Induction of vascular endothelial phenotype and cellular proliferation from human cord blood stem cells cultured in simulated microgravity

Recent studies have demonstrated that stem cells derived from adult hematopoietic tissues are capable of trans-differentiation into non-hematopoietic cells, and that the culture in microgravity (microg) may modulate the proliferation and differentiation. We investigated the application of microg to human umbilical cord blood stem cells (CBSC) in the induction of vascular endothelial phenotype expression and cellular proliferation. CD34+ mononuclear cells were isolated from waste human umbilical cord blood samples and cultured in simulated microg for 14 days. The cells were seeded in rotary wall vessels (RWV) with or without microcarrier beads (MCB) and vascular endothelial growth factor was added during culture. Controls consisted of culture in 1 G. The cell cultures in RWV were examined by inverted microscopy. Cell counts, endothelial cell and leukocyte markers performed by flow cytometry and FACS scan were assayed at days 1, 4, 7 and at the termination of the experiments. Culture in RWV revealed significantly increased cellular proliferation with three-dimensional (3D) tissue-like aggregates. At day 4, CD34+ cells cultured in RWV bioreactor without MCB developed vascular tubular assemblies and exhibited endothelial phenotypic markers. These data suggest that CD34+ human umbilical cord blood progenitors are capable of trans-differentiation into vascular endothelial cell phenotype and assemble into 3D tissue structures. Culture of CBSC in simulated microg may be potentially beneficial in the fields of stem cell biology and somatic cell therapy.     

Altered expression of mRNAs implicated in osteogenesis under conditions of simulated microgravity is regulated by CD200:CD200R

Mouse calvarial cells grown under simulated microgravity conditions (neutral buoyancy) show preferential differentiation towards the osteoclast lineage, as defined by surrogate mRNAs, bone nodule growth and TRAP⁺ cells, when compared with cells cultured under normal gravity conditions. This effect was suppressed in cultures which contained the immunoregulatory molecule CD200, and conversely enhanced by anti-CD200 mAb. Concomitant increases occur in expression of inflammatory cytokines, and their mRNAs, under simulated microgravity conditions. Again cultures containing exogenous CD200 showed suppressed cytokine and cytokine mRNA expression. Further alterations in osteoclastogenesis were seen using cells isolated from cytokine-receptor knockout mice. We conclude that, as assessed by altered expression of mRNAs associated with osteoblast differentiation, CD200:CD200R interactions play an important regulatory role in the enhanced osteoclastogenesis seen under simulated microgravity conditions, with changes in cytokine expression further modulating this effect.     

Evaluation of bleomycin-induced chromosome aberrations under simulated microgravity conditions in human lymphocytes using "FISH" techniques

In the present investigation we report the effects of simulated microgravity conditions (clinostat) on the induction of chromosomal aberrations in human lymphocytes in vitro by (R) Bleomycin. Chromosomal aberrations have been analysed by means of fluorescent in situ hybridisation (FISH) and chromosome-specific composite DNA probes (chromosome painting). The results obtained show that, under simulated microgravity conditions, the levels of both symmetrical and asymmetrical (dicentrics, rings), the number of cells bearing "complex" aberrations and hence the total numbers of aberrations were significantly elevated at any of the dose-levels assayed, compared to the parallel treatments performed as 1g control ("ground"). Furthermore, the ratio symmetrical:asymmetrical translocations was markedly elevated under simulated microgravity conditions, compared to the findings usually observed under "normal" 1g conditions. On these bases, we are much inclined to believe that simulated microgravity, rather than limiting the resealing of DNA double strand breaks (DSB's) induced by genotoxic agents is influencing in terms of enhancement the misrejoining of DSB's which is actually responsible for the fixation of the original lesions to DNA into chromosomal aberrations. In addition, the possible different misrepair processes leading to the formation of symmetrical and asymmetrical translocations might be differentially influenced by microgravity being the symmetrical translocations significantly more represented.