Nitrosative and oxidative stresses contribute to post-ischemic liver injury following severe hemorrhagic shock: the role of hypoxemic resuscitation

Purpose

Hemorrhagic shock and resuscitation is frequently associated with liver ischemia-reperfusion injury. The aim of the study was to investigate whether hypoxemic resuscitation attenuates liver injury.

Methods

Anesthetized, mechanically ventilated New Zealand white rabbits were exsanguinated to a mean arterial pressure of 30 mmHg for 60 minutes. Resuscitation under normoxemia (Normox-Res group, n = 16, PaO₂ = 95–105 mmHg) or hypoxemia (Hypox-Res group, n = 15, PaO₂ = 35–40 mmHg) followed, modifying the FiO₂. Animals not subjected to shock constituted the sham group (n = 11, PaO₂ = 95–105 mmHg). Indices of the inflammatory, oxidative and nitrosative response were measured and histopathological and immunohistochemical studies of the liver were performed.

Results

Normox-Res group animals exhibited increased serum alanine aminotransferase, tumor necrosis factor - alpha, interleukin (IL) -1β and IL-6 levels compared with Hypox-Res and sham groups. Reactive oxygen species generation, malondialdehyde formation and myeloperoxidase activity were all elevated in Normox-Res rabbits compared with Hypox-Res and sham groups. Similarly, endothelial NO synthase and inducible NO synthase mRNA expression was up-regulated and nitrotyrosine immunostaining increased in animals resuscitated normoxemically, indicating a more intense nitrosative stress. Hypox-Res animals demonstrated a less prominent histopathologic injury which was similar to sham animals.

Conclusions

Hypoxemic resuscitation prevents liver reperfusion injury through attenuation of the inflammatory response and oxidative and nitrosative stresses.     

Very low tidal volume ventilation with associated hypercapnia--effects on lung injury in a model for acute respiratory distress syndrome

Background

Ventilation using low tidal volumes with permission of hypercapnia is recommended to protect the lung in acute respiratory distress syndrome. However, the most lung protective tidal volume in association with hypercapnia is unknown. The aim of this study was to assess the effects of different tidal volumes with associated hypercapnia on lung injury and gas exchange in a model for acute respiratory distress syndrome.

Methodology/Principal Findings

In this randomized controlled experiment sixty-four surfactant-depleted rabbits were exposed to 6 hours of mechanical ventilation with the following targets: Group 1: tidal volume = 8–10 ml/kg/PaCO₂ = 40 mm Hg; Group 2: tidal volume = 4–5 ml/kg/PaCO₂ = 80 mm Hg; Group 3: tidal volume = 3–4 ml/kg/PaCO₂ = 120 mm Hg; Group 4: tidal volume = 2–3 ml/kg/PaCO₂ = 160 mm Hg. Decreased wet-dry weight ratios of the lungs, lower histological lung injury scores and higher PaO₂ were found in all low tidal volume/hypercapnia groups (group 2, 3, 4) as compared to the group with conventional tidal volume/normocapnia (group 1). The reduction of the tidal volume below 4–5 ml/kg did not enhance lung protection. However, oxygenation and lung protection were maintained at extremely low tidal volumes in association with very severe hypercapnia and no adverse hemodynamic effects were observed with this strategy.

Conclusion

Ventilation with low tidal volumes and associated hypercapnia was lung protective. A tidal volume below 4–5 ml/kg/PaCO₂ 80 mm Hg with concomitant more severe hypercapnic acidosis did not increase lung protection in this surfactant deficiency model. However, even at extremely low tidal volumes in association with severe hypercapnia lung protection and oxygenation were maintained.     

Protein phosphatase 2A mediates dormancy of glioblastoma multiforme-derived tumor stem-like cells during hypoxia

Purpose

The hypoxic microenvironment of glioblastoma multiforme (GBM) is thought to increase resistance to cancer therapies. Recent evidence suggests that hypoxia induces protein phosphatase 2A (PP2A), a regulator of cell cycle and cell death. The effects of PP2A on GBM tumor cell proliferation and survival during hypoxic conditions have not been studied.

Experimental Design

Expression of PP2A subunits and HIF-α proteins was measured in 65 high-grade astrocytoma and 18 non-neoplastic surgical brain specimens by western blotting. PP2A activity was measured by an immunoprecipitation assay. For in vitro experiments, GBM-derived tumor stem cell-like cells (TSCs) were exposed to severe hypoxia produced by either CoCl₂ or 1% O₂. PP2A activity was inhibited either by okadaic acid or by shRNA depletion of the PP2A C subunit. Effects of PP2A activity on cell cycle progression and cell survival during hypoxic conditions were assessed using flow cytometry.

Results

In our patient cohort, PP2A activity was positively correlated with HIF-1∝ protein expression (P = 0.002). Patients with PP2A activity levels above 160 pMP had significantly worse survival compared to patients with levels below this threshold (P = 0.002). PP2A activity was an independent predictor of survival on multivariable analysis (P = 0.009). In our in vitro experiments, we confirmed that severe hypoxia induces PP2A activity in TSCs 6 hours after onset of exposure. PP2A activity mediated G1/S phase growth inhibition and reduced cellular ATP consumption in hypoxic TSCs. Conversely, inhibition of PP2A activity led to increased cell proliferation, exhaustion of intracellular ATP, and accelerated P53-independent cell death of hypoxic TSCs.

Conclusions

Our results suggest that PP2A activity predicts poor survival in GBM. PP2A appears to reduce the metabolic demand of hypoxic TSCs and enhances tumor cell survival. Modulation of PP2A may be a potential target for cancer therapy.     

Periodontal tissue regeneration using fibroblast growth factor-2: randomized controlled phase II clinical trial

Background

The options for medical use of signaling molecules as stimulators of tissue regeneration are currently limited. Preclinical evidence suggests that fibroblast growth factor (FGF)-2 can promote periodontal regeneration. This study aimed to clarify the activity of FGF-2 in stimulating regeneration of periodontal tissue lost by periodontitis and to evaluate the safety of such stimulation.

Methodology/Principal Findings

We used recombinant human FGF-2 with 3% hydroxypropylcellulose (HPC) as vehicle and conducted a randomized double-blinded controlled trial involving 13 facilities. Subjects comprised 74 patients displaying a 2- or 3-walled vertical bone defect as measured ≥3 mm apical to the bone crest. Patients were randomly assigned to 4 groups: Group P, given HPC with no FGF-2; Group L, given HPC containing 0.03% FGF-2; Group M, given HPC containing 0.1% FGF-2; and Group H, given HPC containing 0.3% FGF-2. Each patient underwent flap operation during which we administered 200 µL of the appropriate investigational drug to the bone defect. Before and for 36 weeks following administration, patients underwent periodontal tissue inspections and standardized radiography of the region under investigation. As a result, a significant difference (p = 0.021) in rate of increase in alveolar bone height was identified between Group P (23.92%) and Group H (58.62%) at 36 weeks. The linear increase in alveolar bone height at 36 weeks in Group P and H was 0.95 mm and 1.85 mm, respectively (p = 0.132). No serious adverse events attributable to the investigational drug were identified.

Conclusions

Although no statistically significant differences were noted for gains in clinical attachment level and alveolar bone gain for FGF-2 groups versus Group P, the significant difference in rate of increase in alveolar bone height (p = 0.021) between Groups P and H at 36 weeks suggests that some efficacy could be expected from FGF-2 in stimulating regeneration of periodontal tissue in patients with periodontitis.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00514657","term_id":"NCT00514657"}}NCT00514657     

Profiling of VEGFs and VEGFRs as prognostic factors in soft tissue sarcoma: VEGFR-3 is an independent predictor of poor prognosis

Background

In non-gastrointestinal stromal tumor soft tissue sarcoma (non-GIST STS) optimal treatment is surgery with wide resection margins. Vascular endothelial growth factors (VEGFs) and receptors (VEGFRs) are known to be key players in the initiation of angiogenesis and lymphangiogenesis. This study investigates the prognostic impact of VEGFs and VEGFRs in non-GIST STS with wide and non-wide resection margins.

Methods

Tumor samples from 249 patients with non-GIST STS were obtained and tissue microarrays were constructed for each specimen. Immunohistochemistry was used to evaluate the expressions of VEGF-A, -C and -D and VEGFR-1, -2 and -3.

Results

In the univariate analyses, VEGF-A (P = 0.040) in the total material, and VEGF-A (P = 0.018), VEGF-C (P = 0.025) and VEGFR-3 (P = 0.027) in the subgroup with wide resection margins, were significant negative prognostic indicators of disease-specific survival (DSS). In the multivariate analysis, high expression of VEGFR-3 (P = 0.042, HR = 1.907, 95% CI 1.024-3.549) was an independent significant negative prognostic marker for DSS among patients with wide resection margins.

Conclusion

VEGFR-3 is a strong and independent negative prognostic marker for non-GIST STSs with wide resection margins.     

Prognostic impacts of angiopoietins in NSCLC tumor cells and stroma: VEGF-A impact is strongly associated with Ang-2

Introduction

Angiopoietins and their receptor Tie-2 are, in concert with VEGF-A, key mediators in angiogenesis. This study evaluates the prognostic impact of all known human angiopoietins (Ang-1, Ang-2 and Ang-4) and their receptor Tie-2, as well as their relation to the prognostic expression of VEGF-A.

Methods

335 unselected stage I-IIIA NSCLC-patients were included and tissue samples of respective tumor cells and stroma were collected in tissue microarrays (TMAs). Immunohistochemistry (IHC) was used to semiquantitatively evaluate the expression of markers in duplicate tumor and stroma cores.

Principal Findings

In univariate analyses, low tumor cell expression of Ang-4 (P = 0.046) and low stromal expressions of Ang-4 (P = 0.009) and Ang-2 (P = 0.017) were individually associated with a poor survival. In the multivariate analysis, low stromal Ang-2 (HR 1.88; CI 95% 1.15-3.08) and Ang-4 (HR 1.47, CI 95% 1.02–2.11, P = 0.04) expressions were independently associated with a poor prognosis. In patients with high tumor cell expression of Ang-2, a concomitantly high tumor VEGF-A expression mediated a dramatic survival reduction (P<0.001). In the multivariate analysis of patients with high Ang-2 expression, high tumor VEGF-A expression appeared an independent poor prognosticator (HR 6.43; CI 95% 2.46–16.8; P<0.001).

Conclusions

In tumor cells, only Ang-4 expression has prognostic impact in NSCLC. In tumor stroma, Ang-4 and Ang-2 are independently associated with survival. The prognostic impact of tumor cell VEGF-A in NSCLC appears strongly associated with a concomitantly high tumor cell expression of Ang-2.     

Overexpression of the HIF hydroxylases PHD1, PHD2, PHD3 and FIH are individually and collectively unfavorable prognosticators for NSCLC survival

Introduction

Hypoxia induced factors (HIFs) are at the heart of the adaptive mechanisms cancer cells must implement for survival. HIFs are regulated by four hydroxylases; Prolyl hydroxylase (PHD)-1,-2,-3 and factor inhibiting HIF (FIH). We aimed to investigate the prognostic impact of these oxygen sensors in NSCLC.

Methods

Tumor tissue samples from 335 resected stages I to IIIA NSCLC patients was obtained and tissue microarrays (TMAs) were constructed. Hydroxylase expression was evaluated by immunohistochemistry.

Principal Findings

There was scorable expression for all HIF hydroxylases in tumor cells, but not in stroma. In univariate analyses, high tumor cell expression of all the HIF hydroxylases were unfavorable prognosticators for disease-specific survival (DSS); PHD1 (P = 0.023), PHD2 (P = 0.013), PHD3 (P = 0.018) and FIH (P = 0.033). In the multivariate analyses we found high tumor cell expression of PHD2 (HR = 2.03, CI 95% 1.20–3.42, P = 0.008) and PHD1 (HR = 1.45, CI 95% 1.01–2.10, P = 0.047) to be significant independent prognosticators for DSS. Besides, there was an additive prognostic effect by the increasing number of highly expressed HIF hydroxylases. Provided none high expression HIF hydroxylases, the 5-year survival was 80% vs. 23% if all four were highly expressed (HR = 6.48, CI 95% 2.23–18.8, P = 0.001).

Conclusions

HIF hydroxylases are, in general, poor prognosticators for NSCLC survival. PHD1 and PHD2 are independent negative prognostic factors in NSCLC. Moreover, there is an additive poor prognostic impact by an increasing number of highly expressed HIF hydroxylases.     

Near Infrared Imaging of EGFR of Oral Squamous Cell Carcinoma in Mice Administered Arsenic Trioxide

Background

The effectiveness of near-infrared imaging (NIR) interrogation of epidermal growth factor receptor (EGFR) expression as a sensitive biomarker of oral squamous cell carcinoma (OSCC) response to arsenic trioxide therapy was studied in mice.

Material and Methods

A431 OSCC in vitro were exposed to 0 µM, 0.5 µM, 2.5 µM, or 5 µM of As₂O₃ for 0 h, 24 h, 48 h and 72 h. Confocal microscopy and flow cytometry confirmed EGFR expression and demonstrated a sensitivity dose-related signal decline with As₂O₃ treatment. Next, mice with pharynx-implanted A431 cells received As₂O₃ i.p. every 48 h at 0.0, 0.5, 2.5, or 5 mg/kg/day (n = 6/group) from day 0 to 10. An intravenous NIR probe, EGF-Cy5.5, was injected at baseline and on days 4, 8, and 12 for dynamic NIR imaging. Tumor volume and body weights were measured three times weekly.

Results

In vitro, A431 EGFR expression was well appreciated in the controls and decreased (p<0.05) with increasing As₂O₃ dose and treatment duration. In vivo EGFR NIR tumor signal intensity decreased (p<0.05) in As₂O₃ treated groups versus controls from days 4 to 12, consistent with increasing dosage. Tumor volume diminished in a dose-related manner while body weight was unaffected. Immunohistochemical staining of excised tumors confirmed that EGFR expression was reduced by As₂O₃ treatment in a dose responsive pattern.

Conclusion

This study demonstrates for the first time that OSCC can be interrogated in vivo by NIR molecular imaging of the EGFR and that this biomarker is effective for the longitudinal assessment of OSCC response to As₂O₃ treatment.     

The prognostic impact of TGF-β1, fascin, NF-κB and PKC-ζ expression in soft tissue sarcomas

Aims

Transforming growth factor-β (TGF-β), fascin, nuclear factor-kappa B (NF-κB) p105, protein-kinase C-zeta (PKC-ζ), partioning-defective protein-6 (Par-6), E-cadherin and vimentin are tumor promoting molecules through mechanisms involved in cell dedifferentiation. In soft tissue sarcomas, their expression profile is poorly defined and their significance is uncertain. We aimed to investigate the prognostic impact of TGF-β1, NF-κB p105, PKC-ζ, Par-6α, E-cadherin and vimentin in non-gastrointestinal stromal tumor soft tissue sarcomas (non-GIST STSs).

Patients and Methods

Tumor samples and clinical data from 249 patients with non-GIST STS were obtained, and tissue microarrays (TMAs) were constructed for each specimen. Immunohistochemistry (IHC) was used to evaluate marker expression in tumor cells.

Results

In univariate analysis, the expression levels of TGF-β1 (P = 0.016), fascin (P = 0.006), NF-κB p105 (P = 0.022) and PKC-ζ, (P = 0.042) were significant indicators for disease specific survival (DSS). In the multivariate analysis, high TGF-β1 expression was an independent negative prognostic factor for DSS (HR = 1.6, 95% CI = 1.1–2.4, P = 0.019) in addition to tumor depth, malignancy grade, metastasis at diagnosis, surgery and positive resection margins.

Conclusion

Expression of TGF-β1 was significantly associated with aggressive behavior and shorter DSS in non-GIST STSs.     

Physiological correlates of endurance time variability during constant-workrate cycling exercise in patients with COPD

Rationale

The endurance time (T_end) during constant-workrate cycling exercise (CET) is highly variable in COPD. We investigated pulmonary and physiological variables that may contribute to these variations in T_end.

Methods

Ninety-two patients with COPD completed a CET performed at 80% of peak workrate capacity (W_peak). Patients were divided into tertiles of T_end [Group 1: <4 min; Group 2: 4–6 min; Group 3: >6 min]. Disease severity (FEV₁), aerobic fitness (W_peak, peak oxygen consumption [ _peak], ventilatory threshold [ _VT]), quadriceps strength (MVC), symptom scores at the end of CET and exercise intensity during CET (heart rate at the end of CET to heart rate at peak incremental exercise ratio [HR_CET/HR_peak]) were analyzed as potential variables influencing T_end.

Results

W_peak, _peak, _VT, MVC, leg fatigue at end of CET, and HR_CET/HR_peak were lower in group 1 than in group 2 or 3 (p≤0.05). _VT and leg fatigue at end of CET independently predicted T_end in multiple regression analysis (r = 0.50, p = 0.001).

Conclusion

T_end was independently related to the aerobic fitness and to tolerance to leg fatigue at the end of exercise. A large fraction of the variability in T_end was not explained by the physiological parameters assessed in the present study. Individualization of exercise intensity during CET should help in reducing variations in T_end among patients with COPD.     

Absence of association between N-acetyltransferase 2 acetylator status and colorectal cancer susceptibility: based on evidence from 40 studies

Background and Objectives

N-Acetyltransferase (NAT) 2 is an important enzyme involved in the metabolism of different xenobiotics, including potential carcinogens, whose phenotypes were reported to be related to individual susceptibility to colorectal cancer (CRC). However, the results remain conflicting. To assess the relationship between NAT2 phenotypes and CRC risk, we performed this meta-analysis.

Methods

A comprehensive literature search was conducted to identify all case-control or cohort studies of NAT2 acetylator status on the susceptibility of CRC by searching of PubMed and EMBASE, up to May 20, 2011. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were used to assess the association.

Results

A total of over 40,000 subjects from 40 published literatures were identified by searching the databases. No significantly elevated CRC risk in individuals with NAT2 slow acetylators compared with fast acetylators was found when all studies pooled (OR = 0.95, 95% CI: 0.87–1.04, I² = 52.6%). While three studies contributed to the source of heterogeneity were removed, there was still null result observed (OR = 0.96, 95% CI: 0.90–1.03, P = 0.17 for heterogeneity, I² = 17.8%). In addition, we failed to detect any associations in the stratified analyses by race, sex, source of controls, smoking status, genotyping methods or tumor localization. No publication bias was observed in this study.

Conclusions

This meta-analysis suggests that the NAT2 phenotypes may not be associated with colorectal cancer development.     

The Association between Histamine 2 Receptor Antagonist Use and Clostridium difficile Infection: A Systematic Review and Meta-analysis

Background

Clostridium difficile infection (CDI) is a major health problem. Epidemiological evidence suggests that there is an association between acid suppression therapy and development of CDI.

Purpose

We sought to systematically review the literature that examined the association between histamine 2 receptor antagonists (H₂RAs) and CDI.

Data source

We searched Medline, Current Contents, Embase, ISI Web of Science and Elsevier Scopus from 1990 to 2012 for all analytical studies that examined the association between H₂RAs and CDI.

Study selection

Two authors independently reviewed the studies for eligibility.

Data extraction

Data about studies characteristics, adjusted effect estimates and quality were extracted.

Data synthesis

Thirty-five observations from 33 eligible studies that included 201834 participants were analyzed. Studies were performed in 6 countries and nine of them were multicenter. Most studies did not specify the type or duration of H₂RAs therapy. The pooled effect estimate was 1.44, 95% CI (1.22–1.7), I² = 70.5%. This association was consistent across different subgroups (by study design and country) and there was no evidence of publication bias. The pooled effect estimate for high quality studies was 1.39 (1.15–1.68), I2 = 72.3%. Meta-regression analysis of 10 study-level variables did not identify sources of heterogeneity. In a speculative analysis, the number needed to harm (NNH) with H₂RAs at 14 days after hospital admission in patients receiving antibiotics or not was 58, 95% CI (37, 115) and 425, 95% CI (267, 848), respectively. For the general population, the NNH at 1 year was 4549, 95% CI (2860, 9097).

Conclusion

In this rigorous systematic review and meta-analysis, we observed an association between H₂RAs and CDI. The absolute risk of CDI associated with H₂RAs is highest in hospitalized patients receiving antibiotics.     

Bioluminescent orthotopic mouse models of human localized non-small cell lung cancer: feasibility and identification of circulating tumour cells

Background

Preclinical models of non-small cell lung cancer (NSCLC) require better clinical relevance to study disease mechanisms and innovative therapeutics. We sought to compare and refine bioluminescent orthotopic mouse models of human localized NSCLC.

Methods

Athymic nude mice underwent subcutaneous injection (group 1-SC, n = 15, control), percutaneous orthotopic injection (group 2-POI, n = 30), surgical orthotopic implantation of subcutaneously grown tumours (group 3-SOI, n = 25), or transpleural orthotopic injection (group 4-TOI, n = 30) of A549-luciferase cells. Bioluminescent in vivo imaging was then performed weekly. Circulating tumour cells (CTCs) were searched using Cellsearch® system in SC and TOI models.

Results

Group 2-POI was associated with unexpected direct pleural spreading of the cellular solution in 53% of the cases, forbidding further evaluation of any localized lung tumour. Group 3-SOI was characterized by high perioperative mortality, initially localized lung tumours, and local evolution. Group 4-TOI was associated with low perioperative mortality, initially localized lung tumours, loco regional extension, and distant metastasis. CTCs were detected in 83% of nude mice bearing subcutaneous or orthotopic NSCLC tumours.

Conclusions

Transpleural orthotopic injection of A549-luc cells in nude mouse lung induces localized tumour, followed by lymphatic extension and specific mortality, and allowed the first time identification of CTCs in a NSCLC mice model.     

The dual PI3K/mTOR inhibitor NVP-BEZ235 induces tumor regression in a genetically engineered mouse model of PIK3CA wild-type colorectal cancer

Purpose

To examine the in vitro and in vivo efficacy of the dual PI3K/mTOR inhibitor NVP-BEZ235 in treatment of PIK3CA wild-type colorectal cancer (CRC).

Experimental Design

PIK3CA mutant and wild-type human CRC cell lines were treated in vitro with NVP-BEZ235, and the resulting effects on proliferation, apoptosis, and signaling were assessed. Colonic tumors from a genetically engineered mouse (GEM) model for sporadic wild-type PIK3CA CRC were treated in vivo with NVP-BEZ235. The resulting effects on macroscopic tumor growth/regression, proliferation, apoptosis, angiogenesis, and signaling were examined.

Results

In vitro treatment of CRC cell lines with NVP-BEZ235 resulted in transient PI3K blockade, sustained decreases in mTORC1/mTORC2 signaling, and a corresponding decrease in cell viability (median IC₅₀ = 9.0–14.3 nM). Similar effects were seen in paired isogenic CRC cell lines that differed only in the presence or absence of an activating PIK3CA mutant allele. In vivo treatment of colonic tumor-bearing mice with NVP-BEZ235 resulted in transient PI3K inhibition and sustained blockade of mTORC1/mTORC2 signaling. Longitudinal tumor surveillance by optical colonoscopy demonstrated a 97% increase in tumor size in control mice (p = 0.01) vs. a 43% decrease (p = 0.008) in treated mice. Ex vivo analysis of the NVP-BEZ235-treated tumors demonstrated a 56% decrease in proliferation (p = 0.003), no effects on apoptosis, and a 75% reduction in angiogenesis (p = 0.013).

Conclusions

These studies provide the preclinical rationale for studies examining the efficacy of the dual PI3K/mTOR inhibitor NVP-BEZ235 in treatment of PIK3CA wild-type CRC.     

Transketolase-like 1 expression is modulated during colorectal cancer progression and metastasis formation

Background

Transketolase-like 1 (TKTL1) induces glucose degradation through anaerobic pathways, even in presence of oxygen, favoring the malignant aerobic glycolytic phenotype characteristic of tumor cells. As TKTL1 appears to be a valid biomarker for cancer prognosis, the aim of the current study was to correlate its expression with tumor stage, probability of tumor recurrence and survival, in a series of colorectal cancer patients.

Methodolody/Principal Findings

Tumor tissues from 63 patients diagnosed with colorectal cancer at different stages of progression were analyzed for TKTL1 by immunohistochemistry. Staining was quantified by computational image analysis, and correlations between enzyme expression, local growth, lymph-node involvement and metastasis were assessed. The highest values for TKTL1 expression were detected in the group of stage III tumors, which showed significant differences from the other groups (Kruskal-Wallis test, P = 0.000008). Deeper analyses of T, N and M classifications revealed a weak correlation between local tumor growth and enzyme expression (Mann-Whitney test, P = 0.029), a significant association of the enzyme expression with lymph-node involvement (Mann-Whitney test, P = 0.0014) and a significant decrease in TKTL1 expression associated with metastasis (Mann-Whitney test, P = 0.0004).

Conclusions/Significance

To our knowledge, few studies have explored the association between variations in TKTL1 expression in the primary tumor and metastasis formation. Here we report downregulation of enzyme expression when metastasis appears, and a correlation between enzyme expression and regional lymph-node involvement in colon cancer. This finding may improve our understanding of metastasis and lead to new and more efficient therapies against cancer.     

A preliminary evaluation of limb salvage surgery for osteosarcoma around knee joint

Objective

To evaluate the effectiveness and drawbacks of diversified procedures of limb salvage surgery (LSS), providing a reference of rational surgical criterion of LSS.

Methods

Fifty eight patients with stage IIB extremity osteosarcoma around knee joint area between 1992 and 2002 were studied retrospectively. Among them, 43 patients were treated by LSS followed by reconstruction. Reconstruction approaches included re-implantation of irradiation-devitalized tumor bone (n = 12), autoclaving-devitalized tumor bone (n = 8), prosthetic replacement (n = 11), allograft transplantation (n = 8) and vascularized fibula autograft implantation (n = 4). Amputations were performed in 15 patients. Patients were followed up for 6–16 years.

Results

There were no significant difference between LSS and amputation groups regarding disease free survival and local recurrence rates. The actuarial 5-year continuous disease free survival and local recurrence rate were 30.0% and 25.0% in patients of devitalized LSS group, whereas those were 56.5% and 8.7% in patients of non-devitalized reconstruction group. The complication rate was significantly higher in LSS group compared to amputation group (P = 0.003).

Conclusion

LSS with non-devitalized procedures is the optimal treatment for osteosarcoma around knee joint area. Prosthesis implantation is the preferred option for bone reconstruction following LSS. Prevention and treatment of post-operative complications should be paid more attention to get good long-term outcomes of surgery.     

Bronchial responsiveness is related to increased exhaled NO (FE(NO)) in non-smokers and decreased FE(NO) in smokers

Rationale

Both atopy and smoking are known to be associated with increased bronchial responsiveness. Fraction of nitric oxide (NO) in the exhaled air (FE_NO), a marker of airways inflammation, is decreased by smoking and increased by atopy. NO has also a physiological bronchodilating and bronchoprotective role.

Objectives

To investigate how the relation between FE_NO and bronchial responsiveness is modulated by atopy and smoking habits.

Methods

Exhaled NO measurements and methacholine challenge were performed in 468 subjects from the random sample of three European Community Respiratory Health Survey II centers: Turin (Italy), Gothenburg and Uppsala (both Sweden). Atopy status was defined by using specific IgE measurements while smoking status was questionnaire-assessed.

Main Results

Increased bronchial responsiveness was associated with increased FE_NO levels in non-smokers (p = 0.02) and decreased FE_NO levels in current smokers (p = 0.03). The negative association between bronchial responsiveness and FE_NO was seen only in the group smoking less <10 cigarettes/day (p = 0.008). Increased bronchial responsiveness was associated with increased FE_NO in atopic subjects (p = 0.04) while no significant association was found in non-atopic participants. The reported interaction between FE_NO and smoking and atopy, respectively were maintained after adjusting for possible confounders (p-values<0.05).

Conclusions

The present study highlights the interactions of the relationship between FE_NO and bronchial responsiveness with smoking and atopy, suggesting different mechanisms behind atopy- and smoking-related increases of bronchial responsiveness.     

Genetic variations in plasma circulating DNA of HBV-related hepatocellular carcinoma patients predict recurrence after liver transplantation

Background

Recurrence prediction of hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) patients undergoing liver transplantation (LT) present a great challenge because of a lack of biomarkers. Genetic variations play an important role in tumor development and metastasis.

Methods

Oligonucleotide microarrays were used to evaluate the genetic characteristics of tumor DNA in 30 HBV-related HCC patients who were underwent LT. Recurrence-related single-nucleotide polymorphism were selected, and their prognostic value was assessed and validated in two independent cohorts of HCC patients (N = 102 and N = 77), using pretransplant plasma circulating DNA. Prognostic significance was assessed by Kaplan-Meier survival estimates and log-rank tests. Multivariate analyses were performed to evaluate prognosis-related factors.

Results

rs894151 and rs12438080 were significantly associated with recurrence (P = .003 and P = .004, respectively). Multivariate analyses demonstrated that the co-index of the 2 SNPs was an independent prognostic factor for recurrence (P = .040). Similar results were obtained in the third cohort (N = 77). Furthermore, for HCC patients (all the 3 cohorts) exceeding Milan criteria, the co-index was a prognostic factor for recurrence and survival (P<.001 and P = .002, respectively).

Conclusions

Our study demonstrated first that genetic variations of rs894151 and rs12438080 in pretransplant plasma circulating DNA are promising prognostic markers for tumor recurrence in HCC patients undergoing LT and identify a subgroup of patients who, despite having HCC exceeding Milan criteria, have a low risk of post-transplant recurrence.     

Tumor site immune markers associated with risk for subsequent basal cell carcinomas

Background

Basal cell carcinoma (BCC) tumors are the most common skin cancer and are highly immunogenic.

Objective

The goal of this study was to assess how immune-cell related gene expression in an initial BCC tumor biopsy was related to the appearance of subsequent BCC tumors.

Materials and Methods

Levels of mRNA for CD3ε (a T-cell receptor marker), CD25 (the alpha chain of the interleukin (IL)-2 receptor expressed on activated T-cells and B-cells), CD68 (a marker for monocytes/macrophages), the cell surface glycoprotein intercellular adhesion molecule-1 (ICAM-1), the cytokine interferon-γ (IFN-γ) and the anti-inflammatory cytokine IL-10 were measured in BCC tumor biopsies from 138 patients using real-time PCR.

Results

The median follow-up was 26.6 months, and 61% of subjects were free of new BCCs two years post-initial biopsy. Patients with low CD3ε CD25, CD68, and ICAM-1 mRNA levels had significantly shorter times before new tumors were detected (p = 0.03, p = 0.02, p = 0.003, and p = 0.08, respectively). Furthermore, older age diminished the association of mRNA levels with the appearance of subsequent tumors.

Conclusions

Our results show that levels of CD3ε, CD25, CD68, and ICAM-1 mRNA in BCC biopsies may predict risk for new BCC tumors.