Risk analysis of breast cancer and breast fibrocystic change in female primary Sjögren's syndrome patients.

Sjögren's syndrome is an autoimmune disease with the chronic inflammation of glandular tissues, typically salivary and lacrimal glands. Since mammary tissue shares the glandular structure, Sjögren's syndrome may also target mammary tissue to cause breast diseases. We therefore determined whether primary Sjögren's syndrome (pSS) is associated with the increased risk of breast cancer (BC) and breast fibrocystic change (FC). Total 282 female patients with pSS were recruited from a medical center in Taiwan, and patients' medical records were reviewed to identify BC and ultrasonographic breast FC. The prevalence, incidence and risk factors for BC and breast FC in pSS patients were determined, and the risk factors for these breast diseases were subsequently analyzed. Our results showed that pSS patients had the increased prevalence and incidence of BC, and high anti-SSA(Ro) antibody titers were found to be associated with the increased risk of BC. Breast FC was also found highly prevalent in these patients. Comorbidity analysis as risk factor for BC in pSS patients showed diabetes mellitus was strongly associated with the increased risk of BC (odds ratio = 10.4, P = .0006), whereas breast FC was inversely associated with the risk of BC (odds ratio = 0.077 P = .0158). These data indicated that pSS is association with the increased risk of BC and with the high prevalence of ultrasonographic breast FC. Our results also suggest that, in pSS patients, the high anti-SSA(Ro) antibody titers and diabetes mellitus confer the increased risk of BC, whereas the absence of ultrasonographic breast FC predicts the higher risk of BC.

Melatonin reduces neuropathic pain behavior and glial activation through MT2 melatonin receptor modulation in a rat model of lysophosphatidylcholine-induced demyelination neuropathy.

In this study, we investigated whether melatonin treatment prevents development of neuropathic pain via suppression of glial mitogen-activated protein kinases (MAPKs) activation in the cuneate nucleus (CN) in a lysophosphatidylcholine (LPC)-induced median nerve demyelination neuropathy model. Rats were fed orally with melatonin once a day at a dose of 37.5, 75, or 150 mg/kg 30 min before until 3 days after LPC treatment. Subsequently, behavioral tests were conducted on these animals, and immunohistochemistry and immunoblotting were used for qualitative and quantitative analysis of glia and MAPKs, including ERK, JNK, and p38, activation. Enzyme-linked immunosorbent assays were applied to measure pro-inflammatory cytokine responses. Furthermore, intra-CN microinjection of S26131 (MT1 receptor antagonist), 4P-PDOT (MT2 receptor antagonist), or prazosin (MT3 receptor antagonist) were performed to investigate the association between melatonin receptor subtypes and effects of melatonin on demyelination neuropathy. LPC treatment of the median nerve induced a significant increase in glial fibrillary acidic protein (GFAP; an astrocyte marker) and ED1 (an activated microglia marker) immunoreactivity in the ipsilateral CN and led to development of neuropathic pain behavior. Inspection of GFAP-immunoreactive astrocytes revealed that astrocytic hypertrophy, but not proliferation, contributed to increased GFAP immunoreactivity. Double immunofluorescence showed that both GFAP-immunoreactive astrocytes and ED1-immunoreactive microglia co-expressed p-ERK, p-JNK, and p-p38 immunoreactivity. Melatonin administration dose-dependently reduced neuropathic pain behavior, decreased glial and MAPKs activation, and diminished the release of pro-inflammatory cytokines in the ipsilateral CN after LPC treatment. Furthermore, 4P-PDOT, but not S26131 or prazosin, antagonized the therapeutic effects of melatonin. In conclusion, administration of melatonin, via its cognate MT2 receptor, inhibited activation of glial MAPKs, production of pro-inflammatory cytokines, and development of demyelination-induced neuropathic pain behavior.

Differential expressions of plasma proteins in systemic lupus erythematosus patients identified by proteomic analysis.

INTRODUCTION: Systemic lupus erythematosus (SLE) is a chronic and complex autoimmune disease with a wide range of clinical manifestations that affects multiple organs and tissues. Therefore the differential expression of proteins in the serum/plasma have potential clinical applications when treating SLE. METHODS: We have compared the plasma/serum protein expression patterns of nineteen active SLE patients with those of twelve age-matched and gender-matched healthy controls by proteomic analysis. To investigate the differentially expressed proteins among SLE and controls, a 2-dimensional gel electrophoresis coupled with high-resolution liquid chromatography tandem mass spectrometry was performed. To further understand the molecular and biological functions of the identified proteins, PANTHER and Gene Ontology (GO) analyses were employed. RESULTS: A total of 14 significantly expressed (p < 0.05, p < 0.01) proteins were identified, and of these nine were up-regulated and five down-regulated in the SLE patients. The functional enrichment analysis assigned the majority of the identified proteins including alpha 2 macroglobulin, complement C4, complement factor H, fibrinogen beta chain, and alpha-1-antitrypsin were part of the complement/coagulation cascade, which is an important pathway that plays a crucial role in SLE pathogenesis. In addition to these proteins the differential expressions of ceruloplasmin, transthyretin, and haptoglobin play a potential role in the renal system abnormalities of SLE. CONCLUSION: Therefore, the identified differentially expressed proteins are relevant to SLE patient's cohort. Most importantly the up-regulated proteins might be the potential candidates for renal system involvement in SLE disease pathogenesis. In order to confirm the diagnostic/therapeutic potential of the identified proteins, future validation studies are required.

Erythropoietin reduces nerve demyelination, neuropathic pain behavior and microglial MAPKs activation through erythropoietin receptors on Schwann cells in a rat model of peripheral neuropathy.

Neuroprotective effects of erythropoietin (EPO) on peripheral nerve injury remain uncertain. This study investigated the efficacy of EPO in attenuating median nerve chronic constriction injury (CCI)-induced neuropathy. Animals received an intraneural injection of EPO at doses of 1,000, 3,000, or 5,000 units/kg 15 min before median nerve CCI. Afterwards, the behavioral and electrophysiological tests were conducted. Immunohistochemistry and immunoblotting were used for qualitative and quantitative analysis of microglial and mitogen-activated protein kinases (MAPKs), including p38, JNK, and ERK, activation. Enzyme-linked immunosorbent assay and microdialysis were applied to measure pro-inflammatory cytokine and glutamate responses, respectively. EPO pre-treatment dose-dependently ameliorated neuropathic pain behavior, decreased microglial and MAPKs activation, and diminished the release of pro-inflammatory cytokines and glutamate in the ipsilateral cuneate nucleus after CCI. Moreover, EPO pre-treatment preserved myelination of the injured median nerve on morphological investigation and suppressed injury-induced discharges. We also observed that EPO receptor (EPOR) expression was up-regulated in the injured nerve after CCI. Double immunofluorescence showed that EPOR was localized to Schwann cells. Furthermore, siRNA-mediated knockdown of EPOR expression eliminated the therapeutic effects of EPO on attenuating the microglial and MAPKs activation, pro-inflammatory cytokine responses, injury discharges, and neuropathic pain behavior in CCI rats. In conclusion, binding of EPO to its receptors on Schwann cells maintains myelin integrity and blocks ectopic discharges in the injured median nerve, that in the end contribute to attenuation of neuropathic pain via reducing glutamate release from primary afferents and inhibiting activation of microglial MAPKs and production of pro-inflammatory cytokines.

Plasma proteomic analysis of systemic lupus erythematosus patients using liquid chromatography/tandem mass spectrometry with label-free quantification.

CONTEXT: Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disease with unknown etiology. OBJECTIVE: Human plasma is comprised of over 10 orders of magnitude concentration of proteins and tissue leakages. The changes in the abundance of these proteins have played an important role in various human diseases. Therefore, the research objective of this study is to identify the significantly altered expression levels of plasma proteins from SLE patients compared with healthy controls using proteomic analysis. The plasma proteome profiles of both SLE patients and controls were compared. METHODS: A total of 19 active SLE patients and 12 healthy controls plasma samples were analyzed using high-resolution electrospray ionization liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) followed by label-free quantification. RESULTS: A total of 19 proteins showed a significant level of expression in the comparative LC-ESI-MS/MS triplicate analysis; among these, 14 proteins had >1.5- to three-fold up-regulation and five had <0.2- to 0.6-fold down-regulation. Gene ontology and DAVID (Database Annotation Visualization, and Integrated Discovery) functional enrichment analysis revealed that these proteins are involved in several important biological processes including acute phase inflammatory responses, complement activation, hemostasis, and immune system regulation. CONCLUSION: Our study identified a group of differentially expressed proteins in the plasma of SLE patients that are involved in the imbalance of the immune system and inflammatory responses. Therefore, these findings may have the potential to be used as prognostic/diagnostic markers for SLE disease assessment or disease monitoring.

Management of gout and hyperuricemia: Multidisciplinary consensus in Taiwan.

Gout is an inflammatory disease manifested by the deposition of monosodium urate (MSU) crystals in joints, cartilage, synovial bursa, tendons or soft tissues. Gout is not a new disease, which was first documented nearly 5,000 years ago. The prevalence of gout has increased globally in recent years, imposing great disease burden worldwide. Moreover, gout or hyperuricemia is clearly associated with a variety of comorbidities, including cardiovascular diseases, chronic kidney disease, urolithiasis, metabolic syndrome, diabetes mellitus, thyroid dysfunction, and psoriasis. To prevent acute arthritis attacks and complications, earlier use of pharmacotherapeutic treatment should be considered, and patients with hyperuricemia and previous episodes of acute gouty arthritis should receive long-term urate-lowering treatment. Urate-lowering drugs should be used during the inter-critical and chronic stages to prevent recurrent gout attacks, which may elicit gradual resolution of tophi. The goal of urate-lowering therapy should aim to maintain serum uric acid (sUA) level <6.0 mg/dL. For patients with tophi, the initial goal can be set at lowering sUA to <5.0 mg/dL to promote tophi dissolution. The goal of this consensus paper was to improve gout and hyperuricemia management at a more comprehensive level. The content of this consensus paper was developed based on local epidemiology and current clinical practice, as well as consensuses from two multidisciplinary meetings and recommendations from Taiwan Guideline for the Management of Gout and Hyperuricemia.

TDAG8, TRPV1, and ASIC3 involved in establishing hyperalgesic priming in experimental rheumatoid arthritis.

Rheumatoid arthritis (RA), characterized by chronic inflammation of synovial joints, is often associated with ongoing pain and increased pain sensitivity. High hydrogen ion concentration (acidosis) found in synovial fluid in RA patients is associated with disease severity. Acidosis signaling acting on proton-sensing receptors may contribute to inflammation and pain. Previous studies focused on the early phase of arthritis (<5 weeks) and used different arthritis models, so elucidating the roles of different proton-sensing receptors in the chronic phase of arthritis is difficult. We intra-articularly injected complete Freund's adjuvant into mice once a week for 4 weeks to establish chronic RA pain. Mice with knockout of acid-sensing ion channel 3 (ASIC3) or transient receptor potential/vanilloid receptor subtype 1 (TRPV1) showed attenuated chronic phase (>6 weeks) of RA pain. Mice with T-cell death-associated gene 8 (TDAG8) knockout showed attenuated acute and chronic phases of RA pain. TDAG8 likely participates in the initiation of RA pain, but all three genes, TDAG8, TRPV1, and ASIC3, are essential to establish hyperalgesic priming to regulate the chronic phase of RA pain.

Analysis of the CD161-expressing cell quantities and CD161 expression levels in peripheral blood natural killer and T cells of systemic lupus erythematosus patients.

Expressed on the cell surface of most of NK cells and some T cells, CD161 has been shown to deliver inhibitory signal in human NK cells. To determine whether the CD161-expressing cell quantities and the cell surface expression levels of CD161 in NK and T cells were altered in systemic lupus erythematosus (SLE) patients, we analyzed the CD3, CD56 and CD161 expression patterns of peripheral blood lymphocytes by flow cytometric analysis to identify different NK and T cell subpopulations. The cell surface expression levels of CD161 were estimated by the mean florescence intensities (MFIs) of CD161. It was found that SLE patients had lower frequencies of CD161+CD56+CD3- and CD161+CD56+CD3+ cells among the lymphocyte population than normal controls, whereas the frequencies of CD161-CD56+CD3- and CD161+CD56-CD3+ cells were not statistically different between two groups. In addition, SLE patients also had decreased absolute counts of all CD161-expressing NK cells and T cells and had reduced frequencies of CD161+ cells in CD56+CD3-, CD56+CD3+ and CD56-CD3+ cell populations. Moreover, SLE patients had reduced MFIs of CD161 in CD161+CD56+CD3+ and CD161+CD56-CD3+, but not CD161+CD56+CD3-, cell populations. Our results indicated that CD161-expressing cell frequency and the CD161 expression levels were reduced in some NK and T cell subpopulations of SLE patients, suggesting possible important role of CD161 and CD161-expressing immune cells in the SLE pathogenesis.

Effect of Antrodia camphorata on inflammatory arterial thrombosis-mediated platelet activation: the pivotal role of protein kinase C.

Antrodia camphorata is a rare Taiwanese medicinal mushroom. Antrodia camphorata extract has been reported to exhibit antioxidant, anti-inflammation, antimetastasis, and anticancer activities and plays a role in liver fibrosis, vasorelaxation, and immunomodulation. Critical vascular inflammation leads to vascular dysfunction and cardiovascular diseases, including abdominal aortic aneurysms, hypertension, and atherosclerosis. Platelet activation plays a crucial role in intravascular thrombosis, which is involved in a wide variety of cardiovascular diseases. However, the effect of Antrodia camphorata on platelet activation remains unclear. We examined the effects of Antrodia camphorata on platelet activation. In the present study, Antrodia camphorata treatment (56-224 µg/mL) inhibited platelet aggregation induced by collagen, but not U46619, an analogue of thromboxane A2, thrombin, and arachidonic acid. Antrodia camphorata inhibited collagen-induced calcium (Ca(2+)) mobilization and phosphorylation of protein kinase C (PKC) and Akt. In addition, Antrodia camphorata significantly reduced the aggregation and phosphorylation of PKC in phorbol-12, 13-dibutyrate (PDBu) activated platelets. In conclusion, Antrodia camphorata may inhibit platelet activation by inhibiting of Ca(2+) and PKC cascade and the Akt pathway. Our study suggests that Antrodia camphorata may be a potential therapeutic agent for preventing or treating thromboembolic disorders.

Brazilin ameliorates high glucose-induced vascular inflammation via inhibiting ROS and CAMs production in human umbilical vein endothelial cells.

Vascular inflammatory process has been suggested to play a key role in the initiation and progression of atherosclerosis, a major complication of diabetes mellitus. Recent studies have shown that brazilin exhibits antihepatotoxic, antiplatelet, cancer preventive, or anti-inflammatory properties. Thus, we investigated whether brazilin suppresses vascular inflammatory process induced by high glucose (HG) in cultured human umbilical vein endothelial cells (HUVEC). HG induced nitrite production, lipid peroxidation, and intracellular reactive oxygen species formation in HUVEC cells, which was reversed by brazilin. Western blot analysis revealed that brazilin markedly inhibited HG-induced phosphorylation of endothelial nitric oxide synthase. Besides, we investigated the effects of brazilin on the MAPK signal transduction pathway because MAPK families are associated with vascular inflammation under stress. Brazilin blocked HG-induced phosphorylation of extracellular signal-regulated kinase and transcription factor NF-κB. Furthermore, brazilin concentration-dependently attenuated cell adhesion molecules (ICAM-1 and VCAM-1) expression induced by various concentrations of HG in HUVEC. Taken together, the present data suggested that brazilin could suppress high glucose-induced vascular inflammatory process, which may be closely related with the inhibition of oxidative stress, CAMs expression, and NF-κB activation in HUVEC. Our findings may highlight a new therapeutic intervention for the prevention of vascular diseases.

Reduced toll-like receptor 9 expression on peripheral CD14+ monocytes of chronic hepatitis B patients and its restoration by effective therapy.

BACKGROUND: Chronic hepatitis B (CHB) patients display Toll-like receptor 9 (TLR9)-dependent defective immune responses. We aimed to study TLR9 expression on CHB patients and its alteration during therapy. METHODS: We compared TLR9 expression on fresh peripheral CD14+ monocytes from a cohort of 97 CHB patients and 35 HBsAg-negative, anti-HCV-negative controls, during pegylated interferon or entecavir therapy. TLR9 expression on liver tissue was also investigated. RESULTS: Compared with controls, peripheral CD14+ monocytes of CHB patients displayed reduced expression of TLR9 mean fluorescence intensity (MFI; 9.90 ±3.64 versus 7.95 ±3.61; P=0.007) independent of age, gender and alanine aminotransferase (ALT; -2.09, 95% CI -3.568, -0.613; P=0.006). Furthermore, age, gender, ALT, HBeAg status, quantitative HBsAg (qHBsAg) or HBV DNA did not predict the TLR9 expression (P=0.863). Hepatic TLR9 messenger RNA (mRNA) was significantly reduced in 54 patients compared with 3 controls (0.45 ±0.32 versus 1-fold). Using response-guided therapy by qHBsAg levels and pretreatment TLR9 MFI as a reference, TLR9 MFI restored to a mean of 1.7- to 2.7-fold in pegylated interferon responders and reduced to a mean of 0.6- to 0.7-fold in non-responders starting from treatment week 12. Among 10 entecavir-treated patients, TLR9 MFI gradually restored to a mean of 1.2- to 2.1-fold starting from treatment week 48. CONCLUSIONS: CHB patients display reduced TLR9 expression on peripheral CD14+ monocytes, which is independent of host and viral markers, and on liver tissue. Responders to pegylated interferon and those under entecavir demonstrate restoration of TLR9 expression. On-treatment TLR9 expression on peripheral monocytes might predict response to pegylated interferon therapy.

Glycan masking of hemagglutinin for adenovirus vector and recombinant protein immunizations elicits broadly neutralizing antibodies against H5N1 avian influenza viruses.

The highly pathogenic avian influenza (HPAI) H5N1 virus, a known trigger of diseases in poultry and humans, is perceived as a serious threat to public health. There is a clear need for a broadly protective H5N1 vaccine or vaccines for inducing neutralizing antibodies against multiple clades/subclades. We constructed single, double, and triple mutants of glycan-masked hemagglutiinin (HA) antigens at residues 83, 127 and 138 (i.e., g83, g127, g138, g83+g127, g127+g138, g83+g138 and g83+g127+g138), and then obtained their corresponding HA-expressing adenovirus vectors and recombinant HA proteins using a prime-boost immunization strategy. Our results indicate that the glycan-masked g127+g138 double mutant induced more potent HA-inhibition, virus neutralization antibodies, cross-clade protection against heterologous H5N1 clades, correlated with the enhanced bindings to the receptor binding sites and the highly conserved stem region of HA. The immune refocusing stem-specific antibodies elicited by the glycan-masked H5HA g127+g138 and g83+g127+g138 mutants overlapped with broadly neutralizing epitopes of the CR6261 monoclonal antibody that neutralizes most group 1 subtypes. These findings may provide useful information in the development of a broadly protective H5N1 influenza vaccine.

Genome-wide association study in NSAID-induced acute urticaria/angioedema in Spanish and Han Chinese populations.

AIM: Acute urticaria/angioedema (AUA) induced by cross-intolerance to NSAIDs is the most frequent clinical entity in hypersensitivity reactions to drugs. In this work, we conducted a genome-wide association study in Spanish and Han Chinese patients suffering from NSAID-induced AUA. MATERIALS & METHODS: A whole-genome scan was performed on a total of 232 cases (112 Spanish and 120 Han Chinese) with NSAID-induced AUA and 225 unrelated controls (124 Spanish and 101 Han Chinese). RESULTS: Although no polymorphism reached genome-wide significance, we obtained suggestive associations for three clusters in the Spanish group (RIMS1, BICC1 and RAD51L 1) and one region in the Han Chinese population (ABI3BP). Five regions showed suggestive associations after meta-analysis: HLF, RAD51L1, COL24A1, GalNAc-T13 and FBXL7. A majority of these genes are related to Ca(2+), cAMP and/or P53 signaling pathways. CONCLUSION: The associations described were different from those related to the metabolism of arachidonic acid and could provide new mechanisms underlying NSAID-induced AUA.

Increased risk of cirrhosis and its decompensation in chronic hepatitis B patients with newly diagnosed diabetes: a nationwide cohort study.

BACKGROUND: The impact of diabetes on cirrhosis, its decompensation, and their time relationship in patients with chronic hepatitis B (CHB) remain unclear. METHODS: We conducted a nationwide cohort study by using the Taiwanese National Health Insurance Research Database, which was comprised of data from >99% of the entire population. Among 1 million randomly sampled enrollees, 14 523 adult CHB patients were identified from 1997 to 2009. Diabetes was defined as newly diagnosed in CHB patients who were given the diagnosis in the years 1998-2001 but not in 1996-1997 and with physician visits of at least twice per year. The cohorts of CHB with newly diagnosed diabetes (n = 351) and without diabetes (n = 7886) were followed up from the diagnosis of diabetes and from 2000 in the patients without diabetes until development of cirrhosis or its decompensation, withdrawal from insurance, or December 2009. RESULTS: Kaplan-Meier survival analysis showed a significantly higher cumulative incidence of cirrhosis (relative risk [RR] = 3.43; 95% confidence interval [CI], 2.62-4.49; P < .001, log-rank test) and decompensated cirrhosis (RR = 4.11; 95% CI, 2.95-5.70; P < .001, log-rank test) among patients with newly developed diabetes compared with those without diabetes. After adjustment for age, sex, CHB treatment, hepatocellular carcinoma, and comorbidity index by Cox proportional hazards model, diabetes was still an independent predictor for cirrhosis (hazard ratio [HR] = 2.015; 95% CI, 1.393-2.915; P < .001) and its decompensation (HR = 1.792; 95% CI, 1.192-2.695; P = .005). CONCLUSIONS: Patients with CHB who develop diabetes are at an increased risk of liver cirrhosis and its decompensation over time.

Heterologous prime-boost immunization regimens using adenovirus vector and virus-like particles induce broadly neutralizing antibodies against H5N1 avian influenza viruses.

Highly pathogenic avian influenza (HPAI) H5N1 viruses continue to trigger severe diseases in poultry and humans, prompting efforts to develop an effective vaccine. Toward that goal, we constructed a recombinant adenovirus vector encoding influenza hemagglutin (rAd-HA) and a flagellin-containing virus-like particle (FliC-VLP). Using a murine model, we investigated a heterologous prime-boost vaccination regimen combining these two vectors. Our results indicate that priming with the rAd-HA vector followed by a FliC-VLP booster induced the highest HA-specific total IgG, IgG1and IgG2a. Maximum neutralizing antibody titers against homologous and heterologous clades of H5N1 virus strains and hemagglutination inhibition resulted from the heterologous vaccination strategy. Our results are likely to contribute to the development of more effective H5N1 vaccines.

Formulation and immunological evaluation of a trivalent vaccine comprising emulsified submicron particles and inactivated virions of H5N1/EV71/JEV.

Combination vaccines can reduce the number of injections and simplify the immunization schedule required to prevent different diseases. Here we assessed the immunogenicity in a mouse model of a vaccine composition comprising inactivated influenza viruses (H5N1/H1N1), enterovirus 71 (EV71), and/or Japanese encephalitis virus (JEV) and investigated whether the vaccine formulations can overcome the immunologic interference between the individual vaccine components. We demonstrated that the antigenic competition happens between H5N1/H1N1 or H5N1/EV71 inactivated virions when the vaccine combinations either formulated with Alum suspensions or without adjuvant. In the presence of PELC emulsified particles, EV71-specific immune responses before and after incorporating H5N1 virus into EV71 vaccine were detected of no significant difference; in addition, H5N1- and EV71-specific immune responses were found at the same level when H5N1/EV71/JEV consolidating into combination vaccine. Emulsified vaccine formulation was represented as a potential tool that is found to reduce the number of injections required to prevent multiple infectious strains causing the same disease (H5N1/H1N1) and/or that protect against different diseases (H5N1/EV71). Combination vaccines can also include a third component to protect against H5N1/EV71/JEV at the same time.

Reduced Toll-like receptor 3 expression in chronic hepatitis B patients and its restoration by interferon therapy.

BACKGROUND: Toll-like receptor (TLR)3 gene variants may correlate with clinical significance of chronic viral infections including HBV. We aimed to investigate the expression of TLR3 in peripheral blood mononuclear cells (PBMCs) and liver cells of chronic hepatitis B (CHB) patients and its response to pegylated interferon or nucleoside analogue therapy. METHODS: We consecutively enrolled 127 CHB patients and 64 hepatitis B surface antigen-negative, anti-HCV-negative healthy individuals as controls. We compared the TLR3 expressions on fresh PBMCs and liver cells from patients and controls, before and during pegylated interferon or nucleoside analogue therapy. RESULTS: Compared to controls, patients had a lower TLR3 mean fluorescence intensity (MFI) on PBMCs (mean ± sd 14.61 ± 13.49 versus 9.70 ± 4.61; P < 0.001), independent of age, gender and alanine aminotransferase (ALT; -13.466, 95% CI -17.202, -9.730; P < 0.001). Patients had limited TLR3 stains on Kupffer cells, whereas controls had diffuse stains on Kupffer and hepatocytes. Hepatic TLR3 messenger RNA was lower in patients than controls (0.47 ± 0.30 versus 1-fold). Using pretreatment TLR3 MFI as a referent, among 5 of 12 pegylated-interferon-treated patients with sustained virological response (SVR), TLR3 MFI was restored to a mean of 1.5- to 1.7-folds immediately after treatment. Among seven non-responders or relapsers, TLR3 MFI reduced to a mean of 0.5- to 0.7-fold. Among 10 entecavir-treated patients with on-treatment virological response, TLR3 MFI gradually was restored to a mean of 1.2-folds during 48-week therapy. CONCLUSIONS: CHB patients have reduced TLR3 expression on PBMCs, independent of age, gender and ALT, and on liver cells. Patients with pegylated-interferon-induced SVR have a more significant restoration of TLR3 expression than those under entecavir.

Different immunity elicited by recombinant H5N1 hemagglutinin proteins containing pauci-mannose, high-mannose, or complex type N-glycans.

Highly pathogenic avian influenza H5N1 viruses can result in poultry and occasionally in human mortality. A safe and effective H5N1 vaccine is urgently needed to reduce the pandemic potential. Hemagglutinin (HA), a major envelope protein accounting for approximately 80% of spikes in influenza virus, is often used as a major antigen for subunit vaccine development. In this study, we conducted a systematic study of the immune response against influenza virus infection following immunization with recombinant HA proteins expressed in insect (Sf9) cells, insect cells that contain exogenous genes for elaborating N-linked glycans (Mimic) and mammalian cells (CHO). While the antibody titers are higher with the insect cell derived HA proteins, the neutralization and HA inhibition titers are much higher with the mammalian cell produced HA proteins. Recombinant HA proteins containing tri- or tetra-antennary complex, terminally sialylated and asialyated-galactose type N-glycans induced better protective immunity in mice to lethal challenge. The results are highly relevant to issues that should be considered in the production of fragment vaccines.

Prognostic value of plasma N-terminal pro B-type natriuretic peptide levels in pneumonia patients requiring intensive care unit admission.

BACKGROUND AND OBJECTIVE: Correct and early risk stratification for critically ill pneumonia patients remains an unmet medical need. This study aimed to test whether N-terminal pro B-type natriuretic peptide (NT-proBNP) can serve as a prognostic marker in this setting. METHODS: This prospective study enrolled 216 pneumonia patients admitted to intensive care unit. Plasma NT-proBNP samples were obtained upon admission and primary outcome was all-cause mortality at 30 days. Meanwhile, Acute Physiology and Chronic Health Evaluation (APACHE) II and Infectious Diseases Society of America/American Thoracic Society (IDSA/ATS) 2007 minor criteria were assessed. RESULTS: Overall 30-day mortality was 30%. NT-proBNP levels were significantly higher in nonsurvivors than survivors (11 938 ± 13 121 vs 5658 ± 9240 pg/mL, P = 0.001). Area under receiver operating characteristic curves of NT-proBNP, APACHE II and IDSA/ATS 2007 minor criteria were not significantly different regarding prediction of mortality (0.715, 0.754 vs 0.654, P = 0.085). Adding NT-proBNP to APACHE II significantly increased the area under receiver operating characteristic curve from 0.754 to 0.794 (P = 0.048). Receiver operating characteristic analysis revealed optimal NT-proBNP and APACHE II cut-offs of 2177.5 pg/mL and 25.5, respectively. In multivariate analysis, both NT-proBNP and APACHE II values above cut-offs had a significantly higher probability of death than those below cut-offs. A categorical approach combining NT-proBNP and APACHE II cut-offs provides additional risk stratification over a single marker approach. CONCLUSIONS: For pneumonia patients admitted to intensive care unit, NT-proBNP strongly and independently predicts mortality, and its prognostic accuracy is comparable with APACHE II and IDSA/ATS 2007 minor criteria.

Development of monocyte Toll-like receptor 2 and Toll-like receptor 4 in preterm newborns during the first few months of life.

BACKGROUND: Although the immaturity of Toll-like receptor 2 (TLR2) and Toll-like receptor 4 (TLR4) at birth in preterm newborns is known, their development during the first few months of life remains unclear. METHODS: Blood monocytes of preterm newborns (gestational age: 24-36 wk) were obtained every 2 wk when possible in order to perform serial measurements of TLR2 and TLR4 surface expression, as well as lipopolysaccharide (LPS)-induced cytokine production. Measurements using monocytes from term newborns and adults were also performed. RESULTS: The monocytes of preterm newborns obtained at birth displayed reduced surface expression of TLR2 and TLR4, and diminished responses of tumor necrosis factor-α (TNF-α) and interleukin (IL)-8 to LPS stimulation. Regardless of gestational age, monocyte expression of TLR2 and TLR4 in preterm newborns increased rapidly within the first 2 wk after birth, quickly reaching those of term newborns. These increases continued for the following 4-6 wk, although the increase began to plateau. By contrast, LPS-induced production of TNF-α and IL-8 did not elevate over this period in preterm newborns. CONCLUSION: The blood monocytes of preterm newborns display rapid increase in TLR2 and TLR4 expression during the first few months of life, whereas LPS-induced cytokine production functionality did not improve in parallel.