OBJECTIVES: To evaluate the protective effect of Schistosoma japonicum cystatin (rSj-Cystatin) in a mouse mode of "two-hit" sepsis. METHODS: Sixty male C57BL/6 mice randomized equally into sham-operated group, protein group, "two-hit" modeling group, and protein intervention group. In the former two groups, the mice received an intraperitoneal injection of 100 μL PBS followed by exposure of the cecum and then by intraperitoneal injection of 100 μL PBS or 25 μg rSj-Cystatin 30 min later; In the latter two groups, 100 μL PBS containing LPS (5 mg/kg) was injected intraperitoneally 24 h before cecal ligation and puncture (CLP), and 100 μL PBS or 25 μg rSj-Cystatin were injected 30 min after CLP. At 12 h after rSj-Cystatin treatment, 6 mice from each group were sacrificed for detection of TNF-α, IL-6, IL-10, TGF-β, iNOS and Arg-1 in the serum, spleen, liver, lung and kidney tissues using ELISA, for examinations of liver, lung and kidney pathologies with HE staining, and for analysis of CD3⁺CD4⁺CD25⁺Foxp3⁺ T cell percentage in the spleen using flow cytometry. The remaining mice were observed for general condition and 72-h survival. RESULTS: The 72-h survival rates in the 4 groups were 100%, 100%, 0% and 20%, respectively, showing significant differences between the latter two groups. The mouse models of "two-hit" sepsis exhibited obvious tissue pathologies and significant elevations of TNF-α and IL-6 in both the serum and tissue homogenate, which were significantly ameliorated by rSj-Cystatin treatment. Treatment with rSj-Cystatin also increased IL-10 and TGF-β levels and spleen CD3⁺CD4⁺CD25⁺Foxp3⁺ T cell percentage. The septic mouse models also showed increased iNOS levels in all the detected tissues and a decreased Arg-1 level in the kidney, and these changes were obviously improved by rSj-Cystatin treatment. CONCLUSIONS rSj-Cystatin has a protective effect against "two-hit" sepsis in mice by regulating the inflammatory microenvironment.
Animals ; Mice ; Sepsis/drug therapy* ; Male ; Schistosoma japonicum/chemistry* ; Mice, Inbred C57BL ; Cystatins/therapeutic use* ; Interleukin-10/metabolism* ; Interleukin-6/blood* ; Tumor Necrosis Factor-alpha/blood* ; Disease Models, Animal ; Transforming Growth Factor beta/metabolism*

Objective: To evaluate the analgesic effect of pregabalin combined with intrathecal sufentanil infusion for the treatment of breakthrough pain in patients with bone metastases. Methods: A total of 60 breakthrough pain patients with bone metastases were randomly divided to 3 groups: group A (pregabalin combined with intrathecal sufentanil infusion group,n=20), group B (placebo combined with intrathecal sufentanil infusion group,n=20) and group C (oral morphine sulfate controlled-release tablet group,n=20). The differences in visual analogue scale (VAS)between background pain and breakthrough pain, the seizure frequency of breakthrough pain, general satisfaction and side effects of the 3 groups were observed. Results: hTe seizure frequency and VAS of breakthrough pain in group A decreased signiifcantly after the treatment (P<0.05) and the general satisfaction was the best among the the 3 groups (P<0.05), with less nausea and vomiting, constipation, drowsiness and fewer other side effects. Conclusion: Pregabalin combined with intrathecal sufentanil infusion can effectively relieve breakthrough pain in patients with bone metastases.

Objective To study the technique of transposon mutagenesis involving the use of Mini-Tn5 on a suicide plasmid for Citrobacter freundii (C. freundii). Methods A conjugation method was used in transposon mutagenesis. The stability of exconjugants was confirmed by series passages under nonselective condition. Detection of anti-ampicillin resistance and purification of the plasmid were conducted for excluding the possibility of plasmid-encoded antibiotic resistance. All exconjugants were screened for auxotrophic mutants and swarming motility mutants. Results Analysis of the generated exconjugants indicated that Mini-Tn5 Km was transposed in C. freundii at a high frequency (ca. 9.2?10 -6). The mutants were stable and retained the transposon-encoded antibiotic resistance after incubation for three passages under nonselective condition. The frequency of auxotrophic mutants in the population confirmed that the insertion of the transposon was random and the C. freundii chromosome did not contain significant insertional hot spots of transposition. No extrachromosomal plasmid DNA was found. Conclusion Transposon mutagenesis of Mini-Tn5 is a suitable technique for the study of differentiation in C. freundii.

Objective To investigate the periodic variety at the germ form, quantity, flagellar expression and activity of breath enzyme etc during the swarming motility of Proteus mirabilis (PM). Methods The 2, 3, 5-tetraphenyltetrazolium chloride (TTC), as indictor of breath enzyme, was added to the culture medium. Electronic microscope was adopted to observe the germ form and flagellar. Quantity of PM was measured by turbidimetry. Results During the PM growth and migration at LB medium, the germ quantity reduced and form changed from short and small to long and thin, flagellar got more and breath enzyme turned into inactive. Conclusion At the LB medium, PM in growth process appeared varieties in length of germ body, flagellar quantity and germs density, etc. PM in solid medium appeared wave motility, resulting from many factors including the environment, the signal transduction in cells and the density of germ.

A strain of P. Aeruginosa,which was seperated from clinical environment ,shows a special characteristic . It keeps normal short rod shape when cultured at 37℃,however,it forms filament without pyocyanin producing when cultured at 25℃ overnight. The filaments will divide and form short rods,simultaneously,produce pyocyanin when culture time is prolonged to over 72h or culture temperature is raised to 37℃. The preliminary study indicates that this phenomenin has nothing to do with nutritive conditions and could the inbluenced by inoculating density and irradiating with ultraviolet rays The absence of pyocyanin was not the cause of filamentous formation by the test results.

Objective To investigate the correlation between bacterial cryptic growth cell(CGC)and irritable bowel syndrome(IBS).Methods The techniques of micro-culture and morphological observation were employed in examination of the CGC in feces specimens of patients suffering from IBS.Furthermore,the animal monitoring tests was applied in the aspects of CGC formation under inducing factors and colonization at local intestinal mucus.Results A great deal of CGC could be observed in the feces of IBS patients and exhibited continuous proliferation characteristics regardless of using or not using antibiotics.And the CGC in feces showed auto-lysis features when the patients suffered from the disease.The monitoring tests demonstrated that the antibiotics often used presently to IBS patients could induce the formation and colonization of intestinal bacterial CGC.Conclusion Based on the experimental and clinical data,we presume that there is a correlation between intestinal bacterial CGC and IBS.