The adipose hormone leptin and its receptor are important for regulation of food intake and energy metabolism. Leptin also is involved in the growth of different tissues. In this study, we show the expression of leptin in primary cultures of normal human osteoblasts (

Anabolic hormones, mechanical loading, and the obese protein leptin play separate roles in maintaining bone mass. We have previously shown that leptin, as well as its receptor, are expressed by normal human osteoblasts. Consequently, we have investigated how leptin affects proliferation, differentiation, and apoptosis of human osteoblasts. Iliac crest osteoblasts, incubated with either leptin (100 ng/ml), calcitriol (1,25(OH)(2)D(3); 10(-9) M) or 1-84 human parathyroid hormone (PTH; 10(-8) M), were cultured for 35 consecutive days and assayed for expression of various differentiation-related marker genes (as estimated by RT-PCR), de novo collagen synthesis, proliferation, in vitro mineralization, and osteoclast signaling. The effects of leptin on protection against retinoic acid (RA; 10(-7) M) induced apoptosis, as well as transition into preosteocytes, were also tested. Leptin exposure enhanced cell proliferation and collagen synthesis over both control condition and PTH exposure. Leptin inhibited in vitro calcified nodule production after 1-2 weeks in culture, however, subsequent to 4-5 weeks, leptin significantly stimulated mineralization. The mineralization profile throughout the entire incubation period was almost undistinguishable from the one induced by PTH. In comparison, 1,25(OH)(2)D(3) generally reduced proliferation and collagen production rates, whereas mineralization was markedly enhanced. Leptin exposure (at 2 and 5 weeks) significantly enhanced the expression of TGFbeta, IGF-I, collagen-Ialpha, ALP, and osteocalcin mRNA. Leptin also protected against RA-induced apoptosis, as estimated by soluble DNA fractions and DNA laddering patterns subsequent to 10 days of culture. The expression profiles of Bax-alpha and Bcl-2 mRNAs indicated that leptin per se significantly protected against apoptosis throughout the entire incubation period. Furthermore, the osteoblast marker OSF-2 was diminished, whereas the CD44 osteocyte marker gene expression was stimulated, indicating a transition into preosteocytes. In terms of osteoclastic signaling, leptin significantly augmented the mRNA levels of both interleukin-6 (IL-6) and osteoprotegerin (OPG). In summary, continuous leptin exposure of iliac crest osteoblasts, promotes collagen synthesis, cell differentiation and in vitro mineralization, as well as cell survival and transition into preosteocytes. Leptin may also facilitate osteoblastic signaling to the osteoclast.

Injuries to articular cartilage are one of the most challenging issues of musculoskeletal medicine due to the poor intrinsic ability of this tissue for repair. Despite progress in orthopaedic surgery, the lack of efficient modalities of treatment for large chondral defects has prompted research on tissue engineering combining chondrogenic cells, scaffold materials and environmental factors. The aim of this review is to focus on the recent advances made in exploiting the potentials of cell therapy for cartilage engineering. These include: 1) defining the best cell candidates between chondrocytes or multipotent progenitor cells, such as multipotent mesenchymal stromal cells (MSC), in terms of readily available sources for isolation, expansion and repair potential; 2) engineering biocompatible and biodegradable natural or artificial matrix scaffolds as cell carriers, chondrogenic factors releasing factories and supports for defect filling, 3) identifying more specific growth factors and the appropriate scheme of application that will promote both chondrogenic differentiation and then maintain the differentiated phenotype overtime and 4) evaluating the optimal combinations that will answer to the functional demand placed upon cartilage tissue replacement in animal models and in clinics. Finally, some of the major obstacles generally encountered in cartilage engineering are discussed as well as future trends to overcome these limiting issues for clinical applications.

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The regeneration of bone tissue depends on the concerted actions of a plethora of signals that recruit mesenchymal stem cells for lineage-specifi c differentiation, with cellular phenotypes serving various functions throughout their life span. The signals are conveyed in hormones, growth factors, and mechanical forces, all of which ensure proper modeling and remodeling. Both processes are secured by indigenous and programmed metabolism in osteoblasts/osteocytes as well as in other stem cell (SC)-derived cell types (e.g., osteoclasts, bone lining cells) involved in the remodeling of the subject tissue. The focus of this review is the concerted action of these signals as well as the regulatory and/or stabilizing control circuits exhibited by a class of small RNAs, designated microRNAs. We discuss an in vitro approach for ensuring proper phenotype acquisition as well as the choice of scaffolds and animal models for in vivo tissue repair. This approach includes selection of SC niches to optimize bone formation in vivo, transcription factors important for osteoblastogenesis, the Wnt and Notch pathways of signaling, selection of delivery systems for gene therapy, use of appropriate matrices and scaffolds, in vivo mechanostimulation, choice of lesions to be repaired, and type of animal to use. We also discuss Wnt-related and SC-based treatment of osteoporosis. Throughout, we offer considerations for the selection of model systems and parameters to assess the entire procedure from initial SC selection to fi nal bone repair, and conclude with a table summarizing our recommendations.

Leptin has been advocated as a centrally acting factor responsible for inhibiting accumulation of bone mass. However, recent investigations unequivocally establish leptin as a local (autocrine) factor expressed by osteoblasts. Exogenously added leptin causes osteoblastic cell proliferation and differentiation, while also rendering osteoblasts more efficacious in terms of mineralization. Leptin acts as an anti-apoptotic agent, and augments messages responsible for the remodelling of bone tissue, i.e., mRNAs for osteoprotegerin (OPG) and the interleukin IL-6. Furthermore, leptin message is readily expressed in osteoblasts subjected to mechanical strain. In this respect, osteoblasts, which are unilaterally stretched proliferate and differentiate, a phenomenon being potentiated by exposure of the cells to differentiating humoral factors. This article discusses a unified model of dually acting leptin through the central nervous system and the mechanostat principle applied to osteoblasts. The proposed model may account for the finely tuned bone homeostasis maintained within rather narrow limits, depending on exposure to humoral factors and the prevailing mechanostat usage mode.

Bone and cartilage are being generated de novo through concerted actions of a plethora of signals. These act on stem cells (SCs) recruited for lineage-specific differentiation, with cellular phenotypes representing various functions throughout their life span. The signals are rendered by hormones and growth factors (GFs) and mechanical forces ensuring proper modelling and remodelling of bone and cartilage, due to indigenous and programmed metabolism in SCs, osteoblasts, chondrocytes, as well as osteoclasts and other cell types (eg T helper cells).This review focuses on the concerted action of such signals, as well as the regulatory and/or stabilizing control circuits rendered by a class of small RNAs, designated microRNAs. The impact on cell functions evoked by transcription factors (TFs) via various signalling molecules, also encompassing mechanical stimulation, will be discussed featuring microRNAs as important members of an integrative system. The present approach to cell differentiation in vitro may vastly influence cell engineering for in vivo tissue repair.

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