Human Reproduction vol.14 no.7 pp.
1791–1793, 1999
Effect of varicocelectomy on the abnormal retention of
residual cytoplasm by human spermatozoa
A.Zini1, M.Buckspan, M.Jamal and K.Jarvi
Division of Urology, Department of Surgery, Mount Sinai Hospital,
University of Toronto, 600 University Avenue, Suite 455, Toronto,
Ontario, Canada, M5G 1X5
1To
whom correspondence should be addressed
Abnormal retention of cytoplasmic residues by human
spermatozoa is associated with the generation of reactive
oxygen species (ROS) in semen and defective sperm func-
tion. We have examined the effect of varicocelectomy on
the retention of residual cytoplasm by human spermatozoa.
Clinical reports of 43 men who underwent microsurgical
varicocelectomy at our institution during a 1 year period
beginning July 1996 were reviewed. Standard semen para-
meters (concentration, motility and morphology) and resid-
ual cytoplasm retention (monitored by Papanicolaou stain)
were assessed before and 6 months after varicocelectomy.
The percentage of spermatozoa with residual cytoplasm
decreased significantly following varicocelectomy compared
to pre-operatively (25.8 versus 18.1% respectively). The
percentages of motile spermatozoa and normal forms
increased significantly (P J 0.0003, P J 0.005 respectively)
following varicocelectomy (22.6 versus 32.9% and 46.4
versus 54.4% respectively). Our data suggest that varico-
celectomy can improve the disposal of residual sperm
cytoplasm by the testis and/or epididymis in infertile men
with varicocele. These data also suggest that varico-
celectomy reduces the potential for ROS generation by
human spermatozoa in these men.
Key words: cytoplasmic droplet/residual cytoplasm/spermato-
zoa/varicocele/varicocelectomy
Introduction
Spermatogenesis is an orderly process whereby male germ cells
in the testis pass through sequential phases of differentiation to
develop subsequently into mature spermatozoa (Clermont,
1972). During the process of spermiogenesis (differentiation
of the haploid spermatid to a mature spermatozoon), extensive
nuclear remodelling and cytoplasmic reduction occur
(Clermont, 1972). Any residual cytoplasm is then normally
removed from spermatozoa prior to their release from the
germinal epithelium of the testis (spermiation) or during early
epididymal transit (Clermont, 1972; Cooper, 1986). Failure to
release excess cytoplasm results in retention of a cytoplasmic
mass in the mid-piece region of the spermatozoon.
Recent work has shown that spermatozoa with retained
excess cytoplasm have defective sperm function (Aitken et al.,
© European Society of Human Reproduction and Embryology
1994; Gomez et al., 1996; Keating et al., 1997). This excess
cytoplasm contains a number of enzymes, many of which are
probably detrimental to normal sperm function (Huszar and
Vigue, 1993, 1994; Gomez et al., 1996). It has been shown
that the retention of residual sperm cytoplasm is negatively
correlated with sperm motility (Gomez et al., 1996; Zini et al.,
1998). It has also been reported (Gomez et al., 1996) that in
a subset of men with unproven fertility, the retention of residual
sperm cytoplasm is positively correlated with reactive oxygen
species (ROS) production in semen (Gomez et al., 1996). This
has important clinical implications because semen ROS have
been clearly shown to impair multiple aspects of sperm
function, including fertilizing capacity (Aitken et al., 1989,
1991; Iwasaki and Gagnon, 1992; Aitken and Fisher, 1994).
The effect of adult varicocelectomy on male fertility remains
controversial. However, it is reported that overall, varicocele
repair results in improved semen quality in 60–80% of infertile
men (Schlesinger et al., 1993). These data suggest that sperma-
togenesis is improved following varicocele repair. However, it
is not known whether varicocele repair can specifically influ-
ence the normal disposal of residual sperm cytoplasm in the
testis and/or epididymis. As such, the purpose of this study
was to examine the possible effect of varicocelectomy on the
retention of residual sperm cytoplasm.
Materials and methods
The clinical reports were reviewed of men presenting for infertility
evaluation at our andrology clinic from July 1996 to July 1997. A
total of 43 consecutive men who underwent varicocelectomy for
clinically detectable varicocele was identified. None of the patients
was azoospermic or had evidence of genital infection. All men
underwent microsurgical subinguinal varicocelectomy as previously
described (Goldstein et al., 1992). All procedures were performed by
the same surgeon (A.Z.). The pre-operative evaluation included a
history, physical examination, scrotal ultrasound and a standard semen
analysis including a Papanicolaou smear. At 6 months (range 4–8)
after varicocelectomy another semen analysis was obtained.
Semen samples were obtained by masturbation after 3–5 days of
sexual abstinence. After liquefaction of semen, standard semen
parameters (volume, concentration, motility, morphology) were
obtained according to World Health Organization (WHO, 1992)
guidelines. Retention of residual cytoplasm was assessed by examina-
tion of the Papanicolaou sperm smears microscopically (magnification
3400) for the presence of light blue-green cytoplasmic staining along
the sperm mid-piece or tail. A mid-piece cytoplasmic droplet greater
than one-third the size of the sperm head was considered significant
(WHO, 1992). A minimum of 200 spermatozoa was examined per
sample. The evaluation of semen analyses (including assessment of
sperm smears) was blinded.
Differences between the pre- and post-varicocelectomy parameters
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A.Zini et al.
Figure 1. Photomicrograph of a Papanicolaou sperm smear
demonstrating one spermatozoon with retained cytoplasm (a: arrow)
and one spermatozoon without residual cytoplasm (b). Bar 5 5
µm.
Table I. Semen parameters before and after microsurgical varicocelectomy
Pre-operative Post-operative P value
% spermatozoa with residual 25.8 6 11.8 18.1 6 9.2 0.0003a
cytoplasm
Sperm concentration (3106/ml) 34.8 6 41.4 33.6 6 55.9 0.89b
% motile spermatozoa 22.6 6 11.0 32.9 6 19.2 0.003a
% normal forms (WHO, 1992) 46.4 6 14.7 54.4 6 14.5 0.005b
Values are expressed as means 6 SD.
aPaired t-test.
bWilcoxon signed-ranks test.
were estimated by parametric and non-parametric tests as appropriate.
Pearson’s correlation coefficient was used to evaluate the association
between individual parameters (SAS Institute, Cary, NC, USA).
Results
Of the 43 men who underwent varicocelectomy, 30 were
evaluable. Of these, 17 underwent left and 13 bilateral varico-
celectomy. The mean age of the 30 men was 34.5 years (range
28–46) with a mean duration of infertility of 2.2 years (range
1–8); 24 men presented with primary infertility and six had
secondary infertility. The 13 men excluded from this study did
not return for their post-varicocelectomy semen analysis.
An example of a spermatozoon with retained cytoplasm is
shown in Figure 1. The mean (6SD) percentages of spermato-
zoa with residual cytoplasm before and at a mean of 6 months
following microsurgical varicocelectomy are shown in Table
I (25.8 6 11.8 versus 18.1 6 9.2% respectively; P 5 0.0003).
The percentage of motile spermatozoa increased significantly
following varicocelectomy (P 5 0.003). In addition, the
percentage of normal forms (WHO, 1992) increased signific-
antly after varicocelectomy (P 5 0.005). However, the sperm
concentration did not change significantly following varico-
celectomy (Table I).
We detected a significant negative correlation between the
percentage of spermatozoa with residual cytoplasm and sperm
motility (r 5 –0.36, P 5 0.02). We also observed a significant
positive correlation between the percentage of spermatozoa
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with residual cytoplasm (prior to varicocelectomy) and the
magnitude of reduction in cytoplasm retention following vari-
cocelectomy (r 5 0.67, P , 0.01).
Discussion
The association between a clinical varicocele and impaired
spermatogenesis is well described (Dubin and Hotchkiss, 1969;
Johnsen and Agger, 1978; Terquem and Dadoune, 1981). Most
studies have reported varying degrees of hypospermatogenesis,
as well as Sertoli cell changes and premature sloughing of germ
cells into the seminiferous tubule lumen. These histological
changes are manifested by the well-described semen abnormal-
ities associated with varicocele (Macleod, 1965). Although
several studies have reported improved semen parameters and
pregnancy rates after varicocele repair (Laven et al., 1992;
Madgar et al., 1995), other studies have not shown a beneficial
effect (Nilsson et al., 1979; Nieschlag et al., 1998). Thus, the
effect of varicocelectomy on male fertility potential remains
controversial.
The present study was designed to evaluate the ability of
varicocelectomy to improve the normal disposal of residual
sperm cytoplasm by the testis and/or epididymis. We have
observed a decrease in the mean percentage of spermatozoa
with residual cytoplasmic droplets following microsurgical
varicocelectomy. This was also accompanied by an increase
in the percentage of sperm motility and the percentage of
normal forms (WHO, 1992). Based on work by Gomez et al.
(1996), the observed reduction in residual sperm cytoplasm
retention would translate into improved sperm function (i.e.
increased sperm motility) and a decreased potential to generate
ROS and other cytotoxic products (Gomez et al., 1996). In
theory, this may also reduce the risk of oxidative spermatozoa
DNA damage and the possible transmission of genetic
mutations (Twigg et al., 1998).
There is preliminary evidence to suggest that varicocele is
associated with increased production of reactive oxygen species
(ROS). It has been reported (Wesse et al., 1993) that fertile
men with varicocele have an increased level of stimulated
ROS production in semen compared to men without varicocele
(Wesse et al., 1993). It has been demonstrated (Lenzi et al.,
1993) that men with varicocele and germ-free genital tract
inflammation can benefit from antioxidant therapy (gluta-
thione), suggesting that these men produce high amounts of
ROS (Lenzi et al., 1993). These authors have also shown that
it is specifically the oxidative sperm membrane damage that
is partially reversed by glutathione therapy in men with
varicocele (Lenzi et al., 1994).
There is indirect and direct evidence to suggest that mon-
itoring the retention of residual sperm cytoplasm may be used
to predict fertility in vitro and in vivo. It has been shown that
the retention of residual sperm cytoplasm is correlated with
ROS in semen, and that these ROS levels are negatively
correlated with sperm fertilizing capacity (Aitken et al., 1991;
Alvarez et al., 1996). It has been demonstrated that semen
ROS levels are negatively correlated with hamster oocyte
penetration test scores and pregnancy rates in vivo (Aitken
et al., 1991). Recently, it has been demonstrated that the
 Varicocelectomy and sperm cytoplasm retention

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