Multi-modal Single-Cell Analysis Reveals Brain Immune Landscape Plasticity during Aging and Gut Microbiota Dysbiosis

Isolation of mouse brain-infiltrating leukocytes for single cell profiling of epitopes and transcriptomes

“…This protocol has been demonstrated to reliably dissociate the murine brain, whether homeostatic, aged, or tumor-burdened, into a single cell suspension, and highly enrich brain-infiltrating leukocytes. We have subsequently performed CyTOF and CITE-seq on leukocytes prepared based on the protocol described herein, achieving high quality results, as shown in the figures of this protocol, and our previous publications ( Golomb et al., 2020 ; Guldner et al., 2020 ).…”

“…This cell yield and viability meet or exceed the minimum requirements of applications such as CyTOF and CITE-seq. Ultimately, leukocyte preparations derived from following this protocol will lead to exquisite single cell profiling in applications such as CITE-seq/REAP-seq and CyTOF, as demonstrated in our recent publications( Golomb et al., 2020 ; Guldner et al., 2020 ). Critically, we observed our brain dissociation protocol preserved many common immune cells identifying or functional surface protein epitopes, as evidenced by positive staining of >40 cell surface epitopes for which we stained in CITE-seq experiments, a subset of which are shown in Figures 3 A–3C.…”

“…The isolation of mouse brain-infiltrating leukocytes described herein has proven successful in our hands in the contexts of homeostasis, aging, and brain metastasis ( Golomb et al., 2020 ; Guldner et al., 2020 ). Importantly, these represent conditions in which the brain has an abundance of myelin (aging) or large epithelial lesions (brain metastasis) that can present challenges to brain dissociation and the generation of single cell suspensions.…”

“…For complete details on the use and execution of this protocol, please refer to Guldner et al. (2020) and Golomb et al. (2020) .…”

The Microbiota-Gut-Brain Axis in Health and Disease and Its Implications for Translational Research

“…Further, GF mice differ in dendrite morphology and show increased volume of the amygdala and Requirement of adult-born neurons for hippocampus-dependent learning. Ogbonnaya et al, 2015 Adult hippocampal neurogenesis is regulated by the microbiome Luczynski et al, 2016 Adult microbiota-deficient mice have distinct dendritic morphological changes: differential effects in the amygdala and hippocampus Möhle et al, 2016 Ly6C(hi) monocytes provide a link between antibiotic-induced changes in gut microbiota and adult hippocampal neurogenesis Swann et al, 2017 Application of 1 H NMR spectroscopy to the metabolic phenotyping of rodent brain extracts: A metabonomic study of gut microbial influence on host brain metabolism Fitzpatrick et al, 2020 Gut-educated IgA plasma cells defend the meningeal venous sinuses Myelination Keogh et al, 2021 Myelin as a regulator of development of the microbiota-gut-brain axis BBB Braniste et al, 2014 The gut microbiota influences blood-brain barrier permeability in mice Microglia maturation Erny et al, 2015 Host microbiota constantly control maturation and function of microglia in the CNS Quigley, 2017 Microbiota-brain-gut axis and neurodegenerative diseases Aging Golomb et al, 2020 Multi-modal single-cell analysis reveals brain immune landscape plasticity during aging and gut microbiota dysbiosis Li et al, 2020 Age-related shifts in gut microbiota contribute to cognitive decline in aged rats Madison and Kiecolt-Glaser, 2021 The gut microbiota and nervous system: Age-defined and age-defying the hippocampus (Luczynski et al, 2016). Moreover, decreased neurogenesis in the hippocampus of adult mice was induced after long-term antibiotic treatment leading to deficits in the novel object recognition task.…”