Chocolate cream labels as novel potential biomaterial? About red herrings in the scientific biomaterial scene

Scientific publications dealing with the biocompatibility analysis of novel potential implant materials are steadily growing (the keyword “biocompatibility” yielded 534,359 publications in PubMed, between august 2018 and 1971). Biocompatibility is a word that is used extensively within biomaterials science since 1971, however, a clear definition of biocompatibility was given by Dave Williams in 2008 [1]. Within these publications the need for the development of new biomaterials for clinical applications with specific requirements are comprehensively described. Often, data are presented which should imply to the reader, that a new biomaterial with physico-chemical properties matching the needs of the surrounding tissue and appropriate cyto- and tissue compatibility was developed. Sometimes far away from the guidelines of the ISO standard (the International Organization for Standardization), statements like perfect biocompatibility are made, though only cytotoxicity tests were performed (without taking the ISO standard into account). As conclusion this material is presented as “holy grain” since the application in the clinic could solve high impact problems in the future of the mankind. Usually, one never hears again from this “ultimate” biomaterials.

The problem of such publications is that although they are selling their cytotoxicity testing according to the ISO standard, this often is in fact not the case. The ISO standards 10993-5 and 10993-12 give a recommendation of sample preparation, cells and test systems which should be applied for the investigation of the cytotoxicity of implant materials [2]. Also, the analysis as well as the evaluation of the applied test systems are clearly described in order to assess the occurrence and the degree of cytotoxic effects. Finally, it is completely clear whether a material is cytotoxic or not.

The journal should not accept such publications unless the authors explain why they have deviated from the ISO standard. Nevertheless, in such cases the term “cytotoxicity” cannot be used. The cytotoxicity of a material is clearly defined in the ISO standard 10993-5. The standard comprises a broad range of suggestions of biological test systems which have to be applied in order to investigate cytotoxic effects of materials on mammalian cells. Data which are generated with missing expertise should no place be given in a scientific journal. Such publications often offer no useful conclusion and are not more than wasting time.

With the following example we would like to drive such an absurd approach to the top. Here, the label material of a chocolate cream was used as a substrate material in order to investigate its “cytotoxicity”.

The label was removed from the glass and sliced in 1×3 cm stripes arbitrarily. The stripes were washed in 70% ethanol for 2.5 h and desiccated under a laminar flow hood. Subsequently, 1.25×10⁶ HCT-116 cells (human colon carcinoma cell line) per well (6-well plate, TCP, Merck) were seeded on the stripes incubated with cell culture medium (DMEM, 10% bovine serum, Biochrom). After three days of incubation cells were stained using the viability dye toluidine blue. Microscopic images were taken using phase contrast microscopy and showed that HCT-116 cells grew excellently on TCP. Also on the label material the cells were of comparable density and had already started to form small colonies (see Fig. 1).

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Fig.1

Single cells and cell colonies (indicated by arrows) on label material and tissue culture plastic.

According to these data one might assume, that the label material is not “cytotoxic” and thereby could be envisioned as implant material for different applications. But what to do with this data now? Does that mean that we should collect all label materials as prospective implant materials? No, that’s not what it means.

The test described above contradicts all recommendations of the ISO standard. Neither suggested cells and test systems were used nor the suggested analysis of the results with subsequent grading of the cytotoxic effects was performed. In addition, an investigation of the bacterial contamination is missing which is clearly recommended in the ISO standard because of the unsterile, only disinfected substrate material which was applied (according to the ISO standard 10993-1 the biological evaluation of candidate implant materials have to be performed in their latter mostly sterile form of application).

With the help of the data presented here, our intention was to invite the reader of scientific publications to look critically at data and studies to questioning the meaningfulness and correctness of data which are presented, the applied test systems and candidate materials which were selected for the analysis and the expertise of the investigators. Beyond that, please questioning whether the conclusion of the story, which was drawn, is really consistent. Finally, we should always have in mind that cytotoxicity testing is performed to protect patients from potential biological risks when using implant materials and should therefore place the highest demands on compliance with scientific, technical and approval requirements.