Quantitative promoter methylation analysis of multiple cancer-related genes in renal cell tumors

Background  

Aberrant promoter hypermethylation of cancer-associated genes occurs frequently during carcinogenesis and may serve as a cancer biomarker. In this study we aimed at defining a quantitative gene promoter methylation panel that might identify the most prevalent types of renal cell tumors.     

Family history and the risk of kidney cancer: a multicenter case-control study in Central Europe.

An elevated familial relative risk may indicate either an important genetic component in etiology or shared environmental exposures within the family. Incidence rates of kidney cancer are particularly high in Central Europe, although no data were available on the familial aggregation or genetic background of kidney cancer in this region. We have, therefore, investigated the role of family history in first-degree relatives in a large multicenter case-control study in Central Europe. A total number of 1,097 cases of kidney cancer and 1,555 controls were recruited from 2000 to 2003 from seven centers in Czech Republic, Poland, Romania, and Russia. The risk of kidney cancer increased with the increasing number of relatives with history of any cancer [odds ratio (OR), 1.15; 95% confidence interval (95% CI), 1.00-1.31 per affected relative], and this association seemed to be more prominent among subjects with young onset (OR, 1.55; 95% CI, 1.09-2.20 per affected relative). Overall, the OR was 1.40 (95% CI, 0.71- 2.76) for the subjects who had at least one first-degree relative with kidney cancer after adjusting for tobacco smoking, body mass index, and medical history of hypertension, and this association was most apparent among subjects with affected siblings (OR, 4.09; 95% CI, 1.09-15.4). Based on the relative risk to siblings in our study population, we estimated that 80% of the kidney cancer cases are likely to occur in 20% of the population with the highest genetic risk, which indicate the importance of further investigation of genetic factors in cancer prevention for kidney cancer.     

High promoter methylation levels of APC predict poor prognosis in sextant biopsies from prostate cancer patients.

PURPOSE: Prostate cancer is a highly prevalent malignancy and constitutes a major cause of cancer-related morbidity and mortality. Owing to the limitations of current clinical, serologic, and pathologic parameters in predicting disease progression, we sought to investigate the prognostic value of promoter methylation of a small panel of genes by quantitative methylation-specific PCR (QMSP) in prostate biopsies. EXPERIMENTAL DESIGN: Promoter methylation levels of APC, CCND2, GSTP1, RARB2, and RASSF1A were determined by QMSP in a prospective series of 83 prostate cancer patients submitted to sextant biopsy. Clinicopathologic data [age, serum prostate-specific antigen (PSA), stage, and Gleason score] and time to progression and/or death from prostate cancer were correlated with methylation findings. Log-rank test and Cox regression model were used to identify which epigenetic markers were independent predictors of prognosis. RESULTS: At a median follow-up time of 45 months, 15 (18%) patients died from prostate cancer, and 37 (45%) patients had recurrent disease. In univariate analysis, stage and hypermethylation of APC were significantly associated with worse disease-specific survival, whereas stage, Gleason score, high diagnostic serum PSA levels, and hypermethylation of APC, GSTP1, and RASSF1A were significantly associated with poor disease-free survival. However, in the final multivariate analysis, only clinical stage and high methylation of APC were significantly and independently associated with unfavorable prognosis, i.e., decreased disease-free and disease-specific survival. CONCLUSIONS: High-level APC promoter methylation is an independent predictor of poor prognosis in prostate biopsy samples and might provide relevant prognostic information for patient management.     

Time dependent alterations on tyrosine hydroxylase, opioid and cannabinoid CB1 receptor gene expressions after acute ethanol administration in the rat brain.

The aim of this study was to examine the differential regulation after acute ethanol administration on tyrosine hydroxylase, proenkephalin and cannabinoid CB(1) receptor gene expressions in selected areas of the rat brain. Rats received an intragastric administration of 3 g/kg ethanol and were killed by decapitation at 1, 2, 4, 8 and 24 h. The results showed an activation of tyrosine hydroxylase gene expression in the ventral tegmental area and the substantia nigra, increased proenkephalin gene expression in the caudate-putamen, nucleus accumbens core and shell, central and medial amygdala, ventromedial hypothalamic nucleus and the paraventricular hypothalamic nucleus. In contrast, a significant decrease in the cannabinoid CB1 receptor gene expression was found in caudate-putamen, central amygdala and ventromedial hypothalamic nucleus. In conclusion, the results suggest that an acute dose of ethanol induces neuroplastic alterations in proenkephalin, tyrosine hydroxylase and cannabinoid CB1 receptor gene expressions that may contribute to trigger the rewarding effects of ethanol consumption.     

Diode laser-induced mitosis in the rabbit retinal pigment epithelium.

BACKGROUND AND OBJECTIVE: The induction of retinal pigment epithelium (RPE) proliferation without damaging the inner layers of the retina might be helpful in patients with RPE atrophic changes and degeneration. This study aimed to induce mitosis in the RPE of the rabbit after subthreshold photocoagulation with the diode laser. MATERIALS AND METHODS: Twenty-five male Dutch rabbits received retinal photocoagulation using an 810-nm diode laser with different power settings and exposure times. The eyes were processed for light microscopy, electron transmission microscopy, and immunohistochemistry. RESULTS: Neither morphological alterations nor mitotic activity was found after 5-mJ energies. Retinal layers were not affected and RPE hyperplasia appeared in the treated areas associated with mitotic activity when 10 mJ was used. Mitosis induction and retinal damage appeared with 20, 50, and 100 mJ and were associated with ophthalmoscopic damage. CONCLUSIONS: The use of subthreshold 810-nm diode laser treatment may induce mitosis in the RPE without causing damage to the neighboring layers.     

Chronic antidepressants induce redistribution and differential activation of alphaCaM kinase II between presynaptic compartments.

Changes in synaptic plasticity are involved in pathophysiology of depression and in the mechanism of antidepressants. Ca(2+)/calmodulin (CaM) kinase II, a protein kinase involved in synaptic plasticity, has been previously shown to be a target of antidepressants. We previously found that antidepressants activate the kinase in hippocampal neuronal cell bodies by increasing phosphorylation at Thr(286), reduce the kinase phosphorylation in synaptic membranes, and in turn its phosphorylation-dependent interaction with syntaxin-1 and the release of glutamate from hippocampal synaptosomes. Here, we investigated the chronic effect of different antidepressants (fluoxetine, desipramine, and reboxetine) on the expression and function of the kinase in distinct subcellular compartments in order to dissect the different kinase pools affected. Acute treatments did not induce any change in the kinase. In total tissue extracts chronic drug treatments induced activation of the kinase; in hippocampus (HC), but not in prefrontal/frontal cortex, this was partially accounted for by increased Thr(286) phosphorylation, suggesting the involvement of different mechanisms of activation. In synaptosomes, all drugs reduced the kinase phosphorylation, particularly in HC where, upon fractionation of the synaptosomal particulate into synaptic vesicles and membranes, we found that the drugs induced a redistribution and differential activation of the kinase between membranes and vesicles. Furthermore, a large decrease in the level and phosphorylation of synapsin I located at synaptic membranes was consistent with the observed decrease of CaM kinase II. Overall, antidepressants induce a complex pattern of modifications in distinct subcellular compartments; at presynaptic level, these changes are in line with a dampening of glutamate release.     

Panretinal photocoagulation versus PRP plus intravitreal bevacizumab for high‐risk proliferative diabetic retinopathy (IBeHi study)

Purpose: To evaluate the effects of panretinal photocoagulation (PRP) compared with PRP plus intravitreal bevacizumab on best corrected visual acuity (BCVA) and total area of fluorescein leakage from active new vessels (NVs) in patients with high‐risk proliferative diabetic retinopathy (PDR). Methods: We carried out a prospective study of patients with high‐risk PDR and no prior laser treatment who were randomly assigned to receive PRP (PRP group) or PRP plus intravitreal injection of 1.5 mg of bevacizumab (PRP‐plus group). In all patients, the PRP was administered at two time‐points (weeks 1 and 3), with the intravitreal bevacizumab delivered at the end of the second laser episode in the PRP‐plus group. Standardized ophthalmic evaluation including Early Treatment Diabetic Retinopathy Study BCVA as well as stereoscopic fundus photography and fluorescein angiography were performed at baseline and at weeks 4, 9 (± 1) and 16 (± 2). Main outcome measures included changes in BCVA and in total area of fluorescein leakage from active NVs. Results: Twenty‐two (n = 30 eyes) consecutive patients completed the 16‐week follow‐up. There was no significant difference between the PRP and PRP‐plus groups with respect to age, gender, type or duration of diabetes, area of fluorescein leakage from active NVs or BCVA. No significant difference in BCVA was observed between the groups throughout the study period. However, the total area of actively leaking NVs was significantly reduced in the PRP‐plus group compared with the PRP group at weeks 4, 9 and 16 (p < 0.001). No major adverse events were identified. Conclusions: In the short‐term, the adjunctive use of intravitreal bevacizumab with PRP was associated with a greater reduction in the area of active leaking NVs than PRP alone in patients with high‐risk PDR.     

Design and In-line Raman Spectroscopic Monitoring of a Protein Batch Crystallization Process

Raman spectroscopy was used to design and monitor a lysozyme protein batch crystallization process in a lab scale study to facilitate the design of a pharmaceutical protein manufacturing process. A D-optimal design that consisted of 18 experiments was performed to elucidate the effect of temperature, concentration of the precipitating agent, time of crystallization, and possible interactions between these three factors on the Raman scattering changes. A polynomial mathematical model was calculated relating the scattering of the lysozyme solutions measured at individual Raman shifts to the significant factors obtained in the previous crystallization experiment. The 2,940-cm⁻¹ band provided the highest correlation values indicative of small prediction errors and good predictive ability for the crystallization model. Raman scattering signals obtained during the experiments were used as input to obtain a response surface for the factors studied and elucidate the relationship between the crystallization process conditions and the crystals obtained. The main factors affecting the crystallization process were the sodium chloride concentration and temperature.