2.The evaluation of efflux transporter model based on RNA interference technology in vitro.
Ling-lei KONG ; Hai-ying YANG ; Mei YUAN ; Xiao-mei ZHUANG ; Hua LI
Acta Pharmaceutica Sinica 2015;50(9):1122-1127
In the present study, the specifically knockdown models of P-gp or MRP2 were constructed by using a series of chemically synthesized small interfering RNA (siRNA) in vitro. The expression of P-gp and MRP2 was measured by real-time PCR and Western blot, and the function was evaluated by applying P-gp and MRP2 substrate, rhodamine and methotrexate. The results showed that MRP2 siRNA-3 or P-gp siRNA-2 significantly decreased the mRNA expression of MRP2 or P-gp, the inhibition ratio was 68% or 84%; MRP2 siRNA-3 or P-gp siRNA-2 at a dose of 80 nmol x L(-1) significantly reduced the protein expression of MRP2 or P-gp at 48 h after treatment, the inhibition ratio was 62% or 70%. Meanwhile, other transporters were not influenced by siRNA. When pretreatment with MRP2 siRNA-3 or P-gp siRNA-2, the efflux of methotrexate or rhodamine decreased significantly and the intra-cellular concentration increased. The results suggested that chemically synthesized siRNA could significantly inhibit the expression and function of MRP2 and P-gp, and the model of RNAi in vitro could be used to evaluate the role of efflux transporters in transportation of drugs.
ATP-Binding Cassette, Sub-Family B, Member 1
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genetics
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Gene Knockdown Techniques
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Multidrug Resistance-Associated Proteins
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genetics
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RNA Interference
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RNA, Small Interfering
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Real-Time Polymerase Chain Reaction
3.CD24 attenuates carbon tetrachloride-induced murine liver fibrosis
Jie XIANG ; Xiongfei LI ; Lei HAI ; Yatong FAN ; Jian ZHENG ; Hua ZHANG ; Xuejun ZHANG
Chinese Journal of Microbiology and Immunology 2017;37(3):171-177
Objective To investigate the effects of CD24 on CCl4-induced murine liver fibrosis and to analyze the possible molecular mechanism.Methods Wild type (WT) and CD24 knockout (CD24-/-) C57BL/6 mice were treated with CCl4 through intraperitoneal injection.Levels of ALT in serum samples were detected and liver tissues were stained with hematoxylin and eosin (HE) to assess liver tissue injury.Sirius Red staining was used to observe liver fibrosis.Real-time PCR was performed to detect the expression of α-SMA (α-smooth muscle actin), Col1a1 (Collagen, typeⅠ, alpha 1), TGF-β1 (transforming growth factor-β1) and CD24 at mRNA level in liver tissues.Western blot was performed to analyze the expression of α-SMA and Col1a1 at protein level.Flow cytometry analysis was used to detect the macrophages in liver tissues.ELISA was used to detect the expression of TGF-β1 in the culture supernatants of M1 and M2 macrophages.Results The expression of CD24 at both mRNA and protein levels were up-regulated in mice with CCl4-induced liver fibrosis.HE staining showed that liver inflammation in CD24-/-mice was more severe than that in WT mice after treated with CCl4.Sirius Red staining of paraffin-embadded liver sections revealed that compared with WT mice, CD24-/-mice presented with more severe liver fibrosis.Moreover, α-SMA and Col1a1, indicators of liver fibrosis, in CD24-/-mice were significantly higher than those in WT mice.Flow cytometry analysis showed that murine hepatic macrophages significantly increased in CD24-/-mice than in WT mice following CCl4 treatment.Real-time PCR analysis also confirmed that significantly enhanced expression of TGF-β1 at mRNA level in liver tissues was observed in CD24-/-mice than in WT mice.TGF-β1 secreted in the culture supernatant of M2 macrophages derived from CD24-/-mice group was more than that of the WT mice group.No significant difference in TGF-β1 secretion in culture supernatant of M1 macrophages was observed between the two groups.Conclusion Taken together, these data suggest that CD24 plays an important role in attenuating CCl4-induced chronic inflammation and hepatic fibrosis in mice.The mechanism of CD24 in alleviating liver fibrosis might be through regulating intrahepatic macrophages, inhibiting the secretion of TGF-β1 by M2 macrophages and suppressing the activation of hepatic stellate cells.
4.Ifosfamide and vinorelbine combined chemotherapy in the treatment of advanced non-small cell lung cancer
Yi LAO ; Shao-Feng CHEN ; Gui-Hua LEI ; De-Ming XU ; Wei WANG ; Hai-Ming ZHONG ;
Chinese Journal of Primary Medicine and Pharmacy 2006;0(08):-
Objective To evaluate therapeutic effects and toxicity of advanced non-small cell lung cancer (NSCLC)treated by combining chemotherapy on ifosfamide(IFO)and vinorelbine(NVB).Methods 107 cases pa- tients with advanced NSCLC were enrolled.IFO was given in a dosage of 1.5g/m~2 on day 1 to 4.and NVB in a dosage of 25mg/m~2 on day 1 and 8.It was repeated every three or four weeks,up to two to four cycles.Results Two patients had complete response and 40 patients had partial response.The overall response rate was 47.7% ,the median survival time 10.3 months,1-year and 2-year survival rate was 42% and 12.3%,respectively.The main toxicity was bone marrow suppression.Conclusion The regimen is effective,sale and tolerable in advanced non- small cell lung cancer therapy.
7.Effects of 5-hydroxytryptamine ascending pathways of dorsal raphe nuclei and habenular nucleus on the respiration and blood pressure of rats.
Lei YU ; Min HUANG ; Hai-yan XU ; Hua ZHAO
Chinese Medical Journal 2007;120(24):2308-2310
Animals
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Blood Pressure
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drug effects
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Electric Stimulation
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Female
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Habenula
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physiology
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Lidocaine
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pharmacology
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Male
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Raphe Nuclei
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physiology
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Rats
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Rats, Wistar
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Respiration
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drug effects
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Serotonin
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physiology
8.Effects of surface roughness of bone cements on histological characteristics of induced membranes.
Hai-Xiao LIU ; Hua-Zi XU ; Yu ZHANG ; Gang HU ; Yue SHEN ; Xiao-Jie CHENG ; Lei PENG
China Journal of Orthopaedics and Traumatology 2012;25(8):662-666
OBJECTIVETo explore surface roughness of bone cement and surround tissue on histological characteristic of induced membranes.
METHODSBone cements with smooth and rough surface were implanted in radius bone defect, intramuscular and subcutaneous sites of rabbits, and formed induced membranes. Membranes were obtained and stained (HE) 6 weeks later. Images of membrane tissue were obtained and analyzed with an automated image analysis system. Five histological parameters of membranes were measured with thickness,area,cell density,ECM density and microvessel density. Double factor variance analysis was used to evaluate the effect of the two factors on histological characteristics of induced membranes.
RESULTSMembranes can be induced by each kind of bone cement and at all the three tissue sites. In histological parameters of thickness,area and micro vessel,there were significant differences among the membranes induced at different tissue sites (P = 0.000, P = 0.000, P = 0.000); whereas, there were no significant differences in histological parameters of cell density and ECM density (P = 0.734, P = 0.638). In all five histological parameters of membranes, there were no significant differences between the membranes induced by bone cements with different surface roughness (P = 0.506, P = 0.185, P = 0.883, P = 0.093, P = 0.918).
CONCLUSIONSurround tissue rather than surface roughness of bone cements can affect the histological characteristics of induced membranes. The fibrocystic number, vascularity, mechanical tension and micro motion of the surround tissue may be closely correlated with the histological characteristics of induced membranes.
Animals ; Bone Cements ; Female ; Membranes ; cytology ; Rabbits ; Radius ; cytology ; Surface Properties ; Tissue Engineering ; methods ; Tissue Scaffolds
9.Study on conditions of seed germination of Cistanche.
Xue-Yi QIAO ; Hua-Lei WANG ; Yu-Hai GUO
China Journal of Chinese Materia Medica 2007;32(18):1848-1850
OBJECTIVETo study the effect of fluridone concentration, stimulating period, temperature and salt on the seed germination of three species of Cistanche.
METHODThe seeds were cultured in Petri dish, and the germination percentage was counted.
RESULTThe highest germination percentage was observed in Cistanche tubulosa, C. deserticola, C. sala seeds pre-treated by 0.1 mg x L(-1) fluridone for 24-29 h. The optimal temperature for the seeds germination of three species of Cistanche was at 20-30 degrees C, and the seeds did not germinate at sub-or supraoptimal temperatures (5 and 35 degrees C). The salt tolerance of C. sala seeds was strong, and the critical value of NaCl concentration was 0.04 mol x L(-1). By contrast, C. tubulosa and C. deserticola seeds were more sensitive to the salt stress, the critical value of NaCl concentration was 0.02 mol x L(-1).
CONCLUSIONThe optimal germination condition and the method of testing germination percentage of three species of Cistanche seeds are as follow: the seeds are pre-treated by 0.1 mg x L(-1) fluridone for 24 h and then cultured at 20-30 degrees C in salt solution which concentration is lower than 0.02 mol x L(-1).
Cistanche ; classification ; growth & development ; Germination ; drug effects ; physiology ; Plants, Medicinal ; growth & development ; Pyridones ; pharmacology ; Seeds ; growth & development ; Sodium Chloride ; pharmacology ; Species Specificity ; Temperature
10.99Tcm-MIBI SPECT in estimation of the treatment response to three-dimensional conformal radiotherapy in patients with non-small cell lung cancer
Li-xin, YAO ; Zhan-zhao, FU ; Tao, GU ; Lei-ming, GUO ; Hai-xia, HUA ; Qing-huai, ZHANG
Chinese Journal of Nuclear Medicine 2010;30(3):155-157
Objective To explore whether the degree of 99Tcm-methoxyisobutylisonitrile (MIBI) uptake in non-small cell lung cancer (NSCLC) could be correlated with the treatment response to three-dimensional conformal radiotherapy.Methods A total of 102 patients with NSCLC were studied with 99Tcm-MIBI SPECT before radiotherapy.The patients were classified by a follow-up CT as responders (complete or partial remission) and non-responders (stable or progressive disease).After intravenous administration of 740 MBq 99Tcm-MIBI, SPECT imaging at 10-30 min (early) and 2-3 h (delayed) were performed.Region of interest (ROI) was placed over the tumors and contralateral normal lung tissue.The uptake ratio of tumor to contralateral normal lung (T/N) was obtained from both early (ER) and delayed (DR) SPECT images.The retention index (RI) was measured as:RI = (DR - ER)/ER×100%.Statistical analysis was performed by two independent-sample t-test and Mann-Whitney U test using software SPSS 13.0.Results 99Tcm-MIBI uptake was significantly higher in responders than in non-responders:2.36 ±0.17 vs 1.82 ±0.14 (ER) and 2.48 ± 0.20 vs 1.94 ± 0.16 (DR), respectively (t = - 13.1,- 12.7, both P< 0.05).The median RI in the responders group was also significantly higher than that in the non-responders group (6.60% vs 5.13%, z = - 6.83, P < 0.05).Conclusion ER, DR and RI of 99Tcm-MIBI SPECT might be useful to estimate the treatment response to three-dimensional conformal radiotherapy in patients with NSCLC.