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 ; genetics ; Gene Knockdown Techniques ; Multidrug Resistance-Associated Proteins ; genetics ; RNA Interference ; RNA, Small Interfering ; Real-Time Polymerase Chain Reaction

OBJECTIVETo observe the auxiliary efficacy and safety of Hebi Recipe (HR)in treating early rheumatoid arthritis (RA).

METHODSTotally 63 early RA patients with Gan-Pi disharmony were randomly assigned to the treatment group [32 cases, treated by HR (one dose per day, taken in two portions for 24 successive weeks) plus Methotrexate (MTX)] and the control group (31 cases, treated by MTX alone). The dosage of MTX was increased from 7.5 mg to 12.5 mg, once per week, 24 weeks as one course of treatment. Efficacy for Chinese medical syndromes, American College of Rheumatology 20 (ACR20) improvement rate, disease activity score in 28 joints (DAS28), laboratory related indices [ESR, rheumatoid factor (RF), C-reactive protein (CRP), anti-cyclic citrullinated peptide (CCP)], and related ultrasonic inspection items (synovium thickness, synovium blood flow classification, effusion of joint), and adverse reactions were observed.

RESULTSThe total effective rate (83.9%, 26/31 cases) and ACR20 improvement rate (74.2%, 23/31 cases) were higher in the treatment group than in the control group [60.7% (17/28 cases), 46.4% (13/28 cases); P < 0.05]. Compared with before treatment in the same group, DAS28 score, ESR, RF, CRP, CCP, synovium thickness, synovium blood flow classification, effusion of joint all decreased in the two groups after treatment (P < 0.01, P < 0.05). Compared with the control group after treatment, ACR20 improvement rate, DAS28 score, ESR, RF, CRP, CCP, synovium thickness, synovium blood flow classification, effusion of joint all decreased in the treatment group (P < 0.01, P < 0.05). Liver dysfunction occurred in 1 case of the treatment group. One leucopenia and 2 liver dysfunction occurred in the control group.

CONCLUSIONHR could effectively improve joints and systemic symptoms of early RA patients with Gan-Pi disharmony.

Arthritis, Rheumatoid ; drug therapy ; C-Reactive Protein ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; therapeutic use ; Humans ; Methotrexate ; Phytotherapy ; Rheumatoid Factor ; Syndrome ; Treatment Outcome

OBJECTIVETo observe therapeutic effects of acupuncture at different acupoints on depression.

METHODSSeventy adult male rats with similar Open-Field score were randomly divided into a normal control group, a model group, a blank control group, a drug treatment group, an acupuncture group I ["Baihui" (GV 20), "Shenting" (GV 24)], II ]"Neiguan" (PC 6), "Sanyinjiao" (SP 6)], and III ("Baihui", "Shenting", "Neiguan" and 'Sanyinjiao"), 10 rats in each group. Separation feeding, long term unpredictability and medium stimulation stress techniques were applied to develop depression model rats. Changes of behaviors were investigated with Open-Field method and sucrose consumption trial.

RESULTSCompared with the normal control group, the score for the behavior and the consumption of sucrose reduced in the depression model group; compared with the blank control group, the score for the behavior and the consumption of sucrose increased in all the treatment groups during treatment. There was no significant difference among the acupuncture groups in the increase of behavior scores 21 days after treatment (P > 0.05), and there were significant differences in the increase of consumption of sucrose in the acupuncture group I and the increase of score for vertical movement in the acupuncture group III 14 days after treatment (P < 0.05, P < 0.01).

CONCLUSIONAcupuncture can change the behavior abnormality induced by separation feeding, long term unpredictability and medium degree stimulation depression model rats and different acupoint selection is a factor in acuounture treatment.

Acupuncture Points ; Acupuncture Therapy ; Animals ; Depression ; therapy ; Depressive Disorder ; Humans ; Problem Behavior ; Rats

OBJECTIVETo explore the effect of Bushen Tiaojing Recipe (BTR) on the quality of oocytes, reproductive hormones, and the expression of bone morphogenetic protein-15 (BMP15) of in vitro fertilization-embryo transfer (IVF-ET) patients.

METHODSSixty infertility patients who prepared for IVF-ET were assigned to two groups according to the treatment order, the treatment group [20 cases, treated with BTR + controlled ovarian hyperstimulation (COH)] and the control group (treated with COH alone, 40 cases). Age, the time limit for infertility, basal follicle-stimulating hormone (bFSH) concentration, usage days and the dosage of gonadotropins (Gn), serum levels of estradiol (E2), luteotropic hormone (LH), and progesterone (P) on the HCG injection day, the number of retrieved occytes, the fertilization rate, the number of embryos, the high quality embryo rate, and the clinical pregnancy rate were compared. Concentrations of follicle-stimulating hormone (FSH), LH, E2, testosterone (T), and P in the follicular fluid were detected via chemiluminescence microparticle immunoassay. The mRNA and protein expression of BMP-15 in mature granulosa cells was detected by real-time fluorescent PCR and Western blot.

RESULTSThirty-two patients were pregnant and the total pregnancy rate was 53.3%. Of them, 19 were pregnant and the total pregnancy rate was 47.5% in the control group, while 20 were pregnant and the total pregnancy rate was 65.0% in the treatment group. But there was no statistical difference between the two groups (P > 0.05). Compared with the control group, the Gn dosage was lower and the high quality embryo rate was higher in the treatment group, showing statistical difference (P < 0.05). There was no statistical difference in serum concentrations of E2, LH, or P on the HCG injection day, the number of retrieved oocytes, or the fertilization rate (P > 0.05). Compared with the control group, FSH concentrations in the follicular fluid were significantly lower and LH concentrations were significantly higher in the treatment group (P < 0.05). The LH concentrations in the follicular fluid were significantly higher in pregnant patients than non-pregnant patients, showing statistical difference (P < 0.05).There was no statistical difference in E2, T, or P concentrations (P > 0.05). The mRNA and protein expression of BMP-15 in granulosa cells was higher in the treatment group than in the control group (P < 0.05). It was also higher in pregnant patients than non-pregnant patients, showing statistical difference (P < 0.05).

CONCLUSIONDuring the IVF-ET process, BTR could elevate the quality of oocytes, and increase the sensitivity of ovarian follicles to exogenous Gn, which was correlated with the mRNA and protein expression of BMP-15 in granulosa cells, and changing concentrations of FSH and LH.

Adult ; Bone Morphogenetic Protein 15 ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Embryo Transfer ; Estradiol ; blood ; metabolism ; Female ; Fertilization in Vitro ; Follicle Stimulating Hormone ; metabolism ; Follicular Fluid ; metabolism ; Humans ; Luteinizing Hormone ; blood ; metabolism ; Oocytes ; drug effects ; Pregnancy ; Progesterone ; blood ; metabolism ; Testosterone ; metabolism ; Young Adult

Objective To determine the effect of mild hypothermia on neonatal piglet cardiac hemodynamic function after hypoxia-ischemia (HI).Method Twenty five 7-day-old piglets were used for hypoxic ischemic brain damage (HIBD) model by the method of temporary occlusion of the bilateral carotid arteries and followed by mechanical ventilation with low concentration of oxygen (FiO_2=6%) for 30 minutes.The piglets were randomly divided into three groups:group A (normothermia with body temperature to 39℃,n=9),group B (body temperature to 36℃for 72 hours,n=8),and group C (body temperature to 34℃for 72 hours,n=8).Mild hypothermia was initiated at 4 hours after HI,the systolic and diastole function were evaluated by Doppler echocardiography at pre-HI,post-Hi 4 hours and post-HI 72 hours.Results There were no significant differences in left ventrieular ejection time/left ventrieular ejection time (LPEP/LVEF),right ventricular ejection acceleration time/right ventricular ejection time (RACT/RVET) and CO at post-HI with hypothermia 72 hours in three groups,but the heart rate decreased in B and group C group.Compared with nonnothermia,mild hypothermia treatment showed no significant differences in MAP,LPEP/LVET,RACT/RVET,CO,SV at post-HI with hypothermia 72 hours.Conclusions Body temperature decreased by 3～5℃for 72 hours will not aggravate hemodynamic abnormity.

Objective To construct a recombinant plasmid pGEX-TSO45W-4B-TSOL18 of Taenia solium,to obtain purified recombinant protein TSO45W-4B-TSOL18 and to prepare rabbit antiserum against the recombinant fusion protein.Methods TSO45W-4B and TSOL18 encoding genes were connected with hydrophobic (Gly4Ser)3 linker.TSO45W-4B-TSOL18 fusion gene was synthesized and cloned into an expression vector pGEX-1λT to construct a recombinant plasmid pGEX-TSO45W-4B-TSOL18.The positive recombinants were transformed into Escherichia(E.) coli ArcticExpress (DE3),and the expression of recombinant protein was induced with isopropylβ-D-thiogalactopyranoside (IPTG) and analyzed by sodium dodecyl sulfate polyacrylamide gelelectrophoresis (SDS-PAGE).The expressed recombinant fusion protein was purified through affinity chromatography.The purified recombinant protein was mixed with complete Freunds adjuvant at a dosage of 0.5 mg.Rabbit was intramuscularly and subcutaneously immunized in the hind leg and the back,respectively.After 2 weeks the rabbit was boosted with purified recombinant protein which was mixed with incomplete Freunds adjuvant at the same dosage.Rabbit can be boosted every 10 days until an adequate response was achieved.At the 6th day after the last immunization,blood was collected from the rabbit heart.Serum was separated to purify and prepare the antiserum.ELISA was applied to determine the titer of the antiserum and Western blot assay was used to determine the specificity of the antiserum.Results The size of the synthesized fusion gene TSO45W-4B-TSOL18 was 789 bp.The results of restriction enzyme digestion showed that the recombinant plasmid pGEX-TSO45W-4B-TSOL18 was successfully constructed.DNA sequencing showed that the size of the fusion gene TSO45W-4B-TSOL18 was 789 bp,which was consistent with expected result.As demonstrated by SDS-PAGE,relative molecular mass of the expressed recombinant fusion protein was approximately 55 × 103,and its purity was 85％ after purification with affinity chromatography.Titer of the antiserum against the recombinant protein was 1 ∶ 512 000 in ELISA assay and the specific rabbit antiserum against the purified recombinant protein TSO45W-4B-TSOL18 could specifically bind to the recombinant fusion protein in Western blot assay.The relative molecular mass of the specific band was about 55 × 103,which was consistent with expected size.Conclusions The recombinant plasmid pGEX-TSO45W-4BTSOL18 of Taenia solium is successfully constructed.High quality recombinant fusion protein TSO45W-4B-TSOL18 and high titer rabbit antiserum are successfully prepared.

Objective To describe the prevalence of aggressive behavior among preschool children and its related family factors.Methods 1234 preschool children in ten kindergartens were rated on their aggressive behavior by their parents,using Child Behavior Checklist(CBCI),Parent Behavior Inventory(PBI)and a general questionnaire.Results The overall prevalence of aggressive behavior among preschool children was 12.9％(95％ CI:11.0-15.0)according to the CBCL assessment,with the rate being slight higher(13.7％,93/680)in boys than in girls(11.9％,66/554).Data from logistic regression analysis showed that parents' hostile/coercive parenting style(OR=2.396,95％CI:1.636-3.510)and inconsistent parenting attitude between parents and grandparents(OR=1.867,95％ CI:1.287-2.710)would lead to more aggressive behaviors in preschool children.Compared with childen without difficulty in falling asleep,those who often(OR=3.415,95％ CI:1.901-6.135)or sometimes(OR=2.147,95％ CI:1.256-3.671)had problem falling asleep at night had more aggressive behaviors.On the other hand,factors as:watching TV less than 1 hour each day (OR=0.252,95％ CI:0.136-0.467),father in older age(OR=0.703,95％ CI:0.503-0.983)and participating in regular outdoor activitiess(OR=0.6 1 7,95％CI:0.399-0.955)were protective factors to the aggressive behaviors of the children.Conclusion The prevalence of aggressive behavior in preschool children was high which called for more attention.Intervention programs targeting the family should consider the influencing factors as ways of parenting,consistent attitude on parenting in the family etc.to reduce the occurrence of aggressive behavior among preschool children.

Objective To study the correlation between interrupter technique to measure airway resistance and routine pulmonary function(Raw).Methods Using Dyn′R spirometer measure Raw and FVC,PEF,MEF_ 75,MEF_ 25,MEF_ 25～75,and calculate correlation coefficient.Results There was a negative correlation between Raw and FVC, PEF, MEF_ 75,MEF_ 25,MEF_ 25～75(r

Objective To explore the effect of carvedilol on ACE2 gene and protein expression in chronic heart failure rats after myocardial infarction.Methods The heart failure model was induced by acute myocardial infarc- tion (AMI) through ligating the left anterior descending coronary artery.One month after operation,rats were randomized to receive placebo or carvedilol 2 mg/(kg?d),by gavage.Sham-operated rats were used as the control group.Hemodynamies,body mass and left ventrieular mass index,plasma and myocardial level of angiotensin Ⅱ were determined.ACE2 gene and protein expression was assessed by using RT-PCR and Western Blot.Results The mortality of placebo and Carvedilol groups were 20%,compared with 0% in sham operated rats.Carvedilol significantly improved LVEDP,LVSP,+dp/dt_(max) and-dp/dt_(min) in CHF rats but all the hemodynamics data were still inferior than that of controls.Plasma and myocardial angiotensin Ⅱ level were increased significantly in CHF placebo rats than those of control rats (plasma Ang Ⅱ:CHF:194?19 vs controls:132?15 ng/L,myocardium Ang Ⅱ:CHF:6.7?0.4 vs control:3.8?0.3 ng/g,P

OBJECTIVE To investigate beta-lactamases production in multi-resistant Citrobacter strains isolated from the First Affiliated Hospital of Anhui Medical University.METHODS Standard agar dilution method was used to determine the minimal inhibitory concentration(MIC) of 52 clinically isolated Citrobacter strains,improved three-dimensional tests were performed to detect the ESBLs-producing,AmpC-producing and MBL-producing strains in 31 Citrobacter isolates,then beta-lactamases-encoding genes were amplified by polymerase chain reaction(PCR).RESULTS Thirty one Citrobacter strains were resistant to many beta-lactams antibiotics.Twenty four strains ESBLs-producing,3 AmpC-producing and 2 ESBLs-producing plus AmpC-producing strains have been detected by improved three-dimensional tests,and not found MBL-producing ones.Twenty strains had blaTEM gene and 1 strain had blaSHV gene,8 strains had blaCTX-M-1 gene,15 strains had blaCTX-M-13 gene and 5 strains had blaCIT gene by PCR.CONCLUSIONS Multi-resistant Citrobacter strains produce one or more types of ESBLs and AmpC beta-lactamase,it may be an important reason of resistance to beta-lactams antibiotics.