No abstract

The recognition of invading microbes followed by the induction of effective innate immune response is crucial for host survival. Human surface epithelial cells are situated at host-environment boundaries and thus act as the first line of host defense against invading microbes. They recognize the microbial ligands via Toll-like receptors (TLRs) expressed on the surface of epithelial cells. TLR2 has gained importance as a major receptor for a variety of microbial ligands. In contrast to its high expression in lymphoid tissues, TLR2 is expressed at low level in epithelial cells. Thus, it remains unclear whether the low amount of TLR2 expressed in epithelial cells is sufficient for mediating bacteria-induced host defense and immune response and whether TLR2 expression can be upregulated by bacteria during infection. Here, we show that TLR2, although expressed at very low level in unstimulated human epithelial cells, is greatly up-regulated by nontypeable Hemophilus influenzae (NTHi), an important human bacterial pathogen causing otitis media and chronic obstructive pulmonary diseases. Activation of an IKK␤-IB␣-dependent NF-B pathway is required for TLR2 induction, whereas inhibition of the MKK3/6-p38␣/␤ pathway leads to enhancement of NTHi-induced TLR2 up-regulation. Surprisingly, glucocorticoids, well known potent anti-inflammatory agents, synergistically enhance NTHi-induced TLR2 up-regulation likely via a negative cross-talk with the p38 MAP kinase pathway. These studies may bring new insights into the role of bacteria and glucocorticoids in regulating host defense and immune response and lead to novel therapeutic strategies for modulating innate immune and inflammatory responses for otitis media and chronic obstructive pulmonary diseases.

Low-intensity electrical current (or mild electrical stimulation; MES) influences signal transduction and activates phosphatidylinositol-3 kinase (PI3K)/Akt pathway. Because insulin resistance is characterized by a marked reduction in insulin-stimulated PI3K-mediated activation of Akt, we asked whether MES could increase Akt phosphorylation and ameliorate insulin resistance. In addition, it was also previously reported that heat shock protein 72 (Hsp72) alleviates hyperglycemia. Thus, we applied MES in combination with heat shock (HS) to in vitro and in vivo models of insulin resistance. Here we show that 10-min treatment with MES at 5 V (0.1 ms pulse duration) together with HS at 42°C increased the phosphorylation of insulin signaling molecules such as insulin receptor substrate (IRS) and Akt in HepG2 cells maintained in high-glucose medium. MES (12 V)+mild HS treatment of high fat-fed mice also increased the phosphorylation of insulin receptor β subunit (IRβ) and Akt in mice liver. In high fat-fed mice and db/db mice, MES+HS treatment for 10 min applied twice a week for 12–15 weeks significantly decreased fasting blood glucose and insulin levels and improved insulin sensitivity. The treated mice showed significantly lower weight of visceral and subcutaneous fat, a markedly improved fatty liver and decreased size of adipocytes. Our findings indicated that the combination of MES and HS alleviated insulin resistance and improved fat metabolism in diabetes mouse models, in part, by enhancing the insulin signaling pathway.

No abstract

Interleukin 15 (IL-15) and IL-2, which promote the survival of memory CD8(+) T cells and regulatory T cells, respectively, bind receptor complexes that share beta- and gamma-signaling subunits. Receptor specificity is provided by unique, nonsignaling alpha-subunits. Whereas IL-2 receptor-alpha (IL-2Ralpha) is expressed together in cis with the beta- and gamma-subunits on T cells and B cells, IL-15Ralpha is expressed in trans on antigen-presenting cells. Here we present a 1.85-A crystal structure of the human IL-15-IL-15Ralpha complex. The structure provides insight into the molecular basis of the specificity of cytokine recognition and emphasizes the importance of water in generating this very high-affinity complex. Despite very low IL-15-IL-2 sequence homology and distinct receptor architecture, the topologies of the IL-15-IL-15Ralpha and IL-2-IL-2Ralpha complexes are very similar. Our data raise the possibility that IL-2, like IL-15, might be capable of being presented in trans in the context of its unique receptor alpha-chain.

Amyloid fibril formation is associated with protein misfolding disorders, including neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's diseases. Familial amyloid polyneuropathy (FAP) is a hereditary disease caused by a point mutation of the human plasma protein, transthyretin (TTR), which binds and transports thyroxine (T(4)). TTR variants contribute to the pathogenesis of amyloidosis by forming amyloid fibrils in the extracellular environment. A recent report showed that epigallocatechin 3-gallate (EGCG), the major polyphenol component of green tea, binds to TTR and suppresses TTR amyloid fibril formation. However, structural analysis of EGCG binding to TTR has not yet been conducted. Here we first investigated the crystal structure of the EGCG-V30M TTR complex and found novel binding sites distinct from the thyroxine binding site, suggesting that EGCG has a mode of action different from those of previous chemical compounds that were shown to bind and stabilize the TTR tetramer structure. Furthermore, EGCG induced the oligomerization and monomer suppression in the cellular system of clinically reported TTR variants. Taken together, these findings suggest the possibility that EGCG may be a candidate compound for FAP therapy.

Transforming growth factor-␤ (TGF-␤

The most common cystic fibrosis transmembrane conductance regulator (CFTR) mutant in cystic fibrosis patients, ⌬F508 CFTR, is retained in the endoplasmic reticulum (ER) and is consequently degraded by the ubiquitin-proteasome pathway known as ER-associated degradation (ERAD). Because the prolonged interaction of ⌬F508 CFTR with calnexin, an ER chaperone, results in the ERAD of ⌬F508 CFTR, calnexin seems to lead it to the ERAD pathway. However, the role of calnexin in the ERAD is controversial. In this study, we found that calnexin overexpression partially attenuated the ERAD of ⌬F508 CFTR. We observed the formation of concentric membranous bodies in the ER upon calnexin overexpression and that the ⌬F508 CFTR but not the wild-type CFTR was retained in the concentric membranous bodies. Furthermore, we observed that calnexin overexpression moderately inhibited the formation of aggresomes accumulating the ubiquitinated ⌬F508 CFTR. These findings suggest that the overexpression of calnexin may be able to create a pool of ⌬F508 CFTR in the ER. INTRODUCTIONCystic fibrosis transmembrane conductance regulator (CFTR) is a polytopic integral membrane protein that functions as a cAMP-dependent Cl Ϫ channel in the apical membrane of epithelial cells (Anderson et al., 1991;Drumm et al., 1991;Tabcharani et al., 1991;Bear et al., 1992). Mutations in the CFTR gene lead to the absence or malfunction of a regulated Cl Ϫ channel in the apical membrane of secretory epithelia, resulting in the clinical symptoms of cystic fibrosis (CF) (Kerem et al., 1989;Riordan et al., 1989;Rommens et al., 1989;Cheng et al., 1990). Therefore, potential CF therapies are aimed at overcoming the functional impairment of various mutant CFTRs, particularly ⌬F508 CFTR, in which a phenylalanine at position 508 is deleted from the first nucleotide-binding fold. This mutation is found in ϳ70% of CF chromosomes and results in a severe form of the disease; Ͼ90% of CF patients have at least one ⌬F508 allele (Tsui, 1992;Sferra and Collins, 1993). Although ⌬F508 CFTR is functionally competent as a cAMP-dependent Cl Ϫ channel in model situations where it reaches the plasma membrane (Drumm et al., 1991;Denning et al., 1992;Li et al., 1993), in mammalian cells ⌬F508 CFTR fails to reach the plasma membrane.CFTR biogenesis is inefficient. During insertion into the ER membrane, CFTR interacts with the cytosolic chaperones Hsc70/Hdj-2 and Hsp90, as well as the ER chaperone calnexin (CNX) (Yang et al., 1993;Pind et al., 1994;Loo et al., 1998;Meacham et al., 1999). After ATP-dependent conformational maturation, assisted by cytosolic and ER chaperones, the chaperones release CFTR, and only ϳ30% of the immature wild-type (wt) CFTR transits to the late secretory pathway, ultimately reaching the plasma membrane (Lukacs et al., 1994). However, most of the immature wt CFTR (ϳ70%), and nearly all of immature ⌬F508 CFTR, fail to mature and do not transit to the late secretory pathway (Lukacs et al., 1994). They are trapped in the ER as core-glycosylated products (ϳ140 kDa) and ar...