Differential Regulation of Degradation and Immune Pathways Underlies Adaptation of the Ectosymbiotic Nematode Laxus Oneistus to Oxic-Anoxic Interfaces

Eukaryotes may experience oxygen deprivation under both physiological and pathological conditions. Because oxygen shortage leads to a reduction in cellular energy production, all eukaryotes studied so far conserve energy by suppressing their metabolism. However, the molecular physiology of animals that naturally and repeatedly experience anoxia is underexplored. One such animal is the marine nematode Laxus oneistus. It thrives, invariably coated by its sulfur-oxidizing symbiont Candidatus Thiosymbion oneisti, in anoxic sulfidic or hypoxic sand. Here, transcriptomics and proteomics showed that, whether in anoxia or not, L. oneistus mostly expressed genes involved in ubiquitination, energy generation, oxidative stress response, immune response, development, and translation. Importantly, ubiquitination genes were also highly expressed when the nematode was subjected to anoxic sulfidic conditions, together with genes involved in autophagy, detoxification and ribosome biogenesis. We hypothesize that these degradation pathways were induced to recycle damaged cellular components (mitochondria) and misfolded proteins into nutrients. Remarkably, when L. oneistus was subjected to anoxic sulfidic conditions, lectin and mucin genes were also upregulated, potentially to promote the attachment of its thiotrophic symbiont. Furthermore, the nematode appeared to survive oxygen deprivation by using an alternative electron carrier (rhodoquinone) and acceptor (fumarate), to rewire the electron transfer chain. On the other hand, under hypoxia, genes involved in costly processes (e.g., amino acid biosynthesis, development, feeding, mating) were upregulated, together with the worm’s Toll-like innate immunity pathway and several immune effectors (e.g., Bacterial Permeability Increasing proteins, fungicides). In conclusion, we hypothesize that, in anoxic sulfidic sand, L. oneistus upregulates degradation processes, rewires oxidative phosphorylation and by reinforces its coat of bacterial sulfur-oxidizers. In upper sand layers, instead, it appears to produce broad-range antimicrobials and to exploit oxygen for biosynthesis and development.

Differential regulation of degradation and immune pathways underlies adaptation of the symbiotic nematode Laxus oneistus to oxic-anoxic interfaces

Eukaryotes may experience oxygen deprivation under both physiological and pathological conditions. Because oxygen shortage leads to a reduction in cellular energy production, all eukaryotes studied so far conserve energy by suppressing their metabolism. However, the molecular physiology of animals that naturally and repeatedly experience anoxia underexplored. One such animal is the symbiotic marine nematode Laxus oneistus. It thrives, invariably coated by its sulfur-oxidizing bacterium Candidatus Thiosymbion oneisti, in anoxic sulfidic or hypoxic sand. Here, transcriptomics and proteomics showed that, whether in anoxia or not, L. oneistus mostly expressed genes involved in ubiquitination, energy generation, oxidative stress response, immune response, development, and translation. Importantly, ubiquitination genes were also highly expressed when the nematode was subjected to anoxic sulfidic conditions, together with genes involved in autophagy, detoxification, chaperone-encoding genes, and ribosome biogenesis. We hypothesize that these degradation pathways were induced to recycle damaged cellular components (mitochondria) and misfolded proteins into nutrients. Remarkably, when L. oneistus was subjected to anoxic sulfidic conditions, lectin genes were also upregulated, potentially to promote the attachment of its thiotrophic anaerobic symbiont. Furthermore, L. oneistus appeared to survive oxygen deprivation by using an alternative electron carrier (rhodoquinone) and acceptor (fumarate), to rewire the electron transfer chain. On the other hand, under hypoxia, genes involved in costly processes (e.g., amino acid biosynthesis, development, feeding, mating) were upregulated, together with the worm's Toll-like innate immunity pathway and several immune effectors (e.g., Bacterial Permeability Increasing proteins, fungicides). In conclusion, we hypothesize that, in anoxic sulfidic sand, L. oneistus survives by overexpressing degradation processes, rewiring oxidative phosphorylation and by reinforcing its coat of bacterial sulfur-oxidizers. In upper sand layers, instead, it appears to produce broad-range antimicrobials and to exploit oxygen for biosynthesis and development.

Oxygen Deprivation Modulates EGFR and PD-L1 in Squamous Cell Carcinomas of the Head and Neck

Abundance and signaling of the epidermal growth factor receptor (EGFR) and programmed cell death protein ligand 1 (PD-L1) in head and neck squamous cell carcinoma (HNSCC) are not only genetically determined but are also subject to the traits of the tumor microenvironment, which has hitherto not been clarified completely. We investigated the impact of hypoxia on the EGFR system and on PD-L1 in six HPV negative HNSCC cell lines in vitro and in FaDu xenografts in vivo. Protein levels of EGFR, AKT, pAKT, ERK1/2, pERK1/2, CA IX, cleaved PARP (apoptosis), LC3B (autophagy), and PD-L1 were quantified by western blot after oxygen deprivation or CoCl2, staurosporine, and erlotinib treatment. In FaDu xenograft tumors the expression of EGFR, CA IX andCD34 staining were analyzed. Reduced oxygen supply strongly downregulated EGFR protein levels and signaling in FaDu cells in vitro and in vivo, and a transient downregulation of EGFR signaling was found in three other HNSCC cell lines. PD-L1 was affected by oxygen deprivation in only one HNSCC cell line showing increased protein amounts. The results of this study indicate a significant impact of the traits of the tumor microenvironment on crucial molecular targets of cancer therapies with high clinical relevance for therapy resistance and response in HNSCC.

Classification of Brain Injury Pathophysiology With GFAP and UCH-L1

Brain injury is pathophysiologically diverse, with many cases presenting with mixed pathologies. Utilizing serum biomarkers to investigate the pathophysiology of injury would help to aid in understanding prognosis and targeting therapeutics. One goal of the study is to develop a traumatic brain injury classification scheme based on two serum biomarkers glial fibrillary acidic protein (GFAP) and ubiquitin carboxy-terminal L1 (UCH-L1). GFAP and UCH-L1 serum marker analysis was performed on patients with isolated traumatic brain injury or healthy, uninjured controls within 32 hours of hospital admission. Machine learning was utilized for classification of brain injury and to develop a novel algorithm capable of classifying the type of brain injury based on GFAP and UCH-L1 concentrations. Each patient’s brain injury was classified using standard clinical and radiographic assessments and stratified into one of four trauma groups: trauma, spontaneous hemorrhage, oxygen deprivation, or a high-velocity trauma with negative radiographic finding. Analysis of prospectively collected serum for GFAP and UCH-L1 was performed on 61 patients and 39 controls. The subjects with trauma, spontaneous hemorrhages and oxygen deprivation could be distinguished from controls with AUC = 1.00. Combination of GFAP and UCH-L1 concentrations distinguished the high-velocity injuries that were negative for radiographic indicators (CT-negative) from controls with AUC of 0.93. Serum biomarker profiles were found to accurately predict etiology across four distinct brain injuries, including CT-negative. Serum markers GFAP and UCHL1 may be helpful for classifying the nature of brain injury, which will aid with prognostication and development of therapeutics.

Preconditioning of Caenorhabditis elegans to Anoxic Insult by Inactivation of Cholinergic, GABAergic, and Muscle Activity

For most metazoans, oxygen deprivation leads to cell dysfunction and if severe, death. Sublethal stress prior to a hypoxic or anoxic insult (“preconditioning”) can protect cells from subsequent oxygen deprivation. The molecular mechanisms by which sublethal stress can buffer against a subsequent toxic insult and the role of the nervous system in the response are not well understood. We studied the role of neuronal activity preconditioning to oxygen deprivation in C. elegans. Animals expressing the histamine gated chloride channels (HisCl1) in select cell populations were used to temporally and spatially inactivate the nervous system or tissue prior to an anoxic insult. We find that inactivation of the nervous system for 3 hours prior to the insult confers resistance to a 48-hour anoxic insult in 4th-stage larval animals. Experiments show that this resistance can be attributed to loss of activity in cholinergic and GABAergic neurons as well as in body wall muscles. These observations indicate that the nervous system activity can mediate the organism’s response to anoxia.

Ischemia-Like Stress Conditions Stimulate Trophic Activities of Adipose-Derived Stromal/Stem Cells

Adipose-derived stromal/stem cells (ASCs) have been shown to exert regenerative functions, which are mainly attributed to the secretion of trophic factors. Upon transplantation, ASCs are facing an ischemic environment characterized by oxygen and nutrient deprivation. However, current knowledge on the secretion capacity of ASCs under such conditions is limited. Thus, the present study focused on the secretory function of ASCs under glucose and oxygen deprivation as major components of ischemia. After exposure to glucose/oxygen deprivation, ASCs maintained distinct viability, but the metabolic activity was greatly reduced by glucose limitation. ASCs were able to secrete a broad panel of factors under glucose/oxygen deprivation as revealed by a cytokine antibody array. Quantification of selected factors by ELISA demonstrated that glucose deprivation in combination with hypoxia led to markedly higher secretion levels of the angiogenic and anti-apoptotic factors IL-6, VEGF, and stanniocalcin-1 as compared to the hypoxic condition alone. A conditioned medium of glucose/oxygen-deprived ASCs promoted the viability and tube formation of endothelial cells, and the proliferation and migration of fibroblasts. These findings indicate that ASCs are stimulated by ischemia-like stress conditions to secrete trophic factors and would be able to exert their beneficial function in an ischemic environment.