Regeneration, tissue remodeling, and organ repair after injury, which rely on the regulated activity of tissue-borne stem cells, become increasingly compromised with advancing age. Mesenchymal stroma cells were isolated from bone of differently aged healthy donors. The rare population of mesenchymal stem cells (MSCs) contained in the primary cell isolates barely declined in number, yet the stem cells displayed diminished long-term proliferation potential relative to the donor age and the expression of vascular cell adhesion molecule-1 (VCAM-1; CD106) was elevated on primary MSCs. In CD106(bright) MSCs, the abundance of a panel of stemness transcription factors remained unchanged. Because the CD106 level could be further enhanced by proinflammatory cytokines, we considered the rate of VCAM-1 expression to be a good reflection of an endogenous inflammatory milieu to which the MSCs are exposed. Treatment of MSCs with increasing doses of interferon-γ exerted no immediate influence on their self-renewal capacity. However, it impacted on the differentiation potential toward the adipogenic or osteogenic lineage. Moderately elevated levels of inflammatory stimuli supported osteoblastogenesis whereas the same treatment reduced adipogenic differentiation in MSCs from young and intermediately aged donors. In MSCs from elderly donors, however, osteoblastogenesis was greatly diminished in an inflammatory environment whereas adipogenic differentiation remained unchanged. Conclusively, moderate levels of inflammatory stimuli are being interpreted by MSCs at a young age as instructive signals for osteoblastogenesis, whereas at old age, an inflammatory milieu may effectively suppress bone remodeling and repair by tissue-borne MSCs while uninterrupted adipogenic differentiation may lead to adipose upgrowth.
Aging as a process is paralleled by a variety of hematological alterations. Characteristic features are a diminished homeostatic control of blood cell production and a decline in immune functions. It is generally accepted that stromal cells play a basal role in hematopoiesis by providing survival and differentiation signals, by secreting cytokines, or through direct contact with hematopoietic stem cells, thereby supporting the generation and replenishment of hematopoi- etic progenitor cells (HPC). Here we demonstrated that HPC-related colony formation is positively influenced by mesenchymal stromal cells (MSCs) when grown in co-culture, in particular regarding the number of primary granulocyte/macrophage colony-forming units as well as with respect to the average size of the formed colonies. These effects were more pronounced when the MSCs originated from young donors than from old ones. Because leukemia inhibitory factor (LIF) plays an important role during hematopoiesis, properties of lin- Sca-1+ cells and MSCs derived from LIF-deficient mice (LIF-/-) were determined both ex vivo and in vitro. LIF-/- animals contain a significantly reduced number of lin- Sca-1+ cells, nevertheless the replating capacity of LIF-/- HPCs was found to be generally unchanged when compared to those from LIF+/+ animals. However, when cocultured with MSCs, LIF-/- lin- Sca-1+ cells exhibited comparable characteristics to HPCs derived from old wild-type animals.
Abstract We have recently described an IL-2/IL-4-producing CD8+CD25+ nonregulatory memory T cell population that occurs in a subgroup of healthy elderly persons who characteristically still have a good humoral response after vaccination. The present study addresses this specific T cell subset and investigates its origin, clonal composition, Ag specificity, and replicative history. We demonstrate that CD8+CD25+ memory T cells frequently exhibit a CD4+CD8+ double-positive phenotype. The expression of the CD8 αβ molecule and the occurrence of signal-joint TCR rearrangement excision circles suggest a thymic origin of these cells. They also have longer telomeres than their CD8+CD25− memory counterparts, thus indicating a shorter replicative history. CD8+CD25+ memory T cells display a polyclonal TCR repertoire and respond to IL-2 as well as to a panel of different Ags, whereas the CD8+CD25− memory T cell population has a more restricted TCR diversity, responds to fewer Ags, and does not proliferate in response to stimulation with IL-2. Molecular tracking of specific clones with clonotypic primers reveals that the same clones occur in CD8+CD25+ and CD8+CD25− memory T cell populations, demonstrating a lineage relationship between CD25+ and CD25− memory CD8+ T cells. Our results suggest that CD25-expressing memory T cells represent an early stage in the differentiation of CD8+ cells. Accumulation of these cells in elderly persons appears to be a prerequisite of intact immune responsiveness in the absence of naive T cells in old age.
AbstractRegeneration, tissue remodeling and organ repair after injury which rely on theregulated activity of tissue-borne stem cells become increasingly compromised withadvancing age.Mesenchymal stroma cells were isolated from bone of differently aged healthydonors. The rare population of mesenchymal stem cells (MSC) contained in theprimary cell isolates, barely declined in number, yet relative to the donor age thestem cells displayed diminished long-term proliferation potential. The expression ofvascular cell adhesion molecule 1 (CD106) was elevated on primary MSC. InCD106bright MSC, the abundance of a panel of stemness transcription factorsremained unchanged. As the CD106 level could be further enhanced by proinflammatorycytokines, we considered the rate of VCAM1 expression a goodreflection of an endogenous inflammatory milieu MSC are exposed to. Treatment ofMSC with increasing doses of interferon gamma exerted no immediate influence ontheir self-renewal capacity. It however impacted on the differentiation potentialtowards the adipogenic or osteogenic lineage. Moderately elevated levels ofinflammatory stimuli supported osteoblastogenesis while the same treatmentreduced adipogenic differentiation in MSC from young and intermediately ageddonors. In mesenchymal stem cells from elderly donors however,osteoblastogenesis was greatly diminished in an inflammatory environment whereasadipogenic differentiation remained unchanged.Conclusively, moderate levels of inflammatory stimuli are being interpreted bymesenchymal stem cells at young age as instructive signals for osteoblastogenesis,whereas at old age, an inflammatory milieu may effectively suppress boneremodeling and repair by tissue-borne mesenchymal stem cells while uninterruptedadipogenic differentiation may lead to adipose upgrowth.
Besides SPAM1 (sperm adhesion molecule 1; formerly named PH-20), further hyaluronidase-like proteins, HYAL5 (hyaluronoglucosaminidase 5) and HYALP1 (hyaluronoglucosaminidase pseudogene 1) are also expressed in murine testicular tissue. As they share a high degree of sequence similarity with known hyaluronidases, all three polypeptides could potentially exhibit hyaluronidase activity, a function that is beneficial for spermatozoa in order to penetrate the hyaluronan-rich cumulus, which surrounds the oocyte. Recently, it was reported that SPAM1-deficient mice are fertile and spermatozoa derived from mutant mice still exhibit hyaluronidase activity [Baba, Kashiwabara, Honda, Yamagata, Wu, Ikawa, Okabe and Baba (2002) J. Biol. Chem. 277, 30310-30314]. We have now recombinantly expressed mouse SPAM1, HYAL5 and HYALP1 in Xenopus laevis oocytes and determined their respective expression pattern in testis. Transcripts of all three genes are expressed in seminiferous tubules in regions where maturing spermatogenic cells reside. SPAM1 and HYAL5 but not HYALP1 proteins exhibit hyaluronidase activity at neutral pH. The two active hyaluronidases are both bound to the cell surface via a glycosylphosphatidylinositol anchor. Furthermore, structural characteristics are discussed that are necessary for hyaluronidases in order to exhibit hyaluronan cleavage.
Mesenchymal stem cells (MSC) are capable of differentiating into bone, fat, cartilage, tendon and other organ progenitor cells. Despite the abundance of MSC within the organism, little is known about their in vivo properties or about their corresponding in vivo niches. We therefore isolated MSC from spongy (cancellous) bone biopsies of healthy adults. When compared with the surrounding marrow, a fourfold higher number of colony-forming units was found within the tight meshwork of trabecular bone surface. At these sites, oxygen concentrations range from 1% to 7%. In MSC cultured at oxygen as low as 3%, rates for cell death and hypoxia-induced gene transcription remained unchanged, while in vitro proliferative lifespan was significantly increased, with about 10 additional population doublings before reaching terminal growth arrest. However, differentiation capacity into adipogenic progeny was diminished and no osteogenic differentiation was detectable at 3% oxygen. In turn, MSC that had previously been cultured at 3% oxygen could subsequently be stimulated to successfully differentiate at 20% oxygen. These data support our preliminary finding that primary MSC are enriched at the surface of spongy bone. Low oxygen levels in this location provide a milieu that extends cellular lifespan and furthermore is instructive for the stemness of MSC allowing proliferation upon stimulation while suppressing differentiation.
d -amino acids are present in some peptides from amphibian skin. These residues are derived from the corresponding l -amino acids present in the respective precursors. From skin secretions of Bombinae , we have isolated an enzyme that catalyzes the isomerization of an l -Ile in position 2 of a model peptide to d -allo-Ile. In the course of this reaction, which proceeds without the addition of a cofactor, radioactivity from tritiated water is incorporated into the second position of the product. The amino acid sequence of this isomerase could be deduced from cloned cDNA and genomic DNA. After expression of this cDNA in oocytes of Xenopus laevis , isomerase activity could be detected. Polypeptides related to the frog skin enzyme are present in several vertebrate species, including humans.
