Byung-Cheol Kang

Inhibition of interleukin-12 production in lipopolysaccharide-activated mouse macrophages by parthenolide, a predominant sesquiterpene lactone in Tanacetum parthenium: involvement of nuclear factor-κB

Pharmacological control of interleukin-12 (IL-12) production may be a key therapeutic strategy for modulating immunological diseases dominated by type-1 cytokine responses. In this study, we investigated the effects of parthenolide, an anti-inflammatory sesquiterpene, on the production of IL-12 from mouse macrophages stimulated with lipopolysaccharide (LPS). Parthenolide potently inhibited the LPS-induced IL-12 production in a dose-dependent manner. The effect of parthenolide on IL-12 p40 promoter activation was analyzed by transfecting RAW264.7 monocytic cells with p40 promoter/luciferase constructs. The repressive effect mapped to a region in the p40 promoter containing a binding site for nuclear factor-kappaB (p40-kappaB). Furthermore, activation of macrophages by LPS resulted in markedly enhanced binding activity to the kappaB site, which significantly decreased upon addition of parthenolide. These results suggest that parthenolide-induced inhibition of IL-12 production in macrophages may explain some of the biological effects of parthenolide including its anti-inflammatory activity.

Hydroquinone, a reactive metabolite of benzene, enhances interleukin-4 production in CD4+ T cells and increases immunoglobulin E levels in antigen-primed mice

Exposure to cigarette smoke is known to increase the risk of the development of allergic disease. The mechanism is not well understood. In this study, we determined the effect of hydroquinone (HQ), a major metabolite of benzene present in large quantities in cigarette tar, on interleukin‐4 (IL‐4) production by CD4+ T cells. HQ significantly enhanced IL‐4 production by keyhole limpet haemocyanin (KLH)‐primed CD4+ T cells in a dose‐dependent manner. The enhancing effect of HQ on IL‐4 production was maximal at a concentration of 50 µm. It increased the level of IL‐4 production approximately 10‐fold. HQ enhanced IL‐4 mRNA expression and also IL‐4 gene promoter activity, suggesting that the enhancing effect of HQ on IL‐4 production may occur at the transcriptional level. Furthermore, the injection of KLH‐primed mice with HQ resulted in a significant increase in the levels of IL‐4 and immunoglobulin E. These findings provide evidence that HQ, a major component of cigarette tar, may enhance allergic immune responses by inducing the production of IL‐4 in CD4+ T cells.

Inhibition of interleukin-12 production by auranofin, an anti-rheumatic gold compound, deviates CD4+ T cells from the Th1 to the Th2 pathway

Interleukin‐12 (IL‐12) may play a central role in the development and progression of rheumatoid arthritis by driving the immune response towards T helper 1 (Th1) type responses characterized by high IFN‐γ and low IL‐4 production. In this study we investigated the effect of auranofin (AF), an anti‐rheumatic gold compound, on IL‐12 production in mouse macrophages and dendritic cells, and studied whether AF‐mediated inhibition of IL‐12 production could regulate a cytokine profile of antigen (Ag)‐primed CD4+ Th cells. Treatment with AF significantly inhibited IL‐12 production in lipopolysaccharide (LPS)‐stimulated macrophages and also in CD40L‐stimulated dendritic cells. AF‐pretreated macrophages reduced their ability to induce IFN‐γ and increased the ability to induce IL‐4 in Ag‐primed CD4+ T cells. AF did not influence the cell surface expression of the class II MHC molecule and the costimulatory molecules CD80 and CD86. Addition of recombinant IL‐12 to cultures of AF‐pretreated macrophages and CD4+ T cells restored IFN‐γ production in Ag‐primed CD4+ T cells. The in vivo administration of AF resulted in the inhibition of IL‐12 production by macrophages stimulated in vitro with LPS or heat‐killed Listeria monocytogenes (HKL), leading to the inhibition of Th1 cytokine profile (decreased IFN‐γ and increased IL‐4 production) in Ag‐primed CD4+ T cells. These findings may explain some known effects of AF including anti‐rheumatic effects and the inhibition of encephalitogenicity, and point to a possible therapeutic use of AF in the Th1‐mediated immune diseases such as autoimmune diseases.

POTENTIATION OF ANTIGEN-SPECIFIC, TH1 IMMUNE RESPONSES BY MULTIPLE DNA VACCINATION WITH AN OVALBUMIN/INTERFERON-GAMMA HYBRID CONSTRUCT

The preferential differentiation of T helper (Th) cells to Th1 or Th2 subsets is important with respect to susceptibility or resistance to particular infections, or to autoimmune diseases and allergic diseases. To more effectively drive immune responses toward antigen‐specific Th1 responses, we constructed a mammalian expression vector (pOVA/IFN‐γ) carrying a hybrid gene in which the ovalbumin (OVA) (a model antigen) cDNA was covalently linked to murine interferon‐γ (IFN‐γ) cDNA. Intramuscular injection of BALB/c mice with the pOVA/IFN‐γ DNA increased both the production of OVA‐specific IFN‐γ by CD4+ T cells and the ratio of anti‐OVA immunoglobulin G (IgG)2a to IgG1 isotypes, while the injection with the pOVA alone, or with the mixture of the pOVA and pIFN‐γ, caused no or little increase. Furthermore, the OVA‐specific, Th1 immune responses were dramatically augmented by multiple injections with the pOVA/IFN‐γ DNA. These studies indicate that the direct linkage of an OVA gene to an IFN‐γ gene in the expression plasmid is required for efficiently confining the Th1 effects of IFN‐γ to the OVA‐specific cells, and the linkage effect of the OVA/IFN‐γ DNA can be potentiated by multiple vaccination.

Sulfasalazine prevents T-helper 1 immune response by suppressing interleukin-12 production in macrophages

Interleukin‐12 (IL‐12) plays a pivotal role in the development of T‐helper 1 (Th1) immune response, which may be involved in the pathogenesis of chronic inflammatory autoimmune disorders. In this study we investigated the effects of sulfasalazine, a drug for treating inflammatory bowel disease and rheumatoid arthritis, on the production of IL‐12 from mouse macrophages stimulated with lipopolysaccharide (LPS). Sulfasalazine potently inhibited the production of IL‐12 in a dose‐dependent manner, in part through the down‐regulation of nuclear factor κB (NFκB) activation in IL‐12 p40 gene. Activation of macrophages by LPS resulted in markedly enhanced binding activities to the κB site, which significantly decreased upon addition of sulfasalazine as demonstrated by an electrophoretic gel shift assay. Importantly, macrophages pretreated with sulfasalazine either in vitro or in vivo reduced their ability to induce interferon‐γ (IFN‐γ) and increased the ability to induce IL‐4 in antigen‐primed CD4+ T cells. From these results, sulfasalazine may induce the Th2 cytokine profile in CD4+ T cells by suppressing IL‐12 production in macrophages, and sulfasalazine‐induced inhibition of IL‐12 production in macrophages may explain some of the known biological effects of sulfasalazine.

Regulation of M2-type pyruvate kinase mediated by the high-affinity IgE receptors is required for mast cell degranulation

M2‐type pyruvate kinase (M2PK) was found to interact directly with the ‘ITAM’ region of the γ chain of the high‐affinity IgE receptor (FcɛRI). Our hypothesis was that mast cell degranulation might require the FcɛRI‐mediated inhibition of M2PK activity.

Chloromethyl ketones inhibit interleukin-12 production in mouse macrophages stimulated with lipopolysaccharide.

Interleukin-12 (IL-12) plays a pivotal role in the development of T-helper type 1 (Th1) immune response, which may be involved in the pathogenesis of chronic inflammatory autoimmune disorders. In this study, we investigated the effects of N-alpha-tosyl-L-phenylalanine chloromethyl ketone (TPCK) and N-alpha-tosyl-L-lysine chloromethyl ketone (TLCK), serine protease inhibitors, on the production of IL-12 from macrophages stimulated with lipopolysaccharide (LPS). TPCK and TLCK potently inhibited this LPS-induced IL-12 production in a dose-dependent manner. The effect of TPCK and TLCK on the IL-12 p40 promoter activation was analyzed by transfecting monocytic RAW264.7 cells with p40 promoter-reporter constructs. The repressive effect maps to a region in the p40 promoter containing a binding site for NFkappaB (p40-kappaB). A linker scan mutant of the p40-kappaB site abrogates the inhibitory effect on the p40 promoter, confirming the functional relevance of the NFkappaB site. Our results show that TPCK and TLCK inhibit NFkappaB-mediated IL-12 production in macrophages. reserved.

Interleukin-12-secreting fibroblasts are more efficient than free recombinant interleukin-12 in inducing the persistent resistance to Mycobacterium avium complex infection.

To determine whether the paracrine secretion of interleukin‐12 (IL‐12) can efficiently stimulate the resistance to Mycobacterium avium complex (MAC) infection, 3T3 fibroblasts were stably transfected to secrete IL‐12 (480 U/106 cells/48 hr) and their effect on MAC infection was investigated in genetically susceptible BALB/c mice, compared with that of free recombinant IL‐12 (rIL‐12). Injection with IL‐12‐secreting fibroblasts (3T3‐IL‐12) during intranasal infection with MAC resulted in a significant decrease in the bacterial load of the lung during the entire 10‐week observation period, while rIL‐12 reduced the bacterial load initially, at 2 weeks, but not by 10 weeks postinfection. Lung CD4+ T cells in mice injected with the 3T3‐IL‐12 cells showed a persistent T helper type 1 (Th1) response throughout the 10‐week period. Furthermore, immunization with the 3T3‐IL‐12 cells induced and maintained significantly higher levels of cytotoxic activity and nitric oxide production by lung cells than did rIL‐12 immunization. This work suggests that IL‐12‐secreting fibroblasts may serve as a vehicle for paracrine secretion of IL‐12 for immunotherapy of MAC infection.

Observer differentiation of mechanical defects versus natural dental caries cavitations on monitor-displayed images with imaging plate readout.

OBJECTIVE The purpose of this study was to compare the ability of dentists to detect mechanically created defects vs natural dental caries cavitations on the proximal surfaces of extracted teeth by means of storage phosphor imaging plate technology. STUDY DESIGN Fifty-two extracted molar and premolar teeth were blocked into sets for bitewing radiographs through use of the DIGORA digital imaging system. Sixteen natural caries cavities and 28 artificial lesions were present in the 80 proximal surfaces included in the study. A group of 16 dentists assessed proximal lesions on unenhanced images on the monitor and 1 month later on contrast-enhanced images. A different group of 16 dentists assessed proximal lesions on contrast-enhanced images and 1 month later on unenhanced images. The Mann-Whitney U test was used to check for a reading-order effect. The Zelen test of odds ratio was used to test for homogeneity, and the Mantel-Haenszel analysis or stratified logistic regression was used for inference about the common odds ratio. Alpha was set at P < .05. RESULTS AND CONCLUSIONS With the DIGORA system, there was little difference between the detection rates of mechanical defects and natural carious cavities with unenhanced images, but the mechanical defects were more readily detected when contrast-enhanced images were used. Cavity depth positively affected the odds of diagnosis of lesions, with deeper lesions being more readily detected than more superficial ones irrespective of whether they were natural or artificial. In comparison with findings of previous studies in which film and a charge-coupled device detector were used, the overall detection rate for natural dental caries was remarkably constant across the modalities.

Deviation of landmarks in accordance with methods of establishing reference planes in three-dimensional facial CT evaluation.

Purpose This study aimed to investigate the deviation of landmarks from horizontal or midsagittal reference planes according to the methods of establishing reference planes. Materials and Methods Computed tomography (CT) scans of 18 patients who received orthodontic and orthognathic surgical treatment were reviewed. Each CT scan was reconstructed by three methods for establishing three orthogonal reference planes (namely, the horizontal, midsagittal, and coronal reference planes). The horizontal (bilateral porions and bilateral orbitales) and midsagittal (crista galli, nasion, prechiasmatic point, opisthion, and anterior nasal spine) landmarks were identified on each CT scan. Vertical deviation of the horizontal landmarks and horizontal deviation of the midsagittal landmarks were measured. Results The porion and orbitale, which were not involved in establishing the horizontal reference plane, were found to deviate vertically from the horizontal reference plane in the three methods. The midsagittal landmarks, which were not used for the midsagittal reference plane, deviated horizontally from the midsagittal reference plane in the three methods. Conclusion In a three-dimensional facial analysis, the vertical and horizontal deviations of the landmarks from the horizontal and midsagittal reference planes could vary depending on the methods of establishing reference planes.