Summary of Study ST004287
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR002323. The data can be accessed directly via it's Project DOI: 10.21228/M8C24H This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST004287 |
| Study Title | Lipidomic profiling of PILRAKO xMG in xenotransplant model after mouse microglia(mMg) depletion |
| Study Summary | To investigate the impact of PILRAKO in healthy and disease contexts, we transplanted WT or PILRAKO human microglial progenitors into xenotransplantation-compatible hCSF1 and 5x-hCSF1 littermates. 5x-hCSF1 mice, which harbor mutant human amyloid precursor protein and presenilin-1 transgenes, have previously been shown to develop robust amyloid plaque pathology by 6 months of age. To enable CNS-wide replacement of endogenous mouse microglia with human microglia, WT and PILRAKO lines were modified to carry a CSF1R-inhibitor resistant mutation (CSF1R-G795A)28. At 6.5 months of age, WT or PILRAKO xMG were isolated from the brains of hCSF1 and 5x-hCSF1 mice (n=3-4/group). WT xMG isolated from 5x-hCSF1 mice exhibited elevated levels of two bis(monoacylglycero)phosphate (BMP) lipids, a signature of endo-lysosomal dysfunction in which ablation of PILRA rescued BMP accumulation in xMG (Fig 6K). These findings are consistent with improved lysosomal function upon PILRA KO. |
| Institute | Denali Therapeutics |
| Last Name | Suh |
| First Name | Jung |
| Address | 161 Oyster Point Blvd, South San Francisco, CA 94080 |
| suh@dnli.com | |
| Phone | +1 06507973837 |
| Submit Date | 2025-09-22 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML |
| Analysis Type Detail | LC-MS |
| Release Date | 2025-10-16 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002323 |
| Project DOI: | doi: 10.21228/M8C24H |
| Project Title: | PILRA regulates microglial neuroinflammation and lipid metabolism as a candidate therapeutic target for Alzheimer’s disease |
| Project Type: | Cellular studies |
| Project Summary: | The Alzheimer’s disease (AD) human genetic landscape identified microglia as a key disease-modifying brain cell type. A common loss-of-function coding variant in paired immunoglobulin-like type 2 receptor alpha (PILRA) associated with reduced AD risk, was found enriched in a cohort of healthy centenarians and rescued APOE4 AD risk, however the mechanisms underlying protection are undefined. Here we identify biological functions of PILRA, an immunoreceptor tyrosine-based inhibitory motif (ITIM)-containing receptor, in human iPSC-derived microglia (iMG) and chimeric AD mice. CRISPR-mediated PILRA knockout (KO) in iMG rescued ApoE4-induced immunometabolic deficits. Moreover, loss of PILRA confers a signature of metabolic resilience in microglia with increased mitochondrial capacity in tandem with elevated antioxidants, reduced ROS and toxic lipid species. Additionally, PILRA KO iMG exhibit improved lysosomal degradation, enhanced migration, and attenuated cytokine responses. We show PPAR and STAT1/3 act as master regulators that mediate downstream signaling and regulate PILRA-dependent microglial functions. Importantly, AD mice transplanted with human PILRA KO microglia showed reduced amyloid pathology and rescued levels of synaptic markers. Finally, we identify a high-affinity human PILRA-specific antagonist antibody that phenocopies PILRA KO. Together, these findings suggest a therapeutic approach to modulate microglial immunometabolism by inhibiting PILRA, thus identifying a pharmacologically tractable target for AD. |
| Institute: | Denali Therapeutics |
| Last Name: | Suh |
| First Name: | Jung |
| Address: | 161 Oyster Point Blvd, South San Francisco, CA 94080 |
| Email: | suh@dnli.com |
| Phone: | +1 6507973837 |
Subject:
| Subject ID: | SU004440 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | cell_line | disease_state | genotype | Sample source |
|---|---|---|---|---|---|
| SA498504 | HSA-000037564 | NA | NA | NA | microglia |
| SA498505 | HSA-000037562 | NA | NA | NA | microglia |
| SA498506 | HSA-000037563 | NA | NA | NA | microglia |
| SA498507 | HSA-000037091 | PILRAKO | 5xFAD | 5x-hCSF1 | microglia |
| SA498508 | HSA-000037093 | PILRAKO | 5xFAD | 5x-hCSF1 | microglia |
| SA498509 | HSA-000037092 | PILRAKO | 5xFAD | 5x-hCSF1 | microglia |
| SA498510 | HSA-000037089 | PILRAKO | WT | hCSF1 | microglia |
| SA498511 | HSA-000037088 | PILRAKO | WT | hCSF1 | microglia |
| SA498512 | HSA-000037087 | PILRAKO | WT | hCSF1 | microglia |
| SA498513 | HSA-000037090 | PILRAKO | WT | hCSF1 | microglia |
| SA498514 | HSA-000037086 | WT | 5xFAD | 5x-hCSF1 | microglia |
| SA498515 | HSA-000037085 | WT | 5xFAD | 5x-hCSF1 | microglia |
| SA498516 | HSA-000037083 | WT | 5xFAD | 5x-hCSF1 | microglia |
| SA498517 | HSA-000037084 | WT | 5xFAD | 5x-hCSF1 | microglia |
| SA498518 | HSA-000037081 | WT | WT | hCSF1 | microglia |
| SA498519 | HSA-000037080 | WT | WT | hCSF1 | microglia |
| SA498520 | HSA-000037082 | WT | WT | hCSF1 | microglia |
| SA498521 | HSA-000037079 | WT | WT | hCSF1 | microglia |
| Showing results 1 to 18 of 18 |
Collection:
| Collection ID: | CO004433 |
| Collection Summary: | Six and half months after HPC transplantation, mice were anesthetized and intracardially perfused with 1X DPBS, half brains were dissected, the cerebellum was removed, and tissue was stored in RPMI 1640 until subsequent perfusions were completed. All steps were performed on ice or at 4°C with ice-cold reagents and all centrifuge steps were performed for 10 minutes at 400xg with full brake and acceleration unless otherwise stated. Brains were manually homogenized using a 7 mL Dounce homogenizer by adding 4 mL of RPMI 1640 and performing 10 strokes with the “loose” pestle followed by 10 strokes with the “tight” pestle. Samples were then run through a pre-soaked 70μM filter then the filter was washed with 10mL of RPMI 1640. The sample was pelleted by centrifugation and myelin was removed by resuspension in 8 L of 30% Percoll overlaid with 2 mL of 1X DPBS centrifuged at 400xg for 20 minutes with acceleration set to 0 and brake set to 4. The myelin band and supernatant were discarded, and cell pellets were resuspended in 70 μL MACS buffer (0.5% bovine serum albumin in 1X DPBS) + 30 μL Mouse Cell Removal beads (Miltenyi) and incubated at 4°C for 15 minutes. Magnetically labeled mouse cells were separated using LS columns and the MidiMACs separator (Miltenyi) while the unlabeled human cells were collected in the flow through. Human microglia cells were pelleted, and washed 1x with 0.9% sodium chloride and collected in Lobind 1.5 mL Eppendorf tubes and snap frozen. |
| Sample Type: | Microglia |
| Storage Conditions: | -80℃ |
| Collection Vials: | Lobind 1.5 mL Eppendorf tubes |
| Storage Vials: | Lobind 1.5 mL Eppendorf tubes |
Treatment:
| Treatment ID: | TR004449 |
| Treatment Summary: | No treatment |
Sample Preparation:
| Sampleprep ID: | SP004446 |
| Sampleprep Summary: | On the day of lipid and metabolite extraction, cell pellets were reconstituted with 400 μL MS grade methanol spiked with internal standards.Samples were vortexed and volumes were adjusted to 800 μL with MS grade H2O, vortexed again, then 600 μL tert-butyl methyl ether (MTBE) was added, vortexed, then centrifuged at 21,000 g for 10 min at 4C. The two phases generated by centrifugation were separated. Each phase (top: non-polar lipids, bottom: polar metabolites) was transferred to glass vials and dried overnight using a Genevac EZ3. Non-polar lipids were resuspended MS grade methanol and polar metabolites were resuspended in 90% methanol:water mixture. |
| Processing Storage Conditions: | On ice |
| Extract Storage: | -20℃ |
Chromatography:
| Chromatography ID: | CH005416 |
| Instrument Name: | Agilent 1290 Infinity II |
| Column Name: | Waters ACQUITY UPLC BEH C18 (100 x 2.1mm,1.7um) |
| Column Temperature: | 55°C |
| Flow Gradient: | 0.0-8.0 min from 45% B to 99% B, 8.0-9.0 min at 99% B, 9.0-9.1 min to 45% B, and 9.1-10.0 min at 45% B. |
| Flow Rate: | 0.25 mL/min |
| Solvent A: | 60% acetonitrile/40% water; 10 mM ammonium formate; 0.1% formic acid |
| Solvent B: | 90% isopropyl alcohol/10% acetonitrile; 10 mM ammonium formate; 0.1% formic acid |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH005417 |
| Instrument Name: | Agilent 1290 Infinity II |
| Column Name: | Waters ACQUITY UPLC BEH C18 (100 x 2.1mm,1.7um) |
| Column Temperature: | 55°C |
| Flow Gradient: | 0.0-8.0 min from 45% B to 99% B, 8.0-9.0 min at 99% B, 9.0-9.1 min to 45% B, and 9.1-10.0 min at 45% B. |
| Flow Rate: | 0.25 mL/min |
| Solvent A: | 60% acetonitrile/40% water; 10 mM ammonium acetate; 0.1% acetic acid |
| Solvent B: | 90% isopropyl alcohol/10% acetonitrile; 10 mM ammonium acetate; 0.1% acetic acid |
| Chromatography Type: | Reversed phase |
Analysis:
| Analysis ID: | AN007127 |
| Analysis Type: | MS |
| Chromatography ID: | CH005416 |
| Num Factors: | 5 |
| Num Metabolites: | 140 |
| Units: | normalized peak area |
| Analysis ID: | AN007128 |
| Analysis Type: | MS |
| Chromatography ID: | CH005417 |
| Num Factors: | 5 |
| Num Metabolites: | 119 |
| Units: | normalized peak area |
