First quantification of alpha‐Gal epitope in current glutaraldehyde‐fixed heart valve bioprostheses

Abstract

Background: Glutaraldehyde fixation does not guarantee complete tissue biocompatibility in current clinical bioprosthetic heart valves (BHVs). Particularly, circulating anti-aGal human antibodies increase significantly from just 10 days after a BHV implantation. The inactivation of such epitope should be mandatory to meet the requirements for a perspectively safe clinical application; nevertheless, its quantitative assessment in commercially available BHVs has never been carried out. Methods: In this investigation, seven different models of BHVs were tested. The number of epitopes was determined with reference to a standard aGal source by an ELISA test. The presence of xenoantigen was subsequently confirmed by immunofluorescence analysis. Porcine tissue, knockout for the aGal epitopes, was used as negative control. Results: Epic TM valve was the only model among those tested, in which the aGal antigen appeared to be completely shielded. Composite Trifecta TM valve exhibited conflicting results: cusps of bovine pericardial tissue were devoid of reactive aGal epitopes, while the stent cover strip of porcine pericardium still maintained 30% of active antigens originally present in native tissue. All other tested BHVs express an aGal amount not significantly different from that exhibited by porcine Mosaic â valve (5.2 AE 0.6 9 10 10 each 10 mg of tissue). Conclusions: For the first time, the quantitative evaluation of the aGal epitope in heart valve bioprostheses, already in clinical practice for about 40 yrs, was finally determined. Such quantification might provide indications of biocompatibility relevant for the selection of bioprosthetic devices and an increase in the confidence of the patient. It might become a major quality control tool in the production and redirection of future investigation in the quest for aGal-free long-lasting substitutes.

Transdentinal Cytotoxicity of Carbodiimide (EDC) and Glutaraldehyde on Odontoblast-like Cells

“…42 GA is used in many different processes as a fixative, cross-linking, and disinfecting agent. As is the case for EDC, GA participates in the construction of bioprosthetic heart valves, 26,43 inhibits proteases, 31 and modifies gelatins and other materials and tissues. 26,44,45 In dental treatments, GA has been used as a desensitizer that reacts with plasma proteins in dentin fluid to precipitate them, blocking dentin tubules.…”

Biocompatibility Evaluation Criteria for Novel Xenograft Materials: Distribution and Quantification of Remnant Nucleic Acid and Alpha-Gal Epitope

“…As previously reported by our group [35], the GLU fixation procedure performed on the TC and EP samples is responsible for aspecific interaction between the epitopes and the antibody used for their detection. In order to avoid such confounding results, previously GLU-treated, TC and EP tissues were subjected to a "GLU inactivation procedures" actually under patent (European Application no.…”

“…Noteworthy, the immune response against these xenogeneic residues has been recognized, together with the presence of nucleic acids materials, as an additional factor leading to tissue mineralization [32]. The amount of available and reactive alpha-Gal epitope evaluated in EP patch resulted in our investigation to be 15% higher than in commercially bovine heart valve substitute [35]. In a hypothetical biocompatibility classification, this assessment puts the EP tissue well behind the bovine and porcine pericardium for high pressure cardiovascular and orthopaedic purposes.…”

“…In fact the low antigenic profile of TC is determined by the complete absence of the alpha-Gal epitope as confirmed by our performed immunofluorescence analysis ( Figure 1) and ELISA test (Table 3). While the lack of such antigen ensures a better biocompatibility, the widespread presence of the nucleic acids material in the GLU-treated biomaterial, will expose the same tissue to a high probability of calcification occurrences similarly to the current biological heart valve substitutes [42] (even if these latter maintain a considerable amount of alpha-Gal epitope able to react with circulating human antibodies [35]). The low amount of DNA content revealed in the tissue (Figure 2) is justified by its low cell density which characterizing this type of matrix.…”

“…Over the last four years, our group has developed a solid experience on assessing the biocompatibility of xenogeneic tissues, based on the quantification and distribution of such xenogeneic epitope [26,31,35]. Accordingly we have investigated the content and distribution of alphaGal as well as of nucleic acid in a variety of novel xenogeneic tissues used or intended for use in clinical practice, that have shown very promising outcomes in different areas of application.…”