Proteomic Evaluation of Inflammatory Proteins in Rat Spleen Interstitial Fluid and Lymph during LPS-Induced Systemic Inflammation Reveals Increased Levels of ADAMST1

Abstract

The spleen is a part of the immune system and is involved in the response to a systemic inflammation induced by blood borne pathogens that may induce sepsis. Knowledge about the protein composition of the spleen microenvironment in a control situation and during systemic inflammation may contribute to our understanding of the pathophysiology of sepsis. To our knowledge, the proteome of the fluid phase of the spleen microenvironment has not previously been investigated. In order to access the proximal fluid surrounding the splenic cells, we collected postnodal efferent spleen lymph from rats by cannulation, and spleen interstitial fluid (IF) by centrifugation. The origin of the isolated spleen IF was assessed by the extracellular tracer (51)Cr-EDTA and the plasma tracer (125)I-HSA. Spleen lymph, IF, and plasma samples were collected during lipopolysaccharide (LPS) induced systemic inflammation and analyzed using a cytokine multiplex assay and, for the first time, using label-free mass spectrometry based proteomics. The concentrations of TNF-α, IL-1β, IL-6, and IL-10 increased severalfold in all fluids after LPS exposure. In total, 281, 201, and 236 proteins were identified in lymph, IF, and plasma, respectively, and several of these were detected after LPS only. A disintegrin and metalloproteinase with thrombospondin motifs 1 (ADAMTS1) was detected by proteomics (the pro- region) in lymph only after LPS. ADAMTS1 was assessed by ELISA (the metalloproteinase domain), and the concentration was significantly higher in IF and lymph than in plasma in a control situation, showing local production in the spleen. A dramatic increase in ADAMTS1 was detected in lymph, IF, and plasma after LPS exposure. In conclusion, the procedures we used to isolate IF and lymph from the spleen during LPS enabled detection of locally produced proteins. Furthermore, we have demonstrated that the inflammatory proteome is different in the spleen microenvironment when compared to that in plasma.

ADAMTS1 protease is required for a balanced immune cell repertoire and tumour inflammatory response

“…Another interesting work reported significant levels of ADAMTS1 in interstitial fluid of spleens, lymph nodes and plasma after a systemic lipopolysaccharide-induced inflammation 48 . At this point, we should recall the original cloning of mouse Adamts1 in a cachexia model 6 , a phenomena with a recognized inflammatory component.…”

Temporal Gene Expression in the Hippocampus and Peripheral Organs to Endotoxin-Induced Systemic Inflammatory Response in Caspase-1-Deficient Mice

“…ADAMTS1 is necessary for normal growth, female fertility, and kidney, adrenal and female reproductive organ structure and function [46]. Recently, it was shown that LPS dramatically increased the synthesis of ADAMTS1 within the splenic lymph and interstitial fluid to a concentration that was much higher than that of plasma, suggesting that the spleen is a major source of ADAMTS1, and that this metalloprotease is involved in the endotoxin-induced disease process [47]. Within the brain, metalloproteases play a key role in controlling blood brain barrier permeability during LPS-induced inflammation, and ADAMTS1 may have a role in regulating neural plasticity, as null female mice have a selective decline of synaptic protein levels in the frontal cortex [48].…”

Lymphatic Vessel Network Structure and Physiology

“…In a rat model of sepsis (cecal ligation and puncture), proteomic analysis of mesenteric lymph revealed 158 proteins significantly changed, including elevations in ApoE, annexin-1, S100A8/9, and the lipocalin NGAL (1208). Analysis of postdnodal lymph from rats subjected to LPS exposure revealed elevations in inflammatory cytokines and the disintegrin ADAMTS1 (827). Findings from these proteomic studies can be influenced by choices of models and methodology.…”