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Article of the Month: The Effect of Smoking on Sperm Functional Quality and Seminal Plasma Proteomic Profile

Every Month the Editor-in-Chief selects an Article of the Month from the current issue of BJUI. The abstract is reproduced below and you can click on the button to read the full article, which is freely available to all readers for at least 30 days from the time of this post.

In addition to the article itself, there is an accompanying editorial written by a prominent member of the urological community. This blog is intended to provoke comment and discussion and we invite you to use the comment tools at the bottom of each post to join the conversation.

If you only have time to read one article this week, it should be this one.

Analysis of the functional aspects and seminal plasma proteomic profile of sperm from smokers

Mariana Pereira Antoniassi*, Paula Intasqui*, Mariana Camargo*, Daniel Suslik
Zylbersztejn*, Valdemir Melechco Carvalho, Karina H. M. Cardozo† and Ricardo
Pimenta Bertolla*

 

*Department of Surgery, Division of Urology, Human Reproduction Section, Sao Paulo Federal University, Fleury Group, and Hospital Sao Paulo, Sao Paulo, Brazil

 

Objective

To evaluate the effect of smoking on sperm functional quality and seminal plasma proteomic profile.

Patients and Methods

Sperm functional tests were performed in 20 non-smoking men with normal semen quality, according to the World Health Organization (2010) and in 20 smoking patients. These included: evaluation of DNA fragmentation by alkaline Comet assay; analysis of mitochondrial activity using DAB staining; and acrosomal integrity evaluation by PNA binding. The remaining semen was centrifuged and seminal plasma was used for proteomic analysis (liquid chromatography-tandem mass spectrometry). The quantified proteins were used for Venn diagram construction in Cytoscape 3.2.1 software, using the PINA4MS plug-in. Then, differentially expressed proteins were used for functional enrichment analysis of Gene Ontology categories, Kyoto Encyclopedia of Genes and Genomes and Reactome, using Cytoscape software and the ClueGO 2.2.0 plug-in.

aotm-nov-1-results

Results

Smokers had a higher percentage of sperm DNA damage (Comet classes III and IV; P < 0.01), partially and fully inactive mitochondria (DAB classes III and IV; P = 0.001 and P = 0.006, respectively) and non-intact acrosomes (P < 0.01) when compared with the control group. With respect to proteomic analysis, 422 proteins were identified and quantified, of which one protein was absent, 27 proteins were under-represented and six proteins were over-represented in smokers. Functional enrichment analysis showed the enrichment of antigen processing and presentation, positive regulation of prostaglandin secretion involved in immune response, protein kinase A signalling and arachidonic acid secretion, complement activation, regulation of the cytokine-mediated signalling pathway and regulation of acute inflammatory response in the study group (smokers).

Conclusion

In conclusion, cigarette smoking was associated with an inflammatory state in the accessory glands and in the testis, as shown by enriched proteomic pathways. This state causes an alteration in sperm functional quality, which is characterized by decreased acrosome integrity and mitochondrial activity, as well as by increased nuclear DNA fragmentation.

Reference: Grey Haze.

Editorial: Semen Proteome Alterations in the Smoking Male

Infertility affects 15% of couples, with male factor infertility influencing half of such couples [1]. Multiple environmental risk factors for male infertility have been identified, including: diet, exposure to pollution, and multiple substance abuses, e.g. alcohol, tobacco, marijuana, opiates [2].

In this issue of BJUI, the study of Antoniassi et al. [3] makes an important contribution to the literature regarding the impact of a particular environmental factor, smoking, on male factor infertility. Standard semen analysis does not have strong sensitivity or specificity for male factor infertility given that it only examines sperm. Guzick et al. [4] demonstrated this in a study that showed extensive overlap in the sperm motility, concentration, and morphology in the male partners of fertile and infertile couples. Given this, multiple aspects of functional sperm testing have been added to standard semen analysis. Proteins added to the semen by the male accessory sex organs play a vital role in male reproductive capacity, providing nourishment and protection to spermatozoa. Batruch et al. [5] identified >2300 proteins in the semen of both fertile and infertile men using mass spectrometry. The physiological role of most of these proteins is unknown. Proteomics identifies the protein complement present in the semen using mass spectrometry. Sharma et al. [6] reported that the proteome of men with reactive oxygen species present is altered and that the absence of certain proteins may impair the neutralization of oxidative stress in semen. Prior studies have shown that the protein complement of the semen is different in men with clinical varicocoeles [7].

While the concern about the impact of smoking on male reproductive health is well documented, the effect that it has on the semen proteome is unknown. Antoniassi et al. [3] compared the semen proteome in both non-smoking and smoking men who presented to their laboratory for semen analysis. Given that patients rather than volunteers were used in this study, one cannot easily generalize the study findings to the general male population. After testing the samples for mitochondrial activity, acrosome function, and DNA integrity, the samples underwent liquid chromatography/mass spectroscopy to analyze the proteome. Sperm from smokers showed lesser sperm integrity, higher DNA fragmentation rates, and a lower percentage of intact mitochondria. The investigators demonstrated that smoking causes changes in the semen proteome. In smoking men one protein was absent, 27 proteins were under-represented, and six were over-represented. This study [3] provides some insight into how certain proteins may impact fertility. The protein S100A9 is overexpressed in smoking men and is associated with chronic inflammation, which corresponds with the fact that smokers frequently exhibit leukocytospermia. Mammoglobin B, the protein absent in men who smoke, is involved in the binding of steroid hormones; therefore, it is conceivable that its absence inhibits the ability of sperm to respond to sex hormones. you can find more info on Theihcc.com around these and other steroids.

This study [3] establishes that, among men who present for fertility evaluation, smokers have an altered proteome compared to non-smokers. This highlights the need for a greater understanding of how the semen proteome interacts with cellular elements. Further studies that examine the proteome of smokers and non-smokers with no history of infertility are warranted.

 

Kenneth J. DeLay and Wayne J.G. Hellstrom

 

Department of Urology, Tulane University School of Medicine, New Orleans, LA, USA

 

References

 

1 Sabanegh E Jr, Agarwal A. Male infertility. In Wein AJ, Kavoussi LR, Novick AC, Partin AW, Peters CA eds, Campbell-Walsh Urology10th edn, Philadelphia, PA: Elsevier Saunders: 2011

 

2 Barazani Y, Katz BF, Nagler HM, Stember DS. Lifestyle, environment, and male reproductive health. Urol Clin North Am 2014; 41: 5566

 

3 AntoniassiMP, Intasqui P, Camargo M et al.Analysisofthefunctionaaspects and seminal plasma proteomic proleofspermfromsmokers. BJU Int 2016; 118: 81422

 

4 GuzickDS, Overstreet JW, Factor-Litvak P et al.Spermmorphologymotility, and concentration in fertile and infertile men. NEnglJMed2001; 345: 138893

 

 

6 Sharma R, Agarwal A, Mohanty G et al. Proteomic analysis of seminal uid from men exhibiting oxidative stress. Reprod Biol Endocrinol 2013; 11: 85.

 

7 Zylbersztejn DS, Andreoni C, Del Giudice PT et al. Proteomic analysis of seminal plasma in adolescents with and without varicocele. Fertil Steril 2013; 99: 928

 

Video: The Effect of Smoking on Sperm Functional Quality and Seminal Plasma Proteomic Profile

Analysis of the functional aspects and seminal plasma proteomic profile of sperm from smokers

Mariana Pereira Antoniassi*, Paula Intasqui*, Mariana Camargo*, Daniel Suslik
Zylbersztejn*, Valdemir Melechco Carvalho, Karina H. M. Cardozo† and Ricardo
Pimenta Bertolla*
*Department of Surgery, Division of Urology, Human Reproduction Section, Sao Paulo Federal University, Fleury Group, and Hospital Sao Paulo, Sao Paulo, Brazil

Objective

To evaluate the effect of smoking on sperm functional quality and seminal plasma proteomic profile.

Patients and Methods

Sperm functional tests were performed in 20 non-smoking men with normal semen quality, according to the World Health Organization (2010) and in 20 smoking patients. These included: evaluation of DNA fragmentation by alkaline Comet assay; analysis of mitochondrial activity using DAB staining; and acrosomal integrity evaluation by PNA binding. The remaining semen was centrifuged and seminal plasma was used for proteomic analysis (liquid chromatography-tandem mass spectrometry). The quantified proteins were used for Venn diagram construction in Cytoscape 3.2.1 software, using the PINA4MS plug-in. Then, differentially expressed proteins were used for functional enrichment analysis of Gene Ontology categories, Kyoto Encyclopedia of Genes and Genomes and Reactome, using Cytoscape software and the ClueGO 2.2.0 plug-in.

aotm-nov-1-results

Results

Smokers had a higher percentage of sperm DNA damage (Comet classes III and IV; P < 0.01), partially and fully inactive mitochondria (DAB classes III and IV; P = 0.001 and P = 0.006, respectively) and non-intact acrosomes (P < 0.01) when compared with the control group. With respect to proteomic analysis, 422 proteins were identified and quantified, of which one protein was absent, 27 proteins were under-represented and six proteins were over-represented in smokers. Functional enrichment analysis showed the enrichment of antigen processing and presentation, positive regulation of prostaglandin secretion involved in immune response, protein kinase A signalling and arachidonic acid secretion, complement activation, regulation of the cytokine-mediated signalling pathway and regulation of acute inflammatory response in the study group (smokers).

Conclusion

In conclusion, cigarette smoking was associated with an inflammatory state in the accessory glands and in the testis, as shown by enriched proteomic pathways. This state causes an alteration in sperm functional quality, which is characterized by decreased acrosome integrity and mitochondrial activity, as well as by increased nuclear DNA fragmentation.

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