Aneuploidies, deletion, and overexpression of TP53 gene in intestinal metaplasia of patients without gastric cancer

Gastric carcinogenesis is attributable to interacting environmental and genetic factors, through a sequence of events including intestinal metaplasia. Using a fluorescence in situ hybridization technique, we investigated the occurrence of ancuploidies of chromosomes 3, 7, 8, 9, and 17, TP53 gene deletion, and expression of p53 in 21 intestinal metaplasia (IM) samples from cancer-free patients and in 20 gastric adenocarcinoma samples. Aneuploidies were found in 71% (15/21) of the IM samples. Trisomy of chromosomes 7 and 9 occurred mainly in complete-type IM; in the incomplete type, trisomy of chromosomes 7 and 8 were more commonly found. The TP53 gene deletion was observed in 60% (3/5) of the IM cases, and immunohistochemistry revealed p53 overexpression in 12% (2/17) of the analyzed IM cases. All gastric adenocarcinoma cases presented higher frequencies of trisomy or tetrasomy of chromosomes 3, 7, 8, 9, and 17. The TP53 deletion was found in all three of the gastric adenocarcinoma analyzed for it, and immunohistochemistry detected overexpression of protein p53 in 80% (12/15) of the analyzed cases. Our study revealed for the first time the presence of aneuploidies of chromosomes 7, 8, 9, and 17 and of TP53 gene deletion and overexpression in IM samples from cancer-free patients. These results suggest that IM and gastric adenocarcinoma may share the same genetic alterations. (C) 2004 Elsevier B.V. All rights reserved.

CD147 overexpression allows an accurate discrimination of bladder cancer patients' prognosis

Background: Urothelial bladder carcinoma (UBC) is a chemo-sensitive tumour, but the response to treatment is heterogeneous. CD 147 has been associated with chemotherapy resistance. We aimed to define tumours with an aggressive phenotype by the combined analysis of clinicopathological and biological parameters.Methods: 77 patients with T1G3 or muscle-invasive UBC treated by radical cystectomy were studied. Immunohistochemistry was performed to detect CD147, heparanase, CD31 (blood vessels identification) and D2-40 (lymphatic vessels identification) expressions. The immunohistochemical reactions were correlated with the clinicopathological and the outcome parameters. 5-year disease-free survival (DFS) and overall survival (OS) rates were estimated using the Kaplan-Meier method. Multivariate analysis was performed by Cox proportional hazards analysis.Results: The 5-year DFS and OS rates were significantly influenced by the classical clinicopathological parameters, and by the occurrence of lymphovascular invasion. CD 147 and heparanase immunoexpression did not affect patients' outcome. However, patients with pT3/pT4 tumours had a median OS time of 14.7 months (95% CI 7.1-22.3, p = 0.003), which was reduced to 9.2 months (95% CI 1.5-17.0, p = 0.008) if the tumours were CD147 positive. We developed a model of tumour aggressiveness using parameters as stage, grade, lymphovascular invasion and CD147 immunoexpression, which separated a low aggressiveness from a high aggressiveness group, remaining as an independent prognostic factor of DFS (HR 3.746; 95% CI 1.244-11.285; p = 0.019) and OS (HR 3.247; 95% CI 1.015-10.388, p = 0.047).Conclusion: CD 147 overexpression, included in a model of UBC aggressiveness, may help surgeons to identify patients who could benefit from a personalized therapeutic regimen. Additional validation is needed. (C) 2011 Elsevier Ltd. All rights reserved.

Galectin-3 Overexpression in Invasive Laryngeal Carcinoma, Assessed by Computer-assisted Analysis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

During the initiation of fermentation overexpression of hexokinase PII in yeast transiently causes a similar deregulation of glycolysis as deletion of Tps1

In the yeast Saccharomyces cerevisiae a novel control exerted by TPS1 (=GGS1=FDP1=BYP1=CIF1=GLC6=TSS1)-encoded trehalose-6-phosphate synthase, is essential for restriction of glucose influx into glycolysis apparently by inhibiting hexokinase activity in vivo. We show that up to 50-fold overexpression of hexokinase does not noticeably affect growth on glucose or fructose in wild-type cells. However, it causes higher levels of glucose-6-phosphate, fructose-6-phosphate and also faster accumulation of fructose-1,6-bisphosphate during the initiation of fermentation. The levels of ATP and Pi correlated inversely with the higher sugar phosphate levels. In the first minutes after glucose addition, the metabolite pattern observed was intermediate between those of the tps1Δ mutant and tile wild-type strain. Apparently, during the start-up of fermentation hexokinase is more rate-limiting in the first section of glycolysis than phosphofructokinase. We have developed a method to measure the free intracellular glucose level which is based on the simultaneous addition of D-glucose and an equal concentration of radiolabelled L-glucose. Since the latter is not transported, the free intracellular glucose level can be calculated as the difference between the total B-glucose measured (intracellular + periplasmic/extracellular) and the total L-glucose measured (periplasmic/extracellular). The intracellular glucose level rose in 5 min after addition of 100 mM-glucose to 0.5-2 mM in the wild-type strain, ± 10 mm in a hxk1Δ hxk2Δ glk1Δ and 2-3 mM in a tps1Δ strain. In the strains overexpressing hexokinase PII the level of free intracellular glucose was not reduced. Overexpression of hexokinase PII never produced a strong effect on the rate of ethanol production and glucose consumption. Our results show that overexpression of hexokinase does not cause the same phenotype as deletion of Tps1. However, it mimics it transiently during the initiation of fermentation. Afterwards, the Tps1-dependent control system is apparently able to restrict Properly up to 50-fold higher hexokinase activity.

Cloning, overexpression, and purification of functional human purine nucleoside phosphorylase

Purine nucleoside phosphorylase (PNP) catalyzes the phosphorolysis of the N-ribosidic bonds of purine nucleosides and deoxynucleosides. A genetic deficiency due to mutations in the gene encoding for human PNP causes T-cell deficiency as the major physiological defect. Inappropriate activation of T-cells has been implicated in several clinically relevant human conditions such as transplant tissue rejection, psoriasis, rheumatoid arthritis, lupus, and T-cell lymphomas. Human PNP is therefore a target for inhibitor development aiming at T-cell immune response modulation. In addition, bacterial PNP has been used as reactant in a fast and sensitive spectrophotometric method that allows both quantitation of inorganic phosphate (Pi) and continuous assay of reactions that generate P i such as those catalyzed by ATPases and GTPases. Human PNP may therefore be an important biotechnological tool for P i detection. However, low expression of human PNP in bacterial hosts, protein purification protocols involving many steps, and low protein yields represent technical obstacles to be overcome if human PNP is to be used in either high-throughput drug screening or as a reagent in an affordable P i detection method. Here, we describe PCR amplification of human PNP from a liver cDNA library, cloning, expression in Escherichia coli host, purification, and activity measurement of homogeneous enzyme. Human PNP represented approximately 42% of total soluble cell proteins with no induction being necessary to express the target protein. Enzyme activity measurements demonstrated a 707-fold increase in specific activity of cloned human PNP as compared to control. Purification of cloned human PNP was achieved by a two-step purification protocol, yielding 48 mg homogeneous enzyme from 1 L cell culture, with a specific activity value of 80 U mg -1. © 2002 Elsevier Science (USA). All rights reserved.

p16INK4A immunohistochemical overexpression in premalignant and malignant oral lesions infected with human papillomavirus

Human papillomavirus (HPV) is believed to promote the oncogenic process, and the correlation between viral oncoproteins and dysfunction of p16 INK4A tumor suppressor protein in oral lesions is controversial. To test the hypothesis that anogenital HPV types participate in disruption of the regulation of p16INK4A suppressor protein in oral lesions, we analyzed 46 oral biopsy specimens for the presence of HPV 6/11 and 16/18 by in situ hybridization (ISH) and for p16INK4A expression by immunohistochemistry (IHC). Eighteen (39%) of the 46 oral lesions were HPV-positive and 28 (61%) were HPV-negative. HPV 6/11 DNA was found in 5 (11%) and HPV 16/18 in 13 (28%) of 46 biopsies. Nine of the 18 HPV-positive oral lesions (50%), assessed by catalyzed signal amplification coupled to ISH (CSA-ISH), gave high-intensity p16INK4A immunostaining. Focal and diffuse patterns were observed in 11/13 (77%) lesions with HPV 16/18, focal immunopositivity in 3/5 (80%) with HPV 6/11, and negative or sporadic p16-labeling in 18/28 (64%) without the presence of HPV DNA. These results showed a strong association between overexpression of p16 protein and malignant oral lesions, mainly those infected by HPV 16/18. We can conclude that high-risk HPV types are associated with p16 overexpression, and p16 may serve as a biomarker in oral cancer related to high-risk HPV infection.

Apoptosis in different gastric lesions and gastric cancer: Relationship with Helicobacter pylori, overexpression of p53 and aneuploidy

Apoptosis has an essential function in maintaining the integrity of the gastrointestinal mucosa. Its deregulation is associated with the occurrence of lesions such as in atrophic gastritis, peptic ulcers, intestinal metaplasia, and stomach tumorigenesis. Thus, the aim of the present study was to investigate the frequency of apoptotic cells (apoptotic index, AI) by using two different immunohistochemical techniques, TUNEL and anti-activated caspase-3 antibody (CPP32), in gastric dyspepsia [chronic gastritis (CG, N = 34), chronic atrophic gastritis (CAG, N = 11), gastric ulcer (GU, N = 17), and intestinal metaplasia (IM, N = 15)], normal gastric mucosae (NM, N = 8), and gastric adenocarcinoma (GC, N = 12). The relationship was investigated between the AI and Helicobacter pylori infection, diagnosed by PCR, overexpression of p53 protein determined by immunohistochemistry, and aneuploidy by fluorescence in situ hybridization, as performed by our laboratory in previous studies. No significant differences were observed in AI between the different groups, whether by the TUNEL technique (F = 1.60; P = 0.1670) or by CPP32 antibody (F = 1.70; P = 0.1420). Nonetheless, CAG and CG groups had AI statistically higher than those of normal mucosae. These two groups (CAG and CG) also showed a higher frequency of apoptosis-positive cases (TUNEL+ or CPP32+). Generally, there was no correlation between the AI detected by the TUNEL and CPP32 techniques in the groups studied, except in the GC group (r = 0.70). Moreover, there was no significant association between apoptosis and H. pylori infection, overexpression of p53 protein and aneuploidy, but the H. pylori-positive cases only of GU (P = 0.0233) and IM (P = 0.0253) groups displayed a statistically higher AI compared to H. pylori-negative NM, when the CPP32 antibody technique was used. Thus, CG and CAG have increased apoptosis, which may occur independent of an association with H. pylori infection, aneuploidy and overexpression of p53 protein. ©FUNPEC-RP.

Overexpression of ANXA1 in Penile Carcinomas Positive for High-Risk HPVs

The incidence of penile cancer varies between populations but is rare in developed nations. Penile cancer is associated with a number of established risk factors and associated diseases including phimosis with chronic inflammation, human papillomavirus (HPV) infection, poor hygiene and smoking. The objective of this study was to identify genes related to this type of cancer. The detection of HPV was analyzed in 47 penile squamous cell carcinoma samples. HPV DNA was detected in 48.9% of penile squamous cell carcinoma cases. High-risk HPV were present in 42.5% of cases and low-risk HPV were detected in 10.6% of penile squamous cell carcinomas. The RaSH approach identified differential expression of Annexin A1 (ANXA1), p16, RPL6, PBEF1 and KIAA1033 in high-risk HPV positive penile carcinoma; ANXA1 and p16 were overexpressed in penile squamous cells positive for high-risk HPVs compared to normal penile samples by qPCR. ANXA1 and p16 proteins were significantly more expressed in the cells from high-risk HPV-positive penile carcinoma as compared to HPV-negative tumors (p<0.0001) independently of the subtype of the carcinoma. Overexpression of ANXA1 might be mediated by HPV E6 in penile squamous cell carcinoma of patients with high-risk HPVs, suggesting that this gene plays an important role in penile cancer. © 2013 Calmon et al.

NF-kB overexpression and decreased immunoexpression of AR in the muscular layer is related to structural damages and apoptosis in cimetidine-treated rat vas deferens

Background: Cimetidine, histamine H2 receptors antagonist, has caused adverse effects on the male hormones and reproductive tract due to its antiandrogenic effect. In the testes, peritubular myoid cells and muscle vascular cells death has been associated to seminiferous tubules and testicular microvascularization damages, respectively. Either androgen or histamine H2 receptors have been detected in the mucosa and smooth muscular layer of vas deferens. Thus, the effect of cimetidine on this androgen and histamine-dependent muscular duct was morphologically evaluated.Methods: The animals from cimetidine group (CMTG; n=5) received intraperitoneal injections of 100 mg/kg b.w. of cimetidine for 50 days; the control group (CG) received saline solution. The distal portions of vas deferens were fixed in formaldehyde and embedded in paraffin. Massońs trichrome-stained sections were subjected to morphological and the following morphometrical analyzes: epithelial perimeter and area of the smooth muscular layer. TUNEL (Terminal deoxynucleotidyl-transferase mediated dUTP Nick End Labeling) method, NF-kB (nuclear factor kappa B) and AR (androgen receptors) immunohistochemical detection were also carried out. The birefringent collagen of the muscular layer was quantified in picrosirius red-stained sections under polarized light. The muscular layer was also evaluated under Transmission Electron Microscopy (TEM).Results: In CMTG, the mucosa of vas deferens was intensely folded; the epithelial cells showed numerous pyknotic nuclei and the epithelial perimeter and the area of the muscular layer decreased significantly. Numerous TUNEL-labeled nuclei were found either in the epithelial cells, mainly basal cells, or in the smooth muscle cells which also showed typical features of apoptosis under TEM. While an enhanced NF-kB immunoexpression was found in the cytoplasm of muscle cells, a weak AR immunolabeling was detected in these cells. In CMTG, no significant difference was observed in the birefringent collagen content of the muscular layer in comparison to CG.Conclusions: Cimetidine induces significant damages in the epithelium; a possible antiandrogenic effect on the basal cells turnover should be considered. The cimetidine-induced muscle cells apoptosis confirms the susceptibility of these cells to this drug. The parallelism between enhanced cytoplasmic NF-kB immunolabeling in the damaged muscular tissue and muscle cell apoptosis suggests that this drug may avoid the translocation of NF-kB to the nucleus and interfere in the control of NF-kB-mediated smooth muscle cell apoptosis. The decreased immunoexpression of ARs verified in the damaged muscular tissue reinforces this possibility. © 2013 Koshimizu et al.; licensee BioMed Central Ltd.

STEAP1 protein overexpression is an independent marker for biochemical recurrence in prostate carcinoma

Aims: To investigate the prognostic value of expression levels of the genes STEAP1 and STEAP2, and of STEAP1 protein, in prostate carcinomas (PCa). Methods and results: STEAP1 and STEAP2 transcript levels were evaluated by RT-qPCR in samples from 35 PCa, 24 adjacent non-neoplastic prostate (AdjP) tissues, five cases of benign prostatic hyperplasia (BPH), and two histologically normal prostates (N). STEAP1 expression was assessed by immunohistochemistry in samples from 198 PCa, 76 AdjP, 22 BPH, and two N. The findings were compared with clinical and pathological parameters and patient outcome. STEAP1 and STEAP2 transcript analysis showed no differences between the groups tested. Although not significant, higher STEAP1 mRNA levels were detected in tumours with high Gleason scores and in patients who presented with biochemical recurrence (BCR). STEAP1 overexpression was detected in PCa, and was significantly associated with high-grade Gleason scores, seminal vesicle invasion, BCR, and worse outcome (metastasis or PCa-specific death). STEAP1 overexpression was significantly associated with shorter BCR-free survival. Multivariate analysis revealed that STEAP1 is an independent marker for BCR. Conclusions: These findings provide evidence that STEAP1 is a biomarker of worse prognosis in PCa patients. © 2013 John Wiley & Sons Ltd.

Overexpression of UCP1 in tobacco induces mitochondrial biogenesis and amplifies a broad stress response

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Overexpression of the activated form of the AtAREB1 gene (AtAREB1 Delta QT) improves soybean responses to water deficit

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Transcriptome response signatures associated with the overexpression of a mitochondrial uncoupling protein (AtUCP1) in tobacco

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The receptor-like kinase SlSOBIR1 is differentially modulated by virus infection but its overexpression in tobacco has no significant impact on virus accumulation

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)