Test conditions can significantly affect the results of in vitro cytotoxicity testing of degradable metallic biomaterials

Abstract

In vitro cytotoxicity testing is an indispensable part of the development of new biomaterials. However, the standard ISO 10993-5 enables variability in the testing conditions, which makes the results of the test incomparable. We studied the influence of media composition on the results of the cytotoxicity test. Solutions of ZnCl2 served as simulated extracts and we also used extracts of three types of Zn-based and Mg-based degradable metals. We incubated the cells with the solutions prepared in two types of media with two concentrations of serum (5 and 10%). We compared the toxic effect of the extracts on L929 murine fibroblast-derived cell line, which is recommended by ISO standard and on “osteoblast-like cells” U-2 OS. We also compared two methods of exposition: solutions were added either to a sub-confluent layer or to the cell suspension. We evaluated the metabolic activity of the cells using the resazurin test. We found out that in vitro cytotoxicity is dramatically influenced by the concentration of serum and by the type of the medium as well as by the type of exposition and type of cells. Therefore, when performing in vitro cytotoxicity testing of biomaterials, the authors should carefully specify the conditions of the test and comparison of different studies should be carried out with caution.

Assessment of the Toxicity of Biocompatible Materials Supporting Bone Regeneration: Impact of the Type of Assay and Used Controls

“…By showing only absorbance or fluorescence values [15,17,18], the results cannot be fully related to other data found in the literature. To fully assess the effects caused by a material or extract, it is necessary to use a control, e.g., cells that have not been in contact with the composite or extract [13,19]; also, the physiological state of the used cell model should be taken into consideration [20], which, in many published studies for various bio composites, is not counted [7,21,22]. For this purpose, the use of tests with a positive control (e.g., cells treated with Triton X-100 solution for an LDH assay) that will allow the assessment of the condition of the cells during the experiments, is recommended.…”

The evaluation of cytotoxicity and cytokine IL-6 production of root canal sealers with and without the incorporation of simvastatin: an invitro study

“…Cultures were kept in an incubator (BBD 6220; Thermo Fisher Scientific, Waltham, MA, USA) at 37 °C under ambient pressure and 5% CO 2 atmosphere. Cells were used from the 3rd passage till the 20th passage [ 26 ]. Confluent cell monolayers were trypsinized, and the cells that were harvested, were used for cytotoxicity experiments.…”

Surface Engineering Strategies to Enhance the In Situ Performance of Medical Devices Including Atomic Scale Engineering

“…Figure 2 represents the evolution of biomaterial from ancient times to the present era. First generation biomaterials were biologically inert [5]. Second generation biomaterials were termed as bioactive biomaterials that formed bonds with tissues, such as glass ceramic [6].…”

“…Second generation biomaterials were termed as bioactive biomaterials that formed bonds with tissues, such as glass ceramic [6]. Third generation biomaterials are bioresorbable material that becomes absorbed/degraded by the body itself [5]. Biomaterials are broadly classified into metallic, ceramic, polymeric, composite, and nanobiomaterials [7].…”

“…There are intelligent (autonomous) biomaterials with the innate ability to acquire knowledge (inherent), and they are classified into metals, ceramics/natural ceramics, polymers/natural polymers, composites, natural nanomaterials, surface engineered materials, synthetic nanomaterials, and responsive (smart) biomaterials for biomedical applications such as brain-machine interface, nanomedicine, tissue engineering, regenerative medicine, natural interfaces, implants, medical devices, and diagnosis (Figure 3). First generation biomaterials were biologically inert [5]. Second generation biomaterials were termed as bioactive biomaterials that formed bonds with tissues, such as glass ceramic [6].…”