Abstract
BACKGROUND: The blood-brain barrier (BBB) expresses transporters, receptors, and tight junction proteins that regulate the exchange of substances between the blood and brain. The differences in the expression of these proteins in the BBB among different species and cultured BBB model cells should be clarified to interpret the BBB function in model animals and cells. This study aimed to elucidate species differences among humans, monkeys, and mice and in vitro-in vivo differences in the BBB proteome using deep proteomic analysis. METHODS: Brain microvessels (BMVs) were isolated from frozen cerebral cortices of human and monkey and frozen mouse cerebrums. BMVs and cultured brain microvascular endothelial cells (BMECs), such as hCMEC/D3, HBMEC/ciβ, and primary BMECs, were analyzed via data-independent acquisition using liquid chromatography-mass spectrometry. RESULTS: Proteomics identified 7,149-8,274 proteins in the BMV fractions and 6,657-7,534 proteins in the brain lysates. Comparative analysis revealed distinct proteomic profiles among the three species, with the human profile being more similar to that of monkeys than that of mice. The expression profile of the solute carrier organic anion transporter family was found to vary among mouse, monkey, and human BMVs. The expression levels of SLC22A6/Slc22a6 and SLC22A8/Slc22a8 were higher in mice than in monkeys and humans, whereas SLC43A3/Slc43a3 expression levels were lower in mice. The expression of amino acid transporters, such as SLC7A5 and SLC3A2/Slc3a2, was higher in BMVs, whereas that of SLC1A5/Slc1a5 and SLC38A9/Slc38a9 was higher in cultured BMECs. MFSD2A/Mfsd2a and SLC27A1/Slc27a1 were highly expressed in BMVs. The expression of tight junction proteins, particularly the claudin family, varied between BMVs and cultured BMECs and among cell lines. Specifically, the expression of claudin-5 was higher in BMVs, and claudin-11 expression was higher in cultured BMECs. CONCLUSIONS: Deep proteomic analysis revealed species-specific differences in transport-related proteins in the BBB. Furthermore, in vitro and in vivo differences were observed in the transporter and claudin protein expression. This study provides a BBB proteome profile dataset and offers insights for a comprehensive understanding of BBB protein expression across species and between in vivo and in vitro conditions.