Yu C.-Y.,Beihua University |
Liu X.,Beihua University |
Yu C.-R.,JOINN Laboratories |
Zhang Y.,Jilin University |
And 3 more authors.
Journal of Jilin University Medicine Edition | Year: 2014
Objective: To establish oxidative stress model of HeLa cells copied by vitamin K3 (VK3), and to investigate the role of external signal-regulated kinase (ERK) pathway in autophagy induced by oxidative stress in HeLa cells. Methods: HeLa cells were divided into control group, VK3 group (30 μmol · L-1), U0126 group (10 μmol · L-1), and VK3 (30 μmol · L-1) + U0126 group (10 μmol · L-1). The survival rate of HeLa cells in each group was measured by MTT assay; the expression levels of external signal-regulated kinase 1/2 (ERK1/2), phosphorylated ERK 1/2 (Phospho-ERK1/2), microtubule-associated protein light chain 3-II (LC3-II), and cysteine aspartic acid specific protease-3 (caspase-3), and its activated fragment cleaved caspase-3 were determined by Western blotting method; the morphological changes of nuclear and apoptotic rate of HeLa cells in each group were detected by Hoechst staining. Results: Compared with VK3 group, the survival rate of HeLa cells in VK3 + U0126 group was decreased (P<0.05). The expressions of Phospho-ERK1/2 in HeLa cells were increased after treated with VK3 for 2, 4, and 8 h (P<0.05). Compared with control group, the expression of LC3-II protein in VK3 group was increased (P<0.05). Compared with VK3 group, the expression of LC3-II in HeLa cells in VK3 + U0126 group was decreased (P<0.05), and the expression of cleaved caspase-3 protein was increased (P<0.05). Compared with VK3 group, the morphological changes of nuclear appeared and the apoptotic rate of HeLa cells in VK3 + U0126 group was increased (P<0.05). Conclusion: Oxidative stress activates autophagy of HeLa cells induced by ERK signal pathway, and the autophagy can inhibit the apoptosis of HeLa cells induced by VK3.
Zou J.,China Agricultural University |
Zou J.,JOINN Laboratories |
Chen Q.,China Agricultural University |
Chen Q.,CAS Institute of Zoology |
And 7 more authors.
Toxicology | Year: 2011
Olaquindox is used in China as feed additive for growth promotion in pigs. Recently, we have demonstrated that olaquindox induced genome DNA damage and oxidative stress in HepG2 cells. The aim of this study was to explore the molecular mechanism of cell cycle arrest and apoptosis induced by olaquindox in HepG2 cells. In the present study olaquindox induced cell cycle arrest to the S phase and dose-dependent apoptotic cell death in HepG2 cells, indicated by accumulation of sub-G1 cell population, nuclear condenstion, DNA fragmentation, caspases activation and PARP cleavage. Meanwhile, the data showed that olaquindox triggered ROS-mediated apoptosis in HepG2 cells correlated with both the mitochondrial DNA damage and nuclear DNA damage, collapse of Δψm, opening of mPTP, down-regulation of Bcl-2 and up-regulation of Bax. Furthermore, we also found that olaquindox increased the expression of p53 protein and induced the release of cytochrome C from mitochondria to cytosol. In conclusion, olaquindox induced apoptosis of HepG2 cells through a caspase-9 and -3 dependent mitochondrial pathway, involving p53, Bcl-2 family protein expression, Δψm disruption and mPTP opening. © 2011 Elsevier Ireland Ltd.
Wang Y.,Peking Union Medical College |
Han Z.-B.,Peking Union Medical College |
Ma J.,JOINN Laboratories |
Zuo C.,JOINN Laboratories |
And 10 more authors.
Stem Cells and Development | Year: 2012
Therapies based on stem cells have shown very attractive potential in many clinical studies. However, the data about the safety of stem cells application remains insufficient. The present study was designed to evaluate the overall toxicology of human umbilical cord mesenchymal stem cells (hUC-MSCs) in cynomolgus monkeys with repeated administrations. hUC-MSCs were administered by intravenous injection once every 2 weeks for 6 weeks. The dose levels employed in this study were 2×10 6, 1×10 7 cells/kg body weight. Toxicity was evaluated by clinical observations (body weight, body temperature, and ophthalmology exams), pathology (blood cell counts, clinical biochemistry, urinalysis, and bone marrow smears), immunologic consequences (lymphoproliferation assay, the secretion of interferon-γ and interleukin-4, the percentage of CD3, CD4, CD8 T cells, and the ratio of CD4 and CD8 T cells) and anatomic pathology. Pharmacodynamics in blood and distribution of hUC-MSCs in the tissues of cynomolgus monkeys were measured by real-time polymerase chain reaction. All animals survived until scheduled euthanasia. No stem cells transplantation-related toxicity was found in this study. hUC-MSCs could be found in the blood of cynomolgus monkeys beyond 8 h. The findings of this 6-week toxicity study showed that the transplantation of hUC-MSCs did not affect the general health of cynomolgus monkeys. © Copyright 2012, Mary Ann Liebert, Inc.
Li J.,Peking University |
Li J.,Beijing Tiantan Biological Products |
Zhang D.,Beijing Tiantan Biological Products |
Ma R.,Beijing Tiantan Biological Products |
And 7 more authors.
Human Vaccines and Immunotherapeutics | Year: 2013
Aims: The current 3-dose regimen of hepatitis B vaccination for infants requiring over 6 mo period may pose the poor rate of compliance and later protection from hepatitis B virus (HBV) infection. This preclinical study is to investigate the feasibility of reducing the number of doses of hepatitis B (HB) vaccine. Results: Eight groups of guinea pigs immunized with two doses of HP-HB vaccines at either 0 and 4 weeks or 0 and 8 weeks elicited geometric titers (GMT) of anti-HBs similar to that of four groups immunized with three doses of controls. The overall GMT of anti-HBs were not significantly different between the E- and C-groups (p > 0.05) of monkeys. Specifically, the anti-HBs titers in the C-group reached the peak of 24857 (938.3-104585) mIU/mL one week after the 3rd dose, which were statistically higher than those of the E-group. However, they were reduced to comparable levels of anti-HBs in the E-group during weeks 9-12, suggesting comparable immune response of both vaccination regimens. Methods: Twelve groups of guinea pigs (four animals in each group) were immunized with 2 experimental recombinant yeast Hansenula polymorpha derived HB vaccines (HP-HB vaccine) and 2 commercial recombinant yeast Saccharomyces cerevisiae vaccines (Temrevac-HB) as controls at 0, 4 and 8 weeks, 0 and 4 weeks, and 0 and 8 weeks respectively. Each guinea pig received 2 μg vaccine. Twelve Cynomolgus monkeys were randomly divided into two groups (six animals in each group). Animals in the experimental group (E-group) were injected with two doses of pilot produced 20 μg HP-HB vaccine. Animals in the control group (C-Group) were immunized with three doses of 10 μg Temrevac-HB. Both vaccines were administered at an interval of 3 weeks for monkeys. Conclusions: The 2-dose regimen of the HP-HB vaccine has comparable HBV immune responses as the 3-dose regimen of Temrevac-HB vaccine in Cynomolgus monkeys. © 2013 Landes Bioscience.
PubMed | Shanghai Institute of Pharmaceutical Industry, Institute for Food and Drug Safety Evaluation, Center for Drug Evaluation, Beijing Institute of Pharmacology and Toxicology and 7 more.
Type: | Journal: Mutation research. Genetic toxicology and environmental mutagenesis | Year: 2014
Although inter-laboratory validation efforts of the in-vivo micronucleus (MN) assay based on flow cytometry (FCM) have taken place in the EU and US, none have been organized in China. Therefore, an inter-laboratory study that included eight laboratories in China and one experienced reference laboratory in the US was coordinated to validate the in-vivo FCM MicroFlow() method to determine the frequency of micro-nucleated reticulocytes (MN-RETs) in rat blood. Assay reliability and reproducibility were evaluated with four known genotoxicants, and the results obtained with the FCM method were compared with the outcome of the traditional evaluation of bone-marrow micronuclei by use of microscopy. Each of the four chemicals was tested at three sites (two in China and the one US reference laboratory). After three consecutive daily exposures to a genotoxicant, blood and bone-marrow samples were obtained from rats 24h after the third dose. MN-RET frequencies were measured in 20,000 RET in blood by FCM, and micro-nucleated polychromatic erythrocyte (MN-PCE) frequencies were measured in 2,000 PCEs in bone marrow by microscopy. For both methods, each genotoxicant was shown to induce a statistically significant increase in the frequency of MN after treatment with at least one dose. Where more doses than one caused an increase, responses occurred in a dose-dependent manner. Spearmans correlation coefficient (rs) for FCM-based MN-RET vs microscopy-based MN-PCE measurements (eight experiments, 200 paired measurements) was 0.723, indicating a high degree of correspondence between methods and compartments. The rs value for replicate FCM MN-RET measurements performed at the eight collaborative laboratories was 0.940 (n=200), and between the eight FCM laboratories with the reference laboratory was 0.933 (n=200), suggesting that the automated method is very well transferable between laboratories. The FCM micronucleus analysis method is currently used in many countries worldwide, and these data support its use for evaluating the in-vivo genotoxic potential of test chemicals in China.
He Y.N.,JOINN Laboratories
Zhonghua bing li xue za zhi = Chinese journal of pathology | Year: 2017
目的：观察SD和Wistar大鼠自发性肿瘤病变及其发生率，为进一步开展致癌实验积累背景数据。 方法：采用4周龄无特定病原体(specific pathogen free，SPF)级SD大鼠180只和Wistar大鼠240只，雌雄各半，在SPF级环境下常规饲养104周后实施安乐死，对死亡动物及安乐死动物进行组织病理学观察，统计自发性肿瘤的类型和发生率。 结果：实际观察了411只大鼠(SD大鼠176只，Wistar大鼠235只)，患瘤动物占57.7%(237/411)。SD大鼠中患瘤动物占55.7%(98/176)，发生良性肿瘤的动物占48.9%(86/176)，发生恶性肿瘤的动物占15.9%(28/176)；Wistar大鼠中患瘤动物占59.1%(139/235)，发生良性肿瘤的动物占51.5%(121/235)，发生恶性肿瘤的动物占14.5%(34/235)。高发的良性肿瘤主要有垂体腺瘤(SD大鼠占23.3%、Wistar大鼠占12.3%)，雌性大鼠的乳腺纤维腺瘤(SD大鼠占21.3%、Wistar大鼠占12.9%)，雌性大鼠的乳腺腺瘤(SD大鼠占16.9%、Wistar大鼠占9.5%)，雄性大鼠的睾丸间质细胞瘤(SD大鼠占0，Wistar大鼠占14.3%)。高发的恶性肿瘤主要有雌性大鼠的乳腺癌(SD大鼠占10.1%、Wistar大鼠占3.4%)，雌性大鼠的子宫平滑肌肉瘤(SD大鼠占0、Wistar大鼠占2.6%)，皮肤鳞状细胞癌(SD大鼠占2.3%、Wistar大鼠占0.9%)，皮下纤维肉瘤(SD大鼠占1.1%、Wistar大鼠占2.1%)，大脑恶性胶质细胞瘤(SD大鼠占1.1%、Wistar大鼠占1.7%)。 结论：饲养104周的SD和Wistar大鼠，自发性肿瘤的发生率均较高。良性肿瘤的发生率高于恶性肿瘤。高发良性肿瘤主要为垂体腺瘤、乳腺纤维腺瘤(雌性动物)、乳腺腺瘤(雌性动物)、睾丸间质细胞瘤(雄性动物)；高发恶性肿瘤主要为乳腺癌(雌性动物)以及某些软组织肉瘤。本实验结果丰富了现有SD和Wisar大鼠自发性肿瘤的数据资料，为开展新药致癌实验提供了背景数据。.Objective: To investigate the spontaneous neoplastic lesions and their incidences in Sprague-Dawley (SD) and Wistar rats, and to accumulate background data for carcinogenicity studies. Methods: One hundred and eighty SD rats and 240 Wistar rats (4-week old) , half in each sex, were used in this study. The rats were housed routinely under specific pathogen-free environment and euthanized after 104 weeks. Histopathological examination was undertaken for all animals including deaths and scheduled euthanasia. The types and incidences of spontaneous tumors were gathered statistically. Results: Total 411 rats (176 SD rats and 235 Wistar rats) were examined in this study. The total tumor incidence of the 411 rats was 57.7%(237/411). The total tumor incidence in SD rats was 55.7%(98/176), benign tumor incidence was 48.9%(86/176) and malignant tumor incidence was 15.9%(28/176). The total tumor incidence in Wistar rats was 59.1%(139/235), benign tumor incidence was 51.5%(121/235) and malignant tumor incidence was 14.5%(34/235). The main benign tumors were pituitary adenoma (23.3% in SD rats, 12.3% in Wistar rats), breast fibroadenoma (21.3% in SD rats, 12.9% in Wistar rats) and breast adenoma (16.9% in SD rats, 9.5% in Wistar rats) in females; testis Leydig cell tumor (0 in SD rats, 14.3% in Wistar rats) in males. The main malignant tumors were breast carcinoma (10.1% in SD rats, 3.4% in Wistar rats) and uterine leiomyosarcoma (0 in SD rats, 2.6% in Wistar rats) in females; squamous cell carcinoma of skin (2.3% in SD rats, 0.9% in Wistar rats); subcutaneous fibrosarcoma (1.1% in SD rats, 2.1% in Wistar rats); brain malignant glioma (1.1% in SD rats, 1.7% in Wistar rats). Conclusions: In the study, a high incidence of spontaneous tumors is reported in both SD and Wistar rats housed for 2 years. The incidence of benign tumors is higher than that of malignant rumors. The benign tumors mainly are pituitary adenoma, breast fibroadenoma and breast adenoma in females, and testis Leydig cell tumor in males. The malignant tumors mainly are breast carcinoma and some soft tissue sarcomas. The results of the study enrich the data of spontaneous tumor in SD and Wisar rats and provide background data for carcinogenicity studies for new drugs.
Li Y.-P.,Peking Union Medical College |
Gao L.-Y.,Peking Union Medical College |
Li K.-T.,JOINN Laboratories |
Meng S.,Peking Union Medical College |
And 5 more authors.
Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences | Year: 2013
A LC-MS/MS method for determining the concentration of the small molecule Hsp90 inhibitor, GM-AMPL, has been developed and validated in rat plasma to support preclinical development. 17-[2-(morpholine-4-yl)ethyl]amino-17-demethoxygeldanamycin (GM-AMPL) and the internal standard 17-allylamino-17-demethoxygeldanamycin (17-AAG) were sufficiently separated on a Venusil MP C18 column that was eluted with 80% methanol in water at 40 °C. Quantification studies were performed with a multiple reaction monitoring (MRM) transition of m/z 657.3→614.3 and 584.3→541.3 for GM-AMPL and IS, respectively, in the negative ion mode. The present method exhibited good linearity (R> 0.999) over the concentration range of 2-600. ng/mL for GM-AMPL in rat plasma with a lower limit of quantification (LLOQ) of 2. ng/mL. The intra-batch and inter-batch assay coefficients of variation (CV) were in range of 1.56-6.84% and 1.62-6.98%, respectively. The plasma samples were extracted with methanol to precipitate protein with extraction recovery in range of 84.09-95.25%. The matrix effect was determined as internal substance (IS) normalized matrix factor of 1.09, 1.18 and 1.05 for samples with three concentration levels of 4, 40 and 400. ng/mL, respectively. This validated method was further applied to successfully determine the pharmacokinetic parameters and oral availability of GM-AMPL in Sprague-Dawley rats following intravenous injection and oral administration. © 2012.
Duong-Ly K.C.,Johns Hopkins University |
Duong-Ly K.C.,Fox Chase Cancer Center |
Woo H.N.,Johns Hopkins University |
Woo H.N.,Case Western Reserve University |
And 8 more authors.
PLoS ONE | Year: 2013
The gene for a Nudix enzyme (SP_1669) was found to code for a UDP-X diphosphatase. The SP_1669 gene is localized among genes encoding proteins that participate in cell division in Streptococcus pneumoniae. One of these genes, MurF, encodes an enzyme that catalyzes the last step of the Mur pathway of peptidoglycan biosynthesis. Mur pathway substrates are all derived from UDP-glucosamine and all are potential Nudix substrates. We showed that UDP-X diphosphatase can hydrolyze the Mur pathway substrates UDP-N-acetylmuramic acid and UDP-N-acetylmuramoyl-L-alanine. The 1.39 Å resolution crystal structure of this enzyme shows that it folds as an asymmetric homodimer with two distinct active sites, each containing elements of the conserved Nudix box sequence. In addition to its Nudix catalytic activity, the enzyme has a 3′5′ RNA exonuclease activity. We propose that the structural asymmetry in UDP-X diphosphatase facilitates the recognition of these two distinct classes of substrates, Nudix substrates and RNA. UDP-X diphosphatase is a prototype of a new family of Nudix enzymes with unique structural characteristics: two monomers, each consisting of an N-terminal helix bundle domain and a C-terminal Nudix domain, form an asymmetric dimer with two distinct active sites. These enzymes function to hydrolyze bacterial cell wall precursors and degrade RNA. © 2013 Duong-Ly et al.
Jiang E.-P.,Beihua University |
Jiang E.-P.,Guangdong Medical College |
Wang S.-Q.,Beihua University |
Wang S.-Q.,Jilin Central General Hospital |
And 4 more authors.
Zhongguo Zhongyao Zazhi | Year: 2014
Objective: To observe the effect of Schisandra chinensis lignans(SCL) on neuronal apoptosis and PI3K/AKT signaling pathway of rats in the cerebral ischemia injury model, and study its possible mechanism. Method: Rats were orally administered SCL high, middle and low dose groups (100, 50, 25 mg·kg-1) for 14 days. The cerebral ischemia injury model was established by using the suture-occluded method to rate the neurological functions. The cerebral infarction area was observed by TTC staining. The pathological changes in brain tissues were determined by HE staining. Bcl-2 and Bax expressions were detected by immunohistochemical assay. The protein expressions of p-AKT and AKT were assayed by Western blotting. Result: Compared with the model group, SCL high, middle and low dose groups showed reduction in the cerebral infarction area to varying degrees, improve the pathological changes in brain tissues, promote the expression of apoptin Bcl-2 and p-AKT, and inhibit the expression of apoptin Bax. Conclusion: SCL shows a protective effect on rats with cerebral ischemia injury. Its mechanism may be related to the increase in p-AKT ability and anti-ischemic brain injury capacity and the inhibition of nerve cells.
Jiang E.-P.,Guangdong Medical College |
Li H.,Beihua University |
Yu C.-R.,JOINN Laboratories |
Yu C.-Y.,Beihua University |
And 6 more authors.
NeuroReport | Year: 2015
Increasing evidence places Schisandrin B (Sch B) at an important position in nerve protection, indicating that Sch B might play a positive role in the therapy of neurodegenerative diseases. However, there is little information on it. Our studies showed that pretreatment with Sch B could reduce lactate dehydrogenase, malondialdehyde, and reactive oxygen species release and significantly increase the cell viability and the superoxide dismutase level. Sch B (10 μM) markedly inhibited cell apoptosis, whereas LY294002 (20 μM), a phosphatidylinositol-3 kinase inhibitor, blocked the antiapoptotic effect. More importantly, Sch B (10 μM) increased the phosphoprotein kinase B/protein kinase B (Akt) and B-cell lymphoma-2/Bcl-2 associated X protein ratios on preincubation with cells for 2 h, which was then inhibited by LY294002 (20 μM). Results indicate that Sch B can protect PC12 cells from apoptosis by activating the phosphatidylinositol-3 kinase/Akt signaling pathway and may emerge as a potential drug for neurodegenerative diseases. Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.