Ulmoplus® increases FGF-2 expression and promote burn wound healing / Ulmoplus® incrementa la expresión de FGF-2 y promueve la cicatrización de heridas por quemadura

SUMMARY: The role of fibroblast growth factor (FGF) in the wound repair process has been described, particularly during the stimulation of angiogenesis in the proliferative phase. Ulmo honey (Eucryphia cordifolia) has demonstrated important bactericidal, fungicidal, antioxidant and healing properties. Ulmoplus®, a honey-based medicinal product, accelerates healing time and promotes fibroblast activation and collagen fiber production. The aim of this study was to evaluate the immunohistochemical expression of endogenous FGF-2 in burns treated with Ulmoplus® vs. the gold standard (hydrogel-tull), and Ulmo honey as the first step to clarifying the regulatory mechanism of this therapeutic option. 15 adult guinea pigs (Cavia porcellus) were used, divided into three groups: C+: positive control treated with hydrogel-tull (gold standard); E1: experimental group treated with Ulmo; and E2: experimental group treated with Ulmoplus®. A deep uniform burn was made under anesthetic that covered 1 cm2 of the skin on the back of each animal. The daily treatments were administered with hydrogel-tull, Ulmo honey or Ulmoplus®, where appropriate. On treatment day 10 the biopsies were taken and processed for histological and immunohistochemical analysis. The immunolabeling was quantified through an integrated optical density (IOD) analysis, expressed as lum/ µm2 using the Image- ProPremier 9.1 software. The endogenous expression of FGF-2 was consistent with the stage of healing presented by the study groups, varying in its localization and IOD intensity. The E2 group presented a significantly higher IOD (60053.55 ± 59877.84 lum/µm2) than the C+ (p = 0.001) and E1 (p = 0.001) groups. Our findings indicated that the medicinal Ulmoplus® honey is effective at facilitating wound closing, and the promoter effect of FGF-2 expression accelerated the healing process compared to the treatments with hydrogel-tull (gold standard) and Ulmo honey.

RESUMEN: Se ha descrito el papel del factor de crecimiento fibroblaástico (FGF) en el proceso de reparación de heridas, particularmente durante la estimulación de la angiogénesis en la fase proliferativa. La miel de Ulmo (Eucryphia cordifolia) ha demostrado importantes propiedades bactericidas, fungicidas, antioxidantes y cicatrizantes. Ulmoplus®, un medicamento a base de miel, acelera el tiempo de cicatrización y promueve la activación de fibroblastos y la producción de fibras colágenas. El objetivo de este estudio fue evaluar la expresión inmunohistoquímica del FGF-2 endógeno en quemaduras tratadas con Ulmoplus® frente al gold standard (hidrogel-tull) y miel de Ulmo como primer paso para esclarecer el mecanismo regulador de esta opción terapéutica. Se utilizaron 15 cobayas adultos (Cavia porcellus), divididos en tres grupos: C +: control positivo tratado con hidrogel-tull (gold standard); E1: grupo experimental tratado con Ulmo; y E2: grupo experimental tratado con Ulmoplus®. Se realizó una quemadura profunda uniforme bajo anestesia que cubrió 1 cm2 de la piel del lomo de cada animal. Los tratamientos diarios se administraron con hidrogel-tull, miel de Ulmo o Ulmoplus®, en cada caso. El día 10 de tratamiento se tomaron biopsias y se procesaron para análisis histológico e inmunohistoquímico. El inmunomarcaje se cuantificó mediante análisis de densidad óptica integrado (DOI), expresado como lum/µm2 utilizando el software Image-ProPremier 9.1. La expresión endógena de FGF-2 fue consistente con la etapa de cicatrización presentada por los grupos de estudio, variando en su localización e intensidad de DOI. El grupo E2 presentó una DOI significativamente mayor (60053,55 ± 59877,84 lum/µm2) que los grupos C + (p = 0,001) y E1 (p = 0,001). Nuestros hallazgos indicaron que la miel de uso médico Ulmoplus® es eficaz para facilitar el cierre de heridas, y el efecto promotor de la expresión de FGF-2 aceleró el proceso de curación en comparación con los tratamientos con hidrogel-tull (gold standard) y miel de Ulmo.

Rol de la osteocalcina más allá del hueso / Role of osteocalcin beyond bone

Los hallazgos osteológicos se intensi!caron en los últimos años. Se demostró que el esqueleto se comporta, además de sus funciones clásicas, como un órgano de secreción endocrina que sintetiza al menos dos hormonas: el factor de crecimiento de !broblastos 23 (FGF-23) y la osteocalcina (Ocn). La Ocn es un péptido pequeño que contiene 3 residuos de ácido glutámico. Estos residuos se carboxilan postraduccionalmente, quedando retenida en la matriz ósea. La forma decarboxilada en el primer residuo de ácido glutámico (GluOcn) fue reportada por poseer efectos biológicos; la resorción ósea es el mecanismo clave para su bioactivación. La presente revisión se centra en los conocimientos actuales sobre la función hormonal de la Ocn. A la fecha se reporta que la Ocn regularía el metabolismo energético aumentando la proliferación de células ` pancreáticas, y la secreción de insulina y de adiponectina. Sobre el músculo esquelético actuaría favoreciendo la absorción y el catabolismo de nutrientes. La función reproductiva masculina estaría regulada mediante el estímulo a las células de Leydig para sintetizar testosterona; en el desarrollo cerebral y la cognición, la Ocn aumentaría la síntesis de neurotransmisores monoaminados y disminuiría el neurotransmisor inhibidor GABA. Si bien son indispensables mayores evidencias para dilucidar los mecanismos reguladores por medio de los cuales actuaría la Ocn, los resultados enumerados en los distintos estudios experimentales establecen la importancia de este novedoso integrante molecular. Dilucidar su rol dentro de estos procesos interrelacionados en seres humanos abriría la posibilidad de utilizar a la Ocn en el tratamiento de enfermedades endocrino-metabólicas. (AU)

Osteological !ndings have intensi!ed in recent years. The skeleton behaves as an endocrine secretion organ that synthesizes at least two hormones: osteocalcin (Ocn) and !broblast growth factor 23 (FGF-23). Ocn is a small peptide that contains 3 glutamic acid residues. After translation, these residues are carboxylated to make possible its retention into the bone matrix. Decarboxylation on the !rst glutamic acid residue (GluOcn) has been reported to have biological effects. Bone resorption is the key mechanism for its bioactivation. This review focuses on current knowledge on Ocn hormonal function. It has been reported that Ocn regulates energy metabolism by increasing the proliferation of pancreatic ` cells, and the secretion of insulin and adiponectin. On the skeletal muscle, it may act by favoring the absorption and catabolism of nutrients. Male reproductive function might be regulated by stimulating Leydig cells to synthesize testosterone. Regarding brain development and cognition, Ocn would increase monoamine neurotransmitters synthesis and decrease inhibitory neurotransmitter GABA. Although more evidence is needed to elucidate the regulatory mechanisms of Ocn, different experimental studies establish the importance of this novel molecular mediator. Clarifying its role within interrelated processes in humans, might open the possibility of using Ocn in different treatments of endocrine-metabolic diseases. (AU)

Pravastatin induces cell cycle arrest and decreased production of VEGF and bFGF in multiple myeloma cell line / Pravastatina induz parada no ciclo celular e diminuição na produção de VEGF e bFGF em linhagem de Mieloma Multiplo

Abstract Multiple myeloma (MM) is a B cell bone marrow neoplasia characterized by inflammation with an intense secretion of growth factors that promote tumor growth, cell survival, migration and invasion. The aim of this study was to evaluate the effects of pravastatin, a drug used to reduce cholesterol, in a MM cell line.Cell cycle and viability were determinate by Trypan Blue and Propidium Iodide. IL6, VEGF, bFGF and TGFβ were quantified by ELISA and qRT-PCR including here de HMG CoA reductase. It was observed reduction of cell viability, increase of cells in G0/G1 phase of the cell cycle and reducing the factors VEGF and bFGF without influence on 3-Methyl-Glutaryl Coenzyme A reductase expression.The results demonstrated that pravastatin induces cell cycle arrest in G0/G1 and decreased production of growth factors in Multiple Myeloma cell line.

Resumo O Mieloma Múltiplo é uma neoplasia de linfócitos B da medula óssea, caracterizada por inflamação com uma intensa secreção de fatores de crescimento que promovem o aumento do volume do tumor, sobrevivência celular, migração e invasão. O objetivo deste estudo foi avaliar os efeitos da pravastatina, uma droga usada para reduzir o colesterol, em um linhagem de MM. O ciclo celular e viabilidade foram determinadas por Trypan Blue e iodeto de propídio. IL6, VEGF, bFGF e TGF foram quantificadas por ELISA e qRT-PCR, incluindo aqui de HMG CoA redutase. Observou-se a redução da viabilidade das células, aumento de células na fase G0/G1 do ciclo celular e redução no VEGF e bFGF, sem influência na expressão da enzima 3-Metil-Glutaril Coenzima A redutase. Os resultados demonstraram que a pravastatina induz parada no ciclo celular em G0/G1 e diminuição da produção de fatores de crescimento em várias linhas de células de Mieloma.

Metabolismo mineral en niños en diálisis peritoneal crónica / Mineral metabolism in patients on chronic peritoneal dialysis

Introduction: Children with chronic kidney disease (CKD) and receiving peritoneal dialysis (PD) have disorders of mineral metabolism that impact their growth, survival and cardiovascular functions. New molecular markers offer a better understanding of the pathophysiology of this disease. Objective: To characterize some components of mineral metabolism, with emphasis on FGF23/Klotho and cardiovascular functions (CV) of these patients. Patients and Method: Prospective observational cohort study. Exclusion criteria: serum 25 (OH) vitamin D < 20 ng/ml, peritonitis within the last two months and active nephrotic syndrome. Calcemia, phosphemia, parathyroid hormone (PTH), 25 (OH) vitD3, 1.25 (OH) vitD3, FGF23 and Klotho in plasma were measured. FGF23 and Klotho were quantified in healthy children as a control group. Echocardiography was performed calculating the left ventricular mass index (LVMI). Descriptive statistics analysis, Pearson correlation coefficient for association among variables and multivariate analysis were conducted. Results: 33 patients, 16 males, aged between 1.2 and 13.4 years were included. Age of onset for PD: 7.3 +/- 5.0 years, time receiving PD: 13.5 +/- 14.5 months. The plasma concentration of 25 (OH) vitD3 was 34.2 +/- 6.3 pg/ml. Calcemia and phosphemia values were 9.8 ± 0.71 and 5.4 +/- 1.0 mg/dl respectively. PTH was 333 +/- 287 pg/ml. FGF23 in plasma was 225.7 +/- 354.3 pg/ml and Klotho 131.6 +/- 72 pg/ml, and in the controls ( n = 16 ), it was 11.9 +/- 7.2 pg/ml and 320 +/- 119 pg/ml, respectively. The residual and total dose of dialysis (KtV) was 1.6 +/- 1.3 and 2.9 +/- 1.6, respectively. FGF23 levels significantly correlated with calcium (p < 0.001, r = 0.85), and inversely with residual KtV, showing no relationship with phosphemia. Klotho level correlated negatively with residual KtV and also, it showed a negative association with chronological age and age at onset of PD. LVMI > 38 g/m² was confirmed in 20/28 patients...

Introducción: Los niños portadores de Enfermedad renal crónica (ERC) en diálisis peritoneal (DP) presentan alteraciones del metabolismo mineral que afectan su crecimiento, estado cardiovascular y sobrevida. Nuevos marcadores moleculares representan una mejor comprensión de la fisiopatología de esta enfermedad. Objetivo: Caracterizar componentes del metabolismo mineral, con énfasis en FGF23/Klotho, y estado cardiovascular (CV) en este grupo de pacientes. Pacientes y Método: Estudio prospectivo observacional. Criterios de exclusión: niveles de 25 (OH) vitamina D < 20 ng/ml, peritonitis hasta 2 meses previos y síndrome nefrótico activo. Se midió calcemia, fosfemia, paratohormona (PTH), 25 (OH) vitD3, 1,25 (OH) vitD3, FGF23 y Klotho en plasma. Se cuantificó FGF23 y Klotho en niños sanos como grupo control. Se efectuó ecocardiografía, calculándose el índice de masa ventricular izquierda (IMVI). Se realizó análisis estadístico descriptivo, coeficiente de correlación de Pearson para asociación entre variables y análisis multivariado. Resultados: Se incluyeron 33 pacientes, 16 varones, edad 1,2 a 13,4 años. Edad de inicio de DP: 7,3 +/- 5,0 años, tiempo en DP: 13,5 +/- 14,5 meses. El nivel plasmático de 25 (OH) vitD3 fue 34,2 +/- 6,3 pg/ml. Los valores de calcemia y fosfemia fueron 9,8 +/- 0,71 y 5,4 +/- 1,0 mg/dl respectivamente. La PTH fue de 333 +/- 287 pg/ml. El FGF23 en plasma fue de 225,7 +/- 354,3 pg/ml y Klotho 131,6 +/- 72 pg/ml, y en los controles (n = 16) fue de 11,9 +/- 7,2 pg/ ml y 320 +/- 119 pg/ml, respectivamente. La dosis de diálisis (KtV) residual y total fue de 1,6 +/- 1,3 y 2,9 +/- 1.6, respectivamente. El nivel de FGF23 se correlacionó significativamente con la calcemia (p < 0,001, r = 0,85), e inversamente con el KtV residual, sin mostrar relación con la fosfemia. El nivel de Klotho...

Light-induced retinal injury enhanced neurotrophins secretion and neurotrophic effect of mesenchymal stem cells in vitro / Lesão de retina induzida por luz aumentou a secreção de neurotrofinas e o efeito neurotrófico das células primordiais mesenquimais in vitro

PURPOSE: To investigate neurotrophins expression and neurotrophic effect change in mesenchymal stem cells (MSCs) under different types of stimulation. METHODS: Rats were exposed in 10,000 lux white light to develop light-induced retinal injury. Supernatants of homogenized retina (SHR), either from normal or light-injured retina, were used to stimulate MSCs. Quantitative real time for polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) were conducted for analysis the expression change in basic fibroblast growth factor (bFGF), brain-derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF) in MSCs after stimulation. Conditioned medium from SHR-stimulated MSCs and control MSCs were collected for evaluation their effect on retinal explants. RESULTS: Supernatants of homogenized retina from light-injured rats significantly promoted neurotrophins secretion from MSCs (p<0.01). Conditioned medium from mesenchymal stem cells stimulated by light-injured SHR significantly reduced DNA fragmentation (p<0.01), up-regulated bcl-2 (p<0.01) and down-regulated bax (p<0.01) in retinal explants, displaying enhanced protective effect. CONCLUSIONS: Light-induced retinal injury is able to enhance neurotrophins secretion from mesenchymal stem cells and promote the neurotrophic effect of mesenchymal stem cells.

OBJETIVO: Investigar a expressão de neurotrofinas e mudança no efeito neurotrófico de células-tronco mesenquimais (MSCs) sob diferentes tipos de estimulação. MÉTODOS: Os ratos foram expostos em 10.000 lux de luz branca para desenvolver a lesão da retina induzida por luz. Os sobrenadantes de homogeneizado de retina (SHR) quer a partir de retina normal ou da lesada por luz, foram usados para estimular as células-tronco mesenquimais. O RT-PCR quantitativa e ELISA foram realizados para análise da alteração de expressão do fator básico de crescimento de fibroblastos (bFGF), do fator neurotrófico derivado do cérebro (BDNF) e do fator neurotrófico ciliar (CNTF) em MSCs após a estimulação. O meio condicionado de células-tronco mesenquimais estimuladas por SHR e controles foram coletadas para avaliação de seu efeito sobre os explantes de retina. RESULTADOS: SHR de retinas de rato lesadas por luz promoveram aumento significativo de secreção de neurotrofinas em MSCs (p<0,01). O meio condicionado de SHR lesado por luz reduziu significativamente a fragmentação do DNA de MSCs (p<0,01), elevação de Bcl-2 (p<0,01) e redução de bax (p<0,01) em explantes de retina, mostrando um aumento do efeito protetor. CONCLUSÕES: A lesão da retina induzidos pela luz é capaz de aumentar a secreção de neurotrofinas e promover o efeito neurotrófico de células-tronco mesenquimais.

Increased fibroblast telomerase expression precedes myofibroblast α-smooth muscle actin expression in idiopathic pulmonary fibrosis

OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast α-smooth muscle actin expression and the tissue expression of inter leu kin-4, transforming growth factor-β, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-β expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast α-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast α-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast α-smooth muscle actin expression predominates in areas of late remodeling. These events seem to be regulated by basic fibroblast growth factor and interleukin-4 tissue expression, respectively.

Klotho and the Aging Process

The klotho gene was originally identified as a putative age-suppressing gene in mice that extends life span when overexpressed. It induces complex phenotypes resembling human premature aging syndromes when disrupted. The gene was named after a Greek goddess Klotho who spun the thread of life. Since then, various functional aspects of the klotho gene have been investigated, leading to the identification of multiple novel endocrine axes that regulate various metabolic processes and an unexpected link between mineral metabolism and aging. The purposes of this review were to overview recent progress on Klotho research and to discuss a novel aging mechanism.

Alteraciones del metabolismo óseo y mineral en enfermedad renal crónica pre-diálisis / Disturbance in mineral and bone metabolism in chronic kidney disease

Las alteraciones del metabolismo mineral y óseo se inician tempranamente en el desarrollo de la enfermedad renal crónica (ERC), aunque las modificaciones en los niveles séricos de calcio y fósforo sólo se pueden apreciar en las etapas finales de la ERC debido a los mecanismos adaptativos que ocurren inicialmente. Con la pérdida de función renal ocurre retención de fósforo, lo que estimula la producción del factor de crecimiento fibroblástico 23 (FGF-23) por el hueso...

Disturbances in mineral and bone metabolism are highly prevalent among chronic kidney disease (CKD) patients and occur early in the development of the disease. However, hyperphosphatemia and hypocalcemia will be present only in the late stage as consequence of adaptative mechanisms. Recent studies indicate that in early CKD, the phosphate retention induces the production of the phosphaturichormone fibroblast growth factor 23 (FGF-23)...

Factores que regulan la morfogénesis y el crecimiento mandibular humano / Factors that regulate the morphogenesis and human mandibular growth

La regulación del crecimiento y desarrollo cráneo-facial está controlada por una serie de interacciones celulares y con la matriz extracelular que estimulan los procesos de proliferación y diferenciación. De fundamental importancia es la cresta neural, una población de células especializadas de células progenitoras que generan los huesos, cartílagos y tejido conectivo de la región. La mandíbula se forma por osificación membranosa en el mesénquima del primer arco faríngeo, pero desarrolla cartílagos secundarios como centros de crecimiento en el cóndilo, en el proceso coronoídeo, en el ángulo mandibular y en la sutura intermaxilar (sínfisis). Estos cartílagos difieren en su origen, su estructura histológica y su respuesta a factores hormonales, metabólicos y mecánicos con respecto a los cartílagos de los huesos largos. Debido a que las células proliferativas son mesenquimáticas y no cartilaginosas, los mecanismos celulares y moleculares que regulan el crecimiento en los cartílagos secundarios, son todavía muy poco conocidos. Los productos génicos BMP (proteina morfogenética de hueso), Ihh (Indian hedgehog), FGF (factor de crecimiento de fibroblastos), Sox-9 y VEGF (factor de crecimiento vascular endotelial) son de gran importancia en el crecimiento mandibular. Este trabajo resume la información reciente acerca de los factores de crecimiento y factores de transcripción, potenciales reguladores del proceso de osificación membranosa y del crecimiento de los cartílagos secundarios de la mandíbula.

Regulation of growth and craniofacial development is controlled by the interactions of cells with each other and with the extracellular environment through signal transduction pathways that control the differentiation process by stimulating proliferation or causing cell death. Of fundamental importance to mandibular development is the neural crest, a specialized population of stem and progenitor cells which generate the bone, cartilage and conjunctive tissue of the first branchial arch. The mandible arises by intramembranous ossification, but develops secondary cartilages as growth centers. Secondary cartilages of the mandible arise in the condylar process, in the coronoid process, angular process of the mandible, and in the intermandibular suture (mental symphysis). These are different, not only in their origins, but in their histologic organization and in their response to hormonal and mechanical factors with articular cartilages of long bones. Because the cells that divide to effect growth and adaptation in these cartilages are of perichondrial/periosteal rather than chondrogenic origin, the cellular and molecular mechanisms that regulate their growth are only beginning to be understood. The main differences of secondary cartilages from cartilages of the limbs and cranial base are, that condylar condroblasts arise from undifferentiated conjuntive cells and the appearance of vascular canals that cross cartilage perpendicularly and connect with the ossification zone. Collagen type I seems to be more important in this process than collagen type II. BMP signaling maintains regulatory roles in skeletons and skeletal growth. Indian hedgehog, Sox-9, fibroblastic growth factor (FGF) and vascular endothelial growth factor (VEGF), are also important in mandibular growth. This article summarizes information regarding growth factors and transcription proteins that are potential growth regulators in these secondary cartilages.

Induction of mitogenesis by synthetic peptides related to the structure of human fibroblast growth factor-1

The synthetic peptide Ac-WFVGLKKNGSSKRGPRT-NH2,related to the human FGF-1 sequence, was shown to be mitogenic upon Balb/c 3T3 fibroblasts in culture (ED50=10-20 muM) and to compete with the growth factor for cellular binding (ID50 = 30-50 muM). The results described suggest that the mitogenic activity of the peptide is dependent on the presence of the residues 122-127 (WFVGLK) in its structure. Also, its affinity for the cellular receptors seems to be dependent on the presence of residues that are important for FGF-heparin binding such as K127, K133 and R137.

Structure and function of fibroblast growth factor

We describe some structural requirements od the fibroblast growth factor (FGF) signaling system for the stimulation of the mitogenic response in terms of the design, synthesis and mitogenic activity of linear peptides related to the human FGF-1 sequence and the structural requirements of heparin for the potentiation of the mitogenic activity of FGF-1. The best mitogenic peptide we have synthesized so far is Ac-WFVGLKKNGSSKRGPRT-NH2, that has been shown: 1)to bind to heparin-Sepharose columns with moderate affinity, requiring about 0.5 M NaCl to be eluted from the resin; 2) to be mitogenic upon BALB/c 3T3 fibroblasts in culture (ED50=10-20 muM) and 3)to compete with human FGF-1 for cellular binding (ID50=30-50 muM). The potentiating activity of heparin upon FGF-1 has shown to be dependent on the oligosaccharide size, degree of sulfation and carboxylation. Apparently, these same requirements hold for the heparan sulfate molecules. Based on the reported studies, ee propose some important requirements of an oligosaccharide to potentiate FGF-1 mitogenic activity: 1) to have a minimum of twelve units, organized as disaccharides where one of the units is a uronic acid and the second is glycosamine; 2) to have at least one iduronic acid sulfated at position 2 and 3) to have N-sulfated glycosamines, preferentially 6-O-sulfated. To have groups of negative charges is not enough: they need to be localized in a correct conformation.