左侧精索静脉曲张对成年大鼠附睾精子结合抗原11C和精子结合抗原11T表达的影响
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摘要
目的明确精子结合抗原11C(SPAG11C)和11T(SPAG11T)在大鼠附睾中的表达定位,探究实验性左侧精索静脉曲张(ELV)对其表达的影响,为研究精索静脉曲张致不育机制提供实验资料。方法 48只成年SD大鼠随机分为4组:假手术2周组(A组)、假手术4周组(B组)和ELV造模2周组(C组)、ELV造模4周组(D组),每组12只。造模组行左肾静脉部分结扎术,建立实验性左侧精索静脉曲张模型;假手术组仅暴露左肾静脉。采用实时定量聚合酶链反应和免疫组织化学的方法,检测SPAG11C和SPAG11T在各组双侧附睾组织中的表达定位及变化。结果 SPAG11C和SPAG11T主要表达在附睾上皮的主细胞胞质内,且其表达呈现区段特异性。在ELV造模2周和4周后,造模组大鼠左侧附睾中SPAG11C和SPAG11T在核酸和蛋白水平上的表达均较同侧假手术组及造模组右侧附睾显著降低(P<0.05),且与ELV造模2周组相比,ELV造模4周组降低更为明显(P<0.05);而ELV造模组右侧附睾与假手术组比较无差异(P>0.05)。结论 SPAG11C和SPAG11T在附睾中的表达具有细胞特异性和区段特异性,且精索静脉曲张可致其表达显著降低,可能是精索静脉曲张导致附睾微环境改变,影响精子成熟,继而导致男性生育力降低的原因之一。
Objective To investigate the expression and distribution of sperm associated antigen 11C and T(SPAG11C and T) in the epididymis of adolescent rats with experimental left varicocele(ELV) model, and to explore the mechanism of varicocele-induced subfertility. Methods The partial ligation of left renal vein was adopted to establish ELV rat model, parallel with the sham treatment meanwhile. Total of 48 SD rats were divided into four groups: ELV-2 weeks and 4 weeks groups, sham-2 weeks and 4 weeks groups. The expressions of SPAG11C and T were detected by the real-time PCR and immunohistochemistry. Results SPAG11C and T were mainly distributed in the cytoplasm of principle cells of the epididymal epithelium. The immunostaining density of SPAG11C was strong in the proximal corpus, and weak gradually from the middle corpus to the cauda. The strong expression of SPAG11 T was detected in the proximal corpus and the cauda, but the weak signal existed in the distal corpus. ELV-2 and 4 weeks later, the expressions of SPAG11C and T stayed normal level in the bilateral epididymides of sham groups and in the right side of ELV groups. Whereas the significant decreases of expressions of SPAG11C and T were found in the left epididymides of ELV groups, compared with sham operation group and ELV right epididymides. The heavier slump was found in the ELV left side at 4 weeks. Conclusion SPAG11C and T are specific proteins expressed in the epididymis, and they involved in the composition of epididymal microenvironment. ELV downregulated their expression levels significantly. Decrease of them may attribute to varicocele-caused male subfertility.
Objective To investigate the expression and distribution of sperm associated antigen 11C and T(SPAG11C and T) in the epididymis of adolescent rats with experimental left varicocele(ELV) model, and to explore the mechanism of varicocele-induced subfertility. Methods The partial ligation of left renal vein was adopted to establish ELV rat model, parallel with the sham treatment meanwhile. Total of 48 SD rats were divided into four groups: ELV-2 weeks and 4 weeks groups, sham-2 weeks and 4 weeks groups. The expressions of SPAG11C and T were detected by the real-time PCR and immunohistochemistry. Results SPAG11C and T were mainly distributed in the cytoplasm of principle cells of the epididymal epithelium. The immunostaining density of SPAG11C was strong in the proximal corpus, and weak gradually from the middle corpus to the cauda. The strong expression of SPAG11 T was detected in the proximal corpus and the cauda, but the weak signal existed in the distal corpus. ELV-2 and 4 weeks later, the expressions of SPAG11C and T stayed normal level in the bilateral epididymides of sham groups and in the right side of ELV groups. Whereas the significant decreases of expressions of SPAG11C and T were found in the left epididymides of ELV groups, compared with sham operation group and ELV right epididymides. The heavier slump was found in the ELV left side at 4 weeks. Conclusion SPAG11C and T are specific proteins expressed in the epididymis, and they involved in the composition of epididymal microenvironment. ELV downregulated their expression levels significantly. Decrease of them may attribute to varicocele-caused male subfertility.
引文
1 Damsgaard J,Joensen UN,Carlsen E,et al.Varicocele Is Associated with Impaired Semen Quality and Reproductive Hormone Levels:A Study of 7035 Healthy Young Men from Six European Countries.Eur Urol 201670(6):1019-1029
2 Gorelick JI,Goldstein M.Loss of fertility in men with varicocele.Fertil Steril 1993;59(3):613-616
3 Nielsen ME,Zderic S,Freedland SJ,et al.Insight on pathogenesis of varicoceles:relationship of varicocele and body mass index.Urology 2006;68(2):392-396
4 Zhang K,Wang Z,Wang H,et al.Hypoxia-induced apoptosis and mechanism of epididymal dysfunction in rats with left-side varicocele.Andrologia 2016;48(3):318-324
5 薛侠,马思敏,邱曙东,等.实验性精索静脉曲张对青春期大鼠睾丸、附睾中CRES蛋白表达的影响.中华男科学杂志2006;12(11):974-978
6 Biswas B,Yenugu S.Transcriptional regulation of the rat sperm-associated antigen 11e(Spag 11e)gene during endotoxin challenge.Mol Genet Genomics 2014;289(5):837-845
7 Wang YJ,Zhang RQ,Lin YJ,et al.Relationship between varicocele and sperm DNA damage and the effect of varicocele repair:a meta-analysis.Reprod Biomed Online 2012;25(3):307-314
8 Ozturk U,Kefeli M,Asci R,et al.The effects of experimental left varicocele on the epididymis.Syst Biol Reprod Med 2008;54(4-5):177-184
9 Lehtihet M,Arver S,Kalin B,et al.Left-sided grade3 varicocele may affect the biological function of the epididymis.Scand J Urol 2014;48(3):284-289
10 Zhang QY,Qiu SD,Ma XN,et al.Effect of experimental varicocele on structure and function of epididymis in adolescent rats.Asian J Androl 2003;5(2):108-112
11 Agarwal A,Hamada A,Esteves SC.Insight into oxidative stress in varicocele-associated male infertility:part 1.Nat Rev Urol 2012;9(12):678-690
12 Zhang QY,Qiu SD,Ma XN,et al.Effect of experimental varicocele on structure and function of epididymis in adolescent rats.Asian J Androl 2003;5(2):108-112
13 Li P,Chan HC,He B,et al.An antimicrobial peptide gene found in the male reproductive system of rats.Science2001;291(5509):1783-1785
14 Yenugu S,Hamil KG,French FS,et al.Antimicrobial actions of human and macaque sperm associated antigen(SPAG)11 isoforms:influence of the N-terminal peptide.Mol Cell Biochem 2006;284(1-2):25-37
15 Hamil KG,Sivashanmugam P,Richardson RT,et al.HE2beta and HE2gamma,new members of an epididymis-specific family of androgen-regulated proteins in the human.Endocrinology 2000;141(3):1245-1253
16 Fr?hlich O,Ibrahim NM,Young LG.EP2 splicing variants in rhesus monkey(Macaca mulatta)epididymis.Biol Reprod 2003;69(1):294-300
17 Fr?hlich O,Po C,Murphy T,et al.Multiple promoter and splicing m RNA variants of the epididymis-specific gene EP2.J Androl 2000;21(3):421-430
18 Avellar MC,Honda L,Hamil KG,et al.Differential expression and antibacterial activity of epididymis protein2 isoforms in the male reproductive tract of human and rhesus monkey(Macaca mulatta).Biol Reprod 2004;71(5):1453-1460
19 Yenugu S,Hamil KG,Grossman G,et al.Identification,cloning and functional characterization of novel sperm associated antigen 11(SPAG11)isoforms in the rat.Reprod Biol Endocrinol 2006;4:23
20 Yenugu S,Hamil KG,Birse CE,et al.Antibacterial properties of the sperm-binding proteins and peptides of human epididymis 2(HE2)family;salt sensitivity,structural dependence and their interaction with outer and cytoplasmic membranes of Escherichia coli.Biochem J2003;372(Pt 2):473-483
21 Yenugu S,Hamil KG,French FS,et al.Antimicrobial actions of the human epididymis 2(HE2)protein isoforms,HE2alpha,HE2beta1 and HE2beta2.Reprod Biol Endocrinol 2004;2:61
22 von Horsten HH,Schafer B,Kirchhoff C.SPAG11/isoform HE2C,an atypical anionic beta-defensin-like peptide.Peptides 2004;25(8):1223-1233
23 von Horsten HH,Derr P,Kirchhoff C.Novel antimicrobial peptide of human epididymal duct origin.Biol Reprod2002;67(3):804-813
24 Biswas B,Yenugu S.Antimicrobial responses in the male reproductive tract of lipopolysaccharide challenged rats.Am J Reprod Immunol 2011;65(6):557-568
25 Biswas B,Yenugu S.Lipopolysaccharide induces epididymal and testicular antimicrobial gene expression in vitro:insights into the epigenetic regulation of spermassociated antigen 11e gene.Immunogenetics 2013;65(4):239-253
26 Liu X,Ju Z,Wang L,et al.Six novel single-nucleotide polymorphisms in SPAG11 gene and their association with sperm quality traits in Chinese Holstein bulls.Anim Reprod Sci 2011;129(1-2):14-21
27 Zhou CX,Zhang YL,Xiao L,et al.An epididymisspecific beta-defensin is important for the initiation of sperm maturation.Nat Cell Biol 2004;6(5):458-464
28 Yudin AI,Tollner TL,Li MW,et al.ESP13.2,a member of the beta-defensin family,is a macaque sperm surfacecoating protein involved in the capacitation process.Biol Reprod 2003;69(4):1118-1128
29 Dohle GR,Colpi GM,Hargreave TB,et al.EAU guidelines on male infertility.Eur Urol 2005;48(5):703-711
30 Hsiao W,Rosoff JS,Pale JR,et al.Varicocelectomy is associated with increases in serum testosterone independent of clinical grade.Urology 2013;81(6):1213-1217
31 刘建军,董强,杨宇如.实验性精索静脉曲张大鼠血清及睾内睾酮的变化.中华男科学杂志2007;13(4):335-337
32 Zheng Y,Zhang X,Zhou J,et al.Effects on the ipsilateral testis during progression of experimental varicocele in rat.Med Sci Monit 2008;14(6):BR122-BR126
33 Zheng YQ,Zhang XB,Zhou JQ,et al.The effects of artery-ligating and artery-preserving varicocelectomy on the ipsilateral testes in rats.Urology 2008;72(5):1179-1184
34 Rajfer J,Turner TT,Rivera F,et al.Inhibition of testicular testosterone biosynthesis following experimental varicocele in rats.Biol Reprod 1987;36(4):933-937
35 Luo DY,Yang G,Liu JJ,et al.Effects of varicocele on testosterone,apoptosis and expression of St AR m RNA in rat Leydig cells.Asian J Androl 2011;13(2):287-291
36 Tanrikut C,Goldstein M,Rosoff JS,et al.Varicocele as a risk factor for androgen deficiency and effect of repair.BJU Int 2011;108(9):1480-1484
37 Soares TS,Fernandes SA,Lima ML,et al.Experimental varicocoele in rats affects mechanisms that control expression and function of the androgen receptor.Andrology 2013;1(5):670-681
38 Jaworski T.Degradation and beyond:control of androgen receptor activity by the proteasome system.Cell Mol Biol Lett 2006;11(1):109-131
39 Anbalagan M,Huderson B,Murphy L,et al.Posttranslational modifications of nuclear receptors and human disease.Nucl Recept Signal 2012;10:e001
40 Lee JH,Lee MJ.Emerging roles of the ubiquitinproteasome system in the steroid receptor signaling.Arch Pharm Res 2012;35(3):397-407
41 杨洁,张秋养,葛玲,等.青春期大鼠实验性精索静脉曲张对附睾雄激素受体的影响.西安交通大学学报·医学版2005;26(1):22-25
2 Gorelick JI,Goldstein M.Loss of fertility in men with varicocele.Fertil Steril 1993;59(3):613-616
3 Nielsen ME,Zderic S,Freedland SJ,et al.Insight on pathogenesis of varicoceles:relationship of varicocele and body mass index.Urology 2006;68(2):392-396
4 Zhang K,Wang Z,Wang H,et al.Hypoxia-induced apoptosis and mechanism of epididymal dysfunction in rats with left-side varicocele.Andrologia 2016;48(3):318-324
5 薛侠,马思敏,邱曙东,等.实验性精索静脉曲张对青春期大鼠睾丸、附睾中CRES蛋白表达的影响.中华男科学杂志2006;12(11):974-978
6 Biswas B,Yenugu S.Transcriptional regulation of the rat sperm-associated antigen 11e(Spag 11e)gene during endotoxin challenge.Mol Genet Genomics 2014;289(5):837-845
7 Wang YJ,Zhang RQ,Lin YJ,et al.Relationship between varicocele and sperm DNA damage and the effect of varicocele repair:a meta-analysis.Reprod Biomed Online 2012;25(3):307-314
8 Ozturk U,Kefeli M,Asci R,et al.The effects of experimental left varicocele on the epididymis.Syst Biol Reprod Med 2008;54(4-5):177-184
9 Lehtihet M,Arver S,Kalin B,et al.Left-sided grade3 varicocele may affect the biological function of the epididymis.Scand J Urol 2014;48(3):284-289
10 Zhang QY,Qiu SD,Ma XN,et al.Effect of experimental varicocele on structure and function of epididymis in adolescent rats.Asian J Androl 2003;5(2):108-112
11 Agarwal A,Hamada A,Esteves SC.Insight into oxidative stress in varicocele-associated male infertility:part 1.Nat Rev Urol 2012;9(12):678-690
12 Zhang QY,Qiu SD,Ma XN,et al.Effect of experimental varicocele on structure and function of epididymis in adolescent rats.Asian J Androl 2003;5(2):108-112
13 Li P,Chan HC,He B,et al.An antimicrobial peptide gene found in the male reproductive system of rats.Science2001;291(5509):1783-1785
14 Yenugu S,Hamil KG,French FS,et al.Antimicrobial actions of human and macaque sperm associated antigen(SPAG)11 isoforms:influence of the N-terminal peptide.Mol Cell Biochem 2006;284(1-2):25-37
15 Hamil KG,Sivashanmugam P,Richardson RT,et al.HE2beta and HE2gamma,new members of an epididymis-specific family of androgen-regulated proteins in the human.Endocrinology 2000;141(3):1245-1253
16 Fr?hlich O,Ibrahim NM,Young LG.EP2 splicing variants in rhesus monkey(Macaca mulatta)epididymis.Biol Reprod 2003;69(1):294-300
17 Fr?hlich O,Po C,Murphy T,et al.Multiple promoter and splicing m RNA variants of the epididymis-specific gene EP2.J Androl 2000;21(3):421-430
18 Avellar MC,Honda L,Hamil KG,et al.Differential expression and antibacterial activity of epididymis protein2 isoforms in the male reproductive tract of human and rhesus monkey(Macaca mulatta).Biol Reprod 2004;71(5):1453-1460
19 Yenugu S,Hamil KG,Grossman G,et al.Identification,cloning and functional characterization of novel sperm associated antigen 11(SPAG11)isoforms in the rat.Reprod Biol Endocrinol 2006;4:23
20 Yenugu S,Hamil KG,Birse CE,et al.Antibacterial properties of the sperm-binding proteins and peptides of human epididymis 2(HE2)family;salt sensitivity,structural dependence and their interaction with outer and cytoplasmic membranes of Escherichia coli.Biochem J2003;372(Pt 2):473-483
21 Yenugu S,Hamil KG,French FS,et al.Antimicrobial actions of the human epididymis 2(HE2)protein isoforms,HE2alpha,HE2beta1 and HE2beta2.Reprod Biol Endocrinol 2004;2:61
22 von Horsten HH,Schafer B,Kirchhoff C.SPAG11/isoform HE2C,an atypical anionic beta-defensin-like peptide.Peptides 2004;25(8):1223-1233
23 von Horsten HH,Derr P,Kirchhoff C.Novel antimicrobial peptide of human epididymal duct origin.Biol Reprod2002;67(3):804-813
24 Biswas B,Yenugu S.Antimicrobial responses in the male reproductive tract of lipopolysaccharide challenged rats.Am J Reprod Immunol 2011;65(6):557-568
25 Biswas B,Yenugu S.Lipopolysaccharide induces epididymal and testicular antimicrobial gene expression in vitro:insights into the epigenetic regulation of spermassociated antigen 11e gene.Immunogenetics 2013;65(4):239-253
26 Liu X,Ju Z,Wang L,et al.Six novel single-nucleotide polymorphisms in SPAG11 gene and their association with sperm quality traits in Chinese Holstein bulls.Anim Reprod Sci 2011;129(1-2):14-21
27 Zhou CX,Zhang YL,Xiao L,et al.An epididymisspecific beta-defensin is important for the initiation of sperm maturation.Nat Cell Biol 2004;6(5):458-464
28 Yudin AI,Tollner TL,Li MW,et al.ESP13.2,a member of the beta-defensin family,is a macaque sperm surfacecoating protein involved in the capacitation process.Biol Reprod 2003;69(4):1118-1128
29 Dohle GR,Colpi GM,Hargreave TB,et al.EAU guidelines on male infertility.Eur Urol 2005;48(5):703-711
30 Hsiao W,Rosoff JS,Pale JR,et al.Varicocelectomy is associated with increases in serum testosterone independent of clinical grade.Urology 2013;81(6):1213-1217
31 刘建军,董强,杨宇如.实验性精索静脉曲张大鼠血清及睾内睾酮的变化.中华男科学杂志2007;13(4):335-337
32 Zheng Y,Zhang X,Zhou J,et al.Effects on the ipsilateral testis during progression of experimental varicocele in rat.Med Sci Monit 2008;14(6):BR122-BR126
33 Zheng YQ,Zhang XB,Zhou JQ,et al.The effects of artery-ligating and artery-preserving varicocelectomy on the ipsilateral testes in rats.Urology 2008;72(5):1179-1184
34 Rajfer J,Turner TT,Rivera F,et al.Inhibition of testicular testosterone biosynthesis following experimental varicocele in rats.Biol Reprod 1987;36(4):933-937
35 Luo DY,Yang G,Liu JJ,et al.Effects of varicocele on testosterone,apoptosis and expression of St AR m RNA in rat Leydig cells.Asian J Androl 2011;13(2):287-291
36 Tanrikut C,Goldstein M,Rosoff JS,et al.Varicocele as a risk factor for androgen deficiency and effect of repair.BJU Int 2011;108(9):1480-1484
37 Soares TS,Fernandes SA,Lima ML,et al.Experimental varicocoele in rats affects mechanisms that control expression and function of the androgen receptor.Andrology 2013;1(5):670-681
38 Jaworski T.Degradation and beyond:control of androgen receptor activity by the proteasome system.Cell Mol Biol Lett 2006;11(1):109-131
39 Anbalagan M,Huderson B,Murphy L,et al.Posttranslational modifications of nuclear receptors and human disease.Nucl Recept Signal 2012;10:e001
40 Lee JH,Lee MJ.Emerging roles of the ubiquitinproteasome system in the steroid receptor signaling.Arch Pharm Res 2012;35(3):397-407
41 杨洁,张秋养,葛玲,等.青春期大鼠实验性精索静脉曲张对附睾雄激素受体的影响.西安交通大学学报·医学版2005;26(1):22-25