The tunica adventitia of human arteries and veins as a source of mesenchymal stem cells.

We previously demonstrated that human pericytes, which encircle capillaries and microvessels, give rise in culture to genuine mesenchymal stem cells (MSCs). This raised the question as to whether all MSC are derived from pericytes. Pericytes and other cells defined on differential expression of CD34, CD31, and CD146 were sorted from the stromal vascular fraction of human white adipose tissue. Besides pericytes, CD34+ CD31- CD146- CD45- cells, which reside in the outmost layer of blood vessels, the tunica adventitia, natively expressed MSC markers and gave rise in culture to clonogenic multipotent progenitors identical to standard bone marrow-derived MSC. Despite common MSC features and developmental properties, adventitial cells and pericytes retain distinct phenotypes and genotypes through culture. However, in the presence of growth factors involved in vascular remodeling, adventitial cells acquire a pericytes-like phenotype. In conclusion, we demonstrate the co-existence of 2 separate perivascular MSC progenitors: pericytes in capillaries and microvessels and adventitial cells around larger vessels.

HAX-1: a family of apoptotic regulators in health and disease.

HAX-1 comprises a family of ubiquitously expressed proteins that play important roles in the regulation of programmed cell death. Herein, we provide a comprehensive review of the expression profile of HAX-1 and its functional implications during health and disease, highlighting its direct involvement in the development of congenital neutropenia and neural abnormalities, when absent, and its contribution to the progression of psoriasis and cancer, when overexpressed. Moreover, we provide new information on the differential expression of the HAX-1 subfamily in three distinct types of epithelial cancers, including breast, skin, and colon. Our results demonstrate a significant up-regulation of the anti-apoptotic HAX-1 variant 001 in skin and colon, but not in breast and cancer cells, indicating tissue-specific differences in its expression pattern and properties during cancer formation and progression. Our findings further reveal a considerable down-regulation, if not abrogation, of three distinct, yet to be characterized, HAX-1 isoforms in breast cancer cells, suggesting that they may function in an opposite manner to the anti-apoptotic variant 001. This study aims to summarize our current knowledge on the physiological implications of the expression profile of the HAX-1 subfamily in health and disease, and provide new information on the differential expression and activities of HAX-1 members in three distinct types of cancer.

HAX-1: a multifaceted antiapoptotic protein localizing in the mitochondria and the sarcoplasmic reticulum of striated muscle cells.

HAX-1 comprises a family of ubiquitously expressed proteins with antiapoptotic properties. In the current study, we investigated HAX-1's temporospatial distribution in rat striated muscles during development and in adulthood. In cardiocytes, HAX-1 is organized at the level of Z-disks throughout embryogenesis and adulthood; however, in skeletal myofibers, it is in register with M-bands during embryonic and early postnatal life and Z-disks during late postnatal and adult life. Immunoelectron microscopy and subcellular fractionation demonstrated that HAX-1 proteins localize at the mitochondrial and sarcoplasmic reticulum (SR) membranes, as well as at sites where the two are closely apposed. Variants I and II selectively concentrate in the mitochondrial membranes, whereas variants III, IV, and V localize in both organelles, albeit to varying extents. Deletion analysis combined with cellular transfections indicated that elimination of HAX-1's NH(2)-terminus abolishes its mitochondrial targeting and attenuates its antiapoptotic capacity, while removal of its binding site for the SR protein phospholamban (PLN) prevents its translocation to the SR. Consistent with this, HAX-1 is preferentially lost from the SR of PLN-deficient hearts. Our findings are the first to present a comprehensive characterization of HAX-1's expression in striated muscles and to provide insights on the mechanisms through which it may modulate apoptosis.

Muscle giants: molecular scaffolds in sarcomerogenesis.

Myofibrillogenesis in striated muscles is a highly complex process that depends on the coordinated assembly and integration of a large number of contractile, cytoskeletal, and signaling proteins into regular arrays, the sarcomeres. It is also associated with the stereotypical assembly of the sarcoplasmic reticulum and the transverse tubules around each sarcomere. Three giant, muscle-specific proteins, titin (3-4 MDa), nebulin (600-800 kDa), and obscurin (approximately 720-900 kDa), have been proposed to play important roles in the assembly and stabilization of sarcomeres. There is a large amount of data showing that each of these molecules interacts with several to many different protein ligands, regulating their activity and localizing them to particular sites within or surrounding sarcomeres. Consistent with this, mutations in each of these proteins have been linked to skeletal and cardiac myopathies or to muscular dystrophies. The evidence that any of them plays a role as a "molecular template," "molecular blueprint," or "molecular ruler" is less definitive, however. Here we review the structure and function of titin, nebulin, and obscurin, with the literature supporting a role for them as scaffolding molecules and the contradictory evidence regarding their roles as molecular guides in sarcomerogenesis.

A perivascular origin for mesenchymal stem cells in multiple human organs.

Mesenchymal stem cells (MSCs), the archetypal multipotent progenitor cells derived in cultures of developed organs, are of unknown identity and native distribution. We have prospectively identified perivascular cells, principally pericytes, in multiple human organs including skeletal muscle, pancreas, adipose tissue, and placenta, on CD146, NG2, and PDGF-Rbeta expression and absence of hematopoietic, endothelial, and myogenic cell markers. Perivascular cells purified from skeletal muscle or nonmuscle tissues were myogenic in culture and in vivo. Irrespective of their tissue origin, long-term cultured perivascular cells retained myogenicity; exhibited at the clonal level osteogenic, chondrogenic, and adipogenic potentials; expressed MSC markers; and migrated in a culture model of chemotaxis. Expression of MSC markers was also detected at the surface of native, noncultured perivascular cells. Thus, blood vessel walls harbor a reserve of progenitor cells that may be integral to the origin of the elusive MSCs and other related adult stem cells.

Purification and culture of human blood vessel-associated progenitor cells.

Multilineage progenitor cells, diversely designated as MSC, MAPC, or MDSC, have been previously extracted from long-term cultures of fetal and adult organs (e.g., bone marrow, brain, lung, pancreas, muscle, adipose tissue, and several others). The identity and location, within native tissues, of these elusive stem cells are described here. Subsets of endothelial cells and pericytes, which participate in the architecture of human blood vessels, exhibit, following purification to homogeneity, developmental multipotency. The selection from human tissues, by flow cytometry using combinations of positive and negative cell surface markers, of endothelial and perivascular cells is described here. In addition, a rare subset of myoendothelial cells that express markers of both endothelial and myogenic cell lineages and exhibit dramatic myogenic and cardiomyogenic potential has been identified and purified from skeletal muscle. The culture conditions amenable to the long-term proliferation of these blood vessel-associated stem cells in vitro are also described.

A reservoir of brown adipocyte progenitors in human skeletal muscle.

Brown adipose tissue uncoupling protein-1 (UCP1) plays a major role in the control of energy balance in rodents. It has long been thought, however, that there is no physiologically relevant UCP1 expression in adult humans. In this study we show, using an original approach consisting of sorting cells from various tissues and differentiating them in an adipogenic medium, that a stationary population of skeletal muscle cells expressing the CD34 surface protein can differentiate in vitro into genuine brown adipocytes with a high level of UCP1 expression and uncoupled respiration. These cells can be expanded in culture, and their UCP1 mRNA expression is strongly increased by cell-permeating cAMP derivatives and a peroxisome-proliferator-activated receptor-gamma (PPARgamma) agonist. Furthermore, UCP1 mRNA was detected in the skeletal muscle of adult humans, and its expression was increased in vivo by PPARgamma agonist treatment. All the studies concerning UCP1 expression in adult humans have until now been focused on the white adipose tissue. Here we show for the first time the existence in human skeletal muscle and the prospective isolation of progenitor cells with a high potential for UCP1 expression. The discovery of this reservoir generates a new hope of treating obesity by acting on energy dissipation.

Prospective identification of myogenic endothelial cells in human skeletal muscle.

We document anatomic, molecular and developmental relationships between endothelial and myogenic cells within human skeletal muscle. Cells coexpressing myogenic and endothelial cell markers (CD56, CD34, CD144) were identified by immunohistochemistry and flow cytometry. These myoendothelial cells regenerate myofibers in the injured skeletal muscle of severe combined immunodeficiency mice more effectively than CD56+ myogenic progenitors. They proliferate long term, retain a normal karyotype, are not tumorigenic and survive better under oxidative stress than CD56+ myogenic cells. Clonally derived myoendothelial cells differentiate into myogenic, osteogenic and chondrogenic cells in culture. Myoendothelial cells are amenable to biotechnological handling, including purification by flow cytometry and long-term expansion in vitro, and may have potential for the treatment of human muscle disease.