Chuang, Y.-H. | bioRxiv Science

bioRxiv2024

Expansion microscopy for super-resolution imaging of collagen-abundant tissues

Expansion microscopy (ExM) is popular for three-dimensional ultrastructural imaging of cultured cells and tissue slices at nanoscale resolution with conventional microscopes via physical expansion of biological tissues. However, the application of this technology to collagen-abundant thick tissues is challenging. We demonstrate a new method, collagen expansion microscopy (ColExM), optimized for expanding tissues containing more than 70% collagen. ColExM succeeded in 4.5-fold linear expansion with minimal structural distortion of corneal and skin tissues. It was also compatible with immunostaining, allowing super-resolution visualization of three-dimensional neural structures innervating hair follicles and corneas. With ColExM, we succeeded in identifying individual mitochondria and previously unrecognized dendritic spine-like structures of corneal nerves. ColExM also enabled fine mapping of structural rearrangement of tight junctions and actin cytoskeletons. Therefore, this method can facilitate the exploration of three-dimensional nanoscale structures in collagen-rich tissues.

bioengineering↗

bioRxiv2022

HLJ1 amplifies endotoxin-induced sepsis severity by promoting IL-12 heterodimerization in macrophages

Heat shock protein (HSP) 40 has emerged as a key actor in both innate and adaptive immunity, whereas the role of HLJ1, a molecular chaperone in HSP40 family, in modulating endotoxin-induced sepsis severity is still unclear. Here, we use single-cell RNA sequencing to characterize mouse liver nonparenchymal cell populations under LPS (lipopolysaccharide) stimulation, and show that HLJ1 deletion affected IFN-{gamma}-related gene signatures in distinct immune cell clusters. HLJ1 deficiency also leads to reduced serum levels of IL-12 in LPS-treated mice, contributing to dampened production of IFN-{gamma} in natural killer cells but not CD4+ or CD8+ T cells, and subsequently to improved survival rate. Adoptive transfer of HLJ1-deleted macrophages into LPS-treated mice results in reduced IL-12 and IFN-{gamma} levels and protects the mice from IFN-{gamma}-dependent mortality. In the context of molecular mechanisms, HLJ1 is an LPS-inducible protein in macrophages and converts misfolded IL-12p35 homodimers to monomers, which maintains bioactive IL-12p70 heterodimerization and secretion. This study suggests HLJ1 causes IFN-{gamma}-dependent septic lethality by promoting IL-12 heterodimerization, and targeting HLJ1 has therapeutic potential in inflammatory diseases involving activating IL-12/IFN-{gamma} axis.

immunology↗