Fig 2: Astrocytes derived from sporadic Alzheimer’s disease iPSCs show higher gelatinase activity. (a) Representative zymograph image using astrocyte-conditioned media (ACM) from control, sporadic and familial iPSC-derived astrocytes. (b) Representative zymograph image of ACMs from control, sporadic and familial iPSC-derived astrocytes pretreated with TNFα and IL1β. (c) Schematics showing experimental design for the detection of MMPs and tau degradation analysis. (d) Representative zymograph image using ACM from control, sporadic and familial AD astrocytes treated with monomeric tau for 0 h, 24 h and 48 h. Note the moderate increase in gelatinase activity at 48 h of ACM from sAD astrocytes (asterisk). (e) Graphs showing difference in MMP activity after exposure to monomeric tau between the ACM from three groups. (f) Representative zymograph image using ACM from NDC, sporadic and familial AD astrocytes treated with aggregated tau for 0 h, 24 h and 48 h. Increase in gelatinase activity at 48 h of ACM from sAD astrocytes was observed (asterisk). (g) Graph showing difference in gelatinase activity of MMPs after exposure to aggregated tau between the three groups. (h) Degradation of aggregated tau by proteases in the ACM from three different iPSC-derived astrocytes. The protease activity is selectively inhibited by MMP2 and MMP9 inhibitors. (i) Graph showing percentage of tau degradation between the groups. One-way ANOVA followed by Tukey’s multiple comparisons tests, n = 3/group; n = 3 experimental repeats, * p < 0.05, ** p < 0.01, and *** p < 0.001.

Fig 3: Schematic showing differential responses in the control and Alzheimer’s disease iPSC-derived astrocytes. (a) Astrocytes generated from Alzheimer’s disease (sAD and fAD) show increased glutamate-induced calcium transients in terms of amplitude and duration when compared to healthy astrocytes (NDC). (b) Extracellular free tau, which is neurotoxic, is taken up by astrocytes via heparin sulphate proteoglycan (HSPG), a process mitigated by heparin. Upon tau exposure, only AD astrocytes exhibit GFAP upregulation and morphological reshaping, attributes of reactive state. Tau is processed in the endo-lysosomal compartment (c) and degraded (d). Meanwhile, the uptake of tau activates ERK pathway with highest levels of pERK, EAAT-2 and pPKC in sporadic AD-derived astrocytes (sAD), leading to the upregulation of astrocytic pro-MMP-2 and MMP-9 (e), which results in differential activation and secretion (f). The capacity to degrade and attenuate extracellular tau was the highest when using the secretome of sAD and NDC astrocytes compared to fAD, and showed a neuroprotective effect accompanied by an increase in the levels of neuronal NMDA, GAP43 and drebrin.