Figure 5. hLf inhibits APP phosphorylation via p38 and PP2Ac interaction-mediated upregulation of PP2A activity. (A) Aβ oligomer treatment increased the expression and secretion of Lf in the mouse primary astrocytes from WT mice. n = 5; one-way ANOVA. (B) Conditional astrocytic medium from Astro-Lf mice inhibited p-APP expression in N2a-sw cells. n = 3; Student’s t -test. (C) hLf treatment reduced the expressions of p-APP and sAPPβ, but not APP in N2a-sw cells. n = 5; one-way ANOVA. (D) hLf treatment diminished the Aβ42 content in the medium. n = 5; one-way ANOVA. (E) Double immunostaining showing the fluorescent intensities of Aβ and p-APP in control N2a-sw cells and hLf-treated N2a-sw cells. The Aβ intensities were both positively correlated with p-APP intensities in control N2a-sw cells (E1) and hLf-treated N2a-sw cells (E2), and the intensities of Aβ (E3) and p-APP (E4) were both decreased in hLf-treated N2a-sw cells. n > 25; Student’s t -test. (F) hLf treatment increased the expression of p-p38 but had no effects on the expressions of p38, p-CDK5, CDK5, p-GSK3α/β, GSK3α/β and PP2Ac in N2a-sw cells. n = 5; one-way ANOVA. (G) The PP2A activity was increased in hLf-treated N2a-sw cells. n = 5; one-way ANOVA. (H) Inhibition of p38 activity promoted p-APP expression, and also abrogated hLf-induced downregulation of p-APP in N2a-sw cells. n = 5; Two-way ANOVA followed by the Bonferroni’s post hoc test. (I) Activation of PP2A activity rescued the p38 inhibition-induced upregulation of p-APP in N2a-sw cells. n = 5; Two-way ANOVA followed by the Bonferroni’s post hoc test. (J) Inhibition of PP2A activity abrogated hLf-induced downregulation of p-APP in N2a-sw cells. n = 5; Two-way ANOVA followed by the Bonferroni’s post hoc test. (K) hLf treatment induced the interaction of PP2Ac and p38, while inhibition of p38 activity inhibited the interaction of PP2Ac and p38, and also abrogated the hLf-induced interaction of PP2Ac and p38 in N2a-sw cells. n = 5; Two-way ANOVA followed by the Bonferroni’s post hoc test. (L) Inhibition of p38 activity reduced the PP2A activity and abrogated the hLf-induced upregulation of PP2A activity in N2a-sw cells. n = 5; Two-way ANOVA followed by the Bonferroni’s post hoc test. (M-N) Overexpression of p38 enhanced the PP2A activity without changing the expression of PP2Ac in N2a-sw cells. n = 5; Student’s t -test. (O) Overexpression of p38 slightly reduced the APP expression, but sharply decreased the expressions of p-APP and sAPPβ in N2a-sw cells. n = 5; Student’s t -test. (P) LRP1 siRNA effectively knocked down the expression of LRP1 in N2a-sw cells. n = 3; Student’s t -test. (Q) LRP1 silence largely inhibited the hLf-induced upregulation of p-p38 and subsequently rescued the hLf-induced downregulation of p-APP in N2a-sw cells. n = 5; Two-way ANOVA followed by the Bonferroni’s post hoc test. *P < 0.05, **P < 0.01.