The redox-sensitive human antioxidant responsive element induces gene expression under low oxygen conditions

Transient transfection studies of human HepG2 and mouse Hepa hepatocarcinoma cells with a reporter gene construct regulated by a human antioxidant responsive element (ARE) from the NQO1 gene demonstrated that the element is responsive to low oxygen conditions. The antioxidant N-acetyl L-cysteine (NAC) strongly inhibited basal aerobic reporter gene activity in HepG2 cells without obviously affecting the hypoxic induction, as is consistent with ARE sensitivity to oxidative stress in aerobic cultures. Electrophoretic mobility shift (EMS) assays of nuclear extracts of HepG2 and Hepa cells lysed under aerobic or hypoxic conditions or after exposure to the phenolic compound 3-(2)-tert-butyl-4-hydroxyanisole (BHA), showed specific and constitutive protein binding to the ARE under all of these conditions. Taken together, these findings show that the ARE can mediate gene expression in response to low oxygen conditions. Co-ordinately regulated expression of ARE-dependent genes, such as phase II detoxification enzymes, may be an important phenotype of solid tumors containing significant regions of pathophysiological hypoxia.      

Antibiotic management of pneumococcal infections in an era of increased resistance

Abstract: Pneumococci are a leading cause of bacterial meningitis and bacteraemia, as well as pneumonia, otitis media and sinusitis in childhood. These organisms recently have shown a dramatic increase in antibiotic resistance. Penicillin-resistant pneumococci are of special concern as they are often resistant to other unrelated antibiotics. This is of particular significance to Aboriginal children who have among the highest rates of pneumococcal infection in the world. Laboratories should now test all invasive pneumococcal isolates for penicillin and third generation cephalosporin resistance. Local treatment guidelines are required for pneumococcal infections, especially for meningitis, taking into account the prevalence of resistant strains within the community. At present, penicillin and amoxycillin remain the drugs of choice for pneumococcal infections, with the exception of meningitis where initial empirical therapy must be with a third generation cephalosporin. Judicious antibiotic use, which avoids over-prescribing and unnecessary use of broad-spectrum agents, improved living standards in underprivileged communities and introduction of an effective conjugate vaccine, able to reduce the rates of pneumococcal infection and hopefully colonization, may limit the spread of resistant strains.     

Sound localization in patients with cochlear implant--preliminary results

OBJECTIVES: To evaluate sound localization ability in patients with unilateral cochlear implant, who do not wear a hearing aid on the contralateral ear, and to try to improve this ability by training. SETTING: Tertiary academic referral center. METHODS: In the initial test, patients were exposed to sound stimuli from different directions and were asked to localize them. Following a training period the patients were re-evaluated by the same test. For each test, the percentage of correct answers and the final test score were calculated. RESULTS: In the initial test, the mean score of the study group of four cochlear implant users was 42.8 (out of a maximal score of 100), the mean rate of correct responses was 27.5%. Following a training period (6.5 sessions on the average), on the final test the mean score of the group was 74.3, while the mean rate of correct responses was 66.5%. CONCLUSIONS: The results demonstrated that patients with unilateral cochlear implant have some ability to localize sound, and that this ability may be improved by regular training.     

Quantification of thrombocyte growth factors in platelet concentrates produced by discontinuous cell separation

Platelet concentrates (PC) are increasingly used to increase bone regeneration in pre-prosthetic surgery. Although it is generally appreciated that certain growth factors (PDGF, TGF, EGF, and ECGF) are present in thrombocyte preparations, relatively little is known about these components in quantitative terms. The study reported here analysed the amounts of growth factors in PC produced under standard conditions from healthy volunteers. All the blood samples (237 in total) were analysed using Quantikine ELISA kits (R and D). The mean +/- SD platelet count in whole blood from these donors was 262,000+/-58,000/microl, while in PC produced by discontinuous cell separation it was 1.419,000+/-333,000/microl. The mean growth factor concentrations in PC preparations in ng/ml were as follows: PDGF-AB 125+/-55 ng/ml; TGF-beta1 221+/-92 ng/ml; IGF-I 85+/-25 ng/ml; PDGF-BB 14+/-9 ng/ml; TGF-beta2 0.4+/-0.3 ng/ml. These growth factor concentrations typically covered a 3-10 fold range: PDGF-AB 29-277ng/ml; PDGF-BB 2-33ng/ml; TGF-beta1 32-397ng/ml; TGF-beta2 0.1-1.2 ng/ml; IGF-I 40-138 ng/ml. Platelet counts in PC were slightly higher for women (Mann-Whitney Test all p < 0.001) than for men, while the concentrations of growth factors in PC exhibited no gender-related difference of any statistical significance.     

Loss of EphB6 expression in metastatic melanoma

Overexpression of various members of the Eph receptor tyrosine kinases and their ephrin ligands has been frequently reported in cancer. In contrast, a loss of EphB6 gene expression has been correlated with a poor prognosis in human neuroblastoma, suggesting a distinct role for this receptor compared to other family members. More recently, an important role of EphB6 signalling in T-cells has been described, suggesting possibly deleterious immunologic effects of a loss of EphB6 in cancer progression. We investigated the expression of EphB6 in melanocytic tumors. EphB6 mRNA of 22 microdissected tissues (7 benign nevi, 7 melanomas, 8 metastases) and 10 different cell lines (normal melanocytes, non-metastatic/metastatic melanoma cell lines) were measured by quantitative real-time RT-PCR. For visualization of EphB6 protein expression, immunohistochemistry of 32 melanocytic lesions were performed. On the mRNA level, the benign nevi revealed the highest EphB6 expression (mean = 1.43), while melanomas (mean = 0.63) and metastases (mean = 0.08; p=0.001) displayed a progressive and significant reduction of EphB6 expression. Accordingly, established melanoma cell lines with metastatic potential showed low EphB6 expression in comparison to normal melanocytes and to most of the melanoma cell lines. Immunohistochemistry revealed homogeneous staining in common nevi, whereas in malignant melanomas and metastases a heterogeneously positive to completely negative EphB6 staining was observed. Remarkably, Spitz nevi stained similarly to ordinary melanocytic nevi. Taken together, we show that melanoma progression to metastatic disease is associated with a significant reduction of EphB6 gene expression which may have considerable consequences for the prognosis of malignant melanoma patients and possible gene-therapeutic approaches.     

Antitumor activity of protein kinase C inhibitors and cisplatin in human head and neck squamous cell carcinoma lines

Protein kinase C (PKC) plays a pivotal role in signal transduction involved in the control of cell proliferation, differentiation and apoptosis. Interference with such signaling pathways may result in altered tumor cell response to antineoplastic drugs. We investigated the effects of two selective PKC inhibitors as single agents and in combination with cisplatin in cell lines derived from squamous cell carcinomas of the head and neck (SCCHN). Safingol (Saf) is directed against the regulatory domain, whereas chelerythrine (Che) interacts with the catalytic domain of PKC. In six SCCHN cell lines (UM-SCC 11B, 14A, 14C and 22B, 8029NA, and a 5-fold cisplatin-resistant subline 8029DDP). PKC activities ranged between 1 and 158 IU/1 x 10(7) cells, and they were inversely proportional to the amount of cellular epidermal growth factor receptor. Using the colorimetric MTT assay, PKC inhibitors Saf and Che showed comparable dose-dependent growth inhibition. The 50% inhibitory concentrations (IC50) were between 3.8-8.6 microM for Saf and 8.5-13.6 microM for Che with no relationship to PKC activity or cisplatin sensitivity of the respective cell lines. Combinations of cisplatin (IC50 = 0.4-5.8 microg/ml) and either PKC inhibitor (5 microM Saf, 10 microM Che) led to a significant decrease of cisplatin IC50 values in most cell lines. However, comparison with theoretical additive dose-response curves showed additive rather than synergistic effects for both PKC inhibitors.