E IRAQI MEDICAL JOURNAL IN THNC ANDPOSTGRADUATE COPPER CONCENTRATIONS

SEMINAL PLASMA

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Status ! "#$% a$& C ''() C $%($t)at# $s #$ S(*#$a+ P+as*a ! Ma+( I$!()t#+#t, a$& T-(#) C ))(+at# $ .#t- Va)# us S'()* Pa)a*(t()s
A/STRACT8
/AC1GROUND8 Human semen contains high concentrations of zinc (Zn) and copper (Cu) in bound and ionic forms .The presence of abnormal levels of these trace elements may affect spermatogenesis with regard to production , maturation , motility , and fertilizing capacity of the spermatozoa. The aim of this study is to evaluate the levels of Zn and Cu in seminal plasma in different groups of male infertility and to correlate their concentrations with various semen parameters. METHODS8 orty primary infertile male individuals ,who had regular unprotected intercourse for at least one year without conception with their partners ,aged !"#$% years were involved in the present study .&fter seminal plasma fluid analyses they were grouped as, azoospermic (n'(!), oligoasthenozoospermic (n'()) ,and teratozoospermic (n'(!) .Twelve fertile males selected from general population and after seminal fluid investigation were ta*en as normospermic control group .Zinc and Copper concentrations in separated seminal plasma of each infertile male and fertile control sub+ect were determined by atomic absorption spectrophotometer. RESULTS8 This study showed significant decrease of seminal plasma Zn mean (,-./) value in oligoasthenozoospermic infertile males than in fertile male controls (p0%.%12) .3n azoospermic males, the mean (,-./) value of seminal plasma Zn levels was decreased (but still beyond the significant level,p'%.%4) when compared with that of fertile males. 5ith regard to seminal plasma copper level there was insignificant decrease in its level in azoospermic males and insignificant increase in its concentration in oligoasthenozoospermic and in teratozoospermic males when compared with that of healthy fertile males.3n addition, there was a significant decrease in seminal plasma copper concentration in azoospermic males than in oligoasthenozoospermic males (p0%.%1").The results also revealed a significant positive correlation between the sperm motility values and the seminal plasma zinc levels in oligoasthenozoospermic group(r'%.)4, p0%.%!!) along with a significant negative correlation between sperm morphology values and zinc seminal plasma levels in teratozoospermic group (r'#%.)1,p0%.%$6). CONCLUSION8 7n the basis of the findings of this study, it seems that the estimation of seminal plasma levels of zinc and copper may aid in investigation and treatment of infertile males. 1E29ORDS8 infertility, azoospermic , oligoasthenozoospermic ,seminal Zinc ,seminal Copper . INTRODUCTION8 3nfertility is comple8 and has many causes and conse9uences depending on the gender, se8ual
:;ept. of <hysiological Chemistry, College of /edicine, =niv. of >aghdad. ::3nstitute of .mberyo ?esearch and 3nfertility,=niv. of &@# Aahrain. :::<oisoning Consultation Center. :3nstitute of .mberyo ?esearch and 3nfertility,=niv. of &@# Aahrain. ::&l#/ussaib Hospital

/as#+ O#(& M -a**(& Sa+(-0 , Na.a+ 1-#), Hussa#$00 , A+# 2a3u4 Ma5#& 000 , /a$ T-a4(t0000, 1-a6)a5 A44as 7a&-#+000 00

history, life style of society and cultural bac*ground of people it affect ((). 3nfertility affects appro8imately 4B to (!B of the worldCs population and in about half of cases male are either the single cause of or contribute the coupleCs infertility (!)./ore than 6%B of male infertility cases are due to low sperm counts ,poor sperm 9uality ,or both .3n 1%#$%B of cases of sperm abnormlities ,the cause is un*nown .3t may be the end result of one or more factors that include chronic illness ,malnutrition ,genetic defects ,structural

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"INC AND COPPER CONCENTRATIONS IN SEMINAL PLASMA abnormalities ,and environmental factors(1) Human semen contains high concentrations of trace elements li*e calcium (Ca) ,magnesium (/g), copper (Cu) ,selenium (-e) ,and zinc (Zn) in bound and free (ionic) forms .These trace elements play very vital role in affecting various parameters of semen ($) . The testicular plasma ,that is ,the fluid composed of the secretions originating in the seminiferous tubules ,tubuli recti ,rete testis ,and ductuli efferents ,and the epididymal plasma serve as nutrient medium in which maturation of the developing spermatozoa ta*es place (",)) .Trace elements have been shown to be essential for testicular development and spermatogenesis (2). Zinc in seminal plasma stabilizes the cell membrane and nuclear chromatin of
6) spermatozoa .Zinc may also be the primary factor responsible for the antibacterial activity of the seminal plasma ((%) and protect the male gonads against the degenerative changes ((() . 3t may also play a regulatory role in the process of capacitation and acrosome reaction ($, (!) .@ow content of this metal (Zn) has been suggested to diminish fertility potential in males ((1). Zinc deficiency leads to gonadal dysfunction, decreases testicular weight, and causes shrin*age of seminiferous tubules (($). Copper is an important element for numerous metalloenzymes and metalloproteins that are involved in energy or antio8idant metabolism. However, in its ionic form (Cu !,) and at high level, this trace element rapidly becomes to8ic to a variety of cells, including human spermatozoa ()). 3t has been identified that Cu is highly to8ic for sperm ((") .3n vitro studies, it has been demonstrated the effect of Cu in intrauterine devices preventing conception (()). The present study was designed to evaluate seminal plasma levels of zinc and copper and to correlate their concentractions with various semen parameters among fertile and infertile male sub+ects. SU/JECTS AND METHODS8 This study was carried out at the 3nstitute of .mberyo ?esearch and 3nfertility, =niv. of &@# Aahrain, >aghdad, during the period from eb.!%%) to &ugust !%%). orty primary infertile male sub+ects ,who had regular unprotected intercourse for at least one year without conception with their partners , aged !"#$% years were included in this study . <atients had no e8ternal ,infections ,traumatic abnormalities which could be implicated in the development of infertility . &t first clinic attendance,

(4,

a detailed bac*ground history and physical e8amination were done on both husband and wife. -emen specimens from all infertile patients were collected into sterile polystyrene +ars after an abstinence period of 1 to " days .& portion of each semen sample was e8amined for sperm count ,motility and morphologic features .3nfertile male patients were then divided into the following three groups Daccording to their sperm count Emotility and Eor morphology ,5H7 criteria ,(66!((2)F G) u' I8 A6 s'()*#% (sperm count 'zero, n'(!), G) u' II 87ligoasthenozoospermic (sperm count 0!%G(%)Em( ,motility0"%B ,n'()), and G) u' III8 Teratozoospermic (sperm count H !%G(%)Eml, motility H"%B, morphology0"%B, n'(!). Twelve fertile males whose partners had conceived within one year and having sperm count more than !% million Em( with motility and morphology more than H"%B were selected from general population and ta*en as normospermic control group . &fter li9uefaction ,the seminal plasma was collected after centrifugation at !%%% rpm for ("#!% minutes .-upernatant ,the seminal plasma ,was transferred in fresh tubes and stored frozen ,at#!%IC until assay .The concentrations of zinc and copper in separated seminal plasma of each infertile patient and fertile control were determined by atomic absorption spectrophotometer, >uc* -cientific, model !(% JK<. RESULTS8 ?esults are e8pressed as mean (, -./) form each parameter. Table ( summarizes the mean (,-./) value on seminal plasma levels of Zn and Cu elements in the three groups of infertile male sub+ects (azoospermia ,oligoasthenozoospermia and teratozoospermia) and in fertile control group . The mean (,-./) value of seminal plasma Zn concentrations was significantly decreased in oligoasthenozoospermic infertile males than in fertile male controls (!.1(,%."6 mgEdl, 1.4",%."6 mgEdl, respectively, p0%.%12, Table (). 3n azoospermic patients ,the mean (,-./) value of seminal plasma Zn level was decreased in comparison with that of controls ,but still beyond the significant value (p'%.%4) .There was no significant differences in seminal plasma Zn concentration between the teratoazoospermic patients and fertile controls as well as among the different groups of infertile males (Table ().Table ( also shows that the mean (,-./) value of seminal plasma Cu levels was insignificantly decreased in azoospermic patients when compared with that of fertile control group .

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"INC AND COPPER CONCENTRATIONS IN SEMINAL PLASMA urthermore ,this element ,Cu ,was insignificantly increased in oligoasthenozoospermic infertile males and in teratozoospermic infertile males than in fertile control sub+ects (Table () . &mong infertile male groups there was a significant decrease in plasma Cu level in azoospermic patients than in oligoasthenozoospermic infertile group (%.1%,%.($ mgEdl, %.2%,%.(1mgEdlL respectively p0%.%1", Table ().

Ta4+( 18"#$% a$& C ''() C $%($t)at# $s #$ S(*#$a+ P+as*a #$ T-)(( G) u's ! I$!()t#+( Ma+(s a$& 7()t#+( C $t) + G) u' .T-( Va+u(s a)( E;')(ss(& as M(a$<= SEM>. &zoospermic (n'(!) !.$% ,%.)$ 7ligoastheno# zoospermic (n'()) !.1( ,%."6 : Teratozoos# permic (n'(!) !.62 ,%.)( Aormospermic (n'(!) 1.4" ,%."6

<arameters -eminal <lasma Zn concentration (mgEdl) -eminal <lasma Cu concentration (mgEdl)

%.1% ,%.($ ::

%.2% ,%.(1

%.$! ,%.($

%.$( ,%.($

:p0%.%" when oligoasthenozoospermic group is compared with normospermic group. ::p0%.%" when azoospermic group is compared with oligoasthenozoosperrmic group.

Correlation coefficient of seminal plasma concentration of Zn and Cu with various seminal parameters in azoospermic ,oligoasthenozoospermic and teratozoospermic males are showed in Table ! . There was a significant positive correlation between seminal plasma Zn concentrations and sperm motility in oligoasthenozoospermic group (r'%.)4L p 0 %.%!!) as well as a significant negative correlation between Zn seminal plasma levels and sperm morphology in treatozoospermic group (r ' #%.)1L p0 %.%$6, Table !).
Ta4+( 28C ))(+at# $ C (!!#%#($t <)> ! S(*#$a+ P+as*a "#$% a$& C ''() C $%($t)at# $s .#t- S(*($ Pa)a*(t()s. <arameters -perm count ((% ) Eml) -perm /orphology (B) /otility (B) &zoospermic(n'(!) Zn (rMp# value) LCu (rMp# value) (%M%) L (%M%) (%M%) L (%M%) (% M %) L (% M %) 7ligoastheno# zoospermic (n'()) Zn (rMp#value) LCu (rMp#value) (#%.1( M%.1")L (%.%($ M %.6) ) (#%.%6" M %.24) L (#%.!6 M %.12) (%.)4 M %.%! :) L (#%.$( M %.(4) Teratoazoos# permic (n'(!) Zn (rMp#value) LCu (rMp#value) (%.(6 M %.)%) L (%."1 M %.(") (#%.)1 M %.%$6 :) L (%.%%" M %.66) (%.%2 M %.4$) L (#%.%2 M %.4))

<#value 0 %.%" was considered significant.

DISCUSSION8 The results of the present study showed that there was significant low concentration of seminal plasma Zn in oligoasthenozoospermic males when compared with that of fertile controls. Zinc level was also decreased in azoospermic infertile males in comparsion with fertile healthy controls, but does not reachs the significant level. 5ith regard to plasma seminal Cu level, it was significantly reduced in azoospermic male patients than in
THE IRAQI POSTGRADUATE MEDICAL JOURNAL

oligoasthenozoospermic infertile males. &li and associates, !%%" ((4) and &li et al, !%%2 ($) observed significant decrease in seminal plasma Zn in oligoospermic and azoospermic infertile males. These authors concluded that Zn concentration in seminal plasma should be considered as one of the factors responsible for decreased testicular function in infertile male sub+ects. 3n contrast , use et al , (666 ((6) have found no significant differences in the
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"INC AND COPPER CONCENTRATIONS IN SEMINAL PLASMA mean values of seminal plasma Zn levels between the entire group of samples from infertile individuals and those from fertile individuals .However, the latter authors observed that Zn concentration of the seminal plasma in azoospermic and oligoasthenozoospermic patients was significantly lower than that in other groups of male infertility .The authors concluded that ade9uate seminal concentration of the Zn is re9uired for normal sperm function . 3t has been demonstrated that Zn in human semen is derived from the prostate ((6) . 7ne important function of the Zn seems to be its regulatory role in the level of dihydrotestosterone by modulating the " N # reductase enzyme activity ,which is located in the microsomal and nuclear fraction of the prostatic cell (!%). 7ther important roles suggested for the Zn include maintenance of the structural integrity of macromolecules (!(), regulation of protein and nucleic acid metabolism (!!), and activation of mitochondrial function and &T< production (!1). & clinical study demonstrated that adult males e8perimentally deprived of Zn showed that the @eydig cell synthesis of testosterone was disturbed (!$). Zinc appears to be a potent scavenger of e8cessive supero8ide anions produced by defective spermatozoa and Eor leu*ocytes in human semen after e+aculation (!"). Thus, it seems that seminal plasma, due to its ade9uate content of Zn, will e8ert protective, antio8idant # li*e activity sufficient to cope with the e8cessive amount of supero8ide anions (!)). The abnormal spermatozoa that infertile men have a higher percentage of it would be a source of supero8ide anions that bind with Zn present in the seminal plasma and thus reduce the Zn levels (2). 7mu and associates, (664 (!2) have demonstated that Zn therapy results in significant improvement in sperm 9uality with increases in sperm density, progressive motility, and improved conception and pregnancy outcome. 7ur study also demonstrated that seminal plasma Zn concentration was significantly correlated with sperm motility. This observation is in accordance with results reported by other authors (2 , (6) and differs from the results of other studies ($). Human prostatic fluid causes the immotile sperm in vesicular fluid to become motile and this change may be induced by Zn in prostatic fluid (!4). Zinc has been detected in human sperm and this suggested its role in motility (!6) . The results of the present study also showed that Cu concentration was insignificantly increased in oligoasthenozoospermic and in teratozoospermic males than in fertile male controls. This result is in compatible with that observed by 5ong et al, !%%( ()) . Copper is an essential trace element that plays an important role in several enzymes such as supero8ide dismutase. Human spermatozoa are particularly susceptible to pero8idative damage because they contain high concentrations of polyunsaturated fatty acids and also possess a significant ability to generate a reactive o8ygen species (?7-), mainly supero8ide anion and hydrogen pero8ide. -upero8ide dismutase (Cu#metalloenzyme) protects human spermatozoa from this pero8idative damage ()). However, this element in its ionic form (Cu !,), particularly, at high concentration rapidly become to8ic to a variety of cells, including human spermatozoa ()). 3n vitro studies, ?oblero et al (()) have demonstrated the effect of Cu in intrauterine devise in preventing conception. CONCLUSION8 7n the basis of the observations of the present study and other reports, seminal zinc and copper may contribute to fertility through their effects on various semen parameters. &de9uate seminal plasma concentration of Zn and Cu are re9uired for normal sperm function and that high to8ic concentration of these elements in seminal plasma is apparently related to defective morphology and Eor motility in infertile males. 3t seems that the estimation of seminal plasma Zn and Cu may help in investigation and treatment of infertile males. RE7ERENCES8 1. Oerry @5. ;efining infertility. amily Health 3nternational. !%%1 L!1 M$#4 2. >arbara >. /en contribute to and suffer from infertility. amily Health 3nternational. !%%1 L!1 M(2#!% ?. Haymond - and Kronows*i & /. ?eproductive related disordersM 3n Tietz Te8tboo* of Clinical Chemistry and /olecular ;iagnostics edited by >urtis C&, &shwood .? and >runs ;., Hsevier -aunders, !%%)L !%26#!("!. @. &li H, &hmed /, >aig /, and &li /. ?elationship of zinc concentrations in blood and seminal plasma with various semen parameters in infertile sub+ects. <a*istan Pournal of /edical -ciences. !%%2L !1M (((#(($. A. /ann T. The biochemistry of semen and of the male reproductive tract. @ondonM /ethuen Q Co @T;, (6)$.

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"INC AND COPPER CONCENTRATIONS IN SEMINAL PLASMA :. 5ong 5 R, li* K, Kroenen </5, -win*els ;5, Thomas C/K, Copius #<eereboom PHP, /er*us H/5/ and -teegers#Theunissen ?</. The impact of calcium, magnesium, zinc and copper in blood and seminal plasma on semen parameters in men. ?eproductive To8icology. !%%(L ("M (1(#(1). 7. CHia -#., 7ng (C#A, Chua @#H, Ho @#/ and Tay -#O. Comparison of zinc concentration in blood and seminal plasma and the various sperm parameters between fertile and infertile men. Pournal of &ndrology. !%%%L !(,"1#"2. 8. Chvapil /. Aew aspects in biological role of zinc. & stabilizer of marcromolecules and biological membrane. @ife -cience. (621L (1, (%$$#(%$6. B. Ovist =. -perm nuclear chromatin decondensation ability. &n in vitro study on e+aculated human spermatozoa. &cta physical -cand -uppl (64%L $4),(#!$. 10. @angley P, Koldsmid P, ;avies A. Jenereal trichomoniasis role in men. Kenitourin /ed. (642L )1, !)$#!)2. 11. >atra A, Aehru >, >ansal /<. ?eproductive potential of male <ortan rats e8posed to various levels of lead with regard to zinc status. >r P Autr. !%%$L 6(, 142#16(. 12. ?iffo /, @evia -, &studillo P. .ffect of zinc on human sperm motility and acrosome reaction. 3nter P &ndrol. (66!L (", !!6#!12. 1?. /armar P@, Oatz -, <raiss ;., ;ebenedicitis TP. -emen zinc level in infertile and postvasectomy patients and patients with prostatitis . ertility. -terility. (62"L !), (%"2. 1@. >edwal ?-, >ahuguna &. Zinc, copper and selenium in reproduction. .8perientia. (66$L "%, )!)#)$%. 1A. -Oandhan O<.?eview on copper in male reproduction and contraception. ?ev r Kynecol 7bstet. (66!L 42M "6$#"64. 1:. ?oblero @, Kuadarrama &, @opez T, and Zegers # Hochschild . .ffect of copper ion on the motility, viability, acrosome reaction and fertilizing capacity of human spermatozoa in vitro . ?eprod ertil ;ev. (66)L 4,42(#42$. 17. 5orld Health 7rganization. 5H7 @aboratory /anual for the .8amination of Human -emen and -emen # cervical /ucus 3nteraction, 1 rd edn. Cambridge =niv. <ress, Cambridge, =O (66!, ). 18. &li H, >aig m, ?ana / , &li /, Sasim ?, and Ohem &O. ?elationship of serum and seminal plasma zinc level and serum testosterone in oligospermic and azoospermic infertile men. P Coll physicians -urg <a*. !%%"L (", )2(#)21. 1B. use H, Oazama T, 7hta - and u+iuchi R. ?elationship between zinc concentrations in seminal plasma and various sperm parameters. 3ntern =rology and Aephrology. (666L 1(, $%(#$%4. 20. @ea* &C, Chisholm K>, Habib O. The effect of zinc on N # reduction of testosterorie by the hyperplastic human prostate gland. P -teroid >iochem. (64$L !%, )"(. 21. Chvapil /. Aew aspects in the biological role of zinc .& stabilizer of macromolecules of biological membranes. @ife -cience. (621L (1,(%$(. 22. >ettger 5P, 7 ;ell >@ .& critical physiological role of zinc in the structure and function of biomembranes. @ife -cience. (64(L !4, ($!". 2?. Ramaguchi /, Oura ,/, 7*ada -. ?ole of zinc as an activator of mitochondrial function in rat liver. >iochem. <harmcol. (64!L 1(,(!46. 2@. Aetter &, Hartoma ?, Aahail O. .ffects of zinc administration on plasma testosterone and dihydrotestosterone and sperm count. &rch &ndrol. (64(L 2, )6#21. 2A. 3rvine ;-. Klutathione as a treatment for male infertility. ?ev ?eprod. (66)L (,)#(!. 2:. Kavella /, @ipovac J. 3n vitro effect of zinc on o8idative changes in human semen. &ndrologia. (664L 1%, 1(2#1!1. 27. 7mu &., ;ashti H, &l#7thman -. Treatment of asthenozoospermic with zinc sulphate M andrological , immunological and obstetric outcome. .ur P 7bstet Kynecol ?eprod >iol. (664L 26, (26#(4$. 28. @indholmer C.The importance of seminal plasma for human sperm motility. >iol. ?eprod. (62$L (%,"11. 2B. >accetti >, <allini J, >urrini & K. The accessory fibers of the sperm tail. 3 3. Their role in binding zinc in mammals and cephalopods. P =ltrastract .?es. (62)L "$,!)(.

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