A Biomarker Panel (Bioscore) Incorporating Monocytic Surface and Soluble TREM-1 Has High Discriminative Value for Ventilator-Associated Pneumonia: A Prospective Observational Study

Introduction

Ventilator-associated pneumonia (VAP) increases mortality in critical illness. However, clinical diagnostic uncertainty persists. We hypothesised that measuring cell-surface and soluble inflammatory markers, incorporating Triggering Receptor Expressed by Myeloid cells (TREM)-1, would improve diagnostic accuracy.

Methods

A single centre prospective observational study, set in a University Hospital medical-surgical intensive Care unit, recruited 91 patients into 3 groups: 27 patients with VAP, 33 ventilated controls without evidence of pulmonary sepsis (non-VAP), and 31 non-ventilated controls (NVC), without clinical infection, attending for bronchoscopy. Paired samples of Bronchiolo-alveolar lavage fluid (BALF) and blood from each subject were analysed for putative biomarkers of infection: Cellular (TREM-1, CD11b and CD62L) and soluble (IL-1β, IL-6, IL-8, sTREM-1, Procalcitonin). Expression of cellular markers on monocytes and neutrophils were measured by flow cytometry. Soluble inflammatory markers were determined by ELISA. A biomarker panel (‘Bioscore’), was constructed, tested and validated, using Fisher’s discriminant function analysis, to assess its value in distinguishing VAP from non VAP.

Results

The expression of TREM-1 on monocytes (mTREM-1) and neutrophils (nTREM-1) and concentrations of IL-1β, IL-8, and sTREM-1 in BALF were significantly higher in VAP compared with non-VAP and NVC (p<0.001). The BALF/blood mTREM-1 was significantly higher in VAP patients compared to non-VAP and NVC (0.8 v 0.4 v 0.3 p<0.001). A seven marker Bioscore (BALF/blood ratio mTREM-1 and mCD11b, BALF sTREM-1, IL-8 and IL-1β, and serum CRP and IL-6) correctly identified 88.9% of VAP cases and 100% of non-VAP cases.

Conclusion

A 7-marker bioscore, incorporating cellular and soluble TREM-1, accurately discriminates VAP from non-pulmonary infection.     

Nebulized Fibrinolytic Agents Improve Pulmonary Fibrinolysis but Not Inflammation in Rat Models of Direct and Indirect Acute Lung Injury

Background

Critically ill patients frequently develop acute lung injury (ALI). Disturbed alveolar fibrin turnover, a characteristic feature of ALI, is the result of both activation of coagulation and inhibition of fibrinolysis. Nebulized fibrinolytic agents could exert lung–protective effects, via promotion of fibrinolysis as well as anti–inflammation.

Methods

Rats were challenged intratracheally with Pseudomonas aeruginosa, resulting in pneumonia as a model for direct ALI, or received an intravenous bolus infusion of lipopolysaccharide, as a model for indirect ALI. Rats were randomized to nebulization of normal saline (placebo), recombinant tissue plasminogen activator (rtPA), or monoclonal antibodies against plasminogen activator inhibitor–type 1 (anti–PAI–1).

Results

Nebulized rtPA or anti–PA1–1 enhanced the bronchoalveolar fibrinolytic system, as reflected by a significant reduction of PAI–1 activity levels in bronchoalveolar lavage fluid, and a consequent increase in plasminogen activator activity (PAA) levels to supranormal values. Both treatments also significantly affected systemic fibrinolysis as reflected by a significant increase in PAA levels in plasma to supranormal levels. Neither nebulized rtPA nor anti–PA1–1 affected pulmonary inflammation. Neither treatment affected bacterial clearance of P. aeruginosa from the lungs in case of pneumonia.

Conclusions

Local treatment with rtPA or anti–PA1–1 affects pulmonary fibrinolysis but not inflammation in models of direct or indirect ALI in rats.     

Timing of Antimicrobial Therapy after Identification of Ventilator-Associated Condition Is Not Associated with Mortality in Patients with Ventilator-Associated Pneumonia: A Cohort Study

Purpose

Delays in antimicrobial therapy increase mortality in ventilator-associated pneumonia (VAP). The more objective ventilator-associated complications (VAC) are increasingly used for quality reporting. It is unknown if delays in antimicrobial administration, after patients meet VAC criteria, leads to worse outcomes.

Materials and Methods

Cohort of 81 episodes of antimicrobial treatment for VAP. We compared mortality, superinfections and treatment failures conditional on the timing of identification of VAC.

Results

60% of patients with VAC had an identifiable episode at least 48 before the initiation of antimicrobials. Antimicrobial administration after the identification of VAC was not associated with intensive care unit (ICU) mortality (OR 0.71, 95% CI 0.11–4.48, p = 0.701) compared to immediate antimicrobial administration. Similarly, the risk of treatment failure or superinfection was not affected by the timing of administration of antimicrobials in VAC (HR 0.95, 95% CI 0.42–2.19, p = 0.914).

Conclusions

We observed no signal of harm associated with the timing to initiate antimicrobials after the identification of a VAC. The identification of VAC should not lead clinicians to start antimicrobials before a diagnosis of VAP can be established.     

Disaturated-phosphatidylcholine and Surfactant protein-B turnover in human acute lung injury and in control patients

Background

Patients with Adult Respiratory Distress Syndrome (ARDS) and Acute Lung Injury (ALI) have low concentrations of disaturated-phosphatidylcholine and surfactant protein-B in bronchoalveolar lavage fluid. No information is available on their turnover.

Objectives

To analyze disaturated-phosphatidylcholine and surfactant protein-B turnover in patients with ARDS/ALI and in human adults with normal lungs (controls).

Methods

²H₂O as precursor of disaturated-phosphatidylcholine-palmitate and 1¹³C-Leucine as precursor of surfactant protein-B were administered intravenously to 12 patients with ARDS/ALI and to 8 controls. Disaturated-phosphatidylcholine and surfactant protein-B were isolated from serial tracheal aspirates, and their fractional synthetic rate was derived from the ²H and ¹³C enrichment curves, obtained by gas chromatography mass spectrometry. Disaturated-phosphatidylcholine, surfactant protein-B, and protein concentrations in tracheal aspirates were also measured.

Results

1) Surfactant protein-B turned over at faster rate than disaturated-phosphatidylcholine both in ARDS/ALI patients and in controls. 2) In patients with ARDS/ALI the fractional synthesis rate of disaturated-phosphatidylcholine was 3.1 times higher than in controls (p < 0.01), while the fractional synthesis rate of surfactant protein-B was not different. 3) In ARDS/ALI patients the concentrations of disaturated-phosphatidylcholine and surfactant protein-B in tracheal aspirates were markedly and significantly reduced (17% and 40% of the control values respectively).

Conclusions

1) Disaturated-phosphatidylcholine and surfactant protein-B have a different turnover both in healthy and diseased lungs. 2) In ARDS/ALI the synthesis of these two surfactant components may be differently regulated.     

Anergic Pulmonary Tuberculosis Is Associated with Contraction of the Vd2+T Cell Population,Apoptosis and Enhanced Inhibitory Cytokine Production

Objective

To study the association of anergic pulmonary tuberculosis with Vδ2⁺ T cells and related cytokine levels.

Methods

82 pulmonary tuberculosis patients were divided into two groups according to their purified protein derivative tuberculin skin test (TST) results: 39 with TST-negative anergic pulmonary tuberculosis and 43 with TST-positive pulmonary tuberculosis, while 40 healthy volunteers were used as control. Based on chest X-ray results, the tuberculosis lesions were scored according to their severity, with a score of ≤ 2.5 ranking as mild, 2.5-6 as moderate and ≥ 6 as severe. The Vδ2⁺ T cell percentage and their expression levels of the apoptosis-related membrane surface molecule FasL in peripheral blood and bronchoalveolar lavage fluids (BALF) were analyzed by flow cytometry, while IL-2, IL-4, IL-6 and IL-10 cytokine and γ-interferon (γ-IFN) concentrations in peripheral blood were determined by ELISA.

Results

Most of the patients with chest X-ray lesion scores higher than 6 belonged to the anergic tuberculosis group (P<0.05). Anergic pulmonary tuberculosis patients displayed reduced peripheral blood Vδ2⁺ T cell counts (P<0.05) and higher FasL expression in peripheral blood Vδ2 ⁺ T cells (P <0.05). The Vδ2⁺ T cell percentages in the BALF of all tuberculosis patients were lower than in their peripheral blood (P <0.05), and IL-4 and IL-10 concentrations in peripheral blood of anergic tuberculosis patients were higher than in TST-positive tuberculosis patients and healthy controls (P <0.05).

Conclusion

Anergic pulmonary tuberculosis is accompanied by reduced Vδ2⁺ T cell percentage, and elevated Vδ2⁺ T cell FasL expression as well as enhanced IL-4 and IL-10 levels in peripheral blood.     

Inhibiting toll-like receptor 4 signaling ameliorates pulmonary fibrosis during acute lung injury induced by lipopolysaccharide: an experimental study

Background

Toll-like receptor 4 (TLR4) is essential in lipopolysaccharide (LPS)-induced fibroblast activation and collagen secretion in vitro. However, its effects on the process of lung fibroblast activation and fibrosis initiation during LPS induced acute lung injury (ALI) remain unknown. The goal of the present study was to determine the effect of inhibiting TLR4 on LPS-induced ALI and fibrosis in vivo.

Methods

The ALI model was established by intraperitoneal injection of LPS in mice. TLR4-small hairpin RNA (shRNA) lentivirus was injected intravenously into the mice to inhibit TLR4 expression. mRNA and protein levels were detected by real-time PCR and Western-blot analysis, respectively. The contents of the C-terminal propeptide of type I procollagen (PICP) in bronchoalveolar lavage fluid (BALF) were detected by ELISA, and the degree of fibrosis was detected by van Gieson collagen staining, the hydroxyproline assay, and alpha smooth muscle actin (α-SMA) immunohistochemical staining.

Results

Overexpression of TLR4, type I procollagen, alpha-SMA, and p-AKT in murine pulmonary tissue after intraperitoneal injection of LPS at 72 hours and 28 days were detected. Moreover, the degree of fibrosis was shown to increase by ELISA analysis of PICP in BALF, van Gieson collagen staining, the hydroxyproline assay, and α-SMA immunohistochemical staining. All of these changes were alleviated by intravenous infection with TLR4-shRNA lentivirus.

Conclusions

Inhibiting TLR4 signaling could ameliorate fibrosis at the early stage of ALI induced by LPS.     

Altered expression of T cell Immunoglobulin-Mucin (TIM) molecules in bronchoalveolar lavage CD4+ T cells in sarcoidosis

Background

Activated T helper (Th)-1 pulmonary CD4⁺ cells and their mediators are essential for the inflammation and granulomatous process in sarcoidosis. Recently, T-cell immunoglobulin and mucin domain (TIM) molecules were suggested to be important regulators of immune function. In this study, we wanted to investigate whether TIM molecules could play a role in sarcoidosis.

Methods

We used real-time polymerase chain reaction to investigate the differential gene expression of TIM-1 and TIM-3 as well as a few Th1 and Th2 cytokines (IL-2, IFN-γ, IL-4, IL-5 and IL-13) in CD4⁺ T cells isolated from bronchoalveolar lavage fluid (BALF) of patients (n = 28) and healthy controls (n = 8). Using flow cytometry, we were also able to analyse TIM-3 protein expression in 10 patients and 6 healthy controls.

Results

A decreased TIM-3 mRNA (p < 0.05) and protein (p < 0.05) expression was observed in patients, and the level of TIM-3 mRNA correlated negatively with the CD4/CD8 T cell ratio in BALF cells of patients. Compared to a distinct subgroup of patients i.e. those with Löfgren''s syndrome, BALF CD4⁺ T cells from non- Löfgren''s patients expressed decreased mRNA levels of TIM-1 (p < 0.05). mRNA expression of IL-2 was increased in patients (p < 0.01) and non-Löfgren''s patients expressed significantly higher levels of IFN-γ mRNA (p < 0.05) versus patients with Löfgren''s syndrome.

Conclusion

These findings are the first data on the expression of TIM-1 and TIM-3 molecules in sarcoidosis. The reduced TIM-3 expression in the lungs of patients may result in a defective T cell ability to control the Th1 immune response and could thus contribute to the pathogenesis of sarcoidosis. The down-regulated TIM-1 expression in non-Löfgren''spatients is in agreement with an exaggerated Th1 response in these patients.     

Apamin Does Not Inhibit Human Cardiac Na+ Current,L-type Ca2+ Current or Other Major K+ Currents

Background

Apamin is commonly used as a small-conductance Ca²⁺-activated K⁺ (SK) current inhibitor. However, the specificity of apamin in cardiac tissues remains unclear.

Objective

To test the hypothesis that apamin does not inhibit any major cardiac ion currents.

Methods

We studied human embryonic kidney (HEK) 293 cells that expressed human voltage-gated Na⁺, K⁺ and Ca²⁺ currents and isolated rabbit ventricular myocytes. Whole-cell patch clamp techniques were used to determine ionic current densities before and after apamin administration.

Results

Ca²⁺ currents (CACNA1c+CACNB2b) were not affected by apamin (500 nM) (data are presented as median [25^th percentile;75^th percentile] (from –16 [–20;–10] to –17 [–19;–13] pA/pF, P = NS), but were reduced by nifedipine to –1.6 [–3.2;–1.3] pA/pF (p = 0.008). Na⁺ currents (SCN5A) were not affected by apamin (from –261 [–282;–145] to –268 [–379;–132] pA/pF, P = NS), but were reduced by flecainide to –57 [–70;–47] pA/pF (p = 0.018). None of the major K⁺ currents (I _Ks, I _Kr, I _K1 and I _to) were inhibited by 500 nM of apamin (KCNQ1+KCNE1, from 28 [20]; [37] to 23 [18]; [32] pA/pF; KCNH2+KCNE2, from 28 [24]; [30] to 27 [24]; [29] pA/pF; KCNJ2, from –46 [–48;–40] to –46 [–51;–35] pA/pF; KCND3, from 608 [505;748] to 606 [454;684]). Apamin did not inhibit the I _Na or I _CaL in isolated rabbit ventricular myocytes (I _Na, from –67 [–75;–59] to –68 [–71;–59] pA/pF; I _CaL, from –16 [–17;–14] to –14 [–15;–13] pA/pF, P = NS for both).

Conclusions

Apamin does not inhibit human cardiac Na⁺ currents, L-type Ca²⁺ currents or other major K⁺ currents. These findings indicate that apamin is a specific SK current inhibitor in hearts as well as in other organs.     

Complementary Role of HCV and HIV in T-Cell Activation and Exhaustion in HIV/HCV Coinfection

Objectives

To investigate whether T-cell activation and exhaustion is linked to HCV- and HIV disease parameters in HIV/HCV infected individuals, we studied T-cell characteristics in HIV/HCV coinfected patients and controls.

Methods

14 HIV/HCV coinfected, 19 HCV monoinfected, 10 HIV monoinfected patients and 15 healthy controls were included in this cross-sectional study. Differences in expression of activation and exhaustion markers (HLA-DR, CD38, PD-1, Tim-3 and Fas) and phenotypic markers on CD4⁺ and CD8⁺ T-cells were analysed by flow cytometry and were related to HCV disease parameters (HCV-viremia, ALT and liver fibrosis).

Results

Frequencies of activated CD4⁺ and CD8⁺ T-cells were higher in HIV/HCV-coinfected compared to healthy controls and HCV or HIV mono-infected individuals. Coinfected patients also showed high expression of the exhaustion marker PD-1 and death receptor Fas. In contrast, the exhaustion marker Tim-3 was only elevated in HIV-monoinfected patients. T-cell activation and exhaustion were correlated with HCV-RNA, suggesting that viral antigen influences T-cell activation and exhaustion. Interestingly, increased percentages of effector CD8⁺ T-cells were found in patients with severe (F3–F4) liver fibrosis compared to those with no to minimal fibrosis (F0–F2).

Conclusions

HIV/HCV coinfected patients display a high level of T-cell activation and exhaustion in the peripheral blood. Our data suggest that T-cell activation and exhaustion are influenced by the level of HCV viremia. Furthermore, high percentages of cytotoxic/effector CD8⁺ T-cells are associated with liver fibrosis in both HCV monoinfected and HIV/HCV coinfected patients.     

Increased Expression of IL-22 Is Associated with Disease Activity in Behcet’s Disease

Objective

Interleukin (IL)-22 has been reported to be involved in the development of autoimmune diseases. This study aimed to analyze the expression and potential role of IL-22 in the pathogenesis of Behcet’s disease (BD).

Methods

The levels of IL-22 in patient sera or supernatants of cultured peripheral blood mononuclear cells (PBMCs) and CD4⁺T cells were detected by enzyme-linked immunosorbent assay (ELISA). Flow cytometry was used to evaluate the frequency of IL-22–producing CD4⁺ T cells. IL-22 mRNA from erythema nodosum skin lesions was examined using real time quantitative RT-PCR.

Results

BD patients with active uveitis showed a significantly higher expression of IL-22 in the supernatants of stimulated PBMCs and CD4⁺T cells compared with BD patients without active uveitis and normal controls. An increased frequency of IL-22-producing CD4⁺T cells was also found in BD patients with active uveitis. IL-22 mRNA expression was elevated in erythema nodosum skin lesions. In BD patients, a high IL-22 level in the supernatant of stimulated PBMCs correlated with the presence of retinal vasculitis and erythema nodosum.

Conclusions

IL-22 was associated with disease activity in BD and correlated with the presence of small vessel inflammation, suggesting that it may be involved in its pathogenesis.     

High levels of memory B cells are associated with response to a first tumor necrosis factor inhibitor in patients with rheumatoid arthritis in a longitudinal prospective study

Introduction

Tumor necrosis factor inhibitor (TNFi) therapy is effective for rheumatoid arthritis (RA). Some researchers have suggested that TNFi therapy affects B-cell homeostasis. We studied the effect of TNFi therapy on the distribution of peripheral B-cell subsets to elucidate B-cell–related biomarkers to predict the TNFi response.

Methods

Peripheral B cells were analyzed for expression of CD19, CD27, CD38 and immunoglobulin D in 31 healthy donors and 96 RA patients, including 21 patients who were followed 3 months after TNFi initiation.

Results

Treatment with steroids significantly altered the distribution of B-cell subsets. After we adjusted for age, sex and steroid dose, we found that patients with RA had B-cell subset proportions similar to controls. B-cell subset distributions did not differ upon use of TNFi at baseline or before or after TNFi introduction. TNFi responders (according to European League Against Rheumatism criteria) at 3 months had significantly higher proportions of CD27⁺ memory B cells at baseline, and ≥26% CD27⁺ cells at inclusion was associated with a relative risk of 4.9 (1.3 to 18.6) for response to TNFi treatment. CD27⁺ cells produced three times more TNFα than did TNFi-naïve B cells and were correlated with interferon γ produced from CD4⁺ cells in patients without TNFi treatment.

Conclusions

In patients with RA, high levels of baseline memory B cells were associated with response to TNFi, which may be related to TNFα-dependent activation of the T helper type 1 cell pathway.     

Phenotype and Functions of Natural Killer Cells in Critically-Ill Septic Patients

Rationale

Natural killer cells, as a major source of interferon-γ, contribute to the amplification of the inflammatory response as well as to mortality during severe sepsis in animal models.

Objective

We studied the phenotype and functions of circulating NK cells in critically-ill septic patients.

Methods

Blood samples were taken <48 hours after admission from 42 ICU patients with severe sepsis (n = 15) or septic shock (n = 14) (Sepsis group), non-septic SIRS (n = 13) (SIRS group), as well as 21 healthy controls. The immuno-phenotype and functions of NK cells were studied by flow cytometry.

Results

The absolute number of peripheral blood CD3–CD56⁺ NK cells was similarly reduced in all groups of ICU patients, but with a normal percentage of NK cells. When NK cell cytotoxicity was evaluated with degranulation assays (CD107 expression), no difference was observed between Sepsis patients and healthy controls. Under antibody-dependent cell cytotoxicity (ADCC) conditions, SIRS patients exhibited increased CD107 surface expression on NK cells (62.9[61.3–70]%) compared to healthy controls (43.5[32.1–53.1]%) or Sepsis patients (49.2[37.3–62.9]%) (p = 0.002). Compared to healthy (10.2[6.3–13.1]%), reduced interferon-γ production by NK cells (K562 stimulation) was observed in Sepsis group (6.2[2.2–9.9]%, p<0.01), and especially in patients with septic shock. Conversely, SIRS patients exhibited increased interferon-γ production (42.9[30.1–54.7]%) compared to Sepsis patients (18.4[11.7–35.7]%, p<0.01) or healthy controls (26.8[19.3–44.9]%, p = 0.09) in ADCC condition.

Conclusions

Extensive monitoring of the NK-cell phenotype and function in critically-ill septic patients revealed early decreased NK-cell function with impaired interferon-γ production. These results may aid future NK-based immuno-interventions.

Trial Registration

NTC00699868.     

Anti-inflammatory effects of novel curcumin analogs in experimental acute lung injury

Background

Acute lung injury (ALI) and its most severe form acute respiratory distress syndrome (ARDS) have been the leading cause of morbidity and mortality in intensive care units (ICU). Currently, there is no effective pharmacological treatment for acute lung injury. Curcumin, extracted from turmeric, exhibits broad anti-inflammatory properties through down-regulating inflammatory cytokines. However, the instability of curcumin limits its clinical application.

Methods

A series of new curcumin analogs were synthesized and screened for their inhibitory effects on the production of TNF-α and IL-6 in mouse peritoneal macrophages by ELISA. The evaluation of stability and mechanism of active compounds was determined using UV-assay and Western Blot, respectively. In vivo, SD rats were pretreatment with c26 for seven days and then intratracheally injected with LPS to induce ALI. Pulmonary edema, protein concentration in BALF, injury of lung tissue, inflammatory cytokines in serum and BALF, inflammatory cell infiltration, inflammatory cytokines mRNA expression, and MAPKs phosphorylation were analyzed. We also measured the inflammatory gene expression in human pulmonary epithelial cells.

Results

In the study, we synthesized 30 curcumin analogs. The bioscreeening assay showed that most compounds inhibited LPS-induced production of TNF-α and IL-6. The active compounds, a17, a18, c9 and c26, exhibited their anti-inflammatory activity in a dose-dependent manner and exhibited greater stability than curcumin in vitro. Furthermore, the active compound c26 dose-dependently inhibited ERK phosphorylation. In vivo, LPS significantly increased protein concentration and number of inflammatory cells in BALF, pulmonary edema, pathological changes of lung tissue, inflammatory cytokines in serum and BALF, macrophage infiltration, inflammatory gene expression, and MAPKs phosphorylation . However, pretreatment with c26 attenuated the LPS induced increase through ERK pathway in vivo. Meanwhile, compound c26 reduced the LPS-induced inflammatory gene expression in human pulmonary epithelial cells.

Conclusions

These results suggest that the novel curcumin analog c26 has remarkable protective effects on LPS-induced ALI in rat. These effects may be related to its ability to suppress production of inflammatory cytokines through ERK pathway. Compound c26, with improved chemical stability and bioactivity, may have the potential to be further developed into an anti-inflammatory candidate for the prevention and treatment of ALI.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-015-0199-1) contains supplementary material, which is available to authorized users.     

Expansion of CD4+CD25+ helper T cells without regulatory function in smoking and COPD

Background

Regulatory T cells have been implicated in the pathogenesis of COPD by the increased expression of CD25 on helper T cells along with enhanced intracellular expression of FoxP3 and low/absent CD127 expression on the cell surface.

Method

Regulatory T cells were investigated in BALF from nine COPD subjects and compared to fourteen smokers with normal lung function and nine never-smokers.

Results

In smokers with normal lung function, the expression of CD25⁺CD4⁺ was increased, whereas the proportions of FoxP3⁺ and CD127⁺ were unchanged compared to never-smokers. Among CD4⁺ cells expressing high levels of CD25, the proportion of FoxP3⁺ cells was decreased and the percentage of CD127⁺ was increased in smokers with normal lung function. CD4⁺CD25⁺ cells with low/absent CD127 expression were increased in smokers with normal lung function, but not in COPD, when compared to never smokers.

Conclusion

The reduction of FoxP3 expression in BALF from smokers with normal lung function indicates that the increase in CD25 expression is not associated with the expansion of regulatory T cells. Instead, the high CD127 and low FoxP3 expressions implicate a predominantly non-regulatory CD25⁺ helper T-cell population in smokers and stable COPD. Therefore, we suggest a smoking-induced expansion of predominantly activated airway helper T cells that seem to persist after COPD development.     

Neoadjuvant oral vs. infusional chemoradiotherapy on locally advanced rectal cancer: Prognostic factors

Aim

To evaluate the prognostic factors and impact on survival of neoadjuvant oral and infusional chemoradiotherapy in patients with locally advanced rectal cancer.

Background

There is still no definitive consensus about the prognostic factors and the impact of neoadjuvant chemoradiotherapy on survival. Some studies have pointed to an improvement in overall survival (OS) and progression-free survival (PFS) in patients with tumor downstaging (TD) and nodal downstaging (ND).

Materials and methods

A set of 159 patients with LARC were treated preoperatively. Group A – 112 patients underwent concomitant oral chemoradiotherapy: capecitabine or UFT + folinic acid. Group B – 47 patients submitted to concomitant chemoradiation with 5-FU in continuous infusion. 63.6% of patients were submitted to adjuvant chemotherapy.

Results

Group A: pathologic complete response (pCR) – 18.7%; TD – 55.1%; ND – 76%; loco-regional response – 74.8%. Group B: pCR – 11.4%; TD – 50%; ND – 55.8%; LRR – 54.5%. The loco-regional control was 95.6%. There was no difference in survival between both groups. Those with loco-regional response had better PFS.

Conclusions

Tumor and nodal downstaging, loco-regional response and a normal CEA level turned out to be important prognostic factors in locally advanced rectal cancer. Nodal downstaging and loco-regional response were higher in Group A. Those with tumor downstaging and loco-regional response from Group A had better OS. Adjuvant chemotherapy had no impact on survival except in those patients with loco-regional response who achieved a higher PFS.     

HLA-A2 and B35 Restricted Hantaan Virus Nucleoprotein CD8+ T-Cell Epitope-Specific Immune Response Correlates with Milder Disease in Hemorrhagic Fever with Renal Syndrome

Background

Hantaan virus (HTNV) infection in humans is a serious public health concern in Asia. A potent T cell activation peptide vaccine from HTNV structure protein represents a promising immunotherapy for disease control. However, the T cell epitopes of the HTNV restricted by the HLA alleles and the role of epitope-specific T cell response after HTNV infection remain largely unexplored.

Methodology/Principal Findings

Five well-conserved novel CD8⁺ T-cell epitopes of the HTNV nucleoprotein restricted by the most popular HLA alleles in Chinese Han population were defined with interferon-γ enzyme-linked immunospot assay in 37 patients infected with HTNV during hospitalization. Two epitopes aa129–aa137 and aa131–aa139 restricted by HLA-A2 and B35, respectively, were selected to evaluate the epitope-specific CD8⁺ T-cell response. HLA-peptide pentamer complex staining showed that the frequency of single epitope-specific CD8⁺ T cell could be detected in patients (95% confidence interval for aa129–aa137: 0.080%–0.208%; for aa131–aa139: 0.030%–0.094%). The frequency of epitope-specific pentamer⁺ CD8⁺ T-cell response was much higher in mild/moderate patients than in severe/critical ones at the acute stage of the disease. Moreover, the frequency of epitope-specific CD8⁺ T cells at acute stage was inversely associated with the peak level of serum creatinine and was positively associated with the nadir platelet counts during the hospitalization. The intracellular cytokine staining and the proliferation assay showed that the effective epitope-specific CD8⁺ T cells were characterized with the production of interferon-γ, expression of CD69 and the strong capacity of proliferation.

Conclusion/Significance

The novel HLA class I restricted HTNV nucleoprotein epitopes-specific CD8⁺ T-cell responses would be closely related with the progression and the severity of the disease, which could provide the first step toward effective peptide vaccine development against HTNV infection in humans.     

Decreased respiratory system compliance on the sixth day of mechanical ventilation is a predictor of death in patients with established acute lung injury

Background

Multiple studies have identified single variables or composite scores that help risk stratify patients at the time of acute lung injury (ALI) diagnosis. However, few studies have addressed the important question of how changes in pulmonary physiologic variables might predict mortality in patients during the subacute or chronic phases of ALI. We studied pulmonary physiologic variables, including respiratory system compliance, P/F ratio and oxygenation index, in a cohort of patients with ALI who survived more than 6 days of mechanical ventilation to see if changes in these variables were predictive of death and whether they are informative about the pathophysiology of subacute ALI.

Methods

Ninety-three patients with ALI who were mechanically ventilated for more than 6 days were enrolled in this prospective cohort study. Patients were enrolled at two medical centers in the US, a county hospital and a large academic center. Bivariate analyses were used to identify pulmonary physiologic predictors of death during the first 6 days of mechanical ventilation. Predictors on day 1, day 6 and the changes between day 1 and day 6 were compared in a multivariate logistic regression model.

Results

The overall mortality was 35%. In multivariate analysis, the PaO₂/FiO₂ (OR 2.09, p < 0.04) and respiratory system compliance (OR 3.61, p < 0.01) were predictive of death on the 6^th day of acute lung injury. In addition, a decrease in respiratory system compliance between days 1 and days 6 (OR 2.14, p < 0.01) was independently associated with mortality.

Conclusions

A low respiratory system compliance on day 6 or a decrease in the respiratory system compliance between the 1^st and 6^th day of mechanical ventilation were associated with increased mortality in multivariate analysis of this cohort of patients with ALI. We suggest that decreased respiratory system compliance may identify a subset of patients who have persistent pulmonary edema, atelectasis or the fibroproliferative sequelae of ALI and thus are less likely to survive their hospitalization.     

Quantitative Proteomic Analysis of Serum from Pregnant Women Carrying a Fetus with Conotruncal Heart Defect Using Isobaric Tags for Relative and Absolute Quantitation (iTRAQ) Labeling

Objective

To identify differentially expressed proteins from serum of pregnant women carrying a conotruncal heart defects (CTD) fetus, using proteomic analysis.

Methods

The study was conducted using a nested case-control design. The 5473 maternal serum samples were collected at 14–18 weeks of gestation. The serum from 9 pregnant women carrying a CTD fetus, 10 with another CHD (ACHD) fetus, and 11 with a normal fetus were selected from the above samples, and analyzed by using isobaric tags for relative and absolute quantitation (iTRAQ) coupled with two-dimensional liquid chromatography-tandem mass spectrometry(2D LC-MS/MS). The differentially expressed proteins identified by iTRAQ were further validated with Western blot.

Results

A total of 105 unique proteins present in the three groups were identified, and relative expression data were obtained for 92 of them with high confidence by employing the iTRAQ-based experiments. The downregulation of gelsolin in maternal serum of fetus with CTD was further verified by Western blot.

Conclusions

The identification of differentially expressed protein gelsolin in the serum of the pregnant women carrying a CTD fetus by using proteomic technology may be able to serve as a foundation to further explore the biomarker for detection of CTD fetus from the maternal serum.     

Small-Size Circulating Endothelial Microparticles in Coronary Artery Disease

Objectives

Several recent lines of evidence indicate that endothelial microparticles are a new biomarker that can be used to monitor endothelial dysfunction in coronary artery disease (CAD). However, data concerning the detection of small microparticles (diameter <0.5 µm) are lacking. The aim of this study was to detect small-size endothelial microparticles (SEMPs) in CAD patients to monitor endothelial dysfunction.

Methods

In total, 19 CAD patients and 14 healthy subjects were recruited. The absolute numbers and percentages of CD31⁺/CD42b⁻ SEMPs and CD62E⁺ SEMPs were determined by flow cytometry. Clinical parameters were also recorded.

Results

The mean percentage of CD62E⁺ SEMPs was higher in the CAD patient group than in the healthy subject group. The area under the receiver operating characteristic curve of the percentage of CD62E⁺ SEMPs was 0.795, and the cut-off value was 1.35. There was no correlation between the percentage of CD62E⁺ SEMPs and various clinical parameters.

Conclusion

The percentage of CD62E⁺ SEMPs is a potential biomarker for monitoring endothelial function in CAD.