Response to “Sperm sorting for selection of healthy sperms: is it safe and useful?“ by Ettore Caroppo, M.D.

21 06 2013

To the Editor:

Although we agree with Dr. Caroppo that the effect of DNA damage of human spermatozoa on fertilization, embryo cleavage, and implantation remains controversial, its detrimental impact on natural conception and miscarriage seems real. Advanced methods for the selection of spermatozoa with healthy DNA will become mandatory, as advocated by Dr. Caroppo (1). As we are now able to significantly reduce the number of spermatozoa with fragmented DNA (2), we will be able to compare prospectively the effect of DNA fragmentation on the outcome of assisted reproductive technology (ART).

Of course, the safety of the process is of concern. However, the safety of the Hoechst dye in combination with fluorescence activated cell sorting (FACS) has been amply documented as much mammalian offspring has been produced with sorting (3). All evidence to date indicates that this dye exerts no detrimental impact. Similarly, the UV laser was described as not having any detectable negative effect on the incidence of DNA damage or mutations in spermatozoa. Only the mechanical aspects of sorting may be detrimental, as the pressure exerted on spermatozoa during their passage through the microfluidic channels has been demonstrated to affect their viability, motility, and velocity in the presence of high pressure (50 psi), but not of lower pressure (30 psi). Here, we used milder pressure (around 20 psi), which can be decreased even further. Due to breakage of the sperm tails, motility is lost during the formation and disruption of little droplets (4). Dr. Caroppo commented that in our study the sperm motility was significantly reduced when compared with the swim-up (3). As mentioned in our report (2), this difference is considerably reduced if we limit the samples to those with lowest quality (concentration ≤10 x 106/ml). In these cases the values are 51.6% +/-14 for sorted spermatozoa and 33% +/-22 in the swim-up. Read the rest of this entry »

Presence of M540 bodies in human semen: techniques to detect them require attention

20 06 2013

To the Editor:

We read with interest the paper by Gomez-Lopez et al., published in Fertility and Sterility (1), in which the authors investigated the presence of merocyanine 540 bodies (M540 bodies) and their impact on the detection of sperm apoptotic markers, including sperm DNA fragmentation.
In this paper the authors report that the incidence of M540 bodies in the semen of infertile men is very low (about 1%) and that their occurrence does not affect the determination of sperm DNA Fragmentation (sDF) by TUNEL coupled to flow cytometry.

These results contrast with previous data by our group that first described M540 bodies (2) and later on demonstrated that they are apoptotic bodies of testicular origin (3). Indeed, we found that M540 bodies can be present in high amount in semen of sub/infertile men (2, 3) and that they heavily flaw the determination of TUNEL positive sperm by flow cytometry (4). We believe that the cause of such discrepancies is the technique that the authors used to reveal M540 bodies in TUNEL processed samples. They first stained by merocyanine 540 and then washed and processed samples by TUNEL assay. By this procedure, merocyanine labeling is washed away from the bodies, since M540 does not bind in a stable (covalent) manner to bodies. As a consequence, they failed to detect M540+ elements, not because of their absence but because of the loss of merocyanine staining. Read the rest of this entry »

Reply to editorial regarding Increased risk of cancer among azoospermic men

13 06 2013

To the Editor:

We thank the distinguished author for bringing attention to our work.

As Dr. Schlegel points out, a natural conclusion to draw from the manuscript is that a male factor evaluation is of critical importance for a man’s reproductive and overall health (1). Indeed, it is estimated that 20% of infertile couples do not receive a male evaluation in the U.S.(2) As is noted, the current estimates may be an underestimate of a man’s lifetime risk. The current analysis followed men for a relatively short period of time (up to 15 years) compared with a man’s complete lifespan (76.3 years) (3, 4). Thus, it is possible that his lifetime risk of cancer would continue to rise as he ages.

Dr. Schlegel does point out some limitations related to the granularity of information on each man. As is noted, infertile men are generally of higher socioeconomic status compared with the general population (5). However, access to care should be independent of semen parameters. Thus it is unlikely that the elevated cancer risk seen in azoospermic men could be entirely explained by socioeconomic factors. We look forward to seeing other groups substantiate our findings.

Michael Louis Eisenberg, M.D. , Stanford University School of Medicine, Stanford, California
Paul Betts, M.S., Cancer Epidemiology and Surveillance Branch, Texas Cancer
Registry, Texas Department of State Health Services, Austin, Texas
Danielle Herder, M.D., Baylor College of Medicine, Houston, Texas
Dolores Lamb, Ph.D., Baylor College of Medicine, Houston, Texas
Larry Lipshultz, M.D., Baylor College of Medicine, Houston, Texas


1. Schlegel PN. The relevance of increased cancer risk in infertile men. Fertil Steril 2013.

2. Eisenberg ML, Lathi RB, Baker VL, Westphal LM, Milki AA, Nangia AK. Frequency of the male infertility evaluation: data from the national survey of family growth. J Urol 2013;189:1030-4.

3. Murphy SL, Xu J, Kochanek KD. Deaths: final data for 2010. Natl Vital Stat Rep 2013;61:1-167.

4. Eisenberg ML, Betts P, Herder D, Lamb DJ, Lipshultz LI. Increased risk of cancer among azoospermic men. Fertil Steril 2013.

5. Hotaling JM, Davenport MT, Eisenberg ML, VanDenEeden SK, Walsh TJ. Men who seek infertility care may not represent the general U.S. population: data from the National Survey of Family Growth. Urology 2012;79:123-7.

Published online in Fertility and Sterility doi:10.1016/j.fertnstert.2013.06.025