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Morphological
changes of spermatozoa in proximalvas deferens after vasectomy
Ren-Qian
WEN, Shi-Qin LI1, Chun-Xiang WANG1, Qing-Hui WANG,
Mei-Yi LIU
Family
Planning Research Institute of Guangdong, Guangzhou 510600, China
1Family Planning Research Institute of Shenzhen, Shenzhen 518006,
China
Asian J Androl 1999 Jun; 1: 61-65
Keywords: spermatozoa; vas deferens; epididymis; vasectomy; vasovasostomy
Abstract
Aim: To investigate the morphological changes of spermatozoa in the proximal vas deferens after vasectomy. Methods: Proximal vas deferens fluids were collected from 79 fertile men (group A) and 64 vasectomized men (group B) during the operations of vasectomy or vasovasostomy. Sperm morphology in the proximal vas deferens was analyzed after staining with the modified Papanicolaou method. Results: The percentage of spermatozoa with a normal oval head from group B (50.7%21.7%) was significantly lower than that of group A (75.2%11.1%). The data in group A was similar to those of normal semen and therefore represents the physiological condition of the proximal vas deferens sperm of fertile men. There were no significant differences in the percentages of normal oval heads in group B with the time since vasectomy. Conclusion: After vasectomy, the spermatozoa in the proximal vas deferens and epididymis were continuously degenerating and being replenished by spermatozoa coming from testis. The obvious morphological degeneration occurred in the sperm tail and spermatozoa with a small oval head and amorphous heads were increased.1 Introduction
After vasectomy the pressure in the lumen of the epididymis and proximal vas deferens may be increased and their internal environment of them may be changed. Spermatozoa stored in the epididymis and proximal vas deferens may be gradually degenerated[1,2]. There is a good relationship between male fertility and the recovery of sperm morphology and other sperm parameters after vasovasostomy. However, the recovery of sperm morphology after vasectomy reversal is not good[3,4], so that the pregnancy rate is in the range of 50% to 58%[5]. Standardized methods for evaluating sperm morphology have been recommended by World Health Organization[6,7,8]. Sperm morphology in the human proximal vas deferens or epididymis is a research project of sperm biology although the spermatozoa from there are difficult to collect. In order to obtain and analyse spermatozoa from the proximal vas deferens, a method of collecting fluids from the proximal vas deferens of fertile men during vasectomy has been developed and data of spermatozoa with normal oval head and other sperm parameters have been published[9]. Changes of sperm parameters were also found in proximal vas deferens from vasectomized men[10]. Data of sperm morphology in semen (ejaculated sperm) from fertile men and infertile men with varicocele had been obtained in our laboratory[11]. Nowadays more studies of sperm biology are focused on ejaculated spermatozoa and the World Health Organization has published aseries of standard methods for sperm morphology analysis. However, there are few reports of spermatozoa before ejaculation (spermatozoa from the epididymis or proximal vas deferens)[9,10]. In this study, the sperm morphology in the proximal vas deferens and their morphological changes after vasectomy have been compared with the sperm morphology from normal fertile men.2 Materials and methods
Seventy-nine fertile men (group A), who were volunteers for vasectomy, aged from 23-35 a and with two or more children, and 64 vasectomized men (group B) who asked for vasovasostomy and aged from 25-40 a, were investigated. All are Chinese peasants (the Han nationality) and there was no fever for 3 months before. A physical examination was performed for each subject and the contents of the scrotum were examined and noted. Testicular size, which was in normal range (15-25 mL), was measured using a Prader orchidometer.
Vasectomy
was performed under local anesthesia and vasovasostomy under local or
continuous anaesthesia. The left and the right proximal vas fluids were
collected with polyethylene tubing (Portex Polyethylene Tubing, UK) according
to the method of Wen et al[9]. Proximal vas fluid containing
spermatozoa before ejaculation in polyethylene tubing was washed out with
Ham's F10 medium. Sperm concentration (20-50)106/mL
was adjusted and spermatozoa were incubated at 37 for 30 min. A small
drop of medium containing
spermatozoa was placed on a cleared slide and a smear made by the feathering
technique. After fixing in a solution of 95% alcohol and 10-15 min air-drying, the slide
was stained using the modified Papanicolaou method recommended by WHO[7].
The slides were examined and phtographed under a microscope (250) and
at least 200 spermatozoa were observed. According to the WHO method[6,7]
the spermatozoa with normal oval head, large oval head, small oval head,
tapering head, pyriform head, duplicate head, amorphous head, midpiece
defect, tail defect, and cytoplasmic droplet were classified.
3 Results
There was no significant difference between the left and the right proximal vas deferens in the sperm morphology of both normal fertile men (before vasectomy, group A) and vasectomized men (group B). The total numbers (left and right) of proximal vas deferens samples obtained from 79 normal fertile men (group A) and 64 vasectomized men (group B) were 128 and 100, respectively. The data of sperm morphology of group A and group B from the proximal vas deferens were shown in Table 1. The percentage of spermatozoa with a normal oval head from group B (50.7%21.7%) was significantly lower than that of group A (75.2%11.1%, t=11.049, P<0.01). In group B the percentage of spermatozoa with a small head, amorphous head and tail defect were significant higher than those in group A. In group B there were no significantly differences in the frequency of abnormal forms except the frequency of spermatozoa with a pyriform head in group B (17/100, 17%) was significantly decreased when compared with that in group A (42/128, 32.8%, 2=6.517, P<0.05). There were no spermatozoa with an abnormal midpiece or spermatozoa with a cytoplasmic droplet in group B. However, in group A there were 3 cases of abnormal midpiece and 2 cases of cytoplasmic droplet. The highest frequency of abnormal sperm forms was amorphous head and tail defects both in group A and group B.
Table
1. Results of sperm morphology in the proximal vas deferens from normal
fertile men (before vasectomy, group A) and vasectomized men (group B).
| |
group A |
group B |
||||
| means |
rang |
frequency# |
means |
rang |
frequency |
|
| Normal oval head (%) |
75.211.1 (N=128) |
(45-96) |
100% |
50.721.7c (N=100) |
(21-91) |
100% |
| abnormal head |
||||||
| large oval head (%) |
3.33.5 (n=15) |
(1-13) |
11.7% |
5.25.8 (n=5) |
(1-13) |
5% |
| Small oval head (%) |
5.03.7 (n=88) |
(1-19) |
68.8% |
15.316.2c (n=68) |
(1-52) |
68% |
| Tapering head (%) |
3.32.3 (n=6) |
(1-6) |
4.7% |
3.02.6 (n=3) |
(1-6) |
3% |
| Pyriform head (%) |
2.41.7 (n=42) |
(1-10) |
32.8% |
3.12.6 (n=17) |
(1-10) |
17%b |
| Duplicate head (%) |
3.35.1 (n=15) |
(1-19) |
11.7% |
4.87.4 (n=5) |
(1-18) |
5% |
| Amorphous head (%) |
11.18.1 (n=124) |
(1-37) |
96.9% |
16.610.8c (n=92) |
(1-57) |
92% |
| Abnormal midpiece (%) |
13.78.5 (n=3) |
(4-20) |
2.3% |
|
|
|
| Cytoplasmic droplet (%) |
1.50.7 (n=2) |
(1-2) |
1.6% |
|
|
|
| Tail defect (%) |
9.36.5 (n=118) |
(1-27) |
92.2% |
29.523.4c (n=92) |
(1-94) |
92% |
bP<0.05, cP<0.01 when compared with group A. # Frequency was the % of samples with this morphological type the number of abnormal forms (n) over the number of normal oval head (N).
Table 2 gives the results of spermatozoa with a normal oval head in the proximal vas deferens from vasectomized men. There were no significant differences between different times since vasectomy (F=1.836, P>005).
Table
2. The results (meanSD) of sperm morphology (normal oval head %) in
the proximal vas deferens from vasectomized men categorized by the time
since vasectomy#.
| <1 year |
1<2 years |
2<3 years |
3<4 years |
4<5 years |
5<8 years |
810 years |
| 64.424.3 |
57.420.1 |
46.519.2 |
45.921.2 |
38.715.3 |
45.821.0 |
50.524.3 |
| (n=11) |
(n=22) |
(n=13) |
(n=8) |
(n=6) |
(n=26) |
(n=14) |
The
morphological graphs of normal fertile men (group A) and vasectomized
men (group B) were shown in Figures 1-3. The spermatozoa showed in Figures
2 and 3 were taken from subjects who had undergone vasectomy for 3
a and 8 a, respectively.
Figure 1. Normal spermatozoa morphology of proximal vas deferens
(250). This picture was taken from a normal subject who asked for vasectomy.
The spermatozoa exhibit a regular oval head with intact midpiece and uncoiled
tail.
Figure 2. Spermatozoa morphology of proximal vas deferens (250).
This picture was taken from a subject who had undergone vasectomy for
3 a. Most of spermatozoa was tail defect (A: no tail; B: a tail leaving
less than one half its probable length); C: spermatozoa with small head;
D: spermatozoa with amorphous head; E: normal spermatozoa. Some of sperm
head had a normal acrosomal region (E).
Figure 3. Spermatozoa morphology of proximal vas deferens (250).
This picture was taken from a subject who had undergone vasectomy for
8 a. Most of sperm head with abnormal acrosomal region (A). Spermatozoa
with tail defect (B: no tail;
C: a tail leaving less than one half its probable length). D: spermatozoa with amorphous head. E: spermatozoa
with small head.
4
Discussion
In
this investigation, the percentage of spermatozoa with a normal oval head
from the proximal vas deferens of group A (75.2%11.1%) was similar to
that of semen from normal fertile men determined in our previous study
(79.7%12.5%)[11], by other author (66%10.2%)[12]
and by WHO (80.5%9.7%)[6]. The
highest percentage of abnormal forms of spermatozoa from the proximal vas
deferens in group A was the spermatozoa with abnormal midpiece in present
study (13.7%8.5%). Although no spermatozoa in this category from normal
semen was reported both by our previous study[11] and by WHO[6],
2.1%3.5% was reported[12]. The spermatozoa in other abnormal
categories in this investigation were similar to those of normal semen
reported by other investigators[6,11,12]. According to these
data, supported by the photograph in Figure 1, it was suggested that the conditions in group A may
be normal for sperm morphology in the human proximal vas deferens.
It
is reported that after vasectomy the epididymal duct and proximal vas
deferens may be distended[1] and there are more degenerated
spermatozoa in the lumen of the cauda epididymis than that in the caput
epididymis[2]. However, there are no reports about the degeneration
of spermatozoa in the epididymis and proximal vas deferens after vasectomy.
There were no significant differences between time since vasectomy in
the percentage of spermatozoa with normal oval head (Table 2). It is known
that after vasectomy spermatozoa may be continually produced in the testis
and stored in the epididymis and proximal vas deferens, where degradation
of spermatozoa occurs. This study suggested that no matter how long the
duration of vasectomy, the morphological normal spermatozoa in the proximal
vas deferens remain at a constant level. Also, after vasectomy (group
B), the percentage of spermatozoa with small head,
amorphous head and tail defects was significantly higher than those of
normal fertile men (group A). The highest frequency in both groups was
spermatozoa with a small head, an amorphous head and tail defects. Their
frequency was similar to those in normal semen[12], 86.4%,
98.4%, 94.6%, respectively. Tail defects were especially prominent. In
group B there were cases of spermatozoa
with no tail in which only the sperm head could be seen (Figures 2
and 3). Most of the tail defects were no tail or a tail of less
than half its probable length (Figure 2),
and that a few spermatozoa had a severely coiled or double tail. As the
degradation of spermatozoa in the epididymis and proximal vas deferens
progressed, spermatozoa with tail defect, with small and amorphous heads
were significantly increased. Pelfrey et al[3] reported
that after vasectomy reversal, ejaculated spermatozoa with a tail defect
was 18.1%, which was higher than in those of male infertility for other
reasons. In this study, the spermatozoa with tail defects in group B was
29.5% and with a frequency was 92%. From our results and those of Pelfrey
et al[3], it is indicated that after vasectomy one of
the obvious changes in sperm morphology in the
epididymis and proximal vas deferens was the degradation of the sperm
tail. Because of the irreversibility
of sperm morphology affected by epididymal dysfunction after vasovasostomy,
infertility may persist in some subjects. This may explain why there was
a lower pregnancy rate from 50% for conventional techniques to 58% for
microsurgical techniques, after vasectomy reversal[5]. The
defect of sperm morphology after vasovasostomy may be one of the causes
of male infertility after vasectomy reversal.
So far, there is no clear association between morphologically abnormal spermatozoa and clinical conditions. An increase of spermatozoa with an amorphous head is due to an allergic reaction; the increase of spermatozoa with abnormal midpiece is related to the infection and spermatozoa with cytoplasmic droplets indicate disturbances of sperm maturation in the epididymis or a shorter period of abstinence[13]. Huang[14] reported that the number of spermatozoa with a tapering head and with an amorphous head is increased by infection, and the increase of spermatozoa with tapering head and with abnormal midpiece is caused by varicocele. It is well known that immuno-reaction after vasectomy results in anti-sperm antibodies in the serum. Our previous report confirmed that, after vasectomy there were antisperm antibodies on the surface of spermatozoa from the proximal vas deferens (79.4% for IgG, 38.2% for IgA)[15]. It is unknown if vasectomy can cause other sperm morphological changes. The frequency of spermatozoa with a pyriform head in group A (32.8%) was similar to that of our previous study in normal fertile men (33.3%)[11], however, the frequency of spermatozoa in this category (17%) was significantly decreased after vasectomy (group B). No spermatozoa with a cytoplasmic droplet were found in group B. Menchini-Fabris et al[13] reported that spermatozoa with a cytoplasmic droplet indicated either that sperm maturation did not occur in the epididymis or that the interval of sexual abstinence was too brief. Our results supported this because spermatozoa stored in the epididymis and proximal vas deferens after vasectomy for a long time, had the appearance of those ejaculated after a long period of abstinences.
In order to explain the clinical significance of the various type of abnormal spermatozoa in the proximal vas deferens, further research works must be performed to follow up the changes of sperm morphology after vasovasostomy and to evaluate the various abnormal sperm in group B and their relationship to pregnancy after vasectomy reversal.
5
Acknowledgements
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The
project was supported by the State Family Planning Committee.
Correspondence to Dr Ren-Qian WEN, Family Planning Research Institute
of Guangdong, 17#, Mei Dong Road, Dong Shan District, Guangzhou 510600,
China.
Tel: +86-20-8767 5290 Fax: +86-20-8777 7331
E-mail: wenrq@163.net
Received 1999-02-08 Accepted 1999-05-08
