Adult ; Drugs, Chinese Herbal ; administration & dosage ; therapeutic use ; Esophageal and Gastric Varices ; drug therapy ; etiology ; Female ; Gastrointestinal Hemorrhage ; drug therapy ; etiology ; Humans ; Infusions, Intravenous ; Liver Cirrhosis ; complications ; drug therapy ; Male ; Middle Aged ; Phytotherapy

Objective To observe the expression of motilin precursor mRNA and location of motilin in human thyroid and to explore the clinical significance.Methods RT-PCR,Sourthern blot and immuno- fluorescence histochemical techniques were applied to detect the expressions of motilin precursor mRNA and motilin in human thyroid and to compare the difference of motilin peptide and motilin mRNA expression among tissues of normal thyroid,nodular goiter and medullary thyroid carcinoama.Results(1)The results of RT-PCR and Southern blot showed that motilin precursor mRNA was expressed in human thyroid.(2)A large number of cells showing collocation of motilin with calcitonin were observed in human thyroid,suggesting that motilin was expressed in C cells.(3)Both the motilin immunoreactive(MTL-IR)and the motilin mRNA were significantly increased in medullary thyroid carcinoma(P0.05).(4)Western blot results showed that the expression of motilin in thyroid medullary carcinoma was significantly higher than that in normal thyroid. However no difference was observed between thyroid nodular goiter and normal thyroid tissue.Conclusion MTL- IR positive cells and motilin mRNA expression are found in human thyroid.Motilin is located in C cells of thyroid. The expressions of motilin and motilin precursor mRNA are significantly increased in medullary thyroid cancer,no difference between thyroid nodular goiter and normal thyroid tissue.

AIM:To observe the effect of docosahexaenoic acid ( DHA) on H2 O2-induced apoptosis in human retinal pigment epithelium cells and its molecular mechanism .METHODS: Human retinal pigment epithelium cell line ARPE-19 was cultured in vitro, and 12.5 mmol/L H2 O2 was used to mimic the oxidative stress condition .The cells were treated with 30~100μmol/L DHA for 4~24 h.The expression of heme oxygenase-1 (HO-1) at mRNA and protein levels was detected by real-time PCR and Western blot , respectively .The enzymic activity of HO-1 was measured by colorimetry . Production of reactive oxygen species ( ROS) was determined by fluorescent probe .Activation of NF-E2-related factor 2 (Nrf2) was examined by immunofluorescence method .Apoptosis of ARPE-19 cells was analyzed by flow cytometry .RE-SULTS:The mRNA and protein expression and the enzymic activity of HO-1 were significantly increased in the ARPE-19 cells after DHA treatment .Meanwhile , nuclear translocation of Nrf 2 was also observed .Apoptosis appeared and ROS was produced upon H2O2 incubation.In contrast, DHA at 100 μmol/L significantly abrogated H2O2-induced apoptosis and ROS production.Furthermore, silencing of HO-1 by specific siRNA, or treatment with ZnPP, an inhibitor of HO-1, partly counteracted the protective effect against H 2 O2-induced apoptosis and ROS production .CONCLUSION: DHA protects retinal pigment epithelial cells against oxidative stress via induction of heme oxygenase -1 expression after Nrf2 activation .

Background and purpose:A large number of studies have showed that retinoblastoma gene 1 (RB1) can inhibit the occurrence and development of many tumors, including neuroblastoma, small cell lung cancer, osteosar-coma, pancreatic cancer, breast cancer and so on. RB1 is also closely related to the regulation of cell cycle, differentia-tion, senescence, apoptosis, growth inhibition, etc. The goal of this article is to elucidate whether miR-222 promotes cell proliferation and invasion by targeting RB1, further to explore the molecular mechanism that miR-222 functions as an oncogene in retinoblastoma cells.Methods:miR-222 (miR-222 mimics) and RB1-wt, miR-NC and RB1-wt, miR-222 and RB1-mut, miR-NC (a controlled miR-222 mimics) and RB1-mut were co-transfected into Y79 cells, and luciferase activity was detected by single photon. Retinoblastoma cells were transfected with miR-222 mimics and miR-NC, and the expressions of RB1 protein were detected by Western blot. Retinoblastoma cell proliferation assays were performed by MTS assay when miR-222, miR-NC, RB1 (pcDNA3.1-RB1), vector (pcDNA3.1), miR-222+RB1 and miR-NC+vec-tor were transfected into Y79 cells. The growth and invasion ability of Y79 cells with ectopic expression of miR-222 were evaluated by MTS and Transwell invasion assays.Results:This study demonstrated that miR-222 could promote the luciferase activity of RB1-wt. The expression levels of luciferase reporter gene activity in Y79 cells after transfection with miR-222+RB1-wt were higher than those in the negative control cells (miR-NC+RB1-wt) (P<0.05). The protein expression levels of RB1 in Y79 cells after transfection with miR-222 were lower than those in miR-NC (P<0.05). Overexpression of RB1 inhibited the proliferation of retinoblastoma cells. miR-222 promoted the prolifera-tion of retinoblastoma cells through targeting RB1 (P<0.05). Moreover, there was no signiifcant difference between the cell survival rates of Y79 which were transfected with miR-222+pcDNA3.1-RB1 and miR-NC+pcDNA3.1 (P>0.05). After transfection with miR-222 mimics for 48 h, Transwell invasion assay showed that the number of cells through the basement membrane was (193±10). Compared with the control group (144±11), it could signiifcantly accelerate the invasion of Y79 cells (P<0.01). There was no signiifcant difference between the number of cells through the basement membrane which were transfected with miR-222+pcDNA3.1-RB1 and miR-NC+pcDNA3.1 (P>0.05).Conclusion:miR-222 promotes cell proliferation and invasion by targeting RB1 expression in retinoblastoma cells.

Background Oxydative stress is an important pathogenesis of age-related macular degeneration.Resent evidences indicate that docosahexaenoic acid (DHA) plays an important role during the development of retinal photoreceptor cells and protect the cells against oxydative stress by inducing the expression of heme oxygenase-1 (HO-1).However,whether DHA can induce the expression of HO-1 in human retinal pigment epithelium (RPE) cells is unelucidated.Objective This study was to investigate the effect of DHA on the expression of HO-1 in RPE cells and its molecular mechanism.Methods Human RPE cell line ARPE-19 was cultured in vitro and treated with 30,50,100 and 120 μmol/L DHA for 4 to 24 hours,respectively,and the cells were cultured without DHA as the control group.The cytotoxicity of DHA was detected by lactate dehydrogenase(LDH),and the expression of HO-1 mRNA and protein were detected by real-time PCR and Western blot assay,respectively.The enzymatic activity of HO-1 was detected by colorimetry.The reactive oxygen species (ROS) proportion in the cells was detected using fluorescence probe H2 DCFDA,and immunofluorescence technology was adopted to detect the nuclear translocation of nuclear facotor-E2-related factor 2 (Nrf2).The expression of Nrt2 protein in the cells was detected by Western blot after intervention of ROS inhibitor N-acetylcysteine (NAC) and transfection of Nrf2 small interfering RNA (siRNA).Results The LDH leakage rate was significantly different after 0,3,50,100 and 120 μmol/L DHA treated the cells for 24 hours (F=8.14,P＜0.05),and the LDH leakage rate in the 120 μmol/L DHA group was significantly higher than that of 0,30,50 and 100 μmol/L DHA group (all at P＜0.05).The relative expression levels of HO-1 mRNA and HO-1 protein or HO-1 enzymatic activity in the cells were significantly different among different concentrations of DHA group in 8 hours after treatment (F=16.24,P＜0.05;F=11.34,P＜0.05;F=11.81,P＜0.05),and the expressions of these factors were considerably higher in the 30,50 and 100 μ mol/L DHA group than those in the 0 μmol/L DHA group (all at P＜0.05).The ROS relative fluorescence intensity and nuclear Nrf2 positive cells proportion were statistically significant among different concentrations of DHA groups (F =11.08,P ＜ 0.05;F=16.42,P＜0.05),and the ROS relative fluorescence intensity and nuclear Nrf2 positive cells proportion were evidently higher in the 30,50 and 100 μmol/L DHA group than those in the 0 μmol/L DHA group (all at P＜0.05).The relative expression levels of HO-1 protein and the proportion of nuclear Nrf2 positive cells were significantly lower in the NAC pretreated 100 μmol/L DHA group than those in the 100 μmol/L DHA group.In addition,the HO-1 relative expression level and the positive cells proportion of nuclear Nrf2 were significantly lower in the of Nrf2 siRNA transfection group than those in the blank siRNA transfection group (both at P＜0.05).Conclusions DHA with concentration below 100 μ mol/L can protect RPE cells from oxidative stress by inducting the expression of HO-1 in the cells via ROS/Nrf2 pathway.

To study the impact of five different origin processing methods, namely natural drying, drying in baking shop, drying by microwave heating, drying in drum and drying with sulphur fumigation, on the quality of Lonicerae Japonicae Flos from Donghai cultivation base in Jiangsu Province, with the contents of chlorogenic acid and galuteolin and the similarity in HPLC fingerprints as the evaluation indicators. The results showed that different origin processing methods had significant impact on the content of chlorogenic acid and the similarity in HPLC fingerprints, but with no significant difference on the content of galuteolin. By means of drying by microwave heating and drying in drum, the samples showed higher contents of chlorogenic acid, respectively 3.67% and 3.39%. The similarities of HPLC fingerprints were 0.815 and 0.793, respectively. By means of the drying in baking shop and the drying with sulphur fumigation, the contents of chlorogenic acid in the samples were 2. 87% and 2. 53% , respectively. The similarities of HPLC fingerprints were 0.964 and 0.765, respectively. The lowest content of chlorogenic acid in naturally dried samples was 1.92%. The similarity of HPLC fingerprints was 0.940. According to the findings as well as the internal control standards for Lonicerae Japonicae Flos herbs of Jiangsu Kanion Pharmaceutical Co. , Ltd. , the optimum processing method for Lonicerae Japonicae Flos from Donghai cultivation base was the drying in baking shop. This study provided a theoretical basis for determining the processing method for Lonicerae Japonicae Flos from Donghai cultivation base of Jiangsu Province.
China ; Desiccation ; methods ; Drugs, Chinese Herbal ; chemistry ; Lonicera ; chemistry ; growth & development ; Quality Control

Objective To observe the radiosensitization effect of Aidi injection on human lung adenocarcinoma A549 cells, and to analyze its possible mechanism.Methods ① A549 cells were treated with different concentrations of Aidi injection (1.875, 3.75, 7.5, 15, 30, 60 mg/mL) for 24 hours, and in the mean time, a blank control group was set up; the effect of Aidi injection on lung adenocarcinoma A549 cells proliferation was detected by methyl thiazolyl tetrazolium (MTT) assay, and the 10% cell growth inhibitor concentration (IC10) was calculated. ② The experiments were divided into blank control, Aidi control, radiation and Aidi pretreatment groups. The Aidi control group was incubated for 24 hours by Aidi injection IC10; the radiotherapy group was given X-ray irradiation of 4 Gy followed by incubation for 24 hours; the Aidi pretreatment group was incubated for 24 hours by Aidi injection IC10 and then given X-ray irradiation of 4 Gy; the blank control group received equal volume of normal saline and was incubated for 24 hours. The survival fraction (SF) value was detected by cell colony formation assay; the protein levels of the serine phosphorylation at 139 locus of histone (γ-H2AX protein), the key protein in homologous recombination repair pathway (Rad51 protein) and the cell autophage characteristic protein (LC3 protein) were detected by Western Blot; the formation of autophagosome was observed by transmission electron microscope.Results Aidi injection possessed the suppression of the growth of human lung adenocarcinoma A549 cells, the proliferation of the cells in various Aidi groups was lower than that in the blank control group, with the increase in drug concentration, the A549 cell growth inhibition ratio (IR) was gradually increased, representing a dose dependent manner, and the IC10 was 3.09 mg/mL. Compared with the blank control group, the SF value in Aidi control group was not significantly different [(94.7±3.85)% vs. (100.0±0.00)%,P > 0.05], the SF value in radiation group was decreased [(71.8±5.9)% vs. (100.0±0.0)%,P < 0.05], and in Aidi pretreatment group, the value was further decreased compared with that in radiation group [(51.9±4.7)% vs. (71.8±5.9)%,P < 0.05]. Compared with the blank control group, the expression of γ-H2AX protein in the three treatment groups was significantly increased, the degree of increase in Aidi pretreatment group was the most obvious, and it was significantly higher than that in radiation group (gray value: 1.44±0.11 vs. 0.93±0.09,P < 0.05). But the expression of Rad51 protein was the highest in radiation group, and it was higher than that in Aidi pretreatment group (gray value: 1.37±0.07 vs. 0.78±0.04, P < 0.05). Compared with the blank control group, the LC3Ⅱ/LC3Ⅰ value in Aidi control group, radiation group and Aidi pretreatment group were increased, and the degree of increase in Aidi pretreatment group was the most significant (0.35±0.06, 0.37±0.07, 0.49±0.06 vs. 0.05±0.04, allP < 0.05). Under transmission electron microscope, compared with the blank control group, the autophagosome in all treatment groups was increased to some extent, and the degree of increase in Aidi pretreatment group was the most remarkable.Conclusion Aidi injection has the enhancing effect of radiosensitization on human lung adenocarcinoma A549 cells, and its mechanism is possibly related to the up-regulation of A549 cell autophagy level.

OBJECTIVETo explore the expression of phospholipase C-gamma 1 (PLC-gamma1) alternative splicing variants in rats.

METHODSAccording to the sequence of human PLCG1 splicing variant, specific primers for rat PLC-gamma1 were designed and synthesized. The rat RNA was reverse transcribed into cDNA, which was amplified using the specific primers, and the PCR products were sequenced and analyzed using BLAST and bioinformatics methods. Totally 21 rat tissue samples were examined, including the heart, liver, lung, kidney, eyeball, and brain obtained in 3 different embryonic stages, 7 different early postnatal stages, and in adulthood.

RESULTSThe result did not show that rat PLC-gamma1 had the same splicing variant (PLC-gamma1a, NM_002660) as human does.

CONCLUSIONSThe same splicing variant of PLC-gamma1 detectable in human may not exist in rats, and the pre-mRNA may undergo splicing resulting predominantly in PLC-gamma1b mRNA. Very likely, the alternative splicing site of rat PLC-gamma1 is not identical to that of human.

Alternative Splicing ; Animals ; Base Sequence ; Molecular Sequence Data ; Phospholipase C gamma ; genetics ; RNA Precursors ; genetics ; RNA Splice Sites ; genetics ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Analysis, DNA

Objective To evaluate the outcome of combination of intensive preconditioning regimen allo-HSCT with imatinib for treatment of Ph chromosome positive acute lymphocyte leukemia (ALL). Methods Between 2009 and 2010, 8 patients diagnosed as Ph+ ALL received allo-HSCT from HLA identical sibling during complete remission. Imatinib was added into the therapies of 5 patients.Seven patients received the intensive preconditioning regimen based on BuCy2, one patient received the regimen of TBI-Cy. A median of 6. 02 × 108/kg mononuclear cells and 3. 14 × 106/kg CD34+ cells were transfused. GVHD prophylaxis included cyclosporine A and methotrexate. Results All patients were well tolerant to the regimen without serious regimen-related toxicity. The median time of ANC≥0. 5 × 109/L was 15. 5 days, and that of PLT≥20 × 109/L was 19 days. Thirty days after allo-HSCT, all patients got donor engraftment successfully. Among 8 cases, 4 cases presented acute GVHD, 2 developed degree Ⅰ , one developed degree Ⅱ , and one developed degree Ⅳ. Seven patients were alive 100 days after allo-HSCT, 3 of whom presented chronic GVHD. At the end of following-up period, 6 patients were alive, among them, 3 patients were alive without relapse; 3 patients relapsed; Two patients died, one from acute GVHD, and one from leukemia relapse. Conclusion Combined intensive preconditioning regimen allo-HSCT with Imatinib was an effective treatment for Ph+ ALL, but the effect of anti-chronic GVHD of imatinib should arouse certain attention.

Objective The levels of Thl cytokines(IL-10 and IL-13)and Th2 cytokines(INF-? and TNF-?)were determined in the sera of patients with Schistosomiasis japonica in order to find the relationship between cytokines and severe hepatic fibrosis(HF)in schistosomiasis.Methods A total of 358 patients with advanced Schistosomiasis japonica were examined by ultrasound.68 HBsAg negative patients were chosen randomly as experimental control.Among them,39 patients were found to have mild HF and 29 were severe HF.The sera levels of Thl and Th2 cytokines were determined with ELISA.Results Among these 358 patients,83(23.2%)were HBsAg positive.Neither earlier nor severer hepatic fibrosis was noted in the patients who had been simultaneously infected with HBV than those only infected with schistosomiasis. There was a significant difference between mild[ 1.60(1.30-12.14)ng/L]and severe[ 4.20(1.43- 52.07)ng/L]HF patients in the level of IL-10(Z=-3.907,P0.05)was found in level of IFN-?,between severe[3.12(1.38-66.14)ng/L]and mild[5.87(1.33-216.33)ng/ L]HF subjects.Our observation did not reveal any obvious difference of TNF-? between severe[ 2.48(0.79 -19.86)ng/L]and mild[ 2.28(0.67-15.72)ng/L]HF groups.Conclusions Patients infected with advanced shistosomiasis may become more susceptible to HBV.The results of the present investigation showed that a high level production of IL-13 was associated with severe HF.