Cyclical heat stress during lactation influences the microstructure of the bovine mammary gland

This study aimed to evaluate the effect of heat stress on mammary epithelial cell (MEC) losses into milk, secretory mammary tissue structure, and mammary epi-thelial cell activity. Sixteen multiparous Holstein cows (632 +/- 12 kg BW) approximately 100 DIM housed in climate-controlled rooms were paired by BW and ran-domly allocated to one of 2 treatments, heat stress (HS) or pair-feeding thermoneutral (PFTN) using 2 cohorts. Each cohort was subjected to 2 periods of 4 d each. In period 1, both treatments had ad libitum access to a common TMR and were exposed to a controlled daily temperature-humidity index (THI) of 64. In period 2, HS cows were exposed to controlled cyclical heat stress (THI: 74-80), while PFTN cows remained at 64 THI and daily DMI was matched to that of the HS cows. Cows were milked twice daily, and milk yield was recorded at each milking. Individual milk samples on the last day of each period were used to quantify MEC losses by flow cytometry using butyrophilin as a cell surface marker. On the final day of period 2, individual bovine mammary tis-sue samples were obtained for histomorphology analysis, assessment of protein abundance, and evaluation of gene expression of targets associated with cellular capacity for milk and milk component synthesis, heat response, cellu-lar proliferation, and autophagy. Statistical analysis was performed using the GLIMMIX procedure of SAS. Milk yield was reduced by 4.3 kg by HS (n = 7) compared with PFTN (n = 8). Independent of treatment, MEC in milk averaged 174 cells/mL (2.9% of total cells). There was no difference between HS and PFTN cows for MEC shed or concentration in milk. Alveolar area was reduced 25% by HS, and HS had 4.1 more alveoli than PFTN. The total number of nucleated MEC per area was greater in HS cows (389 +/- 1.05; mean +/- SE) compared with PFTN (321 +/- 1.05); however, cell number per alveolus was similar between groups (25 +/- 1.5 vs. 26 +/- 1.4). There were no dif-ferences in relative fold expression for GLUT1, GLUT8, CSN2, CSN3, LALBA, FASN, HSPA5, and HSPA8 in HS cows compared with PFTN cows. Immunoblotting analy-ses showed a decrease in abundance for phosphorylated STAT5 and S6K1, and an increase in LC3 II in HS cows compared with PFTN cows. These results suggest that even if milk yield differences and histological changes occur in the bovine mammary gland after 4 d of heat ex-posure, MEC loss into milk, nucleated MEC number per alveolus, and gene expression of nutrient transport, milk component synthesis, and heat-stress-related targets are unaffected. In contrast, the abundance of proteins related to protein synthesis and cell survival decreased signifi-cantly, whereas proteins associated with autophagy were upregulated in HS cows compared with PFTN cow