title: Optical clearing in dense connective tissues to visualize cellular connectivity in situ.
abstract: Visualizing the three-dimensional morphology and spatial patterning of cells embedded deep within dense connective tissues of the musculoskeletal system has been possible only by utilizing destructive techniques. Here we utilize fructose-based clearing solutions to image cell connectivity and deep tissue-scale patterning in situ by standard confocal microscopy. Optical clearing takes advantage of refractive index matching of tissue and the embedding medium to visualize light transmission through a broad range of bovine and whole mount murine tissues, including cartilage, bone, and ligament, of the head and hindlimb. Using non-destructive methods, we show for the first time intercellular chondrocyte connections throughout the bulk of cartilage, and we reveal in situ patterns of osteocyte processes and the lacunar-canalicular system deep within mineralized cortical bone. Optical clearing of connective tissues is expected to find broad application for the study of cell responses in normal physiology and disease pathology.
authors.affiliation: Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana, United States of America.
authors.forename: Sarah
authors.initials: S
authors.lastname: Calve
authors.affiliation: Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana, United States of America.
authors.forename: Andrew
authors.initials: A
authors.lastname: Ready
authors.affiliation: W. Nuhsbaum, Inc., McHenry, Illinois, United States of America.
authors.forename: Christopher
authors.initials: C
authors.lastname: Huppenbauer
authors.affiliation: Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana, United States of America; Department of Basic Medical Sciences, Purdue University, West Lafayette, Indiana, United States of America.
authors.forename: Russell
authors.initials: R
authors.lastname: Main
authors.affiliation: Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana, United States of America.
authors.forename: Corey P
authors.initials: CP
authors.lastname: Neu
mesh_terms: D000818:Animals; D001842:Bone and Bones / Q000166:cytology*; D002356:Cartilage / Q000166:cytology*; D002417:Cattle; D019902:Chondrocytes / Q000166:cytology; D006257:Head / Q000502:physiology; D006614:Hindlimb / Q000166:cytology; D006652:Histological Techniques / Q000379:methods; D008022:Ligaments / Q000166:cytology*; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods; D010011:Osteocytes / Q000166:cytology; D012996:Solutions / Q000737:chemistry
chemical_list: D012996:Solutions