Minimizing the <i>ex vivo</i> confounds of cell-isolation techniques on transcriptomic -profiles of purified microglia

Ocañas, Sarah R.; Pham, Kevin D.; Blankenship, Harris E.; Machalinski, Adeline H.; Chucair-Elliott, Ana J.; Freeman, Willard M.

doi:10.1101/2021.07.15.452509

Cited by 7 publications

(1 citation statement)

References 55 publications

(74 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Tissue samples were immediately stored on ice, with all subsequent procedures performed at 4 ο C. Dissected frontal cortex was passed through a 70 µm cell strainer. Both mechanical tissue homogenization and reduced temperatures (4 ο C) have been shown to limit isolation-associated shifts in microglial gene expression (Ocañas et al, 2021). Homogenates were centrifuged at 800g for 8 min.…”

Section: Methodsmentioning

confidence: 99%

Repeated Activation of Pyramidal Neurons in the Prefrontal Cortex Alters Microglial Phenotype in Male Mice

Bollinger,

Horchar,

Wohleb

2023

J Pharmacol Exp Ther

View full text Add to dashboard Cite

Microglia are highly attuned to fluctuations in neuronal activity. Here, we show that repeated activation of pyramidal neurons in the prefrontal cortex induces broad changes in microglia phenotype; this includes upregulation of pathways associated with microglial proliferation, microglia-neuron interactions, and lysosome induction. Our findings suggest that studies using chemogenetic or optogenetic approaches to manipulate neural circuits should be mindful of indirect effects on non-neuronal cells and their potential contribution to measured outcomes.

show abstract

Section: Methodsmentioning

confidence: 99%

Repeated Activation of Pyramidal Neurons in the Prefrontal Cortex Alters Microglial Phenotype in Male Mice

Bollinger,

Horchar,

Wohleb

2023

J Pharmacol Exp Ther

View full text Add to dashboard Cite

show abstract

Blocking necroptosis reduces inflammation and tumor incidence in a mouse model of diet-induced hepatocellular carcinoma

Mohammed¹,

Thadathil²,

Tran³

et al. 2022

Preprint

View full text Add to dashboard Cite

Background & Aims: Nonalcoholic fatty liver disease (NAFLD) is one of the etiologies that contribute to hepatocellular carcinoma (HCC), and chronic inflammation is one of the proposed mediators of HCC. As necroptosis is a cell death pathway that induces inflammation, we tested whether necroptosis-induced inflammation contributes to the progression of NAFLD to HCC in a mouse model of diet-induced HCC. Methods: Male and female wild-type (WT) mice or mouse models where necroptosis is blocked (Ripk3-/- or Mlkl-/- mice) were fed a control diet or choline-deficient low fat diet (CD-LFD) or CD-high fat diet (CD-HFD) for 6 months. Changes in inflammation, immune cell infiltration, activation of oncogenic pathways, and tumor incidence were assessed by gene expression analysis, western blotting, and flow cytometry. RNA sequencing (RNA-seq) was performed to assess the changes in liver transcriptome. Results: Blocking necroptosis by deleting either Ripk3 or Mlkl reduced markers of inflammation [proinflammatory cytokines (TNFα, IL-6, and IL-1β), F4/80+ve macrophages, CCR2+ve infiltrating monocytes], inflammation associated oncogenic pathways (JNK, PD-L1/PD-1, β-catenin), and HCC in male mice. In female mice, blocking necroptosis reduced HCC independent of inflammation. Blocking necroptosis reduced cell senescence markers in males and females, suggesting a novel cross-talk between necroptosis and cell senescence. Conclusions: Our data show that hepatic necroptosis promotes recruitment and activation of liver macrophages leading to chronic inflammation, which in turn trigger oncogenic pathways leading to the progression of NAFLD to HCC in male mice. In female mice necroptosis contributes to HCC independent of inflammation. Thus, our study suggests that necroptosis is a valid target for NAFLD-mediated HCC.

show abstract

Microglial MHC-I induction with aging and Alzheimer’s is conserved in mouse models and humans

Kellogg

Pham

Machalinski

et al. 2023

Preprint

Self Cite

View full text Add to dashboard Cite

Major Histocompatibility Complex I (MHC-I) function in the CNS is still being determined after previously being thought to be absent from the brain. MHC-I expression increases with brain aging in mouse, rat, and human whole tissue analyses. Neuronal MHC-I expression has been proposed to regulate developmental synapse elimination and tau pathology in Alzheimer's disease (AD). However, the CNS cellular localization of MHC-I expression has been unclear. Across newly generated and publicly available ribosomal profiling, cell sorting, and single cell data, microglia were found to be the primary source of classical and non-classical MHC-I in mice and humans. TRAP-qPCR analysis of 3-6 m.o. and 18-22 m.o. mice revealed significant age-related induction of B2m, H2-D1, H2-K1, H2-M3, H2-Q6, and Tap1 in microglia but not in astrocytes and neurons. Across a timecourse from 12-23 m.o., microglial MHC-I gradually increases until 21 m.o. and then accelerates. MHC-I protein was also enriched in microglia and increased with aging. Expression of MHC-I binding Leukocyte Immunoglobulin-like (Lilr) and Paired immunoglobin-like type 2 (Pilr) receptors in microglia but not astrocytes or neurons opens the possibility of cell-autonomous signaling and are also increased with aging in mice and humans. Increased microglial MHC-I, Lilrs, and Pilrs were observed in mouse AD models and human data across numerous studies and in RNA-Seq of microglia from APP-PSEN1 mice. MHC-I expression occurred concurrently with p16 suggesting an association with cellular senescence. The conserved induction of MHC-I, Lilrs, and Pilrs with aging and AD open the possibility of cell-autonomous signaling to regulate microglial reactivation.

show abstract

Minimizing the ex vivo confounds of cell-isolation techniques on transcriptomic -profiles of purified microglia

Cited by 7 publications

References 55 publications

Repeated Activation of Pyramidal Neurons in the Prefrontal Cortex Alters Microglial Phenotype in Male Mice

Repeated Activation of Pyramidal Neurons in the Prefrontal Cortex Alters Microglial Phenotype in Male Mice

Blocking necroptosis reduces inflammation and tumor incidence in a mouse model of diet-induced hepatocellular carcinoma

Microglial MHC-I induction with aging and Alzheimer’s is conserved in mouse models and humans

Contact Info

Product

Resources

About