Coating PTFE vascular prostheses with a fibroblastic matrix improves cell retention when subjected to blood flow.

An investigation was made into the effect of blood flow on endothelial cells (EC) and mesothelial cells (MC) seeded on a vascular expanded polytetrafluoroethylene (ePTFE) prosthesis coated with a fibroblastic matrix. Endothelial cells were obtained from the external jugular vein and MC from the omentum. To test the performance of prostheses, a custom designed, femoral "ex vivo" circuit was developed in mongrel dogs. Four study groups were established: a control group, A1, where prostheses were uncoated and seeded with EC; a second control group, A2, where prostheses were uncoated and seeded with MC; group B1 where prostheses were coated with a fibroblastic matrix and seeded with EC; and group B2 where coated prostheses were seeded with MC. All cells were labeled with 111Indium oxine (10 microCi/mL) before seeding. After the seeded cells had formed a monolayer on the ePTFE prostheses (which took approximately 24 h) the prostheses were placed in the "ex vivo" circuit. The rates of blood flow to which prostheses were exposed were measured at the point of inflow (117.5 +/- 12.50 mL/min, mean +/- SD) and outflow (72.6 +/- 14.3 mL/min). MC showed a greater baseline radionuclide uptake than did EC. The cells of groups B1 and B2 adhered sufficiently to the fibroblastic matrix and covered enough of the prosthetic surface to be positioned in the "ex vivo" circuit (76.90 +/- 8.24% surface covered in EC-seeded prostheses and 71.65 +/- 6.23% in MC-seeded prostheses). After exposure to blood flow the quantity of radionuclide-labeled cells and the prosthetic surface covered by them were greatly reduced though the fibroblast-coated prostheses showed greater cell retention

Mesothelial versus endothelial cell seeding: evaluation of cell adherence to a fibroblastic matrix using 111In oxine

The aim of this study was to compare the behaviour of mesothelial cells (MC) to that of endothelial cells (EC) when seeded onto a PTFE, prosthesis coated with a fibroblastic matrix. Three study groups were examined: a control group (Control) of PTFE prostheses with a fibroblast matrix (n = 8); Group EC, PTFE prostheses seeded with EC on a fibroblastic matrix (n = 8); and Group MC, PTFE, prostheses seeded with MC on a fibroblastic matrix (n = 8). All cell types were labelled with 111In (100 microCi/ml) 24 h after seeding, when the cells had formed a monolayer on the prosthetic surface. Radioactive levels were measured at 2, 4, 6, and 24 h. Both EC and MC showed optimal adherence. The MC had a better radioactive uptake and retention than the EC. The number of EC and MC cells that remained adherent to the matrix was large enough to ensure complete covering of the prosthetic surface. The use of MC is therefore feasible as an optimal alternative for achieving a natural covering on vascular prostheses prepared with a fibroblastic matrix

Rapid thawing increases the fragility of the cryopreserved arterial wall

To extend present knowledge of the biomechanical and structural changes which occur in the cryopreserved, rapidly thawed arterial wall. Minipig iliac arterial segments were cryopreserved at -196 degrees C in either minimum essential medium or Wisconsin solution. Fresh segments served as the control group. After 1 month, the specimens were rapidly thawed (37 degrees C) and processed for biomechanical, ultrastructural, morphological and immunohistochemical (MMP-1, MMP-2, MMP-3 and MMP-9) analysis. Visualisation of apoptotic cells was performed by TUNEL method. For the mechanical distension analysis, an in vitro circuit was designed. The cryopreserved segments showed a 42% incidence of spontaneous fracture and the appearance of microfractures which affected the endoluminal third of the vessel. An accumulation of liquid in the subelastica was observed. An increased expression of wall-degradative enzymes (mainly MMP-2) was also observed following cryopreservation. No significant differences were detected in the proportional elasticity module or tensile strength of the specimen groups. No differences in mechanical distension were observed between groups after the vessel segments were subjected to the pulsatile circuit flow for 72 h. Cell damage was most intense in the specimens cryopreserved in Wisconsin solution. Cryopreservation in both the solutions employed, followed by rapid thawing, induce changes in the permeability which increase the fragility of the cryopreserved arterial wall. Both increased expression of wall-degradative enzymes and accumulation of liquid may contribute to graft failure after implantation

Evaluation of the smooth muscle cell component and apoptosis in the varicose vein wall

This study was designed to evaluate the role of the smooth muscle cell and the apoptosis in the pathogenesis of the varicose vein. Segments of saphenous vein were obtained from healthy subjects and from those with varicose veins. The vein specimens were subdivided according to subject age (younger or older than 50 years) and according to the varicose vein source (distal or proximal). Morphological, ultrastructural, cell proliferation (anti-PCNA method) and cell death (TUNEL method) analysis were performed. The walls of healthy, control vein specimens acquired a more collagenous and papillomatous appearance with age. A slight increase in the number of TUNEL-positive cells was also observed in specimens from older subjects. The proportion of apoptotic cells was much greater in the varicose veins than in control specimens. Most cellular alterations were seen in proximal varicose segments obtained from young subjects. These specimens showed hypertrophic areas with a high degree of cellularity (both in the media and in the thickened intima). The highest proportion of apoptotic cells and collagenisation were also observed in these areas. The enhanced number of apoptotic cells in varicose veins observed mainly in proximal/young vein specimens could be responsible, at least in part, for the acceleration of the final fibrosclerotic process characteristic of the varicose vein wall

Bioactive bilayered dressing for compromised epidermal tissue regeneration with sequential activity of complementary agents

The article deals with the design, preparation, and evaluation of a new bilayered dressing for application in the healing of compromised wounds. The system is based on the sequential release of two complementary bioactive components to enhance the activation of the regeneration of dermal tissue. The internal layer is a highly hydrophilic and biodegradable film of gelatin and hyaluronic acid (HG), crosslinked with the natural compound genipin, which reacts with the amine groups of gelatin. This film is loaded with the proangiogenic, anti-inflammatory, and antibacterial peptide, proadrenomedullin N-terminal 20 peptide (PAMP), that is released slowly in the wound site. The external layer, more stable and less hydrophilic, is constituted by a biodegradable polyurethane derived from poly(caprolactone) and pluronic L61. This layer is loaded with resorbable nanoparticles of bemiparin (a fractionated low molecular weight heparin), which promotes the activation of growth factors, FGF and VEGF, and provides a good biomechanical stability and controlled permeability of the bilayered dressing. Experiments carried out in mice demonstrate the excellent angiogenic effect of the HG film in the dermal tissue. Application of the bilayered dressing in the wound healing rabbit ear model shows an improved cicatrization of the wound in both ischemic and non-ischemic defects, favoring epithelialization and reducing noticeably the contraction and the inflammation.This work was supported by the CIBER-BBN and a Grant from Spain’s Ministry of Science and Education (SAF2009-13240-C02-01).Peer Reviewe

Tissue remodelling and increased DNA damage in patients with incompetent valves in chronic venous insufficiency

Chronic venous insufficiency (CVI), in which blood return to the heart is impaired, is a prevalent condition worldwide. Valve incompetence is a complication of CVI that results in blood reflux, thereby aggravating venous hypertension. While CVI has a complex course and is known to produce alterations in the vein wall, the underlying pathological mechanisms remain unclear. This study examined the presence of DNA damage, pro-inflammatory cytokines and extracellular matrix remodelling in CVI-related valve incompetence. One hundred and ten patients with CVI were reviewed and divided into four groups according to age (<50 and ≥50 years) and a clinical diagnosis of venous reflux indicating venous system valve incompetence (R) (n = 81) or no reflux (NR) (n = 29). In vein specimens (greater saphenous vein) from each group, PARP, IL-17, COL-I, COL-III, MMP-2 and TIMP-2 expression levels were determined by RT-qPCR and immunohistochemistry. The younger patients with valve incompetence showed significantly higher PARP, IL-17, COL-I, COL-III, MMP-2 and reduced TIMP-2 expression levels and a higher COL-I/III ratio. Young CVI patients with venous reflux suffer chronic DNA damage, with consequences at both the local tissue and systemic levels, possibly associated with ageing.This study (FIS-PI18/00912) was supported by the Instituto de Salud Carlos III (Plan Estatal de I + D+i 2013-2016) and co-financed by the European Development Regional Fund “A way to achieve Europe” (ERDF) and B2017/BMD-3804 MITIC-CM

Histopathological study of JNK in venous wall of patients with chronic venous insufficiency related to osteogenesis process

Chronic venous insufficiency (CVI) is one of the most common vascular pathologies worldwide. One of the risk factors for the development of CVI is aging, which is why it is related to senile changes. The main trigger of the changes that occur in the venous walls in CVI is blood flow reflux, which produces increased hydrostatic pressure, leading to valve incompetence. The cellular response is one of the fundamental processes in vascular diseases, causing the activation of cell signalling pathways such as c-Jun N-terminal kinase (JNK). Metabolic changes and calcifications occur in vascular pathology as a result of pathophysiological processes. The aim of this study was to determine the expression of JNK in venous disease and its relationship with the role played by the molecules involved in the osteogenic processes in venous tissue calcification. This was a cross-sectional study that analyzed the greater saphenous vein wall in 110 patients with (R) and without venous reflux (NR), classified according to age. Histopathological techniques were used and protein expression was analysed using immunohistochemistry techniques for JNK and markers of osteogenesis (RUNX2, osteocalcin (OCN), osteopontin (OPN)). Significantly increased JNK, RUNX2, OCN, OPN and pigment epithelium-derived factor (PEDF) protein expression and the presence of osseous metaplasia and amorphous calcification were observed in younger patients (<50 years) with venous reflux. This study shows for the first time the existence of an osteogenesis process related to the expression of JNK in the venous wall.This study (FIS-PI18/00912) was supported by the Instituto de Salud Carlos III (Plan Estatal de I+D+i 2013-2016) and cofinanced by the European Development Regional Fund ‘‘A way to achieve Europe’’ (ERDF) and B2017/BMD-3804 MITIC-C

Signal Transduction Pathways in Breast Cancer: The Important Role of PI3K/Akt/mTOR

Breast cancer is the cancer with the highest prevalence in women and is the number-one cause of cancer mortality worldwide. Cell transduction is a fundamental process in the development and progression of cancer. Modifications in various cell signalling pathways promote tumour cell proliferation, progression, and survival. The PI3K/Akt/mTOR pathway is an example of that, and it is involved in growth, proliferation, survival, motility, metabolism, and immune response regulation. Activation of this pathway is one of the main causes of cancer cell resistance to antitumour therapies. This makes PI3K/Akt/mTOR signalling a crucial object of study for understanding the development and progression of this disease. Thus, this pathway may have a role as a potential therapeutic target, as well as prognostic and diagnostic value, in patients with breast cancer. Despite the existence of selective PI3K/Akt/mTOR pathway inhibitors and current clinical trials, the cellular mechanisms are not yet known. The present review aims to understand the current state of this important disease and the paths that must be forged.This work was supported by grants from the B2017/BMD-3804 MITIC-CM (Community of Madrid, Spain), cofinanced by the European Development Regional Fund “A Way to Achieve Europe” (ERDF)

Use of a fibroblastic matrix improves the results of mesothelial-cell seeding on vascular prostheses of polytetrafluoroethylene

Mesothelial cells (MC) obtained from the human omentum are a good alternative to the use of endothelial cells (EC) as a covering for vascular prostheses of polytetrafluoroethylene (PTFE), given the antithrombogenic properties and good behaviour in vitro of mesothelial cells. We studied the behaviour of mesothelial cells seeded on PTFE prostheses with an interposed fibroblastic matrix for seeding. The mesothelial cells were extracted from 30-40 g fragments of human omentum by enzymatic digestion with collagenase. The cells extracted were seeded onto small disks of PTFE to which a matrix composed of fibroblastic cells had been fixed with 5% glycerol after the fibroblasts reached convergence. Interposition of a fibroblastic matrix fixed with glycerol notably improved the adherence of the seeded mesothelial cells and the stability and durability of the cell layer formed on the prosthetic surface. The effectiveness of seeding mesothelial cells was confirmed by labelling the cells with l'lIn-oxine. This showed that once the cell layer had formed (24 h after seeding), the fibroblastic matrix favoured the maintenance of a stable layer of mesothelial cells 4 hours after uptake of the radioactive substance