[Molecular mechanism of Bupleuri Radix and Scutellariae Radix drug pair for depression based on integrative pharmacology platform of traditional Chinese medicine].

Xiaochaihu decoction is a classic prescription of traditional Chinese medicine. Modern research has proved its anti-depression effect. However, its pharmacological mechanism for anti-depression effect is difficult to be unveiled because of the complexity of compound Chinese medicines. Bupleuri Radix and Scutellariae Radix is the core drug pair of Xiaochaihu decoction. In this research, Bupleuri Radix and Scutellariae Radix were analyzed by the integrative pharmacology platform to study its molecular mechanism for anti-depression. One hundred and sixteen active ingredients were predicted, 62 for Bupleuri Radix, mainly including saikosaponins, acids, alcohols, and 54 for Scutellariae Radix, mainly including flavonoids and glycosides. Its anti-depression effect was relevant to 118 core targets, including 22 known disease targets, such as serotonin receptor(HTR2C), activating transcription factor(ATF1, ATF2), δ opioid receptor(OPRD1), µ opioid receptor (OPRM1), κ opioid receptor(OPRK1), inositol monophosphatase(IMPA1), Toll-like receptor 4 (TLR4), histamine H1 receptor(HRH1), neurotrophic factor tyrosine kinase receptor1 (NTRK1), Glycogen synthetase kinase 3ß(GSK3ß), etc. The antidepressant effect involved positive regulation of transcription from RNA polymerase Ⅱ promoter, transcription factor binding, cytosol, transcriptional regulation of DNA template, enzyme binding, endocrine system, nervous system, neurotrophin signaling pathway, cell growth and death, signal transduction, thyroid hormone signaling pathway and other related biological processes and metabolic pathways. This study provides a scientific evidence for further study of the anti-depression mechanism of this drug pair.

Correlation of rs9331888 polymorphism with Alzheimer's disease among Caucasian and Chinese populations: a meta-analysis and systematic review.

Clusterin polymorphism (rs9331888) was reported to be associated with the susceptibility to alzheimer's disease (AD). Nevertheless, the results were inconclusive. To derive a more precise estimation of this association, this meta-analysis was conducted. We've conducted a comprehensive search of PubMed, Embase, CNKI and AlzGene database for case-control studies published throughout October, 2016 that evaluated the role of rs9331888 gene variants in AD patients. Odds ratios (ORs) and their 95% confidence intervals (CIs) were calculated to assess the strength of associations between the rs9331888/C > G polymorphism and AD disease. A total of 9 studies were enrolled in the Meta Analysis. The overall analysis revealed a significant association between the rs9331888/C > G polymorphism and AD disease in the recessive model (GG vs. GC + CC: OR = 1.11, 95% CI: 1.05-1.18; P < 0.01). Sub-group analysis revealed that the Caucasian populations which with recessive model (GG vs. GC + CC: OR = 1.12, 95% CI: 1.06-1.2; P < 0.01) were dramatically related to AD, while no significant association was found in the Chinese populations among the five genetic models. Our meta-analysis demonstrated that the rs9331888/C > G polymorphism in the clusterin gene might contribute to AD susceptibility especially in Caucasian populations. Whereas the relationship of the polymorphism to the disease in Chinese populations was still in controversial. Additional well-designed studies, with larger sample sizes, are required to further elucidate this association.

Effects of siRNA specific to the protein kinase CK2α on apoptosis of laryngeal carcinoma cells.

BACKGROUND: The relationship between apoptosis and tumors is a major focus in cancer research. RNA interference (RNAi) technology has emerged as a very potent tool to generate cellular knockdown phenotypes of a desired gene. The aim of this study was to explore the effect of siRNA specific to the protein casein kinase 2α (CK2α) on apoptosis of laryngeal carcinoma cells and to explore possible mechanisms. METHODS: An siRNA expression plasmid specific to CK2α, psiRNA-hH1neo-CK2α, and a non-specific siRNA expression plasmid, psiRNA-hH1neo-cont, were constructed and transfected into Hep-2 cells by a lipofectamine method. The mRNA and protein levels of CK2α in transfected cells were determined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting analysis. Apoptosis was measured by Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) double-staining methods. The morphological changes to Hep-2 cells were observed under transmission electron microscopy (TEM). The levels of Bcl-2 and Bax proteins were measured by Western blotting analysis. RESULTS: Levels of CK2α mRNA and protein were significantly decreased in the psiRNA-hH1neo-CK2α group compared to the other groups (P < 0.05). The apoptotic rate of the psiRNA-hH1neo-CK2α transfected group was significantly higher compared to that in the untransfected group and the siRNA-hH1neo-cont transfected group (25.66% ± 0.83%, 3.66% ± 0.43%, and 5.18% ± 0.22%) (P < 0.05). Compared with the untransfected group and the siRNA-hH1neo-cont transfected group, the psiRNA-hH1neo-CK2α transfected group presented with classical ultrastructural features of apoptosis, such as karyopyknosis, chromatic agglutination adjacent to the nuclear membrane, and apoptotic bodies. Compared with the other two groups, the level of Bcl-2 protein in the psiRNA-hH1neo-CK2α transfected group was decreased (0.20 ± 0.09 vs. 0.72 ± 0.16, 0.56 ± 0.11, P < 0.01), while the Bax protein level was increased (0.81 ± 0.17 vs. 0.26 ± 0.12, 0.33 ± 0.17, P < 0.01) and the ratio of Bcl-2 to Bax was decreased (0.25 ± 0.05 vs. 2.76 ± 0.21, 1.70 ± 0.22, P < 0.01). CONCLUSIONS: The siRNA expression plasmid specific to CK2α could suppress CK2α expression and induce the apoptosis of laryngeal carcinoma cells. This is possibly by decreasing the Bcl-2/Bax ratio. CK2α may provide a potential therapeutic target against human laryngeal carcinoma.

[Effects of protein kinase CK2α on apoptosis and ultrastructure of laryngeal carcinoma cells].

OBJECTIVE: To investigate the effect of protein kinase CK2α on apoptosis and ultrastructure of human laryngeal carcinoma cells and its possible mechanism. METHODS: The siRNA expression plasmid psiRNA-hH1neo-CK2α specific to protein kinase CK2α and non-specific siRNA expression plasmid psiRNA-hH1neo-cont were transfected into Hep-2 cells respectively by lipofectamine method. Western blot was used to detect the expression of kinase CK2α protein. The apoptotic rate was measured by Annexin V-FITC/PI double-staining. The morphological changes of Hep-2 cells were observed under transmission electron microscope (TEM). The expressions of bcl-2 and Bax protein were measured by Western blot. RESULTS: The expression of protein kinase CK2α protein significantly decreased in the Hep-2 cells transfected with psiRNA-hH1neo-CK2α (P < 0.01). Compared with the untransfected cells and siRNA-hH1neo-cont transfected group, psiRNA-hH1neo-CK2α transfected group presented with classical ultrastructural features of apoptosis, such as karyopyknosis, chromatic agglutination adjacent to nuclear membrane and apoptotic body. The apoptotic rate of psiRNA-hH1neo-CK2α transfected group was obviously higher than that in untransfected cells and siRNA-hH1neo-cont transfected group (25.66% ± 0.83% vs 3.66% ± 0.43%, 5.18% ± 0.22%, both P < 0.05). Compared with two other groups, the bcl-2 protein expression of psiRNA-hH1neo-CK2α transfected group decreased (0.20 ± 0.09 vs 0.72 ± 0.16, 0.56 ± 0.11, both P < 0.01), the Bax protein expression increased (0.81 ± 0.17 vs 0.26 ± 0.12, 0.33 ± 0.17, both P < 0.01) while the ratio of bcl-2 to Bax decreased (0.25 ± 0.05 vs 2.76 ± 0.21, 1.70 ± 0.22, both P < 0.01). CONCLUSION: Protein kinase CK2a plays an important role in the apoptosis of human laryngeal carcinoma cells possibly by decreasing bcl-2/Bax. Protein kinase CK2a may provide a potential therapeutic target against human laryngeal carcinoma.

Gene transfer into primary cultures of fetal neural stem cells by a recombinant adenovirus carrying the gene for green fluorescent protein.

OBJECTIVE: To evaluate the transduction efficiency of a recombinant adenovirus carrying the gene for green fluorescent protein (Ad-GFP) into the primary cultures of fetal neural stem cells (NSCs) by the expression of GFP. METHODS: The Ad-GFP was constructed by homologous recombination in bacteria with the AdEasy system; NSCs were isolated from rat fetal hippocampus and cultured as neurosphere suspensions. After infection with the recombinant Ad-GFP, NSCs were examined with a fluorescent microscopy and a flow cytometry for their expression of GFP. RESULTS: After the viral infection, flow cytometry analysis revealed that the percentage of GFP-positive cells was as high as 97.05%. The infected NSCs sustained the GFP expression for above 4 weeks. After differentiated into astrocytes or neurons, they continued to express GFP efficiently. CONCLUSION: We have successfully constructed a viral vector Ad-GFP that can efficiently infect the primary NSCs. The reporter gene was showed fully and sustained expression in the infected cells as well as their differentiated progenies.

[Experimental study on embryonic neural stem cells transplantation into natural rat cochlea via round window].

OBJECTIVE: To investigate the survival of neural stem cell (NSC) infected by recombinant adenovirus with GFP (Ad-GFP) and the expression of GFP in normal rat cochlea and their potential effect on auditory function and cochlea structures via round window transplantation. METHODS: In comparison with the normal rats without any transplantation (Group III), normal rat cochleae were transplanted with NSC infected with Ad-GFP (Group I) or the artificial perilymph (Group II) via round windows. Auditory functions were monitored by thresholds of auditory brain stem responses (ABR); the cochlea structures were examined by HE staining; survivals of implanted NSC were determined by the expression of GFP; survivals of hair cells were accessed by whole mount preparation. RESULTS: Neither at pre-transplantation nor at post-transplantation, there were significant differences in the click-ABR thresholds in rats between Group I and Group II (P > 0.05). There were no significant differences in these values before and after transplantation in the same rats from each group. After transplantation, the cochlea structures were normal in both Group I & Group II. Grafted NSC was visualized by the GFP expression in every turn of the cochlea in all animals of Group I. There were no significant differences in the loss of outer hair cell (OHC) among three groups. The inner hair cell (IHC) and most OHC were normal in every turns of cochleae of all groups. CONCLUSIONS: The embryonic NSC infected with Ad-GFP could survive and express the GFP gene in normal rat cochlea after transplantation via round window, which had not obvious affection to auditory function and inner ear pathology of rat cochlea.

[Inhibitory effects of small interfering RNA specific to protein kinase CK2a on the growth of laryngeal carcinoma cells].

OBJECTIVE: To assess the effect of small interfering RNA (siRNA) specific to protein kinase CK2a on proliferation and apoptosis of Hep-2 cell line. METHODS: siRNA expression plasmid psiRNA-hH1neo-CK2 specific to protein kinase CK2a and non-specific siRNA expression plasmid psiRNA-hH1neo-cont were constructed respectively, and then were transfected into Hep-2 cells by lipofectamine methods. Protein kinase CK2a mRNA and protein of the transfected cells were detected by reverse transcription polymerase chain reaction (RT-PCR) and Western Blot, respectively. Proliferation and apoptosis of the transfected cells were observed by methyl thiazolyl tetrazolium (MTT) method and flow cytometry (FCM), respectively. RESULTS: Protein kinase CK2a mRNA and protein expressions were significantly decreased in the cells transfected with psiRNA-hH1neo-CK2 (P < 0.05). The Hep-2 cells grew slowly after transfected with psiRNA-hH1neo-CK2(P < 0.05). Obvious subdiploid peaks were found in the cells transfected with psiRNA-hH1neo-CK2 (P < 0.05). CONCLUSIONS: siRNA expression plasmid specific to protein kinase CK2a suppressed the protein kinase CK2a expression and the proliferation of Hep-2, and induced apoptosis of Hep-2 cells.