IntroductionThe vertebrate mesoderm gives rise to a diverse variety of tissues, including the heart, vasculature, and blood. It is formed in the primitive streak (PS) which appears first in the posterior epiblast during gastrulation. The nascent PS then elongates to the anterior part, whereas the epiblast cells ingress through the PS, migrate to appropriate location in the embryo, and become either mesoderm or definitive endoderm. 1 Although the process of mesoderm formation is well understood, the underlying molecular mechanisms remain poorly defined.Previous studies have identified several signaling pathways involved in this process. 2 One is the transforming growth factorbeta (TGF-␤) superfamily member, bone morphogenetic protein-4 (BMP-4). Like other TGF-␤ superfamily members, BMP-4 binds to and brings together the type I and type II receptors on the cell surface, which allows receptor II to phosphorylate the receptor I kinase domain. After this activation, the type I receptor phosphorylates certain members of the Smad proteins (Smad 1, 5, 8; R-Smad). Two phosphorylated R-Smads, in combination with a common mediator Smad (Smad4; Co-Smad), form a heterotrimeric complex, which then translocates into the nucleus and cooperates with other transcription factors to modulate target gene expression. [3][4][5][6] Genetic studies in mice have shown that BMP-4 signaling is required for mesoderm formation. [7][8][9] Mice deficient in the BMP-4 ligand, 7 in the BMP type II receptor, 8 or in a BMP type I receptor Alk3, 9 all fail to develop mesoderm. It has also been demonstrated that BMP-4 plays critical roles in mesoderm induction and mesoderm lineage differentiation from mouse embryonic stem cells (mES cells). [10][11][12][13][14][15][16][17][18][19] The key role of BMP-4 in mouse embryos and mES cells has led us to investigate its function in human embryonic stem cells (hES cells), the differentiation of which can, at least in part, mimic the human embryogenesis. 20,21 Xu et al 22 showed that long-term BMP-4 treatment (up to 7 days) induces hES cell differentiation into trophoblast. Later, Pera et al 23 demonstrated that activation of the same signaling pathway by BMP-2 in hES cells results in the formation of extra-embryonic endoderm. Surprisingly, we found that short-term BMP-4 treatment (24 hours) initiated mesoderm induction at a high efficiency in hES cells. In addition, these mesoderm progenitor cells were able to differentiate into various mesoderm lineages. The differential effects of short-term and long-term BMP-4 treatments on hES cell differentiation suggest that the BMP signaling pathway might play a flexible and time-dependent role in human embryonic development and cell fate determination. Methods Cell culture and differentiationThe H1, H7, and H9 hES cell lines were obtained from WiCell Research Institute (Madison, WI). Cells were cultured and passaged on mitomycin C-treated mouse embryonic fibroblast (MEF) feeders in hES cell culture Submitted February 14, 2007; accepted November 20, 2007. Prepublished...

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Bone is a dynamic tissue constantly responding to environmental changes such as nutritional and mechanical stress. Bone homeostasis in adult life is maintained through bone remodeling, a controlled and balanced process between bone-resorbing osteoclasts and bone-forming osteoblasts. Osteoblasts secrete matrix, with some being buried within the newly formed bone, and differentiate to osteocytes. During embryogenesis, bones are formed through intramembraneous or endochondral ossification. The former involves a direct differentiation of mesenchymal progenitor to osteoblasts, and the latter is through a cartilage template that is subsequently converted to bone. Advances in lineage tracing, cell sorting, and single-cell transcriptome studies have enabled new discoveries of gene regulation, and new populations of skeletal stem cells in multiple niches, including the cartilage growth plate, chondro-osseous junction, bone, and bone marrow, in embryonic development and postnatal life. Osteoblast differentiation is regulated by a master transcription factor RUNX2 and other factors such as OSX/SP7 and ATF4. Developmental and environmental cues affect the transcriptional activities of osteoblasts from lineage commitment to differentiation at multiple levels, fine-tuned with the involvement of co-factors, microRNAs, epigenetics, systemic factors, circadian rhythm, and the microenvironments. In this review, we will discuss these topics in relation to transcriptional controls in osteogenesis.

The integrated stress response (ISR) is activated by diverse forms of cellular stress, including endoplasmic reticulum (ER) stress, and is associated with diseases. However, the molecular mechanism(s) whereby the ISR impacts on differentiation is incompletely understood. Here, we exploited a mouse model of Metaphyseal Chondrodysplasia type Schmid (MCDS) to provide insight into the impact of the ISR on cell fate. We show the protein kinase RNA-like ER kinase (PERK) pathway that mediates preferential synthesis of ATF4 and CHOP, dominates in causing dysplasia by reverting chondrocyte differentiation via ATF4-directed transactivation of Sox9. Chondrocyte survival is enabled, cell autonomously, by CHOP and dual CHOP-ATF4 transactivation of Fgf21. Treatment of mutant mice with a chemical inhibitor of PERK signaling prevents the differentiation defects and ameliorates chondrodysplasia. By preventing aberrant differentiation, titrated inhibition of the ISR emerges as a rationale therapeutic strategy for stress-induced skeletal disorders.