(\beta\)-lactolin, a Monoamine Oxidase B Inhibitory Lactopeptide, Suppresses Reactive Oxygen Species Production in Lipopolysaccharide-Stimulated Astrocytes  
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Correspondence: [PERSON]]  
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1 Kirin Central Research Institute, Kirin Holdings Company, Limited, 2-26-1 Muraoka-Higashi, Fujisawa, Kanagawa 251-8555, Japan; [PERSON] (T.A.); [PERSON] (C.T.); [PERSON]DRESS] (R.O.); [PERSON]SS] (Y.A.)  
[PERSON]  
1 Kirin Central Research Institute, Kirin Holdings Company, Limited, 2-26-1 Muraoka-Higashi, Fujisawa, Kanagawa 251-8555, Japan; [PERSON] (T.A.); [PERSON] (C.T.); [PERSON]DRESS] (R.O.); [PERSON]SS] (Y.A.)  
evidence has revealed that DA is crucial for cognitive functions. DA is directly associated with frontal cortex-dependent cognitive functions, including attention, executive function, learning and memory processes [8; 9]. Several studies have shown that DA depletion and neuronal loss in the brain alter cognitive impairment in mice, indicating that restoring the DA system could contribute to the improvement of impaired cognitive performance [10; 11].  
The main enzymes responsible for DA metabolism are monoamine oxidase B (MAO-B) and catechol-O-methyltransferaseanferase (COMT), both of which are highly expressed in astrocytes [12; 13]. MAO-B inhibitors are widely used for the treatment of PD, since MAO-B inhibition increases the synaptic DA concentration [7]. Recent studies have demonstrated that some MAO-B inhibitors have preventive and therapeutic effects on the development of AD [14; 15]. In contrast, MAO-B is known to be an important source of ROS in astrocytes, because it generates hydrogen peroxide (H\({}_{2}\)O\({}_{2}\)) during enzymatic reactions [16; 17]. Several studies have shown that H\({}_{2}\)O\({}_{2}\) production in reactive astrocytes is increased in an MAO-B-dependent manner and an MAO-B inhibitor suppressed ROS production and neuronal loss in transgenic mouse models of AD and PD [14; 18; 19].  
Recently, food-derived MAO-B inhibitory peptides have been identified [20]. \(\beta\)-lactolin, a \(\beta\)-lactopeptide of glycine-threonine-tryptophan-tyrosine (GTWY), is derived from \(\beta\)-lactoglobulin containing tryptophan-tyrosine (WY) sequences, which is abundantly present in fermented dairy products, such as camembert cheese, and in whey proteins digested by certain enzymes. We found that \(\beta\)-lactolin and its core sequence dipeptide, WY, inhibit the enzymatic activity of human recombinant MAO-B [20]. We demonstrated that \(\beta\)-lactolin administration in mice increased DA levels in the hippocampus and improved memory function [20], which was attenuated by knocking down D1-like receptors in the hippocampus [21]. These findings indicated that \(\beta\)-lactolin improves memory function by regulating the dopaminergic system in mice. In addition, \(\beta\)-lactolin prevented neural inflammation in the mouse model of AD [22] and \(\beta\)-lactolin supplementation improved cognitive function in randomized clinical trials [23; 24]. However, the detailed mechanisms of \(\beta\)-lactolin, especially its direct effects on astrocytes, remain unclear.  
In the present study, we investigated the effects of \(\beta\)-lactolin on cultured murine astrocytes. Lipopolysaccharide (LPS) and interferon gamma (IFN-Y) were used to induce reactive astrocytes as previously described [25]. We then evaluated the effects of \(\beta\)-lactolin on DA metabolism in astrocytes by measuring MAO-B enzyme activity and intracellular and extracellular monoamine levels in DA-pretreated astrocytes. This study demonstrated the novel effects of \(\beta\)-lactolin on astrocytes and indicates the involvement of astrocytes in the memory-improving effects of \(\beta\)-lactolin.