The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Häuselmann, H. J.
Right arrow Articles by Evans, C. H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Häuselmann, H. J.
Right arrow Articles by Evans, C. H.
The Journal of Immunology, 1998, 160: 1444-1448.
Copyright © 1998 by The American Association of Immunologists

Differences in Nitric Oxide Production by Superficial and Deep Human Articular Chondrocytes: Implications for Proteoglycan Turnover in Inflammatory Joint Diseases1

H. J. Häuselmann*,{dagger}, M. Stefanovic-Racic{ddagger}, B. A. Michel* and C. H. Evans2,{ddagger}

* Department of Rheumatology, University Hospital, Zürich, Switzerland; {dagger} M. E. Müller Institute for Biomechanics, University of Bern, Bern, Switzerland; and {ddagger} Ferguson Laboratory, Department of Orthopaedic Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
During inflammatory joint diseases, chondrocytes are exposed to cytokines such as IL-1 that induce the synthesis of nitric oxide (NO). Chondrocytes from different zones of the articular cartilage are known to have different metabolic properties. In the present study, we have demonstrated that chondrocytes recovered from the superficial zone of normal, human, articular cartilage synthesize approximately 2 to 3 times as much NO in response to IL-1 as chondrocytes recovered from the deep zone of the same cartilage. Production of NO by normal cartilage in response to IL-1 was also found to decrease with age. Addition of the NO synthase inhibitor NG-monomethyl-L-arginine (L-NMA, 1 mM) blocked NO production by cells of both zones. L-NMA completely reversed the suppression of proteoglycan synthesis imposed by IL-1 in deep chondrocytes, but produced only partial reversal in superficial cells. As noted previously, IL-1 failed to elicit a strong catabolic response in cultures of human cartilage. In the presence of L-NMA, however, IL-1 reduced the metabolic t1/2 of proteoglycans by approximately 50% in both the superficial and deep zones. This suggests that NO has, directly or indirectly, an anticatabolic effect in human cartilage. These data confirm the metabolic heterogeneity of human chondrocytes, and suggest that NO may be involved to different degrees as an endogenous modulator of the turnover of the cartilaginous matrix in different zones of articular cartilage.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Nitric oxide (NO)3 is synthesized in large amounts at sites of inflammation, including human rheumatoid joints (1, 2, 3). One of the most important intraarticular sources of NO is the articular cartilage (4), and the articular chondrocytes of all species yet tested produce very high quantities of this radical when activated by inflammatory mediators such as IL-1. Endogenously generated NO inhibits the synthesis of proteoglycan (5, 6, 7) and collagen (8) by chondrocytes, and thus may compromise the integrity of the cartilaginous surfaces in inflammatory joint diseases.

It is presently controversial whether or not NO is also a mediator of matrix catabolism. Two groups have presented evidence to suggest that NO induces the synthesis of metalloproteinases by monolayer cultures of articular chondrocytes (9, 10), but contrary evidence has been reported for cultures of bovine (11) and lapine (12) cartilage. The effects of NO on cartilage matrix metabolism have been recently reviewed (13, 14).

Recent research is providing increasing evidence that articular cartilage is not a homogenous tissue. Instead, the superficial articular chondrocytes that line the surface of the cartilage have important metabolic differences from the deep cells (15), including greater responsiveness to IL-1 and a greater resistance to the effects of the IL-1R antagonist (16). Such differences have important implications concerning the loss of articular cartilage in arthritis and possibilities for cartilage repair.

In this study, we have compared the ability of normal, human articular chondrocytes from the superficial and deep layers of cartilage to synthesize NO in response to IL-1. Advantage was taken of the alginate culture system within which chondrocytes retain their differentiated phenotype and in vivo metabolic behavior (17). Confirmatory experiments were performed with slices of articular cartilage recovered from the superficial or deep zones of this tissue. Prior research by Fukuda et al. (18), confirmed by Hayashi et al. (19), suggests that the superficial chondrocytes are a major source of NO in bovine cartilage. However, neither human cartilage nor the implications of this finding with regard to matrix turnover have been studied. Our studies, in contrast, include investigation of the differential effects of NO on proteoglycan turnover by superficial and deep chondrocytes, and shed new light on the puzzling observation that human cartilage, unlike the cartilage of experimental animals, does not mount a vigorous catabolic response to IL-1 (16).


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Reagents

Pronase was purchased from Calbiochem (La Jolla, CA); collagenase-P (Clostridium histolyticum, type CLS-2) was from Cooper Biomedicals (Cappel Worthington, Malvern, PA); FBS from HyClone Laboratories (Logan, UT); Ham’s F12/DMEM medium from Life Technologies (Grand Island, NY); and low viscosity alginate (Keltone LV) from Kelco (Chicago, IL). Sephadex G-25 (as prepacked PD-10 columns) was from Pharmacia (Piscataway, NJ). Radiolabeling was performed with [35S]sulfate, 25 to 40 Ci/mg, purchased from Amersham (Arlington Heights, IL). NG-monomethyl-L-arginine (L-NMA) was kindly synthesized by Dr. Paul Dowd and Wei Zhang, Department of Chemistry, University of Pittsburgh (Pittsburgh, PA); human rIL-1ß (hrIL-1ß) was purchased at R&D Systems Europe (Abington, U.K.). Bisbenzimidazole fluorescent dye (Hoechst dye 33258) was purchased from Polysciences (Warrington, PA); papain, L-arginine, and calf thymus DNA were from Sigma Chemical Co. (St. Louis, MO). All other chemicals were reagent grade and purchased from several different companies.

Cartilage sampling and isolation of chondrocytes

Tissues were recovered from 20 fresh human femoral condyles harvested postmortem from 10 donors, with no history of joint disease and macroscopically normal joint cartilage. Tissue was obtained through Examiners Office of University of Bern (Bern, Switzerland), according to their protocol and with institutional approval. The donors were six males (aged 21, 29, 39, 42, 52, and 55 yr) and four females (aged 15, 21, 32, and 50 yr). For four experiments, very thin superficial slices (less than 10% of wet weight of total cartilage) were collected separately from tissue harvested subsequently from the deep layers of articular cartilage of the medial and lateral femoral condyles. The middle layers (approximately 5–10% of the wet weight) were discarded to have a clear separation between superficial and deep chondrocytes. These two pools of tissue were rinsed, blotted, and weighed to determine the ratio of wet weights. For six experiments, articular cartilage representing the entire thickness of uncalcified tissue was cut from the weight-bearing surfaces of the medial and lateral femoral condyles. In eight experiments, the chondrocytes were isolated by proteases, encapsulated and cultured in alginate beads, as recently described by Guo et al. (20), and slightly modified by Häuselmann et al. (21). Briefly, the isolated cells were suspended in sterile 0.15 M NaCl containing low viscosity alginate gel (1.2%) at a density of 4 x 106 cells/ml of gel, then slowly expressed through a 22–1/2-gauge needle in a dropwise fashion into a 102 mM CaCl2 solution. After instantaneous gelation, the beads were allowed to polymerize further for a period of 10 min in the CaCl2 solution. After one wash in 10 vol of 0.15 M NaCl and three washes in 10 vol of Ham’s F12/DMEM medium, the beads finally were placed in 24-well plates.

Results of isolated and cultured chondrocytes from the eight donors were confirmed in part by culturing intact cartilage slices of the corresponding layers from four knee joints (two donors). Cultures were fed daily with F-12/DMEM (1:1) supplemented with 10% FCS and 25 µg/ml ascorbate, and incubated at 37°C in a humidified atmosphere of 5% of CO2 in air.

Treatment of chondrocytes in alginate and cartilage organ cultures with hrIL-1ß, and L-NMA

Six days after the beginning of the culture period, the cells or organ cultures were treated daily for 3 days (proteoglycan synthesis) or 12 days (proteoglycan catabolism) with 1 to 2000 pg/ml hrIL-1ß in the presence or absence of 1 mM L-NMA. Other cultures were maintained as controls without any additions. For each experiment with chondrocytes from cartilage layers, four wells with three alginate beads/well containing cultured chondrocytes or one piece of cartilage (in organ culture experiments) were used. In experiments with a mixed population of cells, five beads or one full-thickness piece of cartilage per well were used.

Measurement of synthesis of proteoglycans

To assess PG synthesis, cultures were treated daily with cytokines for 3 days, followed by incubation for 6 h in the same medium with addition of 50 µCi/ml of [35S]sulfate (sp. act. 25–40 Ci/mg; Amersham Corp., Arlington Heights, IL). After 6 h, PGs were extracted from alginate gel cultures under dissociative condition in 4 M guanidinium chloride containing 20 mM EDTA and protease inhibitors, as described previously (15). Proteoglycans from organ cultures were extracted with 4 M guanidinium chloride after overnight digestion with papain (see DNA measurements of organ cultures). Samples were stored at -70°C until analyzed. 35S-labeled proteoglycans (35S-proteoglycans) in the media and dissociative extracts were quantified by liquid scintillation spectroscopy after chromatography on Sephadex G-25 M in PD 10 columns, as described earlier (15). Parallel cultures of isolated chondrocytes were digested at 60°C with papain, and the DNA content was measured by fluorescence using Hoechst dye 33258 (22).

Measurement of catabolism of newly synthesized 35S-proteoglycans

Cells were labeled with 50 Ci/ml 35SO42- for 12 h. The labeling period then was followed by a chase period of 12 days, with daily replenishment of media and specific treatment of the chondrocytes. The rate of loss (in percentage) of newly synthesized 35S-proteoglycans from cultured chondrocytes and slices was calculated daily by measuring 35S-proteoglycans appearing in the daily medium. The cumulative daily loss of 35S-proteoglycans during the chase period was added to the 35S-proteoglycans remaining in the beads or slices at the end of the experiment and considered as total 35S-proteoglycan-cpm and set to 100% (15, 21). The logarithm of both, the percentage of 35S-proteoglycans remaining in the cultures, and the percentage of 35S-proteoglycans appearing daily in the medium were plotted as a function of time during the chase period. The t1/2 of the newly synthesized 35S-proteoglycans were determined with a curve fit program of sigmaplot from Jandel Corporation (San Rafael, CA) for a single or double exponential decay equation: single exponential decay, f(x) = ae-bx, double exponential decay; f(x) = ae-bx + ce-bx. The single exponential decay starts at a, when x = 0, and decreases with a time constant 2/b to zero with increasing x when b > 0. The double exponential decay starts at a + c when x = 0, and decreases with the two time constants 1/b and 1/d to zero with increasing x. t1/2 is calculated as 0.69/b. Decay of 35S during the experiment was calculated and compensated for (a, amplitude of first exponential; b, rate constant of first exponential; c, amplitude of second exponential; d, rate constant of second exponential; and e, base of natural logarithm = 2718; x, time variable).

Determination of DNA content in chondrocytes and organ cultures

DNA content of the isolated and cultured chondrocytes as well as organ cultures was quantified using the bisbenzimidazole fluorescent dye, Hoechst 33258 (22). Alginate cultures (one well per group) and each slice from organ cultures were digested with papain (125 µg/ml) and evaluated in a spectrofluorometer, as published earlier by Aydelotte and Kuettner (23).

Measurement of nitrite

NO production was measured as NO2- formation using a spectrophotometric assay based upon the Griess reaction (24). We have shown previously that approximately 50% of the NO generated by chondrocytes and cartilage in culture accumulates as NO2- over a wide range of NO concentrations (4, 11). Nitrite concentration was measured daily in conditioned medium of triplicate (organ cultures) and quadruplicate cultures (isolated chondrocytes) per group.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
NO production

Superficial chondrocytes, whether cultured in alginate gels or as organ cultures, reproducibly produced 2 to 3 times as much NO per cell as cells originating from the deeper layers of normal, human, articular cartilage (Fig. 1Go, A and B). The maximum concentration of hrIL-1ß, 2 ng/ml, used in these experiments is well above the concentration of hrIL-1ß that we have shown previously to provoke maximum NO synthesis by human chondrocytes. In each case, 1 mM of L-NMA completely inhibited NO production (Fig. 1Go, C and D).



View larger version (26K):
[in this window]
[in a new window]
  		FIGURE 1. NO2- content of culture supernatants of superficial (left, A and C) and deep (right, B and D) chondrocytes after 1 to 9 days of treatment with or without 2 ng/ml hrIL-1ß in the absence (A and B) or presence (C and D) of 1 mM L-NMA. Open circles = control cultures; open squares = cultures treated with hrIL-1ß. Values given are means ± SD (n = 16).

 
NO synthesis was maximum after 2 days of treatment with hrIL-1ß. The subsequent decline in the rate of NO synthesis occurred despite daily replenishment of the cultures with fresh medium and hrIL-1ß. A similar phenomenon has been noted with bovine (11) and lapine (12) chondrocytes.

Proteoglycan synthesis

The addition of hrIL-1ß to alginate cultures of articular chondrocytes, harvested from throughout the cartilage, strongly inhibited the incorporation of 35SO42- into newly synthesized proteoglycans. This inhibition was partially reversed by the addition of L-NMA (Fig. 2GoA).



View larger version (29K):
[in this window]
[in a new window]
  		FIGURE 2. Inhibition of 35S incorporation into proteoglycan macromolecules by human articular chondrocytes from full-thickness, superficial, and deep cartilage layers treated with hrIL-1ß in the presence or absence of L-NMA. The isolated cells from full-thickness cartilage (A) or cartilage layers (B, open bars = superficial; hatched bars = deep) were treated for 3 days with 0.5 ng/ml hrIL-1ß in the presence or absence of 1 mM L-NMA. Values given are means ± SD (A, n = 12; B, n = 3).

 
The experiment was then repeated with chondrocytes harvested selectively from the superficial or deep layers of the cartilage. As shown in Figure 2GoB, L-NMA completely reversed the inhibitory effects of hrIL-1ß upon proteoglycan synthesis by chondrocytes isolated from the deep layers of the cartilage, but only partially restored proteoglycan synthesis by superficial chondrocytes.

Proteoglycan degradation

The rates of release of 35S-labeled proteoglycans from alginate cultures of superficial and deep chondrocytes are shown in Figure 3Go. As noted previously for human chondrocytes, basal rates of release were low and were not increased by the addition of IL-1. However, catabolism was increased after 6 days of IL-1 treatment by the addition of L-NMA; when added alone, L-NMA did not increase proteoglycan degradation.



View larger version (20K):
[in this window]
[in a new window]
  		FIGURE 3. Effect of L-NMA upon the daily release of 35S-proteoglycans by human adult articular chondrocytes cultured in alginate. A and B, Show the percentage of daily release of 35S-proteoglycan from the alginate cultures established from superficial (A) and deep (C) cells. The values at each time point reflect the mean and SD for the analysis of three separate cultures. The logarithm of the percentage of 35S-proteoglycans released every day in the medium was plotted as a function of time during the chase period (circles, control; squares, IL-1ß, 2 ng/ml; triangles, hrIL-1ß, 2 ng/ml, plus L-NMA, 1 mM; inverted triangles, L-NMA, 1 mM). White crosses in black fields in A and B denote values in which the difference between cultures treated with IL-1ß alone and those treated with IL-1ß + L-NMA differs with a statistical significance of p <= 0.05 (t test).

 
The data were recalculated to provide t1/2 for the newly synthesized proteoglycans (Table IGo). Proteoglycans synthesized by the deep cells had a longer t1/2 than these synthesized by the superficial cells. In each case, L-NMA reduced the t1/2 of the proteoglycans by approximately 50%.


View this table:
[in this window]
[in a new window]
  		Table I. Effect of L-NMA upon the half-lives of newly synthesized 35S-proteoglycans from cultured human chondrocytes

 
Organ cultures

To confirm that the above data were not an artifact of the alginate culture system, key experiments were repeated with slices of otherwise intact cartilage obtained from the superficial and deep zones. In each case, the findings were compatible with those obtained with the alginate cultures (data not shown).

Effect of age

During the course of these experiments, it was noted that NO production by full-thickness cartilage declined as a function of age (Fig. 4Go).



View larger version (46K):
[in this window]
[in a new window]
  		FIGURE 4. Influence of age on maximal IL-1-induced NO production (0.5 ng/ml IL-1ß) by adult human articular chondrocytes cultured in alginate, expressed as percentages of control cultures. Values given are means ± SD (n = 3).

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
These data show that chondrocytes from the superficial zone of normal, human, articular cartilage synthesize much more NO than cells harvested from the deep zone. Endogenously produced NO inhibits the synthesis of proteoglycans in both cases. With the deep cells, NO appears to account entirely for the suppressive effect of hrIL-1ß on proteoglycan biosynthesis. However, in cultures of superficial cells, a substantial component of the inhibition of proteoglycan synthesis by IL-1 is independent of NO. Whether this reflects the different species of proteoglycan synthesized by the superficial and deep cells cannot be determined from the present data. Further, detailed studies comparing the proteoglycans synthesized by superficial and deep cells, and how NO affects each of these are required to clarify this matter. The different responses of the superficial and deep cells help explain our earlier observation (7) that L-NMA alleviates only partially the suppression of proteoglycan synthesis imposed by IL-1 in human chondrocytes derived from full-thickness cartilage.

The age-related decline in NO production by cartilage is intriguing and consistent with the observation that the ability of human chondrocytes to produce NO in response to IL-1 declines with passage (25). Although the cartilages used in this study were grossly normal, there remains the possibility that the decline in NO production is related to increased microdamage with age. An alternative explanation is suggested by the data of Blanco et al. (26), showing that NO induces apoptosis in human articular chondrocytes. This suggests the ironic possibility that the decline in NO production with age reflects the fact that those cells producing the most NO have been eliminated selectively by this radical in an autocrine fashion.

Most specimens of full-thickness human articular cartilage slices in culture are largely insensitive to the catabolic actions of IL-1 (16). Even if basal catabolic levels of newly synthesized 35S-proteoglycan are enhanced in superficial layers of normal adult human cartilage, in contrast to deep layers of full-thickness cartilage, IL-1-induced degradation of newly synthesized 35S-proteoglycan is still low. Therefore, our new finding of significantly enhanced catabolism of newly synthesized proteoglycan if NO synthesis is blocked is an important one. The increased release of proteoglycan from the human slices of both superficial and deep layers starts late, between days 4 and 7 of the experiment, and increases with time up to the last experiment of day 12. This leads to a significant shortening of the t1/2 of newly synthesized 35S-proteoglycans in superficial and deep cartilage layers down to approximately 50% of control cultures. Neither treatment with L-NMA nor IL-1ß alone (as mentioned above) significantly increased daily loss of newly synthesized 35S-proteoglycan compared with control. The complete inhibition of NO production in both layers in these catabolic studies clearly demonstrates that NO, in contrast to its effect in proteoglycan synthesis inhibition, may be a protective agent with respect to proteoglycan catabolism. A similar effect has been demonstrated recently in bovine (11) and lapine (12) cartilage. These results again underline the fact that there might be completely different pathways of synthesis inhibition and catabolism of proteoglycan induced by IL-1 in human articular cartilage.


   Footnotes
 
1 This work was supported in part by National Institutes of Health Grant RO1 AR42025 (C.H.E.) and Swiss National Science Foundation Grant 3200-041558.94/1 (H.J.H.). Back

2 Address correspondence and reprint requests to Dr. H. J. Häuselmann, Department of Rheumatology, Gloria Strasse 25, Zurich, CH8091 Switzerland. Back

3 Abbreviations used in this paper: NO, nitric oxide; hr, human recombinant; L-NMA, NG-monomethyl-L-arginine. Back

Received for publication July 15, 1997. Accepted for publication October 21, 1997.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Farrell, A. J., D. R. Blake, R. M. Palmer, S. Moncada. 1992. Increased concentrations of nitrite in synovial fluid and serum suggest increased nitric oxide synthesis in rheumatic diseases. Ann. Rheum. Dis. 51:1219.[Abstract/Free Full Text]
  2. Ueki, Y., S. Miyake, Y. Tominaga, K. Eguchi. 1996. Increased nitric oxide levels in patients with rheumatoid arthritis. J. Rheumatol. 23:230.[Medline]
  3. Sakurai, H., H. Kohsaka, M. F. Liu, H. Higashiyama, Y. Hirta, K. Kanno, I. Siato, N. Miyasaka. 1995. Nitric oxide production and inducible nitric oxide synthase expression in inflammatory arthritides. J. Clin. Invest. 96:2357.
  4. Stadler, J., M. Stefanovic-Racic, T. R. Billiar, R. D. Curran, L. A. McIntyre, H. I. Georgescu, R. L. Simmons, C. H. Evans. 1991. Articular chondrocytes synthesize nitric oxide in response to cytokines and lipopolysaccharide. J. Immunol. 147:3915.[Abstract]
  5. Taskiran, D., M. Stefanovic-Racic, H. I. Georgescu, C. H. Evans. 1994. Nitric oxide mediates suppression of cartilage proteoglycan synthesis by interleukin-1. Biochem. Biophys. Res. Commun. 200:142.[Medline]
  6. Jarvinen, T. A. H., T. Moilanen, T. L. N. Jarvinen, E. Moilanen. 1995. Nitric oxide mediates interleukin-1 induced inhibition of glycosaminoglycan synthesis in rat articular cartilage. Med. Inflam. 4:107.
  7. Häuselmann, H. J., L. Oppliger, B. A. Michel, M. Stefanovic-Racic, C. H. Evans. 1994. Nitric oxide and proteoglycan biosynthesis by human articular chondrocytes in alginate culture. FEBS Lett. 352:361.[Medline]
  8. Cao, M., A. Westerhausen-Larson, C. Niyibizi, K. Kavalkovich, H. I. Georgescu, C. F. Rizzo, M. Stefanovic-Racic, C. H. Evans. 1997. Nitric oxide inhibits the synthesis of type II collagen without altering Col2A1 mRNA abundance: prolyl hydroxylase as a possible target. Biochem. J. 324:305.
  9. Murrell, G. A. C., D. Jang, R. J. Williams. 1995. Nitric oxide activates metalloproteinase enzymes in articular cartilage. Biochem. Biophys. Res. Commun. 206:15.[Medline]
  10. Tamura, T., T. Nakanishi, Y. Kimura, H. Takako, K. Sasaki, N. Hiromichi, K. Takahashi, M. Takigawa. 1996. Nitric oxide mediates interleukin-1 matrix degradation and basic fibroblast growth factor release in cultured rabbit articular chondrocytes: a possible mechanism of pathological neovascularization in arthritis. Endocrinology 137:3729.[Abstract]
  11. Stefanovic-Racic, M., T. I. Morales, D. Taskiran, L. A. McIntyre, C. H. Evans. 1996. The role of nitric oxide in proteoglycan turnover by bovine articular cartilage organ cultures. J. Immunol. 156:1213.[Abstract]
  12. Stefanovic-Racic, M., M. O. Möllers, L. A. Miller, C. H. Evans. 1997. Nitric oxide and proteoglycan turnover in rabbit articular cartilage. J. Orthop. Res. 15:442.[Medline]
  13. Evans, C. H., S. C. Watkins, M. Stefanovic-Racic. 1996. Nitric oxide and cartilage metabolism. Methods Enzymol. 269:75.[Medline]
  14. Cao, M., H. J. Häuselmann, M. Stefanovic-Racic, and C. H. Evans. 1997. Nitric oxide and cartilage matrix turnover. In Nitric Oxide and Bone Disease. M. Hukkanen, J. Polak, and S. P. F. Hughes, eds. Cambridge University Press, Cambridge, in press.
  15. Aydelotte, M. B., R. R. Greenhill, K. E. Kuettner. 1988. Differences between sub-populations of cultured bovine articular chondrocytes. II. Proteoglycan metabolism. Connect. Tissue Res. 18:223.[Medline]
  16. Häuselmann, H. J., J. Flechtenmacher, L. Michal, E. J. M. A. Thonar, M. Shinmei, K. E. Kuettner, M. B. Aydelotte. 1996. The superficial layer of human articular cartilage is more susceptible to interleukin-1-induced damage than the deeper layers. Arthritis Rheum. 39:478.[Medline]
  17. Häuselmann, H. J., R. J. Fernandes, S. S. Mok, T. M. Schmid, J. A. Block, M. B. Aydelotte, K. E. Kuettner, E. J. M. A. Thonar. 1994. Phenotypic stability of bovine articular chondrocytes after long-term culture in alginate beads. J. Cell Sci. 107:17.[Abstract]
  18. Fukuda, K., F. Kumano, M. Takayama, M. Saito, K. Otani, S. Tanaka. 1995. Zonal differences in nitric oxide synthesis by bovine chondrocytes exposed to interleukin-1. Inflamm. Res. 44:437.
  19. Hayashi, T., H. E. Jasin, E. Abe, T. Yamate, Y. Tagudni. 1997. Nitric oxide production by superficial and deep articular chondrocytes. Arthritis Rheum. 40:261.[Medline]
  20. Guo, J., G. W. Jourdian, K. McCallum. 1989. Culture and growth characteristics of chondrocytes encapsulated in alginate beads. Connect. Tissue Res. 19:277.[Medline]
  21. Häuselmann, H. J., A. B. Aydelotte, B. L. Schumacher, K. E. Kuettner, S. H. Gitelis, E. J. M. A. Thonar. 1992. Synthesis and turnover of proteoglycans by human and bovine adult articular chondrocytes cultured in alginate beads. Matrix 12:116.[Medline]
  22. Kim, Y. J., R. L. Y. Sah, J. Y. H. Doong, A. J. Grodzinsky. 1988. Fluorometric assay of DNA in cartilage explants using Hoechst 33258. Anal. Biochem. 174:168.[Medline]
  23. Aydelotte, M. B., K. E. Kuettner. 1988. Differences between sub-populations of cultured bovine articular chondrocytes. I. Morphology and cartilage matrix production. Connect. Tissue Res. 18:205.[Medline]
  24. Green. L. C., D. A., J. Wagner, P. L. Glogowski, J. S. Skipper, J. S. Wishnok, S. R. Tannenbaum. 1982. Analysis of nitrate, nitrite and [15N] nitrate in biological fluids. Anal. Biochem. 126:131.[Medline]
  25. Blanco, F. J., Y. Geng, M. Lotz. 1995. Differentiation-dependent effects of IL-1 and TGF-ß on human articular chondrocyte proliferation are related to inducible nitric oxide synthase expression. J. Immunol. 154:4018.[Abstract]
  26. Blanco, F. J., R. L. Ochs, H. Schwarz, M. Lotz. 1995. Chondrocyte apoptosis induced by nitric oxide. Am. J. Pathol. 146:75.[Abstract]



This article has been cited by other articles:


Home page
 Proc. Natl. Acad. Sci. USAHome page
N. Taniguchi, B. Carames, Y. Kawakami, B. A. Amendt, S. Komiya, and M. Lotz
Chromatin protein HMGB2 regulates articular cartilage surface maintenance via {beta}-catenin pathway
PNAS, September 29, 2009; 106(39): 16817 - 16822.
[Abstract] [Full Text] [PDF]

Home page
 Reproductive SciencesHome page
G. Chiossi, G. Verocchi, P. Venturini, and F. Facchinetti
Changes in Cervical Nitric Oxide Concentration Correlate With Bishop Score and Cervical Length Modifications in Prostaglandin E2-Mediated Induction of Labor
Reproductive Sciences, April 1, 2006; 13(3): 203 - 208.
[Abstract] [PDF]

Home page
 Rheumatology (Oxford)Home page
N. J. Goodstone and T. E. Hardingham
Tumour necrosis factor {alpha} stimulates nitric oxide production more potently than interleukin-1{beta} in porcine articular chondrocytes
Rheumatology, August 1, 2002; 41(8): 883 - 891.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Cell Physiol.Home page
C. C. Teixeira, K. Mansfield, C. Hertkorn, H. Ischiropoulos, and I. M. Shapiro
Phosphate-induced chondrocyte apoptosis is linked to nitric oxide generation
Am J Physiol Cell Physiol, September 1, 2001; 281(3): C833 - C839.
[Abstract] [Full Text] [PDF]

Home page
 Ann Rheum DisHome page
I Shirazi, I Yaron, Y Wollman, M Blum, T Chernihovsky, R Judovich, A Iaina, and M Yaron
Down regulation of interleukin 1{beta} production in human osteoarthritic synovial tissue and cartilage cultures by aminoguanidine
Ann Rheum Dis, April 1, 2001; 60(4): 391 - 394.
[Abstract] [Full Text] [PDF]

Home page
 Mol Hum ReprodHome page
M.-A. Ledingham, A. J. Thomson, A. Young, L. M. Macara, I. A. Greer, and J. E. Norman
Changes in the expression of nitric oxide synthase in the human uterine cervix during pregnancy and parturition
Mol. Hum. Reprod., November 1, 2000; 6(11): 1041 - 1048.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
R. Gassner, M. J. Buckley, H. Georgescu, R. Studer, M. Stefanovich-Racic, N. P. Piesco, C. H. Evans, and S. Agarwal
Cyclic Tensile Stress Exerts Antiinflammatory Actions on Chondrocytes by Inhibiting Inducible Nitric Oxide Synthase
J. Immunol., August 15, 1999; 163(4): 2187 - 2192.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Häuselmann, H. J.
Right arrow Articles by Evans, C. H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Häuselmann, H. J.
Right arrow Articles by Evans, C. H.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS