Alpha-Lipoic Acid Ameliorates Sperm DNA Damage and Chromatin Integrity in Men with High DNA Damage: A Triple Blind Randomized Clinical Trial

Abstract

Objective: Evidence suggests the contributory role of oxidative stress (OS) to sperm DNA damage and eventually, male infertility. Antioxidant supplementation has exhibited favorable results regarding seminal OS, sperm DNA damage, and chromatin integrity. We aimed to evaluate the effect of alpha-lipoic acid (ALA) supplementation on semen analysis, sperm DNA damage, chromatin integrity, and seminal/intracellular OS in infertile men with high sperm DNA damage.

Materials and methods: In this randomized triple-blind placebo-controlled clinical trial study, we opted for a triple-blind controlled clinical trial design. Considering the study's inclusion criteria for the level of sperm DNA fragmentation (higher than the threshold of 30 and 15%), 70% of participants were selected for this clinical research study. Subjects were divided into case and control groups receiving oral ALA (600 mg/day) and placebo for eighty days, respectively. Sperm parameters and functional tests were examined and compared before and after treatment. The final sample size was 34 and 29 for ALA and placebo receivers, respectively.

Results: No significant differences were observed about anthropometrics and baseline measures of semen analysis, DNA damage, OS, and chromatin integrity between the two groups. Conventional semen parameters were enhanced insignificantly in both groups (P>0.05). DNA damage decreased significantly in the ALA group, as per sperm chromatin structure assay (SCSA, P<0.001). Moreover, chromomycin A3 (CMA3) staining results indicated a decrease in nuclear protamine deficiency post-ALA therapy (P=0.004). Lipid peroxidation decreased significantly after treatment with ALA (P=0.003). Further, seminal antioxidant capacity/activity did not differ significantly in either of the groups (registration number: IRCT20190406043177N1).

Conclusion: An 80-day course of oral ALA supplementation (600 mg/day) alleviates sperm OS, DNA damage, and chromatin integrity in men with high sperm DNA damage.

Keywords: Alpha-lipoic acid; DNA damage; Male infertility; Sperm.

Fig 1

Study design according to CONSORT.

Fig 2

Assessment of sperm functional tests. A. Assessment of sperm morphology with Diff- Quick staining, B. Representative histograms of sperm chromatin structure assay (SCSA) test before and after ALA therapy in an infertile man, C. Chromomycin A3 (CMA3) staining for assessment of sperm protamine deficiency; CMA3 negative or the sperm with normal protamine content and CMA3 positive or protamine deficient sperm, and D. Representative histograms of sperm JC1 staining for assessment of mitochondrial membrane potential before and after alpha-lipoic acid (ALA) therapy in an infertile man (20 μm=100 magnification).

Fig 3

Comparison between the mean measures of seminal analysis parameters before and after intervention (paired t test). A. Semen volume, and sperm B. Concentration, C. Total motility, D. Progressive motility, E. Viability, and F. Normal morphology. ALA; Alpha lipoic acid.

Fig 4

Intragroup comparison between the mean values of sperm DNA damage before and after intervention (paired t test). Examined by A. SCSA and B. TUNEL assays, and the level of sperm chromatin compaction/protamination as indicated by the results of C. Aniline blue and D. CMA3 staining techniques. *; Denotes P<0.05, ALA; Alpha lipoic-acid, SCSA; Sperm chromatin structure assay, TUNEL; Terminal deoxynucleotidyl transferase dUTP nick end labeling, and CMA3; Chromomycin A3.

Fig 5

Comparison between levels of seminal oxidative determinants before and after intervention. A. Total antioxidant capacity, B. Superoxide dismutase, C. Malondialdehyde, and D. Mitochondrial membrane potential. *; Denotes P<0.05, ALA; Alpha lipoic-acid, TAC; Total antioxidant capacity, SOD; Superoxide dismutase, MDA; Malondialdehyde, MMP; Mitochondrial membrane potential, and AU; Arbitrary unit.