Daytime masticatory muscle electromyography biofeedback regulates the phasic component of sleep bruxism.

BACKGROUND: Electromyography (EMG) biofeedback (BF) training is potentially an effective cognitive-behavioural approach to regulate bruxism. OBJECTIVE: This study examined sleep bruxism regulation by daytime clenching control using a single-channel auditory EMG BF device. METHODS: Seventeen male subjects (mean age, 24.4 ± 3.1 years; mean ± SD) with self-reported awake/sleep bruxism were recruited and divided into a BF (n = 10) and a control (CO) group (n = 7). All subjects underwent four EMG recording sessions during both daytime and sleep over 3 weeks. During the daytime, in week 2, the BF group received feedback alert signals when excessive EMG activity with certain burst duration was detected while the subjects performed regular daily activities. The CO group underwent EMG recording sessions without receiving any alerts of parafunctional activity. The number of phasic burst events during sleep was compared between the BF and CO groups. RESULTS: While the number of phasic EMG events was not significantly different between the BF and CO groups at baseline, significantly smaller phasic events were observed in the BF compared to the CO group at the follow-up session (week 3) (P = .006, Tukey's HSD). Since daytime BF training is aimed at raising awareness of awake bruxism, it does not interrupt the sleep sequence or involve associated side effects. CONCLUSION: The present results suggest that EMG BF targeting for tonic EMG events during the daytime can be an effective method to regulate phasic EMG events during sleep.

Inhibition of Asparagine-linked Glycosylation Participates in Hypoxia-induced Down-regulation of Cell-surface MICA Expression.

BACKGROUND/AIM: Hypoxia down-regulates the expression of cell surface major histocompatibility class I-related chain molecule A (MICA) without increasing its shedding. Recently, the inhibition of N-linked glycosylation was also shown to reduce the cell-surface expression of MICA. We investigated the participation of asparagine (Asn)-linked glycosylation in hypoxia-induced down-regulation of cell-surface MICA using osteosarcoma cells. MATERIALS AND METHODS: The cell-surface expression and Asn-N-glycosylation of MICA were estimated by flow cytometry, and western blot analyses, respectively. RESULTS: Hypoxia reduced the expression of N-linked glycosylated MICA, as well as the ratio of N-linked glycosylated to non-glycosylated MICA. 2-Deoxy-D-glucose, which inhibits N-linked glycosylation, reduced the cell-surface expression of MICA under normoxia, while D-Mannose increased N-glycosylated MICA, increasing cell-surface MICA under hypoxia. Cells transfected with wild-type MICA expression vector expressed cell surface MICA more than those transfected with mutant MICA expression vectors designed for abrogation of N-linked glycosylation. CONCLUSION: The inhibition of Asn-N-linked glycosylation participates in hypoxia-induced down-regulation of cell-surface expression of MICA.

Sensitive detection of hemodynamic failure during orthostatic stress in patients with diabetic polyneuropathy using a mini laser Doppler blood flowmeter.

Autonomic dysfunction in diabetes is serious but often underestimated. The purpose of this study was to evaluate hemodynamics within the important initial phase just after standing, which cannot be evaluated by conventional instruments for orthostatic hypotension. Earlobe blood flow (EBF), which indirectly reflects the blood pressure response on standing, was evaluated using a mini laser Doppler flowmeter during standing from the sitting position in 58 healthy controls and 56 diabetic patients categorized as without (11), mild (27), and advanced diabetic polyneuropathy (18). The response area of the EBF waveform within 30 seconds after standing was calculated. An increased response area indicates poor recovery of EBF. Response area increased significantly with the degree of neuropathy (P < .001 for linear trend). Orthostatic hypotension was detected in two patients in the mild neuropathy group. The present approach may be sensitive and practical for detecting autonomic dysfunction not detected with the conventional orthostatic test.

Functional analysis of differences in transcriptional activity conferred by genetic variants in the 5' flanking region of the IL12RB2 gene.

Interleukin 12 receptor ß chain (IL12RB2) is a crucial regulatory factor involved in cell-mediated immune responses, and genetic variants of the gene encoding IL12RB2 are associated with susceptibility to various immune-related diseases. We previously demonstrated that haplotypes with single nucleotide polymorphisms (SNPs) in the 5' flanking region of IL12RB2, including -1035A>G (rs3762315) and -1023A>G (rs3762316), affect the expression of IL12RB2, thereby altering susceptibility to leprosy and periodontal diseases. In the present study, we identified transcription factors associated with the haplotype-specific transcriptional activity of IL12RB2 in T cells and NK cells. The -1023G polymorphism was found to create a consensus binding site for the transcription factor activating protein (AP)-1, and enzyme-linked immunosorbent assay (ELISA)-based binding assays showed that these SNPs enhanced AP-1 binding to this region. In reporter assays, suppression of JunB expression using siRNA eliminated differences in the -1035G/-1023G and -1035A/-1023A regions containing IL12RB2 promoter activity in Jurkat T cells and NK3.3 cells. These results suggested that the -1035/-1023 polymorphisms created differential binding affinities for JunB that could lead to differential IL12RB2 expression. Moreover, the -1035G and -1035A alleles formed binding sites for GATA-3 and myocyte enhancer factor-2 (MEF-2), respectively. Our data indicated that in addition to JunB, the SNP at -1035/-1023 influenced GATA-3 and MEF-2 binding affinity, potentially altering IL12RB2 transcriptional activity. These findings confirm the effects of rs3762315 and rs3762316 on IL12RB2 transcription. These genetic variants may alter cellular activation of T cells and NK cells and modify cell-mediated immune responses.

Orthostatic response of cephalic blood flow using a mini laser Doppler blood flowmeter and hemodynamics of a new active standing test.

PURPOSE: Cephalic hemodynamic assessment is important in initial orthostatic hypotension. We sought to investigate cephalic blood flow (CBF) in the earlobe using a mini laser Doppler flowmeter (LDF) during orthostatic challenge. In addition, we clarified hemodynamic differences during a new active standing protocol using a footstool standing test (FST) with bending of the legs on the footstool in the sitting position to reduce the load of the squatting posture in the conventional squat standing test (SST). METHODS: Ten healthy men (21 ± 0.5 years) performed the SST after a 1 min squat and the FST after a 1 min load consisting of bending the legs on a footstool in the sitting position. Earlobe CBF, beat-to-beat arterial blood pressure (ABP), mean arterial blood pressure (MAP), and heart rate (HR) were recorded during each test. RESULTS: Earlobe CBF showed a transient fall synchronized with the ABP during each test. No significant differences in the recovery times (RTs) of CBF and MAP were observed during the SST (CBF 12.9 ± 0.6 s vs. MAP 12.1 ± 0.5 s, P = 0.313) and FST (CBF 10.6 ± 0.4 s vs. MAP 10.1 ± 0.8 s, P = 0.552). Although the CBF and ABP decreases were not different in each test, the HR increase was significantly lower with the FST (24 ± 2 bpm) than with the SST (31 ± 3 bpm, P < 0.005). CONCLUSIONS: Earlobe CBF reflects the compensatory ABP regulatory response during standing and is potentially useful for estimating the orthostatic ABP response indirectly. Furthermore, the FST is a low-load protocol that can be an effective protocol for a standing test of cardiac function.

Estrogen decreases the expression of claudin-5 in vascular endothelial cells in the murine uterus.

Vascular endothelial (VE)-cadherin and claudin-5 are major components of the adherens and tight junctions of vascular endothelial cells, respectively, and decreases in their expression are associated with increases in endothelial paracellular permeability. In the uterus, estrogen induces endometrial edema. However, the in vivo effect of estrogen on endothelial paracellular permeability is unknown. Therefore, we studied the expression of VE-cadherin and claudin-5 in vascular endothelial cells in murine uteri stimulated by estrogen or progesterone. Ovariectomized mature mice were injected with estradiol-17ß (1 µg/mouse) or progesterone (1 mg/mouse) at intervals of 24 hours for 6 days. The frozen transverse sections of the uteri of these mice and untreated mice were stained for CD31 (vascular endothelial cell marker) plus VE-cadherin or claudin-5 using a double-immunofluorescence method. Then, the percentages of VE-cadherin- or claudin-5-positive vessels among CD31-positive vessels were examined in the uterine endometria. VE-cadherin and claudin-5 were expressed in most CD31-positive vessels in the endometria of the untreated mice. Progesterone did not affect the expression of both VE-cadherin and claudin-5 and estradiol-17ß also did not affect the VE-cadherin expression, but estradiol-17ß significantly decreased the claudin-5 expression. This decreasing effect of estradiol-17ß was detected from 24 hours later when the water content per a uterus significantly increased. The present study indicates that estrogen, but not progesterone, decreases the expression of claudin-5 in vascular endothelial cells in the murine uterine endometrium from 24 hours later, suggesting that the decrease in the claudin-5 expression contributes to the endometrial edema late after the estrogen stimulation.

Regulation of development of CD56 bright CD11c + NK-like cells with helper function by IL-18.

Human γδ T cells augment host defense against tumors and infections, and might have a therapeutic potential in immunotherapy. However, mechanism of γδ T cell proliferation is unclear, and therefore it is difficult to prepare sufficient numbers of γδ T cells for clinical immunotherapy. Recently, natural killer (NK)-like CD56(bright)CD11c(+) cells were shown to promote the proliferation of γδ T cells in an IL-18-dependent manner. In this study, we demonstrated that the NK-like CD56(bright)CD11c(+) cells could directly interact with γδ T cells to promote their sustained expansion, while conventional dendritic cells (DCs), IFN-α-induced DCs, plasmacytoid DCs or monocytes did not. We also examined the cellular mechanism underlying the regulation of CD56(bright)CD11c(+) cells. CD14(+) monocytes pre-incubated with IL-2/IL-18 formed intensive interactions with CD56(int)CD11c(+) cells to promote their differentiation to CD56(bright)CD11c(+) cells with helper function. The development of CD56(bright)CD11c(+) cells was suppressed in an IFN-α dependent manner. These results indicate that CD14(+) monocytes pretreated with IL-2/IL-18, but neither DCs nor monocytes, play a determining role on the development and proliferation of CD56(bright)CD11c(+) cells, which in turn modulate the expansion of γδ T cells. CD56(bright)CD11c(+) NK-like cells may be a novel target for immunotherapy utilizing γδ T cells, by overcoming the limitation of γδ T cells proliferation.

Decreased expression of VE-cadherin and claudin-5 and increased phosphorylation of VE-cadherin in vascular endothelium in nasal polyps.

VE-cadherin and claudin-5 are major components of adherens and tight junctions of vascular endothelial cells and a decrease in their expression and an increase in the tyrosine-phosphorylation of VE-cadherin are associated with an increase in endothelial paracellular permeability. To clarify the mechanism underlying the development of edema in nasal polyps, we studied these molecules in polyp microvessels. Normal inferior turbinate mucosal tissues and nasal polyps from patients treated with or without glucocorticoid were stained for VE-cadherin or claudin-5 and CD31 by a double-immunofluorescence method and the immunofluorescence intensities were graded 1-3 with increasing intensity. To correct for differences in fluorescence intensity attributable to a different endothelial area being exposed in a section or to the thickness of a section, the relative immunofluorescence intensity was estimated by dividing the grade of VE-cadherin or claudin-5 by that of CD31 in each microvessel. Tyrosine-phosphorylation of VE-cadherin was examined by Western blot analysis. The relative intensities of VE-cadherin and claudin-5 in the CD31-positive microvessels significantly decreased in the following order; inferior turbinate mucosa, treated polyps and untreated polyps. The ratio of tyrosine-phosphorylated VE-cadherin to VE-cadherin was significantly higher in untreated polyps than in the inferior turbinate mucosa and treated polyps, between which no significant difference in the ratio was seen. Thus, in nasal polyps, the barrier function of endothelial adherens and tight junctions is weakened, although glucocorticoid treatment improves this weakened barrier function.

Phenotypic characteristics and proliferative activity of hyperplastic ductule cells in cholangiofibrosis induced by thioacetamide in rats.

The oral administration of thioacetamide to rats induces cholangiofibrosis characterized by hyperplasia of ductules surrounded by fibrous tissue. In the present study, we examined the expression of markers of cholangiocyte and hepatocyte phenotypes in these hyperplastic ductule cells and their proliferative activity immunohistochemically. The oral administration of thioacetamide to 21-day-old male Fisher 344 rats for 12 weeks induced multiple areas of various sizes with hyperplastic ductules. The ductules consisted of two types of ductules; ductules composed of cholangiocyte-like cuboidal cells with transparent nuclei and cytoplasm, and of intestinal epithelium-like (IE-like) cells of basophilic nuclei and cytoplasm, and the transition of these two types of cells in the same ductule was sometimes observed. The cholangiocyte-like cells expressed cytokeratin (CK)-7, CK-19 and OV-6 (cholangiocyte phenotype markers) but not Hep Par-1 antigen or HNF4α (hepatocyte phenotype markers). In contrast, the IE-like cells expressed Hep Par-1 antigen and HNF4α but not CK-7, CK-19 or OV-6. The examination of Ki-67 expression showed a much higher proliferative activity for the IE-like cells compared to the cholangiocyte-like cells. The present results show that the hyperplastic ductules induced by thioacetamide are composed of IE-like cells with a high proliferative activity expressing the hepatocyte phenotype markers and of cholangiocyte-like cells with a low proliferative activity expressing the cholangiocyte phenotype markers.

Hypoxia downregulates the expression of cell surface MICA without increasing soluble MICA in osteosarcoma cells in a HIF-1α-dependent manner.

Tumor cells express NKG2D ligands on their cell surface, which are the ligands of the activating receptor, NKG2D, that is expressed on the surface of NK cells. The binding of NK cells to tumor cells through the interaction of NKG2D and its ligands induces the cytolysis of the tumor cells. In the present study, we investigated the effects of hypoxia on the expression of NKG2D ligands on the surface of human osteosarcoma cells using three cell lines. To produce hypoxic and normoxic conditions, the osteosarcoma cell lines were cultured under 1 and 20% O2 conditions, respectively. The osteosarcoma cells expressed NKG2D ligands such as MHC class I-related chain molecules A and B (MICA and MICB) and the UL16-binding proteins 1, 2 and 3 (ULBP 1, 2 and 3). MICA was the most frequently expressed NKG2D ligand in the osteosarcoma cells. Hypoxia decreased the expression of cell surface MICA only without increasing the secretion of soluble MICA, which is produced by proteolytic cleavage of cell surface MICA. Hypoxia consistently decreased the susceptibility of the osteosarcoma cells to the cytotoxicity of the NK cells. Hypoxia induced the expression of hypoxia-inducible factor-1α (HIF-1α), and knockdown of the expression of HIF-1α using small interfering RNA increased the expression of cell surface MICA and concomitantly increased the level of soluble MICA. Hypoxia decreased the production of nitric oxide (NO) metabolites (nitrite and nitrate), thus, indicating a decreasing effect on NO production. However, a NO donor, NOC18, decreased the expression of cell surface MICA without any apparent effects on the expression of HIF-1α under both hypoxic and normoxic conditions. The present results indicate that hypoxia downregulates the expression of cell surface MICA without increasing the level of soluble MICA in a HIF-1α-dependent manner and suggest that the effects of hypoxia are not linked to the hypoxia-induced reduction of NO production.

Downregulation of matrix metalloproteinase-9 mRNA by valproic acid plays a role in inhibiting the shedding of MHC class I-related molecules A and B on the surface of human osteosarcoma cells.

Valproic acid, a histone deacetylase inhibitor, increases the expression of cell surface MHC class I-related chain molecules (MICs) A and B (MICA and B) in osteosarcoma cells and decreases their secretion of soluble MICA and MICB, which are produced by the proteolytic cleavage of cell surface MICs. Osteosarcoma cells have been reported to produce high levels of matrix metalloproteinase (MMP)-2 and -9. In this study, we investigated the involvement of MMP-2 and -9 in the inhibitory action of valproic acid (VPA) on the proteolytic cleavage of cell surface MICs using the U-2 OS and SaOS-2 osteosarcoma cell lines. VPA caused a marked decrease in the expression of MMP-9 mRNA in the U-2 OS and SaOS-2 cells and in the expression of MMP-2 mRNA in the U-2 OS cells, but only a slight decrease in the expression of MMP-2 mRNA in the SaOS-2 cells. The transfection of small interfering RNA (siRNA) for MMP-9 decreased the secretion of soluble MICA and MICB by both U-2 OS and SaOS-2 cells, but that of siRNA for MMP-2 did not. The present study therefore demonstrates that the downregulation of MMP-9 mRNA by VPA plays a role in the inhibitory action of VPA on the secretion of soluble MICA and MICB in osteosarcoma cells.

Valproic acid cooperates with hydralazine to augment the susceptibility of human osteosarcoma cells to Fas- and NK cell-mediated cell death.

We investigated the effects of valproic acid (VPA), a histone deacetylase inhibitor, in combination with hydralazine, a DNA methylation inhibitor, on the expression of cell-surface Fas and MHC-class I-related chain molecules A and B (MICA and B), the ligands of NKG2D which is an activating receptor of NK cells, and on production of their soluble forms in HOS, U-2 OS and SaOS-2 human osteosarcoma cell lines. We also examined the susceptibility of these cells to Fas- and NK cell-mediated cell death. VPA did not increase the expression of Fas on the surface of osteosarcoma cells, while hydralazine did, and the combination of VPA with hydralazine increased the expression of cell-surface Fas. In contrast, the combination of VPA with hydralazine did not increase the production of soluble Fas by osteosarcoma cells. Both VPA and hydralazine increased the expression of cell-surface MICA and B in osteosarcoma cells, and their combination induced a greater increase in their expression. VPA inhibited the production of both soluble MICA and MICB by osteosarcoma cells while hydralazine produced no effect. Both VPA and hydralazine enhanced the susceptibility of osteosarcoma cells to Fas- and NK cell-mediated cell death and the combination of VPA with hydralazine further enhanced the effects. The present results suggest that combined administration of VPA and hydrazine is valuable for enhancing the therapeutic effects of immunotherapy for osteosarcomas.

The promotional effect of IL-22 on mineralization activity of periodontal ligament cells.

OBJECTIVES: Interleukin (IL)-22 acts on non-immune cells to induce anti-microbial responses, protection from tissue damage, and enhance cell regeneration. However, little is known about the involvement of IL-22 in periodontal biology. This study investigated the biological effects of IL-22 on periodontal ligament (PDL) cells as part of studies to assess the involvement of IL-22 in periodontal disease. MATERIALS AND METHODS: Gene expression levels of IL-22 and its receptors in PDL cells and gingival tissue samples were evaluated by real-time PCR. Proliferative responses and mineralized-matrix forming activities of PDL cells were examined in the presence and absence of IL-22. RESULTS: In contrast to the expression of IL-22 receptors detected in PDL tissues and their cell lines, gingival tissues showed modest or no gene expressions of IL-22. The production of several cytokines including IL-11, IL-8 and CCL2 was upregulated by IL-22 treatment of PDL cells in a dose-dependent manner. IL-22 treatment had no effect on the proliferative response in PDL cells. Meanwhile, IL-22 precipitated mineralized nodule formation and induced gene expressions of RUNX2, MSX2 and osteocalcin in PDL cells, suggesting that IL-22 enhances the mineralized matrix-forming activities of PDL cells. CONCLUSION: IL-22 has the potential to promote mineralizing activity in PDL cells and to develop appropriate regenerative therapy.

Small cell osteosarcoma successfully treated by high-dose ifosfamide and methotrexate, combined with carboplatin and pirarubicin.

Small cell osteosarcoma (SCO) is the most rare subtype of osteosarcoma and has a poor prognosis. An 11-year-old boy presented with 2-month history of painful tumefaction in the lower leg. Imaging analysis demonstrated a mixture of osteolytic and osteosclerotic lesions in the proximal tibia and extraskeletal area. Histology of the open biopsy showed small round cells producing mucous matrix. Based on these findings, SCO was suspected. The patient received three cycles of neoadjuvant chemotherapy using high-dose ifosfamide, high-dose methotrexate, pirarubicin and carboplatin. Wide-margin resection was performed followed by tibial lengthening using the Ilizarov method and two cycles of adjuvant chemotherapy with the same drugs as for neoadjuvant chemotherapy. Histology of the resected specimen showed that almost all tumor cells were necrotized. Neither recurrence nor metastasis was found after 4 years. Our experience suggests that neoadjuvant chemotherapy, such as the one used here, would be exceedingly effective for SCO without serious non-hematological toxicities.

Expression of both hepatocellular carcinoma and cholangiocarcinoma phenotypes in hepatocellular carcinoma and cholangiocarcinoma components in combined hepatocellular and cholangiocarcinoma.

Expression of phenotype markers of hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC) in HCC and CC components of 20 combined hepatocellular and cholangiocarcinomas (CHCs) of the liver was investigated immunohistochemically. Both HCC and CC components of all CHCs expressed at least one of the CC phenotype markers [cytokeratin (CK)-7, CK-19, and carbohydrate (CA) 19-9]. HCC components in 90% of CHCs and CC components in 95% of CHCs expressed at least one of these CC phenotype markers in more than 40% of cancer cells. HCC components in all CHCs expressed at least one of the HCC phenotype markers [hepatocyte antigen (HA), α-fetoprotein (AFP), and canalicular carcinoembryonic antigen]. HCC components in 90% of CHCs and CC components in 75% of CHCs expressed HA, AFP, or both. HCC components in 75% of CHCs and CC components in 60% of CHCs expressed HA, AFP, or both in more than 10% of cancer cells. The present results show that both HCC and CC components of most of the CHCs expressed both HCC and CC phenotypes, supporting the hypothesis that CHC originates from a hepatic progenitor cell capable of differentiating into hepatocytes and cholangiocytes.

Enhanced open-loop but not closed-loop cardiac baroreflex sensitivity during orthostatic stress in humans.

The neural interaction between the cardiopulmonary and arterial baroreflex may be critical for the regulation of blood pressure during orthostatic stress. However, studies have reported conflicting results: some indicate increases and others decreases in cardiac baroreflex sensitivity (i.e., gain) with cardiopulmonary unloading. Thus the effect of orthostatic stress-induced central hypovolemia on regulation of heart rate via the arterial baroreflex remains unclear. We sought to comprehensively assess baroreflex function during orthostatic stress by identifying and comparing open- and closed-loop dynamic cardiac baroreflex gains at supine rest and during 60° head-up tilt (HUT) in 10 healthy men. Closed-loop dynamic "spontaneous" cardiac baroreflex sensitivities were calculated by the sequence technique and transfer function and compared with two open-loop carotid-cardiac baroreflex measures using the neck chamber system: 1) a binary white-noise method and 2) a rapid-pulse neck pressure-neck suction technique. The gain from the sequence technique was decreased from -1.19 ± 0.14 beats·min(-1)·mmHg(-1) at rest to -0.78 ± 0.10 beats·min(-1)·mmHg(-1) during HUT (P = 0.005). Similarly, closed-loop low-frequency baroreflex transfer function gain was reduced during HUT (P = 0.033). In contrast, open-loop low-frequency transfer function gain between estimated carotid sinus pressure and heart rate during white-noise stimulation was augmented during HUT (P = 0.01). This result was consistent with the maximal gain of the carotid-cardiac baroreflex stimulus-response curve (from 0.47 ± 0.15 beats·min(-1)·mmHg(-1) at rest to 0.60 ± 0.20 beats·min(-1)·mmHg(-1) at HUT, P = 0.037). These findings suggest that open-loop cardiac baroreflex gain was enhanced during HUT. Moreover, under closed-loop conditions, spontaneous baroreflex analyses without external stimulation may not represent open-loop cardiac baroreflex characteristics during orthostatic stress.

A case of mixed ductal-endocrine carcinoma of the pancreas.

A 66-year-old male patient underwent a stomach-preserving pancreatoduodenectomy procedure because of a tumor located around the lower bile duct under the diagnosis of carcinoma of the lower bile duct. The tumor (3.5 × 2.5 cm) was found at the head of the pancreas and had invaded the papillae of Vater at the duodenum. Histology findings indicated both ductal adenocarcinoma and endocrine tumor. The ductal adenocarcinoma component expressed carcinoembryonic antigen, cytokeratin (CK)-19, CK-20, carbohydrate 19-9, and amylase, whereas the endocrine component, which occupied about one-third of the tumor, expressed glucagon, neuron-specific enolase, and chromogranin A. The Ki-67 labeling indices of the two components were 49.7% and 5.3%, respectively. Herein, we present this case of mixed ductal-endocrine carcinoma of the pancreas. Our findings indicate that its aggressive mass may be ascribable to the adenocarcinoma component with a high proliferative potential.

Involvement of CD56brightCD11c+ cells in IL-18-mediated expansion of human γδ T cells.

γδ T cells are considered to be innate lymphocytes that play an important role in host defense against tumors and infections. We recently reported that IL-18 markedly amplified γδ T cell responses to zoledronate (ZOL)/IL-2. In an extension of this finding, we analyzed the mechanism underlying the IL-18-mediated expansion of γδ T cells. After incubation of PBMCs with ZOL/IL-2/IL-18, the majority of the cells expressed γδ TCR, and the rest mostly exhibited CD56(bright)CD11c(+) under the conditions used in this study. CD56(bright)CD11c(+) cells were derived from a culture of CD56(int)CD11c(+) cells and CD14(+) cells in the presence of IL-2 and IL-18 without the addition of ZOL. They expressed IL-18Rs, HLA-DR, CD25, CD80, CD83, CD86, and CD11a/CD18. In addition, they produced IFN-γ, TNF-α, but not IL-12, when treated with IL-2/IL-18, and they exerted cytotoxicity against K562 cells, thus exhibiting characteristics of both NK cells and dendritic cells. Incubation of purified γδ T cells with CD56(bright)CD11c(+) cells in the presence of ZOL/IL-2/IL-18 resulted in the formation of massive cell clusters and led to the marked expansion of γδ T cells. However, both conventional CD56(-/int)CD11c(high) dendritic cells induced by GM-CSF/IL-4 and CD56(+)CD11c(-) NK cells failed to support the expansion of γδ T cells. These results strongly suggest that CD56(bright)CD11c(+) cells play a key role in the IL-18-mediated proliferation of γδ T cells.

Expression of hepatocyte markers in mass-forming peripheral and periductal-infiltrating hilar intrahepatic cholangiocarcinomas.

In this study, the expression of hepatocyte markers, including α-fetoprotein (AFP), HepPar-1 antigen and arginase-1, was examined immunohistochemically in 14 mass-forming peripheral intrahepatic cholangiocarcinomas (ICCs) that arose from the peripheral portion of the biliary tree, and in 14 periductal-infiltrating hilar ICCs that arose from intrahepatic large bile ducts. Only 2 (14.3%) of the 14 hilar ICCs and 2 (14.3%) of the 14 peripheral ICCs expressed AFP or HepPar-1 antigen. Conversely, arginase-1 was expressed in 8 (57.1%) and 11 (78.6%) of the hilar and peripheral ICCs, respectively, and 4 (28.6%) hilar ICCs and 7 (50%) peripheral ICCs expressed arginase-1 in more than 10% of the cancer cells. The expression of arginase-1 did not differ between peripheral ICCs showing major histology of poorly differentiated adenocarcinoma and those showing other major histologies, including well-or moderately differentiated tubular adenocarcinoma or papillary adenocarcinoma. Results of the present study showed that common hepatocyte markers, including AFP and HepPar-1 antigen, are rarely but definitely expressed in hilar and peripheral ICCs, and that a third hepatocyte marker, arginase-1, is expressed at a high rate in both hilar and peripheral ICCs, irrespective of their histology. These results indicate that care should be taken when using arginase-1 as a hepatocyte marker for distinguishing between a poorly differentiated hepatocellular carcinoma and a mass-forming peripheral ICC showing the histology of poorly differentiated adenocarcinoma.

Clinical and immunological assessment of periodontal disease in Japanese leprosy patients.

Periodontitis is a chronic inflammatory disease caused by the infection of periodontopathic bacteria in dental plaque. However, an individual's susceptibility to this disease appears to be associated with multiple genetic factors, as seen in the case of leprosy. In order to gain a better understanding of the pathophysiology of periodontal disease in subjects with leprosy, we investigated the clinical features of periodontitis and the immunological responses against periodontopathic bacteria in 382 subjects with a history of leprosy and 451 age-matched control subjects. The prevalence of periodontitis and the degree of periodontal pocket depth were found to be significantly higher in leprosy patients than in age-matched controls. Furthermore, a comparison of the clinical parameters of lepromatous leprosy (L-lep) and tuberculoid leprosy (T-lep) patients showed that the probing pocket depth of L-lep patients with periodontal disease was significantly higher than that for T-lep patients. In contrast, serum IgG titers against Porphyromonas gingivalis in L-lep patients were significantly lower than in T-lep patients. These results imply that L-lep patients show more severe periodontal disease than T-lep patients or age-matched control subjects, and that low humoral immunity against P. gingivalis might be one of the genetic factors determining periodontal disease susceptibility in leprosy patients.