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[Objective] The present study was conducted to clone coat protein gene of citrus tristeza virus (CTY) collected from Gannan navel oranges (south of Jiangxi province) and analyze its sequence genetic information, in order to lay the foundation for preventing CTY from spreading by using mild strains cross protection method. [Method] The CP gene of CTY sample was amplified with RT-PCR and sequenced. The recombination and nucleotide sequence composition of CP gene in different CTY samples were detected by using software. The nucleotide genetic distance among collected CTY samples and 10 completed whole genome sequencing CTY isolates were calculated, and followed by sequence homology comparison and phylogenetic tree construction. [Result] Amongst 214 collected citrus leaf samples, 78 samples were infected with CTY, and the disease incidence was 36.4%. Out of 78 CTY samples, only one sample was found to be has genetic recombination. The full-length sequence of CP gene in 78 CTY samples was 672 bp in length, and there were no any base deletion and insertion. The average GC and AT (U) compositions of CP gene were 52.8% and 47.2%, respectively. The CP nucleotide genetic distance amongst 78 CTY samples were 0-0.072, and those between 78 samples and 10 known isolates were 0-0.106, respectively. The nucleotide and its deduced amino acid of CP gene amongst 78 samples shared 89.1%-100.0% and 92.3%-100.0% of homology, and they shared 76.5%-98.9% and 73.6%-99.5% of homologies with 10 known CTY isolates, respectively. The phylogenetic tree analysis results showed that, 78 samples and 10 known CTY isolates could be divided into four groups. The first group (I) contained 49 collected CTY samples and 7 known isolates (6 virulent strains and 1 mild strain). The second group(II) contained 21 collected samples and 2 known isolates Qaha and Mexico-ctv. The third group (III) only contained sample CRJ14 which came from Ruijin region. The fourth group (IV) contained one known isolate HA16-5 and those collected samples from Jiangkou and Ningdu regions of Jiangxi province. [Conclusion] The CP gene of CTY isolates from Gannan navel oranges is highly conserved, and its sequence has variation at a certain extent. There are higher and complex relationship amongst each collected isolate, and most of them are associated with virulent and stem pitting isolates. [ABSTRACT FROM AUTHOR]

An intact L1 define element is 6 kb in length in human genome. The majority of the L1s is truncated and has direction difference, implying that it is interesting to study the effects of different length and directions of L1s on gene. In this work, 7 different segments were obtained from L1-open reading frame 2 (ORF2), each of which was 280 bp in length. Each segment was connected into 8 repeats in head and tail tandem manner and was inserted to downstream of GFP gene in different directions in pEGFP-C1. The inserted ORF2 segments in the sense orientation caused much stronger inhibition on gene transcription and protein expression than antisense sequences did. Among all segments, the first and ninth 280 bp segments of ORF2 in both orientations induced weaker inhibition on gene transcription than other segments in the same orientations and did not induce transcriptional elongation. The distribution of Alu in most regions of genome was inverse ratio with L1. The inserted Alus in both orientations inhibited GFP gene expression, but the inhibition in antisense orientation was stronger than that in sense orientation and the sense Alu was the sequence of inducing transcription elongation. A-rich of ORF2 was probably the molecular basis of its sense orientation with stronger inhibition on gene expression.

Aim: To observe the effect of rhubarb ethanol-extract on hyperlipidemia and liver fatty in rabbits., Methods: Thirty healthy male white rabbits were divided randomly into five groups, six rabbits in each group. The rabbits in control group were fed with common forage. The rabbits in model group were fed with high lipid forage. The rabbits in three different rhubarb groups were fed with high lipid forage and treated with different level rhubarb ethanol-extract (REE). In the process of experiment, periodically measured serology index of the rabbits and observed common physiology index. The rabbits were killed at the end of tenth week, liver fatty degeneration degree and liver coefficient were measured and compared., Results: REE could decrease serum total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C), and increase serum high density lipoprotein cholesterol (HDL-C), and reduce liver fatty de generation and protect liver cell function. And the dose-effect relation was showed among different dose REE groups., Conclusion: REE can significantly reduce blood lipid, prevent and treat hyperlipidemia and liver fatty.

Aim: To observe the synergic action of monkshood polysaccharide (MPS) and adriamycin (ADM) long circulating temperature-sensitive liposome (ALTSL) in targeting therapy for H22 tumor-bearing mice and explore the mechanism., Methods: The anti-tumor activity was evaluated by using the tumor's weight as an index. The life prolongation rate of mice was calculated according to the survival time of the tumor-bearing mice. The killer activity of NK cells and the lymphocyte transformation rate were detected by the LDH release assay and MTT colorimetry, respectively. The apoptosis of tumor cells and the expressions of p53, Fas, Fas-L and caspase-3 were analyzed by flow cytometry. The expressions of IL-2 mRNA and IL-12 mRNA in spleen lymphocytes were determined by RT-PCR. The pathologic changes of tumor, heart, liver and kidney tissues of the tumor-bearing mice were observed under light microscope., Results: Compared with the adriamycin liposome group, the anti-tumor effects were enhanced in (MPS+ALTSL) group with tumor growth inhibitory rate up to 80.4%. The survival time of the tumor-bearing mice in ALTSL and (MPS+ALTSL) groups was significantly prolonged compared with the ADM group (P<0.01). The killer activity of NK cells was higher in ALTSL group than in the NS and ADM groups, and was highest in (MPS+ALTSL) group. The lymphocyte transformation rate of (MPS+ALTSL) group was markedly increased (P<0.01) as compared with the ADM group. The result of RT-PCR indicated that the expressions of IL-2 mRNA and IL-12 mRNA in lymphocytes in the adriamycin long circulating liposome (ALCL) group were significantly higher than those in the ADM group. Expressions of IL-2 mRNA and IL-12 mRNA was much higher in (MPS+ALTSL) group than in ALTSL group. The pathological examination indicated that in (MPS+ALTSL) group, more lymphocytes and monocytes were found in tumor tissue., Conclusion: ALTSL can increase the anti-tumor effect and decrease the side-effects (such as the cytotoxicity) of ADM. MPS combined with ALTSL can enhance killer activity of NK cells and transformation of T cells, supporting their synergic anti-tumor effect.

Aim: To observe the synergistic role between rhIL-2 and adriamycin long circulating temperature-sensitive liposome (ALTSL) in targeting therapy of H22 tumor-bearing mice and explore their anti-tumor mechanism., Methods: The antitumor activity was evaluated by using the tumor's weight as an index. The prolongation rate of mouse life was calculated according to the survival time of the tumor-bearing mice. The killer activity of NK cells and the lymphocyte transformation rate were detected by the LDH and MTT colorimetry, respectively. The apoptosis of tumor cells and the expression of p53, Fas, Fas-L and Caspase-3 were analyzed by flow cytometry (FCM). The expression of IL-2 mRNA and IL-12 mRNA in splenocytes was determined by RT-PCR. The pathologic changes of tumor, heart, liver and kidney tissues of the tumor-bearing mice were observed under light microscope., Results: The tumoristatic rate of rhIL-2+ALTSL (73.5%) was higher than that of adriamycin liposome (ADML) group (67.0%). The survival time of tumor-bearing mice in ALTSL and rhIL-2+ALTSL groups was significantly extended as compared with the NS group (treated with normal saline) and the free ADM group (P <0.01 or P <0.05). The killer activities of NK cells of ALTSL group and rhIL-2+ALTSL group were higher than those of the NS and free ADM groups, and was highest in rhIL-2+ALTSL group. The lymphocyte transformation rate of ALTSL+rhIL-2 group markedly increased ( P <0.01) as compared with the free ADM group. The result of RT-PCR indicated that the expression of IL-2 mRNA and IL-12 mRNA in splenocytes in the adriamycin long circulating liposome (ALCL) group was significantly higher than that in the free ADM group. The enhancement of rhIL-2+ALTSL on expression of IL-2 mRNA and IL-12 mRNA was much stronger than that of ALTSL alone. The pathological examination indicated that in rhIL-2+ALTSL group, the tumor cells were mostly destroyed, and a large amount of lymphocytes and monocytes were found in tumor tissue., Conclusion: ALTSL can increase the anti-tumor effect and decreased the side-effects (such as the cytotoxicity) of ADM. rhIL-2+ALTSL can induce the apoptosis of tumor cells and enhance killer activities of T cells and NK cells. rhIL-2 and ALTSL can synergistically play the antitumor effect.

Aim: To prepare monoclonal antibody(mAb) against human c-erbB2 and identify its specificity., Methods: The epitope of human c-erbB2 antigen was analyzed by using computer software and a immunodominant epitope at the carboxyl-terminal was selected. A peptide consisting of 13 amino acids was synthesized and coupled with keyholelimpet hemocyanin (KLH), and then it was used to immunize BLAB/c mice. The splenocytes of the immunized mice were fused with Sp2/0 cells routinely and the hybridoma cells were selected by HAT selected culture, indirect ELISA, and immunohistochemical staining, and cloned by limiting dilution. The specificity of the mAb was identified by cross-reaction test and blocking test., Results: A hybridoma cell line SC8C1, stably secreting anti-c-erbB2 mAb was obtained. The mAb SC8C1 could react to breast cancer tissue expressing c-erbB2 molecule but did not react to other c-erbB2-negative cells. The mAb will lose the activity after being blocked with synthesized 13 peptide., Conclusion: A anti-c-erbB2 mAb SC8C1 is prepared successfully using synthesized 13 peptide as immunogen.

Background & Objective: Thalidomide may have curative effect on tumor because of its function of anti-angiogenesis. The aim of this research was to observe the effect of thalidomide on tumor homograft growth in mouse H22 model and to investigate the mechanisms involved and its curative possibility to hepatoma., Methods: BALB/c mice were inoculated subcutaneously with H22 cells. The first treatment group was injected intraperitoneal everyday with thalidomide 50 mg/kg from the day of inoculation and the second treatment group was injected from the fourth day of inoculation. The diameters of the tumors were measured everyday. On the twelfth day, the mice were sacrificed and the tumors were weighed. The microvessel densities of the tumors were measured by immunohistochemical staining with anti-CD31 monoclonal antibody; CD31 expression, apoptosis, and proliferation were analyzed by flow cytometry. Expression of vascular endothelial growth factor (VEGF) mRNA was examined by reverse transcription-polymerase chain reaction (RT-PCR)., Results: The tumor weight of the first treatment group (2.27+/-0.33 g) was decreased compared with that of control group (2.70+/-0.61 g) (P >0.05); the ratio of inhibition was 15.93%. The tumor weight of the second treatment group (3.32+/-0.47 g) was decreased compared with that of control group (3.42+/-0.60 g) (P >0.05); the ratio of inhibition was 2.92%. The microvessel count (48.40+/-12.90) and CD31 expression (5.59+/-0.75) of the first treatment group were significantly decreased (P< 0.05) in comparison with those of control groups (81.20+/-26.91,7.04+/-0.50) (P< 0.05). The microvessel count (46.4+/-9.71) and CD31 expression (7.29+/-0.48) of the second treatment group were not statistically significant (P >0.05) compared with those of control groups (74.0+/-32.69, 6.80+/-0.68) (P >0.05). The apoptosis indices of the two treatment groups (17.20+/-7.80,15.33+/-4.10) were significantly increased, compared with control groups (4.37+/-1.98,4.87+/-1.91) (P< 0.05), while there was no significant difference of VEGF mRNA expression between the two treatment groups and control groups (P >0.05). The relative quantities of VEGF mRNA of the two treatment groups were 0.50+/-0.13 and 0.51+/-0.06 (control groups:0.48+/-0.11 and 0.64+/-0.11) (P >0.05), respectively., Conclusion: Thalidomide can significantly induce apoptosis and inhibit angiogenesis in mouse H22 model when it is used at the beginning of carcinogenesis, but it has no obvious anti-angiogenic efficacy on the grown tumor. Thalidomide cannot inhibit VEGF mRNA expression of grafted H22 tumor in mouse.

Aim: To observe the synergistic inhibitory effects of rhIL-2 and adriamycin magnetic albumin microsphere(ADM-MAM) targeting therapy on tumor and to explore their antitumor mechanism., Methods: The antitumor activity was observed using the tumor weight as index. The killer activity of natural killer (NK) cells and the lymphocyte transformation were examined by the LDH release assay and MTT colorimetry, respectively. The apoptosis of tumor cells and the expressions of p53,Fas and FasL were examined by flow cytometry. The expressions of IL-2 and IL-12 were determined by RT-PCR., Results: Compared with the control group, in ADM-MAM targeting therapy group and combination therapy (ADM-MAM and rhIL-2 therapy) group, tumor weight, killer activity of NK cells, and the lymphocyte transformation;were obviously decreased; while the expression of Fas, FasL, IL-2 and IL-12 were markedly increased., Conclusion: ADM-MAM targeting therapy exerts anti-tumor effects with fewer side effects, which can be enhanced when combined with rhIL-2 therapy. Such synergistic anti-tumor effects may be realized by stimulating the proliferation of T cells and growth of NK cell to enhance cellular immunological function.