Research Containing: Receptors

Spaceflight impairs antigen-specific tolerance induction in vivo and increases inflammatory cytokines

by cfynanon 22 August 2016in Biology & Biotechnology No comment

The health risks of a dysregulated immune response during spaceflight are important to understand as plans emerge for humans to embark on long-term space travel to Mars. In this first-of-its-kind study, we used adoptive transfer of T-cell receptor transgenic OT-II CD4 T cells to track an in vivo antigen-specific immune response that was induced during the course of spaceflight. Experimental mice destined for spaceflight and mice that remained on the ground received transferred OT-II cells and cognate peptide stimulation with ovalbumin (OVA) 323-339 plus the inflammatory adjuvant, monophosphoryl lipid A. Control mice in both flight and ground cohorts received monophosphoryl lipid A alone without additional OVA stimulation. Numbers of OT-II cells in flight mice treated with OVA were significantly increased by 2-fold compared with ground mice treated with OVA, suggesting that tolerance induction was impaired by spaceflight. Production of proinflammatory cytokines were significantly increased in flight compared with ground mice, including a 5-fold increase in IFN-gamma and a 10-fold increase in IL-17. This study is the first to show that immune tolerance may be impaired in spaceflight, leading to excessive inflammatory responses.

Modeled microgravity inhibits osteogenic differentiation of human mesenchymal stem cells and increases adipogenesis

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Space flight-induced bone loss has been attributed to a decrease in osteoblast function, without a significant change in bone resorption. To determine the effect of microgravity (MG) on bone, we used the Rotary Cell Culture System [developed by the National Aeronautics and Space Administration (NASA)] to model MG. Cultured mouse calvariae demonstrated a 3-fold decrease in alkaline phosphatase (ALP) activity and failed to mineralize after 7 d of MG. ALP and osteocalcin gene expression were also decreased. To determine the effects of MG on osteoblastogenesis, we cultured human mesenchymal stem cells (hMSC) on plastic microcarriers, and osteogenic differentiation was induced immediately before the initiation of modeled MG. A marked suppression of hMSC differentiation into osteoblasts was observed because the cells failed to express ALP, collagen 1, and osteonectin. The expression of runt-related transcription factor 2 was also inhibited. Interestingly, we found that peroxisome proliferator-activated receptor gamma (PPARgamma2), which is known to be important for adipocyte differentiation, adipsin, leptin, and glucose transporter-4 are highly expressed in response to MG. These changes were not corrected after 35 d of readaptation to normal gravity. In addition, MG decreased ERK- and increased p38-phosphorylation. These pathways are known to regulate the activity of runt-related transcription factor 2 and PPARgamma2, respectively. Taken together, our findings indicate that modeled MG inhibits the osteoblastic differentiation of hMSC and induces the development of an adipocytic lineage phenotype. This work will increase understanding and aid in the prevention of bone loss, not only in MG but also potentially in age-and disuse-related osteoporosis.

Simulated Microgravity Maintains the Undifferentiated State and Enhances the Neural Repair Potential of Bone Marrow Stromal Cells

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Recently, regenerative medicine with bone marrow stromal cells (BMSCs) has gained significant attention for the treatment of central nervous system diseases. Here, we investigated the activity of BMSCs under simulated microgravity conditions. Mouse BMSCs (mBMSCs) were isolated from C57BL/6 mice and harvested in 1G condition. Subjects were divided into 4 groups: cultured under simulated microgravity and 1G condition in growth medium and neural differentiation medium. After 7 days of culture, the mBMSCs were used for morphological analysis, reverse transcription (RT)-polymerase chain reaction, immunostaining analysis, and grafting. Neural-induced mBMSCs cultured under 1G conditions exhibited neural differentiation, whereas those cultured under simulated microgravity did not. Moreover, under simulated microgravity conditions, mBMSCs could be cultured in an undifferentiated state. Next, we intravenously injected cells into a mouse model of cerebral contusion. Graft mBMSCs cultured under simulated microgravity exhibited greater survival in the damaged region, and the motor function of the grafted mice improved significantly. mBMSCs cultured under simulated microgravity expressed CXCR4 on their cell membrane. Our study indicates that culturing cells under simulated microgravity enhances their survival rate by maintaining an undifferentiated state of cells, making this a potentially attractive method for culturing donor cells to be used in grafting.

Related URLs:
<Go to ISI>://WOS:000290255300013

Simultaneous expansion and harvest of hematopoietic stem cells and mesenchymal stem cells derived from umbilical cord blood

by cfynanon 9 June 2015in Biology & Biotechnology No comment

The simultaneous expansion and harvest of hematopoietic stem cells and mesenchymal stem cells derived from umbilical cord blood were carried out using bioreactors. The co-culture of umbilical cord blood (UCB)-derived hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs) was performed within spinner flasks and a rotating wall vessel (RWV) bioreactor using glass-coated styrene copolymer (GCSC) microcarriers. The medium used was composed of serum-free IMDM containing a cocktail of SCF 15 ng center dot mL(-1), FL 5 ng center dot mL(-1), TPO 6 ng center dot mL(-1), IL-3 15 ng center dot mL(-1), G-CSF 1 ng center dot mL(-1) and GM-CSF 5 ng center dot mL(-1). Accessory stromal cells derived from normal allogeneic adipose tissue were encapsulated in alginate-chitosan (AC) beads and used as feeding cells. The quality of the harvested UCB-HSCs and MSCs was assessed by immunophenotype analysis, methylcellulose colony and multi-lineage differentiation assays. After 12 days of culture, the fold-expansion of total cell numbers, colony-forming units (CFU-C), CD34(+)/CD45(+)/CD105(-) (HSCs) cells and CD34(-)/CD45(-)/CD105(+) (MSCs) cells using the RWV bioreactor were (3.7 +/- A 0.3)- , (5.1 +/- A 1.2)- , (5.2 +/- A 0.4)- , and (13.9 +/- A 1.2)-fold respectively, significantly better than those obtained using spinner flasks. Moreover, UCB-HSCs and UCB-MSCs could be easily separated by gravity sedimentation after the co-culture period as only UCB-MSCs adhered on to the microcarriers. Simultaneously, we found that the fibroblast-like cells growing on the surface of the GCSC microcarriers could be induced and differentiated towards the osteoblastic, chondrocytic and adipocytic lineages. Phenotypically, these cells were very similarly to the MSCs derived from bone marrow positively expressing the MSCs-related markers CD13, CD44, CD73 and CD105, while negatively expressing the HSCs-related markers CD34, CD45 and HLA-DR. It was thus demonstrated that the simultaneous expansion and harvest of UCB-HSCs and UCB-MSCs is possible to be accomplished using a feasible bioreactor culture system such as the RWV bioreactor with the support of GCSC microcarriers.

Related URLs:
<Go to ISI>://WOS:000284600400014

Properties of mechano-transduction via simulated microgravity and its effects on intracellular trafficking of VEGFR's

by cfynanon 9 June 2015in Biology & Biotechnology No comment

This study emphasizes the dynamical properties of mechanical loading via simulated microgravity, its effect on acute myeloid leukemia proliferation and hematopoietic stem cell (HSPC) growth and its implications in the area of tissue regeneration. Data presented illustrates that mechanical transduction changes the expression of humoral factors by facilitating paracrine/autocrine signalling, therefore modulating intracellular trafficking of tyrosine kinase receptors. Understanding mechano-transduction in the context of cell and tissue morphogenesis is the major focus of this study. The effects of external physiological stresses, such as blood flow, on several cellular subtypes seem to produce very intricate cellular responses. It is well accepted that mechanical loading plays an intrinsic and extrinsic influence on cell survival. This study shows how microgravity effects hematopoietic stem cells, and human leukemic cell proliferation and expression of its receptors that control cell survival, such as the tyrosine kinase vascular endothelial growth factor receptor-1, receptor-2 and receptor-3.

Detection of renal and urinary tract proteins before and after spaceflight

by cfynanon 9 June 2015in Biology & Biotechnology No comment

BACKGROUND: The recent evolution of genomics and subsequently proteomics offers a major advance in the ability to understand individual human variation in disease and the molecular level changes induced by certain environmental exposures. This original study examines urinary proteome composition to enable the understanding of molecular homeostatic mechanisms in spaceflight and presents the potential for early detection of subclinical disease, microgravity risk mitigation strategies, and countermeasure development for exploration-class missions. METHODS: The urinary proteome composition of six Russian cosmonauts (men, ages 35-51) who flew long-duration missions of 169-199 d was determined 30 d before flight and compared to repeat studies 1 and 7 d postflight. RESULTS: There were 430 proteins identified. Of those, 15 proteins originated in the renal tissues. Of the 15 urinary proteins, 10 were consistently present in the urine. However, the presence of five of the urinary proteins–neutral endopeptidase (NEP), afamin (AFAM), aquaporin-2 (AQP2), aminopeptidase A (AMPE), and dipeptidyl peptidase 4 (DPP4)–was dependent on spaceflight exposure. DISCUSSION: Proteomic investigation of pre- and postflight urine and bioinformation approaches to proteome analysis provide important data relative the mechanism of kidney function in spaceflight. In this initial study, we determined that the evaluation of urinary proteins may help investigators understand changes that are occurring in microgravity. Once additional ground-based and in-flight data are collected, it is feasible to develop targeted studies for tracking specific spaceflight related changes, determine countermeasure and risk-mitigation effectiveness, and possibly detect subclinical disease in flight crewmembers.

The impact of long-term exposure to space environment on adult mammalian organisms: a study on mouse thyroid and testis

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Hormonal changes in humans during spaceflight have been demonstrated but the underlying mechanisms are still unknown. To clarify this point thyroid and testis/epididymis, both regulated by anterior pituitary gland, have been analyzed on long-term space-exposed male C57BL/10 mice, either wild type or pleiotrophin transgenic, overexpressing osteoblast stimulating factor-1. Glands were submitted to morphological and functional analysis.In thyroids, volumetric ratios between thyrocytes and colloid were measured. cAMP production in 10(-7)M and 10(-8)M thyrotropin-treated samples was studied. Thyrotropin receptor and caveolin-1 were quantitized by immunoblotting and localized by immunofluorescence. In space-exposed animals, both basal and thyrotropin-stimulated cAMP production were always higher. Also, the structure of thyroid follicles appeared more organized, while thyrotropin receptor and caveolin-1 were overexpressed. Unlike the control samples, in the space samples thyrotropin receptor and caveolin-1 were both observed at the intracellular junctions, suggesting their interaction in specific cell membrane microdomains.In testes, immunofluorescent reaction for 3beta- steroid dehydrogenase was performed and the relative expressions of hormone receptors and interleukin-1beta were quantified by RT-PCR. Epididymal sperm number was counted. In space-exposed animals, the presence of 3beta and 17beta steroid dehydrogenase was reduced. Also, the expression of androgen and follicle stimulating hormone receptors increased while lutenizing hormone receptor levels were not affected. The interleukin 1 beta expression was upregulated. The tubular architecture was altered and the sperm cell number was significantly reduced in spaceflight mouse epididymis (approx. -90% vs. laboratory and ground controls), indicating that the space environment may lead to degenerative changes in seminiferous tubules.Space-induced changes of structure and function of thyroid and testis/epididymis could be responsible for variations of hormone levels in human during space missions. More research, hopefully a reflight of MDS, would be needed to establish whether the space environment acts directly on the peripheral glands or induces changes in the hypotalamus-pituitary-glandular axis.

Reconstruction of Functional Cortical-like Tissues from Neural Stem and Progenitor Cells

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Neural stem and progenitor cells isolated from embryonic day 13 rat cerebral cortex were immobilized in three-dimensional type I collagen gels, and then the cell-collagen constructs were transferred to rotary wall vessel bioreactors and cultured in serum-free medium containing basic fibroblast growth factor (bFGF) combined with brain-derived neurotrophic factor for up to 10 weeks. Remarkably, the collagen-entrapped cells formed a complex two-layered structure that emulated to a certain extent the cerebral cortex of the embryonic brain in architecture and functionality. The surface layer (layer I) composed primarily of proliferating neural progenitor cells (nestin(+), vimentin(+), and PCNA(+)) predominantly expressed functional neurotransmitter receptors for cholinergic and purinergic agonists while differentiating cells (TuJ1(+) and GFAP(+)) in the deeper layer (layer II) contained differentiated neurons and astrocytes and mainly responded to GABAergic and glutamatergic agonists and to veratridine, which activates voltage-dependent Na(+) channels. An active synaptic vesicle recycling was demonstrated by neuronal networks in the deeper layer using the endocytotic marker FM1-43. Cell polarization forming the characteristic two-layered structure was found to associate with the bFGF and FGF receptor signaling. These engineered functional tissue constructs have a potential use as tissue surrogates for drug screening and detection of environmental toxins, and in neural cell replacement therapy.

Related URLs:
<Go to ISI>://WOS:000259998000009

Microarray analysis of spaceflown murine thymus tissue reveals changes in gene expression regulating stress and glucocorticoid receptors

by cfynanon 9 June 2015in Biology & Biotechnology No comment

The detrimental effects of spaceflight and simulated microgravity on the immune system have been extensively documented. We report here microarray gene expression analysis, in concert with quantitative RT-PCR, in young adult C57BL/6NTac mice at 8 weeks of age after exposure to spaceflight aboard the space shuttle (STS-118) for a period of 13 days. Upon conclusion of the mission, thymus lobes were extracted from space flown mice (FLT) as well as age- and sex-matched ground control mice similarly housed in animal enclosure modules (AEM). mRNA was extracted and an automated array analysis for gene expression was performed. Examination of the microarray data revealed 970 individual probes that had a 1.5-fold or greater change. When these data were averaged (n = 4), we identified 12 genes that were significantly up- or down-regulated by at least 1.5-fold after spaceflight (P < or = 0.05). The genes that significantly differed from the AEM controls and that were also confirmed via QRT-PCR were as follows: Rbm3 (up-regulated) and Hsph110, Hsp90aa1, Cxcl10, Stip1, Fkbp4 (down-regulated). QRT-PCR confirmed the microarray results and demonstrated additional gene expression alteration in other T cell related genes, including: Ctla-4, IFN-alpha2a (up-regulated) and CD44 (down-regulated). Together, these data demonstrate that spaceflight induces significant changes in the thymic mRNA expression of genes that regulate stress, glucocorticoid receptor metabolism, and T cell signaling activity. These data explain, in part, the reported systemic compromise of the immune system after exposure to the microgravity of space.

Regulatory cross talks of bone cells, hematopoietic stem cells and the nervous system maintain hematopoiesis

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Adult hematopoietic stem cells (HSC) continuously replenish the blood with immune and blood cells with a finite life span, from the bone marrow (BM) reservoir of immature and maturing leukocytes. Regulation of HSC migration and development is essential for their function and blood cell production. These diverse and multiple states require a tight regulation to efficiently address host defense and repair requirements. Numerous recent studies disclose a central role for bone related cells in regulation of HSC and hematopoiesis. During ontogeny HSC home and seed the fetal BM in the last gestation period when the bone is already ossified. Ossification involves bone forming osteoblast- and bone degrading osteoclast activity and is considered essential for the formation of BM cavities and hematopoiesis. This synchronized association implies the need for active bone cells and bone turnover for HSC regulation. Osteoblastic cells and SDF- 1+/nestin+ reticular adventitial and CAR cells are crucial for regulation of HSC lodgment, self-renewal and function. Bone resorbing osteoclasts regulate bone turnover and progenitor cell detachment and release from the BM. Sympathetic signals from the nervous system activated by circadian rhythms or stress conditions control both bone turnover and HSC migration and development. In this review we discuss pathways and mechanisms involved in this orchestrated regulatory network. A special focus is made on the pivotal role of the SDF-1/CXCR4 axis as a determinant of HSC fate. Inflammation, DNA damage, cytokine mobilization, microgravity and aging are discussed as specific physiologic and pathologic events entailing dysregulation of the tightly balanced Bone-Brain-Blood triad.

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Research Containing: Receptors

Spaceflight impairs antigen-specific tolerance induction in vivo and increases inflammatory cytokines

Modeled microgravity inhibits osteogenic differentiation of human mesenchymal stem cells and increases adipogenesis

Simulated Microgravity Maintains the Undifferentiated State and Enhances the Neural Repair Potential of Bone Marrow Stromal Cells

Simultaneous expansion and harvest of hematopoietic stem cells and mesenchymal stem cells derived from umbilical cord blood

Properties of mechano-transduction via simulated microgravity and its effects on intracellular trafficking of VEGFR's

The impact of long-term exposure to space environment on adult mammalian organisms: a study on mouse thyroid and testis

Reconstruction of Functional Cortical-like Tissues from Neural Stem and Progenitor Cells

Microarray analysis of spaceflown murine thymus tissue reveals changes in gene expression regulating stress and glucocorticoid receptors

Regulatory cross talks of bone cells, hematopoietic stem cells and the nervous system maintain hematopoiesis