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Human fibroblast-like synoviocyte isolation matter: a comparison between cell isolation from synovial tissue and synovial fluid from patients with rheumatoid arthritis

SUMMARY

OBJECTIVE:

Cell culture technology has become a popular method in the field of cell biology, pharmacology, and medical researches. Primary cells represent the normal physiological condition of human cells. Fibroblasts are the most common native cells of connective tissue that play a crucial role in the entire pathogenesis of various disorders, such as rheumatoid arthritis (RA). Fibroblast-like synoviocytes (FLSs), which overlie the loose connective tissue of the synovial sublining, are known to be the central mediators of joint damage. The most routine approach for the isolation of FLS is an enzymatic digestion of synovial tissue. This experimental study is designed to introduce an easy, fast, and high-throughput method compared with enzymatic digestion for isolation of FLS.

METHODS:

The synovial tissue and synovial fluid (SF) samples were collected from eight patients with RA who underwent routine knee replacement surgery. Synovial tissue was incubated with collagenase VIII enzyme, while SF was washed with a similar volume of phosphate-buffered saline. The cells were further subcultured and stored based on the standard protocols. The purity of isolated synoviocytes was confirmed using flow cytometry analysis.

RESULTS:

Isolation of FLS from SF was more successful with a faster rate, 3–5 days after culture. The morphological assessment and flow cytometry analysis confirmed the purity of SF-derived cells in passage 4.

CONCLUSIONS:

SF could be a more accessible source of FLS than synovial tissue. Obtaining primary FLS from SF is a simple, fast, and cost-effective way to have a large-scale cell during a short time.

KEYWORDS:
Cell culture; Cell isolation; Fibroblast-like synoviocyte; Synovium; Synovial fluid

INTRODUCTION

Rheumatoid arthritis (RA) is a complex and multisystem disorder that primarily affects the synovial joints11. Guo Q, Wang Y, Xu D, Nossent J, Pavlos NJ, Xu J. Rheumatoid arthritis: pathological mechanisms and modern pharmacologic therapies. Bone Res. 2018;27(6):15. https://doi.org/10.1038/s41413-018-0016-9
https://doi.org/10.1038/s41413-018-0016-...
,22. Mellado M, Martínez-Muñoz L, Cascio G, Lucas P, Pablos JL, Rodríguez-Frade JM. T cell migration in rheumatoid arthritis. Front Immunol. 2015;6:384. https://doi.org/10.3389/fimmu.2015.00384
https://doi.org/10.3389/fimmu.2015.00384...
. Fibroblast-like synoviocytes (FLSs) and macrophage-like synoviocytes (MLSs) are two main resident cells in the intimal layer of the synovial membrane, which play a central role in the joint pathology of RA33. Yang X, Chang Y, Wei W. Emerging role of targeting macrophages in rheumatoid arthritis: focus on polarization, metabolism and apoptosis. Cell Prolif. 2020;53(7):e12854. https://doi.org/10.1111/cpr.12854
https://doi.org/10.1111/cpr.12854...
. Nevertheless, FLSs are the most common cell at the bone–pannus interface with a more abundant population than MLSs44. Tu J, Hong W, Zhang P, Wang X, Körner H, Wei W. Ontology and function of fibroblast-like and macrophage-like synoviocytes: how do they talk to each other and can they be targeted for rheumatoid arthritis therapy? Front Immunol. 2018;9:1467. https://doi.org/10.3389/fimmu.2018.01467
https://doi.org/10.3389/fimmu.2018.01467...
,55. Jung SM, Kim KW, Yang CW, Park SH, Ju JH. Cytokine-mediated bone destruction in rheumatoid arthritis. J Immunol Res. 2014;2014:263625. https://doi.org/10.1155/2014/263625
https://doi.org/10.1155/2014/263625...
. The migration and invasion of FLS into cartilage and bone are a key event in synovial hyperplasia, resulting in cartilage destruction in patients with RA66. Bartok B, Firestein GS. Fibroblast-like synoviocytes: key effector cells in rheumatoid arthritis. Immunol Rev. 2010;233(1):233-55. https://doi.org/10.1111/j.0105-2896.2009.00859.x
https://doi.org/10.1111/j.0105-2896.2009...
,77. Ostrowska M, Maśliński W, Prochorec-Sobieszek M, Nieciecki M, Sudoł-Szopińska I. Cartilage and bone damage in rheumatoid arthritis. Reumatologia. 2018;56(2):111-20. https://doi.org/10.5114/reum.2018.75523
https://doi.org/10.5114/reum.2018.75523...
. Activated FLSs produce a wide range of inflammatory mediators, which promote the recruitment and activation of circulating and resident immune cells. Also, the migration of arthritis to unaffected joints has been attributed to the transmigration of FLSs88. Bustamante MF, Garcia-Carbonell R, Whisenant KD, Guma M. Fibroblast-like synoviocyte metabolism in the pathogenesis of rheumatoid arthritis. Arthritis Res Ther. 2017;19(1):110. https://doi.org/10.1186/s13075-017-1303-3
https://doi.org/10.1186/s13075-017-1303-...
,99. Nevius E, Gomes AC, Pereira JP. Inflammatory cell migration in rheumatoid arthritis: a comprehensive review. Clin Rev Allergy Immunol. 2016;51(1):59-78. https://doi.org/10.1007/s12016-015-8520-9
https://doi.org/10.1007/s12016-015-8520-...
. Although there is still much to be learned about their resolution in RA disease, FLSs have sparked a lot of attention in recent studies. The culture and growth of FLSs is the first step for the investigation and analysis of these cells. At present, the synovium is the main and traditional source of FLS, and access to the synovium is feasible only during arthroplasty or arthroscopy surgery. In advanced RA, total joint arthroplasty (TJA) has been suggested as a successful intervention when optimal medical and supportive therapies have failed1010. Lee JK, Choi CH. Total knee arthroplasty in rheumatoid arthritis. Knee Surg Relat Res. 2012;24(1):1-6. https://doi.org/10.5792/ksrr.2012.24.1.1
https://doi.org/10.5792/ksrr.2012.24.1.1...
,1111. Danoff JR, Moss G, Liabaud B, Geller JA. Total knee arthroplasty considerations in rheumatoid arthritis. Autoimmune Dis. 2013;2013:185340. https://doi.org/10.1155/2013/185340
https://doi.org/10.1155/2013/185340...
. It has been reported that early and continuous use of biological and disease-modifying antirheumatic drugs (DMARDs) delays TJA1212. Jämsen E, Virta LJ, Hakala M, Kauppi MJ, Malmivaara A, Lehto MUK. The decline in joint replacement surgery in rheumatoid arthritis is associated with a concomitant increase in the intensity of anti-rheumatic therapy: a nationwide register-based study from 1995 through 2010. Acta Orthop. 2013;84(4):331-7. https://doi.org/10.3109/17453674.2013.810519
https://doi.org/10.3109/17453674.2013.81...
,1313. Ragab OM, Zayed HS, Abdelaleem EA, Girgis AE. Effect of early treatment with disease-modifying anti-rheumatic drugs and treatment adherence on disease outcome in rheumatoid arthritis patients. The Egyptiam Rheumatologist. 2017;39(2):69-74. https://doi.org/10.1016/j.ejr.2016.11.004
https://doi.org/10.1016/j.ejr.2016.11.00...
. Given the importance of FLS in RA pathology, there is a great need to find a reliable and repeatable source for quick access to them. As we know, the extraction of FLSs from synovium obtained during surgery is time-consuming and prone to contamination with microorganisms as well as needs a lot of materials. So, the establishment of an optimized procedure with low cost and shorter time compared with an enzymatic method is essential. In this study, for comparing the efficiencies of different isolation, we aimed to compare two currently being used FLS isolation methods to find a simple and effective procedure.

Materials and reagents

Dulbecco's modified Eagle medium: nutrient mixture F-12 (DMEM/F12)+PSF (penicillin, streptomycin, and fungizone) (Biosera) (for the digestion mixture), fetal bovine serum (FBS) (Biosera), 0.25% trypsin-EDTA (Biosera), collagenase type VIII (Sigma C-2139), fluorochrome-labeled antibodies, cell culture flasks, dishes and tubes, and surgical blade (NO.17) were used.

METHODS

Patients and tissue samples

Human synovial tissue and synovial fluid (SF) were obtained from eight female patients with RA who underwent total knee arthroplasty in the Department of Orthopedics, Shafa Hospital, Sari, Mazandaran, with an average age of 58.8±9.66 years, ranging from 48 to 77 years. All patients fulfilled the 2010 RA classification criteria1414. Kay J, Upchurch KS. ACR/EULAR 2010 rheumatoid arthritis classification criteria. Rheumatology (Oxford). 2012;51 Suppl 6:vi5-9. https://doi.org/10.1093/rheumatology/kes279
https://doi.org/10.1093/rheumatology/kes...
. This study was approved by the Institutional Medical Ethics Committee of Mazandaran University of Medical Sciences (MAZUMS; IR.MAZUMS.REC.1398.1364), and informed consent was acquired from all patients before surgery.

Cell culture

Synovium

The synovial tissues were carried from the surgery department to the cell culture laboratory in a 50-ml falcon tube containing 15-ml DMEM culture medium with 100 U/ml penicillin, 100 U/ml streptomycin, and 1% amphotericin B as transport media and stored at 4°C before processing. Then, the collected tissues were, respectively, washed with phosphate-buffered saline (PBS) (pH 7.3–7.4), alcohol (75%), and finally with PBS+100 U/ml penicillin, 100 U/ml streptomycin, and 1% amphotericin B, each for a period of 2 min. Tissues were minced into 1 mm×1 mm pieces with the help of sterile BP blade No. 17 in a sterilized Petri dish containing DMEM media and then incubated with collagenase VIII for 2 h at 37°C in a shaker incubator to promote isolation of FLS. After incubation time, cell pellets were harvested by centrifugation at 1000 g for 10 min. Then, pellets were suspended in DMEM supplemented with 10% (vol/vol) heat-inactivated FBS, 100 U/ml penicillin, and 100 U/ml streptomycin and placed in a T-25 flask at 37°C with 5% CO2 in a humidified atmosphere. After 48 h, the medium was changed, and the isolation of cells from synovial tissues was checked every day. The medium was changed twice a week.

Synovial fluid

After collection of SF in a 15-ml falcon tube containing anticoagulant, it was diluted twice with sterile PBS and centrifuged at 400g for 10 min. Harvested pellets were suspended in DMEM with 10% FBS and antibiotics containing 100 U/ml penicillin and 100 mg/ml streptomycin. The cells were placed in T-25 flasks and incubated under the standard condition at 37°C with 5% CO2 with saturated humidity. Following the 48 h incubation, fresh DMEM was added to the cells. The culture medium was refreshed every 3–4 days.

SF and synovial tissue culture processing steps are shown in Figure 1.

Figure 1
Schematic illustration of synovial fluid and synovial tissue culture processing steps.

Identification of FLS by flow cytometry

FLSs at passage 4 were identified by flow cytometry. The purity of FLS was checked for the presence of MLS. The cells were stained with the following antibodies: fluorescein isothiocyanate (FITC)-conjugated anti-CD68, phycoerythrin (PE)-conjugated anti-CD14 antibody, and allophycocyanin (APC)-conjugated anti-CD90.

RESULTS

The results showed that three of eight primary cultures of FLS by enzymatic digestion method were successful. The median time to the presence of the first cells was 15 days, and they reached 70–80% of confluency around 30–35 days of culture. The remaining tissue samples were discarded if any cells were observed after 30 days. In contrast, cell isolation of all SF samples was successful, and the first cells were seen after about 3–4 days of culture. Getting 70–80% of confluency was dependent on the initial volume of SF.

Morphology of FLS cells

The cells were monitored for morphological assessment after the first day of their presence in the T-25 flask. Most of the cell population were spindle-shaped fibroblast, even in passage 0; however, few number of round to spherical shape, stellate-shaped, and epithelioid-shaped cells were also observed, especially in initial passages (Figure 2). Overall, isolated FLSs from synovium showed more similarity in shape in passages 1–3 compared with those from SF. It should be mentioned that the homogeneity of cells was different from one sample to another but all get to purity from passages 3 and 4 onward.

Figure 2
Light microscopic features of synovial fluid and synovial tissue culture. Cell outgrowth from synovial tissue on days 15-25 (a) (100×), a mixture of spindle-shaped and spherical shape fibroblasts with different size in passage 0 from synovial tissue (b) (100×) and synovial fluid (b) (400×).

Percentage of FLS

Isolated synovial cells from synovium and SF were evaluated by flow cytometry at passage 4. We found a similar percentage of positive cells in specific markers in both procedures used after passage 4.

Overall comparison between the two methods is summarized in Table 1.

Table 1
Comparison of important characteristics between two isolation methods.

DISCUSSION

Isolation of primary cell lines is required for researchers to investigate the morphological, functional, and cellular characteristics of a special tissue1515. Patil R, Kale AD, Mane DR, Patil D. Isolation, culture and characterization of primary cell lines of human buccal mucosal fibroblasts: a combination of explant enzamytic technique. J Oral Maxillofac Pathol. 2020;24(1):68-75. https://doi.org/10.4103/jomfp.JOMFP_282_19
https://doi.org/10.4103/jomfp.JOMFP_282_...
,1616. Weigand A, Boos AM, Tasbihi K, Beier JP, Dalton PD, Schrauder M, et al. Selective isolation and characterization of primary cells from normal breast and tumors reveal plasticity of adipose derived stem cells. Breast Cancer Res. 2016;18(1):32. https://doi.org/10.1186/s13058-016-0688-2
https://doi.org/10.1186/s13058-016-0688-...
. Nevertheless, a single-standard protocol has not yet been optimized for the isolation of many cells, especially recently discovered ones. FLSs were recently proven to be key players of observed inflammation in RA context1717. Takeuchi Y, Hirota K, Sakaguchi S. Synovial tissue inflammation mediated by autoimmune T cells. Front Immunol. 2019;10:1989. https://doi.org/10.3389/fimmu.2019.01989
https://doi.org/10.3389/fimmu.2019.01989...
,1818. Chang SK, Gu Z, Brenner MB. Fibroblast-like synoviocytes in inflammatory arthritis pathology: the emerging role of cadherin-11. Immunol Rev. 2010;233(1):256-66. https://doi.org/10.1111/j.0105-2896.2009.00854.x
https://doi.org/10.1111/j.0105-2896.2009...
. These cells locate inside joints in the synovium and are involved in pannus formation, a hallmark pathological change in patients with RA1919. You S, Yoo SA, Choi S, Kim JY, Park SJ, Ji JD, et al. Identification of key regulators for the migration and invasion of rheumatoid synoviocytes through a systems approach. Proc Natl Acad Sci U S A. 2014;111(1):550-5. https://doi.org/10.1073/pnas.1311239111
https://doi.org/10.1073/pnas.1311239111...
2121. Cheng L, Wang Y, Wu R, Ding T, Xue H, Gao C, et al. New insights from single-cell sequencing data: synovial fibroblasts and synovial macrophages in rheumatoid arthritis. Front Immunol. 2021;12:709178. https://doi.org/10.3389/fimmu.2021.709178
https://doi.org/10.3389/fimmu.2021.70917...
. Recent advances in the treatment of rheumatologic disorders have resulted in a reduction of access to synovial tissues by investigators2222. Johnson BK, Goodman SM, Alexiades MM, Figgie MP, Demmer RT, Mandl LA. Patterns and associated risk of perioperative use of anti-tumor necrosis factor in patients with rheumatoid arthritis undergoing total knee replacement. J Rheumatol. 2013;40(5):617-23. https://doi.org/10.3899/jrheum.121171
https://doi.org/10.3899/jrheum.121171...
. Accordingly, it is essential to find a more proximal source of FLSs than the replaced joints during arthroplasty. In this study, we readily extracted adherent fibroblast cells from SF compared with synovial tissues. Also, flow cytometry analysis of the SF-derived cells confirmed the phenotype of FLS cells. We found that FLS does not migrate from all synovial tissues, but FLS extraction from all SF samples was successful. However, extracted cells from SF were more heterogeneous in initial passages than those from synovial tissue, but in the following, they reach to a similar phenotype like synovial tissue-derived cells. These results are in the same direction as those obtained by Stebulis et al. who indicated that FLS isolation from SF can facilitate the study of synovial cells when synovial tissues are not available. They found that both SF- and tissue-derived FLS are the same in functional and cellular measures such as the presence of some specific enzymes and production of inflammatory mediators2323. Stebulis JA, Rossetti RG, Atez FJ, Zurier RB. Fibroblast-like synovial cells derived from synovial fluid. J Rheumatol. 2005;32(2):301-6. PMID: 15693092. Moreover, Ahn et al. reported that there is no significant difference between the invasive capacity of the two FLS types and that both of them are identical in surface markers2424. Ahn JK, Kim H, Lee J, Bae EK, Cha HS, Koh EM. Phenotypic characterization and invasive properties of synovial fluid-derived adherent cells in rheumatoid arthritis. Inflammation. 2008;31 (6):365-71. https://doi.org/10.1007/s10753-008-9087-x
https://doi.org/10.1007/s10753-008-9087-...
. Neidhart et al. demonstrated that SF-derived FLSs mediate cartilage destruction similar to tissue-derived ones2525. Neidhart M, Seemayer CA, Hummel KM, Michel BA, Gay RE, Gay S. Functional characterization of adherent synovial fluid cells in rheumatoid arthritis: destructive potential in vitro and in vivo. Arthritis Rheum. 2003;48(7):1873-80. https://doi.org/10.1002/art.11166
https://doi.org/10.1002/art.11166...
. These reports are in agreement with our results regarding the identical characteristics of extracted FLSs from both methods.

CONCLUSIONS

We found that the extraction of FLS from synovial tissues is not 100%, and FLS isolation from SF is the most convenient and effective method. The use of 500-μl SF is sufficient to get a higher number of FLS with a shorter time and lower cost of cultivation than synovial tissues. These data may support the selection of FLS isolation from SF for downstream analysis.

  • Funding: none.

ACKNOWLEDGMENTS

This work was partially supported by grants from Mazandaran University of Medical Sciences (MAZUMS) (IR.MAZUMS.REC.1398.1364) and Kermanshah University of Medical Sciences (KUMS) (990522).

REFERENCES

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    Guo Q, Wang Y, Xu D, Nossent J, Pavlos NJ, Xu J. Rheumatoid arthritis: pathological mechanisms and modern pharmacologic therapies. Bone Res. 2018;27(6):15. https://doi.org/10.1038/s41413-018-0016-9
    » https://doi.org/10.1038/s41413-018-0016-9
  • 2
    Mellado M, Martínez-Muñoz L, Cascio G, Lucas P, Pablos JL, Rodríguez-Frade JM. T cell migration in rheumatoid arthritis. Front Immunol. 2015;6:384. https://doi.org/10.3389/fimmu.2015.00384
    » https://doi.org/10.3389/fimmu.2015.00384
  • 3
    Yang X, Chang Y, Wei W. Emerging role of targeting macrophages in rheumatoid arthritis: focus on polarization, metabolism and apoptosis. Cell Prolif. 2020;53(7):e12854. https://doi.org/10.1111/cpr.12854
    » https://doi.org/10.1111/cpr.12854
  • 4
    Tu J, Hong W, Zhang P, Wang X, Körner H, Wei W. Ontology and function of fibroblast-like and macrophage-like synoviocytes: how do they talk to each other and can they be targeted for rheumatoid arthritis therapy? Front Immunol. 2018;9:1467. https://doi.org/10.3389/fimmu.2018.01467
    » https://doi.org/10.3389/fimmu.2018.01467
  • 5
    Jung SM, Kim KW, Yang CW, Park SH, Ju JH. Cytokine-mediated bone destruction in rheumatoid arthritis. J Immunol Res. 2014;2014:263625. https://doi.org/10.1155/2014/263625
    » https://doi.org/10.1155/2014/263625
  • 6
    Bartok B, Firestein GS. Fibroblast-like synoviocytes: key effector cells in rheumatoid arthritis. Immunol Rev. 2010;233(1):233-55. https://doi.org/10.1111/j.0105-2896.2009.00859.x
    » https://doi.org/10.1111/j.0105-2896.2009.00859.x
  • 7
    Ostrowska M, Maśliński W, Prochorec-Sobieszek M, Nieciecki M, Sudoł-Szopińska I. Cartilage and bone damage in rheumatoid arthritis. Reumatologia. 2018;56(2):111-20. https://doi.org/10.5114/reum.2018.75523
    » https://doi.org/10.5114/reum.2018.75523
  • 8
    Bustamante MF, Garcia-Carbonell R, Whisenant KD, Guma M. Fibroblast-like synoviocyte metabolism in the pathogenesis of rheumatoid arthritis. Arthritis Res Ther. 2017;19(1):110. https://doi.org/10.1186/s13075-017-1303-3
    » https://doi.org/10.1186/s13075-017-1303-3
  • 9
    Nevius E, Gomes AC, Pereira JP. Inflammatory cell migration in rheumatoid arthritis: a comprehensive review. Clin Rev Allergy Immunol. 2016;51(1):59-78. https://doi.org/10.1007/s12016-015-8520-9
    » https://doi.org/10.1007/s12016-015-8520-9
  • 10
    Lee JK, Choi CH. Total knee arthroplasty in rheumatoid arthritis. Knee Surg Relat Res. 2012;24(1):1-6. https://doi.org/10.5792/ksrr.2012.24.1.1
    » https://doi.org/10.5792/ksrr.2012.24.1.1
  • 11
    Danoff JR, Moss G, Liabaud B, Geller JA. Total knee arthroplasty considerations in rheumatoid arthritis. Autoimmune Dis. 2013;2013:185340. https://doi.org/10.1155/2013/185340
    » https://doi.org/10.1155/2013/185340
  • 12
    Jämsen E, Virta LJ, Hakala M, Kauppi MJ, Malmivaara A, Lehto MUK. The decline in joint replacement surgery in rheumatoid arthritis is associated with a concomitant increase in the intensity of anti-rheumatic therapy: a nationwide register-based study from 1995 through 2010. Acta Orthop. 2013;84(4):331-7. https://doi.org/10.3109/17453674.2013.810519
    » https://doi.org/10.3109/17453674.2013.810519
  • 13
    Ragab OM, Zayed HS, Abdelaleem EA, Girgis AE. Effect of early treatment with disease-modifying anti-rheumatic drugs and treatment adherence on disease outcome in rheumatoid arthritis patients. The Egyptiam Rheumatologist. 2017;39(2):69-74. https://doi.org/10.1016/j.ejr.2016.11.004
    » https://doi.org/10.1016/j.ejr.2016.11.004
  • 14
    Kay J, Upchurch KS. ACR/EULAR 2010 rheumatoid arthritis classification criteria. Rheumatology (Oxford). 2012;51 Suppl 6:vi5-9. https://doi.org/10.1093/rheumatology/kes279
    » https://doi.org/10.1093/rheumatology/kes279
  • 15
    Patil R, Kale AD, Mane DR, Patil D. Isolation, culture and characterization of primary cell lines of human buccal mucosal fibroblasts: a combination of explant enzamytic technique. J Oral Maxillofac Pathol. 2020;24(1):68-75. https://doi.org/10.4103/jomfp.JOMFP_282_19
    » https://doi.org/10.4103/jomfp.JOMFP_282_19
  • 16
    Weigand A, Boos AM, Tasbihi K, Beier JP, Dalton PD, Schrauder M, et al. Selective isolation and characterization of primary cells from normal breast and tumors reveal plasticity of adipose derived stem cells. Breast Cancer Res. 2016;18(1):32. https://doi.org/10.1186/s13058-016-0688-2
    » https://doi.org/10.1186/s13058-016-0688-2
  • 17
    Takeuchi Y, Hirota K, Sakaguchi S. Synovial tissue inflammation mediated by autoimmune T cells. Front Immunol. 2019;10:1989. https://doi.org/10.3389/fimmu.2019.01989
    » https://doi.org/10.3389/fimmu.2019.01989
  • 18
    Chang SK, Gu Z, Brenner MB. Fibroblast-like synoviocytes in inflammatory arthritis pathology: the emerging role of cadherin-11. Immunol Rev. 2010;233(1):256-66. https://doi.org/10.1111/j.0105-2896.2009.00854.x
    » https://doi.org/10.1111/j.0105-2896.2009.00854.x
  • 19
    You S, Yoo SA, Choi S, Kim JY, Park SJ, Ji JD, et al. Identification of key regulators for the migration and invasion of rheumatoid synoviocytes through a systems approach. Proc Natl Acad Sci U S A. 2014;111(1):550-5. https://doi.org/10.1073/pnas.1311239111
    » https://doi.org/10.1073/pnas.1311239111
  • 20
    Nygaard G, Firestein GS. Restoring synovial homeostasis in rheumatoid arthritis by targeting fibroblast-like synoviocytes. Nature Rev Rheumatol. 2020;16(6):316-33. https://doi.org/10.1038/s41584-020-0413-5
    » https://doi.org/10.1038/s41584-020-0413-5
  • 21
    Cheng L, Wang Y, Wu R, Ding T, Xue H, Gao C, et al. New insights from single-cell sequencing data: synovial fibroblasts and synovial macrophages in rheumatoid arthritis. Front Immunol. 2021;12:709178. https://doi.org/10.3389/fimmu.2021.709178
    » https://doi.org/10.3389/fimmu.2021.709178
  • 22
    Johnson BK, Goodman SM, Alexiades MM, Figgie MP, Demmer RT, Mandl LA. Patterns and associated risk of perioperative use of anti-tumor necrosis factor in patients with rheumatoid arthritis undergoing total knee replacement. J Rheumatol. 2013;40(5):617-23. https://doi.org/10.3899/jrheum.121171
    » https://doi.org/10.3899/jrheum.121171
  • 23
    Stebulis JA, Rossetti RG, Atez FJ, Zurier RB. Fibroblast-like synovial cells derived from synovial fluid. J Rheumatol. 2005;32(2):301-6. PMID: 15693092
  • 24
    Ahn JK, Kim H, Lee J, Bae EK, Cha HS, Koh EM. Phenotypic characterization and invasive properties of synovial fluid-derived adherent cells in rheumatoid arthritis. Inflammation. 2008;31 (6):365-71. https://doi.org/10.1007/s10753-008-9087-x
    » https://doi.org/10.1007/s10753-008-9087-x
  • 25
    Neidhart M, Seemayer CA, Hummel KM, Michel BA, Gay RE, Gay S. Functional characterization of adherent synovial fluid cells in rheumatoid arthritis: destructive potential in vitro and in vivo. Arthritis Rheum. 2003;48(7):1873-80. https://doi.org/10.1002/art.11166
    » https://doi.org/10.1002/art.11166

Publication Dates

  • Publication in this collection
    13 Dec 2021
  • Date of issue
    Nov 2021

History

  • Received
    22 Aug 2021
  • Accepted
    29 Aug 2021
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