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Objective:To study the effects of Zuogui Pills on the expressions of miR-133b-3p and RhoA in osteoclasts of postmenopausal osteoporosis rats; To discuss its potential mechanism.Methods:SD female rats were randomly divided into normal group, model group, sham-operation group, and Zuogui Pills group using a random number table method, with 6 rats in each group. The model group and Zuogui Pills group were treated with oophorectomy to construct a rat model of osteoporosis. Zuogui Pills group was orally administered with Zuogui Pills decoction at a concentration of 10 g/kg for 12 consecutive weeks. Colorimetric method was used to measure the serum calcium and phosphorus levels of rats, and ELISA method was used to detect ALP levels. Bone density meter was used to measure the bone density of the femurs of rats in each group. The osteoclast of each group were cultured, and the expressions of RANKL and RUNX2 protein were detected by Western blot. MiRNA sequencing and differential expression analysis were performed on bone tissues of rats. Osteoclasts were treated with miR-133b-3p mimic and its negative control. The cell proliferation activity of osteoclasts was detected by cell counting kit-8 (CCK-8). The osteoclast differentiation activity was detected by the tartrate-resistant acid phosphatase staining. The dual-luciferase reporter assay was used to detect the relationship between miR-133b-3p and RhoA. The "rescue" experiment of miR-133b-3p mimic and RhoA co-expression were used to study the molecular regulatory mechanism of Zuogui Pills on osteoclast activity.Results:Compared with the model group, the bone mineral density of Zuogui Pills group significantly increased ( P<0.05, P<0.01), the levels of calcium and phosphorus in serum increased, the level of alkaline phosphatase ALP decreased ( P<0.05), the expression of RANKL protein decreased, and the expression of RUNX2 protein increased. Sequencing results showed that rno-miR-133b-3p was down-regulated in osteoclasts of postmenopausa osteoporosis rats treated with Zuogui Pills with the maximum difference ( P<0.01). Q-PCR results showed that the expression of miR-133b-3p in osteoclasts of Zuogui Pills group was significantly lower than that of the model group. The upregulation of miR-133b-3p could significantly promote the cell proliferation and differentiation of osteoclasts. RhoA overexpression could reverse the excessive proliferation and differentiation of osteoclasts caused by miR-133b-3p overexpression. Conclusions:RhoA is the target gene regulated by miR-133b-3p. Zuogui Pills can inhibit the activity of osteoclasts by regulating miR-133b-3p/RhoA axis, relieving the symptoms of osteoporosis.
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Objective:To investigate the role of peptidyl arginine deiminase 4 (PAD4) in anti-β 2GP1/β 2GP1 complex-induced formation of neutrophil extracellular trapping networks (NETs). Methods:Peripheral blood neutrophils were isolated from healthy humans by density gradient centrifugation. PAD4 expression was detected by Western blot after the neutrophils were incubated with anti-β 2GP1/β 2GP1 complex (100 μg/ml) for a certain period of time. PAD4 inhibitor Cl-amidine (10 μmol/L) was used to pretreat neutrophils. Changes in the expression of citrullinated histone 3 (CitH3) at protein level and the relative content of myeloperoxidase (MPO)-DNA were detected by Western blot and ELISA, respectively. A mouse thrombus model of antiphospholipid syndrome (APS) was established by inferior vena cava stenosis. Intervention experiments were performed by intraperitoneal injection of Cl-amidine (50 mg/kg). The expression of CitH3 at protein level in plasma was detected by Western blot. The concentration of circulating free DNA (cf-DNA) in plasma was measured with fluorescent staining. Thrombus in inferior vena cava was collected and weighted to evaluate whether inhibiting the activity of PAD4 would suppress the APS-IgG-induced formation of NETs and thrombosis. Differences among groups were analyzed by t test or one-way analysis of variance (ANOVA). Results:The expression of PAD4 induced by anti-β 2GP1/β 2GP1 complex was significantly down-regulated in the cytoplasm, but increased in the nucleus [(3.67±0.32) vs (1.47±0.19), t=10.22, P<0.05; (0.57±0.19) vs (2.97±0.31), t=11.49, P<0.05]. Cl-amidine significantly inhibited the anti-β 2GP1/β 2GP1 complex-induced expression of CitH3 protein by neutrophils [(2.46±0.47) vs (0.46±0.13), t=12.24, P<0.01], and reduced the MPO-DNA content in the culture supernatants [(4.09±0.94) vs (2.80±0.57), t=4.23, P<0.05]. In vivo, Cl-amidine significantly inhibited the expression of CitH3 protein [(3.97±0.56) vs (1.09±0.45), t=11.83, P<0.01] and decreased the content of cf-DNA [(2 685.0±735.8) vs (1 784.0±577.0), t=3.93, P<0.05] in plasma of APS mice. Compared with the experimental APS mice in the control group, the weight of thrombus in the APS mice pretreated with Cl-amidine was significantly reduced [(8.22±3.06) vs (4.89±1.90), t=2.27, P<0.05]. Conclusions:PAD4 was involved in the formation of NETs induced by anti-β 2GP1/β 2GP1 complex, which might play an important role in APS thrombosis.
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Objective To detect the levels of neutrophil extracellular traps (NETs) in patients with antiphospholipid syndrome (APS) and to preliminarily explore its formation mechanism.Methods Plasma samples from 27 APS patients and 30 healthy controls were collected.The circulating free DNA (cf-DNA) in plasma was detected by the PicoGreen nucleic acid quantitative assay kit,and the concentration of citrulline histone 3 (CitH3) was analyzed by enzyme-linked immuno sorbent assay (ELISA).The association of cf-DNA/NETs with thrombotic events in APS patients was further analyzed.The neutrophils in healthy controls were separated by density gradient centrifugation and stimulated with anti-β2GPl/β2GPI complex (100 μg/mL) for 4 h,and the cf-DNA/NETs in the culture supernatant was determined.TLR-4 inhibitor-TAK242 (5 μmol/L) was further used to observe whether the stimulation of the anti-β2GPI/β2GPI complex on cells could be intervened.The differences between groups were analyzed by analysis of variance (ANOVA) or rank sum test,Sidak or Dunnett's test were used to compare the mean of multiple samples and the correlation between variables was analyzed by Spearman's correlation test.Results The concentration of cf-DNA/NETs and CitH3 were significantly increased in plasma of APS patients compared with that in healthy controls [175.7(70.6,205.7) ng/ml vs 29.8(7.6,115.7) ng/ml,Z=-3.654,P＜0.05;19.5(7.8,26.4) ng/ml vs 3.3(0.84,10.3) ng/ml,Z=-3.932,P＜0.05],and there was a significant positive correlation between the cf-DNA/NETs and CitH3 (r=0.447,P=0.019).In the APS group,there was no significant difference in cf-DNA/NETs between patients with arterial thrombosis and those with venous thrombosis [177.1(67.8,297.2) ng/ml vs 184.7(82.4,233.9) ng/ml,Z=-0.301,P=0.786],whereas cf-DNA/NETs in the patients who experienced a new thrombotic event in 1 month was significantly higher than those with a history of thrombosis [192.1(83.6,328.8) ng/ml vs 90.0(42.8,184.7) ng/ml,Z=-2.006,P=0.046].In vitro,anti-β2GPI/β2GPI complex (100 μg/ml) stimulated the release of cf-DNA/NETs from neutrophils,which was significantly increased compared with the control group (t=10.39,P＜0.05),while TAK242 significantly inhibited the stimulating effects of anti-β2GPI/β2GPI complex on cells (t=4.22,P＜0.05).Conclusion The level of cf-DNA/NETs in peripheral blood of APS patients is significantly increased,which may play an important role in APS thrombosis.Anti-β2GPI/β2GPI complex induces the formation of cf-DNA/NETs through TLR4 and participates in the pathological process of APS.
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We explored the status of Bartonella infection in rodents and the sequence characteristics of Bartonella in Fujian Province.Rodents in Fujian Province were captured by the night trapping method during 2014-2016.Information of the captured rodents on capturing dates and geographic locations,species,gender and ages were recorded.Heart blood samples were collected,from which the fragments of ghA gene and16S-23S rRNA gene of Bartonella were amplified by polymerase chain reaction (PCR).The PCR products were sequenced and the phylogenetic tree was constructed for homology analysis by biological analysis software.Data on infection rate were analyzed with Chi-square or Fisher exact test to indicate statistical significance.Results showed that 5 917 cages were laid and 381 rodents were captured,density of rodent was 6.44％.The overall Bartonella infection rate in rodents was 12.34 ％,while infection rate in domesticated rodents was 10.61％,with 11.30 ％ in Rattus norvebicus and 10.00％ in R.flavipectus.And the infection rate in wide rodents was 13.86％,with a rate of 22.86％ in Rattus losea and 18.00％ in R.fulvescens,respectively.The infection rate was higher in wild rodents than in domesticated rodents,however,no significant difference was found.The Western Fujian and Northern Fujian region had the higher infection rates of 20.00％ and 25.33％,and no infection was found in Southern Fujian region.The statistical analysis result revealed that a significant difference in infection rate among different region and habitats,but no significant difference in infection rate between male and female rodents,or among different ages.The BLAST results revealed the species to be B.tribocorum,B.elizabethae and B.grahamii.In conclusion,Bartonella infection is found in the rodents in Fujian Province and more attention should be paid on its impact on public health in the province.
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BACKGROUND: Transcranial Doppler(TCD) spectrum analysis techniques cannot measure the diameters of blood vessels and therefore and not be used to calculate the changes of brain blood flow. A formula was designed for regional cerebral blood flow loss index(rCBFLI) following arteriostenosis when arteriostenosis index formula was deduced; the former may provide a reference to assess cerebral function.OBJECTIVE: To introduce the formula of cerebral blood flow loss ratio obtained basing on TCD results.DESIGN: Retrospective controlled study based on patients with cerebral middle arteriostenosis diagnosed with TCD and healthy people.SETTING: At the departments of neurology in a university hospital and in a military medical university affiliated hospital of Chinese PLA.PARTICIPANTS: Between October 1994 and December 2000,43 patients who were confirmed of arteriostenosis with TCD at outpatient or neurological ward of the First Affiliated Hospital of Fujian Medical University were enrolled into this study,and their TCD results were compared with 908 normal controls between March 1992 and September 2000.METHODS: Stenosis index (STI) formula was deduced from TCD results of 31unilateral and 12 bilateral stenoses at middle cerebral artery(MCA) . STI = 1- [ normal average blood velocity (Vm0) ÷ average blood velocity at stenosisSince(PI1 ÷PI0) may be equal to [post-stenosis blood flow(Q1) ÷ normal blood flow(Q0)],regional cerebral blood flow loss index is concluded as:MAIN OUTCOME MEASURES: rCBFLI,relationship between STI and Vm,as well as between rCBFLI and STI.RESULTS: rCBFLI formula was used to calculate the rCBFLI of 31 patients following unilateral MCA stenosis. rCBFLI of these 31 patients was obtained by Pearson statistical analysis. STI was found to be related to the average blood velocity at stenosis with relative coefficient(Vm) of(r) =0.76( P ＜ 0. 001 )and r = 0. 83 ( P ＜ 0. 001 ) . rCBFLI was proved to be correlated with Vm and STI with r = 0. 76( P ＜ 0. 01 ) and r = 0. 81 in 55 side MCA stenoses,and the difference was of statistical significance( P ＜ 0. 001 ).CONCLUSION: rCBFLI can be used to assess the decrement of regional brain blood flow due to the stenosis of supplying artery; also,it can be used to observe the changes of brain hemodynamics if combined with STI.
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The study applied the model of authorized leadership to emergency night nursing in large general hospitals in order to explore the effect of such management. As it turned out, the evaluation of the administrators means of authorization by the three groups of research subjects averaged respectively 20 70,20 62 and 20 05, and the evaluation of the effect of authorization averaged respectively 79 80,78 39 and 83 55. The results indicate that the administrators means of authorization were basically rational and sound and the managerial abilities of those with authorized leadership were appreciated by the emergency night doctors and nurses. Thus management of emergency night nursing by those with authorized leadership may lead to improved efficiency and quality.
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Objective To explore the neuroprotective mechanism of mild hypothermia by studying the effect of mild hypothermia on neuronal apoptosis after transient global cerebral ischemia in rats.Methods A model of transient global cerebral ischemia was made by bilateral common carotid artery occlusion and hypotension in rats. The neuronal demage in the CA 1 section of the hippocampus after cerebral ischemia was observed by nissel body stain mothed;apoptosis was detected by in situ end-labeling of fragmented DNA using the terminal deoxynucleotidyl transferase reaction (TUNEL) and the electrical microscope.Results The hippocampal CA 1 section neurons in normothermic ischemic group were significantly deficit as compared with the false operative group in mild hypothermic ischemic group (P