Objective To study the expression of Toll-like receptor 4(TLR4) in peripheral blood mononuclear cell (PBMCs) and serum interleukin-6(IL-6) in wheezing children under 5 years of age.Methods A total of 224 wheezing children under 5 years of age were divided into API(asthma predictive index)-positive (n=116) and API-negative groups (n=108).Serum level of IL-6 and TLR4 expression on CD14+ monocytes were measured after wheezing was stable for one month.TLR4 expression on CD14+ monoeytes was quantified via flow-cytometry.Serum level of IL-6 was detected by ELISA.Results Serum level of IL-6 and TLR4 expression on CD14+ monocytes of API-positive group were higher than API-negative group [LR4(％):34.9±10.0 vs.30.2± 8.8;IL-6(ng/L):46.4±15.1 vs.40.5±13.6].There was a significant positive correlation between the expression of TLR4 and the content of serum IL-6 in two groups of wheezing children(P＜0.05).Conclusion TLR4 may play a role in the pathogenesis of asthma through promoting the expression of IL-6.TLR4 may be a index to predicting asthma in wheezing children.

Objective To investigate the effects and possible mechanisms of platelet-activating factor (PAF) on the invasion of ovarian cancer cells and to provide a potential target for ovarian cancer therapy.Methods ( 1 ) Serous type ovarian cancer cell line OVCA429 with platelet-activating factor receptor (PAFR) positive and mucinous type cell line RMUG-L (PAFR negative) were treated with 100 nmol/L of the PAF,cell invasion ability was determined by transwell cell migration assay.(2) For determination of the optimal PAF concentration,ovarian cancer cell OVCA429 was treated by 0,0.1,1,10,100,and 1000 nmol/L of PAF for 10 minutes or 24 hour,respectively.To observe the different time point of protein changes,OVCA429 were treated by 100 nmol/L of PAF for 0,5 minutes,10 minutes,30 minutes,1 hour or 12 hours,respectively.The total proteins of treated cells were extracted according to standard protocol.The expression of p38 mitogen-activated protein kinase (p38 MAPK),phosphorylated p38 MAPK (p-p38 MAPK),transcription factor response element-binding protein (CREB),phosphorylated CREB (p-CREB) and matrix metalloproteinase-2 (MMP-2) were detected by western blot.(3) To verify the pathway involved in the PAF induction of the cancer cell invasion,we repeated the experiments by adding the inhibitors when treating cells with PAF.The inhibitors used were as follows,PAFR inhibitor-WEB2076 (50 μmol/L),pp38 MAPK inhibitor-SB203580 (10 μmol/L),CREB binding protein (CBP)-CREB interaction inhibitor217505(25 μ mol/L).All treated cells were divided into 6 groups:control group,PAF group,PAF + WEB2076 group,PAF + SB203580 group,PAF + 217505 group and PAF + SB203580 + 217505group.Results ( 1 ) By transwell assay,100 nmol/L of PAF could improve the invasion ability of OVCA429 cell significantly [ PAF:( 118 ± 23 ) cells vs.control:(36 ± 8 ) cells,P ＜ 0.0l ],while the same treatment had no effect on RMUG-L cells [PAF:(45 t 13) cells vs.control:(53 ±9) cells,P＞0.05].(2) Even a very low concentration of PAF (0.1 nmol/L) could increase the expression of p-CREB and MMP-2,while the most effective concentration of PAF was 100 nmol/L.The highest p-CREB protein expression was detected at 10 minutes after administration of 100 nmol/L PAF,as well as the expression of p-p38 MAPK protein.Even 12 hours after treatment the p-p38 MAPK protein could be detected,while there was no significant difference in the expression of CREB ( P ＞ 0.05 ).(3) As compared with PAF group,both in PAF + WEB2076 group and PAF + SB203580 group,the expressions of p-p38 MAPK,p-CREB and MMP-2 protein were decreased significantly; in PAF + 217505 group,although the expression of p-p38 MAPK and p-CREB protein was significantly higher than the control group,the expression of MMP-2 protein was significantly lower; in PAF + SB203580 + 217505 group,the expression of these three proteins were also significantly lower,but there was no significant difference as compared with that in the PAF + WEB2076 or PAF + SB203580 group.Conclusion PAF could induce MMP-2 expression and contributed to PAFR positive ovarian cancer cell invasion via activation of CREB by phosphorylating of p38 MAPK.

Objective To establish a scald-induced pain model using a constant-temperature electrical scald instrument in rats.Methods Thirty-six pathogen-free male Sprague-Dawley rats, weighing 200-250 g, were randomly divided into 4 groups (n =9 each) using a random number table: control group (group C), scald for 5 s group (group S5), scald for 10 s group (group S10), and scald for 15 s group (group S15).The rats were anesthetized with intraperitoneal chloral hydrate.In S5, S10 and S15 groups, the plantar surface of the left hindpaw of rats were exposed to a constant-temperature electrical scald instrument (85 ℃) for 5, 10 and 15 s, respectively.The plantar surface of the left hindpaw of rats was exposed to an electrical scald instrument (room temperature) for 10 s in group C.At 1 day before treatment (T0),and 1, 3, 5, 7 and 14 days after treatment (T1-5), the mechanical and thermal pain thresholds were measured.Immediately after treatment, and at 24 h after treatment, the total body condition, wound color, and shape of the margin of the wound were observed and recorded.At 24 h after treatment, 3 rats were randomly sacrificed, and the skin from the plantar surface of the left hindpaw was removed for microscopic examination.Results Compared with group C, the thermal pain threshold at T1-2, and the mechanical pain threshold at T1-3 were significantly decreased in group S5, and the thermal pain threshold,and mechanical pain threshold were significantly decreased at T1-4 in group S10 (P＜0.05).The thermal pain threshold ＞ 25 s, and the mechanical pain threshold ＞30 g at T1-5 in group S15.The swelling in foot was bovious, burn blister appeared, and the degree of damage was aggravated in group S10 compared with S5 and S15 groups.Conclusion The scald-induced pain model is successfully established using a constant-temperature electrical scald instrument in rats.

Objective To investigate the expression of Toll-like receptor 4 ( TLR4 ) and nuclear factor-κB ( NF-κB) in patients with critically severe hand, foot and mouth disease ( HFMD) and to evaluate the effects of esmolol intervention on those patients.Methods Fifty-two hospitalized children with critically severe HFMD in the Intensive Care Unit of Xuzhou Children′s Hospital were enrolled in the study from May 2014 to May 2015 and randomly divided into two groups, represented as group A and group B.Children in the group A were given routine treatment, while those in the group B were treated with esmolol in addition to the routine therapy.Thirty children with common HFMD were selected as disease control, and thirty healthy children were set up as normal control.Differences in the expression of TLR4, NF-κB, TNF-αand IL-6 among all children were comparatively analyzed.The levels of TLR4, NF-κB, TNF-αand IL-6 in children from groups A and B were detected after 24 hours, 72 hours and five days of treatment and the differences betweenthetwogroupswereanalyzed.Results (1)Comparedwiththechildrenfromdiseasecontroland normal control groups, those with critically severe HFMD showed significantly increased expression of TLR4, NF-κB, TNF-αand IL-6 (P<0.01).(2)No significant differences in the expression of TLR4, NF-κB, TNF-αand IL-6 were found between the two subgroups of children with critically severe HFMD before treat-ment (all P>0.05).The expression of TLR4, NF-κB, TNF-αand IL-6 in children from both subgroups were significantly decreased after receiving corresponding treatments for 24 hours, 72 hours and five days (all P<0.05).Compared with the children form group A, those from group B showed significantly decreased expression of NF-κB, TNF-αand IL-6 after 24 and 72 hours of corresponding treatments (all P<0.05).No significant difference in the expression of TLR4 was observed between the two subgroups after 24 and 72 hours of corresponding treatments (P>0.05).No significant differences in those observed indicators were found between the two subgroups after five days of treatments (all P>0.05).Conclusion The TLR4/NF-κB/proinflammatory factor pathway might play an important role in the development of critically severe HFMD.Treatment with esmolol could inhibit the expression of NF-κB, reduce the secretion of inflammatory factors and alleviate the inflammatory reaction during critically severe HFMD.

BACKGROUND: Insufficient angiogenesis and the lack of skin appendages are critical challenges in cutaneous wound healing. Stem cell-fabricated cell sheets have become a promising strategy, but cell sheets constructed by a single cell type are inadequate to provide a comprehensive proregenerative microenvironment for wound tissue. METHODS: Based on the communication between cells, in this study, bone marrow mesenchymal stem cells (BMSCs) and hair follicle stem cells (HFSCs) were cocultured to fabricate a composite cell sheet (H/M–CS) for the treatment of fullthickness skin wounds in mice. RESULTS: Experiments confirmed that there is cell–cell communication between BMSCs and HFSCs, which enhances the cell proliferation and migration abilities of both cell types. Cell–cell talk also upregulates the gene expression of proangiogenic-related cytokines in BMSCs and pro-hair follicle-related cytokines in HFSCs, as well as causing changes in the properties of secreted extracellular matrix components. CONCLUSIONS Therefore, the composite cell sheet is more conducive for cutaneous wound healing and promoting the regeneration of blood vessels and hair follicles.

Objective To investigate the effects of inverse ratio ventilation on pulmonary function and ce-rebral oxygen saturation in elderly patients with single lung ventilation.Methods Sixty patients scheduled for elec-tive radical resection of esophageal cancer were divided into 2 groups(n=30 for each group)using a random num-ber table:the experiment group(group A)and the control group(group B). During the two lung ventilation,the ventilator parameters were set as tidal volume(VT)7 mL/kg,inspiratory to expiratory ratio 1:2. During one lung ventilation,the I:E ratio was 1.5:1 in the group A and 1:2 in the group B. At 15 min after two lung ventilation (T1),20 min after one lung ventilation(T2),60 min after one lung ventilation(T3)and 15 min after restarting two-lung ventilation(T4),the blood gas analysis was measured and recorded for the hemodynamics,respiratory me-chanics index and cerebral oxygen saturation respectively. Results Compared with the B group,the Ppeak and VD/VT at T2~T4in the group A were lower while PaO2,Pmean and Cdyn were higher(P<0.05).During one lung ventilation,the incidence of rSO2< 50% or rSO2decreased more than 20% in the group A was lower than that in the group B(P<0.05).The PaO2,Cdyn and rSO2of the two groups at T2~T4were significantly lower and Ppeak, Pmean,PaCO2,VD/VT were significantly higher than the baseline(T1)(P < 0.05). Conclusion During one-lung ventilation,prolonged inspiratory time can improve pulmonary function and lung compliance without increas-ing peak airway pressure,reduce the decline of rSO2 at the same time.