A new approach to sepsis treatment by rasagiline: a molecular,biochemical and histopathological study

Molecular Biology Reports - We aimed to investigate the effects of rasagiline on acute lung injury that develops in the sepsis model induced with the cecal ligation and puncture in rats. The rats...     

Genotoxic evaluation of selective serotonin-reuptake inhibitors by use of the somatic mutation and recombination test in Drosophila melanogaster

This study evaluated different concentrations of selective serotonin-reuptake inhibitors (citalopram and sertraline) for genotoxicity by use of the somatic mutation and recombination test (SMART) in Drosophila melanogaster. Three-day-old larvae, trans-heterozygous for the multiple wing hairs (mwh) and flare (flr3) genes were treated with these two compounds. Two recessive markers were located on the left arm of chromosome 3, i.e. 'multiple wing hairs' (mwh) in map position 0.3 and 'flare-3' (flr3) at 38.8, while the centromere was located in position 47.7. SMART is based on the loss of heterozygosity, which may occur through various mechanisms, such as mitotic recombination, mutation, deletion, half-translocation, chromosome loss, and non-disjunction. Genetic changes occurring in somatic cells of the wing's imaginal discs, cause the formation of mutant clones on the wing blade. The results of this study show that citalopram had a genotoxic effect in the Drosophila SMART. Sertraline, however, did not show any genotoxic effect in balancer heterozygous wings. This study concluded that more information is needed to be certain regarding the mutagenic effects of sertraline.     

Dimer-Dependent Intrinsic/Basal Activity of the Class B G Protein-Coupled Receptor PAC1 Promotes Cellular Anti-Apoptotic Activity through Wnt/β-Catenin Pathways that Are Associated with Dimer Endocytosis

The high expression of PACAP (pituitary adenylate cyclase-activating polypeptide)-preferring receptor PAC1 is associated with nerve injury and tumors. Our previous report (Yu R, et al. PLoS One 2012; 7: e51811) confirmed the dimerization of PAC1 and found that the M-PAC1 mutation in the N-terminal first Cys/Ala lost the ability to form dimers. In this study, Chinese hamster ovary (CHO-K1) cells overexpressing wild-type PAC1 (PAC1-CHO) had significantly higher anti-apoptotic activities against serum withdrawal-induced apoptosis associated with a lower caspase 3 activity and a higher Bcl-2 level in a ligand-independent manner than those of CHO cells overexpressing the mutant M-PAC1 (M-PAC1-CHO). PAC1-CHO had significantly higher β-catenin, cyclin D1 and c-myc levels corresponding to the Wnt/β-catenin signal than did M-PAC1-CHO. In addition, the Wnt/β-catenin pathway inhibitor XAV939 significantly inhibited the anti-apoptotic activities of PAC1-CHO. Top-flash assays demonstrated that PAC1-CHO had a significantly stronger Wnt/β-catenin signal than did M-PAC1-CHO. Acetylcysteine (NAC) as an inhibitor of the dimerization of PAC1 inhibited the anti-apoptotic activities that were endowed by PAC1 and decreased the Wnt/β-catenin signal in Top-flash assays. In the PAC1 Tet (tetracycline)-on inducible gene expression system by doxycycline (Dox), higher expression levels of PAC1 resulted in higher anti-apoptotic activities that were associated with a stronger Wnt/β-catenin signal. A similar correlation was also found with the down-regulation of PAC1 in the Neuro2a neuroblastoma cell. BiFC combined with fluorescence confocal imaging indicated that during serum-withdrawal-induced apoptosis, PAC1 dimers displayed significant endocytosis. These findings indicate that PAC1 has ligand-independent and dimer-dependent intrinsic/basal activity, conferring cells with anti-apoptotic activities against serum withdrawal, which is involved in the Wnt/β-catenin signal and is associated with the endocytosis of PAC1 dimers. The discovery and study of the dimer-dependent basal activity of PAC1 not only help us understand the physiological and pathological role of PAC1 but also promote the development of drugs targeting PAC1.     

The determination of the carbonic anhydrases activators in vitro effect of mixed donor crown ethers

Carbonic anhydrases (CAs) play an important function in various physiological and pathological processes. Therefore, many researchers work in this field in order to design and synthesize new drugs. Both inhibitors and activators of CAs, which are associated with the diagnosis and treatment of many diseases, are very important. The emergence of the use of CA activators in the treatment of Alzheimer has led many scholars to work on this issue. In this study, CA activators and inhibitors are determined. The crown ethers compounds ( 1 , 2 , 3 , 6 , 7 , 8 , and 9 ) were found to cause activation on enzyme activities of hCA I and II. The AC₅₀ values on hCA I and II of the compounds are in the range of 4.6565–374.979 μM. The 4 (IC₅₀; 1.301 and 3.215 μM for hCA I and II) and 5 (IC₅₀; 73.96 and 378.5 μM for hCA I and II) compounds were found to cause inhibition on enzyme activities of hCA I and II.     

Surface Reconstruction through Poisson Disk Sampling

This paper intends to generate the approximate Voronoi diagram in the geodesic metric for some unbiased samples selected from original points. The mesh model of seeds is then constructed on basis of the Voronoi diagram. Rather than constructing the Voronoi diagram for all original points, the proposed strategy is to run around the obstacle that the geodesic distances among neighboring points are sensitive to nearest neighbor definition. It is obvious that the reconstructed model is the level of detail of original points. Hence, our main motivation is to deal with the redundant scattered points. In implementation, Poisson disk sampling is taken to select seeds and helps to produce the Voronoi diagram. Adaptive reconstructions can be achieved by slightly changing the uniform strategy in selecting seeds. Behaviors of this method are investigated and accuracy evaluations are done. Experimental results show the proposed method is reliable and effective.     

Cholesterol transport through the peroxisome-ER membrane contacts tethered by PI(4,5)P_2 and extended synaptotagmins

Most mammalian cells take up cholesterol from low-density lipoproteins(LDLs) via receptor-mediated endocytosis. After reaching lysosomes, LDL-derived cholesterol continues to transport to downstream organelles including the ER for specific structural and functional needs. Peroxisomes are recently found to receive cholesterol from lysosomes through lysosomeperoxisome membrane contacts. However, whether and how cholesterol is conveyed from peroxisomes to the ER remain unknown. Here, by combining high-resolution microscopic analyses and in vitro reconstitution of highly purified organelles or artificial liposomes, we demonstrate that peroxisomes form membrane contacts with the ER through the interaction between peroxisomal PI(4,5)P_2 and ER-resident extended synaptotagmin-1, 2 and 3(E-Syts). Depletion of peroxisomal PI(4,5)P_2 or ESyts markedly decreases peroxisome-ER membrane contacts and induces cholesterol accumulation in lysosomes. Furthermore,we show that cholesterol is delivered from ~3H-labeled peroxisomes or PI(4,5)P_2-containing liposomes to the ER in vitro, and that the presence of peroxisomes augments cholesterol transfer from lysosomes to the ER. Together, our study reveals a new cholesterol transport pathway along the lysosome-peroxisome-ER membrane contacts in the cell.     

Evaluation of 5-HT7 Receptor Trafficking on In Vivo and In Vitro Model of Lipopolysaccharide (LPS)-Induced Inflammatory Cell Injury in Rats and LPS-Treated A549 Cells

This study aimed to investigate the effects of the 5-HT7 receptor agonist (LP44) and antagonist (SB269970) on LPS-induced in vivo tissue damage and cell culture by molecular methods. This study was conducted in two steps. For in vivo studies, 24 female rats were divided into four groups. Group I: healthy; II (2nd h): LPS 5 mg/kg administered intraperitoneally (i.p.); III (4th h): LPS 5 mg/kg administered i.p.; IV (8th h): LPS 5 mg/kg administered i.p. For in vitro studies, we used the A549 cell line. Groups: I control (healthy) (2–4 h); II LPS: 1 µg/ml E. Coli O55:B5 strain (2–4 h); III agonist (LP44) 10^?9 M (2–4 h); IV antagonist (SB269970) 10^?9 M (2–4 h); V LPS+agonist 10^?9 M (LP44 1 µg/ml) (2–4 h); VI LPS+antagonist 10^?9 M (2–4 h). In molecular analyses, we determined increased TNF-α, IL-1β, NF-κB, and 5-HT7 mRNA expressions in rat lung tissues and increased TNF-α, iNOS, and 5-HT7 mRNA expressions in the A549 cell line. In in vitro parameters, LP44 agonist administration-related decrease was observed. Our study showed that lung 5-HT7 receptor expression is increased in LPS-induced endotoxemia. All this data suggest that 5-HT7 receptor overexpression is an important protective mechanism during LPS-induced sepsis-related cell damage.     

B族G蛋白偶联受体PAC1可调控表达体系的建立

PAC1是神经肽垂体腺苷酸环化酶激活多肽(Pituitary adenylate cyclase activating polypeptide,PACAP)的特异受体,属于B族G蛋白偶联受体,介导PACAP的神经递质、神经调质、神经保护、抗神经损伤及调控神经再生等功能,PAC1高表达和神经损伤、肿瘤等生理病理过程密切相关。为了深入了解PAC1的功能,构建PAC1可调控表达的细胞系,通过优化的四环素控制表达系统实现PAC1在中国仓鼠卵巢(Chinese hamster ovary,CHO)细胞的强力霉素(doxycycline,Dox)依赖的可控表达。首先通过双酶切将编码PAC1和增强型黄色荧光蛋白(EYFP,enhanced yellow fluorescent protein)的融和基因PAC1-EYFP克隆到pTRE-Tight载体上,获得重组载体pTRE-PAC1-EYFP;基因测序鉴定正确后将新型的四环素调节元件载体pTet-on advanced和反应元件载体pTRE-PAC1-EYFP分别转入CHO细胞中,G418和潮霉素(Hygromycin)双抗筛选阳性克隆PAC1-Tet-CHO,使用梯度浓度四环素类似物强力霉素Dox诱导PAC1-EYFP表达,48 h后检测受体表达水平,并通过MTT法检测不同PAC1表达水平的细胞增殖活性。荧光检测和Western印迹结果显示,成功获得了具有良好诱导性的Dox依赖的PAC1可控表达的细胞系,这些细胞株在传10代后仍能稳定地可控表达PAC1。MTT结果显示PAC1表达水平越高,细胞增殖活性越强。成功所构建的Dox依赖的PAC1可控表达细胞系,为PAC1的生物学功能的深入研究奠定了基础。