Simultaneous modulation of COX-2, p300, Akt, and Apaf-1 signaling by melatonin to inhibit proliferation and induce apoptosis in breast cancer cells

Melatonin exhibits anti-inflammatory and anticancer effects and could be a chemopreventive and chemotherapeutic agent against cancers, but the precise mechanisms involved remain largely unresolved. In this study, we evaluated the mechanism of action of melatonin in human MDA-MB-361 breast cancer cells. Melatonin at pharmacological concentrations (10(-3) m) significantly suppressed cell proliferation and induced apoptosis in a dose-dependent manner. The observed suppression of proliferation was accompanied by the melatonin-mediated inhibition of COX-2, p300, and NF-κB signaling. Melatonin significantly inhibited COX-2 expression and prostaglandin E(2) (PGE2) production, abrogated p300 histone acetyltransferase activity and p300-mediated NF-κB acetylation, thereby blocking NF-κB binding and p300 recruitment to COX-2 promoter. Pretreatment with a COX-2- or p300-selective inhibitor abrogated the melatonin-induced inhibition of cell proliferation, whereas PGE2 treatment or COX-2 transfection reversed the inhibition by melatonin. Moreover, melatonin markedly inhibited phosphorylation of PI3K, Akt, PRAS40, and GSK-3 proteins, thereby inactivating the PI3K/Akt signaling pathway. Pretreatment with a PI3K- or an Akt-selective inhibitor or an Akt-specific siRNA blocked the melatonin-mediated inhibition of cell proliferation. Conversely, gene delivery of a constitutively active Akt effectively reversed the inhibition by melatonin. Furthermore, melatonin induced Apaf-1 expression, triggered cytochrome C release, and stimulated caspase-3 and caspase-9 activities and cleavage, leading to an activation of the Apaf-1-dependent apoptotic pathway. Pretreatment with an Apaf-1-specific siRNA effectively attenuated the melatonin-induced apoptosis. These results therefore indicate that melatonin inhibits cell proliferation and induces apoptosis in MDA-MB-361 breast cancer cells in vitro by simultaneously suppressing the COX-2/PGE2, p300/NF-κB, and PI3K/Akt/signaling and activating the Apaf-1/caspase-dependent apoptotic pathway.     

Activation of Tetrodotoxin-Resistant Sodium Channel NaV1.9 in Rat Primary Sensory Neurons Contributes to Melittin-Induced Pain Behavior

Tetrodotoxin-resistant (TTX-R) sodium channels Na_V1.8 and Na_V1.9 in dorsal root ganglion (DRG) neurons play important roles in pathological pain. We recently reported that melittin, the major toxin of whole bee venom, induced action potential firings in DRG neurons even in the presence of a high concentration (500 nM) of TTX, indicating the contribution of TTX-R sodium channels. This hypothesis is fully investigated in the present study. After subcutaneous injection of melittin, Na_V1.8 and Na_V1.9 significantly upregulate mRNA and protein expressions, and related sodium currents also increase. Double immunohistochemical results show that Na_V1.8-positive neurons are mainly medium- and small-sized, whereas Na_V1.9-positive ones are only small-sized. Antisense oligodeoxynucleotides (AS ODNs) targeting Na_V1.8 and Na_V1.9 are used to evaluate functional significance of the increased expressions of TTX-R sodium channels. Behavioral tests demonstrate that AS ODN targeting Na_V1.9, but not Na_V1.8, reverses melittin-induced heat hypersensitivity. Neither Na_V1.8 AS ODN nor Na_V1.9 AS ODN affects melittin-induced mechanical hypersensitivity. These results provide previously unknown evidence that upregulation of Na_V1.9, but not Na_V1.8, in small-sized DRG neurons contributes to melittin-induced heat hypersensitivity. Furthermore, melittin-induced biological effect indicates a potential strategy to study properties of TTX-R sodium channels.     

Retinoic Acid and Human Olfactory Ensheathing Cells Cooperate to Promote Neural Induction From Human Bone Marrow Stromal Stem Cells

The generation of induced neuronal cells from human bone marrow stromal stem cells (hBMSCs) provides new avenues for basic research and potential transplantation therapies for nerve injury and neurological disorders. However, clinical application must seriously consider the risk of tumor formation by hBMSCs, neural differentiation efficiency and biofunctions resembling neurons. Here, we co-cultured hBMSCs exposed to retinoic acid (RA) with human olfactory ensheathing cells (hOECs) to stimulate its differentiation into neural cells, and found that hBMSCs following 1 and 2 weeks of stimulation promptly lost their immunophenotypical profiles, and gradually acquired neural cell characteristics, as shown by a remarkable up-regulation of expression of neural-specific markers (Tuj-1, GFAP and Galc) and down-regulation of typical hBMSCs markers (CD44 and CD90), as well as a rapid morphological change. Concomitantly, in addition to a drastic decrease in the number of BrdU incorporated cells, there was a more elevated synapse formation (a hallmark for functional neurons) in the differentiated hBMSCs. Compared with OECs alone, this specific combination of RA and hOECs was significantly potentiated neuronal differentiation of hBMSCs. The results suggest that RA can enhance and orchestrate hOECs to neural differentiation of hBMSCs. Therefore, these findings may provide an alternative strategy for the repair of traumatic nerve injury and neurological diseases with application of the optimal combination of RA and OECs for neuronal differentiation of hBMSCs.     

血浆ox-LDL在阿尔茨海默病患者中的临床研究

目的探讨氧化低密度脂蛋白(oxidized Low density lipoprotein,ox-LDL)与阿尔茨海默病(Alzheimer’s dis-ease,AD)患者认知功能损害的关系。方法运用简易精神状态检查量表(mini mental state examination,MMSE)检测入组患者认知功能,将其分为3组,即正常组(NC组,37例)、血管性痴呆组(VD组,41例)、阿尔茨海默病组(AD组,48例),分别用酶联免疫吸附试验(enzyme-linked immunosorbnent assay,ELSIA)测定3组患者血浆ox-LDL的浓度。结果 NC组、VD组、AD组患者血浆ox-LDL浓度分别为(47.46±10.04)μg/L、(60.95±11.78)μg/L、(112.25±17.81)μg/L,AD组患者血浆ox-LDL浓度显著高于VD组,差异有统计学意义(P<0.01);相关分析显示,AD患者血浆ox-LDL浓度与MMSE得分呈负相关(r=-0.574,P<0.01);对AD患者各个认知领域的分值与血浆ox-LDL水平进行多元线性回归分析显示,血浆ox-LDL水平与AD患者记忆力、定向力、注意力和计算能力呈负相关。结论血浆ox-LDL浓度测定有可能作为判断AD患者认知功能损害的生化指标,亦可作为AD与VD鉴别诊断的生物学指标。     

Research progress in rehabilitation treatment of stroke patients A bibliometric analysis

BACKGROUND: Stroke presents as a transient or chronic brain dysfunction and is associated with high morbidity and high mortality. The doctors and scientists would like to argue how to enhance the validity of the rehabilitation treatment and how to further improve the level of treatment on stroke. OBJECTIVE: The aim of this study was to quantitatively analyze the current worldwide progress in research on stroke rehabilitation treatment based on Web of Science database and ClinicalTrial.gov in the past 10 years. METHODS: We conducted a quantitative analysis of clinical trial articles regarding stroke rehabilitation published in English from 2003 to 2013 and indexed in the National Institutes of Health Clinical Trials registry and Web of Science databases. Data were downloaded on March 15, 2013. RESULTS: (1) From 2003 to 2013, 2 654 clinical trials investigating stroke were indexed in ClinicalTrials.gov. There were only 58 clinical trials registered in 2003, and there was a marked increase from 2005. A total of 605 clinical trials on the rehabilitation of stroke were conducted in the past 10 years. (2) The analysis showed that most of the trials in the field were registered by North American institutions. With respect to the Asian countries, China and Taiwan area of China also published a reasonable proportion of the trials, but comparatively speaking, the number of trials is really rare. Most of the interventions were drugs, followed by the devices, and behavioral interventions were ranked third. (3) In the past 10 years, there were 4 052 studies on stroke indexed by Web of Science database. CONCLUSION: From perspective of research progress, we found that the number of clinical trials and papers on stroke rehabilitation has increased significantly in the past 10 years, between them a remarkable positive correlation exists.     

Isolation and characterization of 113 polymorphic microsatellite loci for the Tibetan frog (Nanorana parkeri) using next generation sequencing

We developed and characterized 113 polymorphic microsatellites in the Tibetan frog (Nanorana parkeri) using 454 GS-FLX next generation sequencing technology. These loci were tested in 46 individuals from two N. parkeri populations from the Tibetan plateau. The average number of alleles per locus was 8.09 (range = 2–20). The average observed heterozygosities (H _O) per locus in the two populations were 0.58 (range = 0.04–1) and 0.29 (range = 0–0.87), respectively. These microsatellites will be useful for population and conservation genetics studies of N. parkeri and other closely related species.     

Probe substrate and enzyme source-dependent inhibition of UDP-glucuronosyltransferase (UGT) 1A9 by wogonin

Background

Drug-metabolizing enzymes (DMEs) inhibition based drug-drug interaction and herb-drug interaction severely challenge the R&D process of drugs or herbal ingredients.

Objective

To evaluate the inhibition potential of wogonin (an important flavonoid isolated from the root of Scutellaria baicalensis) towards one of the most important phase II DMEs, UDP-glucuronosyltransferase (UGT) 1A9.

Methods

Both recombinant UGT1A9-catalyzed 4-methylumbelliferone (4-MU) glucuronidation reaction and human liver microsomes (HLMs)-catalyzed propofol glucuronidation reaction were used as two different probe reactions.

Results

Wogonin noncompetitively inhibited recombinant UGT1A9-catalyzed 4-MU glucuronidation, and exerted competitive inhibition towards HLMs-catalyzed propofol glucuronidation. The inhibition kinetic parameters (K_i) were calculated to be 3.2 µM and 52.0µM, respectively.

Conclusion

Necessary monitoring was needed when wogonin was co-administered with the clinical drugs mainly undergoing UGT1A9-mediated glucuronidation elimination. Additionally, probe reactions-dependent inhibition of wogonin towards the activity of UGT1A9 should be paid attention when translating these in vitro data into in vivo situation.