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Application of Atomic Force Microscopy to Detect Early Osteoarthritis
In This Article
Summary
We present a method to investigate early osteoarthritic changes at the cellular level in articular cartilage by using atomic force microscopy (AFM).
Abstract
Biomechanical properties of cells and tissues not only regulate their shape and function but are also crucial for maintaining their vitality. Changes in elasticity can propagate or trigger the onset of major diseases like cancer or osteoarthritis (OA). Atomic force microscopy (AFM) has emerged as a strong tool to qualitatively and quantitatively characterize the biomechanical properties of specific biological target structures on a microscopic scale, measuring forces in a range from as small as the piconewton to the micronewton. Biomechanical properties are of special importance in musculoskeletal tissues, which are subjected to high levels of strain. OA as a degenerative disease of the cartilage results in the disruption of the pericellular matrix (PCM) and the spatial rearrangement of the chondrocytes embedded in their extracellular matrix (ECM). Disruption in PCM and ECM has been associated with changes in the biomechanical properties of cartilage. In the present study we used AFM to quantify these changes in relation to the specific spatial pattern changes of the chondrocytes. With each pattern change, significant changes in elasticity were observed for both the PCM and ECM. Measuring the local elasticity thus allows for drawing direct conclusions about the degree of local tissue degeneration in OA.
Introduction
Articular cartilage is an avascular, aneural tissue. Sparsely scattered chondrocytes produce, organize, and maintain an expansive extracellular matrix (ECM) into which they are embedded. As a distinct and specialized part of the ECM, chondrocytes are surrounded by a thin layer of specialized matrix known as the pericellular matrix (PCM). The PCM acts as a mechanosensitive cell-matrix interface1 that protects the chondrocytes2 and modulates their biosynthetic response3. As previously described4, in healthy cartilage, chondrocytes are arranged in specific, distinct spatial pa....
Protocol
The human cartilage samples were obtained from patients undergoing total knee arthroplasty in the Department of Orthopaedic Surgery of the University Hospital of Tuebingen, Germany, and the Winghofer-hospital, Rottenburg a.N., Germany, for end-stage OA of the knee. Full departmental, institutional, and local ethical committee approval were obtained before commencement of the study (project number 674/2016BO2). Written informed consent was received from all patients before participation. The methods were carried out in ac.......
Representative Results
Along the physiopathological model from strings to double strings, to small and finally to big clusters, both ECM (Figure 3A) and PCM (Figure 3B) elastic moduli decreased significantly between each pattern change. The only exception was the difference in ECM between strings and double strings (p = 0.072). The results show that the ECM/PCM ratio (Figure 4B) did not change significantly, whereas a marked decrease in the absolute diffe.......
Discussion
Using AFM as a novel and powerful technique to measure the biomechanical properties of biological materials at a nanoscale level, we measured the elastic properties of the ECM and PCM in human osteoarthritic articular cartilage. Cartilage samples were selected according to their predominant spatial pattern of chondrocyte organization as an image-based biomarker for local tissue degeneration. As expected, a strong decline in the values of elasticity of both ECM and PCM was observed along spatial chondrocyte reorganization.......
Acknowledgements
We thank our co-authors from the original publication for their help and support.
....Materials
| Name | Company | Catalog Number | Comments |
| Amphotericin B | Merck | A2942 | |
| Atomic Force Microscope (AFM) | CellHesion 200, JPK Instruments, Berlin, Germany | JPK00518 | |
| AFM head | (CellHesion 200) JPK | JPK00518 | |
| Biocompatible sample glue | JPK Instruments AG, Berlin, Germany | H000033 | |
| Cantilever | tip C, k ¼ 7.4 N/m, All-In-One-AleTl, Budget Sensors, Sofia, Bulgaria | AIO-TL-10 | |
| Dulbecco's modified Eagle's medium (DMEM) | Gibco, Life Technologies, Darmstadt, Germany | 41966052 | |
| Inverted phase contrast microscope (Integrated with AFM) | AxioObserver D1, Carl Zeiss Microscopy, Jena, Germany | L201306_03 | |
| Leibovitz's L-15 medium without L-glutamine | (Merck KGaA, Darmstadt, Germany) | F1315 | |
| Microspheres | Polysciences | 07313-5 | |
| Penicillin-Streptomycin | Sigma | P4333 | |
| Petri dish heater associated with AFM | JPK Instruments AG, Berlin, Germany | T-05-0117 | |
| Scalpel | Feather | 2023-01 | |
| Tissue culture dishes | TPP Techno Plastic Products AG, Trasadingen, Switzerland | TPP93040 | |
| Tissue-tek O.C.T. Compound | Sakura Finetek, Alphen aan den Rijn, Netherlands | SA6255012 |
References
- Guilak, F., et al. The pericellular matrix as a transducer of biomechanical and biochemical signals in articular cartilage. Annals of the New York Academy of Sciences. 1068, 498-512 (2006).
- Peters, H. C., et al.
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