Quantitation of autoradiographic grains in different zones of articular cartilage with image analyzer

Parkkinen, Jyrki; Paukkonen, Kari; Pesonen, Erkki; Markkanen, Seppo; Helminen, Heikki J.; Tammi, Markku

doi:10.1007/bf00266384

Cited by 12 publications

(9 citation statements)

References 18 publications

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“…Aggrecan is the PG that is responsible for most of the elastic properties of cartilage, and its metabolism plays a key role in the health of the joint. PG synthesis rate increases toward the deep zones [90,91], and the synthesis varies in different locations of the joint cartilage; this has been observed especially in the superficial zone of articular cartilage [91]. Factors such as soluble mediators, mechanical stress, aging, and OA can influence the balance and overall rate of turnover and change the extracellular matrix protein gene expressions [92].…”

Section: Proteoglycan Synthesis and Turnover In Young Age Aging And Oamentioning

confidence: 99%

Regular joint loading in youth assists in the establishment and strengthening of the collagen network of articular cartilage and contributes to the prevention of osteoarthrosis later in life: a hypothesis

et al. 2000

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importance of regular joint loading in young age for the development of a well-organized and strong articular cartilage collagen network, which contributes to the prevention of OA later in life. The load-bearing properties of articular cartilage as well as the main features of the metabolism and turnover of proteoglycans (PGs) and collagen are briefly reviewed, especially in relation to joint loading. Prevention of OAThe knowledge of the pathogenesis of OA and its risk factors is today supplemented at an increasing pace, which makes the prospects for prevention of the disease more and more realistic [2][3][4][5][6][7][8]. OA is a disease with a multifactorial etiology. It affects not only the articular cartilage but the whole synovial joint organ. This implies that in addition to articular cartilage, the properties of the subchondral bone and ligaments, the weight-bearing conditions including the biomechanical alignment of the joint, the weakness of the shockabsorbing and weight-bearing muscles, and the integrity and proper function of the neuromuscular system must be taken into regard [9]. Even though the OA process in the joint structures is accomplished by biochemical signals, i.e., cytokines, growth factors, proteins and enzymes, the disease appears to be primarily driven by the local mechanical factors in the joint. Thus, the mechanical etiology of OA deserves ongoing close scrutiny [9].The primary prevention of OA emphasizes hindering the disease from affecting healthy persons [2,5]. Large epidemiological studies on OA have pointed out several risk factors that can be influenced by adjusting our lifestyles. For example, obese persons are instructed to reduce body weight and to carry out exercises to strengthen their lower limb muscles to reduce stress at

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Section: Proteoglycan Synthesis and Turnover In Young Age Aging And Oamentioning

confidence: 99%

Regular joint loading in youth assists in the establishment and strengthening of the collagen network of articular cartilage and contributes to the prevention of osteoarthrosis later in life: a hypothesis

et al. 2000

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“…By correlating grain numbers and stereologically measured structural parameters (cell and nuclear volume, surface area, and radii) from the same regions within loaded disks, we found that changes in aggrecan synthesis correlated strongly with changes in cell and nuclear structure (Buschmann et al, 1995b). Previous investigations have provided automated grain counting procedures either for liquid emulsions or autoradiographic film (Puustjarvi et al, 1993;Parkkinen et al, 1990;Sklarew, 1982). In most cases, the quantitative nature of the method was characterized by comparing the results of automated grain counts to visual counts, the latter considered as the standard (Parkkinen et al, 1990;Sklarew, 1982).…”

Section: Spatial Profiles Of Suvate Incorporationmentioning

confidence: 99%

“…Previous investigations have provided automated grain counting procedures either for liquid emulsions or autoradiographic film (Puustjarvi et al, 1993;Parkkinen et al, 1990;Sklarew, 1982). In most cases, the quantitative nature of the method was characterized by comparing the results of automated grain counts to visual counts, the latter considered as the standard (Parkkinen et al, 1990;Sklarew, 1982). Therefore, the possibility of nonlinear emulsion response or saturation was not assessed.…”

Section: Spatial Profiles Of Suvate Incorporationmentioning

confidence: 99%

A method of quantitative autoradiography for the spatial localization of proteoglycan synthesis rates in cartilage.

Buschmann¹,

Maurer²,

Berger³

et al. 1996

J Histochem Cytochem.

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Incorporation of [35S]-sulfate into cartilage tissue indicates the synthesis of aggrecan, the large aggregating proteoglycan (PG) that endows cartilage with resistance to compression. Scintillation counting of tissue digests provides a quantitative measure of incorporated sulfate but does not provide information on the spatial location of synthesis within the tissue. Such spatially specific information is necessary to determine which cell populations respond to diffusible factors and to correlate local mechanical events (e.g., deformation, interstitial fluid stress) to cellular biosynthetic responses. The aim of this study was to develop and characterize a liquid emulsion autoradiography technique, including an automated grain counting procedure, to derive spatial profiles of aggrecan synthesis rates in cartilage. We chose mature 10-14-month-old bovine humeral head articular cartilage as a model system and applied a liquid emulsion autoradiography technique to [35S]-sulfate-labeled, resin-embedded, and semithin-sectioned tissue explants. High-magnification light microscopy color images were captured on a computer. Automated image analysis for grain number determination included a color thresholding procedure to discriminate grains from the lightly stained structural image and computation of the average area of a single grain from each image. Determination of grain number, whether originating from single grains or grain clusters, was performed by dividing the total grain area in the image by the average area of a single grain in the same image. This procedure largely eliminated the effects of variations of microscope light intensity, camera performance, image focus, section stain intensity, and thresholding on the resulting grain numbers. By altering the specific activity of the medium radiolabel and the emulsion exposure times, we demonstrated a linear dose dependence, without saturation, of grain number on radioactive content in the underlying section. By cutting specimens in half and performing liquid scintillation counting on one half and autoradiography on the other half, we found that each disintegration occurring in the section during exposure resulted in 0.67 +/- 0.21 grains (mean +/- SD; n = 58). Therefore, counted grain numbers can be directly converted to incorporated sulfate, largely reflecting the synthesis of the PG aggrecan. As an example of calculated intratissue profiles of aggrecan synthesis rates, we found for the mature bovine tissue in serum-free medium that aggrecan synthesis increases monotonically from the articular surface to the radial zone by as much as tenfold.

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“…Presently, techniques utilizing computerized image analysis that reduce the subjectivity of histomorphometric assessment of articular cartilage degradation and repair are available (7,8,16). Furthermore, with the development of the appropriate software, semiquantitative histochemical scoring of articular cartilage is now possible (14,23).…”

mentioning

confidence: 98%

“…Presently, techniques utilizing computerized image analysis that reduce the subjectivity of histomorphometric assessment of articular cartilage degradation and repair are available (7,8,16 The purpose of the present study was to incorporate techniques of image analysis in the semiquantitative assessment of glycosaminoglycans in articular cartilage and to demonstrate the efficacy of such techniques in evaluating osteoarthritis in a rabbit model (23).…”

mentioning

confidence: 99%

Method of histomorphometric assessment of glycosaminoglycans in articular cartilage

Shimizu

Coutts

Healey

et al. 1997

Journal Orthopaedic Research

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A method of image analysis has been developed for use in the semiquantitative histomorphometric assessment of glycosaminoglycans in articular cartilage stained with safranin O. The reliability of the methodology is reported along with its application to the assessment of articular cartilage in a model of osteoarthritis, i.e., transection of the anterior cruciate ligament in rabbits. With this system, specimens of normal and osteoarthritic articular cartilage were assessed histomorphometrically for the following parameters: total cartilage area, percentage of safranin O stained area, mean gray scale (average stain intensity), and gray scale index (the relative total amount of glycosaminoglycans). Reproducibility was established for 12 specimens of normal cartilage and found to have a SD of less than 8% of the mean for each parameter that was measured. Image analysis of osteoarthritic cartilage revealed each of the parameters, except for average stain intensity, to be significantly lower than that in control cartilage.

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Quantitation of autoradiographic grains in different zones of articular cartilage with image analyzer

Cited by 12 publications

References 18 publications

Regular joint loading in youth assists in the establishment and strengthening of the collagen network of articular cartilage and contributes to the prevention of osteoarthrosis later in life: a hypothesis

Regular joint loading in youth assists in the establishment and strengthening of the collagen network of articular cartilage and contributes to the prevention of osteoarthrosis later in life: a hypothesis

A method of quantitative autoradiography for the spatial localization of proteoglycan synthesis rates in cartilage.

Method of histomorphometric assessment of glycosaminoglycans in articular cartilage

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