With its abilities of trans-synaptic tracing and self-replication and wide host range, pseudorabies virus (PRV) has been applied in the field of neuroanatomy since the 1970s. Four decades of PRV application have made many advances in researches on neuronal tracing with PRV. Mechanism studies focused on investigating infection of primary neurons and tracing direction in secondary neurons, while application studies focused on development of new pathological strains and innovation of tracing techniques. To date, the mechanism and application of viral tracing are not completely figured out yet. Integration of molecular biology technology will improve the efficiency in related researches.
Animals ; Cell Tracking ; Herpesvirus 1, Suid ; genetics ; physiology ; Humans ; Neurons ; virology ; Pseudorabies ; virology

The rate of corneal graft rejection is still high for high-risk keratoplasty although immune suppression drug is routinely used.The role of traditional Chinese medicine in corneal transplantation is concerned gradually.Heijingpaichitang on the prevention and treatment of rats with high-risk corneal allograft rejection needs further study.Objective This study was to investigate the inhibitory effect of heijingpaichitang on high-risk corneal transplantation immune rejection in rats.Methods Sixteen female SD rats were used as the donors and 32female Wistar rats were served as recipients.The high-risk corneal trasplantation models were established by corneal suture in 32 Wistar rats,and then homogeneity variant SD-Wistar corneal transplantation was performed.The recipients were randomized into model control group,cyclosporinc A (CsA)group,heijingpaichitang group and CsA +heijingpaichitang group.CsA,heijingpaichitang and CsA + heijingpaichitang was orally administered 4 days after operation once per day for 15 days,and normal saline solution was used at the same way in the model control group.Ocular anterior segment reaction was examined under the slit lamp and corneal opacification,edema and neovasculation were scored based on Larkin' s criteria.Rejection index of the corneal graft was recorded and the graft survival time was calculated.The pathological examination of the corneal graft was carried out in all rats,and the inflammatory cells in the corneas and CD4+ cells in the periphery blood were assayed using flow cytometry.The use of the animals complied with ARVO Statement.Results Corneal graft rejection occurred in 10 days after operation in the model control group,12-13 days in the CsA group and heijingpaichitang group and 22 days in the CsA +heijingpaichitang group.Compared with model control group,the scores of the corneal opacification,corneal edema and neovascularization were significantly lower in the CsA group,heijingpaichitang group and CsA+heijingpaichitang group (P＜0.05),and all the scores were declined in the CsA+ heijingpaichitang group compared with CsA group and heijingpaichitang group(P＜0.01),but no significant differences were seen in the scores between the CsA group and heijingpaichitang group(P＞0.05).The mean survival time of grafts was (10.38 ±1.69)days in the model control group,(22.50 ± 3.07) days in the CsA + heijingpaichitang group,with the significant difference (t =-9.790,P =0.000).The pathological examination of graft showed that the lymphocytes and new blood vessels were less in the CsA+heijingpaichitang group compared with CsA group and heijingpaichitang group 15 days after operation.Flow cytometry verified that the number of lymphocytes in graft,CD4+cells and CD4+/CD8+ in periphery blood were significantly lower in the heijingpaichitang group,CsA group and CsA+heijingpaichitang group compared with model control group (P＜0.05).Conclusions Heijingpaichitang can inhibit immune rejection to certain extent in high-risk corneal transplantation rat and has a similar effect to 0.1％ CsA.Heijingpaichitang and 0.1％ CsA have a synergistic effect.

The aerolysin genes (aerA) of BZ and NK isolates were cloned and sequenced. The sequence analysis showed that the partial aerA of BZ and NK isolates consisted of 1393 bp, encoding a protein of 464 amino acids. The similarity of nucleotide and amino acid sequences of aerA between BZ and NK isolates was 97.6% and 98.3% respectively. The nucleotide sequence of aerA of BZ strain exhibited 71.6% to 97.5% homology with other Aeromonas isolates, and the amino acid sequence exhibited 68.0% to 98.9% homology. The phylogenetic tree based on aerA nucleotide sequences from Aeromonas isolates was constructed with neighbor-joining method. It showed that there were three branches of aerolysin genes, and a close relation- ship among Aeromonas hydrophila isolates which were clustered into the same branch.

Objective To explore the effect of vascular endothelial growth factor antisense oligonucleotides(AS-VEGF)on HL60 cell and HL60/VCR multidrug resistance cell and analyze the function of P-gp and the expression of related multidrug resistance genes including Bcl-2,Mcl-1,MDR1,MRP,GST?,TopoⅡ? and TopoⅡ?.Methods A vector AS-VEGF which expressed in eukaryotic cell was established,then transfected the vector into HL60 and HL60/VCR by limposome transfection technology,observed and drew the growth curve by Tapanlan taining,RT-PCR was used to detect the expression of Bcl-2,Mcl-1,MDR1,MRP,GST?,TopoⅡ? and TopoⅡ? in mRNA level after transfected 24 h and 48 h.Western Blot was used to detect the expression of P-gp in proteinum level after transfected 24 h and 48 h.Results The growth of HL60 and HL60/VCR was inhibited by AS-VEGF(1.25 mmol/L).Between HL60 and HL60/VCR,AS-VEGF decreased the expression of MDR1,MRP,GST? and TopoⅡ? but could not influence the expression of Bcl-2,Mcl-1 and TopoⅡ?,and the expression of P-gp was also obviously decreased in 48 h compared with that in 24 h.Conclusions AS-VEGF can inhibite the growth of HL60 and HL60/VCR and reverse multidrug resistance by changing cell microenvironment and the cell membrane correlated protein transportating channel,reduce the cell disintoxicating and the self-repair ability.

OBJECTIVEAn RP-HPLC procedure was established for the determination of danshensu and protocatechuic aldehyde in Guanxinning injection powder.

METHODAn RP-HPLC analytical procedure was developed using Hypersil ODS2 C18 column (4.6 mm x 250 mm, 5 microm) with methanol-0.5% glaceal acetic acid (4.5:95.5) as mobile phase, and a wavelength of 280 nm for UV detection.

RESULTThe linear range was 3.004-45.06 microg x m(L-1) (r = 0.999 5, n = 6) for danshensu, and 0.300-4.509 microg x mL(-1) (r = 0.999 3, n = 6) for protocatechuic aldehyde. The average recoveries were 99.1% and 97.9%, respectively.

CONCLUSIONThe method was stable, accurate, and reproducible, and can be used for the quality control of Guanxinning injection powder.

Benzaldehydes ; analysis ; Catechols ; analysis ; Chromatography, High Pressure Liquid ; methods ; Drug Combinations ; Drugs, Chinese Herbal ; administration & dosage ; chemistry ; isolation & purification ; Injections ; Lactates ; analysis ; Ligusticum ; chemistry ; Plants, Medicinal ; chemistry ; Powders ; Quality Control ; Reproducibility of Results ; Salvia miltiorrhiza ; chemistry

OBJECTIVEThe hypoxic-ischemic encephalopathy caused by asphyxia in peripartum is a serious disease in newborn infants, with a high disability and mortality rate. Lack of regenerative ability in central nervous system after injury is considered as the fundamental cause. However, in recent years many studies have revealed that there are myelin-associated neurite growth inhibitory factors that exert inhibiting effect through the Nogo receptor (NgR). This study aimed to investigate the expression level of NgR and the possible neuroprotective effect of NEP1-40 in newborn rats with hypoxic ischemic brain damage (HIBD).

METHODEighty healthy Wistar rats aged 7 days were randomly divided into 4 groups; 8 in control group, 24 in HIBD model group, 24 in GM-1 group and 24 in NEP1-40 group. The rats of the control group and HIBD group were injected with normal saline (0.25 ml/kg) intraperitoneally, while those in NEP1-40 group and GM-1 group with NEP1-40 12.5 microg/d, GM-1 10 mg/(kg.d) for continuous 3 days of 72-hour group or 7 days of 168-hour group, respectively. In situ hybridization was adopted for detecting the expression of NgR in the brain of the rats at the time point of 24 hours, 72 hours and 7 days. Meanwhile histopathological changes of neurons and axon were detected by transmission electron microscopy (TEM). The SPSS statistical software package for Windows, version 10.0, was used to run Chi-square tests and least significance difference (LSD-t) on the data presented, and P value of less than 0.05 was regarded as statistically significant.

RESULTThe expression level of Nogo-A receptor in the control group was higher than that of the other groups at different time point (t value was 5.48, 6.11, 6.96, 8.24, 5.99 and 5.34, respectively, and all P values were less than 0.05). There were no significant differences in Nogo-A receptor level among the HIBD group, the GM-1 group and the NEP1-40 at 24 hours (t was 1.48, 2.76 and 1.29, respectively, and all P > 0.05), while the expression of Nogo-A receptor of NEP1-40 at 72 hours and 7 days was lower than that of the HIBD group and the GM-1 group at the same time point, respectively (all P < 0.05). Repair of neurons in damaged brain to some extent was found after GM-1 treatment and satisfactory repair of neurons and axon regeneration was obtained with NEP1-40 administration as shown by TEM.

CONCLUSIONHypoxic ischemic brain damage can down-regulate the expression of Nogo-A receptor in the central nervous system. NEP1-40 contributes to the regeneration of axon and repair of brain damage, thus exerts neuroprotective effect.

Animals ; Animals, Newborn ; Brain ; drug effects ; metabolism ; pathology ; GPI-Linked Proteins ; Hypoxia-Ischemia, Brain ; metabolism ; pathology ; Myelin Proteins ; pharmacology ; Nogo Receptor 1 ; Peptide Fragments ; pharmacology ; Rats ; Rats, Wistar ; Receptors, Cell Surface ; Receptors, Peptide ; metabolism

OBJECTIVETo establish chemical pattern recognition method for the identification and evaluation of Atractylodes macrocephala.

METHODThe chemical constituents in methanol extract of 32 samples of A. macrocephala were determined by HPLC. The fingerprints were obtained and were handled by hierarchical clustering analysis and stepwise discriminant analysis.

RESULT AND CONCLUSIONAccording to the result of classification, all samples collected were devided into three Grades--the superior, the ordinary and the fake. Chemical pattern recognition method was established. It may be of practical value for the quality control of A. macrocephala.

Atractylodes ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Cluster Analysis ; Pattern Recognition, Automated ; Plants, Medicinal ; chemistry ; Quality Control ; Rhizome ; chemistry

OBJECTIVEPPAR-gamma is associated with the differentiation, apoptosis, proliferation and cytokine secretion of immunologic cells. This study investigated peripheral blood lymphoblastic PPAR-gamma mRNA expression and its correlation with plasma IL-13 contents in children with acute idiopathic thrombocytopenic purpura (ITP).

METHODSFifty-three children with acute ITP who were in line with the standard test between September 2007 and July 2008 were enrolled. Fifty healthy children during the same period were used as the control group. PPAR-gamma mRNA expression in peripheral blood lymphocytes were detected by RT-PCR. Plasma IL-13 contents were detected using ELISA.

RESULTSPPAR-gamma mRNA expression in peripheral blood lymphocytes from acute ITP children were significantly higher than that in the control group (0.78 +/- 0.03 vs 0.52 +/- 0.05; P< 0.05). Plasma IL-13 contents in children with acute ITP were also significantly higher than those in the control group (160.21 +/- 34.26 pg/mL vs 121.42 +/- 12.69 pg/mL; P< 0.05). There was a positive correlation between plasma IL-13 level and lymphoblastic PPAR-gamma mRNA expression in children with ITP (r=0.89, P< 0.05).

CONCLUSIONSPPAR-gamma mRNA expression in peripheral blood lymphocytes increased and were positively correlated with plasma IL-13 contents in children with acute ITP, suggesting that PPAR-gamma and IL-13 might participate in the pathogenesis of acute ITP.

Acute Disease ; Child ; Child, Preschool ; Female ; Humans ; Interleukin-13 ; blood ; physiology ; Lymphocytes ; metabolism ; Male ; PPAR gamma ; genetics ; physiology ; Purpura, Thrombocytopenic, Idiopathic ; etiology ; immunology ; RNA, Messenger ; analysis

To develop a GFP transgenic cell model under the transcriptional control of TK promoter adjacent to which ARE enhancer was inserted. Synthetic oligonucleotide ARE motif was annealed and purified then inserted into pTK-GFP to construct the vector of pARE-TK-GFP. The TK and ARE-TK fragments were amplified by PCR and cloned into pEGFP-N1 to reconstruct eukaryotic expression vectors of pTK-GFP/Neo and pARE-TK-GFP/Neo. They were transfected into HepG2 cells and clones resistant G418 were isolated. PDTC and Lentinan were used to induce the cell levels of GFP and the fluorescence was measured using a fluorescence plate reader. The results showed that the induced level of GFP is significantly increased and have dose-dependeny in a certain range. This findings indicated that such a cell model offered a potential platform for preliminary screening of all kinds of natural or synthetic chemopreventive agents.
Antineoplastic Agents ; pharmacology ; Base Sequence ; Dose-Response Relationship, Drug ; Drug Screening Assays, Antitumor ; methods ; Enhancer Elements, Genetic ; genetics ; Gene Expression ; drug effects ; Gentamicins ; pharmacology ; Green Fluorescent Proteins ; genetics ; metabolism ; Hep G2 Cells ; Humans ; Lentinan ; pharmacology ; Microscopy, Fluorescence ; Molecular Sequence Data ; Oligonucleotides ; genetics ; Proline ; analogs & derivatives ; pharmacology ; Recombinant Fusion Proteins ; genetics ; metabolism ; Thiocarbamates ; pharmacology ; Transfection

Objective To study the decomposition kinetics of omethoate in blood.Methods The acetonitrile precipitated protein was added into the blood, with the chromatographic column of a Waters BEH C18column (2.1 mm×50 mm, 1.7μm), the mobile phase of 5 mmol/L ammonium acetate aqueous solution-methanol, and the gradient elution with a flow rate of 0.3 mL/min and injection volume of 2μL.With electrospray ionization (ESI) source and positive ion detection, qualitative and quantitative analyses were taken using multi-reaction monitoring mode.Omethoate standard was added into blank human blood to the mass concentrations of 0.78, 1.40, 2.30, 4.50, and 7.20μg/mL, and each mass concentration was preserved at 3 temperatures of-20℃, 4℃, and 20℃, respectively.The content of omethoate was detected at different time points (0, 1, 3, 4, 7, 11, 15, 24, 32, 40, 48, 64, 80, 96, and 120 d).Results Different concentrations of omethoate all showed a descended trend in human blood under different temperature conditions.The decomposition in storage environment of-20℃, 4℃, and 20℃was fit to a one-compartment open model with a first-order kinetic process, which could be expressed as Ct=Coe-αt, with the calculated theoretical values of omethoate concentration close to the measured values.Conclusion All concentrations of omethoate are decomposed in the blood, which vary a lot in different preservation conditions.It is suggested that blood samples should be frozen and detected timely in suspected omethoate poisoning cases.