5 Heat stress during pregnancy compromises intrauterine development and reproductive parameters of female progeny in C57BL/6J mice
Authors: Bento T. F. M. , da Silva J. R. N. , Souza-Fabjan J. M. G. , Silva R. C. , Silva S. B. , Dias I. R. , Moura C. R. F. , Gino I. O. , Batista R. I. T. P. 
Published: January 2021

Heat stress (HS) affects the reproduction of many species, causing subfertility by reducing gametogenesis. This study assessed the effect of HS at different stages of pregnancy in C57BL/6J mice on the somatic development and reproductive parameters of F1 females. A total of 40 females and 40 males aged between 5 and 6 weeks old were mated (1:1). After mating confirmation (vaginal plug presence) the females were subjected to HS during pregnancy in the first half (FP, from Day 1 to 10; n=10), the second half (SP, from Day 11 to delivery; n=10), or the total pregnancy (TP, n=10). A control group (C, n=10) was maintained in normothermic conditions (25°C, 45% relative humidity) throughout the experiment. The HS was induced (41°C for 2h daily) in an environmental chamber heated by 2 red lamps. After delivery, birthweight was recorded and somatic development of the F1 females was monitored weekly until 8 weeks of age. They were superovulated with 5IU of equine chorionic gonadotrophin (eCG) and 5IU of human chorionic gonadotrophin (hCG) 48h later and mated with control F1 males in four groups: FP×C; SP×C; TP×C; and C×C, female and male, respectively. At 72h after mating confirmation, uterine flushing was performed with 0.5mL of phosphate-buffered saline + 0.4% bovine serum albumin and embryos classified. Ovaries were collected for histological analysis of the follicular population with the formula: follicles per ovary×n section×section thickness/n section observed×average diameter of the oocyte nucleus. Pregnancy rate was analysed by chi-squared test. Data of pups born per female, birthweight, somatic development, follicular population, total and viable structures recovered by female were tested for normality by the Shapiro-Wilk test, before ANOVA and Tukey test. Values of P&lt;0.05 were considered to indicate a difference and P&lt;0.10a tendency. No difference (P&gt;0.05) among groups was detected in pregnancy rate (C=80; FP=40; SP=60; TP=60%) or in the number of pups born per female (C=7.0±1.0; FP=6.0±1.5; SP=6.5±1.5; TP=5.1±1.5). Birthweight was lower (P&lt;0.05) for FP (1.1g) and TP (1.2g) than for C (2.2g) and SP (1.8g). However, this difference disappeared (P&gt;0.05) in the third week of development and remained similar until the eighth week (C=21.0; FP=20.4, SP=20.3, TP=20.0g). Similar (P&gt;0.05) follicular population by ovary (total, primordial, primary, secondary, and antral) was observed between the C and HS groups. However, among HS groups, the total number of follicles and number of primordial follicles, respectively, were lower (P&lt;0.05) in the FP (1623; 942) compared with SP (2735; 1918) and TP (2626; 2352); with no difference in primary, secondary, and antral follicles. This resulted in similar (P&gt;0.05) total number of structures recovered by females (C=11.8±5.0, FP=7.6±2.4, SP=10.8±5.5, TP=6.9±3.1), with a tendency (P=0.06) to fewer viable embryos in TP (4.3±2.7) compared with C (9.5±4.6). Considering the increasing global temperature, it is imperative to understand the effects of HS on animal reproductive capacity. In conclusion, gestational HS impaired mice intrauterine development and changed the ovarian follicular population in the F1 generation.