Gene-metabolite profile integration to understand the cause of spaceflight induced immunodeficiency

Abstract

Spaceflight presents a spectrum of stresses very different from those associated with terrestrial conditions. Our previous study (BMC Genom. 15: 659, 2014) integrated the expressions of mRNAs, microRNAs, and proteins and results indicated that microgravity induces an immunosuppressive state that can facilitate opportunistic pathogenic attack. However, the existing data are not sufficient for elucidating the molecular drivers of the given immunosuppressed state. To meet this knowledge gap, we focused on the metabolite profile of spaceflown human cells. Independent studies have attributed cellular energy deficiency as a major cause of compromised immunity of the host, and metabolites that are closely associated with energy production could be a robust signature of atypical energy fluctuation. Our protocol involved inoculation of human endothelial cells in cell culture modules in spaceflight and on the ground concurrently. Ten days later, the cells in space and on the ground were exposed to lipopolysaccharide (LPS), a ubiquitous membrane endotoxin of Gram-negative bacteria. Nucleic acids, proteins, and metabolites were collected 4 and 8 h post-LPS exposure. Untargeted profiling of metabolites was followed by targeted identification of amino acids and knowledge integration with gene expression profiles. Consistent with the past reports associating microgravity with increased energy expenditure, we identified several markers linked to energy deficiency, including various amino acids such as tryptophan, creatinine, dopamine, and glycine, and cofactors such as lactate and pyruvate. The present study revealed a molecular architecture linking energy metabolism and immunodeficiency in microgravity. The energy-deficient condition potentially cascaded into dysregulation of protein metabolism and impairment of host immunity. This project is limited by a small sample size. Although a strict statistical screening was carefully implemented, the present results further emphasize the need for additional studies with larger sample sizes. Validating this hypothesis using an in vivo model is essential to extend the knowledge towards identifying markers of diagnostic and therapeutic value.

Fig. 1

Principal component analysis (PCA). a Gene expressions and b metabolite loads yielded unambiguous group separation. GC ground control, SC spaceflight control, GLPS 4 h LPS treatment on ground, SLPS, 4 h LPS treatment in spaceflight

Fig. 2

Spaceflight-induced stress in conjunction with LPS insult perturbs energy metabolism. The significantly altered genes and metabolites are integrated to construct this network. The shaded ovals are the metabolites and the edges represent their relationships. The arrowheads under each molecule represent their expressions attributed to microgravity (µG) in comparison to terrestrial condition (1 G) (left), and the 4 h LPS assaults in µG in comparison to 1 G (right). ▲ Represents up, ▼ represents down and ▬ represents no change

Fig. 3

Spaceflight-induced stress in conjunction with LPS insult perturbs amino acid metabolism. The significantly altered genes and metabolites are integrated to construct this network. The shaded ovals are the metabolites and the edges represent their relationships. The arrowheads under each molecule represent their expressions attributed to microgravity (µG) in comparison to terrestrial condition (1 G) (left), and the 4 h LPS assaults in µG in comparison to 1 G (right). ▲ Represents up, ▼ represents down and ▬ represents no change

Fig. 4

Correlative network analysis indicates predictive activation and inhibition of energy-associated networks caused by spaceflight-induced stress in conjunction with LPS insult: The arrowheads under µG represent their expressions attributed to microgravity (µG) in comparison to terrestrial condition (1 G) (left), and under LPS represents the 4 h LPS assaults in µG in comparison to 1 G (right). ▲ Represents up, ▼ represents down and ▬ represents no change

Fig. 5

Schematic illustration of the assay conducted simultaneously in spaceflight (STS-135) and on ground. Molecular perturbations caused by LPS assault in both µG and 1 G were assessed in comparison to the no-treatment control in HMVEC-dBL cells. Cell inoculation, treatment, and assay types for two assay variables are noted. The transcriptomic study previously published by us investigated the mRNA–miRNA–protein relationship and how it becomes vulnerable to spaceflight-induced stress. The boxed assay results are used in the present communication