The enhancement of DNA binding ability of a mutated E2 (A338V) protein of HPV-2 / 病毒学报

HPV-2 is a very common type of HPV which causes common warts. The E2 protein of virus can repress the activity of the viral early promoter through binding to the specific binding sites in viral LCR. Previously we reported that the repression of a mutated E2 protein of HPV-2 isolated from a patient with huge common wart on the viral early promoter was obviously decreased, and A338V mutation located at the C terminal DNA binding region of E2 protein. In this study, we expressed and purified the recombinant mutated and prototype E2 fusion proteins, both in the contexts of the C terminal and the full length, by prokaryotic expression system. The electrophoretic mobility shift assay showed E2 protein could bind to double-stranded DNA oligos labeled with biotin that covered two E2 binding sites. The DNA binding abilities of both C terminal and full-length mutated E2 proteins were stronger than the prototype analogs. This result indicates that the enhancement of the mutated E2 DNA binding ability may be the molecular mechanism for its impact on the activity of viral promoter, which correlates with the phenotype of extensive common wart.

Inactivation of bacterial spores using low-temperature plasma / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of low-temperature plasma on inactivation of bacterial spores and explore the mechanism.</p><p><b>METHODS</b>Dielectric barrier discharge (DBD) was employed to generate the atmospheric low-temperature plasma for treatment of B.subtilis var. niger spores with the gas spacing of 3, 4 and 5 and treatment time intervals of 5, 10, 15, 20, 25, 30 and 35 s. The survived colonies was counted with plate counting method, and the killing log value (KLV) at different treatment times was calculated. The inactivation effect of electric field on B.subtilis var.niger spores was also investigated and the spores treated with low-temperature plasma were observed with transmission electron microscope.</p><p><b>RESULTS</b>With the gap spacing of 3, 4 and 5 mm, the KLV of low-temperature plasma on B.subtilis var.niger spores within 25, 30 and 35 s of exposure was more than 5. The germicidal effects of the electric field on B. subtilis var.niger spores were rather poor. Transmission electron microscopy demonstrated total destruction of the surface and interior structure of the spores by low-temperature plasma.</p><p><b>CONCLUSIONS</b>Low-temperature plasma is effective for inactivation of the bacterial spores with a time and dose dependence. The penetrating effect of charged particles and oxygenation effect of the reactive oxygen species might play a dominant role in plasma-induced bacterial spore inactivation, while the role of electric field is negligible.</p>

Oral acute toxicity of (+)-usnic acid in mice and its cytotoxicity in rat cardiac fibroblasts / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe the oral acute toxicity of of (+)-usnic acid in mice and assess its cytotoxicity in rat cardiac fibroblasts.</p><p><b>METHODS</b>The mice with acute poisoning of (+)-usnic acid at different doses by oral administration were observed for toxic manifestations, and the LD(50) was determined. The survival time and survival rate of the mice receiving different doses of (+)-usnic acid were observed. Cultured rat cardiac fibroblasts were inoculated with different concentrations of (+)-usnic acid, and the cell growth inhibition rate was estimated and the IC(50) determined using MTT assay.</p><p><b>RESULTS</b>Higher dose of (+)-usnic acid resulted in more obvious symptoms of poisoning and shorter survival time of the mice. The LD(50) of (+)-usnic acid in mice by oral administration was 388 mg/kg. The manifestations of poisoning such as apathism, pilomotor, chill, dyspnea, torpidity and anorexia was observed. Rat cardiac fibroblasts incubated with (+)-usnic acid showed obvious growth inhibition, which was positively correlated to the dose of (+)-usnic acid, and high dose of (+)-usnic acid caused severe cell injuries. The IC(50) of (+)-usnic acid in rat cardiac fibroblasts was 322 microg/ml.</p><p><b>CONCLUSION</b>(+)-usnic acid is a natural compound of low toxicity in mice, and low to medium dose of (+)-usnic acid dose not produce obvious cytotoxicity.</p>

Molecular epidemiological study on prevalence of human papillomaviruses in patients with common warts in Beijing area / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To study the circulation, distribution, and genomic diversity of HPVs in common warts in Beijing area of China.</p><p><b>METHODS</b>Forty eight patients with pathologically diagnosed common warts were screened for the presence of HPV with HPV type-specific PCR and direct sequencing analysis. The genomic diversity of HPVs prevalent in Chinese patients was analyzed based on LCR.</p><p><b>RESULTS</b>Forty one (85.5%) samples were positive for HPV DNA, 13 (31.7%)--HPV-57, 12 (29.3%)--HPV-1a, 7 (17%)--HPV-27 and 5(12.2%)--HPV-2a. Four cases were infected with two different HPV types, two (4.9%) with HPV-1a and HPV-27, one (2.4%) with HPV-1 and HPV-57 and one (2.4%) with HPV-27 and HPV-57. In contrast to the prevalence of single strain of novel HPV-57 variant and HPV-1 prototype, two HPV-2 and three HPV-27 novel variants were found to circulate in Beijing.</p><p><b>CONCLUSION</b>HPV-1, -2, -27 and -57 are predominantly prevalent in patients with common warts in Beijing.</p>

Transcriptional repressive activity of mutated E2 protein of human papillomavirus 2 (HPV-2) variant / 病毒学报

Common warts are close associated with HPVs infection. In this study, we amplified and sequenced the LCR fragment and E2 gene of HPV-2 that infected the patient of extensive common wart with cutaneous horns, and we constructed the recombinant CAT-reporter plasmids pBLCAT-LCR containing HPV-2 prototype or variant LCR and mammalian expression plasmids pcDNA3. 1-E2 containing prototype or variant E2 ORF individually. The promoter activities of HPV-2 variant and the transcriptional repression activities of the mutated E2 protein were evaluated by transient transfection into HeLa cells. The results showed that there were several mutations in LCR and E2 gene of HPV-2 variant. Compared with the prototype, the viral early promoter activity of variant was significantly increased uder the control of LCR. Compared with the wild type E2 protein, the transcriptional repression activities of the mutated E2 protein was abolished partially. We speculate herein that increased promoter activities and decreased repression effect of the mutated E2 protein are linked, at least partially, with the clinical phenotypes of the uncommon huge common wart.

Bactericidal activity of GLL-37, a novel derivative of the human antimicrobial peptide LL-37 / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To develop and investigate GLL-37, a substitution analogue of the human antimicrobial peptide LL-37 with anti-enzymatic degradation activity and improved efficacy.</p><p><b>METHODS</b>The bactericidal activities of LL-37 and newly developed GLL-37 against 6 Gram-negative and -positive bacteria were determined by Broth microdilution assays. The minimum inhibitory concentrations of LL-37 and GLL-37 against E.coli ATCC 25922 in different NaCl concentration medium were also detected. Both peptides were co-incubated with elastase, and then analyzed by PAGE electrophoresis and bactericidal activity determination.</p><p><b>RESULT</b>GLL-37 showed a stronger elastase resistance ability than LL-37, and was significantly more effective than LL-37 under high-salt condition.</p><p><b>CONCLUSION</b>The antimicrobial peptide GLL-37 derived form LL-37 has the potential as a new therapeutic agent for bacterial infections.</p>

Analgesic effect of interleukin-2 in mouse models of spared nerve injury / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the analgesic effect of interleukin-2 (IL-2) in mice with spared nerve injury (SNI).</p><p><b>METHOD</b>IL-2 was intraperitoneally injected in mice with induced SNI, and von Frey Filaments test and cold plate test were carried out to accesses the analgesic effects of IL-2 and the effect of naloxone in antagonizing the effects of IL-2.</p><p><b>RESULTS</b>IL-2 produced analgesic effects against hyperalgesia and allodynia in mouse models of SNI, and the effect of IL-2 lasted for more than 24 h, showing a double-peak pattern in its action with the two peaks occurring at 30 and 105 min, respectively. The effect of IL-2 could be significantly antagonized by naloxone.</p><p><b>CONCLUSIONS</b>IL-2 has long-lasting analgesic effects in mouse models of SNI model, showing a double-peak pattern of its action. The analgesic effect of IL-2 is probably mediated by opiate receptor.</p>

Study on the relationship between level of CD58 expression in peripheral blood mononuclear cell and severity of HBV infection / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>As one of the intercellular adhesion molecules, CD58 plays important roles in promotion of the adhesion between T cells and target cells, hyperplasia, activation of T cells and natural killer cells, and balance between Th1 and Th2. We studied the relationship between the levels of CD58 expression in peripheral blood mononuclear cells (PBMCs) and severity of HBV infection.</p><p><b>METHODS</b>The levels of CD58 mRNA in PBMCs were detected using quantitative reverse transcription PCR. The percentage of CD58 positive cells was detected by flow cytometry in patients and healthy controls.</p><p><b>RESULTS</b>The levels of CD58 mRNA and the percentage of CD58 positive cells in patients infected with HBV were significantly higher than that in the control. Based on severity of HBV infection, the patients were classified into four groups. The expression of CD58 increased significantly in an order from mild chronic, moderate chronic, severe chronic to severe hepatitis groups. The levels of CD58 mRNA and the percentage of CD58 positive cells in PBMCs from patients with HBV infection were both positively correlated with serum levels of ALT and AST.</p><p><b>CONCLUSION</b>The level of CD58 expression is related with the severity of HBV infection and the degree of liver damage.</p>

Effects of allogeneic hepatic nonparenchymal cell on the survival of grafted skin in mice / 中华烧伤杂志

<p><b>OBJECTIVE</b>To investigate the effect of allogene hepatic nonparenchymal cell (NPC) on the survival of grafted skin in mice and its underlying mechanism.</p><p><b>METHODS</b>Sixty-five C(3)H and fifty-eight C(57)BL/6 mice were employed in the study. Twenty C(3)H mice were used as skin donor and forty as the source of hepatic NPC. The rest five served as the stimulators of mixed lymphocyte culture (MLC) before and on the 7th, 18th, 30th, and 60th day after NPC infusion with 1 at each time point. MLC was determined and expressed as count per minute (CPM). Fifty-eight C(57)BL/6 mice were further divided into experimental (E, n = 50) and control groups (C, n = 8). The mice in C group only underwent skin grafting without NPC infusion. The mice in E group received with 2 x 10(7) NPC via caudal vein, followed by peritoneal injection of cytoxan (200 mg/kg) 48 hours later; They were grafted with skin donated from C(3)H mice 18 days after injections. The survival time of the mice in the two groups was observed. The serum levels of interleukin-4, chimera and MLC in the two groups were determined before and on 7th, 18th, 30th, 60th days after NPC infusion, and micro-chimera were aslo assessed on the 1st and 3rd day after NPC infusion. Five mice were sacrificed at each time point.</p><p><b>RESULTS</b>The survival time of skin graft in E group (70.0 +/- 17.2 day) was obviously longer than that in C group. The serum levels of IL-4, chimera in E group were increased gradually, while MLC response decreased gradually. The serum IL-4 level reached 251.5 +/- 11.0 ng/L and splenic chimera level to 26.30 +/- 1.04% on the 60th day after NPC infusion.</p><p><b>CONCLUSION</b>The high levels of IL-4 and chimera might play important roles in inducing and maintaining immune tolerance.</p>

Influence of ultrasonic processing on the aggregation of PrP-Sc in the brain extracts of the scrapie-infected hamsters / 中华实验和临床病毒学杂志

<p><b>BACKGROUND</b>To evaluate the influence ultrasonic processing on the aggregation of PrP(Sc) in the brain extracts prepared from the scrapie-infected hamsters, and to seek for the way to prepare lower molecular PrP(Sc) polymer.</p><p><b>METHODS</b>The extracts of infected brains were prepared with a lysis solution, and treated with ultrasonics at various conditions during the different phases. The distribution and aggregation state of PrP(Sc) were analyzed by proteinase K treated. Western blot, and afterwards, quantitatively calculated with a commercially supplied software Image Totaltech.</p><p><b>RESULTS</b>The amount of PrP(Sc) in the supernatant of brain homogenates was 1.29-to 1.58-fold increased with appropriate sonication (15 s for 30 times). At the same conditions of ultrasound, the PrP amount in the supernatant prepared from the scrapie-infected hamster brain was significantly increased, whereas that prepared from healthy animal used as normal control showed little change. Comparative analyses of PrP(Sc) pellets prepared by high-speed centrifugation revealed that about 90% PrP(Sc) released into supernatant after ultrasound processing.</p><p><b>CONCLUSION</b>Appropriate sonication of homogenate of scrapie-infected brain increases the extracted amount of PrP(Sc), being favorable to laboratory diagnosis. Larger molecular PrP(Sc) aggregates can be crashed by ultrasonic processing, engendering lower molecular PrP(Sc) polymers.</p>