Monocytes are more susceptible in vitro in rhesus macaques of Chinese origin to recombinant serotype 5 adenovirus with higher preexisting vector specific neutralizing antibody titer / 病毒学报

Human adenovirus serotype 5 (HAd5) infect dendritic cells with low efficiency which restricts the use of HAd5 as an antigen carrying vector in such cells. Aiming to find a novel strategy to detour the traditional method for more convenient clinical use, peripheral blood mononuclear cells isolated from Chinese rhesus macaques were chosen as the target cells for HAd5. In vitro infection protocol was optimized which indicated centrifugation at 1000g could ease the entry of adenovirus. By this protocol, CD14 positive monocytes were infected at high efficiencies (> 80%), and about 10% of natural killer cells were infected; while T and B lymphocytes were rarely infected. Interestingly and importantly, it was the first time to report that in our in vitro study monocytes were more susceptible to HAd5-EGFP in macaques with higher preexisting vector specific neutralizing antibody titers. This phenomenon indicates an expansion of application of adenovirus based vectors for vaccine development and clinical use, especially for the population with preexisting neutralizing antibodies.

A new local ablation for unresectable primary liver tumor: effect of electrothermal and electrochemical therapy on rat liver / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

<p><b>BACKGROUND</b>Electrochemical therapy (ECT) has been used to treat unresectable hepatic tumor. In order to improve its efficacy, we combined ECT with hyperthermia induced by electrothermal needle (ETN) (ETECT). The aim of this study is to investigate the destructive effect of ETECT on normal rat liver.</p><p><b>METHODS</b>Twenty rats were randomized into 4 treatment groups (n=5 in each group): control, ECT alone, hyperthermia alone and ETECT. Following the treatment, sections of the livers were histologically examined by light microscopy and the destructive volumes were measured with micrometer.</p><p><b>RESULTS</b>We found that the destructive volumes in ETECT group were the largest (P<0.01). In ETECT group coagulative necrosis was found in both anode and cathode areas, around which transition zones existed. The transition zones can only be seen when coulomb was increased in ECT group.</p><p><b>CONCLUSION</b>ETECT was demonstrated to enhance the destructive effect of ECT. This study provides theoretical and experimental basis for a new local ablative treatment for unresectable primary liver tumor.</p>

Expression of recombinant human zona pellucida-3 protein (rhZP3) in Pichia pastoris / 生物工程学报

Human Zona Pellucida(ZP), which is a complex matrix surrounding oocytes,is comprised of three immunologically distinct glycoproteins(hZP1, hZP2 and hZP3). Because hZP3 possesses the sperm receptor activity and the acrosome-inducing activity, it has long been used as a candidate antigen to develop an immunocontraceptive vaccine. However, a large amount of native hZP3 protein is unavailable. It is an effective way to express hZP3 protein directly in vitro. Nevertheless, it had been reported that the rhZP3 protein produced in Pichia pastoris was not secreted but accumulated in the cells and could only be purified after being solubilized by strong denaturants. More unfortunately, after purification the final product required 6mol/L urea to maintain solubility. An improved project was advanced with the aim to express secreted and soluble rhZP3 protein in yeast. In this study, the fragment of hZP3 cDNA coding for aa 23 - 408, which the N-terminal leader was removed and most of the C-terminal transmembrane-like domain was reserved, was amplified by two PCR primers including EcoR I and Not I sites respectively and a His6 codon cassette was added to 5'-terminal. The hZP3 insert was incorporated into expression vector pPIC9K. The resulting recombinant yeast expression vector was designated pPIC9K-rhZP3. Linearized pPIC9K-rhZP3 was transformed into Pichia pastoris. After G418 selection, the recombinant Pichia pastoris strains were identified by PCR and the rhZP3 was expressed following the manufacturer' s protocol. Following induction with methanol, the rhZP3 protein was secreted and dissolved into the culture supernatant. SDS-PAGE and Western blot analyses showed that the apparent molecular weight of the expressed rhPZ3 proteins in yeast was smaller and a little size heterogeneity than native ones; after purified with Ni-chelating affinity chromatography, the final product's apparent molecular weight was about 32 - 34KD and their yield more than 20mg/L. We supposed that the C-terminal transmembrane-like domain be useful for secretion of rhZP3 into the culture supernatant and the expressed rhZP3 protein be incompletely digested by proteinases of Pichia into shorter fragments which all were glycosylated inhomogeneously. Fortunately, the fragments of rhZP3 protein can be recognized in Western blot by the polyclonal antibodies to porcine ZP3 which has showed a cross-reactivity with human ZP in vitro. It will be expected that the rhZP3 protein expressed in Pichia pastoris not only has immunogencity, say, it can rise antibodies in vivo to prevent spermatozoa-ovum binding, but also does not contain ovarian factors that might be the cause of undesired side effects, e.g. ovaritis and can be used as a safe immunogen in human antifertility vaccine research.

Antitumor effects of electrothermal and electrochemical therapy on mice with sarcoma180 / 中华口腔医学杂志

<p><b>OBJECTIVE</b>To explore the anti-tumor effects and the mechanism of a new treatment for Sarcoma180 in mice using both hyperthermia (H) and electrochemical therapy (ECT).</p><p><b>METHODS</b>A group of mice with Sarcoma180 (1 cm in diameter) were divided randomly into four groups and given different treatments: Control, ECT alone, H alone and electrothermal & electrochemical therapy (ETECT). The volumes of the tumors were calculated everyday after treatment. The tumor tissues were examined morphologically at 0, 6, 24 and 72 hours after treatment.</p><p><b>RESULTS</b>The tumors treated with ETECT were completely destroyed, and did not recur within a period of ten days by pathological examination. Results of this group were significantly different with the other groups (P<0.01). However, recurred tumors were found in 6 mice of H group and 5 mice of ECT group, respectively.</p><p><b>CONCLUSIONS</b>Based on the results obtained, ETECT induced by ETECD (D-device) is an effective way to eliminate cancer. It is capable of completely destroying cancer cells in a short time. The rate of Sarcoma180 inhibition is 100%, and malignant cells can't recur. The research provided theoretical and experimental bases for the treatment of oral cancer by a new approach to treat oral cancer.</p>