Gingival regeneration aims at restoring the architecture and functionality of oral damaged tissue... more Gingival regeneration aims at restoring the architecture and functionality of oral damaged tissue. Different biomaterials or biological materials have been tested for tissue repair, such as platelet concentrates such as PL. In this article, the use of extracellular vesicles (EVs) derived from platelet lysate (PL) and their combination with hyaluronic acid biomaterials (HA) in an in vitro wound healing assay is investigated. EVs were isolated by size exclusion chromatography from PL. In addition, HA gels were formulated with PL or EVs. EVs or HA combined with EVs (HA-EVs) were tested in vitro in gingival fibroblasts and keratinocytes for biocompatibility (LDH activity and metabolic activity) and by an in vitro wound-healing assay and gene expression analysis. EVs and EVs-HA treatments were biocompatible in gingival fibroblasts and keratinocytes and showed an increase in wound healing in vitro compared to control. Moreover, changes in gene expression related to extracellular matrix re...
The aim of this study was to identify candidate genes for visceral obesity by screening for genes... more The aim of this study was to identify candidate genes for visceral obesity by screening for genes strongly differentially expressed between human subcutaneous and visceral adipose depots. A cDNA microarray with human adipose-derived cDNAs was used as an initial screening to identify genes that are potentially differentially expressed between human subcutaneous and visceral abdominal fat tissues. For the two best candidates, carboxypeptidase E (CPE) and thrombospondin-1 (THBS1) (EST N72406), real-time RT-PCR was performed to confirm their depot specific expression in extremely obese individuals. Both genes appeared to be strongly differentially expressed, having a higher expression in the visceral depot than in the subcutaneous one. For THBS1, the difference in expression between the depots was greater in women than in men. The involvement of CPE and THBS1 in obesity allows us to suggest that the physiological processes controlled by these genes contribute to depot and gender-related differences in the metabolic complications of obesity.
The prevalence of obesity has increased to an extraordinary degree, especially over the last thre... more The prevalence of obesity has increased to an extraordinary degree, especially over the last three decades, so that if these trends persist, practically all of the adult population would be obese in the course of only two generations. On the basis of family and adoption studies, it has been estimated that the genetic component in obesity ranges from 20% to 80%. Thus, the most common forms of obesity would depend on the interaction of multiple genes as well as on the influence of various environmental factors, such as eating behaviour and lifestyle. Although none of these genes potentially involved in the control of body weight seems to be directly responsible for the syndrome, there have been reports of interesting associations between polymorphisms of certain candidate genes and obesity or its metabolic complications. The studies into associations between genotypes and obese phenotype have increased over the last few years and have basically focussed on the genes involved in the co...
To determine whether there are variations in leptin levels according to the β3-adrenoceptor (β3-A... more To determine whether there are variations in leptin levels according to the β3-adrenoceptor (β3-AR) Trp64Arg and uncoupling protein 1 (UCP1) −3826A→G polymorphisms, given the regulatory role of catecholamines through the β3-AR in leptin production and the previously reported association of the UCP1 −3826A→G variant with obesity. A total of 160 men and 172 women randomly chosen from a nationwide population-based
The development of metabolic complications of obesity has been associated with the existence of d... more The development of metabolic complications of obesity has been associated with the existence of depot-specific differences in the biochemical properties of adipocytes. The aim of this study was to investigate, in severely obese men and women, both gender- and depot-related differences in lipoprotein lipase (LPL) expression and activity, as well as the involvement of endocrine and biometric factors and their dependence on gender and/or fat depot. Morbidly obese, nondiabetic, subjects (9 men and 22 women) aged 41.1+/-1.9 years, with a body mass index (BMI) of 54.7+/-1.7 kg/m(2) who had undergone abdominal surgery were studied. Both expression and activity of LPL and leptin expression were determined in adipose samples from subcutaneous and visceral fat depots. In both men and women, visceral fat showed higher LPL mRNA levels as well as lower ob mRNA levels and tissue leptin content than the subcutaneous one. In both subcutaneous and visceral adipose depots, women exhibited higher protein content, decreased fat cell size and lower LPL activity than men. The gender-related differences found in abdominal fat LPL activity could contribute to the increased risk for developing obesity-associated diseases shown by men, even in morbid obesity, in which the massive fat accumulation could mask these differences. Furthermore, the leptin content of fat depots as well as plasma insulin concentrations appear in our population as the main determinants of adipose tissue LPL activity, adjusted by gender, depot and BMI.
Summaryobjetive Given the important role of the β2‐adrenoceptor (β2‐AR) in lipid mobilization an... more Summaryobjetive Given the important role of the β2‐adrenoceptor (β2‐AR) in lipid mobilization and the lack of studies in Southern European countries, the aim of this study was to investigate the role of the glutamine 27 glutamic acid (Gln27Glu) β2‐AR polymorphism in the susceptibility to obesity and its metabolic complications in a population‐based nationwide multicentre study in Spain, especially focusing on the hypothetical influence of gender.design Cross‐sectional population‐based study.patients We studied 666 nonrelated adults (47·9% men and 52·1% women), aged 35–64 years, chosen randomly from a nationwide population‐based survey of obesity, and related conditions including insulin resistance and cardiovascular risk factors.measurements Body mass index (BMI), waist‐to‐hip ratio (WHR), sagittal abdominal diameter (SAD), systolic and diastolic blood pressure, fasting and 2‐h post‐glucose load glycaemic levels, total cholesterol, high density lipoprotein (HDL)‐ and low density...
The present study was performed to analyze in detail gender-and site-related alterations in the a... more The present study was performed to analyze in detail gender-and site-related alterations in the adrenergic signal transduction pathway of lipolysis in fat cells isolated from subcutaneous abdominal and visceral fat depots from severely obese patients. The study group consisted of 30 morbidly obese subjects (9 men and 21 women) aged 41.1±1.9 years, with a body mass index (BMI) of 54.7±1.7 kg/m2, who had undergone abdominal surgery. Protein levels of hormone-sensitive lipase (HSL) and adrenergic receptors (AR), as well as HSL activity and the lipolytic response to adrenergic agents were analyzed. Both fat depots had similar basal lipolysis, but the capacity of catecholamines to activate lipolysis was greater in visceral fat, both at AR and postreceptor levels. Basal lipolysis and lipolytic activity induced by dibutyryl cyclic AMP were higher in men than in women. However, the visceral depot of women showed a higher maximal stimulation by noradrenaline than that of men, in accordance with higher β 1-and β 3-AR protein levels. In conclusion, the main gender-related differences were located in the visceral depot, with women exhibiting a higher sensitivity to catecholamines associated with an increased provision of β-AR, while men showed an enhanced lipolytic capacity at the postreceptor level.
The aim of this study was to identify candidate genes for visceral obesity by screening for genes... more The aim of this study was to identify candidate genes for visceral obesity by screening for genes strongly differentially expressed between human subcutaneous and visceral adipose depots. A cDNA microarray with human adipose-derived cDNAs was used as an initial screening to identify genes that are potentially differentially expressed between human subcutaneous and visceral abdominal fat tissues. For the two best candidates, carboxypeptidase E (CPE) and thrombospondin-1 (THBS1) (EST N72406), real-time RT-PCR was performed to confirm their depot specific expression in extremely obese individuals. Both genes appeared to be strongly differentially expressed, having a higher expression in the visceral depot than in the subcutaneous one. For THBS1, the difference in expression between the depots was greater in women than in men. The involvement of CPE and THBS1 in obesity allows us to suggest that the physiological processes controlled by these genes contribute to depot and gender-related differences in the metabolic complications of obesity.
Extracellular vesicles (EVs) represent a new paradigm, both in cell biology and medicine; specifi... more Extracellular vesicles (EVs) represent a new paradigm, both in cell biology and medicine; specifically, the idea that functional content itself may be delivered directly to cells. EVs are cell-derived membranous structures that work as intercellular communicators exerting their function by transporting their cargo that includes nucleic acids, proteins and lipids. EVs play an essential role in normal physiology, but also in pathological communication, for instance, in cancer, EVs are thought to deliver oncogenic molecules (such as proteins, peptides, RnAs…) to neighboring cells, enhancing propagation of neoplastic cells. Not surprisingly, EV research has become commonplace in every field of biomedicine, being explored as diagnostics and therapeutics. This Collection gathers original Articles that investigate the application of extracellular vesicles on diagnostics and therapeutics, and that report advances in the knowledge of EV biology and the methodological tools for their study.
Hydrogels and extracellular vesicle-based therapies have been proposed as emerging therapeutic as... more Hydrogels and extracellular vesicle-based therapies have been proposed as emerging therapeutic assets in wound closure. The combination of these elements has given good results in managing chronic and acute wounds. The intrinsic characteristics of the hydrogels in which the extracellular vesicles (EVs) are loaded allow for overcoming barriers, such as the sustained and controlled release of EVs and the maintenance of the pH for their conservation. In addition, EVs can be obtained from different sources and through several isolation methods. However, some barriers must be overcome to transfer this type of therapy to the clinic, for example, the production of hydrogels containing functional EVs and identifying long-term storage conditions for EVs. The aim of this review is to describe the reported EV-based hydrogel combinations, along with the obtained results, and analyze future perspectives.
Gelatin methacryloyl (GelMA) hydrogels have been widely used for different biomedical application... more Gelatin methacryloyl (GelMA) hydrogels have been widely used for different biomedical applications due to their tunable physical characteristics and appropriate biological properties. In addition, GelMA could be modified with the addition of functional groups providing inherent antibacterial capabilities. Here, GelMA-based hydrogels were developed through the combination of a GelMA unmodified and modified polymer with quaternary ammonium groups (GelMAQ). The GelMAQ was synthesized from GelMA with a low degree of substitution of methacrylamide groups (DSMA) and grafted with glycidyltrimethylammonium chloride in the free amine groups of the lysine moieties present in the original gelatin. GelMAs with high DSMA and GelMAQ were combined 50/50% or 25/75% (w/w), respectively, and compared to controls GelMA and GelMA with added chlorhexidine (CHX) at 0.2%. The different hydrogels were characterized using 1H-NMR spectroscopy and swelling behavior and tested in (1) Porphyromonas gingivalis t...
A key factor for dental implant success is a good sealing between the implant surface and both so... more A key factor for dental implant success is a good sealing between the implant surface and both soft (gum) and hard (bone) tissues. Surface nanotopography can modulate cell response through mechanotransduction. The main objective of this research was the development of nanostructured titanium (Ti) surfaces that promote both soft and hard tissue integration with potential application in dental implants. Nanostructured Ti surfaces were developed by electrochemical anodization—nanopores (NPs) and nanonets (NNs)—and characterized by atomic force microscopy, scanning electronic microscopy, and contact angle analysis. In addition, nanoparticle release and apoptosis activation were analyzed on cell culture. NP surfaces showed nanoparticle release, which increased in vitro cell apoptosis. Primary human gingival fibroblasts (hGFs) and human bone marrow mesenchymal stem cells (hBM-MSCs) were used to test cell adhesion, cytotoxicity, metabolic activity, and differentiation markers. Finally, cel...
Extracellular vesicles (EVs) present a great potential for the development of new treatments in t... more Extracellular vesicles (EVs) present a great potential for the development of new treatments in the biomedical field. To be used as therapeutics, many different sources have been used for EVs obtention, while only a few studies have addressed the use of platelet-derived EVs (pEVs). In fact, pEVs have been shown to intervene in different healing responses, thus some studies have evaluated their regenerative capability in wound healing or hemorrhagic shock. Even more, pEVs have proven to induce cellular differentiation, enhancing musculoskeletal or neural regeneration. However, the obtention and characterization of pEVs is widely heterogeneous and differs from the recommendations of the International Society for Extracellular Vesicles. Therefore, in this review, we aim to present the main advances in the therapeutical use of pEVs in the regenerative medicine field while highlighting the isolation and characterization steps followed. The main goal of this review is to portray the studi...
Extracellular Vesicles (EVs) are biological nanovesicles that play a key role in cell communicati... more Extracellular Vesicles (EVs) are biological nanovesicles that play a key role in cell communication. Their content includes active biomolecules such as proteins and nucleic acids, which present great potential in regenerative medicine. More recently, EVs derived from Platelet Lysate (PL) have shown an osteogenic capability comparable to PL. Besides, biomaterials are frequently used in orthopedics or dental restoration. Here, we provide a method to functionalize Ti surfaces with PL-derived EVs in order to improve their osteogenic properties. EVs are isolated from PL by size exclusion chromatography, and afterward Ti surfaces are functionalized with PL-EVs by drop casting. Functionalization is proven by EVs release and its biocompatibility by the lactate dehydrogenase (LDH) release assay.
Background: We aimed to evaluate the effect of low doses (LD) bone morphogenetic protein-2 (BMP2)... more Background: We aimed to evaluate the effect of low doses (LD) bone morphogenetic protein-2 (BMP2) and BMP4 micro-immunotherapy (MI) in two in vitro models of periodontal wound healing/regeneration. Methods: We first evaluated the effect of LD of BMP2 and BMP4 MI on a 2D cell culture using human gingival fibroblasts (hGF) under inflammatory conditions induced by IL1β. Biocompatibility, inflammatory response (Prostaglandin E2 (PGE2) release), collagen deposition and release of extracellular matrix (ECM) organization-related enzymes (matrix metalloproteinase-1 (MMP1) and metalloproteinase inhibitor 1 (TIMP1)) were evaluated after short (3 days) and long-term (24 days) treatment with BMP2 or BMP4 MI. Then, given the results obtained in the 2D cell culture, LD BMP4 MI treatment was evaluated in a 3D cell culture model of human tissue equivalent of gingiva (GTE) under the same inflammatory stimulus, evaluating the biocompatibility, inflammatory response and effect on MMP1 and TIMP1 release. Results: LD BMP4 was able to decrease the release of the inflammatory mediator PGE2 and completely re-establish the impaired collagen metabolism induced by IL1β treatment. In the 3D model, LD BMP4 treatment improved tissue viability compared with the vehicle, with similar levels to 3D tissues without inflammation. No significant effects were observed on PGE2 levels nor MMP1/TIMP1 ratio after LD BMP4 treatment, although a tendency to decrease PGE2 levels was observed after 3 days. Conclusions: LD BMP4 MI treatment shows anti-inflammatory and regenerative properties on hGF, and improved viability of 3D gingiva under inflammatory conditions. LD BMP4 MI treatment could be used on primary prevention or maintenance care of periodontitis. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.
In the last years, several studies testing commercial periodontal gels that contain chlorhexidine... more In the last years, several studies testing commercial periodontal gels that contain chlorhexidine (CHX) or other antibacterial agents, have raised concerns regarding their cytotoxicity in periodontal tissues. We aimed at comparing the biocompatibility but also the efficacy as regards to the antibacterial and wound healing ability of different commercial periodontal gels. In vitro human gingival fibroblasts (GF) and a 3D model of human tissue equivalents of gingiva (GTE) were used under inflammatory conditions to evaluate wound closure, cytotoxicity and gene expression. Antibacterial effects were also investigated on Porphyromonas gingivalis growth, viability and gingipain activity. In GF and in the bacterial study, we found cytotoxic effects on GF and a high inhibition on bacterial growth rate in gels containing CHX, asiaticoside, enoxolone, cetylpyridinium chloride, propolis and eugenol. Of the two gels that were non-cytotoxic, Syntoss Biogel (containing chondrontin sulfate) and Em...
Although coronary stents have improved the early and long-term consequences of arterial lesions, ... more Although coronary stents have improved the early and long-term consequences of arterial lesions, the prevention of restenosis and late stent thrombosis is key to prevent a new obstruction of the vessel. Here we aimed at improving the tissue response to stents through surface modification. For that purpose, we used two different approaches, the use of nanostructuration by electrochemical anodization and the addition of a quercitrin (QR) coating to the Ti surface. Four surfaces (Ti, NN, TiQR and NNQR) were characterized by atomic force microscopy, scanning electronic microscopy and contact angle analysis and QR content was evaluated by fluorescent staining. Cell adhesion, cytotoxicity, metabolic activity and nitric oxide (NO) production was evaluated on primary human umbilical cord endothelial cells (HUVECs). Platelet adhesion, hemolysis rate and Staphylococcus epidermidis CECT 4184 adhesion at 30 min were analyzed. Nanostructuration induced an increase on surface roughness, and QR co...
(1) One strategy to improve the outcome of orthopedic implants is to use porous implants with the... more (1) One strategy to improve the outcome of orthopedic implants is to use porous implants with the addition of a coating with an antibacterial biomolecule. In this study, we aimed to produce and test the biocompatibility, the osteopromotive (both under normal conditions and under a bacterial challenge with lipopolysaccharide (LPS)) and antibacterial activities of a porous Ti-6Al-4V implant coated with the flavonoid quercitrin in vitro. (2) Porous Ti-6Al-4V implants were produced by 3D printing and further functionalized with quercitrin by wet chemistry. Implants were characterized in terms of porosity and mechanical testing, and the coating with quercitrin by fluorescence staining. Implant biocompatibility and bioactivity was tested using MC3T3-E1 preosteoblasts by analyzing cytotoxicity, cell adhesion, osteocalcin production, and alkaline phosphatase (ALP) activity under control and under bacterial challenging conditions using lipopolysaccharide (LPS). Finally, the antibacterial properties of the implants were studied using Staphylococcus epidermidis by measuring bacterial viability and adhesion. (3) Porous implants showed pore size of about 500 µm and a porosity of 52%. The coating was homogeneous over all the 3D surface and did not alter the mechanical properties of the Young modulus. Quercitrin-coated implants showed higher biocompatibility, cell adhesion, and osteocalcin production compared with control implants. Moreover, higher ALP activity was observed for the quercitrin group under both normal and bacterial challenging conditions. Finally, S. epidermidis live/dead ratio and adhesion after 4 h of incubation was lower on quercitrin implants compared with the control. (4) Quercitrin-functionalized porous Ti-6Al-4V implants present a great potential as an orthopedic porous implant that decreases bacterial adhesion and viability while promoting bone cell growth and differentiation.
Extracellular vesicles (EVs) have been recently identified as vital components of cell-based ther... more Extracellular vesicles (EVs) have been recently identified as vital components of cell-based therapies based on the observation that conditioned media from cultured stromal cells reproduce some of the beneficial effects of intact cells. In order to obtain clinically active EVs derived from Mesenchymal Stromal Cells (MSCs) different procedures have been reported in the literature. Usually, non-confluent cells are incubated with culture medium for 48 h either with EV-depleted Fetal Bovine Serum (FBS) or without FBS. Our aim was to compare the effects of EVs isolated by ultracentrifugation from human umbilical cord MSC conditioned media obtained using these two conditions: with EV-depleted FBS (UC) or without FBS (UCw/o) on the mRNA expression levels of extracellular matrix related genes using the mouse chondrogenic cell line ATDC-5. We observed a deleterious effect on chondrogenic cells treated with UCw/o, showing higher mRNA expression levels of different metalloproteinases and decor...
Journal of Tissue Engineering and Regenerative Medicine, 2020
The demineralized bone matrix (DBM) is the most widely used bone allograft, which is obtained by ... more The demineralized bone matrix (DBM) is the most widely used bone allograft, which is obtained by removing the mineral component of bone, leading to exposure of the proteins responsible for osteoinduction. For clinical use, DBM shall be formulated with a carrier that provides consistency and improves its osteoinduction. In this study, three DBM formulations with glycerol (Gly), hyaluronic acid (HA) and gelatin methacryloyl (GelMA) were evaluated measuring their physicochemical properties (microstructure, compressive strength and serum cohesivity), and their osteoinductive capacity both in vitro using C2C12 cells and umbilical cord human mesenchymal stem cells and in vivo in an ectopic bone formation model in athymic mice. To assess the effectiveness of DBM in vitro in inducing the differentiation into osteoblasts, alkaline phosphatase (ALP) activity was assessed in combination with a cytotoxicity assay. In vivo, new bone formation was assessed by histological and radiological analysis. In the compression and in the serum cohesive assays, the GelMA DBM formulation showed its superiority over the other formulations. In addition, GelMA showed a more compact structure analyzed by scanning electron microscopy. Higher cell toxicity was observed on Gly formulations in vitro, while GelMa and HA showed very low toxicity. All formulations significantly improved ALP activity compared to control. In the in vivo studies, GelMA showed the greatest osteoinductive potential with a higher percentage of new bone and bone marrow formation. Our results suggest GelMA is useful as a carrier for DBM designed to promote the formation of the new bone.
Gingival regeneration aims at restoring the architecture and functionality of oral damaged tissue... more Gingival regeneration aims at restoring the architecture and functionality of oral damaged tissue. Different biomaterials or biological materials have been tested for tissue repair, such as platelet concentrates such as PL. In this article, the use of extracellular vesicles (EVs) derived from platelet lysate (PL) and their combination with hyaluronic acid biomaterials (HA) in an in vitro wound healing assay is investigated. EVs were isolated by size exclusion chromatography from PL. In addition, HA gels were formulated with PL or EVs. EVs or HA combined with EVs (HA-EVs) were tested in vitro in gingival fibroblasts and keratinocytes for biocompatibility (LDH activity and metabolic activity) and by an in vitro wound-healing assay and gene expression analysis. EVs and EVs-HA treatments were biocompatible in gingival fibroblasts and keratinocytes and showed an increase in wound healing in vitro compared to control. Moreover, changes in gene expression related to extracellular matrix re...
The aim of this study was to identify candidate genes for visceral obesity by screening for genes... more The aim of this study was to identify candidate genes for visceral obesity by screening for genes strongly differentially expressed between human subcutaneous and visceral adipose depots. A cDNA microarray with human adipose-derived cDNAs was used as an initial screening to identify genes that are potentially differentially expressed between human subcutaneous and visceral abdominal fat tissues. For the two best candidates, carboxypeptidase E (CPE) and thrombospondin-1 (THBS1) (EST N72406), real-time RT-PCR was performed to confirm their depot specific expression in extremely obese individuals. Both genes appeared to be strongly differentially expressed, having a higher expression in the visceral depot than in the subcutaneous one. For THBS1, the difference in expression between the depots was greater in women than in men. The involvement of CPE and THBS1 in obesity allows us to suggest that the physiological processes controlled by these genes contribute to depot and gender-related differences in the metabolic complications of obesity.
The prevalence of obesity has increased to an extraordinary degree, especially over the last thre... more The prevalence of obesity has increased to an extraordinary degree, especially over the last three decades, so that if these trends persist, practically all of the adult population would be obese in the course of only two generations. On the basis of family and adoption studies, it has been estimated that the genetic component in obesity ranges from 20% to 80%. Thus, the most common forms of obesity would depend on the interaction of multiple genes as well as on the influence of various environmental factors, such as eating behaviour and lifestyle. Although none of these genes potentially involved in the control of body weight seems to be directly responsible for the syndrome, there have been reports of interesting associations between polymorphisms of certain candidate genes and obesity or its metabolic complications. The studies into associations between genotypes and obese phenotype have increased over the last few years and have basically focussed on the genes involved in the co...
To determine whether there are variations in leptin levels according to the β3-adrenoceptor (β3-A... more To determine whether there are variations in leptin levels according to the β3-adrenoceptor (β3-AR) Trp64Arg and uncoupling protein 1 (UCP1) −3826A→G polymorphisms, given the regulatory role of catecholamines through the β3-AR in leptin production and the previously reported association of the UCP1 −3826A→G variant with obesity. A total of 160 men and 172 women randomly chosen from a nationwide population-based
The development of metabolic complications of obesity has been associated with the existence of d... more The development of metabolic complications of obesity has been associated with the existence of depot-specific differences in the biochemical properties of adipocytes. The aim of this study was to investigate, in severely obese men and women, both gender- and depot-related differences in lipoprotein lipase (LPL) expression and activity, as well as the involvement of endocrine and biometric factors and their dependence on gender and/or fat depot. Morbidly obese, nondiabetic, subjects (9 men and 22 women) aged 41.1+/-1.9 years, with a body mass index (BMI) of 54.7+/-1.7 kg/m(2) who had undergone abdominal surgery were studied. Both expression and activity of LPL and leptin expression were determined in adipose samples from subcutaneous and visceral fat depots. In both men and women, visceral fat showed higher LPL mRNA levels as well as lower ob mRNA levels and tissue leptin content than the subcutaneous one. In both subcutaneous and visceral adipose depots, women exhibited higher protein content, decreased fat cell size and lower LPL activity than men. The gender-related differences found in abdominal fat LPL activity could contribute to the increased risk for developing obesity-associated diseases shown by men, even in morbid obesity, in which the massive fat accumulation could mask these differences. Furthermore, the leptin content of fat depots as well as plasma insulin concentrations appear in our population as the main determinants of adipose tissue LPL activity, adjusted by gender, depot and BMI.
Summaryobjetive Given the important role of the β2‐adrenoceptor (β2‐AR) in lipid mobilization an... more Summaryobjetive Given the important role of the β2‐adrenoceptor (β2‐AR) in lipid mobilization and the lack of studies in Southern European countries, the aim of this study was to investigate the role of the glutamine 27 glutamic acid (Gln27Glu) β2‐AR polymorphism in the susceptibility to obesity and its metabolic complications in a population‐based nationwide multicentre study in Spain, especially focusing on the hypothetical influence of gender.design Cross‐sectional population‐based study.patients We studied 666 nonrelated adults (47·9% men and 52·1% women), aged 35–64 years, chosen randomly from a nationwide population‐based survey of obesity, and related conditions including insulin resistance and cardiovascular risk factors.measurements Body mass index (BMI), waist‐to‐hip ratio (WHR), sagittal abdominal diameter (SAD), systolic and diastolic blood pressure, fasting and 2‐h post‐glucose load glycaemic levels, total cholesterol, high density lipoprotein (HDL)‐ and low density...
The present study was performed to analyze in detail gender-and site-related alterations in the a... more The present study was performed to analyze in detail gender-and site-related alterations in the adrenergic signal transduction pathway of lipolysis in fat cells isolated from subcutaneous abdominal and visceral fat depots from severely obese patients. The study group consisted of 30 morbidly obese subjects (9 men and 21 women) aged 41.1±1.9 years, with a body mass index (BMI) of 54.7±1.7 kg/m2, who had undergone abdominal surgery. Protein levels of hormone-sensitive lipase (HSL) and adrenergic receptors (AR), as well as HSL activity and the lipolytic response to adrenergic agents were analyzed. Both fat depots had similar basal lipolysis, but the capacity of catecholamines to activate lipolysis was greater in visceral fat, both at AR and postreceptor levels. Basal lipolysis and lipolytic activity induced by dibutyryl cyclic AMP were higher in men than in women. However, the visceral depot of women showed a higher maximal stimulation by noradrenaline than that of men, in accordance with higher β 1-and β 3-AR protein levels. In conclusion, the main gender-related differences were located in the visceral depot, with women exhibiting a higher sensitivity to catecholamines associated with an increased provision of β-AR, while men showed an enhanced lipolytic capacity at the postreceptor level.
The aim of this study was to identify candidate genes for visceral obesity by screening for genes... more The aim of this study was to identify candidate genes for visceral obesity by screening for genes strongly differentially expressed between human subcutaneous and visceral adipose depots. A cDNA microarray with human adipose-derived cDNAs was used as an initial screening to identify genes that are potentially differentially expressed between human subcutaneous and visceral abdominal fat tissues. For the two best candidates, carboxypeptidase E (CPE) and thrombospondin-1 (THBS1) (EST N72406), real-time RT-PCR was performed to confirm their depot specific expression in extremely obese individuals. Both genes appeared to be strongly differentially expressed, having a higher expression in the visceral depot than in the subcutaneous one. For THBS1, the difference in expression between the depots was greater in women than in men. The involvement of CPE and THBS1 in obesity allows us to suggest that the physiological processes controlled by these genes contribute to depot and gender-related differences in the metabolic complications of obesity.
Extracellular vesicles (EVs) represent a new paradigm, both in cell biology and medicine; specifi... more Extracellular vesicles (EVs) represent a new paradigm, both in cell biology and medicine; specifically, the idea that functional content itself may be delivered directly to cells. EVs are cell-derived membranous structures that work as intercellular communicators exerting their function by transporting their cargo that includes nucleic acids, proteins and lipids. EVs play an essential role in normal physiology, but also in pathological communication, for instance, in cancer, EVs are thought to deliver oncogenic molecules (such as proteins, peptides, RnAs…) to neighboring cells, enhancing propagation of neoplastic cells. Not surprisingly, EV research has become commonplace in every field of biomedicine, being explored as diagnostics and therapeutics. This Collection gathers original Articles that investigate the application of extracellular vesicles on diagnostics and therapeutics, and that report advances in the knowledge of EV biology and the methodological tools for their study.
Hydrogels and extracellular vesicle-based therapies have been proposed as emerging therapeutic as... more Hydrogels and extracellular vesicle-based therapies have been proposed as emerging therapeutic assets in wound closure. The combination of these elements has given good results in managing chronic and acute wounds. The intrinsic characteristics of the hydrogels in which the extracellular vesicles (EVs) are loaded allow for overcoming barriers, such as the sustained and controlled release of EVs and the maintenance of the pH for their conservation. In addition, EVs can be obtained from different sources and through several isolation methods. However, some barriers must be overcome to transfer this type of therapy to the clinic, for example, the production of hydrogels containing functional EVs and identifying long-term storage conditions for EVs. The aim of this review is to describe the reported EV-based hydrogel combinations, along with the obtained results, and analyze future perspectives.
Gelatin methacryloyl (GelMA) hydrogels have been widely used for different biomedical application... more Gelatin methacryloyl (GelMA) hydrogels have been widely used for different biomedical applications due to their tunable physical characteristics and appropriate biological properties. In addition, GelMA could be modified with the addition of functional groups providing inherent antibacterial capabilities. Here, GelMA-based hydrogels were developed through the combination of a GelMA unmodified and modified polymer with quaternary ammonium groups (GelMAQ). The GelMAQ was synthesized from GelMA with a low degree of substitution of methacrylamide groups (DSMA) and grafted with glycidyltrimethylammonium chloride in the free amine groups of the lysine moieties present in the original gelatin. GelMAs with high DSMA and GelMAQ were combined 50/50% or 25/75% (w/w), respectively, and compared to controls GelMA and GelMA with added chlorhexidine (CHX) at 0.2%. The different hydrogels were characterized using 1H-NMR spectroscopy and swelling behavior and tested in (1) Porphyromonas gingivalis t...
A key factor for dental implant success is a good sealing between the implant surface and both so... more A key factor for dental implant success is a good sealing between the implant surface and both soft (gum) and hard (bone) tissues. Surface nanotopography can modulate cell response through mechanotransduction. The main objective of this research was the development of nanostructured titanium (Ti) surfaces that promote both soft and hard tissue integration with potential application in dental implants. Nanostructured Ti surfaces were developed by electrochemical anodization—nanopores (NPs) and nanonets (NNs)—and characterized by atomic force microscopy, scanning electronic microscopy, and contact angle analysis. In addition, nanoparticle release and apoptosis activation were analyzed on cell culture. NP surfaces showed nanoparticle release, which increased in vitro cell apoptosis. Primary human gingival fibroblasts (hGFs) and human bone marrow mesenchymal stem cells (hBM-MSCs) were used to test cell adhesion, cytotoxicity, metabolic activity, and differentiation markers. Finally, cel...
Extracellular vesicles (EVs) present a great potential for the development of new treatments in t... more Extracellular vesicles (EVs) present a great potential for the development of new treatments in the biomedical field. To be used as therapeutics, many different sources have been used for EVs obtention, while only a few studies have addressed the use of platelet-derived EVs (pEVs). In fact, pEVs have been shown to intervene in different healing responses, thus some studies have evaluated their regenerative capability in wound healing or hemorrhagic shock. Even more, pEVs have proven to induce cellular differentiation, enhancing musculoskeletal or neural regeneration. However, the obtention and characterization of pEVs is widely heterogeneous and differs from the recommendations of the International Society for Extracellular Vesicles. Therefore, in this review, we aim to present the main advances in the therapeutical use of pEVs in the regenerative medicine field while highlighting the isolation and characterization steps followed. The main goal of this review is to portray the studi...
Extracellular Vesicles (EVs) are biological nanovesicles that play a key role in cell communicati... more Extracellular Vesicles (EVs) are biological nanovesicles that play a key role in cell communication. Their content includes active biomolecules such as proteins and nucleic acids, which present great potential in regenerative medicine. More recently, EVs derived from Platelet Lysate (PL) have shown an osteogenic capability comparable to PL. Besides, biomaterials are frequently used in orthopedics or dental restoration. Here, we provide a method to functionalize Ti surfaces with PL-derived EVs in order to improve their osteogenic properties. EVs are isolated from PL by size exclusion chromatography, and afterward Ti surfaces are functionalized with PL-EVs by drop casting. Functionalization is proven by EVs release and its biocompatibility by the lactate dehydrogenase (LDH) release assay.
Background: We aimed to evaluate the effect of low doses (LD) bone morphogenetic protein-2 (BMP2)... more Background: We aimed to evaluate the effect of low doses (LD) bone morphogenetic protein-2 (BMP2) and BMP4 micro-immunotherapy (MI) in two in vitro models of periodontal wound healing/regeneration. Methods: We first evaluated the effect of LD of BMP2 and BMP4 MI on a 2D cell culture using human gingival fibroblasts (hGF) under inflammatory conditions induced by IL1β. Biocompatibility, inflammatory response (Prostaglandin E2 (PGE2) release), collagen deposition and release of extracellular matrix (ECM) organization-related enzymes (matrix metalloproteinase-1 (MMP1) and metalloproteinase inhibitor 1 (TIMP1)) were evaluated after short (3 days) and long-term (24 days) treatment with BMP2 or BMP4 MI. Then, given the results obtained in the 2D cell culture, LD BMP4 MI treatment was evaluated in a 3D cell culture model of human tissue equivalent of gingiva (GTE) under the same inflammatory stimulus, evaluating the biocompatibility, inflammatory response and effect on MMP1 and TIMP1 release. Results: LD BMP4 was able to decrease the release of the inflammatory mediator PGE2 and completely re-establish the impaired collagen metabolism induced by IL1β treatment. In the 3D model, LD BMP4 treatment improved tissue viability compared with the vehicle, with similar levels to 3D tissues without inflammation. No significant effects were observed on PGE2 levels nor MMP1/TIMP1 ratio after LD BMP4 treatment, although a tendency to decrease PGE2 levels was observed after 3 days. Conclusions: LD BMP4 MI treatment shows anti-inflammatory and regenerative properties on hGF, and improved viability of 3D gingiva under inflammatory conditions. LD BMP4 MI treatment could be used on primary prevention or maintenance care of periodontitis. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.
In the last years, several studies testing commercial periodontal gels that contain chlorhexidine... more In the last years, several studies testing commercial periodontal gels that contain chlorhexidine (CHX) or other antibacterial agents, have raised concerns regarding their cytotoxicity in periodontal tissues. We aimed at comparing the biocompatibility but also the efficacy as regards to the antibacterial and wound healing ability of different commercial periodontal gels. In vitro human gingival fibroblasts (GF) and a 3D model of human tissue equivalents of gingiva (GTE) were used under inflammatory conditions to evaluate wound closure, cytotoxicity and gene expression. Antibacterial effects were also investigated on Porphyromonas gingivalis growth, viability and gingipain activity. In GF and in the bacterial study, we found cytotoxic effects on GF and a high inhibition on bacterial growth rate in gels containing CHX, asiaticoside, enoxolone, cetylpyridinium chloride, propolis and eugenol. Of the two gels that were non-cytotoxic, Syntoss Biogel (containing chondrontin sulfate) and Em...
Although coronary stents have improved the early and long-term consequences of arterial lesions, ... more Although coronary stents have improved the early and long-term consequences of arterial lesions, the prevention of restenosis and late stent thrombosis is key to prevent a new obstruction of the vessel. Here we aimed at improving the tissue response to stents through surface modification. For that purpose, we used two different approaches, the use of nanostructuration by electrochemical anodization and the addition of a quercitrin (QR) coating to the Ti surface. Four surfaces (Ti, NN, TiQR and NNQR) were characterized by atomic force microscopy, scanning electronic microscopy and contact angle analysis and QR content was evaluated by fluorescent staining. Cell adhesion, cytotoxicity, metabolic activity and nitric oxide (NO) production was evaluated on primary human umbilical cord endothelial cells (HUVECs). Platelet adhesion, hemolysis rate and Staphylococcus epidermidis CECT 4184 adhesion at 30 min were analyzed. Nanostructuration induced an increase on surface roughness, and QR co...
(1) One strategy to improve the outcome of orthopedic implants is to use porous implants with the... more (1) One strategy to improve the outcome of orthopedic implants is to use porous implants with the addition of a coating with an antibacterial biomolecule. In this study, we aimed to produce and test the biocompatibility, the osteopromotive (both under normal conditions and under a bacterial challenge with lipopolysaccharide (LPS)) and antibacterial activities of a porous Ti-6Al-4V implant coated with the flavonoid quercitrin in vitro. (2) Porous Ti-6Al-4V implants were produced by 3D printing and further functionalized with quercitrin by wet chemistry. Implants were characterized in terms of porosity and mechanical testing, and the coating with quercitrin by fluorescence staining. Implant biocompatibility and bioactivity was tested using MC3T3-E1 preosteoblasts by analyzing cytotoxicity, cell adhesion, osteocalcin production, and alkaline phosphatase (ALP) activity under control and under bacterial challenging conditions using lipopolysaccharide (LPS). Finally, the antibacterial properties of the implants were studied using Staphylococcus epidermidis by measuring bacterial viability and adhesion. (3) Porous implants showed pore size of about 500 µm and a porosity of 52%. The coating was homogeneous over all the 3D surface and did not alter the mechanical properties of the Young modulus. Quercitrin-coated implants showed higher biocompatibility, cell adhesion, and osteocalcin production compared with control implants. Moreover, higher ALP activity was observed for the quercitrin group under both normal and bacterial challenging conditions. Finally, S. epidermidis live/dead ratio and adhesion after 4 h of incubation was lower on quercitrin implants compared with the control. (4) Quercitrin-functionalized porous Ti-6Al-4V implants present a great potential as an orthopedic porous implant that decreases bacterial adhesion and viability while promoting bone cell growth and differentiation.
Extracellular vesicles (EVs) have been recently identified as vital components of cell-based ther... more Extracellular vesicles (EVs) have been recently identified as vital components of cell-based therapies based on the observation that conditioned media from cultured stromal cells reproduce some of the beneficial effects of intact cells. In order to obtain clinically active EVs derived from Mesenchymal Stromal Cells (MSCs) different procedures have been reported in the literature. Usually, non-confluent cells are incubated with culture medium for 48 h either with EV-depleted Fetal Bovine Serum (FBS) or without FBS. Our aim was to compare the effects of EVs isolated by ultracentrifugation from human umbilical cord MSC conditioned media obtained using these two conditions: with EV-depleted FBS (UC) or without FBS (UCw/o) on the mRNA expression levels of extracellular matrix related genes using the mouse chondrogenic cell line ATDC-5. We observed a deleterious effect on chondrogenic cells treated with UCw/o, showing higher mRNA expression levels of different metalloproteinases and decor...
Journal of Tissue Engineering and Regenerative Medicine, 2020
The demineralized bone matrix (DBM) is the most widely used bone allograft, which is obtained by ... more The demineralized bone matrix (DBM) is the most widely used bone allograft, which is obtained by removing the mineral component of bone, leading to exposure of the proteins responsible for osteoinduction. For clinical use, DBM shall be formulated with a carrier that provides consistency and improves its osteoinduction. In this study, three DBM formulations with glycerol (Gly), hyaluronic acid (HA) and gelatin methacryloyl (GelMA) were evaluated measuring their physicochemical properties (microstructure, compressive strength and serum cohesivity), and their osteoinductive capacity both in vitro using C2C12 cells and umbilical cord human mesenchymal stem cells and in vivo in an ectopic bone formation model in athymic mice. To assess the effectiveness of DBM in vitro in inducing the differentiation into osteoblasts, alkaline phosphatase (ALP) activity was assessed in combination with a cytotoxicity assay. In vivo, new bone formation was assessed by histological and radiological analysis. In the compression and in the serum cohesive assays, the GelMA DBM formulation showed its superiority over the other formulations. In addition, GelMA showed a more compact structure analyzed by scanning electron microscopy. Higher cell toxicity was observed on Gly formulations in vitro, while GelMa and HA showed very low toxicity. All formulations significantly improved ALP activity compared to control. In the in vivo studies, GelMA showed the greatest osteoinductive potential with a higher percentage of new bone and bone marrow formation. Our results suggest GelMA is useful as a carrier for DBM designed to promote the formation of the new bone.
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Papers by Joana M Ramis