Tissue requirements for the application of aortic valve neocuspidization - appropriate pericardium properties and homogeneity?

Abstract

Objective: Aortic valve neocuspidization (AVNeo) using autologous pericardium is a promising technique. Expected advantages are reduced immune response, appropriate biomechanics and lower treatment expenses. Nevertheless, autologous pericardium can be affected by patient's age and comorbidities. Usually, glutaraldehyde (GA) - fixed bovine pericardium is the basic material for aortic valve prostheses, easy available and carefully pre-examined in a standardized fabrication process. Aim of the study is the verification of autologous pericardial tissue homogeneity by analysing tissue thickness, biomechanics and extracellular matrix (ECM) composition.

Methods: Segments of human GA-fixed pericardium selected by the surgeon based on visual criteria for cusp pre-cut and remaining after surgical AV replacement were investigated in comparison to bovine standard tissue treated equivalently. Pericardium sampling was performed at up to three positions of each sutured cusp for histological or biomechanical analysis, according to tissue availability.

Results and conclusions: Human pericardia exhibited a higher heterogeneity in collagen content, density of vessel structures and elastic moduli. Thickness, vessel density and collagen and elastin content differed significantly between the species. In contrast, significant interindividual differences were detected in most properties investigated for human pericardial samples but only for tissue thickness in bovine tissues. Higher heterogeneity of human pericardium, differing vessel and collagen content compared to bovine state-of-the-art material might be detrimental for long term AV functionality or deterioration and have to be intensely investigated in patients follow up after autologous cusp replacement.

Fig. 1

Comparison of human vs. bovine pericardium thickness and elastic moduli. Comparison of pericardia thickness A and interindividual differences for both species B. Mean elastic moduli are shown for human vs. bovine tissues C. No differences of elastic moduli of individual bovine pericardia D and interindividual variations observed comparing human samples D. A, B, C and D: Unpaired Student´s t-test and One-way ANOVA, Tukey’s test: *p < 0.5; D human samples: Kruskal-Wallis test, Dunn´s multiple comparison test: *p < 0.5

Fig. 2

Collagen density and hydroxyproline content of human vs. bovine pericardia. A Comparison of distribution and wavelike structure in picrosiriusred stained bovine pericardia compared to human tissues. B Picrosiriusred positivity reflecting collagen distribution in the histological sections is quantified in bovine compared to human pericardium. Coefficient of variation is plotted to visualize heterogeneity of human an bovine individual pericardia C Interindividual comparison in picrosieriusred collagen density of single human and bovine pericardium in relation the section area. D Comparison of hydroxyproline levels according to pericardium weight in human vs. bovine pericardia D and HYP contents of individual human tissues. Unpaired Student´s t-test, One-way ANOVA, Tukey’s test: # 0.05 ≤ p < 0.1; *p < 0.5; ****p < 0.0001. D: Kruskal-Wallis test, Dunn´s multiple comparison test: # 0.05 ≤ p < 0.1

Fig. 3

HE staining based density and heterogeneity of vessels in human vs. bovine pericardial matrix. A Vessels in human and bovine pericardia partially containing erythrocytes (purple). B Vessel count per specimen histological section area is plotted in human and bovine pericardia. C Differences between individual human and individual bovine pericardia samples are visualized. B and C: Unpaired Student´s t-test and One-way ANOVA, Tukey´s test: # 0.05 ≤ p < 0.1; ***p < 0.001

Fig. 4

Quantification of elastin content in histological sections after elastica staining. A Elastic fibers embedded in the collagen matrix are visualized in dark purple. B Comparison of mean and coefficient of variation of human vs. bovine pericardia C Interindividual differences in elastin content comparing individual human pericardia or bovine tissues. B: Unpaired Student´s t-test and One-way ANOVA, Tukey´s test: # 0.05 ≤ p < 0.1; ****p < 0.0001

Fig. 5

Rate and heterogeneity of mesothelial covering visualized by IHC vimentin staining; each point of mesothelial covering disruption is marked by an arrow. A Each sub-region of human pericardial sample 5 (1–10) is shown revealing a range from complete covering (sample 2) to absence of mesothelial cells (sample 4). Samples 2 and 8 are given enlarged in B. Mesothelial cell covering rate is shown comparing different areas of one human pericardium remaining after aortic valve replacement. C: Kruskal-Wallis test, Dunn´s multiple comparison test: *p < 0.5