Does Combined Treatment with Tranexamic Acid and Vancomycin Affect Human Chondrocytes In Vitro?

Abstract

Background: The aim of our study was to examine the combined effects of tranexamic acid (TXA) and vancomycin powder (VP) on chondrocytes in vitro. Despite the use of TXA and VP being linked to a reduced risk of extensive postoperative blood loss and periprosthetic joint infections (PJIs) in TKA, the possible cytotoxic side effects on periarticular cell types remain unclear. Methods: Human chondrocytes were harvested from hyaline cartilage and expanded in monolayer culture before being simultaneously exposed to different concentrations of TXA and VP for varying exposure times. Cell viability and proliferation were assessed using an ATP assay and an Annexin 5 assay, respectively, while changes in the relative expression of chondrogenic marker genes were examined using semiquantitative RT-PCR. Results: The simultaneous exposure of chondrocytes to TXA and VP for more than 48 h led to a reduction in both cell viability and proliferation rates. When exposing chondrocytes to the lowest examined concentrations of both TXA (10 mg/mL) and VP (3 mg/mL), the observed effects were delayed until 96 h. However, our study found no dependencies of the observed effects on the concentrations tested. Further, we found no effects on the expression of chondrogenic marker genes. Conclusions: Consequently, limiting the exposure time of chondrocytes to TXA and VP in an in vitro setting to 24 h may be considered safe and could help to further improve the understanding of the safe use of substances in vivo. However, further in vitro research is required to develop a comprehensive understanding of the effects of both VP and TXA on important periarticular cell types in TKA, including chondrocytes, osteocytes, and tenocytes.

Keywords: chondrocytes; osteoarthritis; total joint replacement; toxicity; tranexamic acid; vancomycin powder; viability.

Figure 3

After treatment of chondrocytes with varying concentrations of TXA and VP for different exposure times, treated cultures and untreated controls isolated from five different donors (n = 5) were compared to examine changes in the relative expression of chondrogenic marker genes by semiquantitative RT-PCR. Bar charts represent mean values and standard deviations for the expression of chondrogenic marker genes (a) ACAN, (b) COL2A1, (c) SOX9 and (d) COMP. Primer details can be found in Table 1. Statistically significant differences between the varying TXA and VP concentrations and exposure times were analyzed using an independent t-test or the Mann–Whitney U test in the case of non-normally distributed data. d = days; TXA = tranexamic acid; VP = vancomycin powder; h = hours.

Figure 1

After treatment of chondrocytes with varying concentrations of TXA and VP for different exposure times, cell proliferation and viability were analyzed using ATP assays. Bar charts show mean values and standard deviations from results derived from all five donors. Statistically significant differences between varying exposure times and concentrations used were assessed using either an independent t-test or the Mann–Whitney U test for non-normally distributed data. * Denotes a significant difference (p < 0.05) compared to untreated control samples. TXA = tranexamic acid; VP = vancomycin powder; h = hours.

Figure 2

After treatment of chondrocytes with varying concentrations of TXA and VP for different exposure times, cell apoptosis was evaluated using Annexin 5 assays. After exposure to TXA and VP, chondrocyte cultures were double-stained with 6-carboxyfluorescein diacetate (CFDA) and Annexin 5. While CFDA-stained living cells appear green, Annexin 5-Cy3-stained apoptotic cells appear red. Early apoptotic cells are stained by both CFDA and Annexin 5. Within the figure, representative fluorescence microscopy images are shown (scale bar = 200 μm). CFDA = 6-carboxyfluorescein diacetate; TXA = tranexamic acid; VP = vancomycin powder; h = hours.