Anti-HSP70 monoclonal antibody, clone 5F8

During chronic obstructive pulmonary disease (COPD) exacerbations, interplay between pathogen-associated molecular patterns (PAMPs; e.g. lipopolysaccharide (LPS) and lipoteichoic acid (LTA)) and damage-associated molecular patterns (DAMPs; e.g. extracellular heat shock protein 70 (eHsp70) and adenosine triphosphate (ATP)) might influence patient's outcome. Nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome might have a role in dysfunctional immune system in COPD. We hypothesized that LPS, LTA, eHsp70 and their combinations induce NLRP3 inflammasome activation, and we aimed to explore this assumption. We used monocytic (monocyte-derived macrophages (MDMs) and THP-1) and bronchial epithelial cells (NHBE and NCI-H292) to represent systemic and local airway compartments that could be affected in COPD. Bacterial components and eHsp70 stimulated NLRP3 and interleukin (IL)-1 beta gene expression as well as IL-1 beta and ATP release from all cells compared to non-treated cells. LDH secretion was induced in cell lines only. eHsp70 had inhibitory (NCI-H292) or stimulatory (NHBE) effects on eATP levels compared to PAMP alone. Regarding NLRP3 inflammasome activation, eHsp70 had mostly antagonistic effects. We demonstrated that bacterial components and eHsp70 activate NLRP3 inflammasome and increase ATP secretion. We suggest that extracellular Hsp70 might modulate immune responses provoked by bacterial infections and affect COPD patients' outcome during acute exacerbations.

In their role as molecular chaperones, heat shock proteins (Hsps) mediate protein folding thereby mitigating cellular damage caused by physiological and environmental stress. Nauplii of the crustaceanArtemia franciscanarespond to heat shock by producing Hsps; however, the effects of cold shock on Hsp levels inA. franciscanahave not been investigated previously. The effect of cold shock at 1 degrees C followed by recovery at 27 degrees C on the amounts ofArHsp90,Hsp70,ArHsp40, andArHsp40-2mRNA and their respective proteins inA. franciscananauplii was examined by quantitative PCR (qPCR) and immunoprobing of western blots. The sameHspmRNAs and proteins were also quantified during incubation of nauplii at their optimal growth temperature of 27 degrees C. qPCR analyses indicated that the abundance ofArHsp90,Hsp70, andArHsp40mRNA remained relatively constant during both cold shock and recovery and was not significantly different compared with levels at optimal temperature. Western blotting revealed that ArHsp90, ArHsp40, and ArHsp40-2 were generally below baseline, but at detectable levels during the 6 h of cold shock, and persisted in early recovery stages before declining. Hsp70 was the only protein that remained constant in quantity throughout cold shock and recovery. By contrast, all Hsps declined rapidly during 6 h when nauplii were incubated continuously at 27 degrees C optimal temperature. Generally, the amounts of ArHsp90, ArHsp40, and ArHsp40-2 were higher during cold shock/recovery than those during continuous incubation at 27 degrees C. Our data support the conclusion that low temperature preserves Hsp levels, making them available to assist in protein repair and recovery after cold shock.