Human Reproduction vol.5 no. 1 pp.89-93, 1990
Quantitative evaluation of spermatogenesis by testicular histology in men with congenital absence of the vas deferens undergoing epididymal sperm aspiration
Sherman J.Silber1, Pasquale Patrizio2 and Ricardo H.Asch2 St Luke's Hospital, St Louis, MO and 2Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, University of California at Irvine, Orange, CA, USA 'To whom correspondence should be addressed
A quantitative histological evaluation of testicular biopsy in patients undergoing microsurgical epididymal sperm retrieval for in-vitro fertilization revealed no correlation between the quantity or quality of spermatozoa retrieved and the actual testicular sperm production. Poor retrieval of sperm was associated with blockage induced by secondary pressure damage in the rete testis collecting area. Key words: aspiration/azoospermia/obstnicUve/speiTn/spermatogenesis Introduction Clinical experience has confirmed that spermatogenesis appears to be 'normal' in men with chronic obstructive azoospermia (Silber, 1984, 1986, 1989a,b). Nonetheless, despite technically accurate microsurgical bypass operations, many men fail to recover adequate sperm counts. Furthermore, in patients with a congenital absence of the vas, when sperm are aspirated directly from the epididymis for the purpose of in-vitro fertilization, some patients yield high numbers of motile spermatozoa, and others yield very low numbers of motile or non-motile spermatozoa (Silber et al.y 1987, 1988; Silber, 1989a). We wished to determine whether quantitative production of spermatozoa as evaluated by testicular histology could, in any way, predict those patients who are more likely to have a successful microsurgical vasoepididymostomy, or yield high numbers of motile spermatozoa for subsequent in-vitro fertilization procedures (Silber, 1989a,b; Silber etai, 1987, 1988). It is now well documented that quantitative evaluation of die number of mature spermatids per tubule by testicular histology is a very accurate reflection of sperm production and, indeed, sperm count in the unobstructed human (Heller and Clermont, 1964; Steinberger and Tjioe, 1968; Zukerman et al., 1978; Silber and Rodriguez-Rigau, 1981). An exponential power curve equation y = a(x?) yields a simple graph to determine sperm production and expected sperm count based upon die number of mature spermatids per tubule (Figure 1). In die present study, we wished to compare the quantitative production of spermatozoa based on a testicular biopsy in patients with congenital absence of the vas deferens undergoing © Oxford University Press
Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
epididymal sperm aspiration for in-vitro fertilization, with the following two objectives. Firsdy, to determine whether long-term obstruction results in any diminution in overall spermatogenesis, and secondly, to determine whether the quantitative testicular sperm production correlates with the amount or quality of spermatozoa obtained from these patients. Methods Thirty-two patients witJi congenital absence of the vas deferens underwent direct microsurgical aspiration of epididymal or vasa efferentia spermatozoa for attempted in-vitro fertilization of their wife's oocytes. Fluid was aspirated microsurgically from die distal epididymis first and then more and more proximally until > 10% sperm motility was observed. Fertilization was attempted with the distal-most sperm that exhibited motility. Much greater motility was always observed in the proximal-most caput epididymis (or even the vasa efferentia) than in the distal epididymis. The technique for microsurgical aspiration of epididymal sperm involved a standard scrotal incision opening the tunica albuginea and exposing the entire length of congenitally obstructed epididymis. Under 40 X magnification, an opening was made in the epididymal tunic and the tubule exposed. A tiny longitudinal slit was then made in the epididymal tubule and a small pipette was used to aspirate fluid from the opening. Fluid was aspirated from successively more proximal segments of the epididymal tubule which in most cases led to the proximal caput or vasa efferentia. The total quantity of spermatozoa obtained, as well as the percentage motility and the grade of motility, were accurately recorded. Sperm motility was graded with a classical 0 to 4 system, and verified using computerized video-monitoring (CELL-SOFT Series 300 System, CRYO Resources Ltd, New York). Grade 0 spermatozoa had no motility. Grade 1 spermatozoa vibrated in place only, with no forward progression and grade 2 spermatozoa demonstrated slow forward progression with poor linearity. Grade 3 spermatozoa demonstrated forward progression with good linearity, but low velocity and grade 4 sperm showed good linearity and rapid velocity. A linearity (as defined by CELLSOFT criteria) of < 3.0 correlated with what was viewed as grade 1 motility and a linearity of > 3.0 was viewed as grade 2 motility. Of the motile spermatozoa observed, sperm velocities ranged from 15.9 fimJs to 57.6 /tm/s and linearity from 2.1 to 5.75. Twenty-two of these patients randomly underwent testicular biopsies as part of the procedure to determine quantitatively total spermatogenesis. A technique for quantitative evaluation of the 89
SJ.SUber, P.Patrizio and R.H.Asch QUANTITATIVE TESTICLE BIOPSY AND SPEflM COUNT ttttn
bfamv
rnimbtr of nature
tubule CorrttUtai Rt
60
SO 40 30 20
* Ofioosponnic non-oluUiuctBd •PotJ-openflw obttrudsd pjflintj — R t g n u t a i BIM
10
• I •I • I i I i I • I i I i I i I • I • I 10 I
20
ipwrn count i10*/ct
30
40
50
60
70
90
100 110
I
Fig. 1. Exponential curve correlating the count of mature spermatids in the testicle biopsy to the sperm count in the semen analysis (Silber and Rodriguez-Rigau, 1981).
Fig. 2. Section of seminiferous tubule with over 50 mature spermatids per tubule (250 x).
90 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
Evaluation of spermatogenesis
Fig. 3. Section of seminiferous tubules with five mature spermatids and six pachytene spermatocytes seen at upper left, and over 30 mature spermatids and 13 pachytene spermatocytes at right (400x). biopsy has been previously reported (Heller and Clermont, 1964; Steinberger and Tjioe, 1968; Zukerman et al., 1978; Silber et al., 1981). The total number of mature spermatids (ScSd) from stages I, II, V and VI were counted in a total of 20 or more tubules. The total number of mature spermatids (oval forms) was then divided by the number of tubules counted in order to determine the number of mature spermatids per tubule (see Figures 2 and 3). The total number of pachytene spermatocytes per tubule was also determined and the ratio of mature spermatids to pachytene spermatocytes was calculated in order to ascertain if any deviation from the normal maturation ratio had occurred in these chronically obstructed cases. Results It is intriguing to note that the frequency distribution of sperm production as determined from testicular biopsy was considerably lower than what one would expect in a normal population of patients (MacLeod and Gold, 1951; Smith et al., 1977; Zukerman et al., 1977). None the less, all of the 22 testicle biopsies examined demonstrated adequate spermatogenesis that would generally be classified within a 'normal range' for each individual patient, despite up to 40 years of congenital epididymal obstruction.
TaWe I. Lack of relationship of testicular sperm production (determined by quantitative testicle biopsy) to quality of epididyma] spermatozoa or• to the fertilization rate Sperm production (determined from biopsy) 5-10 11-20 21-30 31-40 >40
x x x x x
10* 10* 10* 10* 10*
No. of patients
Sperm with forward progression recovered
Fertilization
Pregnancy
3 8 7 2 2
3 (100%) 3 (38%) 5 (71%) 1 (50%) 1 (50%)
2 (67%) 4 (50%) 4 (57%) 1 (50%) 1 (50%)
0 (0%) 2 (25%) 4 (57%) 1 (50%) 0 (0%)
Table I demonstrates the lack of relationship between testicular sperm production and the quality of epididymal spermatozoa which could be retrieved, or to the fertilization rate obtained with those spermatozoa. There was no significant improvement in the percentage of spermatozoa with forwardly progressive motility recovered from the epididymis in patients with greater quantitative sperm production. Greater sperm production observed on testicular biopsy had no correlation with the fertilization rate or pregnancy rate in these patients. Table II gives detailed examples of quantitative sperm production compared to the quantity of spermatozoa retrievable
91 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
SJ.SUber, P.Patririo and R.H.Asch
from the epididymis. It is evident that there is no correlation between the testicular rate of sperm production and the number of spermatozoa retrievable from epididymal aspiration. There is also no correlation between the percentage or quality of motility and total testicular sperm production. Furthermore, as indicated
Table n . Lack of relationship of testicular sperm production rate to quantity of epididymal sperm retrieved Mature spermatid per tubule(s)
Quantity Pachytene P/S spermatocytes (P) sperm production per tubule (X10 6 )
17.4 20.2 22.5 21.9 23.4 23.0 23.6 23.0 26.9 22.3 28.1 27.3 27.3 27.9
7 11 16 16 17 18 19 20 23 25 26 28 28 29
25.4 20.3 20.3 25.3 23.9 23.4 22.8 24.7 31.4 26.1 23.0 32.0 22.4 25.2
1.46 1.00 0.90 1.10 1.02 0.97 0.97 0.93 1.17 0.85 0.82 1.17 0.82 0.90
No. of spermatozoa retrieved Motility Prog. Count (xlO 6 ) 10% 1% 5% 10% 30% 30% 1% 1% 1% 30% 10% 10% 10% 20%
10 13 13 24
480 52 47 1 59 36 31 37 101 119
by the ratio of pachytene spermatocytes to mature spermatids, there is no deviation from normal maturation patterns. Figure 4 demonstrates that in cases where no spermatozoa, or poor quantities of non-motile spermatozoa were all that could be retrieved from the epididymis, sectioning of the proximal vasa efferentia and rete testis revealed leakage of sperm into the interstitium with proximal secondary obstruction. Thus, poor sperm retrieval was not associated with reduced spermatogenesis but rather with secondary pressure-induced inspissation and obstruction in the rete testis collecting system.
Discussion As has been previously demonstrated, using an exponential curve summarized in Figure 1, the number of mature spermatids per tubule correlates very closely with the expected sperm count in non-obstructed patients (Silber and Rodriguez-Rigau, 1981; Silber, 1989). The reason that a simple count of the number of mature spermatids per tubule on a histological section can give an accurate indication of the number of spermatozoa expected in the semen is that the rate of spermatogenesis is constant in any species, including man. When men have low sperm counts or high sperm counts, it is not because they are producing spermatozoa at a slower or faster rate; there are simply fewer cells. Thus, by counting the number of spermatids in a fixed tissue specimen, accurate kinetic information can be obtained that
Fig. 4. Section through vasa efferentia packed with chronically obstructed spermatozoa, and showing leakage into the interstitium resulting in secondary blockage (lOOx).
92 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
Evaluation of spermatogeoesis
correlates well with the patient's expected sperm count if he were not obstructed. This correlation was maintained in our series of patients undergoing vasectomy-reversal for obstructive azoospermia (Silber, 1989a,b). It seemed likely, therefore, that this method of quantifying spermatogenesis in men with congenital absence of the vas deferens might give some prediction of the quantity of spermatozoa retrievable from the epididymis. This, however, was not the case. Ironically, it is the scattered, mosaic arrangement of the various stages of spermatogenesis in the human seminiferous tubule (as opposed to the orderly wave of the spermatogenesis moving across the seminiferous tubule in most other species) that allows such simple quantitation of the human testicular biopsy. In rats, a cut through any particular seminiferous tubule will show only one particular stage of spermatogenesis (Zukerman et al., 1978; Steinberger and Tjioe, 1968). In the human, however, a single cut through one seminiferous tubule will demonstrate several different stages of spermatogenesis. Thus, in humans, very few tubules are required to obtain a good statistical sample of the total range of spermatogenesis in the entire testicle. In every case in which we have performed epididymal sperm aspiration, either no spermatozoa or poor-quality spermatozoa were retrieved from the distal epididymis and better quality and greater numbers of spermatozoa were obtained from more proximal locations. In many cases, aspiration had to be carried out as far proximally as the vasa efferentia. Despite going to this most proximal location, we frequently found a discrepancy between the number of motile spermatozoa which could be retrieved and the testicular sperm production as determined by testicular biopsy. This discrepancy in the number of spermatozoa retrievable from the epididymis or vasa efferentia appeared not to be related to sperm production but rather to pressure-induced perforations and secondary congestion and inspissation in the proximal drainage area of the testicle, i.e. the rete testis. This finding was consistent with our previous findings in cases of vasoepididymostomy (Silber, 1978). Decreased retrieval of spermatozoa in such cases is related to secondary, proximal blockages in the rete testis or vasa efferentia region.
Silber,S.J. (1978) Epididymal extravasation following vasectomy as a cause for failure of vasectomy reversal. Fertil. Sterii, 31, 309—315. Silber.S.J. (1984) Microsurgery for vasectomy reversal and vasoepididymostomy. Urology, 23, 505—524. Silber,S.J. (1986) Diagnosis and treatment of obstructive azoospermia. In Santen.R.J. and Swerdloff,R.S. (eds), Male Reproductive Dysfunction. Marcel Dekker, New York, pp. 479-517. Silber.S.J. (1989a) Results of microsurgical vasoepididymostomy: role of epididymis in sperm maturation. Hum. Reprod, 4, 298—303. Silber.S.J. (1989a) Results of microsurgical vasoepididymostomy: role of epididymis in sperm maturation. Hum. Reprod., 4, 298—303. Silber.S.J. (1989b) Pregnancy after vasovasostomy: a study of factors affecting long-term return of fertility in 282 patients followed for 10 years. Hum. Reprod., 4, 318-322. Silber.S.J. and Rodriguez-Rigau.L.J. (1981) Quantitative analysis of testicle biopsy: determination of partial obstruction and prediction of sperm count after surgery for obstruction. Fertil SteriL, 36,480—495. Silber.S.J., Ord.T., Borrero.C, BalmacedaJ. and Asch.R.H. (1987) New treatment for infertility due to congenital absence of the vas deferens. Lancet, ii, 850-851. Silber.S.J., Balmaceda.J., Borrero.C., Ord.T. and Asch.R.H. (1988) Pregnancy with sperm aspiration from the proximal head of the epidklymis: a new treatment for congenital absence of the vas deferens. Fertil. SteriL, 50, 525-528. Smith.K.D., Rodriguez-Rigau.L.J. and Steinberger.E. (1977) Relation between indices of semen analyses and pregnancy rates in infertile couples. Fertil. SteriL, 28, 1314-1319. Steinberger.E. and Tjioe.D.Y. (1968) A method of quantitative analysis of human seminiferous epithelium. Fertil. SteriL, 19, 960. Zukerman,Z., Rodriguez-Rigau.L.J., Smith.K.D. and Steinberger.E. (1977) Frequency distribution in sperm counts in fertile and infertile males. Fertil. SteriL, 28, 1310-1313. Zukerman,Z., Rodriguez-Rigau.L.J., Weiss,D.B., Chowdhury.L.J., Smith.K.D. and Steinberger.E. (1978) Quantitative analysis of the seminiferous epithelium in human testicle biopsies and the relation of spermatogenesis to sperm density. Fertil. SteriL, 30, 448. Received on April 11, 1989; accepted on September 14, 1989
The reason for doing such a study is that there is an obvious correlation between quantitative production of sperm as determined by testicular biopsy and the daily sperm production rate, as well as with the sperm count in men without obstructive azoospermia. The striking paucity of spermatozoa in the epididymis of some of these men with long-term obstruction must be due either to a diminished sperm production or a secondary rete blockage caused by pressure build-up. Our study shows that although there is a general decrease in sperm production with long-term congenital blockage, it is not as significant a problem in these patients as is secondary obstruction in the rete testis.
References Heller.C.G. and Clermont.Y. (1964) Kinetics of the germinal epithelium in man. Rec. Prog. Hormone Res., 20, 545. MacLeod,J. and Gold.R.Z. (1951) The male factor in fertility and infertility. II. Spermalozoan counts in 1000 men of known fertility and in 1000 cases of infertile marriage. J. UroL, 66, 436.
93 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
Quantitative evaluation of spermatogenesis by testicular histology in men with congenital absence of the vas deferens undergoing epididymal sperm aspiration
Sherman J.Silber1, Pasquale Patrizio2 and Ricardo H.Asch2 St Luke's Hospital, St Louis, MO and 2Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, University of California at Irvine, Orange, CA, USA 'To whom correspondence should be addressed
A quantitative histological evaluation of testicular biopsy in patients undergoing microsurgical epididymal sperm retrieval for in-vitro fertilization revealed no correlation between the quantity or quality of spermatozoa retrieved and the actual testicular sperm production. Poor retrieval of sperm was associated with blockage induced by secondary pressure damage in the rete testis collecting area. Key words: aspiration/azoospermia/obstnicUve/speiTn/spermatogenesis Introduction Clinical experience has confirmed that spermatogenesis appears to be 'normal' in men with chronic obstructive azoospermia (Silber, 1984, 1986, 1989a,b). Nonetheless, despite technically accurate microsurgical bypass operations, many men fail to recover adequate sperm counts. Furthermore, in patients with a congenital absence of the vas, when sperm are aspirated directly from the epididymis for the purpose of in-vitro fertilization, some patients yield high numbers of motile spermatozoa, and others yield very low numbers of motile or non-motile spermatozoa (Silber et al.y 1987, 1988; Silber, 1989a). We wished to determine whether quantitative production of spermatozoa as evaluated by testicular histology could, in any way, predict those patients who are more likely to have a successful microsurgical vasoepididymostomy, or yield high numbers of motile spermatozoa for subsequent in-vitro fertilization procedures (Silber, 1989a,b; Silber etai, 1987, 1988). It is now well documented that quantitative evaluation of die number of mature spermatids per tubule by testicular histology is a very accurate reflection of sperm production and, indeed, sperm count in the unobstructed human (Heller and Clermont, 1964; Steinberger and Tjioe, 1968; Zukerman et al., 1978; Silber and Rodriguez-Rigau, 1981). An exponential power curve equation y = a(x?) yields a simple graph to determine sperm production and expected sperm count based upon die number of mature spermatids per tubule (Figure 1). In die present study, we wished to compare the quantitative production of spermatozoa based on a testicular biopsy in patients with congenital absence of the vas deferens undergoing © Oxford University Press
Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
epididymal sperm aspiration for in-vitro fertilization, with the following two objectives. Firsdy, to determine whether long-term obstruction results in any diminution in overall spermatogenesis, and secondly, to determine whether the quantitative testicular sperm production correlates with the amount or quality of spermatozoa obtained from these patients. Methods Thirty-two patients witJi congenital absence of the vas deferens underwent direct microsurgical aspiration of epididymal or vasa efferentia spermatozoa for attempted in-vitro fertilization of their wife's oocytes. Fluid was aspirated microsurgically from die distal epididymis first and then more and more proximally until > 10% sperm motility was observed. Fertilization was attempted with the distal-most sperm that exhibited motility. Much greater motility was always observed in the proximal-most caput epididymis (or even the vasa efferentia) than in the distal epididymis. The technique for microsurgical aspiration of epididymal sperm involved a standard scrotal incision opening the tunica albuginea and exposing the entire length of congenitally obstructed epididymis. Under 40 X magnification, an opening was made in the epididymal tunic and the tubule exposed. A tiny longitudinal slit was then made in the epididymal tubule and a small pipette was used to aspirate fluid from the opening. Fluid was aspirated from successively more proximal segments of the epididymal tubule which in most cases led to the proximal caput or vasa efferentia. The total quantity of spermatozoa obtained, as well as the percentage motility and the grade of motility, were accurately recorded. Sperm motility was graded with a classical 0 to 4 system, and verified using computerized video-monitoring (CELL-SOFT Series 300 System, CRYO Resources Ltd, New York). Grade 0 spermatozoa had no motility. Grade 1 spermatozoa vibrated in place only, with no forward progression and grade 2 spermatozoa demonstrated slow forward progression with poor linearity. Grade 3 spermatozoa demonstrated forward progression with good linearity, but low velocity and grade 4 sperm showed good linearity and rapid velocity. A linearity (as defined by CELLSOFT criteria) of < 3.0 correlated with what was viewed as grade 1 motility and a linearity of > 3.0 was viewed as grade 2 motility. Of the motile spermatozoa observed, sperm velocities ranged from 15.9 fimJs to 57.6 /tm/s and linearity from 2.1 to 5.75. Twenty-two of these patients randomly underwent testicular biopsies as part of the procedure to determine quantitatively total spermatogenesis. A technique for quantitative evaluation of the 89
SJ.SUber, P.Patrizio and R.H.Asch QUANTITATIVE TESTICLE BIOPSY AND SPEflM COUNT ttttn
bfamv
rnimbtr of nature
tubule CorrttUtai Rt
60
SO 40 30 20
* Ofioosponnic non-oluUiuctBd •PotJ-openflw obttrudsd pjflintj — R t g n u t a i BIM
10
• I •I • I i I i I • I i I i I i I • I • I 10 I
20
ipwrn count i10*/ct
30
40
50
60
70
90
100 110
I
Fig. 1. Exponential curve correlating the count of mature spermatids in the testicle biopsy to the sperm count in the semen analysis (Silber and Rodriguez-Rigau, 1981).
Fig. 2. Section of seminiferous tubule with over 50 mature spermatids per tubule (250 x).
90 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
Evaluation of spermatogenesis
Fig. 3. Section of seminiferous tubules with five mature spermatids and six pachytene spermatocytes seen at upper left, and over 30 mature spermatids and 13 pachytene spermatocytes at right (400x). biopsy has been previously reported (Heller and Clermont, 1964; Steinberger and Tjioe, 1968; Zukerman et al., 1978; Silber et al., 1981). The total number of mature spermatids (ScSd) from stages I, II, V and VI were counted in a total of 20 or more tubules. The total number of mature spermatids (oval forms) was then divided by the number of tubules counted in order to determine the number of mature spermatids per tubule (see Figures 2 and 3). The total number of pachytene spermatocytes per tubule was also determined and the ratio of mature spermatids to pachytene spermatocytes was calculated in order to ascertain if any deviation from the normal maturation ratio had occurred in these chronically obstructed cases. Results It is intriguing to note that the frequency distribution of sperm production as determined from testicular biopsy was considerably lower than what one would expect in a normal population of patients (MacLeod and Gold, 1951; Smith et al., 1977; Zukerman et al., 1977). None the less, all of the 22 testicle biopsies examined demonstrated adequate spermatogenesis that would generally be classified within a 'normal range' for each individual patient, despite up to 40 years of congenital epididymal obstruction.
TaWe I. Lack of relationship of testicular sperm production (determined by quantitative testicle biopsy) to quality of epididyma] spermatozoa or• to the fertilization rate Sperm production (determined from biopsy) 5-10 11-20 21-30 31-40 >40
x x x x x
10* 10* 10* 10* 10*
No. of patients
Sperm with forward progression recovered
Fertilization
Pregnancy
3 8 7 2 2
3 (100%) 3 (38%) 5 (71%) 1 (50%) 1 (50%)
2 (67%) 4 (50%) 4 (57%) 1 (50%) 1 (50%)
0 (0%) 2 (25%) 4 (57%) 1 (50%) 0 (0%)
Table I demonstrates the lack of relationship between testicular sperm production and the quality of epididymal spermatozoa which could be retrieved, or to the fertilization rate obtained with those spermatozoa. There was no significant improvement in the percentage of spermatozoa with forwardly progressive motility recovered from the epididymis in patients with greater quantitative sperm production. Greater sperm production observed on testicular biopsy had no correlation with the fertilization rate or pregnancy rate in these patients. Table II gives detailed examples of quantitative sperm production compared to the quantity of spermatozoa retrievable
91 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
SJ.SUber, P.Patririo and R.H.Asch
from the epididymis. It is evident that there is no correlation between the testicular rate of sperm production and the number of spermatozoa retrievable from epididymal aspiration. There is also no correlation between the percentage or quality of motility and total testicular sperm production. Furthermore, as indicated
Table n . Lack of relationship of testicular sperm production rate to quantity of epididymal sperm retrieved Mature spermatid per tubule(s)
Quantity Pachytene P/S spermatocytes (P) sperm production per tubule (X10 6 )
17.4 20.2 22.5 21.9 23.4 23.0 23.6 23.0 26.9 22.3 28.1 27.3 27.3 27.9
7 11 16 16 17 18 19 20 23 25 26 28 28 29
25.4 20.3 20.3 25.3 23.9 23.4 22.8 24.7 31.4 26.1 23.0 32.0 22.4 25.2
1.46 1.00 0.90 1.10 1.02 0.97 0.97 0.93 1.17 0.85 0.82 1.17 0.82 0.90
No. of spermatozoa retrieved Motility Prog. Count (xlO 6 ) 10% 1% 5% 10% 30% 30% 1% 1% 1% 30% 10% 10% 10% 20%
10 13 13 24
480 52 47 1 59 36 31 37 101 119
by the ratio of pachytene spermatocytes to mature spermatids, there is no deviation from normal maturation patterns. Figure 4 demonstrates that in cases where no spermatozoa, or poor quantities of non-motile spermatozoa were all that could be retrieved from the epididymis, sectioning of the proximal vasa efferentia and rete testis revealed leakage of sperm into the interstitium with proximal secondary obstruction. Thus, poor sperm retrieval was not associated with reduced spermatogenesis but rather with secondary pressure-induced inspissation and obstruction in the rete testis collecting system.
Discussion As has been previously demonstrated, using an exponential curve summarized in Figure 1, the number of mature spermatids per tubule correlates very closely with the expected sperm count in non-obstructed patients (Silber and Rodriguez-Rigau, 1981; Silber, 1989). The reason that a simple count of the number of mature spermatids per tubule on a histological section can give an accurate indication of the number of spermatozoa expected in the semen is that the rate of spermatogenesis is constant in any species, including man. When men have low sperm counts or high sperm counts, it is not because they are producing spermatozoa at a slower or faster rate; there are simply fewer cells. Thus, by counting the number of spermatids in a fixed tissue specimen, accurate kinetic information can be obtained that
Fig. 4. Section through vasa efferentia packed with chronically obstructed spermatozoa, and showing leakage into the interstitium resulting in secondary blockage (lOOx).
92 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
Evaluation of spermatogeoesis
correlates well with the patient's expected sperm count if he were not obstructed. This correlation was maintained in our series of patients undergoing vasectomy-reversal for obstructive azoospermia (Silber, 1989a,b). It seemed likely, therefore, that this method of quantifying spermatogenesis in men with congenital absence of the vas deferens might give some prediction of the quantity of spermatozoa retrievable from the epididymis. This, however, was not the case. Ironically, it is the scattered, mosaic arrangement of the various stages of spermatogenesis in the human seminiferous tubule (as opposed to the orderly wave of the spermatogenesis moving across the seminiferous tubule in most other species) that allows such simple quantitation of the human testicular biopsy. In rats, a cut through any particular seminiferous tubule will show only one particular stage of spermatogenesis (Zukerman et al., 1978; Steinberger and Tjioe, 1968). In the human, however, a single cut through one seminiferous tubule will demonstrate several different stages of spermatogenesis. Thus, in humans, very few tubules are required to obtain a good statistical sample of the total range of spermatogenesis in the entire testicle. In every case in which we have performed epididymal sperm aspiration, either no spermatozoa or poor-quality spermatozoa were retrieved from the distal epididymis and better quality and greater numbers of spermatozoa were obtained from more proximal locations. In many cases, aspiration had to be carried out as far proximally as the vasa efferentia. Despite going to this most proximal location, we frequently found a discrepancy between the number of motile spermatozoa which could be retrieved and the testicular sperm production as determined by testicular biopsy. This discrepancy in the number of spermatozoa retrievable from the epididymis or vasa efferentia appeared not to be related to sperm production but rather to pressure-induced perforations and secondary congestion and inspissation in the proximal drainage area of the testicle, i.e. the rete testis. This finding was consistent with our previous findings in cases of vasoepididymostomy (Silber, 1978). Decreased retrieval of spermatozoa in such cases is related to secondary, proximal blockages in the rete testis or vasa efferentia region.
Silber,S.J. (1978) Epididymal extravasation following vasectomy as a cause for failure of vasectomy reversal. Fertil. Sterii, 31, 309—315. Silber.S.J. (1984) Microsurgery for vasectomy reversal and vasoepididymostomy. Urology, 23, 505—524. Silber,S.J. (1986) Diagnosis and treatment of obstructive azoospermia. In Santen.R.J. and Swerdloff,R.S. (eds), Male Reproductive Dysfunction. Marcel Dekker, New York, pp. 479-517. Silber.S.J. (1989a) Results of microsurgical vasoepididymostomy: role of epididymis in sperm maturation. Hum. Reprod, 4, 298—303. Silber.S.J. (1989a) Results of microsurgical vasoepididymostomy: role of epididymis in sperm maturation. Hum. Reprod., 4, 298—303. Silber.S.J. (1989b) Pregnancy after vasovasostomy: a study of factors affecting long-term return of fertility in 282 patients followed for 10 years. Hum. Reprod., 4, 318-322. Silber.S.J. and Rodriguez-Rigau.L.J. (1981) Quantitative analysis of testicle biopsy: determination of partial obstruction and prediction of sperm count after surgery for obstruction. Fertil SteriL, 36,480—495. Silber.S.J., Ord.T., Borrero.C, BalmacedaJ. and Asch.R.H. (1987) New treatment for infertility due to congenital absence of the vas deferens. Lancet, ii, 850-851. Silber.S.J., Balmaceda.J., Borrero.C., Ord.T. and Asch.R.H. (1988) Pregnancy with sperm aspiration from the proximal head of the epidklymis: a new treatment for congenital absence of the vas deferens. Fertil. SteriL, 50, 525-528. Smith.K.D., Rodriguez-Rigau.L.J. and Steinberger.E. (1977) Relation between indices of semen analyses and pregnancy rates in infertile couples. Fertil. SteriL, 28, 1314-1319. Steinberger.E. and Tjioe.D.Y. (1968) A method of quantitative analysis of human seminiferous epithelium. Fertil. SteriL, 19, 960. Zukerman,Z., Rodriguez-Rigau.L.J., Smith.K.D. and Steinberger.E. (1977) Frequency distribution in sperm counts in fertile and infertile males. Fertil. SteriL, 28, 1310-1313. Zukerman,Z., Rodriguez-Rigau.L.J., Weiss,D.B., Chowdhury.L.J., Smith.K.D. and Steinberger.E. (1978) Quantitative analysis of the seminiferous epithelium in human testicle biopsies and the relation of spermatogenesis to sperm density. Fertil. SteriL, 30, 448. Received on April 11, 1989; accepted on September 14, 1989
The reason for doing such a study is that there is an obvious correlation between quantitative production of sperm as determined by testicular biopsy and the daily sperm production rate, as well as with the sperm count in men without obstructive azoospermia. The striking paucity of spermatozoa in the epididymis of some of these men with long-term obstruction must be due either to a diminished sperm production or a secondary rete blockage caused by pressure build-up. Our study shows that although there is a general decrease in sperm production with long-term congenital blockage, it is not as significant a problem in these patients as is secondary obstruction in the rete testis.
References Heller.C.G. and Clermont.Y. (1964) Kinetics of the germinal epithelium in man. Rec. Prog. Hormone Res., 20, 545. MacLeod,J. and Gold.R.Z. (1951) The male factor in fertility and infertility. II. Spermalozoan counts in 1000 men of known fertility and in 1000 cases of infertile marriage. J. UroL, 66, 436.
93 Downloaded from https://academic.oup.com/humrep/article-abstract/5/1/89/743651 by University of California, Santa Barbara user on 07 March 2018
