title: Nonlinear optical microscopy of articular cartilage.
abstract: OBJECTIVE
  To assess the ability of nonlinear optical microscopy (NLOM) to image ex vivo healthy and degenerative bovine articular cartilage.
  
  
  METHODS
  Fresh bovine femoral-tibial joints were obtained from an abattoir. Articular cartilage specimens were harvested from the tibial plateau. Normal and degenerative specimens were imaged by NLOM and subsequently fixed and processed for histological examination.
  
  
  RESULTS
  NLOM provided high resolution images of articular cartilage at varying depths with high sensitivity to tissue morphology and high specificity to tissue components without fixing, sectioning or staining. Spectroscopic segmentation of nonlinear optical signals isolated the collagen matrix from the chondron (chondrocyte and non-collagen pericellular matrix). Images from the superficial zone were consistent with the presence of a matrix composed of both elastin-like and collagen fibers distributed in a depth-dependent morphological arrangement, whereas only collagen was demonstrated in the middle and deep zones. Alterations of collagen matrix associated with advanced degenerative joint disease (fibrocartilage) were observed with NLOM. Individual chondrocytes were imaged and demonstrated intracellular fluorescence consistent with the presence of products of intracellular biochemical processes.
  
  
  CONCLUSIONS
  Thin images of living articular cartilage using NLOM may be obtained with (sub-)cellular resolution at varying depths without fixing, sectioning or staining. Extracellular matrical collagen and chondron may be imaged separately in native tissue using spectrally distinct, endogenous, nonlinear optical signals. NLOM was sensitive to macromolecular composition and pathologic changes in articular cartilage matrix. Advances in instrumentation may lead to the application of NLOM to study articular cartilage in vivo.
authors.affiliation: Department of Biomedical Engineering, Texas A&M University, College Station, TX 77843, USA.
authors.forename: Alvin T
authors.initials: AT
authors.lastname: Yeh
authors.forename: Marie J
authors.initials: MJ
authors.lastname: Hammer-Wilson
authors.forename: David C
authors.initials: DC
authors.lastname: Van Sickle
authors.forename: Hilary P
authors.initials: HP
authors.lastname: Benton
authors.forename: Aikaterini
authors.initials: A
authors.lastname: Zoumi
authors.forename: Bruce J
authors.initials: BJ
authors.lastname: Tromberg
authors.forename: George M
authors.initials: GM
authors.lastname: Peavy
mesh_terms: D000818:Animals; D002357:Cartilage Diseases / Q000378:metabolism / Q000473:pathology* / Q000662:veterinary*; D002358:Cartilage, Articular / Q000033:anatomy & histology* / Q000737:chemistry / Q000473:pathology; D002417:Cattle; D002418:Cattle Diseases / Q000378:metabolism / Q000473:pathology*; D003094:Collagen / Q000032:analysis; D018613:Microscopy, Confocal; D011509:Proteoglycans / Q000032:analysis; D013048:Specimen Handling / Q000379:methods
chemical_list: D011509:Proteoglycans; D003094:Collagen