The Effect of Different Irrigation Solutions on the Cytotoxicity and Recovery Potential of Human Osteoblast Cells In Vitro

Abstract

Background: Surgeons use various irrigation solutions to minimize the risk of prosthetic joint infection after total joint arthroplasty. The toxicity of these solutions is an important consideration in their use. This study investigates the effect of irrigation solutions Bacitracin, Clorpactin (sodium oxychlorosene), and Irrisept (chlorhexidine) on osteoblast cytotoxicity and proliferation. Methods: Four replicates of 6 conditions at 3 time points (1, 2, and 4 min) were tested: control (normal saline), Bacitracin (33 IU/ml), Clorpactin (0.05%, 0.1%, 0.2%), and Irrisept (0.05% chlorhexidine gluconate). Human osteoblasts were cultured at 37 C and 5% CO 2 until confluent monolayers were obtained. The treatment solution was applied, and cells were washed 3x with warm phosphate-buffered saline and then supplemented with a fresh medium. Phase-contrast images were taken before and after treatment. The cytotoxicity and proliferation of the treated cells was measured for all conditions on day 3 and day 5 after treatment using the alamarBlue assay. Results: All test conditions showed morphological changes to cells after treatment; controls did not. Cells demonstrated curling and detachment. This effect was the worst and permanent with Irrisept, whereas other treatments showed a return to normal morphology after 1 week. All treatments showed increased %alamarBlue reduction after 5 days except Irrisept, which showed decreased reduction. There was no statistically significant time or dose dependence with Clorpactin treatment. Conclusions: Clorpactin and Bacitracin are damaging to human osteoblast cells in vitro as compared with normal saline. This damage is at least partially reversible as shown by morphology and cell viability assay. Irrisept caused more damage than either Clorpactin or Bacitracin, and the damage was not reversible.

Irrigation Solutions Negatively Affect the Viability and Function of Human Fibroblasts: An in vitro Study

“…Liu et al [12] also studied the effects of differing CHG dilutions on fibroblasts, myoblasts, and osteoblasts and concluded that the clinically used concentration of CHG (2%) was toxic to these cells in vitro. Same toxic effect of Irrisept on osteoblasts was shown by Markel, et al [21]. Another past study used CHG at concentrations 5–2,400 times below those used clinically on in vitro fibroblasts as well and found that the CHG totally suppressed cell proliferation [32].…”

Common Wound Irrigation Solutions Produce Different Responses in Infected vs Sterile Host Tissue: Murine Air Pouch Infection Model

“…We previously showed similar effects on human osteoblasts' viability and recovery after an in vitro exposure. Irrisept, Clorpactin, and Bacitracin all demonstrated some toxic effects on the cells; the damage was to some extent reversible with solutions other than Irrisept [ 21 ]. Difference in testing method (in vitro vs in vivo) should be taken into consideration when comparing studies.…”

“…Bacitracin (33 IU/ml) and Clorpactin (sodium oxychlorosene, 0.2% dilution) were freshly prepared as aqueous solutions. The Bacitracin concentration was based on the available solution at our institution and its use in our previous in vitro study on human osteoblasts [ 21 ], while the Clorpactin concentration was based on the recommendation by original solution development [ 10 ]. Irrisept (chlorhexidine gluconate 0.05% in sterile water) and Bactisure (ethanol 1%, acetic acid 0.6%, sodium acetate 0.2%, benzalkonium chloride 0.013%, and water) were from manufactures.…”

“…Our previous in vitro work demonstrated that some irrigation solutions had toxic effects on osteoblasts, fibroblasts, and tenocytes, and cells could either recover over time or had severe, permanent damage [ 21 ]. This in vivo study, approved by the Ascension Animal Committee, applied a validated murine air pouch model for 2 primary goals: to investigate the effectiveness of irrigation solutions used in practice to minimize bacterial infection and to evaluate the extent of tissue damage caused by the solutions in the presence or absence of infection [ 22 ].…”

The Effects of Three Chlorhexidine-Based Mouthwashes on Human Osteoblast-Like SaOS-2 Cells. An In Vitro Study

“…The low viability levels registered in this last work could have been ascribed to the relatively short culture time, masking the regenerative capacity of the cells over time. Therefore, in a recent study investigating the effect of different antiseptic solutions, a longer observation time was selected as in our paper [ 46 ]. The cytotoxic effect of CHX was confirmed also at a low concentration (0.05%), with human osteoblast cells failing to recover over the course of 5 days.…”