Expression profiles of immune genes in milk somatic cells and MIR predicted mineral contents in milk as indicators of mastitis

Despite rigorous management practices, mastitis remains a problem in high producing dairy cows. To identify early indicators of mastitis, the following parameters were evaluated: 1. expression profiles in milk somatic cells of L-Selectin (SELL), Interleukin 8 (IL8), and Gelsolin (GSN) genes; 2. MIR predicted mineral contents in milk. We used milk somatic cell samples collected at 3 lactation stages (DIM 7, 14, 21) from seventeen Holstein cows. Total RNA was extracted by using Maxwell 16 RNA purification kit and DNAse treated. RNA quality and quantity were assessed before performing RT and qPCR. Expression analyses, normalized on two reference genes selected by geNorm (ATPase and RPS9), were performed by using the qBasePLUS software. Furthermore, individual milk MIR spectra were collected at the same DIM, and prediction equations (Soyeurt et al., 2009) were used to predict Ca, K, Mg, Na, and P milk contents. Least square means for gene expressions and predicted mineral contents in healthy vs mastitis cows (Positive California Mastitis Test and SCC > 100,000 cells/ml in each of three samplings) were computed by Proc GLM in SAS. Correlations among all response variable and regressions of gene expressions on mineral contents were estimated by CORR and REG procedures in SAS. The 3 considered genes showed no differential expression at the 3 lactation stages, while their expressions were significantly different in mastitis vs healthy cows (p<0.05) in all of the 3 considered lactation stages. Mineral contents were significantly more concentrated at 7 day than in the other two days (p<0.05) and in mastitis vs healthy cows (p<0.05). A significant correlation was found between GSN expression and Na, Mg and K contents (R2=0.47, 0.34, 0.30, p<0.05). Also regressions of GSN expression on mineral contents were significant (p<0.05). Gene expression results corroborate our hypothesis of SELL, IL8 and GSN expression as indicators of mastitis, because of their roles in innate immunity. Results about mineral concentrations confirmed their concentration changes in milk during a mastitis. Therefore they could be reliable indicators of subclinical mastitis.