Objective To determine the influence of protein fusion on the biological characteris- tics of hymidine kinase(TK)and human immunodeficiency virus(HIV)Tat recombinant protein. Methods By utilizing polymerase chain reaction(PCR)technique,different fragments containing two,four or six glycines(Gly)were inserted between the HIV Tat gene and TK,and cloned into PBK vector.After testified by sequencing,the vectors were transfected into E coli.After induced by iso- propyl thiogalactose(IPTG),bacilli were collected and destructed by ultrasonic,the fusion proteins were determined by monoclonal antibody against HIV protein.HepG2 cells were incubated in DMEM supplement with 10?g/mL HIV-Gly(n)-TK(n=0,2,4,6)fusion protein,TK-HIV Tat and only HIV Tat.HepG2 cells in different groups were detected by immunofluorescence assay 24 hours after transduction with HIV Tat monoclonal antibody.The rate of apoptosis after cells were incubated with gencilovir(10?g/mL)for 3 days was determined by cell flow cytometry,while survival cell ratio was recorded by trypan blue.The data were analyzed by statistics(t-test).Results The Tat-Gly(n)-TK (n= 0,2,4,6)recombinant genes were constructed and inserted into PBK vectors,which were expressed in E coli and then purified.Cells in different groups,which were incubated with Tat-Gly (n)-TK(n=0,2,4,6)fusion proteins,Tat-TK fusion protein,TK-Tat fusion proteins or only Tat proteins respectively,were detected by immunofluorescence assay.The intensities of fluorescence in different groups were almost same,but the ratios of cell survival or apoptosis were different.The highest ratio of cells apoptosis(14.77%)was in the group that cellular culture medium was mixed with Tat-Gly(4)-TK fusion protein,followed by the groups containing 6,2 glycines or no TK gene in genes(4.30%,12.69% and 1.03%,respectively).There were significant differences between each 2 groups among the all groups(t-test,P

OBJECTIVETo explore the effect of negative emotions on serum levels of adrenocorticotropic hormone (ACTH) and neuropeptide Y (NYP) in hepatitis B liver cirrhosis (HBLC) patients.

METHODSTotally 617 HBLC patients were assigned to the negative emotion group (415 cases) and the non-negative emotion group (202 cases) judged by negative emotions. Case numbers of various grading Child-Pugh were recorded in the two groups. Their liver functions were compared between the two groups. Serum levels of ACTH and NPY were detected using double antibody sandwich enzyme-linked immunosorbent assay (ELISA) in the two groups.

RESULTSThere was no statistical difference in Child-Pugh grading between the two groups (χ2 = 0.65, P = 0.72). Compared with the non-negative emotional group, serum ACTH levels decreased significantly in the negative emotion group with statistical difference (P < 0.05). There was no statistical difference in serum ACTH levels between the two groups (P > 0.05).

CONCLUSIONThe negative emotion of HBLC patients was not related to the serum ACTH level, but to relatively lower-concentration serum NPY levels.

Adrenocorticotropic Hormone ; blood ; Emotions ; Hepatitis B ; blood ; psychology ; Humans ; Liver Cirrhosis ; blood ; psychology ; Neuropeptide Y ; Serum

This study was designed to determine the impact of chrysoeriol on proliferation and cell cycle progression in the human multiple myeloma cell lines RPMI 8226 and KM3, and its related molecular mechanisms. Chryseoriol was identified by using the phosphorylated AKT-specific cytoblot high throughput assay. CCK-8 assay was employed to examine the growth inhibition rate and IC(50) (48 h) in peripheral blood mononuclear cells (PBMNCs), RPMI 8226 and KM3 cells treated with chrysoeriol at various concentrations. Cells were labeled with 5-6-carboxyfluorescein diacetate succinimidyl ester (CFSE), and the proliferation dynamics was detected by flow cytometry and analyzed with ModFit software. The cell cycles of RPMI 8226 and KM3 cells were measured by flow cytometry when the IC(50) concentration of chrysoeriol was adopted. The alterations in cell-cycle related proteins (Cyclin B1, Cyclin D1, p21) and proteins in PI3K-AKT-mTOR pathway were determined by Western blot analysis. The results showed the proliferation of multiple myeloma cells was significantly inhibited by chrysoeriol, resulting in cell cycle arrest in G(2)/M phase. Chrysoeriol could significantly reduce the expression of p-AKT (s473) and p-4eBP1 (t37/46) protein, meanwhile enhanced Cyclin B1 and p21 protein expression. Similar effects were not observed in PBMNCs from normal donors. It was concluded that chrysoeriol was a selective PI3K-AKT-mTOR pathway inhibitor. It restrained the proliferation of human multiple myeloma cells, but didn't affect proliferation of PBMNCs from normal donors. It might exhibit the cell cycle regulatory effect via the inhibition of PI3K-AKT-mTOR signal pathway.

Objective:To observe the effect of An-pressing manipulation on biceps brachii with delayed onset muscle soreness (DOMS) in healthy male volunteers.Methods:A total of 30 male college student volunteers were randomly divided into a blank group,a model group and a treatment group,10 cases in each group.Subjects in the blank group did not receive any intervention;subjects in the model group received active weight-bearing eccentric exercise on the non-favored side of the upper limb to establish the models,while not receiving any treatment;subjects in the treatment group received both the same modeling and An-pressing manipulation treatment.The subjective rating of perceived exertion (RPE),subjective soreness sensation threshold and soreness grade were evaluated before modeling,immediately after modeling,and 24,48,72,96 and 120 h after modeling.Serum total antioxidant capacity (T-AOC) was measured before modeling,immediately after modeling,and 24,48 and 72 h after modeling.Serum creatine kinase MM isoenzyme (CK-MM) was measured before modeling and 24,48 and 72 h after modeling.Results:At 24,48,72 and 120 h after treatment,the soreness grades of the treatment group were lower than those of the model group (all P＜0.05).The RPE scores of the treatment group were lower than those of the model group (all P＜0.05) immediately after modeling,at 24,48,72,96 and 120 h after modeling.The subjective soreness sensation threshold of the treatment group was higher than that of the model group immediately after modeling,at 24,48,72 and 96 h after modeling (all P＜0.05).Immediately after modeling,T-AOC value in the treatment group was higher than that in the model group and blank group (both P＜0.05).CK-MM of the treatment group was lower than that of the model group at 48 h and 72 h after modeling (P＜0.05).Conclusion:An-pressing manipulation shows a certain therapeutic effect on biceps brachii with DOMS by strengthening the body's antioxidant and anti-damage abilities,which can effectively reduce the pain and accelerate the recovery from fatigue damage.

Objective:To observe the influence of pressing force and time on the thermal effect of An-pressing manipulation.Methods:Eight healthy volunteers were recruited to receive An-pressing manipulation at Xinshu (BL 15) on the right side.The pressing force and time were both divided into five levels:the force described as extremely mild,mild,moderate,strong and extremely strong and time given by 2.5 min,5.0 min,7.5 min,10.0 min and 15.0 min.The real-time change in local acupoint temperature as well as the change during 1.0-15.0 min after the manipulation were observed.Results:Compared with the baseline data,the real-time changes in the temperature after An-pressing Xinshu (BL 15) on the right side with different levels of force (from mild to strong) were respectively (1.88t0.64) ℃,(2.05±0.68) ℃,(2.25±0.59) ℃,(2.35±0.61) ℃ and (2.32±0.69) ℃;the changes in 15.0 min after the manipulation were respectively (-0.11±0.11) ℃,(0.03±0.14) ℃,(0.59±0.58) ℃,(1.38±0.70) ℃ and (2.09±0.98) ℃.The real-time temperature changes after the manipulation for different durations (from short to long) were respectively (1.94±0.37) ℃,(2.33±0.29) ℃,(2.49±0.31) ℃,(2.51±0.39) ℃ and (2.41±0.55) ℃;the changes in 15.0 min after the manipulation were respectively (0.53±0.49) ℃,(0.33±0.30) ℃,(0.52±0.33) ℃,(0.55±0.38) ℃ and (0.76±0.36) ℃.Conclusion:The thermal effect presented an increasing tendency with the extension of pressing time,and the temperature reached the top at 7.5 min;the thermal effect showed an increasing tendency with the rise of pressing force,and the temperature reached the top upon a moderate level of force.The pressing time can produce a greater influence on the real-time temperature than the pressing force;the pressing force can produce a greater influence on maintaining the temperature than the pressing time.

Objective:To explore the optimal thermal effect parameter combination of An-pressing manipulation based on the pressing force, time and frequency, and to compare the thermal effect differences between the rhythmic and the continuous An-pressing manipulations. Methods:Three levels of light, moderate and heavy pressing forces were determined according to the An-pressing forces of the clinical tuina physicians; the pressing time and frequency parameters were determined according to the literatures about An-pressing manipulation. The volunteers were stimulated by the homemade An-pressing manipulation stimulator on the right Xinshu (BL 15), and then the three-factor and three-level orthogonal tests were carried out according to the test sequence specified by the L9(34) orthogonal table, and the temperature before and after pressing was recorded by an infrared thermal imaging system to screen the best parameters for the thermal effect of the An-pressing manipulation, thus to determine the optimal pressing parameters. The optimal parameters were then used for both continuous and rhythmic An-pressing manipulations to stimulate the bilateral Xinshu (BL 15). The temperature changes after pressing and the duration of the thermal effect (temperature difference ≤0.5℃ on both sides) were recorded by the infrared thermal imaging system, to explore the differences in the thermal effects of different An-pressing manipulations. Results:Among the three factors of pressing force, time and frequency, the influences of different pressing forces on temperature were significantly different (F=32.843,P=0.030), and the influence of 2.5 kg pressing force was the most significant; the effects of different pressing time on temperature were significantly different (F=54.102,P=0.018), and the pressing time of 7.5 min was the most significant; the influences of different pressing frequencies on temperature were not statistically significant (F=2.181,P=0.314), though the influence of 10 times/min pressing frequency was the largest. The influences on temperature difference of the rhythmic and the continuous An-pressing manipulations were significantly different (P=0.031 on the left side andP=0.045 on the right side), but there was no statistical difference in the duration of the thermal effect (P=0.690). Conclusion:The An-pressing manipulation parameters that significantly affect the temperature difference are pressing force and time. The optimal combination of thermal effect parameters is pressing force of 2.5 kg, time of 7.5 min, and frequency of 10 times/min. The local thermal effect of the rhythmic An-pressing manipulation is significantly greater than of the continuous An-pressing manipulation.

OBJECTIVETo observe the effectiveness and safety of external applying Compound Tripterygium wilfordii Hook F. (TwHF) in relieving joint pain in rheumatoid arthritis (RA) patients.

METHODSIn this double-blinded, randomized multicenter trial, a total of 174 moderately active RA patients were enrolled and randomly assigned to the treatment group (treated with Compound TwHF, 87 cases) and the placebo control group (87 cases). Compound TwHF or placebo was externally applied in painful joints, 20 g each time, once per day for 8 weeks. Self-reported joint pain relief was taken as a primary effective indicator. Visual analogue scale for pain (VAS), disease activity score of 28 joints (DAS28), VAS for general health (GH) were evaluated before treatment, at week 4 and after treatment. Erythrocyte sedimentation rate (ESR) and hypersensitive C reactive protein (hs-CRP) were tested before and after treatment. Menstrual changes in females were observed during treatment. Skin irritation occurred during the recording process was assessed using skin irritation strength. Intention to treat (ITT) was statistically analyzed.

RESULTSThe joint pain relief rate in the treatment group was 90.8% (79/87 cases), higher than that in the placebo control group (69.0%, 60/87 cases; P = 0.001). VAS pain score, DAS28, VAS for GH score were significantly improved in the two groups at week 4 of treatment and after treatment, as compared with before treatment (P < 0.01). ESR and hs-CRP levels significantly decreased in the treatment group after treatment (P < 0.05, P < 0.01). No difference was found in post-treatment VAS pain score, DAS28, VAS for GH score, ESR, or hs-CRP between the two groups (P > 0.05). Eight adverse events occurred in the treatment group (5 skin allergy, 1 intolerance of medical odor, and 2 mild liver injury), while 3 adverse events occurred in the placebo control group (2 skin allergy, 1 mild liver injury). There was no statistical difference in adverse event between the two groups (P > 0.05). No menstrual change occurred in the treatment group.

CONCLUSIONExternal applying Compound TwHF was an effective and safe way to relieve-joint pain of RA patients, which could be taken as an adjuvant therapy.

Arthralgia ; drug therapy ; Arthritis, Rheumatoid ; drug therapy ; Blood Sedimentation ; C-Reactive Protein ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Female ; Humans ; Phytotherapy ; Treatment Outcome ; Tripterygium

OBJECTIVETo study the effect of angiogenesis inhibitor SU6668 on the growth and metastasis of gastric cancer in SCID mice.

METHODSMetastatic model was established by orthotopic implantation of histologically intact human tumor tissue into the gastric wall of SCID mice. Forty-eight mice were randomly divided into four groups, and saline, 5-FU, SU6668, and 5-FU plus SU6668 were administered by i.p. every day for 6 weeks after tumor implantation. The mice were killed and tumor weight, tumor inhibition rate, intratumoral microvessel density(MVD), apoptotic index(AI) and metastasis inhibition were evaluated.

RESULTSCompared with the control, tumor growth was significantly inhibited in mice treated respectively with 5-FU, SU6668 and 5-FU plus SU6668 with inhibition rates of 47.5%, 64.1% and 69.2% respectively. Decreased MVD and increased AI were noted in the mice treated with SU6668 and 5-FU plus SU6668. The incidences of liver and peritoneal metastases was significantly inhibited and decreased to 62.5%, 69.9% in SU6668 group, and 74.9%, 90% in 5-FU plus SU6668 group. The growth and metastasis of human gastric cancer implanted in SCID mice were significantly inhibited in SU6668 group and combined group, especially in combined group.

CONCLUSIONAngiogenesis inhibitor SU6668 has a strong inhibitory effect on tumor growth and metastasis of human gastric cancer transplanted in SCID mice, and has synergistic effect combined with cytotoxic agents.

Angiogenesis Inhibitors ; pharmacology ; therapeutic use ; Animals ; Apoptosis ; drug effects ; Cell Line, Tumor ; Drug Synergism ; Fluorouracil ; pharmacology ; therapeutic use ; Humans ; Indoles ; pharmacology ; therapeutic use ; Liver Neoplasms ; prevention & control ; secondary ; Male ; Mice ; Mice, SCID ; Neoplasm Transplantation ; Neoplasms, Experimental ; Neovascularization, Pathologic ; drug therapy ; Pyrroles ; pharmacology ; therapeutic use ; Stomach Neoplasms ; drug therapy ; pathology

Objective To study the influence of fluoride on the expression of osteoprotegerin(OPG) mRNA and protein in suckling rat osteoblasts. Methods Osteoblasts obtained from calvarial of suckling Wistar rats were cultured in vitro in the media supplemented with NaF at a series of doses[O(control), 1,2 and 4 mg/L groups], and OPG mRNA expression and protein were evaluated by RT-PCR and ELISA methods, respectively. Results OPG mRNA expression in suckling rat osteoblasts cultured in vitro significantly increased after exposure to NaF for 48 h and 72 h(F=333.48,808.34,P<0.05). OPG mRNA expression in suckling rat osteoblasts cultured in vitro after exposure to NaF for 48 h at different doses(0.810±0.003, 0.819±0.031 and 0.870±0.044 for 1,2 and 4 mg/L groups, respectively) compared with that of control (0.800±0.040, all P<0.05). OPG mRNA expression further increased for 72 h exposure to NaF(0.933±0.047,1.031±0.051,1.240±0.062 for 1,2 and 4 mg/L, respectively), significantly higher than that of the control (0.805±0.020,all P<0.05) and corresponding groups at 48 h. NaF doses and time exposure exhibited a significant synergistic effect on OPG mRNA expression(F=2004.16, P<0.05). NaF also enhanced OPG protein expression in suckling rat osteoblasts cultured in vitro. Significant differences were observed only in 4 mg/L group(0.228±0.014,0.277±0.048) and control(0.205±0.012,0.229±0.010) at 48 h and 72 h (P<0.05). In addition, OPG protein expression at 72 h post-exposure was higher than that at 48 h,but there was no synergistic effect between concentration and time(F=1.21,P>0.05). Conclusions The results suggested that NaF could increase OPG mRNA and protein expression in suckling rat osteoblasts with a synergistic effect between the doses and exposure time.

The infection status of anisakid larvae was examined in 290 marine fish of 25 species and in 108 cephalopods of 3 species purchased in Bayuquan region, Yingko city nearby the coast of the Bohai Sea from may to August 1992. A total of 7,327 larvae were collected from 156 fish of 19 species and 8 squids of one species. The 3rd-stage larvae of Anisakis simplex were collected from 121 fish (63.4%) of 15 species (N = 191) and from 8 squids (14.8%) of one species (N = 54), and they were total, 5,992 (81.8%). Out of remaining 1,335 larvae, 154 (2.1%) were classified as Thynnascaris type B from 23 fish of 4 species, 1,013 (13.8%) as Thynnascaris type C from 79 fish of 13 species. 164 (2.2%) as Hysterothylacium China type V from 20 fish of 4 species, 3 (0.04%) as Raphidascaris from 3 fish of 2 species and one was Pseudoterranova decipiens larva.
Animal ; Anisakiasis/veterinary* ; Anisakiasis/parasitology ; Anisakiasis/epidemiology ; Anisakis/isolation & purification ; Anisakis/classification* ; China ; Fish Diseases/parasitology* ; Fish Diseases/epidemiology ; Fishes ; Larva ; Seawater ; Squid/parasitology*