T cell activation responses are differentially regulated during clinorotation and in spaceflight

Abstract

Studies of T lymphocyte activation with mitogenic lectins during spaceflight have shown a dramatic inhibition of activation as measured by DNA synthesis at 72 h, but the mechanism of this inhibition is unknown. We have investigated the progression of cellular events during the first 24 h of activation using both spaceflight microgravity culture and a ground-based model system that relies on the low shear culture environment of a rotating clinostat (clinorotation). Stimulation of human peripheral blood mononuclear cells (PBMCs) with soluble anti-CD3 (Leu4) in clinorotation and in microgravity culture shows a dramatic reduction in surface expression of the receptor for IL-2 (CD25) and CD69. An absence of bulk RNA synthesis in clinorotation indicates that stimulation with soluble Leu4 does not induce transition of T cells from G0 to the G1 stage of the cell cycle. However, internalization of the TCR by T cells and normal levels of IL-1 synthesis by monocytes indicate that intercellular interactions that are required for activation occur during clinorotation. Complementation of TCR-mediated signaling by phorbol ester restores the ability of PBMCs to express CD25 in clinorotation, indicating that a PKC-associated pathway may be compromised under these conditions. Bypassing the TCR by direct activation of intracellular pathways with a combination of phorbol ester and calcium ionophore in clinorotation resulted in full expression of CD25; however, only partial expression of CD25 occurred in microgravity culture. Though stimulation of purified T cells with Bead-Leu4 in microgravity culture resulted in the engagement and internalization of the TCR, the cells still failed to express CD25. When T cells were stimulated with Bead-Leu4 in microgravity culture, they were able to partially express CD69, a receptor that is constitutively stored in intracellular pools and can be expressed in the absence of new gene expression. Our results suggest that the inhibition of T cell proliferative response in microgravity culture is a result of alterations in signaling events within the first few hours of activation, which are required for the expression of important regulatory molecules.

Signal Transduction in Primary Human T Lymphocytes in Altered Gravity During Parabolic Flight and Clinostat Experiments

“…Furthermore, CD3 surface expression reconstituted after T cell activation during 1g conditions but not in microgravity [27]. Investigations of TCR internalization in stimulated human PBMCs using rotating wall vessel system revealed that CD3 internalization was delayed in simulated microgravity [30] an effect in accordance with our findings of gravity-dependence of CD3 surface expression.…”

“…Under our chosen experimental conditions (60 rpm, 4 mm pipette diameter), a maximal residual acceleration of 2x10 -3 g was calculated at the outer radius of the pipette and decreases towards the center. 2D clinorotation with this experimental setup is an established model system for the simulation of microgravity using T lymphocytes and other cell types [13,30,31,32] A recent study (ESA Ground-based facilities / GBF project) compared different ground based microgravity platforms to real microgravity and identified fast 2D clinorotation as a good simulation of microgravity suitable for cell culture studies. In accordance to recommendations from the same study we will refer to clinorotation-induced microgravity as simulated microgravity [33].…”

Spaceflight Transcriptomes: Unique Responses to a Novel Environment

“…And while the divergence of plants and animals largely prevents specific gene comparisons in spaceflight responses, certain classes of genes seem similarly affected by spaceflight. Calcium-mediated signal transduction is impacted by spaceflight in both plants and animals (Hughes-Fulford et al, 1998;Hashemi et al, 1999;Paul et al, 2001;Salmi and Roux, 2008), and genes affected by calcium signaling are differentially expressed. Heat shock proteins are highly conserved in both kingdoms, and heat shock genes are upregulated in some spaceflight studies.…”

Modeled gravitational unloading triggers differentiation and apoptosis in preosteoclastic cells

“…Several investigations conducted in space laboratories and with instruments averaging the gravity vector, and thus simulating conditions of reduced gravity, have shown that isolated cells may change their behavior under altered gravitational conditions. Loss of cell activation [Cogoli et al, 1984], perturbation of signal transduction [Schmitt et al, 1996;Hashemi et al, 1999] and modification of genetic expression [Walther et al, 1998;Hammond et al, 1999] have been described. This suggests that some of the mechanisms at the roots of the systemic effects observed in weightlessness, such as depression of the immune system [Gmü nder and Cogoli, 1996] …”