Since cancer stem cells (CSCs) were first identified in leukemia in 1994, they have been considered promising therapeutic targets for cancer therapy. These cells have self-renewal capacity and differentiation potential and contribute to multiple tumor malignancies, such as recurrence, metastasis, heterogeneity, multidrug resistance, and radiation resistance. The biological activities of CSCs are regulated by several pluripotent transcription factors, such as OCT4, Sox2, Nanog, KLF4, and MYC. In addition, many intracellular signaling pathways, such as Wnt, NF-κB (nuclear factor-κB), Notch, Hedgehog, JAK-STAT (Janus kinase/signal transducers and activators of transcription), PI3K/AKT/mTOR (phosphoinositide 3-kinase/AKT/mammalian target of rapamycin), TGF (transforming growth factor)/SMAD, and PPAR (peroxisome proliferator-activated receptor), as well as extracellular factors, such as vascular niches, hypoxia, tumor-associated macrophages, cancer-associated fibroblasts, cancer-associated mesenchymal stem cells, extracellular matrix, and exosomes, have been shown to be very important regulators of CSCs. Molecules, vaccines, antibodies, and CAR-T (chimeric antigen receptor T cell) cells have been developed to specifically target CSCs, and some of these factors are already undergoing clinical trials. This review summarizes the characterization and identification of CSCs, depicts major factors and pathways that regulate CSC development, and discusses potential targeted therapy for CSCs.Signal Transduction and Targeted Therapy (2020) 5:8; https://doi.

Although mesenchymal progenitor cells can be isolated from periodontal ligament (PDL) tissues using stem cell markers STRO-1 and CD146, the proportion of these cells that have the capacity to differentiate into multiple cell lineages remains to be determined. This study was designed to quantify the proportions of primary human PDL cells that can undergo multi-lineage differentiation, and to compare the magnitude of these capabilities relative to bone marrow-derived mesenchymal stem cells (MSC) and parental PDL (PPDL) cells. PDL mesenchymal progenitor (PMP) cells were isolated from PPDL cells using the markers STRO-1 and CD146. The colony-forming efficiency and multi-lineage differentiation potential of PMP, PPDL, and MSCs under chondrogenic, osteogenic and adipogenic conditions were determined. Flow cytometry revealed that on average 2.6% of PPDL cells were STRO-1+/CD146+, whereas more than 63% were STRO-1-/CD146-. Colony-forming efficiency of STRO-1+/CD146+ PMP cells (19.3%) and MSCs (16.7%) was significantly higher than that of PPDL cells (6.8%). Cartilage specific genes, early markers of osteoblastic differentiation, and adipogenic markers were significantly up-regulated under appropriate conditions in PMP cells and MSCs compared to either their non-induced counterparts or induced PPDL cells. Consistent with these findings, immunohistochemistry revealed substantial accumulation of cartilaginous macromolecules, mineralized calcium nodules, and lipid vacuoles under chondrogenic, osteogenic or adipogenic conditions in PMP and MSC cultures, respectively, compared to non-induced controls or induced PPDL cells. Thus, STRO-1+/CD146+ PMP cells demonstrate multi-lineage differentiation capacity comparable in magnitude to MSCs, and could potentially be utilized for regeneration of the periodontium and other tissues.

We characterized the temporal changes in chondrogenic genes and developed a staging scheme for in vitro chondrogenic differentiation of human mesenchymal stem cells (hMSCs) in three-dimensional (3D) alginate gels. A time-dependent accumulation of glycosaminoglycans, aggrecan, and type II collagen was observed in chondrogenic but not in basal constructs over 24 days. qRT-PCR demonstrated a largely characteristic temporal pattern of chondrogenic markers and provided a basis for staging the cellular phenotype into four stages. Stage I (days 0-6) was defined by collagen types I and VI, Sox 4, and BMP-2 showing peak expression levels. In stage II (days 6-12), gene expression for cartilage oligomeric matrix protein, HAPLN1, collagen type XI, and Sox 9 reached peak levels, while gene expression of matrilin 3, Ihh, Homeobox 7, chondroadherin, and WNT 11 peaked at stage III (days 12-18). Finally, cells in stage IV (days 18-24) attained peak levels of aggrecan; collagen IX, II, and X; osteocalcin; fibromodulin; PTHrP; and alkaline phosphatase. Gene profiles at stages III and IV were analogous to those in juvenile articular and adult nucleus pulposus chondrocytes. Gene ontology analyses also demonstrated a specific expression pattern of several putative novel marker genes. These data provide comprehensive insights on chondrogenesis of hMSCs in 3D gels. The derivation of this staging scheme may aid in defining maximally responsive time points for mechanobiological modulation of constructs to produce optimally engineered tissues.

Maturation of epiphyseal growth plate chondrocytes plays an important role in endochondral bone formation. Previously, we demonstrated that retinoic acid (RA) treatment stimulated annexin-mediated Ca 2؉ influx into growth plate chondrocytes leading to a significant increase in cytosolic Ca 2؉ , whereas K-201, a specific annexin Ca 2؉ channel blocker, inhibited this increase markedly. The present study addressed the hypothesis that annexin-mediated Ca 2؉ influx into growth plate chondrocytes is a major regulator of terminal differentiation, mineralization, and apoptosis of these cells. We found that K-201 significantly reduced up-regulation of expression of terminal differentiation marker genes, such as cbfa1, alkaline phosphatase (APase), osteocalcin, and type I collagen in RA-treated cultures. Maturation of epiphyseal growth plate chondrocytes, which plays an important role during endochondral ossification, is accompanied by major changes of chondrocyte morphology, biosynthetic activities, and energy metabolism. These processes involve an ordered progression of various cell differentiation stages, including proliferation, hypertrophic differentiation, terminal differentiation, and ultimately programmed cell death (apoptosis) (1, 2). During normal development these sequential events are under the strict control of local and systematic factors such as hormones and growth factors. If these processes, however, occur during pathological conditions, they can result in serious cartilage or bone defects. Evidence of endochondral ossification is also seen during osteophyte formation in osteoarthritic cartilage (3, 4). Terminal differentiation of growth plate chondrocytes is an essential process, which primes the cartilage skeleton for its subsequent invasion by osteoblasts and its replacement by a bone matrix. Despite the obvious importance of these terminal differentiation events still little is known about mechanisms regulating these processes.cbfa1, a member of the runt domain family of transcription factors, was originally discovered as a key transcription factor, which controls osteoblast differentiation. In cbfa1-null mice no endochondral and intramembranous bone formation occurs due to an arrest in osteoblast differentiation (5-8). Recent studies have indicated that cbfa1 also plays an important regulative role in terminal chondrocyte maturation. Transgenic mice, which overexpress cbfa1 in non-hypertrophic chondrocytes, display an acceleration of endochondral ossification. Overexpression of cbfa1 in chondrocytes of cbfa1-null mice partially rescued the abnormalities of cbfa1-null mutant mice. In particular, it rescued hypertrophic chondrocyte differentiation in the humerus and femur (9). Thus, cbfa1 seems to play dual functions in endochondral bone formation; it plays a key role in osteoblast differentiation from mesenchymal precursor cells, and it has the ability to stimulate hypertrophic and terminal chondrocyte differentiation.Chondrocyte hypertrophy and terminal differentiation are accompanied by an increase ...

Success of cloning using adult somatic cells has been reported in sheep, mice and cattle. The report that 'Dolly' the sheep, the first clone from an adult mammal, inherited shortened telomeres from her cell donor and that her telomeres were further shortened by the brief culture of donor cells has raised serious scientific and public concerns about the 'genetic age' and potential developmental problems of cloned animals. This observation was challenged by a recent report that showed calves cloned from fetal cells have longer telomeres than their age-matched controls. The question remains whether Dolly's short telomeres were an exception or a general fact, which would differ from the telomeres of fetal-derived clones.

Mineralization of growth plate cartilage is a critical event during endochondral bone formation, which allows replacement of cartilage by bone. Ankylosis protein (Ank), which transports intracellular inorganic pyrophosphate (PPi) to the extracellular milieu, is expressed by hypertrophic and, especially highly, by terminally differentiated mineralizing growth plate chondrocytes. Blocking Ank transport activity or ank expression in terminally differentiated mineralizing growth plate chondrocytes led to increases of intra- and extracellular PPi concentrations, decreases of alkaline phosphatase (APase) expression and activity, and inhibition of mineralization, whereas treatment of these cells with the APase inhibitor levamisole led to an increase of extracellular PPi concentration and inhibition of mineralization. Ank-overexpressing hypertrophic nonmineralizing growth plate chondrocytes showed decreased intra- and extracellular PPi levels; increased mineralization-related gene expression of APase, type I collagen, and osteocalcin; increased APase activity; and mineralization. Treatment of Ank-expressing growth plate chondrocytes with a phosphate transport blocker (phosphonoformic acid [PFA]) inhibited uptake of inorganic phosphate (Pi) and gene expression of the type III Na+/Pi cotransporters Pit-1 and Pit-2. Furthermore, PFA or levamisole treatment of Ank-overexpressing hypertrophic chondrocytes inhibited APase expression and activity and subsequent mineralization. In conclusion, increased Ank activity results in elevated intracellular PPi transport to the extracellular milieu, initial hydrolysis of PPi to Pi, Pi-mediated upregulation of APase gene expression and activity, further hydrolysis and removal of the mineralization inhibitor PPi, and subsequent mineralization

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We demonstrate that in Drosophila melanogaster the histone H3.3 replacement variant is encoded by two genes, H3.3A and H3.3B. We have isolated cDNA clones for H3.3A and cDNA and genomic clones for H3.3B. The genes encode exactly the same protein but are widely divergent in their untranslated regions (UTR). Both genes are expressed in embryos and adults; they are expressed in the gonads as well as in somatic tissues of the flies. However, only one of them, H3.3A, shows strong testes expression. The 3' UTR of the H3.3A gene is relatively short (approximately 250 nucleotides (nt)). H3.3B transcripts can be processed at several polyadenylation sites, the longest with a 3' UTR of more than 1500 nt. The 3' processing sites, preferentially used in the gonads and somatic tissues, are different. We have also isolated the Drosophila hydei homologues of the two H3.3 genes. They are quite similar to the D. melanogaster genes in their expression patterns. However, in contrast to their vertebrate counterparts, which are highly conserved in their noncoding regions, the Drosophila genes display only limited sequence similarity in these regions.