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. 2024 Apr 24;14(9):1275.
doi: 10.3390/ani14091275.

SLC34A2 Targets in Calcium/Phosphorus Homeostasis of Mammary Gland and Involvement in Development of Clinical Mastitis in Dairy Cows

Xueying Wang  1   2 , Bohao Zhang  1   2 , Weitao Dong  1   2 , Yu Zhao  3 , Xingxu Zhao  1   2   3 , Yong Zhang  1   2   3 , Quanwei Zhang  1   2   3
Affiliations

SLC34A2 Targets in Calcium/Phosphorus Homeostasis of Mammary Gland and Involvement in Development of Clinical Mastitis in Dairy Cows

Xueying Wang et al. Animals (Basel). .

Abstract

The type II Na/Pi co-transporter (NaPi2b), encoded by the solute carrier (SLC) transporter 34A2 (SLC34A2), is responsible for calcium (Ca) and phosphorus (P) homeostasis. Unbalanced Ca/P metabolism induces mastitis in dairy cows. However, the specific role of SLC34A2 in regulating this imbalance in Holstein cows with clinical mastitis (CM) remains unclear. The aim of this study was to investigate the role of SLC34A2 and identify differentially expressed proteins (DEPs) that interact with SLC34A2 and are associated with Ca/P metabolism in dairy cows with CM. Immunohistochemical and immunofluorescence staining results showed that SLC34A2 was located primarily in the mammary epithelial cells of the mammary alveoli in both the control (healthy cows, Con/C) and CM groups. Compared to the Con/C group, the relative expression of the SLC34A2 gene and protein were significantly downregulated in the CM group. We identified 12 important DEPs included in 11 GO terms and two pathways interacting with SLC34A2 using data-independent acquisition proteomics. The PPI (protein-and-protein interaction) network results suggested that these DEPs were associated with ion metabolism and homeostasis, especially SLC34A2. These results demonstrate that SLC34A2 downregulation is negatively correlated with the occurrence and development of CM in Holstein cows, providing a basis for exploring the function and regulatory mechanism of SLC34A2 in Ca/P metabolism and homeostasis in Holstein cows with CM.

Keywords: SLC34A2; calcium/phosphorus homeostasis; mastitis; proteomics.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Distribution and co-localization analysis of SLC34A2 protein in MGs. (A1–B2) IHC staining shows SLC34A2 distribution in MGs of Con/C (A1,A2) and CM (B1,B2) groups. (C1–D2) SLC34A2 negative controls for Con/C (C1,C2) and CM (D1,D2) groups. (E1–F2) Localization in MGs: nuclei (blue, E1,F1), CK18 (red, E2,F2), SLC34A2 (green, E3,F3), with merged CK18 and SLC34A2 (E4,F4) for Con/C and CM, respectively. Con/C represents healthy Holstein cows; CM indicates cows with clinical mastitis. NC: negative control; MA: mammary alveoli; MECs: mammary epithelial cells; NEUT: neutrophil. Scale bars: 50 nm (200× magnification) and 100 nm (400× magnification).
Figure 2
Figure 2
Expression pattern analysis of SLC34A2 mRNA and protein in MGs. (A) mRNA levels of SLC34A monitored by qRT-PCR assay. (B) Protein bands of SLC34A monitored by Western blot assay; the whole Western blots are shown in Figures S1 and S2. (C) Optical density of bands. Con/C, control. CM, clinical mastitis. Data are presented as means ± SEM. ** represents p < 0.01.
Figure 3
Figure 3
Identification of candidate DEPs and GO terms related to Ca/P metabolism and involved with SLC34A2 between control and clinical mastitis groups based on GO enrichment data. (A) Candidate DEPs and GO terms with significance of p < 0.01 and p.adjust < 0.05 related to Ca/P metabolism and involving SLC34A2. X-axis represents the number of DEPs. Y-axis represents GO terms including biological process (BP), molecular function (MF), and cellular component (CC). (B) Venn diagram of candidate DEPs in BP, MF, and CC groups. (C) Volcano plots of 18 DEPs, including 5 downregulated and 13 upregulated DEPs. (D) Heat map of 18 candidate DEPs. (E) PPI network analysis of 18 DEPs and 11 GO terms related to Ca/P metabolism.
Figure 4
Figure 4
Identification of candidate DEPs and pathways related to Ca/P metabolism and involved with SLC34A2 between the control and clinical mastitis groups based on pathways. (A) Candidate DEPs and pathways related to Ca/P metabolism and involving SLC34A2. (B) PPI network and heat map analysis of mineral absorption and PTH synthesis, secretion, and action pathways related to Ca/P metabolism involving SLC34A2. (C) Volcano plots of 14 DEPs: 1 downregulated and 13 upregulated.
Figure 5
Figure 5
Identification of candidate DEPs related to Ca/P metabolism and involved with SLC34A2 between the control and clinical mastitis groups based on the GO enrichment and pathway data. (A) Venn diagram of candidate DEPs in the merged GO terms and pathways. (B) Relative expression levels of 12 DEPs quantified by DIA proteomics; the y-axis represents the log2 (FC) values. (C) Sankey diagram of the 11 GO terms, 12 shared DEPs, and two pathways related to Ca/P metabolism and ion homeostasis.
Figure 6
Figure 6
PPI network analyses of the candidate DEPs, GO terms, and pathways in ion metabolism and homeostasis in the MGs between the control and clinical mastitis groups.
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