Journal: Cells

Article Title: Neurodegeneration Induced by Anti-IgLON5 Antibodies Studied in Induced Pluripotent Stem Cell-Derived Human Neurons

doi: 10.3390/cells10040837

Figure Lengend Snippet: Graphical illustration of the experimental setup. Short-term autoantibody exposure was performed on human Neuronal Stem cells (hNSC) and the experimental setup is shown in panel ( A ). Long-term autoantibody exposure was performed on human-induced Pluripotent Stem Cell (hiPSC)-derived neurons and the experimental setup is shown in panel ( B ). Black arrows and text specify differentiation procedures. Colored arrows and text boxes specify the time points at which different analyses were performed. LDH: lactate dehydrogenase, MEA: multi electrode array.

Article Snippet: For long-term exposure experiments, a predifferentiated human-induced pluripotent stem cell line (hiPSC, XCell Science, Novato, CA, USA) was propagated in Geltrex (Gibco, Thermo Fisher, Waltham, MA, USA)-coated 6-well plates [ ].

Techniques: Derivative Assay

Journal: Cells

Article Title: Neurodegeneration Induced by Anti-IgLON5 Antibodies Studied in Induced Pluripotent Stem Cell-Derived Human Neurons

doi: 10.3390/cells10040837

Figure Lengend Snippet: Anti-IgLON5 antibodies reduce cell surface IgLON5 clusters, synaptic proteins, and neuronal activity. Indirect immunofluorescent staining of human induced pluripotent stem cell (hiPSC) cultures treated with control IgG or anti-IgLON5 IgG ( A ). Quantification of IgLON5-positive clusters in hiPSC cultures revealed a significant reduction after 21 and 35 days of antibody exposure ( B ) ( n = 9). Immunostaining for the synaptic proteins synaptophysin and PSD95 revealed a decrease in synaptophysin clusters after 21 days of exposure and in both synaptophysin and PSD95 clusters after 35 days of exposure ( C , D ) ( n = 6). Multi electrode array (MEA) analysis of hiPSC cultures showed the expected increase in neuronal activity as they matured, however, the spike rate of neurons treated with anti-IgLON5 IgG was significantly lower than that of untreated neurons ( E ) ( n = 5). Example of a MEA data sample is shown in ( F ) (top). When the signal crosses the threshold (horizontal lines) a spike is registered. Registered spikes from a single electrode in a well with an untreated culture (F, middle) and an anti-IgLON5 IgG-treated culture (F, bottom) during a 15 s period. Statistical analysis: two-tailed unpaired t -test or Two-way ANOVA followed by Sidak’s multiple comparisons test (*: p < 0.05, ***: p < 0.001, ****: p < 0.0001). Scale bar length: 5 μm.

Article Snippet: For long-term exposure experiments, a predifferentiated human-induced pluripotent stem cell line (hiPSC, XCell Science, Novato, CA, USA) was propagated in Geltrex (Gibco, Thermo Fisher, Waltham, MA, USA)-coated 6-well plates [ ].

Techniques: Activity Assay, Staining, Control, Immunostaining, Two Tailed Test

Journal: Cells

Article Title: Neurodegeneration Induced by Anti-IgLON5 Antibodies Studied in Induced Pluripotent Stem Cell-Derived Human Neurons

doi: 10.3390/cells10040837

Figure Lengend Snippet: Anti-IgLON5 antibodies increase cell death in induced pluripotent stem cell (hiPSC)-derived neural cultures. Cultures treated with anti-IgLON5 IgG showed an increase in phosphorylated-tau (p-tau) (T205) expression after 7 and 21 days of exposure ( A – C ) ( n = 7–8). Quantification of nuclei with unhealthy morphology revealed an increased percentage of dying cells after 21 and 35 days of exposure ( D ) ( n = 7–18). A lactate dehydrogenase (LDH)-assay (a marker of necrosis) showed increased LDH release to the culture medium of anti-IgLON5 IgG exposed cultures after 30 days ( E ) ( n = 8). Statistical analysis: two-way ANOVA followed by Sidak’s multiple comparisons test. (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001). Scale bars: 50 μm. β-tub III: β-tubulin III.

Article Snippet: For long-term exposure experiments, a predifferentiated human-induced pluripotent stem cell line (hiPSC, XCell Science, Novato, CA, USA) was propagated in Geltrex (Gibco, Thermo Fisher, Waltham, MA, USA)-coated 6-well plates [ ].

Techniques: Derivative Assay, Expressing, Lactate Dehydrogenase Assay, Marker