University of Edinburgh, Large Animal Hospital, Easter Bush Veterinary Centre, Midlothian, UK.
Equine Vet J. 2012 May;44(3):355-60. doi: 10.1111/j.2042-3306.2011.00437.x. Epub 2011 Sep 1.
To investigate the effects of commonly used antibiotics on cell viability and gene expression of equine bone marrow-derived mesenchymal stromal cells (MSC) in vitro.
Bone marrow-derived MSC were cultured in media containing gentamicin, amikacin, penicillin, enrofloxacin or ceftiofur at concentrations of 50, 100, 200 and 500 µg/ml. The alamarBlue fluorescence assay was used to assess cell viability over 48 h. After 5 days the cells were released and lysed prior to RNA extraction and reverse transcription. RNA levels were assessed using spectrophotometry and quantitative PCR was used to analyse gene expression of COL1A2, COL5A1, TNC, TNFα, CASP3, BCl2 and TGFβR2 relative to the reference gene GAPDH.
Enrofloxacin produced a significant concentration-dependent reduction in cell viability at 200 µg/ml and higher concentrations (P = 0.009). Amikacin significantly reduced cell viability at 500 µg/ml (P = 0.002). Penicillin had no effect on cell viability at the concentrations tested (P = 0.18). Gentamicin and ceftiofur showed some interaction with the assay but had no overall effect on cell viability. At 500 µg/ml gentamicin (P<0.001), amikacin (P = 0.03), enrofloxacin (P<0.001) and ceftiofur (P<0.001) caused significant reductions in RNA levels. At 50 µg/ml gentamicin (P<0.001) and amikacin (P = 0.019) reduced BCl2 expression. Enrofloxacin produced a significant increase in COL1A2 expression (P<0.001).
Enrofloxacin reduced MSC viability in vitro and may require cautious use in clinical situations. Penicillin has minimal detrimental effects on MSC in vitro and its use in conjunction with MSC at implantation appears safe. Further work is needed to fully investigate the effects of gentamicin, amikacin and ceftiofur.
Clinicians using local antibiotic administration should consider the potential for local toxicity as well as the need for effective concentrations of the antibiotic.
研究常用抗生素对体外培养的马骨髓间充质基质细胞（MSC）活力和基因表达的影响。
将骨髓来源的 MSC 在含庆大霉素、阿米卡星、青霉素、恩诺沙星或头孢噻呋的培养基中培养，浓度分别为 50、100、200 和 500μg/ml。使用 alamarBlue 荧光法评估 48 小时内的细胞活力。5 天后，细胞释放并裂解，然后提取 RNA 并进行逆转录。使用分光光度法评估 RNA 水平，并使用定量 PCR 分析 COL1A2、COL5A1、TNC、TNFα、CASP3、BCl2 和 TGFβR2 相对于参考基因 GAPDH 的基因表达。
恩诺沙星在 200μg/ml 及更高浓度时表现出显著的浓度依赖性细胞活力降低（P=0.009）。阿米卡星在 500μg/ml 时显著降低细胞活力（P=0.002）。青霉素在测试浓度下对细胞活力无影响（P=0.18）。庆大霉素和头孢噻呋与测定有一定的相互作用，但对细胞活力无总体影响。在 500μg/ml 时，庆大霉素（P<0.001）、阿米卡星（P=0.03）、恩诺沙星（P<0.001）和头孢噻呋（P<0.001）导致 RNA 水平显著降低。在 50μg/ml 时，庆大霉素（P<0.001）和阿米卡星（P=0.019）降低 BCl2 表达。恩诺沙星使 COL1A2 表达显著增加（P<0.001）。
恩诺沙星降低了 MSC 的体外活力，在临床情况下使用时需要谨慎。青霉素对 MSC 的体外作用最小，在植入时与 MSC 联合使用似乎是安全的。需要进一步研究以充分调查庆大霉素、阿米卡星和头孢噻呋的影响。
使用局部抗生素给药的临床医生应考虑局部毒性的可能性以及抗生素有效浓度的必要性。
