Autophagy dysregulation, mitochondrial dynamic abnormality and cell cycle re-entry are implicated in the vulnerable neurons of patients with Alzheimer's disease. This study was designed to testify the association among autophagy, mitochondrial dynamics and cell cycle in dividing neuroblastoma (N2a) cells. The N2a cells were cultured in vitro and treated with different concentrations of 3-methyladenine (3-MA). The cell viability was detected by methyl thiazolyl tetrazolium (MTT) assay. They were randomly divided into control group (cells cultured in normal culture medium) and 3-MA group (cells treated with 10 mmol/L 3-MA). The cell cycle was analyzed in the two groups 3, 6, 12, and 24 h after treatment by flow cytometry. Western blotting was used to evaluate the expression levels of mitofission 1 (Fis1), mitofusin 2 (Mfn2), microtubule-associated protein 1 light chain 3 (LC3), cell cycle-dependent kinase 4 (CDK4) and cdc2. The flow cytometry revealed that the proportion of cells in G(2)/M was significantly increased, and that in G0/G1 was significantly reduced in the 3-MA group as compared with the control group. Western blotting showed that the expression levels of Fis1, LC3, and CDK4 were significantly up-regulated in the 3-MA group at the four indicated time points as compared with the control group. Mfn2 was initially decreased in the 3-MA group, and then significantly increased at 6 h or 12 h. Cdc2 was significantly increased in the 3-MA group at 3 h and 6 h, and then dropped significantly at 12 h and 24 h. Our data indicated that 3-MA-induced suppressed autophagy may interfere with the cell cycle progression and mitochondrial dynamics, and cause cell death. There are interactions among cell cycle, mitochondrial dynamics and autophagy in neurons.

Autophagy dysregulation, mitochondrial dynamic abnormality and cell cycle re-entry are implicated in the vulnerable neurons of patients with Alzheimer's disease. This study was designed to testify the association among autophagy, mitochondrial dynamics and cell cycle in dividing neuroblastoma (N2a) cells. The N2a cells were cultured in vitro and treated with different concentrations of 3-methyladenine (3-MA). The cell viability was detected by methyl thiazolyl tetrazolium (MTT) assay. They were randomly divided into control group (cells cultured in normal culture medium) and 3-MA group (cells treated with 10 mmol/L 3-MA). The cell cycle was analyzed in the two groups 3, 6, 12, and 24 h after treatment by flow cytometry. Western blotting was used to evaluate the expression levels of mitofission 1 (Fis1), mitofusin 2 (Mfn2), microtubule-associated protein 1 light chain 3 (LC3), cell cycle-dependent kinase 4 (CDK4) and cdc2. The flow cytometry revealed that the proportion of cells in G(2)/M was significantly increased, and that in G0/G1 was significantly reduced in the 3-MA group as compared with the control group. Western blotting showed that the expression levels of Fis1, LC3, and CDK4 were significantly up-regulated in the 3-MA group at the four indicated time points as compared with the control group. Mfn2 was initially decreased in the 3-MA group, and then significantly increased at 6 h or 12 h. Cdc2 was significantly increased in the 3-MA group at 3 h and 6 h, and then dropped significantly at 12 h and 24 h. Our data indicated that 3-MA-induced suppressed autophagy may interfere with the cell cycle progression and mitochondrial dynamics, and cause cell death. There are interactions among cell cycle, mitochondrial dynamics and autophagy in neurons.
Adenine ; administration & dosage ; analogs & derivatives ; Apoptosis ; drug effects ; Autophagy ; drug effects ; genetics ; CDC2 Protein Kinase ; Cell Cycle ; drug effects ; genetics ; Cell Division ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cell Survival ; drug effects ; Cyclin B ; biosynthesis ; Cyclin-Dependent Kinases ; Gene Expression Regulation ; drug effects ; Humans ; Membrane Proteins ; biosynthesis ; Microtubule-Associated Proteins ; biosynthesis ; Mitochondrial Dynamics ; drug effects ; genetics ; Mitochondrial Proteins ; biosynthesis ; Neuroblastoma ; Signal Transduction ; drug effects

OBJECTIVETo explore the status of clenbuterol pollution in swine products in Beijing city in 2002.

METHODSEuropean Union method (EUR 15127-EN Cy2.3) was adopted to examine the samples. Samples were screened by enzyme-linked immunosorbent assay (ELISA) and confirmed by GC-MS. Detected limit of the method was 0.5 micro g/kg. Samples, including lung, liver, pork, kidney and urine of swine, were collected from slaughterhouses, refrigeratories and markets in 11 districts of Beijing.

RESULTSThe results indicated that 185 out of 1 379 samples were positive with an annual positive rate of 13.4%. The highest was 15.7% in lung of swine, followed by urine 15.2% and pork liver 14.0%.

CONCLUSIONRates of detection had decreased from 30.0% to 2.7% during 2002.

Animals ; China ; Clenbuterol ; analysis ; Drug Residues ; analysis ; Food Contamination ; analysis ; Humans ; Meat ; analysis ; Meat Products ; Swine

OBJECTIVETo study the clinicopathological features of the Chinese hereditary non-polyposis colorectal cancer and its germline mutation of hMLH(1) and hMSH(2).

METHODSThirteen typical Chinese hereditary non-polyposis colorectal carcinoma (HNPC)C kindreds and 19 non-typical HNPCC families were registered and followed up. The germline mutation of the hMLH(1) and hMSH(2) of 12 index cases of 6 typical and 6 non-typical HNPCC were screened by PCR-SSCP. Samples with abnormal mobility were sequenced directly.

RESULTSThe average age of typical HNPCC was 47, no difference existed between sexes. Location of the tumors of typical HNPCC represented 44.7% on the right half colon and non-typical HNPCC 65.8% on the rectum. The rate of the metachronos cancer was 11.5%. The 3-, 5-and 10-year survival rate was 64.0%, 45.3% and 31.2% respectively. Among 12 cases, 8 showed abnormal mobility. Except for an intron polymorphism, six exons abnormalities were found in 5 of 12 proband. Sequencing showed 4 missense, 7 insertion and a nonsense mutations.

CONCLUSIONSChinese HNPCC is early onset, more common on proximal colon and better prognosis. Mutation of hMSH(2) is dominant in the Chinese typical HNPCC, but mutation of hMLH(1) is more common in the non-typical group.

Adaptor Proteins, Signal Transducing ; Adult ; Asian Continental Ancestry Group ; genetics ; Carrier Proteins ; genetics ; Colorectal Neoplasms, Hereditary Nonpolyposis ; genetics ; pathology ; DNA Mutational Analysis ; Female ; Follow-Up Studies ; Germ-Line Mutation ; Humans ; Male ; Middle Aged ; MutL Protein Homolog 1 ; MutS Homolog 2 Protein ; genetics ; Mutation, Missense ; Nuclear Proteins ; genetics ; Pedigree ; Polymerase Chain Reaction

OBJECTIVETo investigate the clinicopathological features of Chinese hereditary nonpolyposis colorectal cancer (HNPCC).

METHODSPatients who met the Amsterdam criteria were enrolled in this study from several hospitals in China. Clinicopathological features of patients with HNPCC were compared between the patients with suspected HNPCC and sporadic colorectal cancer.

RESULTSOne hundred and sixty-seven individuals from 31 families met the Amsterdam criteria. The average age was 48.6 (22-78) years old. There were 43 cases (31.9%) with ascending colon cancer and 52 cases (38.5%) with rectal cancer. The 3-, 5-, 10- years survival rate was 70.3%, 49.9% and 39.7% respectively. The incidence of multiple primary neoplasms was 20.4% .

CONCLUSIONSChinese HNPCC is characterized by early disease onset. Rectal cancer and ascending colon cancer are the first and the secondly common cancer for Chinese HNPCC. Gastric cancer is the most common parenteral cancer in Chinese HNPCC families.

Adult ; Aged ; Asian Continental Ancestry Group ; China ; epidemiology ; Colorectal Neoplasms, Hereditary Nonpolyposis ; epidemiology ; genetics ; pathology ; Female ; Humans ; Incidence ; Male ; Middle Aged ; Prognosis ; Survival Rate ; Young Adult

OBJECTIVETo explore methods of preventing and reversing rejection after simultaneous pancreas-kidney transplantation (SPK).

METHODSSeventeen patients performed SPK operation from Sep, 1999 to Sep, 2003 were reviewed retrospectively. Immunosuppression was achieved by triple regimen consisting of cyclosporine, mycophenolate mofetil (MMF)/azathioprine and steroid. 2 patients were treated with Dalizumab, the other three patients used OKT3 as immune induction.

RESULTS1 patient experienced the accelerated rejection, the pancreas and kidney grafts were resected because of failure of conservative therapy. 8 patients experienced renal acute rejection, 2 cases suffered from pancreas acute rejection at the same time. All these patients received daily high dose pulse steroid for 3 days. OKT3 was administered in 2 patients with steroid resistance rejection. All the grafts were successfully rescued.

CONCLUSIONSReasonable application of immunosuppression after SPK operation and adoption of systemic measures which can reduce sensitivity of high risk receptor before SPK operation are the effective methods of preventing and treating rejection.

Administration, Oral ; Adult ; Azathioprine ; administration & dosage ; Cyclosporine ; administration & dosage ; Diabetic Nephropathies ; surgery ; Drug Therapy, Combination ; Female ; Glucocorticoids ; administration & dosage ; Graft Rejection ; prevention & control ; Humans ; Immunosuppressive Agents ; administration & dosage ; Kidney Transplantation ; immunology ; Male ; Middle Aged ; Pancreas Transplantation ; immunology ; Prednisolone ; administration & dosage ; Retrospective Studies ; Transplantation, Homologous

OBJECTIVETo investigate the specificity and sensitivity of the immunohistochemistry for hMLH1 and hMSH2 with detection of microsatellite instability (MSI) to identify the kindreds with hereditary nonpolyposis colorectal cancer and to analyse its value in clinical practice.

METHODSpecimens of 16 cases with HNPCC and 16 cases with sporadic colorectal cancer were detected by immunostaining with hMLH1 and hMSH2 and MSI was also detected.

RESULTSThe specificity and sensitivity of the immunohistochemistry for hMLH1 and hMSH2 were 91.7% and 87.5% respectively. The specificity and sensitivity of MSI were 100% and 75.0%. By combining two methods, the specificity and sensitivity were 91.7% and 93.8% respectively.

CONCLUSIONSBy combination of the immunohistochemistry for hMLH1 and hMSH2 and detection of MSI to identify the kindreds with HNPCC, the specificity and sensitivity are improved which is better than to use either of them alone. And it is very easy and cheap that it can be used in clinics.

Adaptor Proteins, Signal Transducing ; Adult ; Aged ; Carrier Proteins ; Colorectal Neoplasms, Hereditary Nonpolyposis ; genetics ; DNA-Binding Proteins ; analysis ; Female ; Genomic Instability ; Humans ; Immunohistochemistry ; Male ; Microsatellite Repeats ; Middle Aged ; MutL Protein Homolog 1 ; MutS Homolog 2 Protein ; Neoplasm Proteins ; analysis ; Nuclear Proteins ; Proto-Oncogene Proteins ; analysis ; Sensitivity and Specificity

AIM To evaluate the quality of Beidougen Formula Granules.METHODS Fifteen batches of standard decoctions and three batches of formula granules were prepared,after which paste rate and contents,transfer rates of magnoflorine,daurisoline,dauricine were determined.HPLC specific chromatograms were established,and cluster analysis was adopted in chemical pattern recognition.RESULTS For three batches of formula granules,the paste rates were 15.1%-16.6%,the contents of magnoflorine,daurisoline,dauricine were 18.93-19.39,9.42-9.60,6.79-6.85 mg/g with the transfer rates of 34.42%-35.25%,43.81%-44.65%,27.27%-27.51%from decoction pieces to formula granules,respectively,and there were seven characteristic peaks in the specific chromatograms with the similarities of more than 0.95,which demonstrated good consistence with those of standard decoctions and accorded with related limit requirements.Fifteen batches of standard decoctions were clustered into two types,and the medicinal materials produced from Jilin,Hebei,Shangdong could be used for the preparation of formula granules.CONCLUSION This reasonable and reliable method can provide references for the quality control and clinical application of Beidougen Formula Granules.

OBJECTIVETo Explore a two-step culture system to generate a large number of dendritic cells (DC) differentiated from cord blood (CB) CD(34)(+) cells.

METHODSEnriched CB CD(34)(+) cells with immunoadsorption were primarily cultured in the presence of stem cell factor (SCF), Flt-3 ligand (FL), thrombopoietin (Tpo) and interleukin-3 (IL-3) for 7 (group I), 10 (group II) or 14 days (group III) respectively, and then further cultured with GM-CSF, IL-4 and TNF-alpha for 5 - 8 days to induce DC. The expansion and cell function were evaluated by flow cytometry (FCM) and mix-lymphocyte reaction (MLR), and detection of IL-12 in the supernatant by using ELISA.

RESULTSThe total nucleated cells with 53.39 +/- 20.59-, 307.17 +/- 119.59- and 1117.25 +/- 335.49-folds expansion could be respectively obtained after 7 - 14 days of expansion culture. After DC induction, CD(1a)(+) cells were 21.40 +/- 16.70-, 143.2 +/- 60.35- and 150.8 +/- 42.16-fold increase as compared to the initial nucleated cells. Comparing with that in group I, the CD(1a)(+) cells were much more in groups II and III; but there was no difference between the latter two groups (P > 0.05). The cultured cells in the three groups showed almost the same allo-stimulatory capability and IL-12 excretion when the second culture duration maintained 8 days, while the capability and excretion were greatly decreased when the duration shortened to 5 days (P < 0.05).

CONCLUSIONA plenty of functionally mature DC could be obtained from the CD(34)(+) cells in the two-step culture system of 7 - 10 days HSC expansion followed by 8 days DC induction.

Antigens, CD34 ; analysis ; Cell Differentiation ; Cells, Cultured ; Dendritic Cells ; cytology ; physiology ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; Humans ; Interleukin-12 ; biosynthesis ; Lymphocyte Activation

In order to discuss the "entropy weight method" for weighting various indicators in the comprehensive evaluation of Angelicae Sinensis Radix slices(ASR), the quality of ASR was comprehensively evaluated by entropy weight-based gray systematic theory and cluster analysis. In this study, the contents of ferulic acid, volatile oil, polysaccharide, alcohol extract, water extract, moisture, total ash and acid-insoluble ash in 44 batches of ASR from different sources were determined. The entropy weight method was used for objective weighting. With relative correlation(r_i) as a measure, a multi-index comprehensive evaluation model was constructed for the quality of ASR. The results showed that the relative correlation value of 44 batches of ASR ranged from 0.301 9 to 0.662 9. There were certain differences in the quality of ASR from different sources. The ASR S1-S8, traceable and standardized in processing techno-logy, showed a high relative correlation degree and high quality ranking, indicating that the implementation of systemic management of the production chain of Chinese herbal pieces was beneficial to the quality control of ASR. The quality evaluation results of 44 batches of ASR were consistent with those of traditional geo-authentic habitats for ASR and the mainstream varieties of ASR on market, and basically consistent with the results of cluster analysis. This study suggests that the gray systematic theory based on the entropy weighting method can be used for the quality evaluation of ASR. The objective weighting of the entropy weight method improves the reliability of the gray correlation method and the scientificity of ASR quality evaluation.
Angelica sinensis ; Drugs, Chinese Herbal ; Entropy ; Oils, Volatile ; Plant Roots ; Reproducibility of Results