Objective To investigate the changes of expression of stromal cell-derived factor 1α (SDF-1α) in retinas after partial optic nerve injury in rats. Methods Models with injury of partial optic nerve were induced in rats. Retinal tissues were collected 1,2,3,5,7,10 and 14 d after injury. Enzyme linked immunosorbent assay and Real-time quantitative PCR were employed to detect the expression of SDF-1α protein and mRNA in retinal tissues respectively in injury group (n=28), sham operated group (n=28) and normal control group (n=12). Results The expression of SDF-1α protein and mRNA in retinas was higher than that in sham operated group and normal control group at different time points after injury (P<0.01), and it reached the peak at the 5th day after injury. The expression of SDF-1α protein and mRNA maintained a high level at the 14th day after injury. Conclusion The expression of SDF-1α protein and mRNA is up-regulated after partial optic nerve injury, and may last for a long time.

OBJECTIVETo observe the differential effect of joint ultrasound on the syndrome differentiation of rheumatoid arthritis (RA) by observing the high frequency ultrasound performances among inactive stage and different syndromes in active stage.

METHODSTotally 83 RA patients in the active stage were assigned to the dampness heat syndrome group (DHS, 59 cases)and the cold dampness syndrome group (CDS, 24 cases) according to Chinese medicine (CM) syndrome typing. Besides, 20 RA patients in the remission stage were recruited as the control group (abbreviated as the remission group). By using high frequency ultrasound and power Doppler ultrasound technology, a comparative observation of synovitis, tenosynovitis, synovial blood flow, and bone erosion in the 2nd-5th metacarpophalangeal (MCP) joints, proximal interphalangeal (PIP) joints, wrist joints, knee joints, the second and the fifth metatarsophalangeal (MTP) joints (a total of 24 joints) was performed in all patients. Correlation analyses were performed between the ultrasound performance, laboratory indices, and the disease activity. Ultrasound data of each RA patient were analyzed by their total scores. Χ2 test was used for enumeration data. The measurement data was expressed as x ± s. One-way ANOVA was used for data of normal distribution, while non- parametric test was used for data of non-normal distribution. Correlation analysis of two variables was performed for clinical indicators and ultrasound indicators. Its significance was detected using Pearson correlation.

RESULTSCompared with the remission group, the severity degree of synovitis, tenosynovitis, synovial blood flow, and bone erosion significantly increased in the DHS group (P < 0.01). There was statistical difference in ESR, CRP, anti-CCP, DAS28 score, and the positive rate of RF (P < 0.05, P < 0.01). There was statistical difference in the severity degree of synovitis and synovial blood flow, and DAS28 score in the CDS group (P < 0.05). Compared with the CDS group, there was statistical difference in the four ultrasound indices (P < 0.05, P < 0.01), ESR, CRP, anti-CCP, DAS28 score, and the positive rate of RF in the DHS group (P < 0.05, P < 0.01). There was no statistical difference in G, IgG, IgA, or IgM among the three groups (P > 0.05). There existed positive correlation between ESR and the synovitis degree, synovial blood flow, and bone erosion in the DHS group (r = 0.444, 0.397, 0.486, P < 0.05).There existed positive correlation between ESR and the synovitis degree, bone erosion, and synovial blood flow in the DHS group (r = 0.378, 0.270, P < 0.05). There existed positive correlation between the DAS28 score and the synovitis degree and synovial blood flow in the DHS group (r = 0.304, 0.351, P < 0.05).

CONCLUSIONSThe inflammation degree was the most severe in RA patients of DHS. High frequency ultrasound could provide better evidence for Chinese medical syndrome differentiation of RA patients.

Arthritis, Rheumatoid ; diagnostic imaging ; Humans ; Medicine, Chinese Traditional ; Metacarpophalangeal Joint ; ultrastructure ; Syndrome ; Synovitis ; diagnostic imaging ; Ultrasonography

Objective:To use polyamidoamine(PAMAM)dendrimer as gene delivery system for survivin gene anti- sense oligodeoxynucleotide(asODN)transfection for inhibition of HepG2 cancer cell growth.Methods:The first to the fifth generation of PAMAM and asODN were used to prepare a complex:PAMAM-asODN.The morphology of PAMAM- asODN was observed using agrose electrophoresis and atomic force microscope(AFM).PAMAM-asODN was then used to transfect HepG2 cells and cells transfected with asODN served as control.The transfection efficacy of PAMAM-asODN into HepG2 cells was observed under confocol microscope,the surviving mRNA expression was analyzed by RT-PCR,and the inhibition of HepG2 cell growth was determined by MTT assay.Results:Agrose electrophoresis showed strong complexing action between PAMAM and asODN and they formed a complex with a diameter of 25 nm.Confocol microscope showed the transfection efficacy of PAMAM-asODN was higher than that of asODN.RT-PCR showed a decreased expression of sur- vivin mRNA in PAMAM-asODN transfected cells.MTF results demonstrated that the growth of HepG2 cell was obviously inhibited after transfection of PAMAM-asODN and the inhibition rate increased with culture time,concentration of com- plex,the generation of PAMAM.PAMAM-asODN at 6.0?mol/L G4.0 resulted in a 55% inhibition of HepG2 cells 96 h after culture.Conclusion:PAMAM dendrimers can efficiently mediate the entry of survivin asODN into HepG2 cells,re- sulting in inhibition of HepG2 cells.PAMAM might be a promising gene carrier for potential molecular therapy of cancer.

OBJECTIVETo observe the effect of Cordyceps sinensis (CS) powder on renal oxidative stress and mitochondria functions in 5/6 nephrectomized rats, and to primarily explore its possible mechanisms.

METHODSTotally 30 male Sprague-Dawley rats were divided into the sham-operation group, the model group, and the treatment group by random digit table, 10 in each group. A chronic kidney disease (CKD) rat model was prepared by one step 5/6 nephrectomy. Rats in the treatment group were intragastrically administered with CS powder solution at the daily dose of 2 g/kg, once per day. Equal volume of double distilled water was intragastrically administered to rats in the sham-operation group and the model group. All medication lasted for 12 weeks. The general condition of rats, their body weight, blood pressure, 24 h proteinuria, urinary N-acetyl-β-D-glucosaminidase (NAG), serum creatinine (SCr) , and blood urea nitrogen (BUN) were assessed before surgery, at week 2, 4, 6, 8, 10, and 10 after surgery. Pathological changes of renal tissues were observed under light microscope. Morphological changes of mitochondria in renal tubular epithelial cells were observed under transmission electron microscope. Activities of antioxidant enzymes including reduced glutathione (GSH), manganese superoxide dismutase (MnSOD), and malondialdehyde (MDA) in fresh renal tissue homogenate were detected. Mitochondria of renal tissues were extracted to detect levels of mitochondrial membrane potential and changes of reactive oxygen species (ROS). And expressions of cytochrome-C (Cyto-C) and prohibitin in both mitochondria and cytoplasm of the renal cortex were also measured by Western blot.

RESULTS(1) Compared with the sham-operation group, body weight was significantly decreased at week 2 (P <0. 01), but blood pressure increased at week 4 (P <0. 05) in the model group. Compared with the model group, body weight was significantly increased at week 12 (P <0. 01), but blood pressure decreased at week 8 (P < 0. 01) in the treatment group. (2) Compared with the sham-operation group, 24 h proteinuria, urinary NAG, blood SCr and BUN significantly increased in the model group (all P <0. 01). Compared with the model group, blood and urinary biochemical indices all significantly decreased in the treatment group (all P <0. 01). (3) Results of pathological renal scoring: Glomerular sclerosis index, scoring for tubulointerstitial fibrosis, degree of tubulointerstitial inflammatory infiltration were all obviously higher in the model group than in the sham-operation group (all P <0. 01). All the aforesaid indices were more obviously improved in the treatment group than in the model group (all P <0. 01). (4) Compared with the sham-operation group, activities of MnSOD and GSH-Px were significantly reduced, but MDA contents obviously increased in the renal cortex of the model group (all P <0. 01). Compared with the model group, activities of MnSOD and GSH-Px obviously increased (P <0. 05, P <0. 01), but MDA contents obviously decreased in the renal cortex of the treatment group (P <0. 01). (5) Compared with the sham-operation group, the mitochondrial membrane potential significantly decreased, but ROS levels significantly increased in the model group (all P <0.01). Compared with the model group, mitochondrial transmembrane potential increased in the treatment group, thereby inhibiting the tendency of increased production of ROS (both P < 0. 01). (6) Results of Western blot showed that, compared with the sham-operation group, expression levels of mitochondrial Cyto-C and Prohibitin were significantly reduced in the renal cortex (P <0. 01), but significantly elevated in the cytoplasm of the model group (P <0. 01). Compared with the model group, each index was obviously improved in the treatment group with statistical difference (P <0. 05, P <0. 01).

CONCLUSIONCS powder had renal protection, and its mechanism might partially depend on in- hibition of oxidative stress and protection for mitochondria.

Acetylglucosaminidase ; metabolism ; Animals ; Blood Urea Nitrogen ; Cordyceps ; Drugs, Chinese Herbal ; pharmacology ; Kidney ; Kidney Cortex ; Kidney Diseases ; Kidney Function Tests ; Male ; Malondialdehyde ; metabolism ; Mitochondria ; Nephrectomy ; Oxidative Stress ; drug effects ; Proteinuria ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism

OBJECTIVETo investigate the significance of Rac subfamily members in the gastrointestinal carcinogenesis and progression.

METHODSThe mRNA expression of Rac1, Rac2 and Rac3 in 12 kinds of gastrointestinal cancer cell lines was examined by semi-quantitative RT-PCR. The activities of Rac1 protein in 5 kinds of gastric cancer cell lines were tested by pull-down assay.

RESULTSCompared with the normal gastric mucosa and intestinal epithelial cell line, the mRNA expression of Rac1 and Rac3 was up-regulated in most of gastrointestinal cancer cell lines. The activities of Rac1 protein increased markedly in gastric cancer cell lines.

CONCLUSIONThe increased mRNA expression of Rac1 and Rac3 in gastrointestinal cancer cell lines and the abnormal activation of Rac1 protein in gastric cancer cell lines might be correlated with the carcinogenesis of gastrointestinal cancer.

Cell Line, Tumor ; Gastrointestinal Neoplasms ; metabolism ; Humans ; RNA, Messenger ; analysis ; Reverse Transcriptase Polymerase Chain Reaction ; rac GTP-Binding Proteins ; genetics ; rac1 GTP-Binding Protein ; analysis ; genetics

OBJECTIVEThe combination of oxaliplatin (L-OHP), 5-fluorouracil (5-Fu) and folinic acid (FA), being one of the effective regimens for advanced gastric cancer, is used in form of chronomodulated chemotherapy for advanced gastric cancer to investigate its efficacy and safety.

METHODSTwenty-six patients received a 4-day chronomodulated infusion of L-OHP, 5-Fu and FA. L-OHP (25 mg.m(-2).d(-1)) infused from 10:00 am to 22:00 pm, and followed by 5-Fu (600 mg.m(-2).d(-1)) and FA (300 mg.m(-2).d(-1)) from 22:00 pm to 10:00 am for 4 days using a multichannel programmable pump, every 2 weeks as an cycle for at least 2 cycles.

RESULTSTwenty-six patients with previously untreated advanced gastric cancer were eligible. Two complete and 13 partial remissions were observed with an overall response rate of 57.7%. Stable disease was observed in 6 patients (23.1%) and progressive disease in five (19.2%). Four of these patients underwent surgery. The median remission time was 3.5 months and time to tumor progression (TTP) was 4.5 months. The median overall survival time was 8 months. A total of 80 cycles were given without any grade 4 toxicity observed, but grade 3 thrombocytopenia (1.3%) and mucositis (1.3%) in one patient, two grade 3 neutropenia (2.5%) and nausea/vomiting (2.5%) in 2.

CONCLUSIONChronomodulated intravenous chemotherapy of oxaliplatin, 5-fluorouracil and folinic acid is effective and safe in the treatment of advanced gastric cancer.

Adenocarcinoma ; drug therapy ; Adult ; Aged ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Chronotherapy ; Drug Administration Schedule ; Female ; Fluorouracil ; administration & dosage ; Humans ; Leucovorin ; administration & dosage ; Male ; Middle Aged ; Organoplatinum Compounds ; administration & dosage ; Stomach Neoplasms ; drug therapy ; Treatment Outcome

OBJECTIVETo evaluate the inhibitory effects of survivin antisense oligonucleotide (survivin-ASODN) mediated by polyamidoamine dendrimer (PAMAM) against the growth of subcutaneously transplanted colorectal cancer in nude mice.

METHODSNude mouse models bearing colorectal cancer was established by subcutaneous injection of SW620 cells. Survivin- OSADN (300 microg/L) was mixed with 4.06 microg/L PAMAM or liposome to prepare two transfection complexes, and their morphologies were observed by transmission electron microscope. The particle size of the prepared complexes was determined by laser particle size analyzer, and the zeta potential was measured. The encapsulation efficiency and the DNA release rate in vitro were determined by ultraviolet spectrophotometer. The transfection complexes were then directly injected into the xenografts of the tumor-bearing nude mice. The tumor volume changes were observed, and the expression of survivin in the transplanted tumor was measured by Western blotting.

RESULTSThe PAMAM-survivin-ASODN complex had a significantly smaller diameter and greater zeta potential than liposome-survivin-ASODN (P<0.01 and 0.05, respectively). The encapsulation efficiency was comparable between the two complexes. In in vitro condition, PAMAM-survivin-ASODN allowed sustained survivin-ASODN release for as long as 14 days, as compared with the 5 days for the liposome complex. After injection into the tumor xenografts, PAMAM-survivin- ASODN resulted in significantly lower expression of survivin protein in the transplanted tumors (P<0.05), and also in significantly greater reduction of the tumor volume than the liposome complex (P<0.05).

CONCLUSIONPAMAM can effectively deliver survivin-ASODN into transplanted colorectal tumor cells to reduce the expression of survivin and inhibit the tumor growth.

Animals ; Cell Proliferation ; drug effects ; Colorectal Neoplasms ; pathology ; Dendrimers ; Humans ; Inhibitor of Apoptosis Proteins ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Microtubule-Associated Proteins ; genetics ; pharmacology ; Neoplasm Transplantation ; Oligonucleotides, Antisense ; pharmacology ; Polyamines ; pharmacology ; Repressor Proteins ; Tumor Cells, Cultured

OBJECTIVETo study the expression and possible function of RhoA in human gastric cancer cell lines.

METHODSThe expression of RhoA in human gastrointestinal cancer cell lines was detected by Western blot. Antisense plasmid of RhoA was constructed by pGEFL and transferred into gastric cancer cell line AGS by lipofectamine. Cell survival was examined by MTT assays, and cell cycle was detected by flow cytometry.

RESULTSThe expression of RhoA protein in 10 different kinds of human cancer cell lines was much higher than that in immortalized human intestinal epithelial cell line. After being transfected with antisense RhoA, with the decrease in RhoA protein expression, the growth rate of AGS was inhibited, and the number of cells in S phase was increased by 14%.

CONCLUSIONRhoA is overexpressed in many human cancer cell lines. Some of the malignant characteristics of a gastric cancer cell line can be partially reversed by inhibiting RhoA expression.

Antisense Elements (Genetics) ; pharmacology ; Cell Cycle ; Cell Line, Tumor ; Genetic Therapy ; Humans ; Stomach Neoplasms ; chemistry ; pathology ; therapy ; rhoA GTP-Binding Protein ; analysis ; antagonists & inhibitors ; physiology

Objective To investigate the effect ofstromal cell derived factor-1 (SDF-1) on the regulation of neural stem cells (NSCs) migration.Methods NSCs were obtained from the cerebral cortex of embryonic rats and cultured in serum-free medium,and their stem cell properties were assessed by means of induced differentiation in vitro into neurons and astrocytes.After in vitro cell culture,the purity of NSCs and the co-expression rate of CXCR4/nestin were detected by flow cytometry.Blind-well chambers were employed to detect the chemotactic effects of SDF-1 by counting the cells which had crossed a 8 μm pore membrane when confronted with varying concentrations of SDF-1 (0,1,10,50,100,500 and 1000 ng/mL),and the distribution of cells migrated out of the same neurosphere was overviewed by μ-slides in the persistent concentration gradient of SDF-1.Results Neurospheres were formed by persistent proliferation of NSCs, which were capable of differentiating into neurons (β-tubulin+) and astrocytes (GFAP+) in media without mitogens,and flow cytometry analyses showed that most of the cultured cells expressed nestin and the co-expression rate of CXCR4/nestin was nearly 80%.SDF-1 showed great chemotaxis to NSCs,and the amount of cells having migrated through the membrane in 500 ng/ml SDF-1 group was higher than that in other groups (P<0.05).When the cells were confronted with a linear concentration gradient (from 500 to 0 ng/mL),which was generated by diffusion and stable for at least 48 h,the cells migrated out ofa neruosphere could distribute irregularly with more cells locating in the region of higher concentration of SDF-1 and longer migration distance away from the center of the neurosphere than the opposite.Conclusion SDF-1 binding to its specific receptor CXCR4 was capable of inducing NSCs migrating directionally to the source of SDF-1.

To further explore the mechanism of congenital pyrimidine 5'-nuleotidase I (P5'N-I) deficiency, on the basis of purification of the protein, the molecular weight and amino acid composition were analysed by mass-spectrograph and amino-acid analyzer, microsequencing and bioinformation analysis of P5'N-I were performed after it was hydrolysed by trypsin. The results showed that three fractions were found in the purified P5'N-I and their molecular weights were 26,952.9, 55,476 and 110,938, respectively. The sequence from one of the peptide fragments was I-E-G-P-T-I-R-Q-I-E. The homologous sequence was not found after comparision with the ten-amino-acid sequence in GenBank by blast procedure. Amino acid analysis indicated that P5'N-I was composed of 18 amino acids at least, and 243 amino acid residues. In conclusion, the enzyme might be an allosteric enzyme, there might be homologous dimer or tetramer in physiological status of normal human erythrocyte, the microsequence could be designed as the probe for fishing the genes of interest. The composition of amino acid might be an important information in determination of its protein primary structure.
5'-Nucleotidase ; blood ; chemistry ; isolation & purification ; Amino Acid Sequence ; Amino Acids ; analysis ; Chromatography, High Pressure Liquid ; Electrophoresis, Polyacrylamide Gel ; Erythrocytes ; enzymology ; Humans ; Mass Spectrometry ; Molecular Weight ; Peptide Fragments ; chemistry ; Sequence Analysis, Protein