Genetic evidence for a protective role for heat shock factor 1 and heat shock protein 70 against colitis

Inflammatory bowel disease (IBD) involves infiltration of leukocytes into intestinal tissue, resulting in intestinal damage induced by reactive oxygen species (ROS). Pro-inflammatory cytokines and cell adhesion molecules (CAMs) play important roles in this infiltration of leukocytes. The roles of heat shock factor 1 (HSF1) and heat shock proteins (HSPs) in the development of IBD are unclear. In this study, we examined the roles of HSF1 and HSPs in an animal model of IBD, dextran sulfate sodium (DSS)-induced colitis. The colitis worsened or was ameliorated in HSF1-null mice or transgenic mice expressing HSP70 (or HSF1), respectively. Administration of DSS up-regulated the expression of HSP70 in colonic tissues in an HSF1-dependent manner. Expression of pro-inflammatory cytokines and CAMs and the level of cell death observed in colonic tissues were increased or decreased in DSS-treated HSF1-null mice or transgenic mice expressing HSP70, respectively, relative to control wild-type mice. Relative to macrophages from control wild-type mice, macrophages prepared from HSF1-null mice or transgenic mice expressing HSP70 displayed enhanced or reduced activity, respectively, for the generation of pro-inflammatory cytokines in response to lipopolysaccharide stimulation. Suppression of HSF1 or HSP70 expression in vitro stimulated lipopolysaccharide-induced up-regulation of CAMs or ROS-induced cell death, respectively. This study provides the first genetic evidence that HSF1 and HSP70 play a role in protecting against DSS-induced colitis. Furthermore, this protective role seems to involve various mechanisms, such as suppression of expression of pro-inflammatory cytokines and CAMs and ROS-induced cell death.     

Programmed cell death is not a necessary prerequisite for fusion of the fetal mouse palate

It has been widely accepted that programmed cell death (PCD) is an essential event in palatogenesis and that its failure can result in cleft palate, one of the most common birth defects in the human. However, some conflicting results have been reported concerning the timing of cell death occurring in the fusing palate and therefore the role of PCD in palatal fusion is controversial. In order to clarify whether cell death is indispensable for mammalian palatogenesis, we cultivated the palates of day-13 mouse fetuses in vitro and prevented cell death by treating them with the inhibitors of caspases-1 and -3 or with aurintricarboxylic acid which inhibits the activity of caspase-activated DNase. Even when cell death was almost completely inhibited, palatal fusion took place successfully. Histological examination revealed that in the absence of apoptotic cell death, the medial edge epithelia of opposing palatal shelves adhered to each other and subsequently, the midline epithelial seam was disrupted and disappeared to bring about mesenchymal confluence across the palate. It seems that cell death is not a necessary prerequisite for palatal fusion but it may help to efficiently eliminate unnecessary cells which failed to migrate or differentiate properly.     

Activation of heat shock factor 1 by pyrrolidine dithiocarbamate is mediated by its activities as pro-oxidant and thiol modulator

Pyrrolidine dithiocarbamate (PDTC) is known to inhibit NF-kappa B, which plays a critical role(s) as an immediate early mediator of immune and inflammatory responses. Here we show that PDTC induces heat shock factor 1 (HSF1) activation and heat shock protein expression, while other antioxidants such as butylated hydroxytoluene (BHT), n-propylgallate (PG), ascorbic acid (AA), and N-acetyl-L-cysteine (NAC) do not. Since PDTC exerts other functions than antioxidant, e.g., a pro-oxidant, metal chelator, and thiol group modulator, we examined which of these activities is responsible for the PDTC-induced HSF1 activation. PDTC-induced HSF1 activation was not prevented by metal chelators, EDTAs, indicating that the metal chelating effect of PDTC is not linked to the HSF1 activation. PDTC increased intracellular GSSG level. In addition, PDTC-induced activation of HSF1 was significantly inhibited by NAC and a thiol-reducing agent dithiothreitol (DTT), while it was partially prevented by other antioxidants, AA, BHT, and PG. These results suggest that the activation of HSF1 by PDTC may be due to its activities as pro-oxidant and thiol group modulator rather than anti-oxidant.     

Establishment of thermotolerance in maize by exposure to stresses other than a heat shock does not require heat shock protein synthesis

Maize (Zea mays) seedlings were pretreated prior to heat shock with either a progressive water stress of −0.25 megapascal PEG/hour from 0 to −1.25 megapascal over a 6-hour time period, or various concentrations of copper, cadmium, or zinc for 4 days. When the subsequent heat shock of 40 or 45°C was administered for 3 hours, the seedlings showed an induced thermotolerance to these temperatures, which were otherwise lethal to control (water grown) seedlings. Thermotolerance was exhibited by both the root and the shoot of pretreated seedlings, even though the water and heavy metal stresses were applied only to the roots. Neither of these pretreatments had induced the synthesis of detectable levels of heat shock proteins (Hsps) at the time of heat shock. Pretreatment of seedlings with a progressive heat shock of 2°C/hour from 26 to 36°C, which did induce Hsps 18, 70, and 84, resulted in tolerance of a severe water stress of −1.5, −1.75, or −2.0 megapascal for 24 hours. But these seedlings producing Hsps were no better protected against water stress than those pretreated with a progressive water stress which did not produce Hsps. Hsps appear not to act as general stress proteins and their presence is not always required for the establishment of thermotolerance.     

Endogenous thrombospondin-1 is not necessary for proliferation but is permissive for vascular smooth muscle cell responses to platelet-derived growth factor.

We have reexamined the role of endogenous thrombospondin-1 (TSP1) in growth and motility of vascular smooth muscle cells (SMCs). Based on the ability of aortic-derived SMCs isolated from TSP1 null mice and grown in the absence of exogenous TSP1 to grow at comparable rates and to a slightly higher density than equivalent cells from wild-type mice, TSP1 is not necessary for their growth. Low concentrations of exogenous TSP1 stimulate growth of TSP1 null SMCs, but higher doses of TSP1 or its C-terminal domain are inhibitory. However, SMCs from TSP1 null mice are selectively deficient in chemotactic and proliferative responses to platelet-derived growth factor and in outgrowth in three-dimensional cultures. Recombinant portions of the N- and C-terminal domains of TSP1 stimulate SMC chemotaxis through different integrin receptors. Based on these data, the relative deficiency in SMC outgrowth during an ex vivo angiogenic response of muscle tissue from TSP1 null mice is probably due to restriction of platelet-derived growth factor dependent SMC migration and/or proliferation.