Detection of embryonic stem cell markers in adult human adipose tissue-derived stem cells.

BACKGROUND: Bone marrow transplantation is already an established therapy, which is now widely used in medicine to treat leukemia, lymphoma, and several inherited blood disorders. The culture of multilineage cells from easily available adipose tissue is another source of multipotent mesenchymal stem cells, and is referred to as adipose tissue-derived stem cells (ADSCs). While ADSCs are being used to treat various conditions, some lacuna exists regarding the specific proteins in these. It was therefore decided to analyze the specific proteins of embryonic cells in ADSCs. AIMS: To analyze the specific protein of embryonic stem cells (ESCs) in ADSCs. MATERIALS AND METHODS: Adult human adipose tissue-derived stem cells (ADSCs) were harvested from 13 patients after obtaining patients' consent. The specific markers of ESCs included surface proteins CD10, CD13, CD44, CD59, CD105, and CD166, and further nucleostemin, (NS) NANOG, peroxisome proliferator-activated receptor-gγ, collagen type 1 (Coll1), alkaline phosphate, (ALP) osteocalcin (OC), and core binding factor 1 (Cbfa1) were analyzed using by reverse transcription-polymerase chain reaction, (RT-PCR) immunofluorescence (IF), and western blot. RESULTS: All the proteins were expressed distinctly, except CD13 and OC. CD13 was found individually with different expressions, and OC expression was discernable. CONCLUSIONS: Although the ESC with its proven self-renewal capacity and pluripotency seems appropriate for clinical use, the recent work on ADSCs suggests that these adult stem cells would be a valuable source for future biotechnology, especially since there is a relative ease of procurement.

Leptin increases extracellular matrix mineralization of human osteoblasts from heterotopic ossification and normal bone.

Heterotopic ossification (HO) is the pathologic formation of bone in soft tissue. The exact pathomechanism is unknown but probably involves a disturbed osteoblast differentiation. Leptin, known as the obesity gene, may regulate normal osteoblast function in vitro. The aim of the present in vitro study was to further analyze the pathomechanisms of HO, including a possible role of leptin in ectopic bone formation. Human osteoblasts were cultivated either from normal bone or from resected HO. Both groups were incubated with increasing doses of leptin. Phenotype expression and mineralization of extracellular matrix were measured after 7, 14, and 21 days. In both groups, leptin increased both the formation of bone nodules and Ca-45 incorporation. This is the first study to analyze the effect of leptin on bone cells from ectopic ossification. Similar to the in vitro behavior of normal osteoblasts, cells from HO respond to leptin exposure with an increased mineralization of the extracellular matrix. This mechanism may be involved in the pathogenesis of ectopic bone formation in vivo.

A comparative analysis of phenotype expression in human osteoblasts from heterotopic ossification and normal bone.

BACKGROUND AND AIMS: Heterotopic ossification (HO) is a pathological bone formation process in which ectopic bone is formed in soft tissue. The formation of bone depends on the expression of the osteoblast phenotype. Earlier studies have shown conflicting results on the expression of phenotype markers of cells originating from HO and normal bone. The hypothesis of the present study is that cells from HO show an altered expression of osteoblast-specific phenotype markers compared to normal osteoblasts. The aims of the study were to further characterize the expression of osteoblast phenotypemarkers and to provide a comparison with other study results. PATIENTS AND METHODS: Using an in vitro technique, reverse transcription polymerase chain reaction (RT-PCR), real-time PCR and immunohistochemistry, we compared the phenotype gene expression (type I collagen, alkaline phosphatase, Cbfa-1, osteocalcin) of osteoblasts from resected HO and normal bone (iliac crest). RESULTS: Cells from HO expressed the osteoblast phenotype (type I collagen, alkaline phosphatase) but were characterized by a depleted osteocalcin expression. The expression of Cbfa-1 (osteocalcin transcription gene) showed a large variety in our study. Preoperative radiotherapy had no effect on phenotype expression in cells from HO. CONCLUSION: Our results provide a characterization of cells originating from HO and support the thesis of an impaired osteoblast differentiation underlying the formation of HO. The transcription axis from Cbfa-1 to osteocalcin could be involved in the pathogenesis of HO.

Influence of brain injury on early posttraumatic bone metabolism.

BACKGROUND: Various clinical studies and observations demonstrate enhanced osteogenesis in patients sustaining traumatic brain injury. It is presumed that the induction of this process starts early after trauma. The purpose of our study was to investigate humoral markers of bone metabolism during the early posttraumatic period, with special regard to traumatic brain injury. METHODS: Serum concentrations of biochemical markers of bone metabolism (calcium, inorganic phosphorus, carboxyl-terminal propeptide of type 1 procollagen, pyridinoline cross-linked telopeptide domain of type 1 collagen, Ostase, osteocalcin, intact parathyroid hormone, and calcitonin) were measured in three different groups of 80 patients during the first posttraumatic week. Patients were categorized into three groups: group I, fractures only; group II, isolated traumatic brain injury; and group III, traumatic brain injury in combination with fractures. RESULTS: Osteocalcin levels were significantly lower in the presence of traumatic brain injury (p < .05). Elevated pyridinoline cross-linked telopeptide domain of type 1 collagen levels expressed enhanced bone resorption in all groups, but levels were significantly higher in the absence of traumatic brain injury (p < .05). Intact parathyroid hormone levels were significantly higher on days 0 and 1 in the combined presence of traumatic brain injury plus fractures. CONCLUSION: These results demonstrate an imbalance of bone formation and resorption parameters in patients with traumatic brain injury during the early posttraumatic period, suggesting a central regulation in bone formation. The lower levels of osteocalcin detected in this study may play an important role in patients with brain injury and the later development of posttraumatic heterotopic ossification.

Osteoblasts response to allogenic and xenogenic solvent dehydrated cancellous bone in vitro.

The present in vitro study investigates the cellular interaction of primary human osteoblasts with human and bovine solvent dehydrated cancellous bone (SDCB) discs. These are bio-implants from solvent dehydrated, gamma-irradiated preserved human and bovine cancellous bone, pre-treated to remove all cells, genetic components and water soluble proteins. Primary human osteoblasts were harvested from cancellous chips of trauma patients undergoing osteosynthesis with bone grafting from the iliac crest. All patients provided informed consent. The present investigation tested proliferation, synthesis of phenotypic marker, and morphology of primary cultured human osteoblasts on SDCB in vitro. The total protein and collagen type 1 content could not be revealed, due to the inherent naturally occurring protein content in these two bio-implants. In conclusion, our in vitro results suggest that SDCB may be a suitable bone substitute which provides a well structured and biocompatible scaffold for ingrowing human osteoblasts.

Vigilancia sanitaria nacional de vacuna antisarampionosa: ensayo de potencia / National sanitary surveillance of measles vaccines: potency assay

América se perfila como el continente más promisorio en la lucha para la erradicación del sarampión. En este sentido, uno de los puntos clave es la calidad de las vacunas utilizadas. En Venezuela, la Gerencia de Control Nacional de Productos Biológicos (GCNPB) del INH "Rafael Rangel", es la encargada de verificar la calidad, seguridad y eficacia de ésta y todas las demás vacunas empleadas a nivel nacional. Uno de los ensayos más importantes en la evaluación de la calidad de la vacuna antisarampionosa, es la determinación de su potencia, para lo cual se emplean medios y cultivos celulares susceptibles. El objetivo del presente trabajo es comparar los medios y células empleadas por la GCNPB para este ensayo, con respecto a las utilizadas por el Instituto Nacional de Salud Pública y Protección Ambiental (RIVM) Bilthoven, Holanda, centro certificado por la OPS/OMS. En los resultados obtenidos no se detectaron diferencias estadísticamente significativas, al emplear células y medios de la GCNPB con respecto a las utilizadas por el RIVM, lo cual avala la calidad de los resultados emitidos, fortalece el Sistema Nacional de Vigilancia Sanitaria y apoya al Programa Nacional de Inmunización, al asegurar la calidad de las vacunas que utiliza en la lucha para la erradicación del sarampión

Avances en el estudio de la toxicidad específica de la vacuna pertussis elaborada por el INH "RR" / Advances in the study of specific toxicity of the pertussis vaccine elaborated by the INH \"RR\"

La vacuna Pertussis, normalmente administrada bajo la forma de vacuna triple o DPT (Difteria, Tétanos y Pertussis), es conocida como una de las vacunas más reactogénicas, causante de la mayoría de las reacciones secundarias presentadas después de la administración de la vacuna triple. Su aprobación es una relación de compromiso entre su toxicidad y un nivel de potencia adecuado para garantizar su capacidad inmunogénica con el menor grado posible de reacciones secundarias. A nivel de control de calidad, para determinar la toxicidad específica de la vacuna pertussis tradicionalmente se ha empleado la prueba de ganancia de peso en el ratón, sin embargo, la variable biológica involucrada, así como todos los factores que pueden incidir sobre la respuesta del animal pueden conllevar a la obtención de resultados contradictorios. Por otra parte, considerando la diversidad de agentes potencialmente tóxicos que posee esta vacuna, es primordial la aplicación de diversas metodologías que permitan la amplia evaluación de su toxicidad específica. Razón por la cual se diseñó este Trabajo de Investigación, con miras a estudiar por una parte la toxicidad específica de la vacuna pertussis elaborada por el Instituto Nacional de Higiene Rafael Rangel INH "RR", tanto en su forma individual como integrando la vacuna triple, (Difteria, Pertussis y Tétanos) y por la otra, evaluar conjuntamente estos resultados con los obtenidos en la prueba de ganancia de peso, con el objeto de realizar una mejor evaluación de la toxicidad específica y así permitir tomar una decisión de aprobación o rechazo sobre bases más sólidas y seguras