Oral niacin mitigates heat-induced reproductive impairments in male mice

Abstract

Heat stress is a major environmental factor that impairs male fertility by disrupting hormonal regulation, increasing oxidative stress, and inducing germ cell apoptosis. Niacin (vitamin B3) has antioxidant and thermoregulatory properties that may counteract these effects. This study investigated whether oral niacin could protect against chronic heat stress-induced reproductive damage in adult BALB/c male mice. Mice were randomly assigned to Control, Heat Control, and two heat-stressed groups receiving niacin at 100 or 200 mg/kg/day for 30 days, with heat stress applied daily at 36 ± 2 °C for 4 h. Heat stress reduced body weight gain, testis and accessory gland weights, sperm count and motility, acrosome and DNA integrity, testosterone and thyroid hormones, and increased lipid peroxidation and pro-apoptotic gene expression. Niacin supplementation dose-dependently mitigated these effects: total sperm count increased from 9.8 × 10⁵ ± 1.85 × 10⁵ to 3.66 × 10⁶ ± 4.62 × 10⁵, total motility from 49.0 ± 13.6% to 81.7 ± 5.8%, testosterone from 0.89 ± 0.21 to 4.44 ± 0.70 ng/mL, and Bcl2 expression increased 2.7-fold. Fertility improved, with pregnancy risk rising from 50% to 100%, litter size from 3.7 ± 0.3 to 7.2 ± 0.3 pups, and blastocyst formation from 37.8 ± 2.7% to 68.0 ± 2.6%. Niacin also normalized stress-related gene expression, preserved testicular histology, and did not affect liver enzyme activities, highlighting its potential as a dietary supplement to protect against heat-induced male reproductive dysfunction.

Keywords: Heat stress; Male fertility; Niacin; Oxidative stress; Sperm quality; Thermoregulation.

Fig. 1

A one-way ANOVA followed by LSD confidence interval adjustments was employed to compare values between various study groups on each distinct time points. Asterisks denote statistically significant differences (P ≤ 0.05) between each niacin group and the heat stress group.

Fig. 2

A one-way ANOVA followed by LSD confidence interval adjustments was employed to compare values between various study groups on each distinct time points. Asterisks denote statistically significant differences (P ≤ 0.05) between each niacin group and the heat stress group. RBT, Rectal Body Temperature; SB, Surface Body Temperature. 10, 20, and 30 indicate the days after the start of the heat stress period.

Fig. 3

Representative images of mouse gametes and embryos. (A) Bulged oviducts after superovulation containing expanded cumulus–oocyte complexes (arrowheads). (B) Early and expanded blastocysts on day 5 of in vitro culture compared with retarded or degenerated embryos. (C) 4-cell stage embryos on day 2 of culture. (D,E) Chlortetracycline-stained sperm showing non-capacitated (F pattern) and capacitated (B pattern) cells. (F–H) Acridine orange-stained sperm highlighting mature (green) versus immature or protamine-deficient (yellow/reddish) nuclei. (I,J) Eosin–nigrosin-stained sperm illustrating live (colorless heads) versus dead (red heads) cells. (K) Coomassie Brilliant Blue-stained sperm showing acrosome-intact (blue line, arrowheads) versus acrosome-reacted/damaged spermatozoa. (L) Sperm chromatin dispersion assay displaying non-fragmented DNA with large halos around nuclei.

Fig. 4

A one-way ANOVA followed by LSD confidence interval adjustments was employed to compare values between various study groups on each distinct time points. Asterisks denote statistically significant differences (P ≤ 0.05) between each niacin group and the heat stress group.

Fig. 5

Effects of chronic heat stress and oral niacin supplementation on histological and morphometric parameters of testicular tissue in mice at Days 15 and 30. (A) Spermatogenesis index (%), (B) Tubular differentiation index (%), (C) Number of interstitial cells, (D) Number of spermatocytes, (E) Diameter of seminiferous tubules (µm), (F) Epithelial thickness (µm), and (G) Johnsen’s score were evaluated in four groups: Control (black), Heat Control (red), Niacin 1 (green; lower dose), and Niacin 2 (blue; higher dose).Bars represent mean ± SEM. Columns with different letters (a, b, c) are significantly different at p < 0.05.

Fig. 6

Histological structure of the testis in different groups during the 15-day study period (H&E staining). Degenerated seminiferous tubules (arrows), disruption of cellular structural organization, and a marked reduction in epithelial thickness and seminiferous tubule diameter are evident in the HC group compared to the other study groups.

Fig. 7

Histological structure of the testis in different groups during the 30-day study period (H&E staining). Degenerated seminiferous tubules (arrows), disruption of cellular structural organization, and a marked reduction in epithelial thickness and seminiferous tubule diameter are evident in the H group compared to the other study groups.