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Normal
and varicocele testis in adolescents
G.
Santoro1, C. Romeo2
1Department
of Biomorphology and Biotechnologies, 2Department of Medical
and Surgical Paediatric Sciences, University of Messina, 98125
Messina, Italy
Asian J Androl 2001 Dec; 3: 259-262
Keywords:
varicocele;
testis; seminiferous tubules; adolescence
Abstract
The authors reviewed the results of their research on the structure and composition of normal and varicocele seminiferous tubules in adolescents. They give new evidences of normal structure of adolescent testis and demonstrate, for the first time, the ultrastructural and immunohistochemical modifications of the lamina propria and basal lamina in the adolescent varicocele patients, which are similar to those observed in adults, but less severe, and of the adherence junctions in seminiferous tubules. They also report the presence of oxidative stress in adolescents limited to testis and not generalised as in the adults. These data are well correlated to different clinical studies that support the hypothesis of a progressive course of varicocele and the need for surgical treatment in adolescent varicocele patients.The
adolescent age plays a key role in the development of testicular
diseases, since the lesions observed are often progressive. For this reason
we have recently focused our attention on the structure and composition
of normal and varicocele seminiferous tubules in the adolescents. The
varicocele is a common disease of the testis and represents one of the
most common causes of male infertility[1].
The
human seminiferous tubules are formed by a very complex stratified epithelium
containing spermatogenic cells in different stages of development, and
supporting cells, the Sertoli cells. The epithelium is surrounded by a
lamina propria composed of a basal layer and 5-7 external cellular layers.
The cellular layers are constituted of 3-5 inner layers of myofibroblasts
and one or more outer layers of fibroblasts. The basal layer, composed
of extracellular matrix (ECM) components, is formed by two layers: the
inner electron-transparent layer, the lamina lucida or rara, is situated
close to the plasma membrane of the germinal epithelium, and the outer
electron-dense layer, the lamina densa. The external cellular layers
are separated by laminae of ECM consisting of glycosaminoglycans (GAGs), proteoglycans
and collagen fibers. The myofibroblasts are individual flat cells with
a diameter of 40-60 m; they do not form continuous cell layers and are
not completely covered by a basal lamina. In the angular interstices between
various seminiferous tubules, there are Leydig cells, which represent
the endocrine component of the testis[2-5].
The
lamina propria of normal adolescent seminiferous tubule showed a maximum
thickness of 10 m and a surface area between 3970 and 4472 m2,
while the tubular diameter
was between 158 and 200 m2. In adolescent affected by varicocele,
we showed different
degrees of thickening of the lamina propria till a maximum value of 35
m. It was the result of an increased deposition of extracellular components,
starting from the innermost layer of collagen fibers extending to the
outer extracellular layer[2]. This condition was responsible
for the formation of deep invaginations facing the germinal epithelium,
which, in other testicular pathologies, were ascribed to tubular damage
caused by a blockage in the mediation of the lamina propria between the
interstitium and the germinal epithelium[6].The increased thickness
of the lamina propria corresponded to a progressive increase in its surface
area and a reduction in the tubular diameter[2]. This damage
was not as severe as that described in adult varicocele, in which a diffuse
sclerosis of the lamina propria was observed[7].
Despite
the modifications of the extracellular layers, the myofibroblasts still maintained
their morphological features with only mild alterations[3].
Moreover, using -smooth muscle isoactin as specific marker of myofibroblasts,
we do not observe
any myofibroblast transformation into fibroblast, as already demonstrated
in adult varicocele, that corresponds to the reported progressive sclerosis
of the lamina propria[7]. The myofibroblasts are responsible
for the contraction of
the seminiferous tubules necessary for the transport of testicular spermatozoa
and fluid and take part in the regulation of spermatogenesis and the creation of
the blood testis barrier[8, 9]. The integrity of myofibroblasts
may be an important
feature of adolescent varicocele.
In
adolescent varicocele patients, the peritubular basal lamina also showed
ultrastructural changes characterized by an uneven profile with a variable
thickness[10]. Similar observations have been reported in adult
varicocele[7] and in other testicular pathology[11,12].
The peritubular basal lamina was also altered in
two of its major components: laminin and collagen type IV[10].
In normal testis laminin is localized in the lamina lucida, while collagen
type IV in both layers
of the peritubular basal lamina; both are present as a continuous and
uniform line[5]. These two molecules, apart from a mechanical
linkage, play a key
role in the regulation of major biological cellular functions[13-15].
Through the relationship with the adhesion receptors of the integrin family
localized in the cellular membrane, and consequently with the intracellular
actin-associated proteins, messages are transmitted from the ECM to the
nuclear compartment, thus controlling the ubiquitous process of differentiation,
proliferation, adhesion, migration, gene expression[16,17]
and spermatogenesis[18,19]. In adolescent varicocele patients,
laminin immunereaction displayed an irregular line with a wavy profile
that sometimes appeared to be interrupted. Collagen type IV showed areas
of annular thickening of the immunereaction alternating with areas of
interrupted and reduced immunopositivity. Moreover, its immunofluorescence
distribution appeared to be irregular and wavy, following the morphology
of the basal lamina observed by transmission electron microscopy along
the deep invaginations[4]. The observed modification in the
morphology and composition of the peritubular basal
lamina could represent one of the mechanisms responsible for the lesions
characteristic of varicocele.
Another
pivotal role in the testicular function is played by two actin-associated
proteins, i.e., vinculin and talin; they are important in cell activity
as previously described[16-19].
Vinculin
and talin have been identified in different tissues and cell types. Particularly,
vinculin is localized at cell-cell and cell-ECM adhesion sites, while talin
is present only at cell-ECM junction[20-22]. They frequently
co-localize within
cells[20] taking part in the formation of the adherens junction.
Miyamoto et al[23] have demonstrated that talin,-actinin
and vinculin represent a unique subset of three cytoskeletal proteins
whose membrane accumulation requires both integrin aggregation and ECM
occupancy, but not the tyrosine-kinase activity needed for many other
integrin-responsive proteins. Alterations of one of these components of
the adherens junction could negatively influence the others. We have demonstrated,
for the first time, that in normal human adolescent testes, these
two actin-associated proteins had the same pattern as that observed in
other tissues. Particularly, in Sertoli and Leydig cells vinculin was
expressed at the cell-cell
and the cell-ECM adherence junctions, while talin was present only at
the cell-ECM adherence junctions level[24]. In the adolescent
varicocele patients, there were slight alterations of vinculin and talin
pattern only in the Sertoli cells, while the Leydig cells presented an
expression and distribution similar to the normal testes. Occasionally,
in the same sample, altered seminiferous tubules were found adjacent to
normal ones. These evidences could represent a not well consolidated and
generalized tubular damage[24]. On the basis of our results,
we speculate that the modifications in two components of the adherence junction,
i. e., basal lamina and actin-associated protein, could negatively influence
the spermatogenesis in varicocele patients. In this regard, Pelletier
et al
have reported an altered peritubular basal lamina and changes in Sertoli
cell junctions in tubular regions where germ cells were depleted[25].
Recently
nitric oxide (NO) has been identified in the testis where it should play
certain roles. NO has been reported to be important in the regulation
of male reproductive function and fertility, Leydig cell function, myofibroblast
contraction and hence tubular peristalsis[26,27]. NO acts also
on the peritubular lamina propria modulating its permeability and regulates
the activity of muscle cells and pericytes of testicular vessels[28].
Nevertheless experimental studies have shown that NO at supra-physiological
levels can be harmful for both testicular and sperm function[29,30].
In normal testis, two isoforms of NO synthases, i.e., endothelial and
inducible nitric oxide synthase (eNOS and iNOS) were demonstrated by immunohistochemistry
and western blot analysis[31]. In adolescent varicocele patients,
the two isoforms were also identified but iNOS appeared to be up-regulated[31].
This important finding could explain the significant increase of
NO recorded in the spermatic veins of adolescent varicocele patients.
It is possible that varicocele causes an up-regulation of the iNOS that
produces increased amount of NO, causing an oxidative stress, which if
long continued will result in testis and sperm dysfunction[32].
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Correspondence
to: Dr.
Giuseppe Santoro, Department of Biomorphology and Biotechnologies,
1st Floor Torre Biologica, Policlinico Universitario, Viale Gazzi, 98125
Messina, Italy.
Tel: +39-90-221 3637
Fax: +39-90-692 449
E-mail:
santorof@unime.it
Received 2001-09-29 Accepted 2001-11-22
