Recruitment of CCR6+ Foxp3+ regulatory gastric infiltrating lymphocytes in Helicobacter pylori gastritis.

Helicobacter pylori (H. pylori) infection is associated with an inflammatory response in the gastric mucosa, leading to chronic gastritis, peptic ulcers, and gastric cancer. Increased T-cell infiltration is found at sites of H. pylori infection. The CCR6+ subset of CD4+ regulatory T cells (Tregs), a newly characterized subset of Tregs, has been reported to contribute to local immune inhibition. However, whether CCR6+ Tregs are present in H. pylori gastritis, and what their relationship is to disease prognosis, remains to be elucidated. In this study, gastric infiltrating lymphocytes were isolated from endoscopic biopsy specimens of H. pylori gastritis patients and analyzed. We found that in gastric infiltrating lymphocytes, CCR6+  CD4+  CD25high Tregs, which express high levels of CD45RO, are positively associated with more severe inflammation in gastric mucosa during H. pylori infection. Furthermore, the frequency of CCR6+ Tregs in gastric infiltrating lymphocytes, but not CCR6- Tregs, is significantly increased in inflamed gastric tissues, which is inversely correlated with significantly lower expression of IFN-γ+  CD8+ T cells. We also found that the frequency of CCR6+ Tregs is positively correlated with the frequency of CD4+  IFN-γ+ T cells. In addition, the frequency of CCR6+ Tregs, but not that of CCR6- Tregs, is significantly correlated with increased inflammation in H. pylori gastritis. This study demonstrates that immunosuppression in H. pylori gastritis might be related to the activity of CCR6+ Tregs, which could influence disease prognosis.

GSK-3 inhibitors enhance TRAIL-mediated apoptosis in human gastric adenocarcinoma cells.

Resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis has been reported in some cancer cells, including AGS human gastric adenocarcinoma cells. Reducing this resistance might shed light on the treatment of human gastric adenocarcinoma. In this study, we examined whether glycogen synthase kinase-3 (GSK-3) inhibitors can restore TRAIL responsiveness in gastric adenocarcinoma cells. The effect of two GSK-3 inhibitors, SB-415286, and LiCl, on apoptosis signaling of TRAIL in human gastric adenocarcinoma cell lines and primary gastric epithelial cells was analyzed. Both inhibitors can sensitize gastric adenocarcinoma cells, but not primary gastric epithelial cells, to TRAIL-induced apoptosis by increasing caspase-8 activity and its downstream signal transmission. Adding p53 siRNA can downregulate GSK-3 inhibitor-related sensitization to TRAIL-induced apoptosis and caspase-3 activity. GSK-3 inhibitors strongly activate the phosphorylation of JNK. Inhibition of JNK leads to earlier and more intense apoptosis, showing that the activation of JNK may provide anti-apoptotic equilibrium of pro-apoptotic cells. Our observations indicate that GSK-3 inhibitors can sentize AGS gastric adenocarcinoma cells to TRAIL-induced apoptosis. Therefore, in certain types of gastric adenocarcinoma, GSK-3 inhibitor might enhance the antitumor activity of TRAIL and mightbe a promising candidate for the treatment of certain types of gastric adenocarcinoma.

Interleukin-32 plays an essential role in human calcified aortic valve cells.

Interleukin-32 (IL-32) is an inflammatory cytokine produced mainly by T, natural killer, and epithelial cells. Previous studies on IL-32 have primarily investigated its proinflammatory properties. The IL-32 also has been described as an activator of the p38 mitogen-activated protein kinase (MAPK) and NF-κB, and induces several cytokines. In this study, we hypothesized that the inflammatory regulators NF-κB, MAP kinase, STAT1, and STAT3 are associated with the expression of the IL-32 protein in human calcified aortic valve cells. This study comprised aortic valve sclerotic patients and control group patients without calcified aortic valve. Increased IL-32 expression in calcified aortic valvular tissue was shown by immunohistochemical staining and western blotting. There was an increase in NF-κB p65 level, p-ERK, p-JNK, and p-p38 MAPK activation underlying IL-32 expression in the study. The level of p-STAT3 but not p-STAT1 was found to be increased in calcified aortic valve tissue. In cultured primary human aortic valve interstitial cells, inhibition of NF-κB or MAPK kinase pathways results in a decrease of IL-32 expression. Treatment of recombinant IL-32 induced the levels of TNF-α, IL-6, IL-1ß, and IL-8. Our findings demonstrate that IL-32 may be an important pro-inflammatory molecule involved in calcific aortic valve disease.

A paeonol derivative, YPH-PA3 promotes the differentiation of monocyte/macrophage lineage precursor cells into osteoblasts and enhances their autophagy.

Previous studies have indicated that paeonol inhibits RANKL-induced osteoclastogenesis by inhibiting the ERK, p38, and NF-κB pathway. We modified paeonol to form a new compound, YPH-PA3, and found that it promoted osteoclastogenesis rather than inhibited it the way paeonol does. The aim of this study is to investigate the mechanisms involved in YPH-PA3-promoted osteoclastogenesis. YPH-PA3-promoted differentiation of RAW264.7 cells (human monocytes) into osteoclasts is activated through ERK/p38/JNK phosphorylation, affecting c-FOS, NF-κB, and NFATc2. Real-time quantitative PCR and western blot revealed an increased expression of autophagy-related markers during YPH-PA3-induced osteoclastogenesis. We also demonstrated the relationship between p62/LC3 localization and F-actin ring formation by double-labeling immunofluorescence. Knockdown of p62 small-interfering RNA (siRNA) attenuated YPH-PA3-induced expression of autophagy-related genes. Our study results indicated that p62 may play a role in YPH-PA3-induced autophagy and osteoclastogenesis, which may help to develop a novel therapeutic strategy against osteoclastogenesis-related diseases.

Radix Paeoniae Rubra stimulates osteoclast differentiation by activation of the NF-κB and mitogen-activated protein kinase pathways.

BACKGROUND: Radix Paeoniae Rubra (RPR), a traditional Chinese herb, has anti-inflammatory and immuno-regulatory properties. This study explored the effects of RPR on stimulation of osteoclast differentiation in RAW264.7 cells and peripheral blood mononuclear cells (PBMC)s. METHODS: The mature osteoclasts were measured by bone resorption assays and TRAP staining. JNK, ERK, p38 and NF-κB inhibitors were used applied in order to verify their contribution in RPR-induced osteoclast differentiation. The NF-κB and MAPK pathways were evaluated by western blotting, RT-PCR and luciferase assay. RESULTS: RPR induced osteoclast differentiation in a dose-dependent manner and induced the resorption activity of osteoclasts differentiation of RAW264.7 cells and PBMCs. Western blotting showed that RPR treatment induced phosphorylation of JNK, ERK, and p38 in RAW 264.7 cells. Treatment of JNK, ERK, and p38 MAP kinase inhibitors verified the contribution of JNK, ERK and p38. RPR treatment induced c-Fos and NFATc1 protein expression; NF-κB inhibitor treatment and luciferase assay verified the contribution of the NF-κB pathway. CONCLUSIONS: This study demonstrated the interesting effect, in which RPR stimulated osteoclast differentiation in murine RAW264.7 cells and human monocytes.

Gallic Acid Induces Apoptosis in Human Gastric Adenocarcinoma Cells.

BACKGROUND/AIM: Gastric cancer is one of the most common malignant cancers with a poor prognosis and high mortality rate worldwide. Current treatment of gastric cancer includes surgery and chemotherapy as the main modalities, but the potentially severe side-effects of chemotherapy present a considerable challenge. Gallic acid is a trihydroxybenzoic acid found to exert an anticancer effect against a variety of cancer cells. The purpose of this study was to determine the anti-cancer activity of Galla chinensis and its main component gallic acid on human gastric adenocarcinoma cells. MATERIALS AND METHODS: MTT assay and cell death ELISA were used to determine the apoptotic effect of Gallic Chinensis and gallic acid on human gastric adenocarcinoma cells. To determine the pathway and relevant components by which gallic acid-induced apoptosis is mediated through, cells were transfected with siRNA (Fas, FasL, DR5, p53) using Lipofectamine 2000. Reults: Gallic Chinensis and gallic acid induced apoptosis of human gastric adenocarcinoma cells. Gallic acid induced up-regulation of Fas, FasL, and DR5 expression in AGS cells. Transfection of cells with Fas, FasL, or DR5 siRNA reduced gallic acid-induced cell death. In addition, p53 was shown to be involved in gallic acid-mediated Fas, FasL, and DR5 expression as well as cell apoptosis in AGS cells. CONCLUSION: These results suggest that gallic acid has a potential role in the treatment of gastric cancer.

PD-1 and PD-L1 Up-regulation Promotes T-cell Apoptosis in Gastric Adenocarcinoma.

BACKGROUND/AIM: The programmed death 1 (PD-1) receptor and its ligand (PD-L1) play pivotal roles in regulating host immune responses. However, the inhibitory effects of this pathway on the function of tumor infiltrating T lymphocytes in gastric adenocarcinoma patients are not well-defined. MATERIALS AND METHODS: We characterized the expression of PD-1 and PD-L1 in peripheral blood and tumor infiltrating cells and analyzed the association between PD-1/PD-L1 expression and disease progression in a cohort of 60 patients with Helicobacter pylori infection, including 18 with gastric adenocarcinoma, 23 with gastritis, and 19 asymptomatic controls. RESULTS: Relative to controls, the expression of PD-1 on peripheral blood and tumor infiltrating T cells increased with disease progression. In vitro, T cells induced PD-L1 expression on primary gastric adenocarcinoma epithelial cells in an IFN-γ-dependent manner, which in turn promoted T cells apoptosis. Blocking of PD-L1 reversed this effect. CONCLUSION: This study provides evidence for a new therapeutic target in gastric adenocarcinoma patients.

Expression of CD25(high) regulatory T cells and PD-1 in gastric infiltrating CD4(+) T lymphocytes in patients with Helicobacter pylori infection.

We observed by flow cytometry that the frequency of both gastric infiltrating Tregs and PD-1-positive CD4 T cells is correlated with the density of Helicobacter pylori, suggesting that cellular immunity against this pathogen is inhibited.

Increased PD-1 and decreased CD28 expression in chronic hepatitis B patients with advanced hepatocellular carcinoma.

BACKGROUND/AIMS: Hepatitis B infection is a well-known cause of hepatocellular carcinoma (HCC). This study aims to investigate the role that the co-stimulatory molecule CD28 and co-inhibitory molecule programmed death-1 (PD-1) play in compromising the function of tumour-infiltrating lymphocytes (TIL) in hepatitis B virus (HBV)-related HCC. METHODS: A total of 45 patients with HBV-related HCC were enrolled during the period February 2008 to March 2010. The immune phenotype and the expression of PD-1, CD28 and CD127 in TIL in biopsy specimens and in peripheral blood lymphocytes (PBL) from the same patients were analysed by flow cytometry. RESULTS: Among the 45 patients, there was a male predominance (80%) and the mean age was 50 ± 13.68 years (range: 29-71). The majority of TIL were CD45RO(+) CD69(+). PD-1 expression was higher and CD28 and CD127 expression levels were lower in TIL than in PBL. The prevalence of portal vein thrombosis was 40%. Furthermore, tumour thrombosis invasion into the portal vein correlated with the expression level of the PD-1 co-inhibitory molecule. CONCLUSION: PD-1(+) tumour-infiltrating lymphocytes correlate with portal vein thrombosis and might serve as a potential prognostic marker of and a novel therapeutic target for HBV-related HCC.

Clinical course and outcome of children with status asthmaticus treated in a pediatric intensive care unit: a 15-year review.

BACKGROUND AND PURPOSE: The purpose of this study was to analyze the clinical course and outcomes of children with status asthmaticus treated in a pediatric intensive care unit (PICU) in Taiwan. METHODS: The medical charts of all patients aged from 2 to 18 years with status asthmaticus who were admitted to the PICU National Taiwan University Hospital, Taipei, Taiwan, from 1990 to 2006 were reviewed retrospectively. RESULTS: Twenty eight children were admitted on 33 occasions; 4 patients had more than 1 admission. The mean age was 6.23 years (range, 2-17 years). No control medication was regularly used prior to admission for 81.8% of the episodes. The mean PICU stay was 2.55 days (range, 1-13 days) and, for 78.8% of episodes, the patients were discharged from the ICU within 2 days. For 11 episodes (33.3%), the patients required mechanical ventilation for a mean duration of 3.6 days. The mortality rate was 3.3% (n = 1). Complications included bronchopneumonia (57.6%), hemodynamic compromise (9.1%), neurologic symptoms (6.1%), and air leak (3.3%). Patients requiring mechanical ventilation had significantly lower initial pulse oximeter oxygen saturation (<90%), lower blood gas pH (<7.25), higher partial pressure of carbon dioxide, and longer duration of PICU and hospital stay (p < 0.05) when compared with patients not requiring mechanical ventilation. For patients with a history of repeated PICU admissions for asthma, the intubation rate was 77.7%. CONCLUSIONS: The prognosis for childhood status asthmaticus is favorable. The most common trigger factors were respiratory tract infection and poor asthma control.

Early detection of glutaric aciduria type I by newborn screening in Taiwan.

BACKGROUND/PURPOSE: Glutaric aciduria type 1 (GA1) is an inborn error of lysine and tryptophan metabolism. There is a lack of initial diagnostic signs of the disease, but late treatment often results in severe neurologic impairment. In this study, we analyzed the results of screening for GA1 in a Chinese population. METHODS: Dry blood spots were obtained at about 3 days of age from 357,307 newborns and tested for elevation of glutaryl (C5DC)-carnitine by tandem mass spectroscopy. A second sample of blood spots was required from those cases with abnormal elevation of C5DC-carnitine (higher than the cut-off value) (recall). If the results remained abnormal, those cases were referred for confirmation of the diagnosis and treatment. RESULTS: Between August 2001 and February 2005, there were 40 cases with C5DC-carnitine more than 0.13 microM (the cut-off value), from whom a second sample of blood spots was obtained (recall rate, 0.02%); two cases were confirmed to be affected by GA1. Because of the low positive prediction rate using this cut-off value, we elevated the cut-off value slightly. Between February 2005 and August 2006, there were eight cases with C5DC-carnitine more than 0.22 microM from whom a second sample of blood spots was obtained (recall rate, 0.01%); three cases were confirmed to be affected by GA1. All five cases with persistent elevation of C5DC-carnitine were referred and diagnosis was confirmed in each, giving an incidence of 1 in 71,461 newborns. There were no false negatives. Magnetic resonance imaging studies obtained from four cases showed frontotemporal atrophy at the time of diagnosis. Two cases were followed for over 1 year, and under treatment with dietary control and carnitine supplementation, both had normal development and neither exhibited a frank episode of encephalopathic crisis. CONCLUSION: With properly established cut-offs, GA1 can be successfully screened for in populations with a low incidence of the disease. Early treatment is likely to improve the outcome of cases discovered by screening.

Seizure associated with total parenteral nutrition-related hypermanganesemia.

The trace element manganese is usually supplied when total parenteral nutrition is used. However, long-term parenteral administration of manganese, which bypasses the normal regulatory mechanism, may cause hypermanganesemia. Manganese poisoning presents clinically with parkinsonian-like symptoms and psychological changes. Seizures are a rare presentation of this disease. This report describes a 10-year-old female who had received total parenteral nutrition for 3 months because of short bowel syndrome, and presented with tonic-clonic seizure, decreased level of consciousness, and fever. The serum electrolytes, glucose and the cerebrospinal fluid examination were normal. The blood culture grew Pantoea agglomerans. The brain magnetic resonance imaging disclosed no evidence of central nervous system infection. However, symmetric high-intensity signal on T1-weighted images was documented in the basal ganglia, especially in the globus pallidus. Her whole blood manganese level was 3.7 microg/dL, which was significantly higher than the normal range (0.4-1.4 microg/dL). Diagnosis of hypermanganesemia related to total parenteral nutrition was made.