Application of light sheet microscopy for qualitative and quantitative analysis of bronchus-associated lymphoid tissue in mice

Abstract

Bronchus-associated lymphoid tissue (BALT) develops at unpredictable locations around lung bronchi following pulmonary inflammation. The formation and composition of BALT have primarily been investigated by immunohistology that, due to the size of the invested organ, is usually restricted to a limited number of histological sections. To assess the entire BALT of the lung, other approaches are urgently needed. Here, we introduce a novel light sheet microscopy-based approach for assessing lymphoid tissue in the lung. Using antibody staining of whole lung lobes and optical clearing by organic solvents, we present a method that allows in-depth visualization of the entire bronchial tree, the lymphatic vasculature and the immune cell composition of the induced BALT. Furthermore, three-dimensional analysis of the entire lung allows the qualitative and quantitative enumeration of the induced BALT. Using this approach, we show that a single intranasal application of the replication-deficient poxvirus MVA induces BALT that constitutes up to 8% of the entire lung volume in mice deficient in CCR7, in contrast to wild type mice (WT). Furthermore, BALT induced by heat-inactivated E. coli is dominated by a pronounced T cell infiltration in Cxcr5-deficient mice, in contrast to WT mice.

The chemokine receptor CCR7 is a promising target for rheumatoid arthritis therapy

“…This finding was not expected since different groups have reported delayed but exacerbated immune responses in Ccr7 −/− and plt/plt mice using cutaneous contact hypersensitivity (CHS) models and spontaneous formation of ectopic lymphoid tissue in Ccr7 −/− mice. 13,14,21,22 These aberrant responses were attributed to impaired LN migration and function of regulatory T cells (Tregs). In the CIA model of the present study, we did not observe such reactions in Ccr7 −/− mice, even at late time points, and anti-CII Ab titers were strongly reduced compared to WT mice.…”

Quantitative whole-brain 3D imaging of tyrosine hydroxylase-labeled neuron architecture in the mouse MPTP model of Parkinson's disease

“…Among the various techniques available, immunolabeling-enabled three-dimensional imaging of solvent-cleared organs (iDISCO) has emerged as a simple and rapid method for achieving good antibody penetration and tissue clearing (Liebmann et al, 2016; Renier et al, 2014; Renier et al, 2016). iDISCO–LSFM is particularly useful for CNS research as the low brain anatomical variation between individual mice allows scanned brains to be mapped onto a common coordinated framework (CCF) for subsequent unbiased quantitative image analysis of immune-labeled molecular targets (Detrez et al, 2019; Hasegawa et al, 2019; Henning et al, 2019; Kjaergaard et al, 2019; Liebmann et al, 2016; Merz et al, 2019; Mzinza et al, 2018; Renier et al, 2016).…”

Tissue clearing technique: Recent progress and biomedical applications

“…Mzinza et al introduced 3DISCO/iDISCO and LSFM to assess bronchus‐associated lymphoid tissue (BALT) in mice. They demonstrated qualitative and quantitative analysis of BALT induced by modified vaccinia virus Ankara (MVA) in whole mouse lung lobes, and characterized the development and organization of heat‐inactivated E. coli‐induced BALT (Mzinza et al ., 2018). Vargas's team combined optimized BABB clearing and large‐scale 3D microscopy to study murine lung fibrosis.…”