Microglia and neuroinflammation play an important role in the development and progression of Alzheimer's disease (AD). Inositol polyphosphate-5-phosphatase D (INPP5D/SHIP1) is a myeloid-expressed ...gene genetically-associated with AD. Through unbiased analyses of RNA and protein profiles in INPP5D-disrupted iPSC-derived human microglia, we find that reduction in INPP5D activity is associated with molecular profiles consistent with disrupted autophagy and inflammasome activation. These findings are validated through targeted pharmacological experiments which demonstrate that reduced INPP5D activity induces the formation of the NLRP3 inflammasome, cleavage of CASP1, and secretion of IL-1β and IL-18. Further, in-depth analyses of human brain tissue across hundreds of individuals using a multi-analytic approach provides evidence that a reduction in function of INPP5D in microglia results in inflammasome activation in AD. These findings provide insights into the molecular mechanisms underlying microglia-mediated processes in AD and highlight the inflammasome as a potential therapeutic target for modulating INPP5D-mediated vulnerability to AD.
SORL1 is implicated in the pathogenesis of Alzheimer’s disease (AD) through genetic studies. To interrogate the roles of SORL1 in human brain cells, SORL1-null induced pluripotent stem cells (iPSCs) ...were differentiated to neuron, astrocyte, microglial, and endothelial cell fates. Loss of SORL1 leads to alterations in both overlapping and distinct pathways across cell types, with the greatest effects in neurons and astrocytes. SORL1 loss induces a neuron-specific reduction in apolipoprotein E (APOE) and clusterin (CLU) and altered lipid profiles. Analyses of iPSCs derived from a large cohort reveal a neuron-specific association between SORL1, APOE, and CLU levels, a finding validated in postmortem brain. Enhancement of retromer-mediated trafficking rescues tau phenotypes observed in SORL1-null neurons but does not rescue APOE levels. Pathway analyses implicate transforming growth factor β (TGF-β)/SMAD signaling in SORL1 function, and modulating SMAD signaling in neurons alters APOE RNA levels in a SORL1-dependent manner. Taken together, these data provide a mechanistic link between strong genetic risk factors for AD.
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•Loss of SORL1 induces a reduction of APOE and CLU levels in neurons•APOE levels and CLU levels are associated with SORL1 levels in human brain in neurons•Modulation of SMAD signaling regulates APOE levels in neurons in a SORL1-dependent manner•SORL1 KO and SORL1 G511R neurons display altered lipid profiles
Lee et al. use SORL1-null iPSCs to identify genes and pathways altered by loss of SORL1 across brain cell types. These studies reveal that SORL1 has a neuron-specific role in regulating APOE and CLU levels and implicate TGF-β-mediated SMAD signaling in this process.
Trisomy 21 (T21) causes Down syndrome and an early-onset form of Alzheimer's disease (AD). Here, we used human induced pluripotent stem cells (hiPSCs) along with CRISPR-Cas9 gene editing to ...investigate the contribution of chromosome 21 candidate genes to AD-relevant neuronal phenotypes. We utilized a direct neuronal differentiation protocol to bypass neurodevelopmental cell fate phenotypes caused by T21 followed by unbiased proteomics and western blotting to define the proteins dysregulated in T21 postmitotic neurons. We show that normalization of copy number of APP and DYRK1A each rescue elevated tau phosphorylation in T21 neurons, while reductions of RCAN1 and SYNJ1 do not. To determine the T21 alterations relevant to early-onset AD, we identified common pathways altered in familial Alzheimer's disease neurons and determined which of these were rescued by normalization of APP and DYRK1A copy number in T21 neurons. These studies identified disruptions in T21 neurons in both the axonal cytoskeletal network and presynaptic proteins that play critical roles in axonal transport and synaptic vesicle cycling. These alterations in the proteomic profiles have functional consequences: fAD and T21 neurons exhibit dysregulated axonal trafficking and T21 neurons display enhanced synaptic vesicle release. Taken together, our findings provide insights into the initial molecular alterations within neurons that ultimately lead to synaptic loss and axonal degeneration in Down syndrome and early-onset AD.
Background
SORL1 is a neuronal sorting receptor that has been strongly implicated in the pathogenesis of Alzheimer’s disease (AD). While SORL1 has structural similarities to low‐density lipoprotein ...(LDL) receptors, it is unclear if it plays a role in regulating lipoprotein levels in the brain. We aimed to utilize human induced pluripotent stem cells (iPSCs) to understand the intersection of SORL1 and APOE biology and interrogate the potential role of SORL1 as an LDL receptor.
Method
We differentiated iPSC lines derived from the ROS and MAP cohorts that span the cognitive and neuropathological spectrum of aging to neuron and astrocyte fates and performed unbiased proteomic profiling, Western blotting, and ELISA to quantify protein levels of SORL1 and APOE. We next used CRISPR/Cas9 to generate SORL1 null iPSC lines and differentiated to neuron, astrocyte, and microglial fates. In neurons and astrocytes, AD‐relevant phenotypes such as APOE, Abeta, and phospho‐tau were assessed. Rescue experiments were performed in SORL1 KO neurons using pharmacological perturbations and shRNA to elucidate the pathways downstream of loss of SORL1 that lead to altered Abeta, tau and APOE.
Result
We observed a strong correlation between SORL1 and APOE protein expression in neurons derived from ROSMAP cohort individuals that was not apparent in astrocytes. Next, we validated previously reported findings that loss of SORL1 in neurons results in an elevation in Abeta levels. We expanded upon these findings to show that SORL1 null neurons display elevated phospho‐tau levels and a significant reduction of extracellular and intracellular APOE. While enhancing retromer and autophagy function rescued the elevation of phosphorylated tau in SORL1 null neurons, these treatments did not rescue reduced APOE levels. Rather, loss of SORL1 resulted in a reduction in APOE RNA expression, suggesting that loss of SORL1 leads to a downregulation of APOE at the transcriptional level. Integration of profiling data from ROSMAP iNs and SORL1 knock out iNs implicate the transcription factors NFkB and REV‐ERB as potentially mediating the effect of loss of SORL1 on APOE RNA levels.
Conclusion
SORL1 has a neuron‐specific function in regulating APOE levels that is separable from its role in regulating Abeta and phospho‐tau levels.
Background
Herpes simplex virus 1 (HSV‐1) has been associated with pathogenesis of Alzheimer’s Disease (AD) through several landmark studies. We explored the transcriptomics landscape of HSV‐1 ...infection and anti‐viral treatment using acyclovir on AD‐associated genes using large‐scale bulk RNA sequencing to identify biomarkers, cell types and cell type specific interactions.
Method
We used recent technological advances in human 3D brain organoids and transcriptomics sequencing tools.
Result
We found that HSV‐1 infection in cerebral organoids from a single individual resulted in transcriptomic perturbations were enriched for AD‐associated genes. ACV treatment restored HSV‐1 induced host transcript perturbations in cerebral organoids for most genes in a dosage‐dependent manner, demonstrating that immediate anti‐viral treatment of HSV‐1 infection is effective in our human cellular model. However, we also observed off‐target transcriptomic effects from ACV treatment that were enriched for AD‐associated genes and genes associated with autoimmune diseases.
HSV‐1 infection in cerebral organoids also resulted in significantly decreased Abeta42/40 and Abeta42/38 ratios detected from the supernatant, compared to uninfected control cerebral organoids. HSV‐1 infection followed by immediate ACV treatment rescued the Abeta42/40 ratios detected from the supernatant, but did not rescue the Abeta42/38 ratios. We found that influenza A (IAV) infection did not result in an enrichment of AD‐associated transcriptomic perturbations and both Abeta42/40 and Abeta42/38 ratios were not significantly changed in the supernatant between IAV‐infected cerebral organoids and uninfected cerebral organoids.
We explored cell type specific effects of HSV‐1 infection and ACV treatment in cerebral organoids through cell type enrichment analyses of the bulk RNA sequence data and flow cytometry experiments. We found complex cell type interactions perturbed by HSV‐1 infection with varying degrees of rescue by ACV, such as increased proportions of reactive astrocytes and decreased proportions of live neurons.
Finally, we correlated the transcriptomic signature of HSV‐1 infected cerebral organoids with transcriptomes of post‐mortem brain samples from LOAD patients and discovered a high correlation with subtype A (31% of patients).
Conclusion
Our work is an unbiased study to identify biomarkers and novel therapeutic pathways, as well as potential off‐target effects associated with HSV‐1 infection and ACV treatment in AD.
