dynamic changes depending on the stage
of spermatogenesis and some structural variations of seminiferous tubules.
Although in some tubules both vimentin and NF-H immunoreactivity was presen
t at high levels, in the Sertoli cells from most individuals an inverse rel
ationship in the staining intensity of vimentin and NF-H was observed. The
strongest NF-H immunoreactivity was detected in Sertoli cells associated wi
th stage 3 spermatids, whereas vimentin immunoreactivity was most abundant
in association with stage 5 spermatids. The leydig cells did not show funct
ional changes of the NFP immunoreactivity. The results obtained provide new
evidence for the heterogeneous phenotype of human Sertoli cells and raise
the question of their exact nature and origin.