Mechanisms of tissue factor induction by Porphyromonas gingivalis in human endothelial cells

Associations between periodontal infection and cardiovascular disease have been documented. Porphyromonas gingivalis is a well-established periodontal pathogen, and tissue factor (TF) is a key initiator of the coagulation cascade. In this context, P. gingivalis has been reported to enhance TF expression in human endothelial cells. The present study investigated the underlying mechanisms of TF induction by P. gingivalis in human umbilical vein endothelial cells. P. gingivalis increased TF expression in a dose- and time-dependent manner. Not only live bacteria but also glutaraldehyde-fixed bacteria increased TF expression to the same extent. However, sonicates of P. gingivalis did not induce TF expression. Cytochalasin D and SMIFH2, which are inhibitors of actin polymerization and actin nucleation, respectively, inhibited the TF expression induced by P. gingivalis. Finally, TF production was decreased or increased in the presence of various signaling inhibitors, including mitogen-activated protein kinases. These results suggest that P. gingivalis induces endothelial TF expression by a bacterial internalization-dependent mechanism and through diverse signal transduction mechanisms.

Mechanisms of tissue factor induction by Porphyromonas gingivalis in human endothelial cells

Associations between periodontal infection and cardiovascular disease have been documented. Porphyromonas gingivalis is a well-established periodontal pathogen, and tissue factor (TF) is a key initiator of the coagulation cascade. In this context, P. gingivalis has been reported to enhance TF expression in human endothelial cells. The present study investigated the underlying mechanisms of TF induction by P. gingivalis in human umbilical vein endothelial cells. P. gingivalis increased TF expression in a dose- and time-dependent manner. Not only live bacteria but also glutaraldehyde-fixed bacteria increased TF expression to the same extent. However, sonicates of P. gingivalis did not induce TF expression. Cytochalasin D and SMIFH2, which are inhibitors of actin polymerization and actin nucleation, respectively, inhibited the TF expression induced by P. gingivalis. Finally, TF production was decreased or increased in the presence of various signaling inhibitors, including mitogen-activated protein kinases. These results suggest that P. gingivalis induces endothelial TF expression by a bacterial internalization-dependent mechanism and through diverse signal transduction mechanisms.

Green tea polyphenol (–)-epigallocatechin-3-gallate prevents ultraviolet-induced apoptosis in PC12 cells

Green tea polyphenol (–)-epigallocatechin-3-gallate (EGCG) is a potent antioxidant with protective effects against neurotoxicity. However, it is currently unclear whether EGCG protects neuronal cells against radiation-induced damage. Therefore, the objective of this study was to investigate the effects of EGCG on ultraviolet (UV)-induced oxidative stress and apoptosis in PC12 cells. The effects of UV irradiation included apoptotic cell death, which was associated with DNA fragmentation, reactive oxygen species (ROS) production, enhanced caspase-3 and caspase-9 activity, and poly (ADP-ribose) polymerase cleavage. UV irradiation also increased the Bax/Bcl-2 ratio and mitochondrial pathway-associated cytochrome c expression. However, pretreatment with EGCG before UV exposure markedly decreased UV-induced DNA fragmentation and ROS production. Furthermore, the UV irradiationinduced increase in Bax/Bcl-2 ratio, cytochrome c upregulation, and caspase-3 and caspase-9 activation were each ameliorated by EGCG pretreatment. Additionally, EGCG suppressed UV-induced phosphorylation of p38 and rescued UV-downregulated phosphorylation of ERK. Taken together, these results suggest that EGCG prevents UV irradiationinduced apoptosis in PC12 cells by scavenging ROS and inhibiting the mitochondrial pathways known to play a crucial role in apoptosis. In addition, EGCG inhibits UV-induced apoptosis via JNK inactivation and ERK activation in PC12 cells. Thus, EGCG represents a potential neuroprotective agent that could be applied to prevent neuronal cell death induced by UV irradiation.

Green tea polyphenol (–)-epigallocatechin-3-gallate prevents ultraviolet-induced apoptosis in PC12 cells

Green tea polyphenol (–)-epigallocatechin-3-gallate (EGCG) is a potent antioxidant with protective effects against neurotoxicity. However, it is currently unclear whether EGCG protects neuronal cells against radiation-induced damage. Therefore, the objective of this study was to investigate the effects of EGCG on ultraviolet (UV)-induced oxidative stress and apoptosis in PC12 cells. The effects of UV irradiation included apoptotic cell death, which was associated with DNA fragmentation, reactive oxygen species (ROS) production, enhanced caspase-3 and caspase-9 activity, and poly (ADP-ribose) polymerase cleavage. UV irradiation also increased the Bax/Bcl-2 ratio and mitochondrial pathway-associated cytochrome c expression. However, pretreatment with EGCG before UV exposure markedly decreased UV-induced DNA fragmentation and ROS production. Furthermore, the UV irradiationinduced increase in Bax/Bcl-2 ratio, cytochrome c upregulation, and caspase-3 and caspase-9 activation were each ameliorated by EGCG pretreatment. Additionally, EGCG suppressed UV-induced phosphorylation of p38 and rescued UV-downregulated phosphorylation of ERK. Taken together, these results suggest that EGCG prevents UV irradiationinduced apoptosis in PC12 cells by scavenging ROS and inhibiting the mitochondrial pathways known to play a crucial role in apoptosis. In addition, EGCG inhibits UV-induced apoptosis via JNK inactivation and ERK activation in PC12 cells. Thus, EGCG represents a potential neuroprotective agent that could be applied to prevent neuronal cell death induced by UV irradiation.

Comparison of inflammatory cytokine-inducing activity of lipopolysaccharides from major periodontal bacteria

Porphyromonas gingivalis (Pg), Aggregatibacter actinomycetemcomitans (Aa), Tannerella forsythia (Tf), Prevotella intermedia (Pi), and Fusobacterium nucleatum (Fn) are major periodontal pathogens. Lipopolysaccharides (LPSs) from periodontal bacteria play an important role in periodontal pathogenesis by stimulating host cells to produce inflammatory cytokines. In this study, highly pure LPSs from the five major periodontopathogens were prepared, and their monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α)-inducing activities were compared in human umbilical vein endothelial cells (HUVECs) and THP-1 macrophagic cells, respectively. In HUVECs, LPSs from Aa and Fn were potent stimulators for MCP-1 induction; however, LPSs from Pg, Pi, and Tf were much weaker MCP-1 inducers. In THP-1 cells, LPSs from Pg, Aa, and Fn were relatively strong inducers of TNF-α, whereas LPSs from Pi and Tf produced little activity. The Toll-like receptor (TLR)2/TLR4 dependency of various LPSs was also determined by measuring NF-κB reporter activity in TLR2- or TLR4-expressing 293 cells. LPSs from Aa, Fn, and Tf stimulated only TLR4; however, LPSs from Pg and Pi stimulated both TLR2 and TLR4. These results suggest that LPSs from major periodontal bacteria differ considerably in their cell-stimulating activity.

Induction of Prostaglandin E₂ by Porphyromonas gingivalis in Human Dental Pulp Cells

Cyclooxygenase-2 (COX-2)-mediated prostaglandin E₂ (PGE₂) plays a key role in development and progression of inflammatory responses and Porphyromonas gingivalis is a common endodontic pathogen. In this study, we investigated induction of COX-2 and PGE₂ by P. gingivalis in human dental pulp cells (HDPCs). P. gingivalis increased expression of COX-2, but not that of COX-1. Increased levels of PGE₂ were released from P. gingivalis-infected HDPCs and this PGE₂ increase was blocked by celecoxib, a selective COX-2 inhibitor. P. gingivalis activated all three types of mitogen-activated protein kinases (MAPKs). P. gingivalis-induced activation of nuclear factor-κB (NF-κB) was demonstrated by the results of phosphorylation of NF-κ B p65 and degradation of inhibitor of κB-α (IκB-α). Pharmacological inhibition of each of the three types of MAPKs and NF-κB substantially attenuated P. gingivalis induced PGE2 production. These results suggest that P. gingivalis should promote endodontic inflammation by stimulating dental pulp cells to produce PGE₂.

Inhibition of Lipopolysaccharide-stimulated Inflammatory Cytokine Production by LY303511 in Human Macrophagic THP-1 Cells

We have previously shown that the specific phosphatidylinositol 3-kinase inhibitor LY294002 (LY29), and its inactive analog LY303511 (LY30), inhibit a monocyte chemoattractant protein-1 (MCP-1) expression in human umbilical vein endothelial cells; these results suggest the potential of LY30 as an anti-inflammatory drug. In this study, we determined the effects of LY30 on the production of various inflammatory cytokines in human macrophagic THP-1 cells which were stimulated with lipopolysaccharide (LPS). LY30 selectively suppressed the mRNA expression of IL-12 p40, TNF-α, and MCP-1 without affecting the expression of IL-1α, IL-6, and IL-8. Inhibition of the production of IL-12 and TNF-α by LY30 was also demonstrated using ELISA assays. In order to elucidate the mechanisms of the action of LY30, we examined the role played by the mitogen-activated protein kinases and the key transcription factors, AP-1 and NF-κB in LPS-stimulated THP-1 cells. The results revealed that LY30 inhibited LPS-induced activation of ERK, but not p38 or JNK. Furthermore, the AP-1 DNA binding activity was suppressed by LY30 based upon the dosage, whereas NF-κB DNA binding was not affected. These results suggest that LY30 selectively inhibits cytokine production in the LPS-stimulated macrophagic THP-1 cells by downregulating the activation of ERK and AP-1.

Evaluation of antibacterial activity and osteoblast-like cell viability of TiN, ZrN and (Ti(1-x)Zr(x))N coating on titanium

PURPOSE: The aim of this study was to evaluate antibacterial activity and osteoblast-like cell viability according to the ratio of titanium nitride and zirconium nitride coating on commercially pure titanium using an arc ion plating system. MATERIALS AND METHODS: Polished titanium surfaces were used as controls. Surface topography was observed by scanning electron microscopy, and surface roughness was measured using a two-dimensional contact stylus profilometer. Antibacterial activity was evaluated against Streptococcus mutans and Porphyromonas gingivalis with the colony-forming unit assay. Cell compatibility, mRNA expression, and morphology related to human osteoblast-like cells (MG-63) on the coated specimens were determined by the XTT assay and reverse transcriptase-polymerase chain reaction. RESULTS: The number of S. mutans colonies on the TiN, ZrN and (Ti(1-x)Zr(x))N coated surface decreased significantly compared to those on the non-coated titanium surface (P<0.05). CONCLUSION: The number of P. gingivalis colonies on all surfaces showed no significant differences. TiN, ZrN and (Ti(1-x)Zr(x))N coated titanium showed antibacterial activity against S. mutans related to initial biofilm formation but not P. gingivalis associated with advanced periimplantitis, and did not influence osteoblast-like cell viability.

Evaluation of antibacterial activity and osteoblast-like cell viability of TiN, ZrN and (Ti(1-x)Zr(x))N coating on titanium

PURPOSE: The aim of this study was to evaluate antibacterial activity and osteoblast-like cell viability according to the ratio of titanium nitride and zirconium nitride coating on commercially pure titanium using an arc ion plating system. MATERIALS AND METHODS: Polished titanium surfaces were used as controls. Surface topography was observed by scanning electron microscopy, and surface roughness was measured using a two-dimensional contact stylus profilometer. Antibacterial activity was evaluated against Streptococcus mutans and Porphyromonas gingivalis with the colony-forming unit assay. Cell compatibility, mRNA expression, and morphology related to human osteoblast-like cells (MG-63) on the coated specimens were determined by the XTT assay and reverse transcriptase-polymerase chain reaction. RESULTS: The number of S. mutans colonies on the TiN, ZrN and (Ti(1-x)Zr(x))N coated surface decreased significantly compared to those on the non-coated titanium surface (P<0.05). CONCLUSION: The number of P. gingivalis colonies on all surfaces showed no significant differences. TiN, ZrN and (Ti(1-x)Zr(x))N coated titanium showed antibacterial activity against S. mutans related to initial biofilm formation but not P. gingivalis associated with advanced periimplantitis, and did not influence osteoblast-like cell viability.

Clathrin and Lipid Raft-dependent Internalization of Porphyromonas gingivalis in Endothelial Cells

Porphyromonas gingivalis is one of the most important periodontal pathogens and has been to known to invade various types of cells, including endothelial cells. The present study investigated the mechanisms involved in the internalization of P. gingivalis in human umbilical vein endothelial cells (HUVEC). P. gingivalis internalization was reduced by clathrin and lipid raft inhibitors, as well as a siRNA knockdown of caveolin-1, a principal molecule of lipid raft-related caveolae. The internalization was also reduced by perturbation of actin rearrangement, while microtubule polymerization was not required. Furthermore, we found that Src kinases are critical for the internalization of P. gingivalis into HUVEC, while neither Rho family GTPases nor phosphatidylinositol 3-kinase are required. Taken together, this study indicated that P. gingivalis internalization into endothelial cells involves clathrin and lipid rafts and requires actin rearrangement associated with Src kinase activation.

Aggregatibacter actinomycetemcomitans Strongly Stimulates Endothelial Cells to Produce Monocyte Chemoattractant Protein-1 and Interleukin-8

Aggregatibacter actinomycetemcomitans is the most important etiologic agent of aggressive periodontitis and can interact with endothelial cells. Monocyte chemoattractant protein-1 (MCP-1) and interleukin-8 (IL-8) are chemokines, playing important roles in periodontal pathogenesis. In our current study, the effects of A. actinomycetemcomitans on the production of MCP-1 and IL-8 by human umbilical vein endothelial cells (HUVEC) were investigated. A. actinomycetemcomitans strongly induced the gene expression and protein release of both MCP-1 and IL-8 in a dose- and time-dependent manner. Dead A. actinomycetemcomitans cells were as effective as live bacteria in this induction. Treatment of HUVEC with cytochalasin D, an inhibitor of endocytosis, did not affect the mRNA up-regulation of MCP-1 and IL-8 by A. actinomycetemcomitans. However, genistein, an inhibitor of protein tyrosine kinases, substantially inhibited the MCP-1 and IL-8 production by A. actinomycetemcomitans, whereas pharmacological inhibition of each of three members of mitogen-activated protein (MAP) kinase family had little effect. Furthermore, gel shift assays showed that A. actinomycetemcomitans induces a biphasic activation (early at 1-2 h and late at 8-16 h) of nuclear factor-kappaB (NF-kappaB) and an early brief activation (0.5-2 h) of activator protein-1 (AP-1). Activation of canonical NF-kappaB pathway (IkappaB kinase activation and IkappaB-alpha degradation) was also demonstrated in these experiments. Although lipopolysaccharide from A. actinomycetemcomitans also induced NF-kappaB activation, this activation profile over time differed from that of live A. actinomycetemcomitans. These results suggest that the expression of MCP-1 and IL-8 is potently increased by A. actinomycetemcomitans in endothelial cells, and that the viability of A. actinomycetemcomitans and bacterial internalization are not required for this effect, whereas the activation of protein tyrosine kinase(s), NF-kappaB, and AP-1 appears to play important roles. The secretion of high levels of MCP-1 and IL-8 resulting from interactions of A. actinomycetemcomitans with endothelial cells may thus contribute to the pathogenesis of aggressive periodontitis.

Mechanisms Underlying Enterococcus faecalis-Induced Tumor Necrosis Factor-alpha Production in Macrophages

Enterococcus faecalis, a gram-positive bacterium, has been implicated in endodontic infections, particularly in chronic apical periodontitis. Proinflammatory cytokines, including tumor necrosis factor-alpha (TNF-alpha), are involved in the pathogenesis of these apical lesions. E. faecalis has been reported to stimulate macrophages to produce TNF-alpha. The present study investigated the mechanisms involved in TNF-alpha production by a murine macrophage cell line, RAW 264.7 in response to exposure to E. faecalis. Both live and heat-killed E. faecalis induced high levels of gene expression and protein release of TNF-alpha. Treatment of RAW 264.7 cells with cytochalasin D, an inhibitor of endocytosis, prevented the mRNA up-regulation of TNF-alpha by E. faecalis. In addition, antioxidant treatment reduced TNF-alpha production to baseline levels. Inhibition of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein (MAP) kinase also significantly attenuated E. faecalis-induced TNF-alpha expression by RAW 264.7 cells. Furthermore, activation of NF-kappaB and AP-1 in RAW 264.7 cells was also stimulated by E. faecalis. These results suggest that the phagocytic uptake of bacteria is necessary for the induction of TNF-alpha in E. faecalis-stimulated macrophages, and that the underlying intracellular signaling pathways involve reactive oxygen species, ERK, p38 MAP kinase, NF-kappaB, and AP-1.

Mechanisms Underlying Enterococcus faecalis-Induced Tumor Necrosis Factor-alpha Production in Macrophages

Enterococcus faecalis, a gram-positive bacterium, has been implicated in endodontic infections, particularly in chronic apical periodontitis. Proinflammatory cytokines, including tumor necrosis factor-alpha (TNF-alpha), are involved in the pathogenesis of these apical lesions. E. faecalis has been reported to stimulate macrophages to produce TNF-alpha. The present study investigated the mechanisms involved in TNF-alpha production by a murine macrophage cell line, RAW 264.7 in response to exposure to E. faecalis. Both live and heat-killed E. faecalis induced high levels of gene expression and protein release of TNF-alpha. Treatment of RAW 264.7 cells with cytochalasin D, an inhibitor of endocytosis, prevented the mRNA up-regulation of TNF-alpha by E. faecalis. In addition, antioxidant treatment reduced TNF-alpha production to baseline levels. Inhibition of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein (MAP) kinase also significantly attenuated E. faecalis-induced TNF-alpha expression by RAW 264.7 cells. Furthermore, activation of NF-kappaB and AP-1 in RAW 264.7 cells was also stimulated by E. faecalis. These results suggest that the phagocytic uptake of bacteria is necessary for the induction of TNF-alpha in E. faecalis-stimulated macrophages, and that the underlying intracellular signaling pathways involve reactive oxygen species, ERK, p38 MAP kinase, NF-kappaB, and AP-1.

Comparison of Antibacterial effect of Listerine(R) with Various root canal irrigants / 대한치과보존학회지

The purpose of this study is to compare the antibacterial effect of Listerine(R) on two microorganisms (P. gingivalis and E. faecalis) with various root canal irrigants (NaOCl, CHX, EDTA) and to identify possibility of using Listerine(R) as a root canal irrigant. Porphyromonas gingivalis ATCC 3327 and Enterococcus faecalis ATCC 29212 were used in this experiment. For the test irrigants, 0.5%, 1%, 2.5%, 5.25% NaOCl, 0.1%, 0.2%, 1%, 2% CHX, 0.5M EDTA (18.6% EDTA) and Listerine(R) were prepared. Distiled water was used as control. Two methods-1) Comparison of turbidity in broth and 2) Agar diffusion test-were used to determine the extent of antibacterial effect of Listerine(R) and to compare it with that of NaOCl, CHX, and EDTA. All solutions tested were effective against two bacterial strains compared with control (p<0.001). Any concentration of NaOCl, CHX, and EDTA showed similarly high effectiveness against all bacterial strains. In all experiment, Listerine(R) showed significantly low antibacterial effect compared with the other root canal irrigants (p<0.05). In conclusion, the results reflect remarkably low antibacterial effect of Listerine(R) as compared with root canal irrigants in general so it is not suitable for the root canal irrigant.

Effect of rotational speed of Protaper(TM) rotary file on the change of root canal configuration / 대한치과보존학회지

This study was conducted to evaluate canal configuration after shaping by ProTaper(TM) with various rotational speed in J-shaped simulated resin canals. Forty simulated root canals were divided into 4 groups, and instrumented using by ProTaper(TM) at the rotational speed of 250, 300, 350 and 400 rpm. Pre-instrumented and post-instrumented images were taken by a scanner and those were superimposed. Outer canal width, inner canal width, total canal width, and amount of transportation from original axis were measured at 1, 2, 3, 4, 5, 6, 7 and 8 mm from apex. Instrumentation time, instrument deformation and fracture were recorded. Data were analyzed by means of one-way ANOVA followed by Scheffe's test. The results were as follows 1. Regardless of rotational speed, at the 1~2 mm from the apex, axis of canal was transported to outer side of a curvature, and at 3~6 mm from the apex, to inner side of a curvature. Amounts of transportation from original axis were not significantly different among experimental groups except at 5 and 6 mm from the apex. 2. Instrumentation time of 350 and 400 rpm was significantly less than that of 250 and 300 rpm (p < 0.01). In conclusion, the rotational speed of ProTaper(TM) files in the range of 250~400 rpm does not affect the change of canal configuration, and high rotational speed reduces the instrumentation time. However, appearance of separation and distortion of Ni-Ti rotary files can occur in high rotational speed.

A comparative study of the canal configuration after shaping by protaper rotary and hand files in resin simulated canals / 대한치과보존학회지

The purpose of this study was to compare the canal configuration after shaping by ProTaper rotary files and ProTaper hand files in resin simulated canals. Forty resin simulated canals with a curvature of J-shape and S-shape were divided into four groups by 10 blocks each. Simulated root canals in resin block were prepared by ProTaper rotary files and ProTaper hand files using a crown-down pressureless technique. All simulated canals were prepared up to size #25 file at end-point of preparation. Pre- and post-instrumentation images were recorded with color scanner. Assessment of canal shape was completed with an image analysis program. Measurements were made at 0, 1, 2, 3, 4, 5, 6 and 7 mm from the apex. At each level, outer canal width, inner canal width, total canal width, and amount of transportation from original axis were recorded. Instrumentation time was recorded. The data were analyzed statistically using independent t-test. The result was that ProTaper hand files cause significantly less canal transportation from original axis of canal body and maintain original canal configuration better than ProTaper rotary files, however ProTaper hand files take more shaping time.

Increased Growth of Human Leukemic HL-60 Cells by Dimethyl Sulfoxide and Dimethylformamide / 대한면역학회지

A recent study has proposed that dimethyl sulfoxide (DMSO) increases the number of HL-60 cells in high cell density conditions by inhibiting cell density-dependent apoptosis. We observed that dimethylformamide (DMF), a DMSO-related polar compound, also increased the concentration of HL-60 cells. The effective dose range of DMSO and DMF was 0.5-1% and 0.2-0.6% respectively. DMF, like DMSO, inhibited density-dependent apoptosis of HL-60 cells. The flow cytometric PKH26 cell proliferation assay showed DMSO and DMF actively increased cell division. However, the difference in the distribution of cell cycle phase was not noted between the control and the DMSO- or DMF-treated HL-60 cultures. Finally, DMSO and DMF stimulated HL-60 growth even in low density conditions. These results suggest that DMSO and OMF at appropriate concentrations increase the number of HL-60 cells by both apoptosis inhibition and cell division augmentation.

The effects of oxygen radicals on the activity of nitric oxide synthase and guanylate cyclase

Reactive oxygen species such as superoxides, hydrogen peroxide (H2O2) and hydroxyl radicals have been suggested to be involved in the catalytic action of nitric oxide synthase (NOS) to produce NO from L-arginine. An examination was conducted on the effects of oxygen radical scavengers and oxygen radical-generating systems on the activity of neuronal NOS and guanylate cyclase (GC) in rat brains and NOS from the activated murine macrophage cell line J774. Catalase and superoxide dismutase (SOD) showed no significant effects on NOS or GC activity. Nitroblue tetrazolium (NBT, known as a superoxide radical scavenger) and peroxidase (POD) inhibited NOS, but their inhibitory actions were removed by increasing the concentration of arginine or NADPH respectively, in the reaction mixture. NOS and NO-dependent GC were inactivated by ascorbate/FeSO4 (a metal-catalyzed oxidation system), 2'2'-azobis-amidinopropane (a peroxy radical producer), and xanthine/xanthine oxidase (a superoxide generating system). The effects of oxygen radicals or antioxidants on the two isoforms of NOS were almost similar. However, H2O2 activated GC in a dose-dependent manner from 100 microM to 1 mM without significant effects on NOS. H2O2-induced GC activation was blocked by catalase. These results suggested that oxygen radicals inhibited NOS and GC, but H2O2 could activate GC directly.

Effect of Ionizing Radiation on the Host Resistance Against Listeria Monocytogenes Infection and the Cytokine Production in Mice / 대한치료방사선과학회지

PURPOSE: To evaluate the qualitative immunologic changes by ionizing radiation, we studied the altered capacities of the macrophages and lymphocytes to produce cytokines in conjunction with resistance to Listeria monocytogenes (LM) infection in mice. MATERIALS AND METHODS: BALB/c mice and Listeria monocytogenes were used. The mice were infected intraperitoneally with 105LM at 1 day after irradiation (300cGy) and sacrificed at 1, 3, 5 days after infection, and then the numbers of viable LM per spleen in the irradiated and control group were counted. Tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and nitric oxide (NO) were assessed after irradiation. RESULTS: Under gamma-ray irradiation with a dose range of 100-850cGy, the number of total splenocytes decreased markedly in a dose-dependent manner, while peritoneal macrophages did so slightly. Cultured peritoneal macrophages produced more TNF-alpha in the presence of lipopolysaccharide (LPS) during the 24 hours after in vitro irradiation, but their capacity of TNF-alpha production showed a decreased tendency at 5 days after in vivo total body irradiation. With 100cGy and 300cGy irradiation, cultured peritoneal macrophages produced more NO in the presence of LPS during the 24 hours after in vitro irradiation than without irradiation. Activated splenocytes from irradiated mice (300cGy) exhibited a decreased capacity to produce IL-2 and IFN-gamma with Concavalin-A stimulation at 3 days after irradiation. When BALB/c mice were irradiated to the total body with a dose of 300cGy, they showed enhanced resistance during early innate phase, but a significant inhibition of resistance to LM was found in the late innate and acquired T-cell dependent phases. CONCLUSION: These results suggest that increased early innate and decreased late innate and acquired immunity to LM infection by ionizing radiation (300cGy) may be related to the biphasic altered capacity of the macrophages to produce TNF-alpha and the decreased capacities of the lymphocytes to produce IL-2 and IFN-gamma in addition to a marked decrease in the total number of cells.

Effect of Ultraviolet B Irradiation on the TNF-alpha /IFN-gamma Production and Immunity to Listeria monocytogenes Infection in Mice

The ultraviolet radiation (UVR) is known to be a potent modulator of many host immune functions and the exposure of experimental animals to the inflammatory effects of UVR induces depressions in their ability to initiate and effectuate various types of cellular immune responses. In this study, the effects of UV-B (280 320 nm) radiation on resistance to a facultative intracellular bacterium, Listeria monocytogenes (LM), were examined at the cellular level. The numbers of cultivable LM recovered from the spleens of UV-B-irradiated mice were decreased at 2 days postinfection compared with those of untreated control mice. However, the acquired immunity, developed 7 days after immunization with streptomycin (SM)-sensitive LM, in either UV-irradiated, LPS- or IL-1-pretreated mice was less stronger than that developed in untreated, control mice. To elucidate the possible mechanisms underlying the observation that UVR did increase innate immunity but decreased acquired immunity of mice to the infection with LM, the effects of UVR of mice on the production of IFN-r by activated splenocytes and TNF-a by peritoneal macrophages were assessed. Activated splenocytes from UV-irradiated mice exhibited a reduced capacity to produce IFN-r and cultured peritoneal macrophages produced more TNF-a in the presence of LPS during 24 hours after UV radiation. Though TNF-r activity was not detected in the sera of LM-infected mice, intravenous LPS injection induced TNF-r production and UVR decreased TNF activity in sera obtained from LM-infected mice with LPS induction 9 days after irradiation. Although Ia-negative macrophages were predominant in the peritoneal macrophages from untreated control mice, the infection of mice with LM caused a marked increase in Ia expression on peritoneal macrophages. However, UVR resulted in decreased expression of Ia molecule on the peritoneal macrophages during the LM infection. These findings suggest that the dual effects of UVR on the innate and acquired immunity of mice to the LM infection may be associated with altered capacities of splenocytes and peritoneal macrophages of the mice to produce cytokines, in addition to decrease of la molecule expression on the macrophages.