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J Pablo Rodríguez

Publications and source records attributed to J Pablo Rodríguez.

7 recordsLinked to original sources
PubMed2006

Aromatase activity of human mesenchymal stem cells is stimulated by early differentiation, vitamin D and leptin.

Human mesenchymal stem cells (hMSCs) are multipotent cells present in bone marrow, which differentiate into osteoblasts and adipocytes, among other lineages. Oestrogens play a critical role in bone metabolism; its action may affect the adipocyte to osteoblast ratio in the bone marrow. In hMSCs, oestrogens are synthesized from C19 steroids by the enzyme aromatase cytochrome P450. In this study, we assessed whether aromatase enzymatic activity varied through early osteogenic (OS) and adipogenic (AD) differentiation. Also, we studied the effect of leptin and 1,25 dihydroxyvitamin D3 (1,25(OH)2D3) on aromatase cell activity. Finally, we analysed whether conditions that modify oestrogen generation by cells affected hMSCs differentiation. For these purposes, hMSCs derived from post-menopausal women (65-86 years old) were cultured under basal, OS or AD conditions, in the presence or the absence of leptin and 1,25(OH)2D3. Aromatase activity was measured by the tritiated water release assay and by direct measurement of steroids synthesized from 3H-labelled androstenedione or testosterone. Our results showed that different OS and AD patterns of aromatase activity developed during the first period of differentiation (up to 7 days). A massive and sharp surge of aromatase activity at 24 h characterized early OS differentiation, while increased but constant aromatase activity was increased through adipogenesis. Both leptin and vitamin D increased aromatase activity during osteogenesis, but not during adipogenesis; finally, we showed that favourable aromatase substrates concentration restrained MSCs adipogenesis but improved osteogenesis. Thus, it could be inferred that a high and early increase of local oestrogen concentration in hMSCs affects their commitment either restraining AD or facilitating OS differentiation, or both.

Adipocytes↗
PubMed2005

High affinity leptin receptors are present in human mesenchymal stem cells (MSCs) derived from control and osteoporotic donors.

There are disparate observations on central and peripheral effects of leptin, but several studies consistently support its role as a link between fat and bone. Bone marrow stroma contains mesenchymal stem cells (MSCs), which differentiate into osteoblasts and adipocytes, among others. In this study we assessed the expression of leptin receptors protein in MSCs from control and osteoporotic postmenopausal donors and their change during osteogenic and adipogenic differentiation. Also, we assessed the effects of leptin on osteogenic and adipogenic differentiation of these cells. We demonstrated high affinity leptin binding (KD = 0.36 +/- 0.02 nM) in both types of cells. Binding was very low under basal, but increased significantly (2-3 times) through osteogenic and adipogenic differentiation. Osteoporotic MSCs showed lower leptin binding capacity than control cells at an early osteogenic and adipogenic differentiation time, which could restrict cell sensitivity to the protective action of leptin. In this regard, we observed that leptin significantly inhibited adipocyte differentiation in control but not in osteoporotic MSCs, while it exerted a low stimulatory effect on calcium deposition (10%-20%) in both types of MSCs cells. In summary, we report the presence of high affinity leptin receptors on control and osteoporotic MSCs, which were modified distinctly by osteogenic and adipogenic stimulation and a direct and distinct effect of leptin on both type of cells.

Aged↗
PubMed2004

Cytoskeletal organization of human mesenchymal stem cells (MSC) changes during their osteogenic differentiation.

Human MSCs have been studied to define the mechanisms involved in normal bone remodeling and the regulation of osteogenesis. During osteogenic differentiation, MSCs change from their characteristic fibroblast-like phenotype to near spherical shape. In this study, we analyzed the correlation between the organization of cytoskeleton of MSCs, changes in cell morphology, and the expression of specific markers (alkaline phosphatase activity and calcium deposition) of osteogenic differentiation. For osteoblastic differentiation, cells were cultured in a culture medium supplemented with 100 nM dexamethasone, 10 mM beta- glycerophosphate, and 50 microg/ml ascorbic acid. The organization of microfilaments and microtubules was examined by inmunofluorescence using Alexa fluor 594 phalloidin and anti alpha-tubulin monoclonal antibody. Cytochalasin D and nocodazole were used to alter reversibly the cytoskeleton dynamic. A remarkable change in cytoskeleton organization was observed in human MSCs during osteogenic differentiation. Actin cytoskeleton changed from a large number of thin, parallel microfilament bundles extending across the entire cytoplasm in undifferentiated MSCs to a few thick actin filament bundles located at the outermost periphery in differentiated cells. Under osteogenic culture conditions, a reversible reorganization of microfilaments induced by an initial treatment with cytochalasin D but not with nocodazole reduced the expression of differentiation markers, without affecting the final morphology of the cells. The results indicate that changes in the assembly and disassembly kinetics of microfilaments dynamic of actin network formation may be critical in supporting the osteogenic differentiation of human MSCs; also indicated that the organization of microtubules appears to have a regulatory role on the kinetic of this process.

Aged↗
PubMed2004

Differential activation of ERK1,2 MAP kinase signaling pathway in mesenchymal stem cell from control and osteoporotic postmenopausal women.

Osteoblasts, the cells responsible for bone formation, derive from mesenchymal stem cells (MSCs) in bone marrow. To acquire a new cell phenotype, uncommitted MSCs must undergo several proliferation and differentiation changes. Although, it is known that extracellular signal-regulated protein kinases (ERKs) mitogen-activated protein (MAP) kinase pathway signaling is involved in the proliferation and differentiation processes, the role of ERKs in osteogenic differentiation it is controversial, at present. In addition, the function that ERK could play in MSCs derived from osteoporotic patients it is not well documented. In this study, we analyze whether previously observed differences in the dynamic response of MSCs from normal and osteoporotic postmenopausal women can be explained by changes in the activation of this signal transduction pathway. Levels of ERK phosphorylation and their correlation with osteogenic differentiation were evaluated in cultures of MSCs derived from osteoporotic postmenopausal women and "healthy" controls. The results show that, under basal conditions, MSCs derived from osteoporotic donors show a level of ERK phosphorylation 2.5 times higher than MSCs derived from control donors. The addition of the osteogenic stimulus only slightly increases the p-ERK level in cells derived from osteoporotic donors, and is higher in cells derived from control women. Important differences in the ability of PD98059 to inhibit phosphorylation of ERK in both types of cells were also observed, as well as the effect that this inhibition produced on calcium deposition. We conclude that the MAP kinase pathway signaling is differentially activated in MSCs derived from osteoporotic postmenopausal women. The high p-ERK levels in MSC derived from osteoporotic donors could determine the unresponsiveness of these cells to the osteogenic differentiation stimulus.

Aged↗
PubMed2002

Modulation of the proliferation and differentiation of human mesenchymal stem cells by copper.

Copper plays important functional roles in bone metabolism and turnover. It is known that it is essential for normal growth and development of the skeleton in humans and in animals. Although at present the exact role that copper plays in bone metabolism is unknown, bone abnormalities are a feature of severe copper deficiency. Osteoblasts are derived from mesenchymal stem cells (MSCs) present in bone marrow stroma, which are able to differentiate into bone, adipocytes, and other cell phenotypes. Excess adipogenesis in postmenopausal women may occur at the expense of osteogenesis and, therefore, may be an important factor in the fragility of postmenopausal bone. The purpose of this study was to evaluate whether an increase of the extracellular concentration of copper affects the ability of MSCs to differentiate into osteoblasts or adipocytes. The results showed that copper modified both the differentiation and the proliferative activity of MSCs obtained from postmenopausal women. Copper (50 microM) diminished the proliferation rate of MSCs, increasing their ability to differentiate into the osteogenic and the adipogenic lineages. Copper induced a 2-fold increase in osteogenic differentiation of MSCs, measured as a increase in calcium deposition. Copper (5 and 50 microM) diminished the expression of alkaline phosphatase (50 and 80%, respectively), but induced a shift in the expression of this enzyme to earlier times during culture. Copper also induced a 1.3-fold increase in the adipogenic differentiation of MSCs. It is concluded that copper stimulates MSC differentiation, and that this is preferentially towards the osteogenic lineage.

Adipocytes↗