An Assessment of Blood Vessel Remodeling of Nanofibrous Poly(ε-Caprolactone) Vascular Grafts in a Rat Animal Model.

The development of an ideal vascular prosthesis represents an important challenge in terms of the treatment of cardiovascular diseases with respect to which new materials are being considered that have produced promising results following testing in animal models. This study focuses on nanofibrous polycaprolactone-based grafts assessed by means of histological techniques 10 days and 6 months following suturing as a replacement for the rat aorta. A novel stereological approach for the assessment of cellular distribution within the graft thickness was developed. The cellularization of the thickness of the graft was found to be homogeneous after 10 days and to have changed after 6 months, at which time the majority of cells was discovered in the inner layer where the regeneration of the vessel wall was found to have occurred. Six months following implantation, the endothelialization of the graft lumen was complete, and no vasa vasorum were found to be present. Newly formed tissue resembling native elastic arteries with concentric layers composed of smooth muscle cells, collagen, and elastin was found in the implanted polycaprolactone-based grafts. Moreover, the inner layer of the graft was seen to have developed structural similarities to the regular aortic wall. The grafts appeared to be well tolerated, and no severe adverse reaction was recorded with the exception of one case of cartilaginous metaplasia close to the junctional suture.

Using virtual microscopy for the development of sampling strategies in quantitative histology and design-based stereology.

Only a fraction of specimens under study are usually selected for quantification in histology. Multilevel sampling or tissue probes, slides and fields of view (FOVs) in the regions of interest (ROIs) are required. In general, all parts of the organs under study should be given the same probability to be taken into account; that is, the sampling should be unbiased on all levels. The objective of our study was to provide an overview of the use of virtual microscopy in the context of developing sampling strategies of FOVs for stereological quantification. We elaborated this idea on 18 examples from multiple fields of histology, including quantification of extracellular matrix and muscle tissue, quantification of organ and tumour microvessels and tumour-infiltrating lymphocytes, assessing osseointegration of bone implants, healing of intestine anastomoses and osteochondral defects, counting brain neurons, counting nuclei in vitro cell cultures and others. We provided practical implications for the most common situations, such as exhaustive sampling of ROIs, sampling ROIs of different sizes, sampling the same ROIs for multiple histological methods, sampling more ROIs with variable intensities or using various objectives, multistage sampling and virtual sampling. Recommendations were provided for pilot studies on systematic uniform random sampling of FOVs as a part of optimizing the efficiency of histological quantification to prevent over- or undersampling. We critically discussed the pros and cons of using virtual sections for sampling FOVs from whole scanned sections. Our review demonstrated that whole slide scans of histological sections facilitate the design of sampling strategies for quantitative histology.

Vancomycin-Loaded Collagen/Hydroxyapatite Layers Electrospun on 3D Printed Titanium Implants Prevent Bone Destruction Associated with S. epidermidis Infection and Enhance Osseointegration.

The aim of the study was to develop an orthopedic implant coating in the form of vancomycin-loaded collagen/hydroxyapatite layers (COLHA+V) that combine the ability to prevent bone infection with the ability to promote enhanced osseointegration. The ability to prevent bone infection was investigated employing a rat model that simulated the clinically relevant implant-related introduction of bacterial contamination to the bone during a surgical procedure using a clinical isolate of Staphylococcus epidermidis. The ability to enhance osseointegration was investigated employing a model of a minipig with terminated growth. Six weeks following implantation, the infected rat femurs treated with the implants without vancomycin (COLHA+S. epidermidis) exhibited the obvious destruction of cortical bone as evinced via a cortical bone porosity of up to 20% greater than that of the infected rat femurs treated with the implants containing vancomycin (COLHA+V+S. epidermidis) (3%) and the non-infected rat femurs (COLHA+V) (2%). The alteration of the bone structure of the infected COLHA+S. epidermidis group was further demonstrated by a 3% decrease in the average Ca/P molar ratio of the bone mineral. Finally, the determination of the concentration of vancomycin released into the blood stream indicated a negligible systemic load. Six months following implantation in the pigs, the quantified ratio of new bone indicated an improvement in osseointegration, with a two-fold bone ingrowth on the COLHA (47%) and COLHA+V (52%) compared to the control implants without a COLHA layer (27%). Therefore, it can be concluded that COLHA+V layers are able to significantly prevent the destruction of bone structure related to bacterial infection with a minimal systemic load and, simultaneously, enhance the rate of osseointegration.

Mesenchymal Stromal Cell Therapy in Novel Porcine Model of Diffuse Liver Damage Induced by Repeated Biliary Obstruction.

In liver surgery, biliary obstruction can lead to secondary biliary cirrhosis, a life-threatening disease with liver transplantation as the only curative treatment option. Mesenchymal stromal cells (MSC) have been shown to improve liver function in both acute and chronic liver disease models. This study evaluated the effect of allogenic MSC transplantation in a large animal model of repeated biliary obstruction followed by partial hepatectomy. MSC transplantation supported the growth of regenerated liver tissue after 14 days (MSC group, n = 10: from 1087 ± 108 (0 h) to 1243 ± 92 mL (14 days); control group, n = 11: from 1080 ± 95 (0 h) to 1100 ± 105 mL (14 days), p = 0.016), with a lower volume fraction of hepatocytes in regenerated liver tissue compared to resected liver tissue (59.5 ± 10.2% vs. 70.2 ± 5.6%, p < 0.05). Volume fraction of connective tissue, blood vessels and bile vessels in regenerated liver tissue, serum levels of liver enzymes (AST, ALT, ALP and GGT) and liver metabolites (albumin, bilirubin, urea and creatinine), as well as plasma levels of IL-6, IL-8, TNF-α and TGF-ß, were not affected by MSC transplantation. In our novel, large animal (pig) model of repeated biliary obstruction followed by partial hepatectomy, MSC transplantation promoted growth of liver tissue without any effect on liver function. This study underscores the importance of translating results between small and large animal models as well as the careful translation of results from animal model into human medicine.

The time has come to extend the expiration limit of cryopreserved allograft heart valves.

Despite the wide choice of commercial heart valve prostheses, cryopreserved semilunar allograft heart valves (C-AHV) are required, and successfully transplanted in selected groups of patients. The expiration limit (EL) criteria have not been defined yet. Most Tissue Establishments (TE) use the EL of 5 years. From physiological, functional, and surgical point of view, the morphology and mechanical properties of aortic and pulmonary roots represent basic features limiting the EL of C-AHV. The aim of this work was to review methods of AHV tissue structural analysis and mechanical testing from the perspective of suitability for EL validation studies. Microscopic structure analysis of great arterial wall and semilunar leaflets tissue should clearly demonstrate cells as well as the extracellular matrix components by highly reproducible and specific histological staining procedures. Quantitative morphometry using stereological grids has proved to be effective, as the exact statistics was feasible. From mechanical testing methods, tensile test was the most suitable. Young's moduli of elasticity, ultimate stress and strain were shown to represent most important AHV tissue mechanical characteristics, suitable for exact statistical analysis. C-AHV are prepared by many different protocols, so as each TE has to work out own EL for C-AHV.

Inflammatory cell infiltrates, hypoxia, vascularization, pentraxin 3 and osteoprotegerin in abdominal aortic aneurysms - A quantitative histological study.

Information about the tissue characteristics of abdominal aortic aneurysms (AAAs), some of which may be reflected in the serum, can help to elucidate AAA pathogenesis and identify new AAA biomarkers. This information would be beneficial not only for diagnostics and follow-up but also for potential therapeutic intervention. Therefore, the aim of our study was to compare the expression of structural proteins, immune factors (T and B lymphocytes, macrophages, neutrophils and pentraxin 3 (PTX3)), osteoprotegerin (OPG), microvessels and hypoxic cells in AAA and nonaneurysmal aortic walls. We examined specimens collected during surgery for AAA repair (n = 39) and from the abdominal aortas of kidney donors without AAA (n = 8). Using histochemical and immunohistochemical methods, we quantified the areas positive for smooth muscle actin, desmin, elastin, collagen, OPG, CD3, CD20, MAC387, myeloperoxidase, PTX3, and hypoxia-inducible factor 1-alpha and the density of CD31-positive microvessels. AAA samples contained significantly less actin, desmin, elastin and OPG, more collagen, macrophages, neutrophils, T lymphocytes, B lymphocytes, hypoxic cells and PTX3, and a greater density of vasa vasorum (VV) than those in non-AAA samples. Hypoxia positively correlated with actin and negatively correlated with collagen. Microvascular density was related to inflammatory cell infiltrates, hypoxia, PTX3 expression and AAA diameter. The lower OPG expression in AAAs supports the notion of its protective role in AAA remodeling. AAA contained altered amounts of structural proteins, implying reduced vascular elasticity. PTX3 was upregulated in AAA and colocalized with inflammatory infiltrates. This evidence supports further evaluation of PTX3 as a candidate marker of AAA. The presence of aortic hypoxia, despite hypervascularization, suggests that hypoxia-induced neoangiogenesis may play a role in AAA pathogenesis. VV angiogenesis of the AAA wall increases its vulnerability.