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Biomedical subjects

Wei Zhou

Publications and source records attributed to Wei Zhou.

At least 91 records · Page 5Linked to original sources

IMA clipping for a type ii endoleak: combined laparoscopic and endovascular approach.

We describe herein a combined laparoscopic and endovascular approach to treat a type II endoleak due to retrograde flow in the patent inferior mesenteric artery (IMA). A 61-year-old gentleman presented with enlarging aneurysm sac confirmed on computed tomography scan evaluation after elective endovascular repair of an infrarenal abdominal aortic aneurysm. A combined laparoscopic and endovascular approach was used. After distal IMA was identified and marked with a clip laparoscopically, on-table angiography showed a proximal left colic branch and persistent flow in the IMA. Therefore, further laparoscopic exploration was performed by dissection along the distal branch. The origin of IMA was then located and subsequently sealed with 2 surgical clips. The completion angiography confirmed the proper position of the surgical clips and absence of endoleak. Our case demonstrated useful role of endovascular techniques in identifying the origin of IMA during laparoscopic approach for treating type II endoleak.

Aortic Aneurysm, Abdominal↗

The effect of venue sampling on estimates of HIV prevalence and sexual risk behaviors in men who have sex with men.

OBJECTIVE: The objective of this study was to investigate differences in HIV prevalence and sexual risk behaviors among men who have sex with men (MSM) according to their gay venue visit patterns. METHODS: In a cross-sectional survey, a population-based sample of men aged 18 to 64 years who self-identified as gay or bisexual were interviewed by telephone regarding their sexual behaviors, HIV serostatus, and gay venue visit patterns. RESULTS: A total of 398 men were recruited for the study. The results showed that frequent gay venue visitors were more likely to engage in high-risk sexual behaviors. Among gay venue attendees who visited different types of gay venues, men who visited sex clubs/bathhouses reported the highest rates of 5 or more male sexual partners and unprotected anal intercourse (UAI) with secondary partners (62.6% and 34.6%, respectively), gay bar/club attendees and cruisers reported higher rates of having sex with women (8.5% and 14.8%, respectively), and circuit party attendees reported the highest HIV prevalence (40.4%) and serodiscordant UAI (30.2%). CONCLUSIONS: MSM who visited different types of gay venues and with varied visit frequency showed marked differences in sexual risk behaviors, and the differences suggest the importance of weighting procedure to obtain unbiased estimates in venue-based studies.

Adolescent↗

The technical development of crystals and oscillators in China and their market situation.

In recent years the manufacture of and demand for crystals and oscillators in China have been increasing quickly because of the development of industries, communications, science, transportation, and electronic techniques. This paper introduces the manufacture, research, and characteristics of the technical development of crystals and oscillators in the last ten years in China. To meet the demand from industry, science, and other markets, the number of companies that produce and research crystals and oscillators has increased from several to more than 300 in the last 20 years. Manufacturing technology and equipment have also improved. For marketing and technical reasons, more than 75% of the companies manufacture mainly 49 micro, 49 micros, and microm crystals. Now more and more companies plan to build production lines to make cold-weld crystals and other precision crystals, and at least two companies have already built their production lines. Most of the precision crystals are AT-cut but some are SC-cut. Integrated TCXOs and DTCXOs have been made by different companies, and the frequency-temperature stability of them is from +/- 2 ppm to +/- 0.5 ppm in a wide temperature range. Specifications of some OCXOs are almost the same as those of foreign OCXOs. The main design techniques in China are experimental. However, in recent years, specially designed software has been developed for oscillator design, and advanced instrumentation and equipment are used. The market situation for crystals and oscillators has increased.

Journal Article↗

The leader sequence triggers and enhances several functions of clusterin and is instrumental in the progression of human prostate cancer in vivo and in vitro.

OBJECTIVE: To investigate the role of the leader sequence (which during clusterin biosynthesis facilitates its proper post-translational processing and secretion) in the functional activities of clusterin, a ubiquitous secretory glycoprotein with many biological functions, reported to be pro-apoptotic and anti-apoptotic in target cells, but for which the dual mechanism remains unclear. MATERIALS AND METHODS: We designed an expression vector starting from the second in-frame ATG on the full-length human clusterin cDNA that was capable of driving the expression of both the full-length and the truncated isoforms of clusterin. We established stable expression clones of the androgen-dependent prostate cancer line LNCaP expressing clusterin with and without the leader sequence. This induced expression provided an opportunity to evaluate both the in vivo and in vitro actions of clusterin expression. RESULTS: The LNCaP cells expressing clusterin with the leader sequence resisted apoptosis induced by tumour necrosis factor (TNF)-alpha, but clones with no leader sequence were highly susceptible to TNF-alpha-induced apoptosis. Furthermore, in the absence of the leader sequence, the expressed clusterin had a molecular weight consistent with that of the predicted holoprotein (40 kDa), suggesting a compromised post-translational processing with diffuse distribution throughout the cytoplasm. However, cells transfected with the full-length vector expressed clusterin of 60 and 35 kDa variants, and located exclusively in the Golgi apparatus. In vivo, only the overexpression of the full-length clusterin is anti-apoptotic and stimulates the proliferation of tumour. CONCLUSION: The leader sequence is important in determining the functions of clusterin, which include anti-apoptotic and anti-necrotic properties. The lack of the leader sequence allowed the incompletely processed clusterin to induce apoptosis in target cells; without the leader sequence, clusterin functions differently. Thus, the leader sequence is a trigger for many functions of clusterin in the progression of human prostate cancer cells.

Apoptosis↗

New nematicidal azaphilones from the aquatic fungus Pseudohalonectria adversaria YMF1.01019.

Two new azaphilone metabolites, named pseudohalonectrin A (1) and B (2), were isolated from the culture of the aquatic fungus Pseudohalonectria adversaria YMF1.01019, originally separated from submerged wood in Yunnan Province, China. Pseudohalonectrin A and B were assessed for their nematicidal activity against the pine wood nematode Bursaphelenchus xylophilus and their structures were defined after spectral analysis. This is the first report of secondary metabolites from any member of the genus Pseudohalonectria.

Animals↗

Differential properties of GTP- and Ca(2+)-stimulated exocytosis from large dense core vesicles.

Many cells utilize a GTP-dependent pathway to trigger exocytosis in addition to Ca(2+)-triggered exocytosis. However, little is known about the mechanism by which GTP triggers exocytosis independent of Ca(2+). We used dual-color evanescent field microscopy to compare the motion and fusion of large dense core vesicles stimulated by either mastoparan (Mas) in Ca(2+)-free conditions or high K(+) in the presence of Ca(2+). We demonstrate that Mas is hardly effective in triggering the fusion of the predocked vesicles but predominantly mobilizes cytosolic vesicles. In contrast, Ca(2+)-dependent exocytosis is largely due to predocked vesicles. Fusion kinetics analysis and carbon-fiber amperometry reveal that Mas induces a brief 'kiss-and-run' fusion and releases only a small amount of the cargo, whereas Ca(2+) stimulates a more persistent opening of the fusion pore and larger release of the contents. Furthermore, we show that Mas-released vesicles require a much shorter time to reach fusion competence once they approach the plasma membrane. Our data suggest the involvement of different mechanisms not only in triggering and fusion but also in the docking and priming process for Ca(2+)- and GTP-dependent exocytosis.

Animals↗

Extracellular ATP stimulates exocytosis via localized Ca(2+) release from acidic stores in rat pancreatic beta cells.

Three different methods, membrane capacitance (C(m)) measurement, amperometry and FM dye labeling were used to investigate the role of extracellular ATP in insulin secretion from rat pancreatic beta cells. We found that extracellular application of ATP mobilized intracellular Ca(2+) stores and synchronously triggered vigorous exocytosis. No influence of ATP on the readily releasable pool of vesicles was observed, which argues against a direct modulation of the secretory machinery at a level downstream of Ca(2+) elevation. The stimulatory effects of ATP were greatly reduced by intracellular perfusion of BAPTA but not EGTA, suggesting a close spatial association of fusion sites with intracellular Ca(2+) releasing sites. ATP-induced Ca(2+) transients and exocytosis were not blocked by thapsigargin (TG), by a ryanodine receptor antagonist or by dissipation of pH in acidic stores by monensin alone, but they were greatly attenuated by IP(3) receptor inhibition as well as ionomycin plus monensin, suggesting involvement of IP(3)-sensitive acidic Ca(2+) stores. Taken together, our data suggest that extracellular ATP triggers exocytosis by mobilizing spatially limited acidic Ca(2+) stores through IP(3) receptors. This mechanism may explain how insulin secretion from the pancreas is coordinated through diffusible ATP that is co-released with insulin.

Adenosine Triphosphate↗

Recombinant neural protein PrP can bind with both recombinant and native apolipoprotein E in vitro.

The most essential and crucial step during the pathogenesis of transmissible spongiform encephalopathy is the conformational change of cellular prion protein (PrP(C)) to pathologic isoform (PrP(Sc)). A lot of data revealed that caveolae-like domains (CLDs) in the cell surface were the probable place where the conversion of PrP proteins happened. Apolipoprotein E (ApoE) is an apolipoprotein which is considered to play an important role in the development of Alzheimer's disease and other neurodegenerative diseases by forming protein complex through binding to the receptor located in the clathrin-coated pits of the cell surface. In this study, a 914-bp cDNA sequence encoding human ApoE3 was amplified from neuroblastoma cell line SH-SY5Y. Three human ApoE isomers were expressed and purified from Escherichia coli. ApoE-specific antiserum was prepared by immunizing rabbits with the purified ApoE3. GST/His pull-down assay, immunoprecipitation and ELISA revealed that three full-length ApoE isomers interact with the recombinant full-length PrP protein in vitro. The regions corresponding to protein binding were mapped in the N-terminal segment of ApoE (amino acid 1-194) and the N-terminal of PrP (amino acid 23-90). Moreover, the recombinant PrP showed the ability to form a complex with the native ApoE from liver tissues. Our data provided direct evidence of molecular interaction between ApoE and PrP. It also supplied scientific clues for assessing the significance of CLDs on the surface of cellular membrane in the process of conformational conversion from PrP(C) to PrP(Sc) and probing into the pathogenesis of transmissible spongiform encephalopathy.

Apolipoproteins E↗

Mechanism of rat mesenteric arterial KATP channel activation by 14,15-epoxyeicosatrienoic acid.

Recently, we reported that 11,12-epoxyeicosatrienoic acid (11,12-EET) potently activates rat mesenteric arterial ATP-sensitive K(+) (K(ATP)) channels and produces significant vasodilation through protein kinase A-dependent mechanisms. In this study, we tried to further delineate the signaling steps involved in the activation of vascular K(ATP) channels by EETs. Whole cell patch-clamp recordings [0.1 mM ATP in the pipette, holding potential (HP) = 0 mV and testing potential (TP) = -100 mV] in freshly isolated rat mesenteric smooth muscle cells showed small glibenclamide-sensitive K(ATP) currents (19.0 +/- 7.9 pA, n = 5) that increased 6.9-fold on exposure to 5 microM 14,15-EET (132.0 +/- 29.0 pA, n = 7, P < 0.05 vs. control). With 1 mM ATP in the pipette solution, K(ATP) currents (HP = 0 mV and TP = -100 mV) were increased 3.5-fold on exposure to 1 microM 14,15-EET (57.5 +/- 14.3 pA, n = 9, P < 0.05 vs. baseline). In the presence of 100 nM iberiotoxin, 1 microM 14,15-EET hyperpolarized the membrane potential from -20.5 +/- 0.9 mV at baseline to -27.1 +/- 3.0 mV (n = 6 for both, P < 0.05 vs. baseline), and the EET effects were significantly reversed by 10 microM glibenclamide (-21.8 +/- 1.4 mV, n = 6, P < 0.05 vs. EET). Incubation with 5 microM 14,15-epoxyeicosa-5(Z)-enoic acid (14,15-EEZE), a 14,15-EET antagonist, abolished the 14,15-EET effects (31.0 +/- 11.8 pA, n = 5, P < 0.05 vs. 14,15-EET, P = not significant vs. control). The 14,15-EET effects were inhibited by inclusion of anti-G(s)alpha antibody (1:500 dilution) but not by control IgG in the pipette solution. The effects of 14,15-EET were mimicked by cholera toxin (100 ng/ml), an exogenous ADP-ribosyltransferase. Treatment with the ADP-ribosyltransferase inhibitors 3-aminobenzamide (1 mM) or m-iodobenzylguanidine (100 microM) abrogated the effects of 14,15-EET on K(ATP) currents. These results were corroborated by vasodilation studies. 14,15-EET dose-dependently dilated isolated small mesenteric arteries, and this was significantly attenuated by treatment with 14,15-EEZE or 3-aminobenzamide. These results suggest that 14,15-EET activates vascular K(ATP) channels through ADP-ribosylation of G(s)alpha.

8,11,14-Eicosatrienoic Acid↗

Polymorphisms of vitamin D receptor and survival in early-stage non-small cell lung cancer patients.

Our previous analysis suggested that surgery season in the summer time and high vitamin D intake are associated with improved survival in early-stage non-small cell lung cancer (NSCLC) patients. Here, we investigated the associations of vitamin D receptor (VDR) polymorphisms of Cdx-2 G>A, FokI C>T, and BsmI C>T with overall survival (OS) and recurrence-free survival (RFS) in 373 early-stage NSCLC patients. The data were analyzed using log-rank test and Cox proportional hazards models. The median follow-up time was 71 months (range, 0.1-140 months), with 186 deaths and 127 recurrences. There was no association between VDR polymorphisms and survival, overall or among adenocarcinoma patients. Among squamous cell carcinoma (SCC) patients, the G/A+A/A genotype group of the Cdx-2 polymorphism was associated with better OS: the 5-year OS rates were 41% [95% confidence interval (95% CI), 28-53] for the G/G and 55% (95% CI, 39-71) for the G/A+A/A genotypes, respectively (P = 0.04, log-rank test), with the adjusted hazard ratio of 0.56 (95% CI, 0.33-0.95) for G/A+A/A versus G/G. For the joint effects of the three polymorphisms, subjects with two or more "protective" alleles have better OS among SCC patients, with the adjusted hazard ratios of 0.20 (95% CI, 0.09-0.48), 0.40 (95% CI, 0.19-0.87), and 0.43 (95% CI, 0.19-0.97), respectively, for subjects with two, three, and four or more "protective" alleles when compared with subjects with zero or one "protective" allele (P(trend) = 0.71). Similar associations were found in haplotype analysis and for RFS among SCC patients. In conclusion, VDR polymorphisms may be associated with improved survival among SCC patients of early-stage NSCLC.

Adult↗

Effect of experimental glaucoma in primates on oscillatory potentials of the slow-sequence mfERG.

PURPOSE: To determine the effect of experimental glaucoma in macaque monkeys on oscillatory potentials (OPs) in the slow-sequence multifocal electroretinogram (mfERG). METHODS: Photopic slow-sequence mfERGs were recorded from anesthetized adult macaque monkeys and normal human subjects. The stimulus consisted of 103 equal-sized hexagons within 17 degrees of the fovea. The m-sequence was slowed, with 14 blank frames, approximately 200 ms, interleaved between flashes for monkeys and 7 blank frames, approximately 100 ms, for humans, to produce waveforms similar to the photopic full-field flash ERG. Recordings were made under control conditions (24 monkey eyes, 7 human) and after laser-induced experimental glaucoma in monkeys (n = 8). A Fourier fast transform [FFT] was used to determine the frequency ranges of the major OPs. OP amplitudes were quantified by using root mean square (RMS) for two-frequency bands in five horizontal and four vertical locations. Visual field defects were assessed using behavioral static perimetry. Full-field photopic flash ERGs also were recorded. RESULTS: OPs in two distinct frequency bands were discriminated in the monkey mfERG: fast OPs, with a peak frequency of 143 +/- 20 Hz, and slow OPs, with a peak at 77 +/- 8 Hz. There were similar findings in humans and with the flash ERG in monkeys. The fast OP RMS in monkey control eyes was significantly larger in temporal than nasal retina (P < 0.01) and in superior versus inferior retina (P < 0.05) as reported previously. The slow OP RMS was largest in the foveal region. Experimental glaucoma reduced fast OP RMS in all locations studied, even when visual field defects were moderate (MD = -5 to -10 dB; P < 0.05), whereas the slow OP RMS was reduced significantly primarily in the foveal region when field defects were severe (MD < -10 dB; P < 0.05). The fast OP RMS showed a moderate correlation with local visual field sensitivity and with local ganglion cell density (calculated from visual field sensitivity). For the slow OPs the correlation was much poorer. Consistent with previous studies, the photopic negative response (PhNR) amplitude was significantly reduced when the visual sensitivity was minimally affected. CONCLUSIONS: OPs in the ERG of primates fall in two frequency bands: fast OPs with a peak frequency around 143 Hz and slow OPs, with a peak frequency around 77 Hz. The fast OPs, which rely more on the integrity of retinal ganglion cells and their axons than do the slow OPs, have potential utility for monitoring the progression of glaucoma and the effects of treatment.

Animals↗

Lixazinone stimulates mitogenesis of Madin-Darby canine kidney cells.

Polycystic kidney diseases (PKD) are characterized by excessive proliferation of renal tubular epithelial cells, development of fluid-filled cysts, and progressive renal insufficiency. cAMP inhibits proliferation of normal renal tubular epithelial cells but stimulates proliferation of renal tubular epithelial cells derived from patients with PKD. Madin-Darby canine kidney (MDCK) epithelial cells, which are widely used as an in vitro model of cystogenesis, also proliferate in response to cAMP. Intracellular cAMP levels are tightly regulated by phosphodiesterases (PDE). Isoform-specific PDE inhibitors have been developed as therapeutic agents to regulate signaling pathways directed by cAMP. In other renal cell types, we have previously demonstrated that cAMP is hydrolyzed by PDE3 and PDE4, but only PDE3 inhibitors suppress proliferation by inhibiting Raf-1 activity (Cheng J, Thompson MA, Walker HJ, Gray CE, Diaz Encarnacion MM, Warner GM, Grande JP. Am J Physiol Renal Physiol 287:F940-F953, 2004.) A potential role for PDE isoform(s) in cAMP-mediated proliferation of MDCK cells has not previously been established. Similar to what we have previously found in several other renal cell types, cAMP hydrolysis in MDCK cells is directed primarily by PDE4 (85% of total activity) and PDE3 (15% of total activity). PDE4 inhibitors are more effective than PDE3 inhibitors in increasing intracellular cAMP levels in MDCK cells. However, only PDE3 inhibitors, and not PDE4 inhibitors, stimulate mitogenesis of MDCK cells. PDE3 but not PDE4 inhibitors activate B-Raf but not Raf-1, as assessed by an in vitro kinase assay. PDE3 but not PDE4 inhibitors activate the ERK pathway and activate cyclins D and E, as assessed by histone H1 kinase assay. We conclude that mitogenesis of MDCK cells is regulated by a functionally compartmentalized intracellular cAMP pool directed by PDE3. Pharmacologic agents that stimulate PDE3 activity may provide the basis for new therapies directed toward reducing cystogenesis in patients with PKD.

3',5'-Cyclic-AMP Phosphodiesterases↗

Differential effects on BAFF and APRIL levels in rituximab-treated patients with systemic lupus erythematosus and rheumatoid arthritis.

The objective of this study was to investigate the interaction between levels of BAFF (B-cell activation factor of the tumour necrosis factor [TNF] family) and APRIL (a proliferation-inducing ligand) and B-cell frequencies in patients with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) treated with the B-cell-depleting agent rituximab. Ten patients with SLE were treated with rituximab in combination with cyclophosphamide and corticosteroids. They were followed longitudinally up to 6 months after B-cell repopulation. Nine patients with RA, resistant or intolerant to anti-TNF therapy, treated with rituximab plus methotrexate were investigated up to 6 months after treatment. The B-cell frequency was determined by flow cytometry, and serum levels of BAFF and APRIL were measured by enzyme-linked immunosorbent assays. BAFF levels rose significantly during B-cell depletion in both patient groups, and in patients with SLE the BAFF levels declined close to pre-treatment levels upon B-cell repopulation. Patients with SLE had normal levels of APRIL at baseline, and during depletion there was a significant decrease. In contrast, patients with RA had APRIL levels 10-fold higher than normal, which did not change during depletion. At baseline, correlations between levels of B cells and APRIL, and DAS28 (disease activity score using 28 joint counts) and BAFF were observed in patients with RA. In summary, increased BAFF levels were observed during absence of circulating B cells in our SLE and RA patient cohorts. In spite of the limited number of patients, our data suggest that BAFF and APRIL are differentially regulated in different autoimmune diseases and, in addition, differently affected by rituximab treatment.

Adrenal Cortex Hormones↗

Hydroxyethyl starch exhibits antiinflammatory effects in the intestines of endotoxemic rats.

We performed the present in vivo study to investigate the effect of hydroxyethyl starch (HES) on intestinal production of inflammatory mediators and activation of transcription factors during endotoxemia. Rats with endotoxemia induced by lipopolysaccharide (LPS) (5 mg/kg, IV) were treated with HES (16 mL/kg, IV) or saline (64 mL/kg, IV). At 2, 3, or 6 h after the LPS challenge, the rat ileal tissues were collected. Various ileal inflammatory mediator levels (tumor necrosis factor-alpha, interleukin [IL]-6, cytokine-induced neutrophil chemoattractant-1, and IL-10), inflammatory mediator messenger RNAs (mRNAs), activities of nuclear factor (NF)-kappaB and activator protein (AP)-1, and ileal myeloperoxidase-positive cells were determined in each group. HES significantly reduced the increased intestinal levels of tumor necrosis factor-alpha, IL-6, cytokine-induced neutrophil chemoattractant-1, and the mRNAs in the endotoxemic rats. Similarly, HES could decrease the myeloperoxidase-positive cells induced by LPS and also inhibit ileal NF-kappaB and AP-1 activations. Our results suggest that during endotoxemia HES may down-regulate intestinal inflammatory mediator production, and this antiinflammatory effect of HES may act through suppression of NF-kappaB and AP-1 activations.

Animals↗

In situ reconstruction with cryopreserved arterial allografts for management of mycotic aneurysms or aortic prosthetic graft infections: a multi-institutional experience.

We designed this study to evaluate a multi-institutional experience regarding the efficacy of cryopreserved aortic allografts in the treatment of infected aortic prosthetic grafts or mycotic aneurysms. We reviewed clinical data of all patients from 4 institutions who underwent in situ aortic reconstruction with cryopreserved allografts for either infected aortic prosthetic graft or mycotic aneurysms from during a 6-year period. Relevant clinical variables and treatment outcomes were analyzed. A total of 42 patients (37 men; overall mean age 63 +/- 13 years, range 41-74 years) were identified during this study period. Treatment indications included 34 primary aortic graft infections (81%), 6 mycotic aneurysms (22%), and 2 aortoenteric erosions (5%). Transabdominal and thoracoabdominal approaches were used in 38 (90%) and 4 patients (10%), respectively. Staphylococcus aureus was the most commonly identified organism (n=27, 64%). Although there was no intraoperative death, the 30-day operative mortality was 17% (n=7). There were 21 (50%) nonfatal complications, including local wound infection (n=8), lower-extremity deep venous thrombosis (n=5), amputation (n=6), and renal failure requiring hemodialysis (n=2). The average length of hospital stay was 16.4 +/- 7 days. During a mean follow-up period of 12.5 months, reoperation for allograft revision was necessary in 1 patient due to graft thrombosis (6%). The overall treatment mortality rate was 21% (n=9). In situ aortic reconstruction with cryopreserved allografts is an acceptable treatment method in patients with infected aortic prosthetic graft or mycotic aneurysms. Our study showed that mid-term graft-related complications such as reinfection or aneurysmal degeneration were uncommon.

Adult↗

[An investigation on AIDS knowledge of Chinese peacekeepers in Liberia and effects of relevant health education].

OBJECTIVE: To investigate and analyze AIDS knowledge of the Chinese peacekeepers in Liberia and to evaluate the effects of AIDS education in order to conduct more effective health education of AIDS and increase AIDS knowledge among the Chinese peacekeepers in the areas with a high incidence of AIDS. METHODS: A questionnaire was designed on the basis of that recommended by WHO to investigate the knowledge of AIDS among the 528 Chinese peacekeepers in Liberia respectively before and after relevant health education, and the results were compared. RESULTS: There was an obvious deficiency in AIDS knowledge among the subjects before the education and an evident increase after it, and the difference was significant (P <0.01). CONCLUSION: The Chinese peacekeepers are deficient in AIDS knowledge, and the deficiency can be effectively improved by relevant health education, which is essential for the health of the Chinese peacekeepers in the areas with a high incidence of AIDS.

Acquired Immunodeficiency Syndrome↗

Establishment of cell free conversion system with biotin-labelled recombinant PrPsen expressed in E. coli.

OBJECTIVE: To report a protocol using biotin-labelled PrP protein in cell free conversion assay instead of isotope. METHODS: A hamster PrP protein (HaPrP) was expressed in E. coli and purified with HIS-tag affinity chromatograph. After being labelled with biotin, HaPrP was mixed with PrPSc preparation from scrapie strain 263K. RESULTS: Protease-resistant bands were detected after four-day incubation. CONCLUSION: The new conversion model provides a reliable, easily handling, and environment-friendly method for studies of prion and transmissible spongiform encephalopathies.

Animals↗

[Inhibitory effects of deoxyribozyme on the expression of Period1 gene in vitro and on morphine-induced psychic dependence in mice].

OBJECTIVE: To investigate the regulatory effects of deoxyribozyme on the expression of Period 1 gene in vitro and on the morphine-induced psychic dependence in mice. METHODS: The specific deoxyribozymes toward Period 1 mRNA was designed by MFold analysis and synethsized chemically. By LipofectAMINE mediated DNA transfection technique, DRz164 and pcDNA3-Per1 were introduced into NIH3T3 cells. The effects of deoxyribozyme on Period 1 gene were studied by reverse transcript-polymerase chain reaction (RT-PCR) and flow cytometry(FCM). The morphine-induced reward in mice was observed in a conditioned place preference test after pretreatment of the mice with the intracerebroventricular administration of deoxyribozyme. RESULTS: After NIH3T3 cells were transfected by pcDNA3-Per1 and DRz164, the Period 1 mRNA was reduced by 42.4%. And PERIOD proteins were decreased by 57.5%. After being pretreated with deoxyribozyme, the mice did not show morphine-induced place preference. CONCLUSION: DRz164 can highly block the expression of Period 1 gene, which cleaves the Period 1 mRNA in the transfected cells specifically. The suppression of morphine-induced place preference can be effected by pretreating the mice with alleviating their psychic dependence on morphine.

Animals↗