Real-time detection of antibiotics cytotoxicity in rabbit periosteal cells using microfluidic devices with comparison to conventional culture assays.
BMC Musculoskelet Disord
; 20(1): 339, 2019 Jul 26.
Article
em En
| MEDLINE
| ID: mdl-31349830
ABSTRACT
BACKGROUND:
Local antibiotic application has been widely used in orthopedic surgery. The dose-related toxicity of antibiotics towards periosteal tissues and resulting effects on osteogenic expression are yet to be studied.METHODS:
Periosteal cells harvested from the medial tibia of New Zealand White rabbits were used. A seeding density of 5 × 103 cells/cm2 was determined to be optimal for testing in the pilot study; the cells were cultured in xCELLigence 96-well plates. Microfluidic impedance analyzers were used to monitor cellular proliferation in microfluidic culture systems with exposure to three different concentrations (10 µg/mL, 100 µg/mL, and 1000 µg/mL) of cefazolin, ciprofloxacin, and vancomycin, respectively. The correlation of cell index at day 7 with optical density values from WST-1 assays using conventional cultures was evaluated by calculating the Pearson's coefficient. RNA analysis was performed to investigate the expression of osteogenic markers in the cultured cells, including core-binding factor alpha 1 (Cbfa1), osteopontin (OPN), and osteopontin promoter (OPNp), relative to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as the endogenous control.RESULTS:
A significant dose-related inhibition of cell index was found for all the 3 antibiotics, whereas the WST-1 assays showed a significant dose-related inhibition of cellular proliferation only at a high dose of cefazolin (1000 µg/mL) and medium-to-high dose of ciprofloxacin (100 µg/mL and 1000 µg/mL). Pearson's coefficient analysis indicated a high correlation between the cell index and optical density values of WST-1 assays only for medium and high doses of ciprofloxacin (100 µg/mL and 1000 µg/mL); a moderate correlation was seen for cefazolin, and a low dose of ciprofloxacin (10 µg/mL). RNA analysis confirmed significant dose-related inhibition of cfba1, OPN, and OPNp expression by all three antibiotics.CONCLUSION:
With optimal seeding amounts, rabbit periosteal cells can be dynamically monitored in the xCELLigence microfluidic system. Dose-related inhibition of cellular proliferation and osteogenic expression was found after exposure to cefazolin and ciprofloxacin. By providing real-time detection and exhibiting comparable correlation, microfluidic impedance-based analyzer is a feasible alternative to the conventional WST-1 assays.Palavras-chave
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Base de dados:
MEDLINE
Assunto principal:
Osteogênese
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Periósteo
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Testes de Toxicidade Aguda
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Dispositivos Lab-On-A-Chip
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Antibacterianos
Idioma:
En
Ano de publicação:
2019
Tipo de documento:
Article