Alterations in BDNF and trkB mRNAs following acute or sensitizing cocaine treatments and withdrawal

In the present study, we used in situ hybridization to examine the influence of acute or repeated cocaine administrations and withdrawal from repeated cocaine treatment on the level of brain-derived neurotrophic factor (BDNF) and its receptor trkB mRNAs in rat brain. Cocaine (10 mg/kg i.p.) injected acutely produced locomotor hyperactivation, while repeated (single injection for 5 days) administrations of cocaine (10 mg/kg) induced a two-fold increases in the locomotor activity in rats in response to a challenge cocaine dose (10 mg/kg) on day 10, as compared to the saline-treated animals (sensitization). Cocaine treatments induced a brain-region-specific decrease in the levels of trkB mRNA. On the other hand, BDNF mRNA in the rat hippocampus was increased only in the group of rats subjected to cocaine withdrawal. Animals under cocaine withdrawal demonstrated a significant increase in the immobility time measured by the use of modified forced swimming test. Therefore, the increases in the levels of BDNF mRNA in the rat hippocampus seem to be correlated with "depressive-like" behavioral effects during withdrawal from repeated cocaine treatment. In the shell (but not in the core) of the nucleus accumbens, the levels of BDNF mRNA were significantly increased following acute and repeated cocaine treatment as well as during cocaine withdrawal, which indicates that the alterations in the neurotrophin level in the brain region important for the expression of cocaine-induced sensitization involve other mechanisms.     

Recruitment and activation of mRNA decay enzymes by two ARE-mediated decay activation domains in the proteins TTP and BRF-1

In human cells, a critical pathway in gene regulation subjects mRNAs with AU-rich elements (AREs) to rapid decay by a poorly understood process. AREs have been shown to directly activate deadenylation, decapping, or 3'-to-5' exonucleolytic decay. We demonstrate that enzymes involved in all three of these mRNA decay processes, as well as 5'-to-3' exonucleolytic decay, associate with the protein tristetraprolin (TTP) and its homolog BRF-1, which bind AREs and activate mRNA decay. TTP and BRF-1 each contain two activation domains that can activate mRNA decay after fusion to a heterologous RNA-binding protein, and inhibit ARE-mediated mRNA decay when overexpressed. Both activation domains employ trans-acting factors to trigger mRNA decay, and the N-terminal activation domain functions as a binding platform for mRNA decay enzymes. Our data suggest that the TTP protein family functions as a molecular link between ARE-containing mRNAs and the mRNA decay machinery by recruitment of mRNA decay enzymes, and help explain how deadenylation, decapping, and exonucleolytic decay can all be independently activated on ARE-containing mRNAs. This describes a potentially regulated step in activation of mRNA decay.     

DNA- versus RNA-based methods for human papillomavirus detection in cervical neoplasia

OBJECTIVE: To compare DNA-based and mRNA-based methods for detection of high-grade cervical neoplasia in Norway. METHODS: HPV prevalence was analyzed in 383 women with positive index cytology, selected from gynecology clinics. All patients were investigated by a new PAP smear, histology, and two commercially available HPV tests: Hybrid Capture II (Digene, Gaithersburg, MD) and the Pre Tect HPV-Proofer (NorChip AS). Cases with positive DNA test and negative mRNA test and cases with high-grade histology and negative HPV tests were retested with PCR and sequencing. We regarded the infection as latent or transient if sequencing revealed an HPV type included in both assays. RESULTS: High-risk HPV was detected in 99.7% of the histological confirmed high-grade lesions (CIN2+) (290/291). The DNA test was positive in 95% (275/291), and the mRNA test was positive in 77% (225/291) of the histological confirmed high-grade lesions. All invasive carcinomas were mRNA positive. The DNA test was significantly more often positive in benign and low-grade lesions, some of which were found to be false positive due to cross-contamination with unrelated types. High-grade histology was detected in 83% of women with normal cytology and positive mRNA test. Latent or transient infections were detected in 11 low-grade and 12 high-grade preinvasive lesions. Sequencing revealed high-risk HPV types included only in the DNA test in 35 high-grade preinvasive lesions, HPV 52 and 58 were the most prevalent HPV types. CONCLUSIONS: These HPV tests have the potential to improve the detection rate of high-grade cervical neoplasia, with some limitations. The mRNA test seems to be more appropriate for risk-evaluation. Larger scale, population based studies are necessary to evaluate the predictive values of HPV testing in Norway.     

Role of AFP mRNA expression in peripheral blood as a predictor for postsurgical recurrence of hepatocellular carcinoma： A systematic review and meta-analysis

AIM: To identify the role of alpha-fetoprotein (AFP) mRNA expression in peripheral blood one week after surgery as a predictor for recurrence of hepatocellular carcinoma (HCC). METHODS: Published studies fulfilling the selection criteria were identified by searching several databases online. After a methodology assessment using a quality scale designed by European Lung Cancer Working Party, data in each research were aggregated by means of meta-analysis. RESULTS: Altogether 368 cases were included in the 9 selected studies, which fulfilled the selection criteria. The quality scores ranged from 35% to 84% with a median score of 55%. The 'design' subscore had the lowest median value (38%). By aggregating the data, a high x2 value (77.576) was presented. The fail-safe number was 136 and 64 for P= 0.05 and 0.01, respectively. CONCLUSION: AFP mRNA expression in peripheral blood 1 wk after surgery correlated with the recurrence of HCC and was a good predictor for tumor recurrence.     

Effects of arginine supplementation on splenocyte cytokine mRNA expression in rats with gut-derived sepsis

AIM: To investigate the effects of arginine (Arg)-enriched diets before sepsis and/or Arg-containing total parenteral nutrition (TPN) after sepsis or both on cytokine mRNA expression levels in splenocytes of rats with gut-derived sepsis. METHODS: Rats were assigned to four experimental groups. Groups 1 and 2 were fed with a semipurified diet, while groups 3 and 4 had part of the casein replaced by Arg which provided 2% of the total calories. After the rats were fed with these diets for 10 d, sepsis was induced by cecal ligation and puncture (CLP), at the same time an internal jugular vein was cannulated. All rats were maintained on TPN for 3 d. Groups 1 and 3 were infused with conventional TPN, while groups 2 and 4 were supplemented with Arg which provided 2% of the total calories in the TPN solution. All rats were killed 3 d after CLP to examine their splenocyte subpopulation distribution and cytokine expression levels. RESULTS: Plasma interleukin (IL)-2, IL-4, tumor necrosis factor-α(TNF-α) and interferon (IFN-γ) were not detectable 3 d after CLP. There were no differences in the distributions of CD45Ra+, CD3+, CD4+, and CD8+ cells in whole blood and splenocytes among the four groups. The splenocyte IL-2 mRNA expression in the Arg-supplemented groups was significantly higher than that in group 1. IL-4 mRNA expression in groups 3 and 4 was significantly higher than that in groups 1 and 2. The mRNA expression of IL-10 and IFN-γ was significantly higher in group 4 than in the other three groups. There was no difference in TNF-α mRNA expression among the four groups. CONCLUSION: The influence of Arg on the whole blood and splenic lymphocyte subpopulation distribution is not obvious. However, Arg administration, especially before and after CLP, significantly enhances the mRNA expression levels of Th1 and Th2 cytokines in the spleen of rats with gut-derived sepsis.     

Immunocytochemical mapping and quantification of expression of a putative type 1 serotonin receptor in the crayfish nervous system

Serotonin is an important neurotransmitter that is involved in modulation of sensory, motor, and higher functions in many species. In the crayfish, which has been developed as a model for nervous system function for over a century, serotonin modulates several identified circuits. Although the cellular and circuit effects of serotonin have been extensively studied, little is known about the receptors that mediate these signals. Physiological data indicate that identified crustacean cells and circuits are modulated via several different serotonin receptors. We describe the detailed immunocytochemical localization of the crustacean type 1 serotonin receptor, 5-HT1crust, throughout the crayfish nerve cord and on abdominal superficial flexor muscles. 5-HT1crust is widely distributed in somata, including those of several identified neurons, and neuropil, suggesting both synaptic and neurohormonal roles. Individual animals show very different levels of 5-HT1crust immunoreactivity (5-HT(1crust)ir) ranging from preparations with hundreds of labeled cells per ganglion to some containing only a handful of 5-HT(1crust)ir cells in the entire nerve cord. The interanimal variability in 5-HT(1crust)ir is great, but individual nerve cords show a consistent level of labeling between ganglia. Quantitative RT-PCR shows that 5-HT1crust mRNA levels between animals are also variable but do not directly correlate with 5-HT(1crust)ir levels. Although there is no correlation of 5-HT1crust expression with gender, social status, molting or feeding, dominant animals show significantly greater variability than subordinates. Functional analysis of 5-HT1crust in combination with this immunocytochemical map will aid further understanding of this receptor's role in the actions of serotonin on identified circuits and cells.     

Levothyroxin restores hypothyroidism-induced impairment of LTP of hippocampal CA1: electrophysiological and molecular studies

Hypothyroidism impairs synaptic plasticity as well as learning and memory. Clinical reports are conflicting about the ability of thyroid hormone replacement therapy to fully restore the hypothyroidism-induced learning and memory impairment. Recently, we have shown that hypothyroidism impairs LTP and cognition in adult rats. We have studied the effect of thyroxin replacement therapy on hypothyroidism-induced LTP impairment using electrophysiological and molecular approaches. Recording from CA1 region of the hippocampus in anesthetized adult rat indicated that 6 weeks of thyroxin replacement therapy (20 microg/kg/day) fully restored LTP impaired by hypothyroidism. Western blotting showed reduction in phosphorylated (P)-CAMKII, total-CaMKII, neurogranin, and calmodulin basal levels in the CA1 region of the hippocampus of hypothyroid rats. The levels of these molecules were normalized by thyroxin replacement therapy. The hypothyroid-induced elevation of basal calcineurin levels and activity was also normalized by thyroxin treatment. However, thyroxin replacement therapy did not restore hypothyroidism-induced reduction in PKCgamma basal protein levels. Additionally, real-time PCR, showed a reduction in basal neurogranin mRNA level that was normalized by thyroxin replacement therapy. In the sham (control) rats, induction of LTP by high-frequency stimulation increases P-CaMKII, and total CaMKII levels as well as CaMKII phosphotransferase activity. However, in hypothyroid rats, the same stimulation protocol induced an increase only in total-CaMKII. Thyroxin treatment normalized the levels and activity of these molecules. The results demonstrated that thyroxin therapy normalized the electrophysiological and molecular effects of hypothyroidism on the CA1 region and emphasized the critical role P-CaMKII plays in hypothyroidism-induced LTP impairment.     

应用组织芯片研究CD44v9 mRNA在乳腺癌中的表达及与Ki-67相关性

目的:探讨CD44v9 mRNA和Ki-67在乳腺癌中表达的关系。方法:应用组织芯片分别结合原位杂交和免疫组化方法检测乳腺癌中CD44v9 mRNA和Ki-67的表达。结果:乳腺癌组织CD44v9 mRNA和Ki-67表达水平明显高于正常乳腺组织(P<0.05)。乳腺癌组织中CD44v9 mRNA的表达与淋巴结转移、远处转移具有显著相关性,Ki-67的表达与组织病理分级、淋巴结转移均有显著相关性。它们的表达与肿瘤大小、年龄等无关(P>0.05)。两者表达成正相关(r=0.367,P<0.01)。结论:CD44v9 mRNA和Ki-67与乳腺癌的恶性生物学行为密切相关,可作为预测肿瘤转移潜能的指标。组织芯片技术是病理学研究的一种新方法,具有高效、简便及标准化的特点。