Rita Kandel MD

Pathology and Laboratory Medicine,

Sinai Health System,

University of Toronto,

Toronto ,ON Canada

HISTOLOGY OF CARTILAGE

REPAIR

AMERICAN SOCIETY FOR TESTING

AND MATERIALS (ASTM) GUIDELINES

- Gross evaluation - Histologic evaluation (validated scoring system)

- Biochemical evaluation (quantity and quality of collagen and proteoglycans)

- Biomechanical assessment

- Appropriate statistical analyses

GROSS ASSESSMENT

ICRS grading system

HISTOLOGICAL ANALYSIS OF

REPAIR

Implant

1) Conventional histology

Arzi B et al., Acta Biomater 2015; 1 (23):72-81

Synovium

HISTOLOGICAL APPEARANCE

OF JOINT SURFACE

Hyaline (non-

calcified)

Cartilage

Bone

Calcified

cartilage

SAMPLE PROCESSING FOR HISTOLOGY

1) EXCISE DEFECT

2) FIXATION: 10% buffered formalin duration dependent on tissue size

3) DECALCIFICATION: 0.5M EDTA duration dependent on tissue size at room temperature

4) EMBEDDING: paraffin vs plastic vs frozen section

5) SAMPLING OF DEFECT: standardize location

HISTOLOGICAL EVALUATION OF

REPAIR: tissue quality

Kandel RA et al. Biomaterials 2006

H&E

Toluidine blue

CPP

In vitro formed cartilage In vivo cartilage

Bone

1mm

Safranin O

EVALUATION OF REPAIR: interfaces

Hoemann C et al. Cartilage 2011

Safranin O

Native

Repair

200um

100um

Interface

LATERAL DEEP

one

one

Toluidine blue

1mm

Kandel RA et al. Biomaterials 2006

CLASSIFICATION SYSTEMS

Orth P et al Tiss Engineering Part C, 2012

100 μm

Hyaline cartilage

Calcified cartilage

ARTICULAR CARTILAGE: depth dependent

zonal architecture

10 -

20%

40 -

60%

30 -

40%

Co

llagen

Adapted from Klein TJ. et al., 2009

PRG4

Proteoglycan

Alkaline phosphatase

ALKALINE PHOSPHATASE

Bone

X X Xx

Hoemann C et al., Cartilage 2011

X X X x

HISTOLOGICAL ANALYSIS OF

REPAIR

1) Histology

2) Immunohistochemical staining

aggrecan

type II collagen type I collagen PRG4

PRG4

HISTOLOGICAL ANALYSIS OF

REPAIR

1) Histology 2) Immunohistochemical staining 3) Polarized light microscopy

REPAIR NORMAL

SAMPLING PITFALLS

Hoemann C et al. Cartilage 2011; 2:153 -172

Orth P et al. Progress in Histochemistry and Cytochemistry 2015

REPAIR

SAMPLING PITFALLS

HISTOLOGY PITFALLS

1) Fixation duration

2) Method of decalcification

3) Grading expertise

4) Poor quality images

European Cells and Materials 2016; 31:79-94

DOES HISTOLOGY PREDICT

FUNCTION?

HISTOLOGICAL APPEARANCE OF JOINT

SURFACE

Hyaline

Cartilage

Calcified

cartilage

Bone

Cells

Matrix

Normal

HISTOLOGICAL APPEARANCE autologous chondrocyte implantation at 1 year

Safranin O stain

Ref. Sancho-Tello M et al., Int J Artif Organs 2015; 38(4):210-223

Ref. Sancho-Tello M et al., Int J Artif Organs 2015; 38(4):210-223