Induction of ornithine decarboxylase in T/C-28a2 chondrocytes by lysophosphatidic acid: signaling pathway and inhibition of cell proliferation

Among several extracellular messengers tested, lysophosphatidic acid (LPA) was able to cause the most marked induction of ornithine decarboxylase (ODC) in serum-starved human T/C-28a2 chondrocytes. LPA also induced the activation/phosphorylation of Src, Akt and p44/42 MAPK, and the translocation of PKC-delta from cytosol to membrane coupled to its tyrosine phosphorylation. Experiments with selective signaling inhibitors indicate that LPA leads to Src activation through Gi protein-coupled receptors. In turn Src can activate PI3K and PKC-delta, and all these signaling proteins are required for ODC induction. In conclusion these results show that chondrocytes may be a novel target for LPA action. However, although LPA is considered a mitogen for several cell types and ODC induction is generally correlated to cell growth, LPA was not able to stimulate chondrocyte growth, but rather exerted an anti-proliferative effect.     

Spontaneous expression of neural phenotype and NGF, TrkA, TrkB genes in marrow stromal cells

Marrow stromal cells (MSCs) have the ability to provide growth factors and differentiate into neural-like cells on treating with EGF, bFGF and other factors. We wanted to explore whether growth factors secreted by MSCs itself could induce self-differentiation into neural-like cells. Here, we show that even in the absence of inducing factors, rMSCs spontaneously differentiate into neural-like cells expressing neural markers, such as nestin, beta-tubulin III, Doublecortin (DCX), microtubule-associated protein 2 (MAP2) and neuron-specific enolase (NSE). Furthermore, some cells become neurosphere-like growing in suspension. Compared with control and neural-like rMSCs induced by epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF), we found using real-time PCR that self-differentiating rMSCs (SDrMSCs) expressed significantly higher levels of neurotrophic high-affinity receptors (TrkA and TrkB). Coincident with neural marker expression, nerve growth factor (NGF) mRNA was significantly higher than controls despite lower protein levels in the supernatant. Our study suggests that rMSCs have the potential to differentiate into neural cells spontaneously in culture and may contribute towards the natural function of MSCs for neural system in vivo.     

Effect of the cyclic stretch on the expression of osteogenesis genes in human periodontal ligament cells

Periodontal ligament cells can potentially differentiate into osteoblast-like cells and influence the remodeling of periodontal tissues under mechanical strain conditions. In the present study, Gene chip technology was adopted to investigate the effect of the cyclic stretch on the expression of osteogenic-related genes in human periodontal ligament cells (HPDLCs). Cultured HPDLCs were subjected to 12% elongation cyclic stretch for 24 h using a Flexercell Strain Unit, and then GEArray Q series human osteogenesis gene expression profile chip with 96 spot array numbers was used to conduct parallel analyses on the change of the related gene expression in the osteogenic differentiation of HPDLCs stimulated by cyclic stretch. The results show that after the HPDLCs were stimulated by the cyclic stretch, the expression of 21 osteogenic-related genes was significantly upregulated, including 10 growth factor genes and their associated molecules, 10 extracellular matrix genes and their associated proteins, and 1 cell adhesion molecule. Two genes were significantly downregulated, including one growth factor gene and one cell adhesion molecule. Then the expressions of 10 candidate genes were validated using Real-time RT-PCR. These results indicate that cyclic stretch with 12% deformation can stimulate or inhibit some gene expression which was associated with the process of HPDLCs differentiation.     

Glucocorticoids exert context-dependent effects on cells of the joint in vitro

Introduction

Glucocorticoids are known to attenuate bone formation in vivo leading to decreased bone volume and increased risk of fractures, whereas effects on the joint tissue are less characterized. However, glucocorticoids appear to have a reducing effect on inflammation and pain in osteoarthritis. This study aimed at characterizing the effect of glucocorticoids on chondrocytes, osteoclasts, and osteoblasts.

Experimental

We used four model systems to investigate how glucocorticoids affect the cells of the joint; two intact tissues (femoral head- and cartilage-explants), and two separate cell cultures of osteoblasts (2T3-pre-osteoblasts) and osteoclasts (CD14⁺-monocytes). The model systems were cultured in the presence of two glucocorticoids; prednisolone or dexamethasone. To induce anabolic and catabolic conditions, cultures were activated by insulin-like growth factor I/bone morphogenetic protein 2 and oncostatin M/tumor necrosis factor-α, respectively. Histology and markers of bone- and cartilage-turnover were used to evaluate effects of glucocorticoid treatment.

Results

Prednisolone treatment decreased collagen type-II degradation in immature cartilage, whereas glucocorticoids did not affect collagen type-II in mature cartilage. Glucocorticoids had an anti-catabolic effect on catabolic-activated cartilage from a bovine stifle joint and murine femoral heads. Glucocorticoids decreased viability of all bone cells, leading to a reduction in osteoclastogenesis and bone resorption; however, bone morphogenetic protein 2-stimulated osteoblasts increased bone formation, as opposed to non-stimulated osteoblasts.

Conclusions

Using highly robust in vitro models of bone and cartilage turnover, we suggest that effects of glucocorticoids highly depend on the activation and differential stage of the cell targeted in the joint. Present data indicated that glucocorticoid treatment may be beneficial for articular cartilage, although detrimental effects on bone should be taken into account.     

PI3K/Akt is involved in brown adipogenesis mediated by growth differentiation factor-5 in association with activation of the Smad pathway

We have previously demonstrated promotion by growth differentiation factor-5 (GDF5) of brown adipogenesis for systemic energy expenditure through a mechanism relevant to activating the bone morphological protein (BMP) receptor/mothers against decapentaplegic homolog (Smad)/peroxisome proliferator-activated receptor gamma co-activator 1α (PGC-1α) pathway. Here, we show the involvement of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in brown adipogenesis mediated by GDF5. Overexpression of GDF5 in cells expressing adipocyte protein-2 markedly accelerated the phosphorylation of Smad1/5/8 and Akt in white and brown adipose tissues. In brown adipose tissue from heterozygous GDF5^Rgsc451 mutant mice expressing a dominant-negative (DN) GDF5 under obesogenic conditions, the basal phosphorylation of Smad1/5/8 and Akt was significantly attenuated. Exposure to GDF5 not only promoted the phosphorylation of both Smad1/5/8 and Akt in cultured brown pre-adipocytes, but also up-regulated Pgc1a and uncoupling protein-1 expression in a manner sensitive to the PI3K/Akt inhibitor Ly294002 as well as retroviral infection with DN-Akt. GDF5 drastically promoted BMP-responsive luciferase reporter activity in a Ly294002-sensitive fashion. Both Ly294002 and DN-Akt markedly inhibited phosphorylation of Smad5 in the nuclei of brown pre-adipocytes. These results suggest that PI3K/Akt signals play a role in the GDF5-mediated brown adipogenesis through a mechanism related to activation of the Smad pathway.     

Neurogenic differentiation of human adipose-derived stem cells: Relevance of different signaling molecules,transcription factors,and key marker genes

Since numerous diseases affect the central nervous system and it has limited self-repair capability, a great interest in using stem cells as an alternative cell source is generated. Previous reports have shown the differentiation of adipose-derived stem cells in neuron-like cells and it has also been proved that the expression pattern of patterning, proneural, and neural factors, such as Pax6, Mash1, Ngn2, NeuroD1, Tbr2 and Tbr1, regulates and defines adult neurogenesis. Regarding this, we hypothesize that a functional parallelism between adult neurogenesis and neuronal differentiation of human adipose-derived stem cells exists. In this study we differentiate human adipose-derived stem cells into neuron-like cells and analyze the expression pattern of different patterning, proneural, neural and neurotransmitter genes, before and after neuronal differentiation. The neuron-like cells expressed neuronal markers, patterning and proneural factors characteristics of intermediate stages of neuronal differentiation. Thus we demonstrated that it is possible to differentiate adipose-derived stem cells in vitro into immature neuron-like cells and that this process is regulated in a similar way to adult neurogenesis. This may contribute to elucidate molecular mechanisms involved in neuronal differentiation of adult human non-neural cells, in aid of the development of potential therapeutic tools for diseases of the nervous system.     

Cellular interplay in pulmonary arterial hypertension: Implications for new therapies

Pulmonary arterial hypertension (PAH) is a complex and multifactorial disease characterized by vascular remodeling, vasoconstriction, inflammation and thrombosis. Although the available therapies have resulted in improvements in morbidity and survival, PAH remains a severe and devastating disease with a poor prognosis and a high mortality, justifying the need of novel therapeutic targets. An increasing number of studies have demonstrated that endothelial cells (ECs), smooth muscle cells (SMCs) and fibroblasts of the pulmonary vessel wall, as well as platelets and inflammatory cells have a role in PAH pathogenesis. This review aims to integrate the interplay among different types of cells, during PAH development and progression, and the impact of current therapies in cellular modulation. The interplay among endothelial cells, smooth muscle cells and fibroblasts present in pulmonary vessels wall, platelets and inflammatory cells is regulated by several mediators produced by these cells, contributing to the pathophysiologic features of PAH. Current therapies are mainly focused in the pulmonary vascular tone and in the endothelial dysfunction. However, once they have not been effective, novel therapies targeting other PAH features, such as inflammation and platelet dysfunction are emerging. Further understanding of the interplay among different vascular cell types involved in PAH development and progression can contribute to find novel therapeutic targets, decreasing PAH mortality and morbidity in the future.     

Endometrial stromal cells and decidualized stromal cells: Origins,transformation and functions

Decidualization of endometrium, which is characterized by endometrial stromal cell (ESC) decidualization, vascular reconstruction, immune cell recruitment, and plentiful molecule production, is a crucial step for uterus to become receptive for embryo. When implantation takes place, ESCs surround and directly interact with embryo. Decidualized stromal cells (DSCs) are of great importance in endometrial decidualization, having a broad function in regulating immune activity and vascular remodeling of uterus. DSCs are shown to have a higher metabolic level and looser cytoskeleton than ESCs. What's the origin of ESCs and how ESCs successfully transform into DSCs had puzzled scientists in the last decades. Breakthrough had been achieved recently, and many studies had elucidated some of the characters and functions of DSCs. However, several questions still remain unclear. This paper reviews current understanding of where ESCs come from and how ESCs differentiate into DSCs, summarizes some characters and functions of DSCs, analyzes current studies and their limitations and points out research areas that need further investigation.