Objective To explore the clinical efficacy and side effect of hepatocyte growth - promoting factor in treatment of infant hepatitis syndrome. Methods Sixty one cases of infant hepatitis syndrome were chased as the treatment group who hospitalized in our hospital from March 2002 to Feb 2003,and 54 cases of infant hepatitis syndrome as control group who hospitalized during March 2001 to Feb 2002. The treatment group were administrated with hepatocyte growth - promoting factor for 2 weeks. We obser-ved the recovery of patient's liver function (TBIL.ALT, AST) and the side effect of hepatocyte growth- promoting factor after two weeks of the treatment. Results After the treatment,TBIL and ALT decreased significantly in the treatment group of infant hepatitis syndrome. The treatment group was superior to the control group (P

NAC transcription factors involved in plant growth and development, as well as responses to biotic and abiotic stress. RNAi Vectors for SmNAC transcription factors of Salvia miltiorrhiza was constructed by using Gateway cloning technology, in order to further study the function of SmNAC1 transcription factor. According to Gateway cloning technology, the specific fragments of SmNAC1 containing attB adapter was amplified by PCR using ultra-fideling phusion polymerase of NEB. By the BP recombination reaction, the PCR product containing attB was transferred to an donor vector (pENTR/SD/D-TOPO). Finally, SmNACi specific gene was cloned into pK7GWIWG2D plant expression vectors by LR recombination reaction. Experimental results showed that Gateway cloning technology provide a rapid and highly efficient way to clone the interested gene.
Cloning, Molecular ; methods ; Genetic Vectors ; genetics ; Plant Proteins ; genetics ; Polymerase Chain Reaction ; RNA Interference ; Reproducibility of Results ; Salvia miltiorrhiza ; genetics ; Transcription Factors ; genetics

According to the designed specific primers of gene fragment based on the Salvia miltiorrhiza transcriptome data, a full-length cDNA sequence of SQS2 from S. miltiorrhiza f. alba was cloned by the method of reverse transcription polymerase chain reaction (RT-PCR). The SmSQS2 cDNA sequence was obtained, this sequence is named SmSQS2 and its GenBank registration number is KM244731. The full length of SmSQS2 cDNA was 1245 bp, encoding 414 amino acids including 5'UTR 115 bp and 3'UTR 237 bp. Sequence alignment and phylogenetic analysis demonstrated that SmSQS2 had relative close relationship to the SQS2 of S. miltiorrhiza. The induction of E. coli [pET28-SQS2] in different temperature, induction time, IPTG concentrations and density of inducing host bacterium (A600) were performed, Shaking the culture at 30 degrees C until the A600 is approximately 0.6 and add IPTG to final concentration of 0.2 mmol x L(-1), and then the optimal expression of SmSQS2 recombinant protein were accumulated after the induction time of 20 h. The research provided important base for the study of sterol and terpene biosynthesis of SQS2 in S. miltiorrhiza f. alba.
Cloning, Molecular ; Farnesyl-Diphosphate Farnesyltransferase ; chemistry ; genetics ; metabolism ; Models, Molecular ; Molecular Sequence Data ; Phylogeny ; Plant Proteins ; chemistry ; genetics ; metabolism ; Salvia miltiorrhiza ; chemistry ; classification ; enzymology ; genetics ; Sequence Alignment

AIMTo study the characteristics of the interaction between insulin and liposome.

METHODSThe interaction between insulin and liposome was studied by fluorescence spectra and microcalorimetry methods. The sample of insulin liposome interaction after separation by supper-centrifugalization or gel filtration was determined by fluorescence and HPLC.

RESULTSThe results indicate that there was only little increase in fluorescence intensity and no blue shift of peak in fluorescence spectrum. Fluorescence quenching experiments with NaI and acrylamide as quenchers showed that the KSVs (the slope of Strm-Volmer equation) of insulin were more similar to that with added liposome, indicating low interaction between insulin with liposome. The microcalorimetric results indicate that the heat released during the mixture of liposome with insulin, was 1.98 kcal.mol-1, suggesting that the reaction belongs to weak reaction. The quantity of insulin in the insulin-liposome mixture sample after separation by ultracentrifuge or by Sephadex G-75 determined by HPLC, the combination percent was only 0.2%, indicating low interaction between insulin and liposome.

CONCLUSIONThe interaction between insulin and liposome was weak.

Chemistry, Pharmaceutical ; Chromatography, High Pressure Liquid ; Drug Carriers ; Drug Delivery Systems ; Hypoglycemic Agents ; administration & dosage ; chemistry ; Insulin ; administration & dosage ; chemistry ; Liposomes ; Spectrometry, Fluorescence

This study was conducted to investigate the in vitro characteristics of cationic liposomes composed of 3beta-[N-[2-(N', N'-dimethylamino) ethyl] carbamoyl] cholesterol (DC-Chol) and dipalmitoylphosphatidylcholine loaded with doxorubicin (DXR), and to provide useful information for the in vivo tumor vascular targeting of cationic liposomes. Cationic liposomes composed of different amounts of DC-Chol (0 mol%, 10 mol%, 25 mol%, 50 mol%) were loaded with the conventional anti-cancer drug doxorubicin. Their size, zeta potential, encapsulation efficiency, and DXR release in vitro were investigated. Moreover, their uptake by rat aortic endothelial cells (RAECs) was observed at 15 min, 30 min, 1 h, and 4 h of incubation. FITC-Dextran was i.v. injected to the H22 tumor-bearing KM mice to stain the neovasculature. The characteristics of resulting DXR-loaded cationic liposomes were in stable characteristics with particle sizes around 100 - 200 nm and capsulation efficiency greater than 90%. Increased cationic lipid led to enhanced zeta potential, and meanwhile it also resulted in quick release of the loaded drug, indicating increased slits or pores on the membrane upon the addition of DC-Chol. RAECs could more avidly take up DXR-loaded cationic liposomes when the content of DC-Chol increased in the liposomes, and DXR were quickly released in the cytoplasm and transported to the nuclei. The neovasculature stained by FITC-Dextran was clearly observed. DXR-cationic liposomes composed of DC-Chol could be used for tumor vascular targeting in vivo for further study.
Animals ; Aorta ; cytology ; Cell Line, Tumor ; Cholesterol ; analogs & derivatives ; chemistry ; pharmacokinetics ; Doxorubicin ; administration & dosage ; pharmacokinetics ; Drug Carriers ; Drug Delivery Systems ; Endothelial Cells ; metabolism ; Female ; Liposomes ; chemistry ; pharmacokinetics ; Liver Neoplasms, Experimental ; pathology ; Male ; Mice ; Neovascularization, Pathologic ; metabolism ; Particle Size ; Rats ; Rats, Sprague-Dawley

A cationic liposome was prepared with a ligand directed at folate receptor in cancer cells to improve selectivity and facilitate its access to the cancer cells. Folate-polyethyleneglycol-distearoylphosphatidylethanolamine (F-PEG-DSPE) was synthesized by reacting folic acid (F), polyoxyethylene-bis-amine (NH2-PEG-NH2), succinic anhydride (SUC) with distearoylphosphatidylethanolamine (DSPE). Folate receptor-targeted liposomes composed of DPPC/DC-Chol/F-PEG-DSPE (10:10:0.75, molar ratio) were prepared by film dispersion method. A negative charged dextran fluorescein anionic (DFA) was used as a model to explore the in vitro properties and cell uptake efficiencies of liposomal DFA on KB and HpeG2 cells. The formulations were investigated by orthogonal experiment using encapsulation efficiency as the optimized indexes. The size, 4 potential, entrapment efficiency and DFA release in vitro were investigated. The results showed that DFA loaded folate receptor-targeted liposomes had high encapsulation efficiency and the mean size approximately 144 nm. The cationic liposomes had some toxicity to HepG2 cells, and at low concentration (0.0125-0.1 micromol x L(-1)) , the toxicity was linear with the concentration of DC-chol. The folate receptor-targeted liposomes showed great effects on increasing liposome cellular uptake of DFA. In summary, the method of film dispersion method is suitable for producing DFA loaded lipsomes with high entrapment efficiency, small size and slow release. The folate receptor-targeted liposomes can efficiently deliver DFA into cells in vitro. This may represent a promising option for researches on cancer gene therapy.
Carrier Proteins ; metabolism ; Dextrans ; chemistry ; Drug Carriers ; Drug Delivery Systems ; Fluoresceins ; chemistry ; Folate Receptors, GPI-Anchored ; Folic Acid ; chemistry ; Genetic Therapy ; Hep G2 Cells ; Humans ; KB Cells ; Liposomes ; administration & dosage ; chemical synthesis ; metabolism ; Particle Size ; Phosphatidylethanolamines ; chemistry ; Polyethylene Glycols ; chemistry ; Receptors, Cell Surface ; metabolism ; Transfection

The paper is aimed to study the enzymatic hydrolysis of the activatable cell-penetrating peptide (ACPP) that was designed and synthesized. The ACPP was composed of three parts, polyanionic sequence peptide, peptide sequence that specifically cleaved by matrix metalloproteinase (MMP) and cell penetrating peptide (CPP). The ACPP was hydrolyzed by type IV collagenase (MMP-2/9) under the condition of 37 degrees C and was monitored by reversed-phase high performance liquid chromatography (RP-HPLC). The efflux of peak was collected and detected by matrix assisted laser desorption ionization orthogonal time of flight mass spectrometry (MALDIO-TOF-MS) to speculate the sequences of the peptide fragments. The results indicated that the ACPP could be cleaved by type IV collagenase at target site as predicted, released CPP. The half life of the cleavage was about 4 h. Meanwhile, the peptide fragments may be cleaved again at other sites by type IV collagenase.
Amino Acid Sequence ; Cell-Penetrating Peptides ; chemical synthesis ; chemistry ; Chromatography, High Pressure Liquid ; Hydrolysis ; Matrix Metalloproteinase 2 ; chemistry ; Matrix Metalloproteinase 9 ; chemistry ; Peptide Fragments ; chemistry ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Cell-penetrating peptide (CPP) can be used in pharmaceutics as a highly efficient drug delivery transporter. In this study, four tumor cell lines (MCF-7, MDA-MB-231, C6, and B16F10) were used to observe the uptake of fluorescein isothiocyanate (FITC) labeled CPP and the effects of time and concentration of CPP on cell penetration was studied. The CPP exocytosis on C6 cell line was observed, and its exocytosis kinetics was described by zero order equation. In addition, low-temperature condition (4 degrees C) and endocytosis inhibitors were utilized to investigate the mechanism of CPP uptake by cells. Low-temperature condition did not show significantly inhibition on CPP uptake. Heparin, a membrane glycoprotein receptor inhibitor, showed strong inhibition effect (only 3%-10% of the control) on CPP uptake. Chlorpromazine, chloroquine and 5-(N-ethyl-N-isopropyl)-amiloride (EIPA) showed little effect on CPP uptake. This study indicated that CPP penetration had little selectivity on cell type, but the amount and rate of CPP penetration into cells were related to the type of cell lines. The adsorption of CPP on cell membrane induced by sulfate proteoglycan plays an important role on CPP penetration.
Adsorption ; Amiloride ; analogs & derivatives ; pharmacology ; Cell Line, Tumor ; Cell Membrane ; metabolism ; Cell-Penetrating Peptides ; administration & dosage ; metabolism ; pharmacokinetics ; Chloroquine ; pharmacology ; Chlorpromazine ; pharmacology ; Dose-Response Relationship, Drug ; Exocytosis ; Heparin ; analogs & derivatives ; metabolism ; pharmacology ; Humans ; Proteoglycans ; metabolism ; Temperature ; Time Factors

Cell-penetrating peptides (CPPs) offer a non-selective and receptor-independent mode to promote cellular uptake. Although the non-specificity of CPP-mediated internalization allows this approach applicable to a wide range of tumor types potentially, their universality is a significant obstacle to their clinical utility for targeted delivery of cancer therapeutics and imaging agents. Accordingly, many reports have focused on selective switching of systemically delivered inert CPPs into their active form in lesions (tumor). In this review, our attention is mainly confined to such an enzyme-sensitive domain incorporated delivery system with activatable CPPs (ACPPs), which have displayed the exciting strength in balancing the CPPs' pros and cons, and potential in the treatment and diagnosis of some diseases.
Cell-Penetrating Peptides ; chemistry ; Drug Delivery Systems ; Enzymes ; chemistry ; Humans ; Neoplasms ; drug therapy

This study was performed to investigate the chemical constituents in the twigs and leaves of Harrisonia perforate. Six compounds were isolated from the 95% EtOH extract of the twigs and leaves of Harrisonia perforate by silica gel, ODS, Sephadex LH-20 column chromatographies and preparative HPLC. On the basis of chemical properties and spectra data, these compounds were identified as harriperfin E (1), kihadanin A (2), kihadanin B (3), 6α-acetoxyobacunol acetate (4), gardaubryone C (5), and β-sitosterol methyl ether (6), respectively. Compound 1 is a new chromone, and compounds 2-6 are isolated from this plant for the first time.
Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; Phytochemicals ; chemistry ; isolation & purification ; Plant Leaves ; chemistry ; Simaroubaceae ; chemistry