ObjectiveTo study the effects of intrathecal injection of cyclin dependent kinase 5 inhibitor Roscovitine on the hyperalgesia induced by remifentanil in a rat model of incisional pain.MethodsForty-five SD rats were randomly divided into 5 groups ( n =9 in each group):control group ( C ),incisional pain group ( Ⅰ ),Roscovitine group(ROS),remifentanil group(R) and Roscovitine + remifentanil group ( ROS + R).Roscovitine (50μg/10μl) was injected intrathecally at 30 min before plantar incision in groups ROS and ROS + R,other groups were injected with 20％ DMSO(10μl).All groups except for C group needed to be made the model of incisional pain.In group R and ROS + R,remifentanil(0.04 mg/kg) was infused subcutaneously with a pump for 30min at the moment of surgical incision.The paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency(PWTL) were used to evaluate the behavioral changes and measured at 24 h before incision and at 2 h,6 h,24 h and 48 h after incision.ResultsCompared with group C,the PWMT and PWTL of group Ⅰ were significantly decreased after incision (P＜0.01).Compared with group Ⅰ,the PWMT and PWTL of group R were significantly decreased after incision (P ＜ 0.01 ) ; however,the PWTL was significantly increased (2h:( 20.26 ± 1.33)s,(17.97 ±0.47)s;48h:(22.15 ±0.660)s,(19.89 ±1.27) s),P＜0.05)in ROS group.Compared with group R,at 2h after incision,the significant increase of PWTL ( ( 19.13 ± 1.72)s,( 14.41 ± 2.30) s) and PWMT ( ( 10.4 ± 1.95 ) g,(6.38 ± 0.91 ) g) were observed,then lasted up to 48 h ( ( 19.24 ± 2.8 ) s,( 14.87 ± 1.95 ) s )and 24h ( (8.88 ± 1.41 ) g,( 6.83 ± 0.80) g) respectively in group ROS + R (P ＜ 0.05 ).ConclusionIn a rat model of postoperative pain,Roseovitine could reduce the thermal hyperalgesia purely induced by incision,and also could prevent the development of hyperalgesia induced by remifentanil.

A new phenolic glycoside was isolated from the spikes of Prunella vulgaris. Its structure was elucidated as gentisic acid 5-O-beta-D-(6'-salicylyl)-glucopyranoside by spectroscopic evidence and chemical analysis.

Objective To ascertain the significance of qualitative and quantitative detections of urine neutrophil gelatinase-associ-ated lipocalin(NGAL)in the early diagnosis of diabetic nephropathy.Methods 74 patients with diabetes mellitus were divided into the normoalbuminuria group[(DN1 ,n =26,urinary albumin excretion rate(UAER)<30 mg/24 h],the microalbuminuria group (DN2 ,n=24,UAER 30-300 mg/24 h)and the macroalbuminuria group(DN3 ,n=24,UAER>300 mg/24 h)according to the 24 h UAER,and at the same time the control group(n=25)was set up.Both the activity and levels of urine NGAL in the above groups were examined and compared.Results The activity band of urine NGAL in each group were more distinct than that of the control group(P =0.000).The NGAL activity in the macroalbuminuria group was higher than that in the normoalbuminuria group(P <0.05).No difference in the NGAL activity was found between the micro-and normal-albuminuria groups(P >0.05);the quantitative detection demonstrated that differences of the NGAL content existing between the macroalbuminuria group and the other three groups had statistical significance(P =0.000),the urinary NGAL excretion in the macroalbuminuria group was higher than that in the microalbuminuria group,normoalbuminuria group and the control group(P =0.000);No differences were found between the control group and the normoalbuminuria group and between the control group and the microalbuminuria group(P >0.05).Conclu-sion For the patients with diabetic nephropathy,the activity detection of urine NGAL is more sensitive than UAER and the NGAL quantitative detection and could become a new type index in the early diagnosis of diabetic nephropathy.

Objective To investigate the role of Ca2+/calmodulin-dependent protein kinase Ⅱ on pain behavior in a mouse model of bone cancer pain. Methods 40 male C3H/HeN mice were divided randomly into 5 groups:sham group (S group, n=8) ,control group (C group, n=8) and KN93 treat group (T1, n=8;T2, n=8;T3, n = 8 ). Group C and T were induced mouse models of bone cancer pain by intra-left-femur inoculations of osteolytic NCTC2472 cells while group S were injected only α-MEM. On the 14 d after inoculations,group S and C received intrathecal injection of 20% DMSO 5 μl . While group T1, T2, T3 received intrathecal injection of KN93 15nmol,30nmol,60nmol which dissolved in 5 μl 20% DMSO respectively. Mice received pain behavior tests including quantification of spontaneous flinches, paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) before and at 0.5 h,2 h,4 h,8 h after administration. Results Treatment with KN93(15 nmol) have no effect on bone cancer pain,while treatment with KN93(30 nmol,60 nmol) can dose-dependently reverse quantification of spontaneous flinches, mechanical allodynia and thermal hyperalgesia which were induced by bone cancer pain, At 0. 5h after administration, the quantification of spontaneous flinches of the two groups ( (7.25 + 1.49 ), (4. 12 + 1.36 ) ) were decreased when compared with control group ( 11.62 + 1.92 ),PWMT((1.28 +0.14)g;(1.75 +0.46)g),PWTL((14.64 +2.12) s; (16.85 + 1.61)s)were increased when compared with control group ( (0.47 + 0. 16) g, ( 11.32 + 1.68 ) s) (P ＜ 0.05 ＝. The effect lasts for at least 4 h and disappears at 8 h. Conclusion CaMK Ⅱ may play an important role in the mechanism of bone cancer pain. Intrathecal KN93 injection can effectively attenuated bone cancer pain.

Objective To investigate the effects of repeated intrathecal cyclic AMP response elementbinding protein (CREB) antisense oligodeoxynucleotide (ODN) on the expression of NR2A in spinal cord in mice with neuropathic pain produced by chronic constrictive injury of the sciatic nerve (CCI).Methods Forty C57BL/6 male mice in which intrathecal catheter was successfully implanted were randomly divided into 4 groups ( n =10 each):normal saline group (group NS),CREB sense ODN group (group S),CREB missense ODN group (group M),and CREB antisense ODN group (group A).In groups NS,S,M and A,normal saline 5μl,sense ODN 5 μg/5 μl,missense ODN 5 μg/5 μl and antisense ODN 5 μg/5 μl were injected intrathecally once a day for 6 days,starting from the 1st day after CCI,respectively.Paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) were measured on day 1 before CCI and on day 1,3,5 and 7 after CCI.Five mice from each group were sacrificed on day 7 and 14 after CCI and the lumbar segment of the spinal cord (L3-5 )was removed for determination of NR2A expression using Western blot and RT-PCR.Results Compared with the baseline value,no significant change was found in PWMT and PWTL on day 1-7 after CCI in group A ( P ＞0.05),while PWMT and PWTL were significantly decreased on day 1-7 after CCI in groups NS,S and M (P ＜0.05).Compared with groups NS,S and M,the expression of NR2A mRNA and protein was significantly downregulated on day 7 and 14 after CCI in group A ( P ＜ 0.05).The expression of NR2A mRNA and protein was significantly up-regulated on day 14 after CCI compared with that on day 7 after CCI in all the groups.Conclusion Intrathecal CREB antisense ODN during the development of neuropathic pain can attenuate neuropathic pain and inhibition of the expression of NR2A in mouse spinal cord may be involved in the mechanism.

ObjectiveTo investigate the effects of cannabinoid receptor 2 (cannabinoid receptor 2,CB2R) agonist JWHO15 on the hyperalgesia induced by remifentanil in a rat model of postoperative pain.Methods Sixty SD rats were randomly divided into 10 groups ( n =6 each ):control groups ( C1 and C2 ),incisional pain groups (I1 and I2),incisional pain plus JWHO15 groups (QI and FI),remifentanil groups (R1 and R2),and JWHO15 plus remifentanil groups ( QR and FR).Rats in group QL/QR and FI/FR were intrathecal injection with 10μg JWHO15 ( diluted in 10μl 20％ DMSO solution) and intraperitoneal administration with 100μg JWHO15 ( diluted in 10μl 4％ DMSO solution) respectively 30 min before plantar incision while rats in group C,I and R were received with the same volume of DMSO solution.Plantar incision surgery was operated in rats of group I,R,QI/FI,and QR/FR.In group R and QR/FR,remifentanil (0.04 mg/kg) was infused subcutaneously to rats with a pump for 30 min at the moment of surgical incision.The paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency (PWTL) at 24 h before incision and at 2 h,6 h,24 h and 48 h after incision were tested to evaluate the behavioral changes.ResultsCompared with group C and baseline,the level of PWMT and PWTL decreased at 2 h,6 h,24 h and 48h after incision in group Ⅰ (P＜ 0.01 ) ;Compared with group Ⅰ,the significant decrease of PWMT and PWTL were observed after incision in group R (P ＜ 0.05 ) ; Compared with group R,the significant increase of PWMT (7.78 ± 1.09) and PWTL ( 17.28 ± 1.58) were observed from 6 h after incision in group QR(P＜0.05).And the increase of PWMT (7.79 ±0.72,9.50 ± 1.17,7.86 ± 1.16) and PWTL ( 16.23 ± 1.50,19.53 ± 1.63,18.10 ± 0.93) were observed at 6 h,24 h and 48 h after incision in group FR(P＜0.05).ConclusionIntrathecal and intraperitoneal administration of JWHO15 in this investigation dose could relief remifentanil-induced hyperalgesia in a rat model of postoperative pain.

Objective To investigate the effects of intrathecally cyclic AMP response element-binding protein(CREB) antisense oligodeoxynucleotide (ODN) on neuropathic pain behaviors.Methods Using mouse model of neuropathic pain induced by chronic constriction injury of sciatic nerve (CCI),24 male C57BL/6 mice successfully received intrathecal catheter implantation and without motor dysfunction were randomly divided into 4groups(n=6):Saline group(NS),CREB sense ODN group(S),CREB missense ODN group(M),CREB antisense ODN group(A).Mice in NS,S,M and A were intrathecally treated with Saline 5μ l,Sense ODN 5μg/5μl,Missense ODN 5μg/5μl and Antisense ODN 5μg/Sμl once daily on day 1 ～6 after CCI respectively.Paw withdrawal mechanical threshold (PWMT) and paw withdrawal thermal latency(PWTL) were tested on day 1 before CCI and day 1,3,5,7,10,14,17,21 after CC(I).Results Mice in A group maintained the pain thresholds in the baseline and lasted at least 7 days after CCI ( 7 d,PWMT:( 0.81 ± 0.20 ) g vs ( 1.00 ± 0.19 ) g,P ＞ 0.05 ;PWTL:(5.96 ± 0.69) s vs (6.93 ± 1.08 ) s,P ＞ 0.05 ).The withdrawal thresholds in the ipsilateral hind paws of the mouse were significantly lower than baseline in A group on day 10 after CCI( 10 d,PWMT:(0.56 ±0.19)g vs (1.00±0.19)g,P＜0.05; PWTL:(3.93 ±0.28)s vs (6.93 ± 1.08)s,P＜0.05).Compared with NS group ( 10 d,PWMT:(0.56 ±0.19)g vs (0.37 ±0.08)g,P＜0.05; PWTL:(3.93 ±0.28)s vs (3.14 ±0.45)s,P＜0.05),S group,M group,the withdrawal thresholds of A group was significantly elevated on day 10 after CCI.These effects lasted up to at least day 21 after CCI.Conclusion Intrathecally treated with CREB antisense ODN in the development of neuropathic pain induced by CCI completely improved pain behaviors during the course of injection,and the effects of relief pain lasted at least 15d after no injection.

Objective To investigate the mechanism related to reduced antibiotic sensitivity of Acinetobacter baumannii inducted by meropenem in vitro.Methods Three strains of clinically isolated carbapenems-sensitive Acinetobacter baumannii were induced by meropenem in vitro, and the mutant strains (MS1, MS2 and MS3) were obtained.Minimal inhibitory concentrations (MICs) of antimicrobial agents to strains before and after induction were determined by automatic drug sensitivity analyzer .The homology of strains was analyzed by Enterobacterial repetitive intergenic consensus -polymerase chain reaction ( ERIC-PCR).Modified Hodge test and EDTA-Na2-double disk synergy test were used to detect carbapenemase and metallo-β-lactamase (MBL), respectively.Main carbapenemase genes were detected by PCR and followed by DNA sequencing.Expressions of adeB and outer membrane proteins in strains before and after induction were detected with fluorescence quantitative PCR and SDS -polyacrylamide gel electrophoresis , respectively.t test was used for data analysis .Results The sensitivity of mutant Acinetobacter baumannii strains to meropenem and most antibiotics was reduced , except to imipenem, amikacin and polymyxin; and the reduced sensitivity to meropenem in MS2 and MS3 was of genetic stability.ERIC-PCR showed 100%homology between the mutant strains and parental strains .Both carbapenemase and metallo -β-lactamase were negative in mutant strains and parental strains , and only OXA-51 gene was found.The expressions of adeB gene in mutant strains were 24.26 ±0.91, while those in parental strains were 22.81 ±0.38, and the difference was not significant (t =2.534, P >0.05).Outer membrane protein with molecular weight 54 000 was missing in MS1, while that with molecular weight 47 000 was missing in MS2 and MS3.Conclusion Reduced antibiotics sensitivity in meropenem -induced Acinetobacter baumannii may be correlated with the deficiency of outer membrane protein with molecular weight 47 000.

Objective To investigate the clinical value of ultrasonic diagnosis for neonatal annular pancreas,analyze the reasons of missed diagnosis and misdiagnosis,and improve diagnostic accuracy of ultrasonography for this disease.Methods Clinical data of 98 newborns with annular pancreas confirmed by gastrointestinal contrast and surgery were analyzed retrospectively.Preoperative ultrasonogram were compared with the result of gastrointestinal contrast and surgery.Ultrasound images failed to be correctly dignosed were further studied to summarize diagnostic points for this disease.Results Among the 98 cases, 75 were correctly diagnosed by ultrasound with a diagnostic accordance rate of 76.5%,1 8 were missed diagnosed and 5 were misdiagnosed with a total misdiagnosis rate of 23.5%.Ten cases associated with other congenital gastrointestinal tract anomalies were missed diagnosed due to ignoring scanning pancreas.Five cases were missed diagnosed due to obvious intestinal cavity flatulence.Three cases were missed diagnosed due to lack of awareness of the disease.Five cases were misdiagnosed for duodenal stenosis or duodenal atresia.Conclusions Ultrasound has important diagnostic value for neonatal annular pancreas,providing the dignostic evidences for clinical treatment.Thus it can be used as the preferred auxiliary examination of the disease.Since annular pancreas is often accompanied by other gastrointestinal malformations and can be interfered by abdominal gas,missed diagnosis and misdiagnosis occurred easily.To improve the accuracy of ultrasonic diagnosis,all causes of neonatal gastrointestinal tract obstructions should be considered during the examination.

Objective To identify the drug resistance-related genes in a clinically isolated strain of Enterobacter cloacae.Methods A strain of Enterobacter cloacae was isolated from sputum of a patient with chronic obstructive pulmonary disease from the Affiliated Hospital of Xuzhou Medical University in March 2013.Modified Hodge test and metal enzyme inhibition test were performed for drug-resistant phenotype screening.Carbapenemase genes blaMUS-1, blaVIM-1, blaVIM-2, blaIMP, blaKPC-2, blaNDM-1, blaOXA-48 and blaGESwere amplified by polymerase chain reaction (PCR), and the positive products were sequenced and analyzed.Plasmid conjugation and transformation experiments were used to confirm that the resistance gene mediated by plasmids.Agar dilution method was used for antibiotic susceptibility test.Results Both modified Hodge test and metal enzyme inhibition test were positive in this strain of Enterobacter cloacae.blaNDM-1 gene and blaKPC-2 gene were detected by PCR, and further confirmed by sequencing.blaNDM-1 gene was carried by IncX plasmid with 54×103 bp, KPC-2 gene was carried by untyping plasmid with 42×103 bp.The strain was only sensitive to tetracycline (MIC=2 μg/mL) and tigecycline (MIC=1 μg/mL).The symptoms were improved after the patient was treated by tigecycline combined with Piperacillin/Tazobactam.Conclusion blaNDM-1 and blaKPC-2 genes in Enterobacter cloacae can be mediated by plasmids, and appropriate therapy for its infection should be based on the result of antibiotic susceptibility test.