In order to study the effects and the possible mechanisms of Daxx overexpressed in HepG2 to hydrogen peroxide treatment, and to search new targets for cancer chemotherapy, HepG2cells were transfected using lipofectamine 2000, and selected by treatment with G418. Stable cell lines were confirmed by reverse transeriptase polymerase chain reaction (RT-PCR) targeting vector gene. Experiments include the following groups: (1) control group (non-transfected cells); (2) transfected with empty vector (HepG2/GFP cells); and (3) transfected with pEGFP-C1-Daxx (HepG2/GFP-Daxx cells). After incubation with hydrogen peroxide (H2O2) for 24 h, cellular viability was analyzed by MTT, and cellular apoptosis was measured by flow cytometric analysis. Gene expression at protein level was detected by Western blot. The RT-PCR results showed that Daxx RNA in cells transfected with pEGFP-C1-Daxx was increased significantly compared with that in the HepG2/GFP cells. Fluorescence microscopy revealed that Daxx protein was localized in the nuclei. Hydrogen peroxide was used to induce apoptosis of HepG2 cells and observed that the hydrogen peroxide decreased the viability of HepG2 cells in concentration-dependent pattern. The IC50 values in three groups (Normal cells, HepG2/GFP cells and HepG2/GFP-Daxx cells) were 0.72, 0.76, and 0.49 mmol/L respectively. The apoptotic ratio was significantly higher in HepG2/GFP-Daxx cells as compared to the other two groups. HepG2/GFP-Daxx cell incubated with hydrogen peroxide, showed a significant increase in the activation of caspase-3 and JNK as compare with the other groups. Over-expression of Daxx facilitated HepG2 cells apoptosis induced by hydrogen peroxide. Furthermore, there may be a synergetic relation with apoptosis and increase of JNK activity.

Aim To investigate the effect of intestinal cholesterol absorption inhibitor Ezetimibe on lipid accumulation in RAW264.7 cells and identify the underlying mechanism.Method RAW264.7 cells were pretreated with the indicated concentrations of Ezetimibe (0,0.003,0.01 and 0.03 mol·L~(-1))for 24 hours or pretreated with the optimal concentration(0.03 mol·L~(-1))of Ezetimibe for different periods (0,6,12 and 24 h),followed by incubation with 50 mg·L~(-1) oxLDL for 24 hours,then the number of intracellular lipid droplets and lipid content were measured by using oil red O staining and HPLC; the expression of NPC1L1 was measured by Western blot.Results Pretreatment with indicated concentrations of Ezetimibe caused a concentration-dependent inhibition of intracellular lipid accumulation;pretreatment with 0.03 mol·L~(-1) Ezetimibe caused a time-dependent inhibition of intracellular lipid accumulation.It was noted that pretreatment with 0.03 mol·L~(-1) Ezetimibe for 24 hours inhibited CE by about 47%+0.1% compared with control group(oxLDL alone).Immunoblotting results showed that NPC1L1 was expressed in RAW264.7 cells and it was down-regulated after Ezetimibe treatment.Conclusions Ezetimibe causes concentration-dependent and time-dependent inhibition of lipid accumulation in RAW264.7 cells;it also reduces NPC1L1 expression in RAW264.7 cells.

Aim To explore the anti-proliferation effects of curcumin trinicotinate ( CurTn ) on vascular smooth muscle cell ( VSMC ) and its mechanism. Methods The cells were cultured in DMEM supple-mented with 10% fetal bovine serum. MTT assay was used to examine cell proliferation. FCM was used to observe cell cycle. The expressions of PCNA, Cy-clinD1 and p-ERK1/2 were analyzed using Western blot. Results CurTn could inhibit the proliferation of VSMC and showed a certain amount-time relationship. What’ s more, CurTn could increase the G1 phase pro-portion of cell, decrease the S phase proportion and the expression level of PCNA protein. It was also found that CurTn significantly inhibit the protein expression of p-ERK1/2 and Cyclin D1 . Conclusion CurTn may inhibit the proliferation of VSMC via downregulating the expression of CyclinD1 and p-ERK1/2 .

Objective:To determine the equilibrium solubility and the apparent oil/water partition coefficients of nicotinate-curcu-min ester,so as to provide basis for the new formula design. Methods:Nicotinate-curcumin ester was dissolved in buffer solution with pH of 1.2-7.8. HPLC was used to detect the equilibrium solubility of nicotinate-curcumin ester in various solutions. The apparent oil/water partition coefficients in octanol-water and octanol-buffer solution system were measured by a shaking-flask method. Results:The solubility of nicotinate-curcumin ester in pH 6.8 was the highest. The apparent oil-water partition coefficients of nicotinate-curcumin ester in pH 1.2,5.8,6.5 and 7.8 were lgPapvalues within the range(lgPap=1.69-1.98) beneficial to the absorption in vivo. But in pH 2,5 and 6.8,the lgPapvalues were greater than 2 with strong lipophilicity. Conclusion:The equilibrium solubility as well as ap-parent oil/water partition coefficients of nicotinate-curcumin ester is greatly influenced by the pH value of media. In the pH value with relatively high solubility,lipophilicity is stronger,suggesting it is difficult to be absorbed by the body and needs to be further studied on dosage forms.

Objective:To explore the effects of the process parameters of film dispersion-ultrasonic method on particle diameters and encapsulation efficiency of solid lipid nanoparticles of nicotinate-curcumin in order to obtain the process parameters resulting in smaller particle diameters and higher encapsulation efficiency .Methods: The content of nicotinate-curcumin was determined by HPLC.With the particle diameters and encapsulation efficiency of solid lipid nanoparticles of nicotinate -curcumin as the evaluation in-dicators , the process parameters of film dispersion-ultrasonic method were optimized by orthogonal experimental design and 95％confi-dence interval overlap method as the statistical analysis .Results:The optimum technological parameters of the solid lipid nanoparticles were as follows:the water bath temperature was 40℃and the rotation speed of eggplant bottle was 80 r· min-1 .The average particle size of solid lipid nanoparticles was 107.8 nm, polymer dispersity index ( PDI) was 0.583, and the encapsulation efficiency was 68.91％.Conclusion:The rotation speed of eggplant shaped bottle has notable influence on the particle diameters of solid lipid nanop -articles of nicotinate-curcumin prepared by film dispersion-ultrasonic method , while the water-bath temperature shows great influence on the encapsulation efficiency .Under the optimum conditions , it is possible to obtain the solid lipid nanoparticles with relatively smaller average particle diameters and higher encapsulation efficiency .

OBJECTIVE: To construct a recombinant lentiviral expression vector pCDH-Daxx-EGFP to investigate the effect of Daxx on the proliferation of vascular smooth muscle cells (VSMCs). METHODS: The recombinant lentiviral expression vector pCDHDaxx-EGFP was constructed using PCR-based accurate synthesis method. After identification by sequencing and enzyme digestion, the recombinant lentiviral vector was contransfected into 293T cells with lentivirus packaging vector. The recombinant lentivirus particles were collected and purified to infect VSMCs, whose expression of Daxx was detected with Western boltting. The cells infected with the empty vector pCDH-EGFP or pCDH-Daxx-EGFP were incubated in serum-free medium or in the presence of angiotensin Ⅱ (AngⅡ). The cell viability was determined with MTT assay, and the cell cycle changes were analyzed with flow cytometry. The cell migration ability was assessed using a scratch wound healing assay. The expression of p-Akt protein in the cells was detected using Western blotting. RESULTS: Double enzyme digestion and sequencing confirmed successful construction of the recombinant plasmid. Compared with the cells infected with the empty vector, the cells infected with pCDH-Daxx-EGFP exhibited significantly increased expressions of Daxx protein ( < 0.05). AngⅡ treatment of the cells infected with the pCDH-Daxx-EGFP, as compared with the cells infected with the empty vector, significantly lowered the cell viability, S phase cell ratio and cell migration ability ( < 0.05), and significantly decreased the expression level of p-Akt protein ( < 0.05). CONCLUSIONS We successfully constructed the recombinant lentiviral vector pCDH-Daxx-EGFP and overexpressed Daxx in primary cultured VSMCs using this vector. Daxx overexpression can inhibit AngⅡ-induced proliferation and migration in VSMCs probably by regulating p-Akt protein.

The theory of "liver opens at the eyes" was first seen in Yellow Emperor's Internal Canon of Medicine， which is the ancient people's summary of the connection between the liver and the eyes. The theory of "liver opens at the eyes" suggests the characteristic of "co-damage and co-recover of liver and eyes". It has been found in clinical practice that liver diseases and eye diseases often occur together， and "liver and eyes co-recover" is an ideal choice. The key to achieving "liver and eyes co-recover" is to analyze its pharmacological material basis and mechanism. With the development of modern medicine， more and more evidence indicates that the liver and eyes have complex and close relationships in physiological and pathological aspects. In a pathological state， there is a phenomenon of "liver and eyes co-damage"， and after the intervention of traditional Chinese medicine， "liver and eyes co-recover" occurs. "Liver and eyes co-damage and co-recover" can be explained through the "co-material basis and co-action mechanism". On this basis， the research group tentatively proposed that the liver and eyes had "four common characteristics" （4CCs）， namely "co-damage， co-recover， co-material basis， and co-action mechanism" from the theoretical connotation of traditional Chinese medicine， clinical practice， and molecular biology. Additionally， the group also took the intervention of Prunella vulgaris， traditional Chinese medicine， for removing liver fire and improving eyesight on immune liver injury （ILI） and allergic conjunctivitis （AC） as examples to analyze 4CCs. This project aims to deeply analyze the scientific connotation of the theory of "liver opens at the eyes"， reveal the common characteristics and biological essence of liver and eyes， explore a new research paradigm of "liver and eyes co-recover"， and provide a reference for the study of common problems of multi-organ associated diseases.

Inflammation is involved in the development of various acute and chronic diseases in the body. Sustained inflammatory responses are key driving factors for diseases such as cancer， neurodegenerative diseases， cardiovascular diseases， metabolic syndrome， inflammatory bowel disease， and arthritis. Therefore， finding anti-inflammatory drugs is crucial for the prevention and treatment of various diseases. In recent years， there has been increasing attention to finding natural drugs with minimal toxic side effects. Lonicerae Japonicae Flos and Lonicerae Flos， as traditional Chinese medicines potent in clearing heat and removing toxins， have strong biological activity and multiple pharmacological effects. They are widely distributed in the plant world and have significant medicinal value. With the continuous advancement of the research on Lonicerae Japonicae Flos and Lonicerae Flos， they have been widely used in the medical field and possess great development potential. Currently， research mainly focuses on the anti-inflammatory mechanisms of Lonicerae Japonicae Flos and Lonicerae Flos， while systematic summaries of their anti-inflammatory active ingredients are rare. Therefore， this paper focuses on the differential analysis of the anti-inflammatory active components of Lonicerae Japonicae Flos and Lonicerae Flos. In addition， it reviewed the possible mechanisms by which extracts and active ingredients of Lonicerae Japonicae Flos and Lonicerae Flos may exert anti-inflammatory effects through various pathways， such as influencing the release of cellular inflammatory factors， regulating inflammatory signaling pathways such as nuclear factor-κB （NF-κB）， mitogen-activated protein kinase （MAPK）， signal transducer and activator of transcription 3 （STAT3）， MAPK/extracellular signal-regulated kinase （ERK）/c-Jun N-terminal kinase （JNK）， phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/NF-κB， and Janus kinase （JAK）/signal transducer and activator of transcription （STAT） signaling pathways， increasing antioxidant stress capacity， enhancing immune defense capabilities， and improving intestinal microbiota， aiming to provide a theoretical basis for the rational clinical application of Lonicerae Japonicae Flos and Lonicerae Flos.

The successful retrieval of ancient mitochondrial DNA (mtDNA) from Neanderthals provides powerful experimental evidence that clarifies the arguments between the out-of-Africa and multiregional models of evolution. However, the lack of nuclear DNA from Neanderthal fossils and mtDNA of early modern human fossils dating back to approximately the same time in the Pleistocene constitutes a limitation that may compromise the significance of mtDNA phylogenetic analysis. In this report, we introduce a mitochromic analysis using Neanderthal mtDNA as a foreign transgene and humans as a naturally occurring transgenic species. Forty Neanderthal mtDNA retrievable nuclear fragments were identified by blasting human genome data with Neanderthal mtDNA. Five of the 40 fragments exhibited higher correlation with Neanderthal mtDNA than those with modern human mtDNA. Furthermore, these five nuclear fragments harbor Neanderthal mtDNA-unique haplotypes. Based on the 98%+ identity between Neanderthal and modern human mtDNA when compared by groups, we suggest that some of the modern human nuclear fragments retrieved using Neanderthal mtDNA may aid in decoding Neanderthal genetic information, and also may simultaneously demonstrate a close genetic evolutionary relationship between modern humans and Neanderthals.