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Assays useful for measuring sperm chromatin quality


Routine semen analysis does not absolutely predict the quality of sperm chromatin, which may help to explain why it often fails to predict reproductive outcomes in infertile couples.

The study's authors suggested that sperm chromatin quality evaluation could provide additional information that can facilitate decision making in male fertility management during their presentation at the AUA annual meeting in Anaheim, CA.

"Routine semen analysis using World Health Organization reference values is the most commonly ordered evaluation in this population, and decisions on patient management rely heavily on its results," said Peter T. Chan, MD, director of male reproductive medicine in the department of urology at McGill University Health Centre, Montreal. "Unfortunately, other than at extreme values, semen analyses fail to predict reproductive outcomes.

A total of 21 men in the study presented with idiopathic infertility, while 19 others were healthy community volunteers. In ad-dition, the study included 12 men with advanced testicular cancer (>2 months post-orchiectomy) and 11 others with Hodgkin's lymphoma.

"The choice of idiopathic infertility patients as subjects and healthy volunteers as controls was obvious," Dr. Chan said. "But we felt that broadening our population to include those with some of the most common cancers affecting men at reproductive age-who are often facing the decision of whether or not they should bank their sperm-might help to generate research data that can help in counseling them regarding fertility preservation."

Patients ranged in age from 21 to 48 years. Each underwent a detailed history and physical examination, as well as a morning serum hormonal profile (follicle-stimulating hormone, luteinizing hormone, and total testosterone), and answered a panel of standardized questions to control for psychosocial stress levels.

After patients' semen parameters were analyzed according to WHO criteria, their sperm chromatin quality was assessed with four complementary flow cytometry-based tests: acridine orange (AO) assay, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL), chromomycin A3 (CMA3), and monobromobimane thiol labeling (mBBr) assay.

"These assays have been routinely used in our laboratory for years for basic science research," he said. "Many research laboratories are capable of performing them. They are costly for clinical use simply because they aren't yet routine clinical tests, but I imagine that if their clinical value is soon established, they will become more popular and not as costly."

The assays measure sperm chromatin quality in different ways, but their results were closely coordinated in the McGill study. TUNEL assay results, for example, paralleled those from AO (p<.01) and mBBr (p<.005) across the entire range of semen parameters from all subjects.

On the other hand, there was no correlation found within any of the four patient groups among motile sperm density, normal morphology sperm density, and the four sperm chromatin assays.

Still, Dr. Chan noted, that does not mean there is no role for semen analysis in normal clinical practice.

"Routine semen analysis is still an important initial step for fertility evaluation, since it is inexpensive and can often be informative," he said. "But the clinician must realize that it does have significant limitations for the majority of patients with histories of clinical infertility.

"In the era of assisted reproductive technology, it would be difficult to base the management decision solely on the results of semen analysis, as it fails to fully disclose the quality of sperm."

Dr. Chan's group has already expanded its patient population to further evaluate the correlation of sperm chromatin assay results with routine semen parameters. Eventually, he said, the researchers will factor reproductive outcomes (natural or through assisted reproduction) into the equation.

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