Detection of superoxide radical in all biological systems by Thin Layer Chromatography.

The study presents a new method that detects O2•-, via quantification of 2-hydroxyethidium (2-ΟΗ-Ε+) as low as ∼30 fmoles by High-Performance Thin Layer Chromatography (HPTLC). The method isolates 2-ΟΗ-Ε+ after its extraction by the anionic detergent SDS (at 18-fold higher than its CMC) together with certain organic/inorganic reagents, and its HPTLC-separation from di-ethidium (di-Ε+) and ethidium (Ε+). Quantification of 2-OH-E+ is based on its ex/em maxima at 290/540 nm, and of di-E+ and E+ at 295/545 nm. The major innovations of the present method are the development of protocols for (i) efficient extraction (by SDS) and (ii) sensitive quantification (by HPTLC) for 2-OH-E+ (as well as di-E+ and E+) from most biological systems (animals, plants, cells, subcellular compartments, fluids). The method extracts 2-ΟΗ-Ε+ (by neutralizing the strong binding between its quaternary N+ and negatively charged sites on phospholipids, DNA etc) together with free HE, while protects both from biological oxidases, and also extracts/quantifies total proteins (hydrophilic and hydrophobic) for expressing O2•- levels per protein quantity. The method also uses SDS (at 80-fold lower than its CMC) to extract/remove/wash 2-ΟΗ-Ε+ from cell/organelle exterior membrane sites, for more accurate internal content quantification. The new method is applied on indicative biological systems: (1) artificially stressed (mouse organs and liver mitochondria and nuclei, ±exposed to paraquat, a known O2•- generator), and (2) physiologically stressed (cauliflower plant, exposed to light/dark).

Effect of rutin on anxiety-like behavior and activity of acetylcholinesterase isoforms in specific brain regions of pentylenetetrazol-treated mice.

The aim of the present study was to investigate the effect of rutin administration (100 mg/kg/day) to pentylenetetrazol (PTZ)-treated Balb-c mice (60 mg/kg/day), with respect to anxiety-like behavior using both open-field and elevated plus-maze (EPM) tests, and acetylcholinesterase (AChE) activity in salt-soluble (SS) fraction and detergent-soluble (DS) fraction of the cerebral cortex, hippocampus, striatum, midbrain, and diencephalon. Our results demonstrated that the administration of PTZ in 3 doses and the induction of seizures increased significantly anxiety behavior of mice and reduced significantly DS-AChE activity in all brain regions examined, while the reduction in the SS fraction was brain region-specific. Rutin administration to normal mice did not affect their behavior, while it induced a brain region-specific reduction in SS-AChE and a significant decrease in DS-AChE in all brain regions. We demonstrated for the first time that pretreatment of PTZ-mice with rutin (PTZ + Rutin group) prevented the manifestation of anxiety and induced interestingly a further significant reduction on the SS- and DS-AChE activities only in the cerebral cortex and striatum, in comparison with PTZ group. Our results show that rutin exhibits an important anxiolytic effect and an anticholinesterase activity in specific brain areas in the seizure model of PTZ.

In vivo quantification and pharmacokinetic studies of cotinine in mice after smoke exposure by LC-MS/MS.

A sensitive analytical method was developed and validated for the quantification of cotinine in mouse plasma after exposure to smoke of 0.5, 1.0, and 1.5 commercially available cigarettes, using liquid chromatography tandem mass spectrometry. The method was validated over a linear concentration range of 0.075-20.0 ng/mL with the R2 value being higher than 0.99. Both the precision (coefficient of variation; %) and accuracy (relative error; %) were within acceptable criteria of <15%. The lower limit of quantification (LLOQ) for cotinine was 0.075 ng/mL with sufficient specificity, accuracy, and precision. Following exposure to 0.5, 1.0, and 1.5 cigarette smoke, it was observed that the AUC and the Cmax increased linearly as the doses increased. The pharmacokinetics of cotinine was found linear for the range of 0.5-1.5 commercial cigarette smoke. The quantification of the concentration of cotinine in mouse plasma after smoke exposure will facilitate future behavioral and toxicological experiments in animals and may prove useful in predicting cotinine levels in humans during smoking.

Assessment of the long-term outcome of TVT procedure for stress urinary incontinence in a female population: results at 17 years' follow-up.

INTRODUCTION AND HYPOTHESIS: To assess the outcome of the tension-free vaginal tape (TVT) procedure in female patients with urodynamic stress urinary incontinence at 17 years follow-up. METHODS: We carried out a prospective study at the 2nd Department of Obstetrics and Gynecology, Aretaieio Hospital, University of Athens, Greece. Patients who had undergone a TVT procedure 17 years ago. The follow-up assessment included gynecological examination, urinalysis, cough stress test in the lithotomy and/or upright position, filling and voiding cystometry, and uroflow. Also, all patients were required to complete the Patient Satisfaction Questionnaire (PSQ). RESULTS: Out of the 61 initial patients, 56 were available for follow-up. Objective cure rate was 83.9% (47/56) at 17 years follow-up. Subjective cure rate was 78.6% (44/56), subjective improvement was 8.9%, and failure rate was 12.5%. Frequency was present in 39.3% of patients, overactive bladder symptoms were present in 30.3% of patients and urge urinary incontinence was reported by 12.5% of patients. Difficulty emptying the bladder was reported by 10 patients (17.8%) and recurrent urinary tract infection was seen in 3.5% of patients. There was one case of TVT erosion to the vaginal mucosa, which was managed conservatively. CONCLUSIONS: The TVT procedure for the management of stress urinary incontinence in women maintains its efficacy in the long term, having an objective cure rate of 83.9% and a subjective cure rate of 78.6% at 17 years' follow-up, with a very low complications rate.

Are Extensive Open Lung Resections for Elderly Patients with Lung Cancer Justified?

BACKGROUND: Older patients with malignancies are more comorbid than younger ones and are usually undertreated only because of their age. The aim of this study is to investigate the safety of open anatomical lung resections for lung cancer in elderly patients. METHODS: We retrospectively analyzed all patients who underwent lung resection for lung cancer in our institution and categorized them into two groups: the elderly group (≥70 years old) and the control (<70). RESULTS: In total, 135 patients were included in the elderly group and 375 in the control. Elderly patients were more frequently diagnosed with squamous cell carcinoma (59.3% vs. 51.5%, p = 0.037), higher differentiated tumors (12.6% vs. 6.4%, p = 0.014), and at an earlier stage (stage I: 55.6% for elderly vs. 36.6%, p = 0.002). Elderly patients were more vulnerable to postoperative pneumonia (3.7% vs. 0.8%, p = 0.034), lung atelectasis (7.4% vs. 2.9%, p = 0.040), and pleural empyema (3.2% vs. 0%, p = 0.042), however, with no increased 30-day-mortality (5.2% for elderly vs. 2.7%, p = 0.168). Survival was comparable in both groups (43.4 vs. 45.3 months, p = 0.579). CONCLUSIONS: Elderly patients should not be excluded from open major lung resections as the survival benefit is not reduced in selected patients.

Genetic Variations of CD40 and LTßR Genes Are Associated With Increased Susceptibility and Clinical Outcome of Non-Small-Cell Carcinoma Patients.

BACKGROUND: Immune system-related receptors CD40 (tumor necrosis factor receptor superfamily member 5), BAFFR (tumor necrosis factor receptor superfamily member 13C), and LTßR (tumor necrosis factor receptor superfamily member 3) play a pivotal role in non-small-cell lung cancer (NSCLC). To further evaluate their role in NSCLC, CD40 rs1883832 (T>C), BAFFR rs7290134 (A>G), and LTßR rs10849448 (A>G) single-nucleotide polymorphisms (SNPs) were investigated regarding their impact in risk and clinical outcome of NSCLC patients. METHODS: The three selected SNPs were evaluated in 229 NSCLC patients and 299 healthy controls, while CD40, BAFFR, and LTßR protein expression was assessed by immunohistochemistry in 96 tumor specimens from NSCLC patients. RESULTS: In total, CD40 rs1883832 was associated with NSCLC risk, with the T allele, after adjusting for cofactors, being related to increased risk (p = 0.007; OR 1.701). Moreover, the CT genotype was associated with increased risk (p = 0.024; OR 1.606) and poorer 5-year overall survival (OS) after adjusting for cofactors (p = 0.001, HR 1.829), while CC was associated with higher CD40 expression in tumorous cells (p = 0.040) and in stromal cells (p = 0.036). In addition, AA homozygotes for the LTßR rs10849448 had increased risk for NSCLC in multivariate analysis (p = 0.008; OR, 2.106) and higher LTßR membranous expression (p = 0.035). Although BAFFR rs7290134 was associated with BAFFR membranous expression (p = 0.039), BAFFR rs7290134 was not associated with neither the disease risk nor the prognosis of NSCLC patients. CONCLUSIONS: In conclusion, CD40 rs1883832 and LTßR rs10849448 seem to be associated with increased risk for NSCLC, while CD40 rs1883832 is also associated with OS of patients with NSCLC.

High expression of BCL-2 predicts favorable outcome in non-small cell lung cancer patients with non squamous histology.

BACKGROUND: Bcl-2 promotes cell survival by inhibiting adapters needed for the activation and cleavage of caspases thus blocking the proteolytic cascade that ultimately dismantles the cell. Bcl-2 has been investigated as a prognostic factor in non small cell lung cancer (NSCLC) patients with conflicting results. METHODS: Here, we quantitatively assessed Bcl-2 expression in two large and independent cohorts to investigate the impact of Bcl-2 on survival. AQUA(R), a fluorescent-based method for analysis of in situ protein expression, was used to measure Bcl-2 protein levels and classify tumors by Bcl-2 expression in a cohort of 180 NSCLC patients. An independent cohort of 354 NSCLC patients was used to validate Bcl-2 classification and evaluate outcome. RESULTS: Fifty % and 52% of the cases were classified as high expressers in training and validation cohorts respectively. Squamous cell carcinomas were more likely to be high expressers compared to adenocarcinomas (63% vs. 45%, p = 0.002); Bcl-2 was not associated with other clinical or pathological characteristics. Survival analysis showed that patients with high BCL-2 expression had a longer median survival compared to low expressers (22 vs. 17.5 months, log rank p = 0.014) especially in the subset of non-squamous tumors (25 vs. 13.8 months, log rank p = 0.04). Multivariate analysis revealed an independent lower risk for all patients with Bcl-2 expressing tumors (HR = 0.53, 95% CI 0.37-0.75, p = 0.0003) and for patients with non-squamous tumors (HR = 0.5, 95% CI 0.31-0.81, p = 0.005). CONCLUSIONS: Bcl-2 expression defines a subgroup of patients with a favorable outcome and may be useful for prognostic stratification of NSCLC patients.

Histologic-type specific role of cell cycle regulators in non-small cell lung carcinoma.

BACKGROUND: Lung cancer is the most lethal type of cancer in humans. Cell cycle alterations have commonly been encountered in lung cancer and may have prognostic value. MATERIALS AND METHODS: This study investigates the immunohistochemical expression of the important cell cycle regulators phosphatase and tensin homolog deleted on chromosome 10 (PTEN), p27, Cks1, and Skp2 in 128 non-small cell lung carcinomas (64 adenocarcinomas, 46 squamous cell carcinomas, and 18 large cell undifferentiated carcinomas) and adjacent non-neoplastic lung tissue. RESULTS: PTEN and p27 were always highly expressed in non-neoplastic lung whereas Cks1 and Skp2 were not expressed in normal tissue. Decreased PTEN expression was noted in 19/64 adenocarcinomas, 15/46 squamous cell carcinomas, and 7/18 undifferentiated large cell carcinomas. Reduced expression of p27 was noted in 28/64, 19/46, and 6/18 of the tumors, respectively. Increased expression of Cks1 was seen in 38/64, 26/46, and 11/18 and increased expression of Skp2 in 29/64, 30/46, and 14/18 of the tumors, respectively. An inverse relationship between p27 and Skp2 levels was found in adenocarcinomas and between p27 and Cks1 levels in squamous cell carcinomas. Decreased PTEN and p27 expression were associated with advanced tumor stage in squamous cell carcinomas. Univariate analysis showed that high p27 and PTEN and low Cks1 expression correlated with increased survival in patients with squamous cell carcinoma independently of tumor stage. CONCLUSIONS: Aberrant expression of PTEN, p27, Cks1, and Skp2 is a common feature of all three major types of non-small cell lung cancer NSCLC, but seems to be involved in the progression of squamous cell carcinoma alone.

Vulvo-Perineal and Perianal Paget Disease. Radical Excision and Reconstruction with Singapore Flap.

Extramammary Paget disease (EMPD) is an uncommon intraepithelial malignancy, affecting the vulvo-perineal and perianal region, occurring in 6.5% of all Paget diseases. Usually, an underlying invasive adenocarcinoma denotes a more aggressive behaviour of the disease. We present the multidisciplinary approach in a 75-year old patient with this rare disease. The patient underwent a radical surgical excision and, subsequently, a Singapore flap was used for primary closure. The final histology confirmed the presence of a non-invasive Paget tumor, but a focus of high-grade invasive adenocarcinoma was noted in a perineal nodule. The histological margins were free of tumor. The patient did not undergo any adjuvant treatment because of severe chronic medical problems, although, eighteen months after treatment, she remains well, with no signs of recurrence. In conclusion, radical surgical excision, often necessitating reconstruction techniques, remains the gold standard of care and further adjuvant treatment should be individualised.

Sclerosing Mediastinitis Causing Unilateral Pulmonary Edema Due to Left Atrial and Pulmonary Venous Compression. A Case Report and Literature Review.

Sclerosing mediastinitis (SM), previously named chronic fibrosing mediastinitis, is an inflammatory process that in its end-stage results to sclerosis around the mediastinal structures. SM is quite rare and has been correlated with inflammatory and autoimmune diseases, as well as malignancy. SM may either present in a mild form, with minor symptoms and a benign course or in a more aggressive form with severe pulmonary hypertension and subsequent higher morbidity and mortality. The diagnosis of SM may be difficult and quite challenging, as symptoms depend on the mediastinal structure that is mainly involved; quite often the superior vena cava. However, practically any mediastinal structure may be involved by the fibrotic process, such as the central airways, as well as the pulmonary arteries and veins, leading to obstruction or total occlusion. The latter may be impossible to undergo proper surgical excision of the lesion, and is considered to be a real challenge to the surgeon. We herein report a case of SM that presented with arterial and venous compression. The imaging appearance was that of unilateral pulmonary edema, associated with lung collapse. The case is supplemented by a non-systematic review of the relevant literature.

Gestational Diabetes and T-cell (Th1/Th2/Th17/Treg) Immune Profile.

BACKGROUND/AIM: Gestational diabetes mellitus (GDM) is a common pregnancy complication, characterized by insulin resistance and low-grade systemic inflammation with a pro-inflammatory immune system response. Our objective was to study the peripheral Th1, Th2, Th17 and Treg response in GDM compared to normal pregnancy. MATERIALS AND METHODS: Th1, Th2, Th17 and Treg subsets was determined by flow cytometry based on staining for specific intracellular cytokines, as well as C-reactive protein (CRP) and total IgE circulating levels. The health status of all offspring was also assessed 6 months post-delivery. RESULTS: A total of 49 Caucasian adult pregnant women were enrolled into a GDM (n=26) and Control (n=23) group. At the third trimester of pregnancy, the GDM group had a higher proportion of Th2, Th17 and Treg cells compared to control. Contrary to the control group, the GDM group exhibited no significant change in the Th1/Th2/Th17/Treg profile postpartum. Furthermore, higher circulating CRP and total IgE levels were noted in the GDM group compared to controls. At the 6-month post-delivery assessment, 30.8% of the offspring from the GDM group were found to have developed atopic dermatitis, food allergy or allergic proctocolitis compared to none from the control group. CONCLUSION: Compared to an uncomplicated pregnancy, GDM exhibits a significantly different peripheral T-cell profile at the third pregnancy trimester characterized by higher proportion of Th2, Th17 and Treg cells which persist six months post-delivery, while the increased high sensitivity CRP (hsCRP) levels stressed the low-grade inflammatory profile of this disease.

Expression of Immune System-Related Membrane Receptors CD40, RANK, BAFFR and LTßR is Associated with Clinical Outcome of Operated Non-Small-Cell Lung Cancer Patients.

An increasing number of studies implicates the NF-κB (Nuclear Factor of kappa light chain gene enhancer in B cells) alternative pathway in non-small-cell lung cancer (NSCLC). We assessed the clinical significance of CD40 (Tumor necrosis factor receptor superfamily member 5, TNFRSF5), BAFFR (B-cell activating factor receptor), RANK (Receptor activator of NF-κB) and LTßR (lymphotoxin ß receptor) receptors, which activate the alternative pathway of NF-κB, in NSCLC. Evaluation of CD40, BAFFR, RANK and LTßR expression was performed based on the Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) datasets, while protein expression was assessed by immunohistochemistry in specimens from 119 operated NSCLC patients. CD40 gene overexpression was correlated with improved five-year overall survival (OS) (p < 0.001), while increased BAFFR and LTßR mRNA levels were associated with worse OS in patients with adenocarcinomas (p < 0.001 and p < 0.001, respectively). Similarly, patients with adenocarcinomas exhibited a negative correlation between membranous BAFFR protein expression in carcinoma cells and three- and five-year survival (p = 0.021; HR, 4.977 and p = 0.030; HR, 3.358, respectively) as well as between BAFFR protein overexpression in cancer-associated fibroblasts (CAFs) and two-year survival (p = 0.036; HR, 1.983). Patients with increased LTßR nuclear protein staining or stage II patients with lower cytoplasmic LTßR protein expression had worse five-year OS (p = 0.039 and p = 0.008, respectively). Moreover, CD40 protein expression in tumor infiltrating lymphocytes (TILs) and CAFs was positively associated with metastatic spread while BAFFR protein expression in CAFs was negatively associated with bone metastasis (p = 0.041). Our data suggests that CD40, BAFFR, RANK and LTßR play an important role in NSCLC and further supports the role of NF-κB alternative pathway in NSCLC.

Expression Of Intracellular Components of the NF-κB Alternative Pathway (NF-κB2, RelB, NIK and Bcl3) is Associated With Clinical Outcome of NSCLC Patients.

A growing number of studies has shed light on the role of the NF-κΒ in non-small-cell lung cancer (NSCLC). To address the significance of major effectors of the NF-κΒ alternative pathway, we investigated the relationship between NF-κΒ2, RelB, NIK and Bcl3 expression (mRNA and protein) and the clinical outcome of NSCLC patients. NF-κΒ2, RelB, NIK and Bcl3 protein expression levels were assessed by immunohistochemistry in tissue samples from 151 NSCLC patients who had curative resection. mRNA levels were also evaluated in 69 patients using quantitative real-time PCR. Although all studied proteins were overexpressed in NSCLC (P < 0.001 for all), only RelB mRNA levels were strongly increased in cancerous specimens compared to tumor-adjacent non-neoplastic tissues (P = 0.009). Moreover, NF-κB2, RelB and Bcl3 expression was associated with overall survival (OS). In particular, cytoplasmic and mRNA expression of RelB was related to 5-year OS (P = 0.014 and P = 0.006, respectively). Multivariate analysis also showed that Bcl3 expression (nuclear and cytoplasmic) was associated with increased 5-year OS (P = 0.002 and P = 0.036, respectively). In addition, higher Bcl3 mRNA levels were associated with inferior OS in stages I & II and improved OS in stages III and IV after 5-year follow-up (P = 0.004 and P = 0.001, respectively). Furthermore, stage I patients with lower NF-κB2 mRNA levels had better 5-year survival in univariate and multivariate analysis (P = 0.031 and P = 0.028, respectively). Interestingly, RelB expression (cytoplasmic and mRNA) was inversely associated with relapse rates (P = 0.027 and P = 0.015, respectively), while low NIK cytoplasmic expression was associated with lower relapse rates (P = 0.019). Cytoplasmic NIK expression as well as NF-κB2/ Bcl3 detection was associated with lymph node infiltration (P = 0.039 and P = 0.014, respectively). The present study confirms the deregulation of the NF-κB alternative pathway in NSCLC and also demonstrates the importance of this pathway in prognosis, recurrence and infiltration of regional lymph nodes.

Influence of blood transfusions and preoperative anemia on long-term survival in patients operated for non-small cell lung cancer.

It has been postulated that transfusions have immunosuppressive effects that promote tumor growth and metastasis. Moreover perioperative anemia is considered an independent prognostic factor on outcome in patients operated for malignancy. We evaluated the influence of red blood cell (RBC) transfusions and perioperative anemia on survival in non-small cell lung carcinoma (NSCLC) patients. From 1999 through 2005, 331 consecutive patients, male/female=295/36 (mean age 64+/-9 years), who underwent radical surgery for NSCLC were prospectively enrolled in this cohort and followed up for a mean of 27.2 months. The overall survival of patients was analyzed in relation to RBC transfusions and perioperative anemia. These parameters were analyzed in the whole cohort of patients and separately for stage I patients. Patients were divided according to perioperative transfusion, into Group A (transfused) and Group B (non-transfused) and according to the preoperative haemoglobin (Hb) level into Group 1(Hb<12g/dl) and Group 2(Hb> or =12g/dl), respectively. The overall transfusion rate was 25.7%. Univariate analysis showed that in the whole cohort of patients overall survival was significantly shorter in Group A (mean 33.6 months, 5-year survival 25.1%) compared to Group B (mean 48.0 months, 5-year survival 37.3%) (p=0.001). It also showed that patients with preoperative Hb level <12g/dl (Group 1), (mean of 33.0 months, 5-year survival 21.3%) had shorter survival compared to Group 2 patients (mean 49.3 months and 5-year survival 40.0%), respectively (p=0.002). Multivariate analysis in the whole cohort of patients showed that preoperative anemia was an independent risk factor for survival while RBC transfusion was not. In particular for stage I patients, it was shown that RBC transfusion was an independent prognostic factor for long-term survival as detected by multivariate analysis (p=0.043), while anemia was not. RBC transfusions affect adversely the survival of stage I NSCLC patients, while do not exert any effect on survival of patients with surgically resectable more advanced disease, where preoperative anemia is an independent negative prognostic factor. These findings indicate that RBC transfusion might exert an immunomodulatory effect on patients with early disease while in more advanced stages this effect is not apparent.

NF-kB2 Genetic Variations are Significantly Associated with Non-Small Cell Lung Cancer Risk and Overall Survival.

During the last decade, a growing number of publications implicate NF-kB2 in NSCLC pathogenesis. Here, we investigated the clinical relevance of NF-kB2 single nucleotide polymorphisms (SNPs) rs7897947, rs11574852 and rs12769316 in NSCLC and their association with NF-kB2 protein and mRNA levels. Our data show that TT (rs7897947T >G) and AA (rs12769316G >A) genotypes were strongly associated with an increased risk for NSCLC (P = 0.019 and P = 0.003, respectively). Additionally, in multivariate analysis, TT (rs7897947T >G) homozygosity was associated with worse 2- and 3-year survival rates (P = 0.030 and P = 0.028, respectively), especially among patients with stages III/IV, who had worse 2, 3 and 5-year survival (P = 0.001, P = 0.022 and P = 0.035, respectively). In chemotherapy-treated patients, TT (rs12769316G >A) homozygosity was also associated with worse 2- and 3-year survival compared to G allele carriers (P = 0.006 and P = 0.014, respectively). Furthermore, rs12769316 was correlated with survival outcome of stage I and II patients (P = 0.031 and P = 0.006, respectively). Interestingly, amongst the patients who developed metastases, A allele carriers had better 5-year survival (P = 0.020). In addition, rs12769316 was associated with NF-kB2 protein (P = 0.001) and mRNA expression (P = 0.017) as well as with tumor maximum diameter (P = 0.025). Overall, this study suggests that rs7897947 and rs12769316 are involved in NSCLC susceptibility, in treatment response and in clinical outcome.

BNN-20, a synthetic microneurotrophin, strongly protects dopaminergic neurons in the "weaver" mouse, a genetic model of dopamine-denervation, acting through the TrkB neurotrophin receptor.

Neurotrophic factors are among the most promising treatments aiming at slowing or stopping and even reversing Parkinson's disease (PD). However, in most cases, they cannot readily cross the human blood-brain-barrier (BBB). Herein, we propose as a therapeutic for PD the small molecule 17-beta-spiro-[5-androsten-17,2'-oxiran]-3beta-ol (BNN-20), a synthetic analogue of DHEA, which crosses the BBB and is deprived of endocrine side-effects. Using the "weaver" mouse, a genetic model of PD, which exhibits progressive dopaminergic neurodegeneration in the Substantia Nigra (SN), we have shown that long-term administration (P1-P21) of BNN-20 almost fully protected the dopaminergic neurons and their terminals, via i) a strong anti-apoptotic effect, probably mediated through the Tropomyosin receptor kinase B (TrkB) neurotrophin receptor's PI3K-Akt-NF-κB signaling pathway, ii) by exerting an efficient antioxidant effect, iii) by inducing significant anti-inflammatory activity and iv) by restoring Brain-Derived Neurotrophic Factor (BDNF) levels. By intercrossing "weaver" with NGL mice (dual GFP/luciferase-NF-κΒ reporter mice, NF-κΒ.GFP.Luc), we obtained Weaver/NGL mice that express the NF-κB reporter in all somatic cells. Acute BNN-20 administration to Weaver/NGL mice induced a strong NF-κB-dependent transcriptional response in the brain as detected by bioluminescence imaging, which was abolished by co-administration of the TrkB inhibitor ANA-12. This indicates that BNN-20 exerts its beneficial action (at least in part) through the TrkB-PI3K-Akt-NF-κB signaling pathway. These results could be of clinical relevance, as they suggest BNN-20 as an important neuroprotective agent acting through the TrkB neurotrophin receptor pathway, mimicking the action of the endogenous neurotrophin BDNF. Thus BNN-20 could be proposed for treatment of PD.

Thiol redox state and oxidative stress in midbrain and striatum of weaver mutant mice, a genetic model of nigrostriatal dopamine deficiency.

In this study we measured thiol redox state (TRS) and the oxidative stress indicator lipid peroxidation in midbrain and striatum of adult (4 months old) male control (+/+) and weaver (wv/wv) mice in order to relate them with oxidative stress in conditions of progressive and severe (approximately 70%) nigrostriatal dopaminergic neurodegeneration. Specifically, we measured the specific TRS components glutathione (GSH), glutathione disulfide (GSSG), cysteine (CSH), and the general classes of TRS components. The latter are the protein thiols (PSH) and the disulfides between (a) protein (P) and protein thiols (PSSP), (b) protein and non-protein (NP/R) thiols (PSSR, PSSC) and (c) non-protein and non-protein thiols (NPSSR, NPSSC). In addition, the main product of lipid peroxidation malonyl dialdehyde (MDA) was estimated. In the midbrain of wv/wv, GSH and NPSSC levels are decreased (44% and 64%, respectively) and GSSG, NPSSR, CSH, PSH, PSSP, PSSR and MDA levels are increased (23%, 660%, 110%, 51%, 68%, 18% and 44%, respectively). In the striatum of male wv/wv, protein and non-protein thiol/disulfide and MDA levels do not change, possibly due to the high decrease in striatal dopamine level versus midbrain. Our data show that the high degeneration of the dopaminergic nigrostriatal neurons in male adult wv/wv mice is accompanied by significant changes in TRS and an increase in lipid peroxidation in the midbrain, suggesting involvement of oxidative stress in the degeneration of dopaminergic neurons. They also strengthen the possible use of thiol antioxidants for the development of new neuroprotective therapeutic strategies for neurodegenerative diseases, such as Parkinson's disease.

Variant of BCL3 gene is strongly associated with five-year survival of non-small-cell lung cancer patients.

OBJECTIVES: BCL3, a known atypical IκB family member, has been documented to be upregulated in hematological malignancies and in some solid tumors, functioning as a crucial player in tumor development. Recently, rs8100239, a tag-Single Nucleotide Polymorphism (SNP) in BCL3 (T>A) has been identified, but there are no data regarding its involvement in non-small-cell lung cancer (NSCLC) initiation and progression. MATERIALS AND METHODS: To study the possible association of BCL3 with NSCLC, 268 patients and 279 healthy controls were genotyped for rs8100239. Moreover, BCL3 protein expression was also investigated in 112 NSCLC cases through an immunohistochemical analysis. RESULTS: NSCLC patients with AA genotype displayed significantly worse prognosis compared to T allele carriers (P<0.001), who had less frequent intermediate nuclear BCL3 expression (P=0.042). In addition, overexpression of BCL3 was detected in tumor specimens, compared to normal tissue (P<0.001). Furthermore, BCL3 protein levels were associated with five-year survival (P=0.039), maximum diameter of lesion (P=0.012), grade (P=0.002) and relapse frequency (P=0.041). CONCLUSIONS: The present study is the first to show a relationship between the genetic variation rs8100239 of BCL3 and cancer patients' survival. It also represents the first quantitative evaluation of BCL3 expression in NSCLC. Our findings indicate that rs8100239 may be considered as a novel prognostic indicator, demonstrating also the overexpression of BCL3 protein in NSCLC and implicating this pivotal molecule in the pathogenesis of NSCLC.

Thiol redox state (TRS) and oxidative stress in the mouse hippocampus after pentylenetetrazol-induced epileptic seizure.

In this study we evaluated oxidative stress (lipid peroxidation and protein oxidation) and thiol redox state [TRS: glutathione (GSH), glutathione disulfide (GSSG), cysteine (CSH), protein (P) thiols (PSH) and protein and non-protein (NP) mixed/symmetric disulfides (PSSR, NPSSR, NPSSC, PSSP)] in hippocampus after pentylenetetrazol (PTZ) administration at convulsive and subconvulsive dose. The significant decrease in PSH, CSH and NPSSC, as well as the increase in PSSP, NPSSR, lipid peroxidation and protein oxidation levels after PTZ-induced seizure indicate increased oxidative damage in hippocampus, although the levels of GSH and GSSG do not change significantly.

Effect of pentylenetetrazol-induced epileptic seizure on thiol redox state in the mouse cerebral cortex.

In the present study we examined the effects of pentylenetetrazol (PTZ) administration on the thiol redox state (TRS), lipid peroxidation and protein oxidation in left and right mouse cerebral cortex in order (a) to quantitate the major components of the thiol redox state and relate them with oxidative stress and cortical laterality, and (b) to investigate whether neuronal activation without synchronization, induced by subconvulsive doses of PTZ, can cause similar qualitative effects on the thiol redox state. Specifically, we examined the TRS components [glutathione (GSH), glutathione disulfide (GSSG), cysteine (CSH), protein (P) thiols (PSH) and protein and non-protein (NP) mixed/symmetric disulfides (PSSR, NPSSR, NPSSC, PSSP)]. At 15 min after seizure, GSH, GSSG, CSH, NPSSC, PSSR and PSSC levels are decreased in left (14-50%) and right (11-53%) cortex while PSSP levels are increased in both left (1400%) and right (1600%) cortex. At 30 min after seizure, GSSG, CSH, NPSSC, PSSR and PSSC levels are decreased in left (14-51%) and right (18-56%) cortex while PSSP and protein carbonyl levels are increased in left (2300% and 20%, respectively) and right (2800% and 21%, respectively) cortex. At 24 h after seizure, the TRS components return to normal and protein carbonyl levels are decreased in left (16%) and right (20%) cortex. The significant decrease in GSH, GSSG, CSH, NPSSC, PSSR and PSSC, as well as the increase in protein carbonyl and the high increase in PSSP levels after PTZ-induced seizure indicate increased oxidative stress in cerebral cortex of mice, and of similar magnitude and TRS-component profiles between left and right cerebral cortex.