A Single Nucleotide Polymorphism in the RASGRF2 Gene Is Associated with Alcoholic Liver Cirrhosis in Men

Background Genetic polymorphisms in the RAS gene family are associated with different diseases, which may include alcohol-related disorders. Previous studies showed an association of the allelic variant rs26907 in RASGRF2 gene with higher alcohol intake. Additionally, the rs61764370 polymorphism in the KRAS gene is located in a binding site for the let-7 micro-RNA family, which is potentially involved in alcohol-induced inflammation. Therefore, this study was designed to explore the association between these two polymorphisms and susceptibility to alcoholism or alcoholic liver disease (ALD). Methods We enrolled 301 male alcoholic patients and 156 healthy male volunteers in this study. Polymorphisms were genotyped by using TaqMan® PCR assays for allelic discrimination. Allelic and genotypic frequencies were compared between the two groups. Logistic regression analysis was performed to analyze the inheritance model. Results The A allele of the RASGRF2 polymorphism (rs26907) was significantly more prevalent among alcoholic patients with cirrhosis (23.2%) compared to alcoholic patients without ALD (14.2%). This difference remained significant in the group of patients with alcohol dependence (28.8% vs. 14.3%) but not in those with alcohol abuse (15.1% vs. 14.4%). Multivariable logistic regression analysis showed that the A allele of this polymorphism (AA or GA genotype) was associated with alcoholic cirrhosis both in the total group of alcoholics (odds ratio [OR]: 2.33, 95% confidence interval [CI]: 1.32–4.11; P = 0.002) and in the group of patients with alcohol dependence (OR: 3.1, 95% CI: 1.50–6.20; P = 0.001). Allelic distributions of the KRAS polymorphism (rs61764370) did not differ between the groups. Conclusions To our knowledge, this genetic association study represents the first to show an association of the RASGRF2 G>A (rs26907) polymorphism with ALD in men, particularly in the subgroup of patients with AD. The findings suggest the potential relevance of the RAS gene family in alcoholism and ALD

Técnicas y tecnologías en hidrología médica e hidroterapia

El objetivo del presente informe es difundir entre los profesionales médicos la información que contribuya a orientarles en la materia, conocer las aplicaciones terapéuticas o rehabilitadoras que pueden ofrecer los distintos centros termales y explorar su interacción con los tratamientos médicos habituales a los que suelen estar sometidos los usuarios de estos programas de termalismo

Molecular and histological studies of bladder wound healing in a rodent model

The field of regenerative medicine encounters different challenges. The success of tissue-engineered implants is dependent on proper wound healing. Today, the process of normal urinary bladder wound healing is poorly characterized. We aspired to explore and elucidate the natural response to injury in an in vivo model in order to further optimize tissue regeneration in future studies. In this study, we aimed to characterize histological and molecular changes during normal healing in a rat model by performing a standardized incisional wound followed by surgical closure. We used a rodent model (n = 40) to follow the healing process in the urinary bladder for 28 days. Surgical exposure of the bladder without incision (n = 40) was performed in controls. Histological characterization and western blot analyses of proteins was carried out using specific staining and markers for inflammation, proliferation, angiogenesis, and tissue maturation. For the molecular characterization of gene expression total RNA was collected for RT2-PCR in wound healing pathway arrays. Analysis of histology revealed distinct, but overlapping, phases of healing with a local inflammatory response (days 1-8) simultaneous with a rapid formation of granulation tissue and proliferation (days 2-8). We also identified significant changes in gene expression related to inflammation, proliferation, and extracellular matrix formation. Healing of an incisional wound in a rodent urinary bladder demonstrated that all the classical phases of wound healing: hemostasis, inflammation, proliferation followed by tissue maturation were present. Our data suggest that the bladder and the skin share similar molecular signaling during wound healing, although we noted differences in the duration of each phase compared to previous studies in rat skin. Further studies will address whether our findings can be extrapolated to the human bladder

Queratinocitos derivados de piel humana modificados por el vector retroviral FOCH 29-NeoR

<p class="MsoNormal"><span style="font-size: 8.5pt; font-family: Arial">En este protocolo se evaluará la eficiencia de la transducción mediada por el vector retroviral FOCH 29-NeoR derivado del virus de Friend; éste ha mostrado una alta eficiencia en la transducción, tanto de células madres hematopoyéticas como de otras líneas celulares. Se medirá su eficiencia de transducción en cultivos primarios de queratinocitos, derivados de biopsias de piel humana o de sobrantes de procedimientos quirúrgicos como circuncisiones, mastectomías y cirugía cosmética de pacientes que consultan el Hospital Universitario San Vicente de Paul, Hospital la María, la Clínica del Rosario y la Clínica León XIII.</span></p> <p class="MsoNormal"><span style="font-size: 8.5pt; font-family: Arial">Las muestras de piel se procesarán en un lapso no superior a 12 horas, se eliminará el exceso de dermis y tejido conectivo por digestión con dispasa (0.6-2.4 U/ml a 37</span><span style="font-size: 8.5pt; font-family: Symbol">°</span><span style="font-size: 8.5pt; font-family: Arial">C) durante 1 hora. Las muestras serán lavadas con PBS, antibiótico (penicilina + estreptomicina) y se cortarán en fragmentos de 1-2 mm; después de 2-3 horas de digestión con tripsina-EDTA (0.25%) las células serán resuspendidas en KGM (Medio de crecimiento para queratinocitos) y se sembrarán a una concentración de 10</span><span style="font-size: 5pt; font-family: Arial">5 </span><span style="font-size: 8.5pt; font-family: Arial">- 3x10</span><span style="font-size: 5pt; font-family: Arial">5 </span><span style="font-size: 8.5pt; font-family: Arial">células por plato de 100 mm; se incubarán a 37</span><span style="font-size: 8.5pt; font-family: Symbol">°</span><span style="font-size: 8.5pt; font-family: Arial">C, 5% CO</span><span style="font-size: 5pt; font-family: Arial">2 </span><span style="font-size: 8.5pt; font-family: Arial">con cambios de medio 2-3 veces por semana. Se harán subcultivos con el fin de expandirlos y congelar una parte de las células (banco).</span></p> <p class="MsoNormal"><span style="font-size: 8.5pt; font-family: Arial">Posteriormente, los queratinocitos en cultivo serán modificados con el vector retroviral anfotrópico FOCH 29-NeoR ya sea por cocultivo con la línea productora del virus Clon 26, o por infección con sobrenadante filtrado (0.45mm) producido por la misma, en presencia de polybren. </span></p> <p class="MsoNormal"><span style="font-size: 8.5pt; font-family: Arial">La eficiencia de la transducción se determinará mediante reacción en cadena de la polimerasa (PCR) con el ADN de las células post transducción, con cebadores específicos del gen de resistencia a la neomicina (NeoR); se incluirán tanto controles negativos (células no transducidas ) como positivos (plásmidos que contienen el gen NeoR o células de la línea productora del virus). Para demostrar funcionalmente la expresión del gen vectorizado se harán ensayos de resistencia a la G418 (500-1000 </span><span style="font-size: 8.5pt; font-family: Symbol">m</span><span style="font-size: 8.5pt; font-family: Arial">g/ml).</span></p> <p class="MsoNormal"><span style="font-size: 8.5pt; font-family: Arial">Confirmar que el vector FOCH29-NeoR transluce eficientemente los queratinocitos, permitirá utilizarlo posteriormente para vectorizar genes de interés para el tratamiento de algunas enfermedades de la piel o bien factores de crecimiento tales como el factor de crecimiento epidérmico (EGF) y el factor de crecimiento derivado de plaquetas (PDGF), potenciadores de los procesos de epitelialización.</span></p> <p class="MsoNormal"><span style="font-size: 8.5pt; font-family: Arial">Este trabajo sentará las bases para el desarrollo paulatino de esta tecnología en nuestro medio.</span><span style="font-size: 10pt; font-family: Arial"></span></p&gt

A perioperative layered autologous tissue expansion graft for hollow organ repair

Tissue engineering has not been widely adopted in clinical settings for several reasons, including technical challenges, high costs, and regulatory complexity. Here, we introduce the Perioperative Layered Autologous Tissue Expansion graft (PLATE graft), a composite biomaterial and collagen-reinforced construct with autologous epithelium on one side and smooth muscle tissue on the other. Designed to mimic the structure and function of natural hollow organs, the PLATE graft is unique in that it can be produced in a standard operating theatre and is cost-effective. In this proof-of-principle study, we test its regenerative performance in eight different organs, present biomechanical and permeability tests, and finally explore its in vivo performance in live rabbits.</p

A perioperative layered autologous tissue expansion graft for hollow organ repair

Tissue engineering has not been widely adopted in clinical settings for several reasons, including technical challenges, high costs, and regulatory complexity. Here, we introduce the Perioperative Layered Autologous Tissue Expansion graft (PLATE graft), a composite biomaterial and collagen-reinforced construct with autologous epithelium on one side and smooth muscle tissue on the other. Designed to mimic the structure and function of natural hollow organs, the PLATE graft is unique in that it can be produced in a standard operating theatre and is cost-effective. In this proof-of-principle study, we test its regenerative performance in eight different organs, present biomechanical and permeability tests, and finally explore its in vivo performance in live rabbits

Insights into cellular behavior and micromolecular communication in urothelial micrografts

Abstract Autologous micrografting is a technique currently applied within skin wound healing, however, the potential use for surgical correction of other organs with epithelial lining, including the urinary bladder, remains largely unexplored. Currently, little is known about the micrograft expansion potential and the micromolecular events that occur in micrografted urothelial cells. In this study, we aimed to evaluate the proliferative potential of different porcine urothelial micrograft sizes in vitro, and, furthermore, to explore how urothelial micrografts communicate and which microcellular events are triggered. We demonstrated that increased tissue fragmentation subsequently potentiated the yield of proliferative cells and the cellular expansion potential, which confirms, that the micrografting principles of skin epithelium also apply to uroepithelium. Furthermore, we targeted the expression of the extracellular signal-regulated kinase (ERK) pathway and demonstrated that ERK activation occurred predominately at the micrograft borders and that ERK inhibition led to decreased urothelial migration and proliferation. Finally, we successfully isolated extracellular vesicles from the micrograft culture medium and evaluated their contents and relevance within various enriched biological processes. Our findings substantiate the potential of applying urothelial micrografting in future tissue-engineering models for reconstructive urological surgery, and, furthermore, highlights certain mechanisms as potential targets for future wound healing treatments