ASO Author Reflections: Technical Insights into Inferior Vena Cava Reconstruction with Tubularized Bovine Pericardium.

Many different grafting materials have been proposed in myringoplasty. The aim of this study was to evaluate the results obtained in transmeatal underlay myringoplasty using bovine and equine pericardium. The results were compared with those obtained by using autologous temporalis fascia. The study group consisted of 52 patients with tympanic perforation. Twenty-nine patients were randomly selected for treatment with bovine pericardium and 23 for equine pericardium. A group of 14 patients was treated with autologous temporalis fascia. Closure of the perforation was achieved in 19 of 29 patients (66%) treated with bovine pericardium, in 19 of 23 (83%) treated with equine pericardium, and in 13 of 14 (93%) treated with autologous fascia. The best functional results in patients who gained closure of the perforation were obtained by means of equine pericardium. The overall long-term tympanic closure rate demonstrates that equine pericardium has a greater take rate than bovine pericardium. The results obtained are inferior to those obtained with autologous fascia, but this technique is less aggressive. The higher success rate with equine pericardium may be due to the fact that it is thinner and easier to handle and model than bovine pericardium.

This study comparatively evaluates the characteristics of glutaraldehyde-treated acellular bovine and donkey pericardium using histological and electronic microscopic observation techniques, shrinkage temperature, and mechanical properties, as well as determining calcium and phosphorus content at 4 and 8 weeks after the subcutaneous implantation of donkey and bovine pericardium in Wistar rats. Donkey pericardium was significantly thinner compared with bovine pericardium (1.622 ± 0.161 mm vs. 4.027 ± 0.401 mm, P < 0.0001) and was associated with significantly greater tensile strength (14.21 ± 3.81 MPa vs. 3.78 ± 1.20 MPa, P = 0.001) and elastic modulus (81.67 ± 20.41 MPa vs. 21.67 ± 11.69 MPa, P < 0.0001) over bovine pericardium. Shrinkage temperature of donkey pericardium was similar to that of bovine pericardium (87.43 ± 0.55°C vs. 87.50 ± 0.36°C, P = 0.810). No differences between groups were observed for maximum load (donkey: 21.64 ± 7.02 KN/m vs. bovine: 15.05 ± 4.50 KN/m, P = 0.082) and tear strength (donkey: 11.54 ± 5.33 MPa vs. bovine: 10.69 ± 3.77 MPa, P = 0.757). Calcium content was significantly lower in donkey pericardium compared with bovine pericardium at 4 weeks (690.15 ± 191.27 µg/g vs. 1381.73 ± 62.52 µg/g, P = 0.001) and 8 weeks (205.24 ± 62.40 µg/g vs. 910.48 ± 398.29 µg/g, P = 0.037). This preliminary study has confirmed that glutaraldehyde-tanned donkey pericardium, demonstrating reduced calcification and increased tensile strength, may provide a suitable bioprosthetic valve substitute.

Objective: To validate the hypothesis that camel pericardium could be more protected than bovine pericardium against calcification process according to the huge difference in their respective lifestyle and lifetime. Methods: Glutaraldehyde (GA) fixed bovine and camel pericardium samples (BP and CP respectively) were both implanted in 30 New Zealand white rats (2 BP and 2 CP matched specimens in each animal) and explanted after 60 days. Unimplanted GA-fixed samples of both species served as control. Matched implanted samples and unimplanted samples were randomly submitted to elemental analysis by spectroscopy, phospholipid extraction, macroscopic and X-ray examination and histology. Results: At 60 days, calcium and phosphorus content were respectively 9.54% ± 3.1% and 4.79% ± 1.4% of tissue dry weight in BP, and 12.52% ± 2.7% and 6.14% ± 1.3% of tissue dry weight in CP (ns). In X-ray analysis, the calcification score was 1.28 ± 0.45 and 2.14 ± 0.98 in BP and CP samples respectively without significant difference (p < 0.08). In histology, calcifications were lower in BP than in CP: 1.37 ± 0.85 vs 2.28 ± 0.83 (ns); collagen fibers were better conserved in BP than in CP: 2.4 ± 0.48 vs 1.87 ± 0.78 (ns), and less disoriented: 25% vs 62% (ns). In unimplanted samples, there was a higher but not significant rate of extracted lipids in CP: 5.7 ± 1.8 vs 9.5 ± 3.8 nanomoles in PS fraction and 11.3 ± 3.7 vs 19 ± 7.7 nanomoles in total fatty acids, in BP and CP samples respectively. All results were in conjunction and demonstrated a higher but not significant rate of mineralization in camel pericardium after implantation, which could be related to a higher but not significant basic rate of phospholipid and fatty acids. Conclusion: This experiment study in a subcutaneous rat model has failed to valid our hypothesis. Because the differences observed between bovine and camel pericardium did not reach the significance, at the best, there is no difference between both species and at the worst, camel pericardium has a higher rate of the phosphatidylserine fraction of phospholipid, and is more sensitive and prompt to calcification.

Surgical site infection (SSI) and polypropylene mesh (PPM) infections are recurrent problems in abdominal hernia surgeries, highlighting the need for a new antimicrobial material for surgical repair. The aim of this study was to evaluate the in vivo antimicrobial effect of a new biological mesh made of decellularized bovine pericardium (BP), added with vancomycin (VAN) or silver nanoparticles (AgNPs), as prevention for SSI. Thirty-five Wistar rats were divided into four groups: BP C+ (n=9) with BP without additions; PP C+ (n=8) with PPM; BP AgNPs (n=9) with BP added with silver nanoparticles; and BP VAN (n=9) with BP added with vancomycin. The 1 cm² meshes were stitched into the muscle fascia under the subcutaneous tissue of the rats' backs, followed by inoculation with methicillin-resistant Staphylococcus aureus. The animals were observed for 7 days, with subsequent euthanasia, and histological and bacteriological analysis. The BP VAN group had better infection control compared to the PP C+ and BP AgNPs groups (1x10¹ vs. 1.4x10³CFU/g, p=0.0303; 1x10¹ vs. 1.5x104CFU/g, p<0.0001, respectively). BP AgNPs showed less bacterial reduction compared to BP C+ (p=0.042). In the histological analysis, there was a mild inflammatory reaction in BP VAN, moderate in BP C+, and intense in PP C+ and BP AgNPs. BP added with vancomycin showed promising antimicrobial action, while the use of silver nanoparticles did not demonstrate efficacy in this study.

Glutaraldehyde (GA) crosslinked bovine or porcine pericardium tissues exhibit high cell toxicity and calcification in the construction of bioprosthetic valves, which accelerate the failure of valve leaflets and motivate the exploration for alternatives. Polyphenols, including curcumin, procyanidin and quercetin, etc, have showed great calcification inhibition potential in crosslinking collagen and elastin scaffolds. Herein, we developed an innovative phenolic fixing technique by using curcumin as the crosslinking reagent for valvular materials. X-ray photoelectron spectroscopy and Fourier transform infrared spectrometry assessments confirmed the hydrogen bond between curcumin and acellular bovine pericardium. Importantly, the calcification inhibition capability of the curcumin-crosslinked bovine pericardium was proved by the dramatically reduced Ca2+ content in the curcumin-fixed group in in vitro assay, a juvenile rat subcutaneous implants model, as well as an osteogenic differentiation model. In addition, the results showed that the curcumin-fixed bovine pericardium exhibited better performance in the areas of mechanical performance, hemocompatibility and cytocompatibility, in comparison with the GA group and the commercialized product. In summary, we demonstrated that curcumin was a feasible crosslinking reagent to fix acellular bovine pericardium, which showed great potential for biomedical applications, particularly in cardiovascular biomaterials with calcification inhibition capacity.

A Biomechanical and Microstructural Analysis of Bovine and Porcine Pericardium for Use in Bioprosthetic Heart Valves

Freeze-drying and gamma irradiation are the techniques widely use in tissue banking for preservation and sterilization of tissue grafts respectively. However, the effect of these techniques on biomechanical properties of bovine pericardium is poorly known. A total of 300 strips of bovine pericardium each measured 4 cm x 1 cm were used in this study to evaluate the effect of freeze-drying on biomechanical properties of fresh bovine pericardium and the effect of gamma irradiation on biomechanical properties of freeze-dried bovine pericardium. The strips were divided into three equal groups, which consist of 100 strips each group. The three groups were fresh bovine pericardium, freeze-dried bovine pericardium and irradiated freeze-dried bovine pericardium. The biomechanical properties of the pericardial strips were measured by a computer controlled instron tensiometer while the strips thickness was measured by Mitutoyo thickness gauge. The results of the study revealed that freeze-drying has no significant (p > 0.05) effect on the tensile strength, Young's modulus (stiffness) and elongation rate of fresh bovine pericardium. Irradiation with 25 kGy gamma rays caused significant decreased in the tensile strength, Young's modulus and elongation rate of the freeze-dried pericardium. However, gamma irradiation has no significant effect on the thickness of freeze-dried bovine pericardium, while freeze-drying caused significant decreased in the thickness of the fresh bovine pericardium. The outcome of this study demonstrated that freeze-drying has no significant effect on the biomechanical properties of fresh bovine pericardium, and gamma irradiation caused significant effect on the biomechanical properties of freeze-dried bovine pericardium.

Evaluation of transcatheter heart valve biomaterials: Biomechanical characterization of bovine and porcine pericardium.

Bioprosthetic heart valves (BHV) fabricated from glutaraldehyde pretreated heterograft materials, porcine aortic valves or bovine pericardium (BP), are widely used in cardiac surgery. BHV progressively fail in clinical use due to structural degeneration. Previously we reported that dityrosine, an oxidized amino acid, was present in failed clinical BP-BHV explants; unimplanted BP had no detectable dityrosine. In the same studies BP were demonstrated in vitro to be susceptible to oxidative damage, that could be mitigated with BP covalently modified with the antioxidant, 3-(4-hydroxy-3,5-di-tert-butylphenyl)propyl amine (DBP). The present studies compared in rat subdermal implants glutaraldehyde pretreated BP to BP modified with either DBP or the chemical reactions used to link DBP. All BP explants regardless of DBP demonstrated reduced hydroxyproline and increased digestibility by collagenase. However, the DBP-BP explants showed significant inhibition of reduced explant shrink temperatures (an index of crosslinking) as compared with control BP. Significant mitigation of calcification was observed in both the BP-DBP and chemically modified explants as compared with BP. Dityrosine was not detectable in the 90 day explants. It is concluded that rat subdermal BP implants undergo both calcific and noncalcific structural degeneration, but without the formation of dityrosine, unlike clinical BP explants.

Glutaraldehyde-stabilized bovine pericardium is used for clinical application since 1970s because of its desirable features such as less immunogenicity and acceptable durability. However, a propensity for calcification is reported on account of glutaraldehyde treatment. In this study, commercially available glutaraldehyde cross-linked bovine pericardium was evaluated for its in vitro cytotoxic effect, macrophage activation, and in vivo toxic response in comparison to decellularized bovine pericardium. Glutaraldehyde-treated bovine pericardium and its extract were observed to be cytotoxic and it also caused significant inflammatory cytokine release from activated macrophages. Significant antibody response, calcification response, necrotic, and inflammatory response were noticed in glutaraldehyde-treated bovine pericardium in comparison to decellularized bovine pericardium in a rat subcutaneous implantation model. Glutaraldehyde-treated bovine pericardium also failed in acute systemic toxicity testing and intracutaneous irritation testing as per ISO 10993. With respect to healing and implant remodeling, total lack of host tissue incorporation and angiogenesis was noticed in glutaraldehyde-treated bovine pericardium compared to excellent host fibroblast incorporation and angiogenesis within the implant in decellularized bovine pericardium. In conclusion, using in vitro and in vivo techniques, this study has demonstrated that glutaraldehyde-treated bovine pericardium elicits toxic response compared to decellularized bovine pericardium which is not congenial for long-term implant performance.

Inhibition of advanced glycation end product formation and serum protein infiltration in bioprosthetic heart valve leaflets: Investigations of anti-glycation agents and anticalcification interactions with ethanol pretreatment

Introduction: Bioprosthetic heart valves (BHV) fabricated from glutaraldehyde treated bovine pericardium (BP) have limited durability due to structural valve degeneration (SVD) resulting from both calcification and advanced glycation end product (AGE) deposition. Hypothesis: We investigated the hypothesis that PYR modification of BHV leaflets mitigates AGE-serum protein uptake, oxidation, and calcification mechanisms in model studies, both in vitro and in vivo. Methods: To model AGE accumulation in BHV, we used well-established in vitro and in vivo models. AGE and serum albumin (SA) uptake in BP were quantitated per immunohistochemistry (IHC) staining intensity. PYR uptake was quantified using a novel fluorescent assay. BP collagen was examined by second-harmonic generation (SHG) microscopy. Calcification studies investigated the activity of BP alkaline phosphatase (ALP), and BP calcium levels were quantitated using a colorimetric assay. In vivo studies used a juvenile rat subdermal BP implantation model with PYR oral administration or BP-PYR modification. Endpoints were calcification, SA, and AGE accumulation per IHC and ALP activity per enzyme histochemistry. Results: PYR modification of BP resulted in high-affinity binding of PYR with 2.0±0.2 μg/mg of PYR binding to BP that was stable for 28 days in physiological conditions. Modification had an inhibitory effect on SA uptake in vitro with 60±5% (p&lt;0.001) reduction in IHC staining intensity; PYR modification of BP resulted in increased protection from oxidative damage in vitro with collagen structure preservation (by SHG) after ten days, while control tissues were degraded. After 28-day of in vivo subdermal BP implantation in rats, a significant 70±5% (p&lt;0.001) decrease and 80±5% (p&lt;0.001) decrease in SA IHC staining and AGE staining, respectively, as well as ALP activity was documented in BP-PYR. Quantitative analysis of calcium content demonstrated a reduction from 108±5 μg/mg in control to 77±5 μg/mg in BP-PYR explants (p&lt;0.001). Conclusions: Our study demonstrated that PYR mitigates SVD mechanisms in vitro and in vivo, limiting both AGE-serum protein accumulation and calcification with associated inhibition of ALP.

In Vivo Evaluation of a Decellularized Pericardium-polymer Biohybrid Composite for Use as a Cardiovascular Tissue Substitute

Bovine pericardium (BP) exhibits distinct biochemical and biomechanical properties that are dominant by the structural protein collagen. The enzymatic degradation of collagen molecules is critical for in vivo incorporation and remodeling of BP in tissue engineering applications. A non-destructive method for monitoring BP during degradation would provide a valuable tool for quantifying functional changes initially in vitro and ultimately in vivo. In this study, we demonstrated the sensitivity of multi-spectral fluorescence lifetime imaging system (ms-FLIm) developed by our group to collagen content and compressive modulus of BP during collagenase degradation. A pairwise study was performed using bacterial collagenase to partially digest BP. We measured the biomaterials properties with ms-FLIm and destructive conventional measurements including collagen assay, compressive test and histology. A single factor study design was utilized. Test group samples were digested by bacterial collagenase for 0, 8, 16 and 24 hours, while control group samples were prepared in the Hank’s balanced salt solution to control for time in solution. Statistical analysis was performed using the Kendall τB correlation test. The results demonstrate that fluorescence parameters measured by ms-FLIm are significantly correlated with collagen content and compressive modulus (|τB| > 0.45, p < 0.05). Based on these findings, we aim to predict BP’s collagen content and mechanical properties using fluorescence metrics, and ultimately apply ms-FLIm for non-destructively monitoring of in vivo remodeling of BP.

Outcomes of carotid endarterectomy in the Vascular Quality Initiative based on patch type