Objective To investigate the effects of KLT on apoptosis of HepG2 cells and expression of Bcl-2 and Capase-8. Methods The cell fine HepG2 was induced by diverse density of KLT, and HepG2 cell was collected respectively after induction of 12 h, 24 h, 48 h. The control group was installed simulta-neon]y. The cell apoptosis and the expression of Bcl-2 and Caspase-8 were detected by flow cytometry (FCM). Results KLT can induce the apoptosis of HepG2 cell significantly, and the longer time past, the more apoptosis of HepG2 was. KLT can increase the expression of Caspase-8, but ineffective to Bcl-2. Con-clusion KLT can significantly induce the apoptosis of HepG2 cell through regulating the expression of Caspase-8.

Objective Through studying the apoptosis induced by stichopus japonicus acid mucopoly saccharide in the hepatocellular carcinoma cell line HepG2 in vitro, analysing the expression of Bcl-2 and nm-23in HepG2, to provide the theory foundation and its feasibility on whether it can be used for the chemotherapy of hepetocellular carcinoma. Methods The cells of HepG2 were cultured in vitro and treated with SJAMP at different doses(0.25,0. 5,1.0,2.0,4.0 g/L). MTT was used to observe the inhibitory effects of SJAMP on cell growth, Western blotting was used to detect apoptosis, and the apoptosis related change of expression of protein Bcl-2 and nm23-H1. Results (1) MTT identified that SJAMP produced an obvious time-and-dose-dependent inhibitory effect on the HlepG2 cells. (2) Western blot showed that SJAMP could induce the apoptosis of HepG2 cells through changing the expression of the protein of Bcl-2 and nn23-H1 (P<0.05). Conclusion (1)SJAMP produced obvious inhibitory effects on HepG2 cells and induce HepG2 apoptosis. (2)SJAMP can enduce the anti-tumor function in the method of changing the expression of protein Bcl-2 and nm23-H1.

Objective To study the cause of death and mechanism after(TACE)in China during the past 14 years.Methods Related repots in Chinese Medical Current Content(CBM)and National Knowledge lnfrastruc ture(CNKI)from January 1994 to June 2008 were retrieved.The cause of death and mechainsm after TACE wer e analyzed.Results A total of 150 patients who died after TACE were reposed in China during the past 14 ye ar s.84%eases were caused by liver lunction failure,upper gastrointestinal bleeding and rupture of liver cancer. 78.7%cases died one month postoperation.Conclusion Liver function failure.upper gastrointestinal bleeding and rupture of liver cancer are the main complications which Can cause death and the majority cases died early.

BACKGROUND:Distraction osteogenesis is one of the most important tissue engineering technologies. However, the exact signaling pathway controling mesenchymal stem cel-osteoblast lineage (MSC-OB) migration during distraction osteogenesis has not yet been elucidated. More efforts should be paid to make a ful understanding of the mechanism on MSC-OB lineage migration, which can improve the clinical efficacy of distraction osteogenesis.
OBJECTIVE:To evaluate the effects of mechanical stretch on the ability of MSC-OB mobility and expression of mammalian target of rapamycin (mTOR) signaling pathway as wel as matrix metaloproteinases (MMPs) in MSC-OB, and to make clear the mechanism by which controls MSC-OB migration during distraction osteogenesis.
METHODS:Twelve Sprague-Dawley rats were randomized into two groups: experimental group (n=6), anin vivo rat mandibular distraction osteogenesis model was established on the right side of rats; non-stretch group (n=6), only the mandibular resection was done but with no distraction osteogenesis. Immunohistochemical staining was used to detect phosphorylated mTOR expression in new osteotylus at 15 days after operation. In addition, an in vitro cel stretch model was made in the mandibular mesenchymal stem cels from healthy Sprague-Dawley rats under resting tension force (6%, 4 hours); no distraction was done in control group. The ability of MSC-OB mobility, the expression of mTOR, Raptor, p70S6K and MMPs were evaluated using experiment methods including immunohistochemistry staining, real-time PCR and scratch assay.
RESULTS AND CONCLUSION: The expression of phosphorylated mTOR in MSC-OB was upregulated in the mandibular bone calus of the stretch group than the non-stretch group (P < 0.05). In thein vitro experiments, MSC-OB applied with mechanical stretch (6%, 4 hours) showed elevated gene expression levels of mTOR, Raptor, p70S6K, MMP-2, MMP-9 and MMP-13 compared with the control group (0%, 4 hours). Meanwhile, MSC-OB in the experiment group (6%, 4 hours) showed a greater ability of mobility, as demonstrated by a farther distance after 48 hours of observation (P < 0.05). The present study suggests that the enhancement of MSC-OB mobility correlates with increase of the gene expression of MMPs and mTOR signaling pathway. Mechanical stretch may promote MSC-OB migration through activation of mTOR/MMPs signaling pathway.

OBJECTIVE:To observe the outcomes of umbilical cord blood mesenchymal stem call transplantation for treating neural function of multiple system atrophy (MSA) patients.METHODS:A total of 20 MSA patients were selected at the Beijing Wu Stem Cells Medical Center from January to October 2008.All patients received treatment of vessel distention,anti-free radical,trophic nerve and call membrane stabilization,as well as umbilical cord blood mesenchymal stem call transplantation via intrathecal injection.Patients at left-lateral position,and body bent at hips,knees and necks.Acupuncture was conducted at the space of lumbar vertebra 3 and 4.Following local anesthesia,No.9 needle was directly pricked into the subarachnoid cavity.2 mg dexamethasone was slowly infused,and 5 mL (5×106 stem cells) umbilical cord blood mesenchymal stern call injection was obtained and slowly infused into the subarachnoid cavity within 10 minutes,once per week,four times as a course,totally one course.We adopted Unified Multiple System Atrophy Rating Scale (UMSARS) to evaluate those MSA patients.The higher score represented a severe pathogenetic condition.RESULTS:Compared with pretransplantation,the UMSARS score was significantly decreased in 20 patients 4 weeks follwing transplantation (P ＜ 0.01).After the treatment,patient's clinical symptoms such as slow movement,balance disturbance,orthostatic hypotension,urinary and bowel disorders had full obvious improvement.Graft versus host disease was not found.CONCLUSION:It is indicated that mesenchymal stem call transplantation is effective,can partly improve MSA patients' clinical symptoms,and improve patients' life quality.

To establish human beta-defensin-3 gene transgenic cell lines as competent donor cells for the production of transgenic animals using somatic cell nuclear transfer (SCNT). Firstly, we obtained human beta-defensin-3 by RT-PCR from human placenta, and subsequently inserted the fragment hBD into the corresponding site of the plasmid pBCP. Then we moved the combined fragment BCD (including 5' and 3' regulating region of beta-casein and hBD) into the corresponding site of the plasmid pEGFP-C1. Finally we successfully constructed mammary-specific expression vector pEBCD. We transected pEBCD into Holstein Fetal fibroblast cells by Lipofectamine TM-2000 and selected in medium with G418 for three to four weeks. We identified G418 resistant transfectants by PCR, RT-PCR and EGFP detection. Our results indicated that human beta-defensin-3 gene stably was integrated into the open region of the chromatin in G418 resistant fibroblast cells. Meanwhile we identified the expression of human beta-defensin-3 in the supernatant of stable transfected mammary epithelial cells by Western blotting. This study may provide competent transgenic donor cells for the production of transgenic animals by SCNT and improve the efficiency of transgenic cloning.
Animals ; Animals, Genetically Modified ; Caseins ; genetics ; Cattle ; Epithelial Cells ; metabolism ; Genes, erbB-1 ; genetics ; Genetic Vectors ; genetics ; Humans ; Mammary Glands, Animal ; cytology ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; beta-Defensins ; biosynthesis ; genetics

To discuss the effect of co-culture on the quality of mouse blastocysts and their epigenetic modification. We divided mouse zygotes into three co-culture experiment groups : with granular cells (group I ), oviduct epithelium cells (group II) and oviduct tissue (group III). Meanwhile, we set up control A (cultured in vitro, only KSOM (KCl+ simplex optimized medium)) and control B (cultured in vivo). Then we compared cleavage rate and blastocyst rate among different groups. After that we evaluated the quality of blastocysts by using ICM/TE (Inner cell mass/Trophectoderm cells) ratio via staining with propidium iodide and Hoechest333258, and analyzed the level of genome methylation and histone acetylation by immunofluorescence. Compared with the control group A, the co-culture groups had increased cleavage rate and blastocyst rate (P < 0.05), blastocyst cells and the ICM/TE ratio of co-culture groups were higher (P < 0.05), the level of genome methylation and histone acetylation had no significant difference between groups in vitro (P > 0.05), but the level of genome methylation in vivo was significantly higher than that of in vitro (P < 0.05). The co-culture methods can successfully promote the development rate of embryos in vitro, and improve the quality of the blastocyst. However, the methods have drawbacks in changing the abnormal genome methylation with in vitro culture.
Animals ; Blastocyst ; cytology ; Coculture Techniques ; DNA Methylation ; Epigenesis, Genetic ; genetics ; Epithelial Cells ; cytology ; Fallopian Tubes ; cytology ; Female ; Male ; Mice

Objective To compare the anesthetic effects of interscalene brachial plexus combined with ulnar nerve and axillary brachial plexus block guided by nerve stimulator. Methods Eighty patients belonging to ASA ⅠorⅡ and undergoing replantation of severed palm or wrist were divided randomly into 2 groups, Each group had 40 patients. Nerve stimulator guided nerve block. Patients in groupⅠreceived interscalene brachial plexus combined with ulnar nerve block, and those in groupⅡreceived axillary brachial plexus block. The onset time, hold time, tourniquet tolerance of radial nerve, median nerve and ulnar nerve of two groups was recorded. The phrenic nerve block, Horner′s syndrome and recurrent laryngeal nerve block was compared between two groups. Results The onset time of radial nerve, median nerve and ulnar nerve in group Ⅰwas (5.13 ± 0.76), (7.13 ± 1.04), (3.23 ± 0.62) min , in group Ⅱ was (9.23 ± 1.61), (12.35 ± 1.76), (8.83 ± 1.13) min, and there were significant differences (P<0.05). The excellent rates of sensory block of radial nerve, median nerve and ulnar nerve in group Ⅰ were 90.0%(36/40), 85.0%(34/40), 97.5%(39/40), in group Ⅱ were 72.5%(29/40), 65.0%(26/40), 70.0%(28/40), and there were significant differences (P<0.05). The full rates of motor block of radial nerve, median nerve and ulnar nerve in groupⅠwere 75.0%(30/40), 37.5%(27/40), 80.0%(32/40), in groupⅡ were 47.5%(19/40), 40.0%(16/40), 45.0%(18/40), and there were significant differences (P < 0.05). The tourniquet tolerance rate in group Ⅰwas significantly higher than that in groupⅡ:90.0%(36/40) vs. 62.5%(25/40) , P<0.05. In groupⅠ, phrenic nerve block occurred in 2 patients, and Horner syndrome occurred in 1 patient. None had laryngeal recurrent nerve block in both group. Conclusions The interscalene brachial plexus combined with ulnar nerve block guided by nerve stimulator is more suitable for a long time microsurgery of the palm or wrist, because it takes action faster, has better sensory and motor block effects, improves the rate of tourniquet tolerance without increasing untoward reaction.

OBJECTIVE: Based on metabonomics technology of high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) and hydrogen nuclear magnetic resonance spectroscopy (¹H NMR), the pharmacokinetic characteristics and therapeutic mechanism of Rhei Radix et Rhizoma (RhRR, Dahuang in Chinese), Eupolyphaga Steleophaga (EuS, Tubiechong in Chinese) combined with RhRR acting on acute liver injury were explored. METHODS: Models of acute liver injury were established, and the pharmacokinetic methods of five components of RhRR-EuS in rats were found by HPLC-MS/MS. The liver tissues of different groups of mice were analyzed by ¹H NMR spectroscopy combined with multivariate statistical analysis to investigate the metabolomics of RhRR-EuS and RhRR. RESULTS: Pharmacokinetic results showed there were different levels of bimodal phenomenon in different groups, and the absorption of free anthraquinone in RhRR increased after compatibility with EuS. In addition, the pathological state of acute liver injury in rats can selectively promote the absorption of emodin, chrysophanol, physcion and aloe emodin. Through 15 differential metabolites in the liver tissue of acute liver injury mice, it was revealed that RhRR-EuS and RhRR could protect the liver injury by regulating the metabolism of glutamine and glutamic acid, alanine, aspartic acid and glutamic acid, and phosphoinositide. However, the regulation of RhRR was weaker than that of RhRR-EuS. CONCLUSION For the first time, we studied the pharmacokinetics and metabolomics differences of RhRR-EuS and RhRR in rats and mice with acute liver injury, in order to provide theoretical reference for clinical treatment of liver disease by DHZCP.

Objective Mitochondrial reactive oxygen species(mtROS)could cause damage to pancreatic β-cells,rendering them susceptible to oxidative damage.Hence,investigating the potential of the mitochondria-targeted antioxidant(Mito-TEMPO)to protect pancreatic β-cells from ferroptosis by mitigating lipid peroxidation becomes crucial. Methods MIN6 cells were cultured in vitro with 100 μmol/L sodium palmitate(SP)to simulate diabetes.FerroOrange was utilized for the detection of Fe2+fluorescence staining,BODIPY581/591C11 for lipid reactive oxygen species,and MitoSox-Red for mtROS.Alterations in mitophagy levels were assessed through the co-localization of lysosomal and mitochondrial fluorescence.Western blotting was employed to quantify protein levels of Acsl4,GPX4,FSP1,FE,PINK1,Parkin,TOMM20,P62,and LC3.Subsequently,interventions were implemented using Mito-TEMPO and Carbonyl cyanide 3-chlorophenylhydrazone(CCCP)to observe changes in ferroptosis and mitophagy within MIN6 cells. Results We found that SP induced a dose-dependent increase in Fe2+and lipid ROS in MIN6 cells while decreasing the expression levels of GPX4 and FSP1 proteins.Through bioinformatics analysis,it has been uncovered that mitophagy assumes a crucial role within the ferroptosis pathway associated with diabetes.Additionally,SP decreased the expression of mitophagy-related proteins PINK1 and Parkin,leading to mtROS overproduction.Conversely,Mito-TEMPO effectively eliminated mtROS while activating the mitophagy pathways involving PINK1 and Parkin,thereby reducing the occurrence of ferroptosis in MIN6 cells.CCCP also demonstrated efficacy in reducing ferroptosis in MIN6 cells. Conclusion In summary,Mito-TEMPO proved effective in attenuating mtROS production and initiating mitophagy pathways mediated by PINK1 and Parkin in MIN6 cells.Consequently,this decreased iron overload and lipid peroxidation,ultimately safeguarding the cells from ferroptosis.