Objective To investigate the expressions of calcium sensitive receptor (CaSR) and tight junction protein (Claudin)-14 in renal calcium oxalate stone rat model. Methods Thirty male SD rats were randomly divided into control group (n=15) and model group (n=15). The rat model of renal calcium oxalate stone was established by gavaging 1%glycol (2 mL/d) and 2% ammonium chloride. The expression of CaSR protein was detected using immunohistochemical assay. RT-PCR was used to detect the Claudin-14 mRNA expression. The expression levels of CaSR and Claudin-14 protein were de-tected by Western blot assay respectively. The full automatic biochemical analyzer was used to detect rat renal function and changes of blood and urine biochemical indices. Results A large stone crystallization was observed under light microscope in model group. The serum levels of Cr, BUN, 24-h urine calcium and urine volume were significantly higher in model group than those of control group (P<0.01). There were no significant differences in serum calcium level and urinary pH value be-tween two groups. The expression levels of Claudin-14 mRNA and CaSR protein were significantly higher in model group than those of control group (P<0.01). The Claudin-14 protein expression was specifically higher in renal tissues of model group. There was no Claudin-14 protein expression in control group. There was a positive correlation between CaSR and Claudin-14 protein expression in renal tissues of model group. Also, CaSR and Claudin-14 protein expressions were posi-tively correlated with the 24-h urine volume. Conclusion The increased expressions of CaSR and Claudin-14 involved in the formation of kidney stone by increasing the urinary calcium excretion.

Objective To identify the putative CpG-N ODNs in adenovisus 2 DNA (Adv2 DNA) and Adv5 DNA by comparing the sequence difference among Adv2, 5, 12 DNA and E.coli (EC) DNA. Methods Sequences of Adv2, 5, 12 DNA and EC DNA were obtained from the Entrez Nucleotides database at NCBI. The specific CpG motifs of Adv2 DNA and Adv5 DNA were identified after above sequences were analyzed and compared by softwares such as DNATools, BioEdit, and so on. All the 12-ODNs with specific CpG motif core were searched from Adv2 DNA and Adv5 DNA. Results 19 specific CpG motifs were ascertained and 504 12-ODNs were detected in Adv2 DNA and Adv5 DNA. Conclusion 504 12-ODNs were putative CpG-N ODNs in Adv2 DNA and Adv5 DNA.

Objective To obtain a potent bacterial DNA antagonizing CpG oligonucleotides (CpG-N ODN) from the structures of Adv2, 5 DNA. Methods Ten putative CpG-N ODNs were synthesized and investigated. Their abilities to inhibit the TNF-? release from hPBMC were observed. Based on the above results, the putative CpG-N ODN was redesigned according to the relationship between the structure and free energy using RNA structure software (version 3.71). Eleven putative CpG-N ODNs were synthesized and screened. Results Out of the ten initial CpG ODNs, ODN101 was the only CpG-N ODN with weak activity to inhibit TNF-? release from hPBMC induced by CpG-N ODN. After redesign, five CpG-N ODNs with strong activity were confirmed. Conclusion Six CpG-N ODNs were obtained with activity to inhibit TNF-? release from hPBMC induced by CpG-N ODN.

Objective To study the correlation of NF-?B expression with the grade, the stage, the recurrence, the metastasis and PKC expression of bladder transitional cell carcinoma. Methods The expression of NF-?B and PKC was examined with SP immunohistochemical method in 54 cases of paraffin-embedded bladder transitional cell carcinomas tissues and 13 cases of normal bladder tissues. Results The positive expression rate of NF-?B P65 and PKC in bladder transitional cell carcinomas was 88 9% and 81 5% respectively. The expression rate of NF-?B decreased along with pathological grade and clinical stage increased, while its expression increased along with the rate of recurrence and metastasis elevated. There was a positive relationship between NF-?B and PKC expressions. Conclussion NF-?B may play a role at early stage of bladder transitional cell carcinoma. It probably serves as an index to evaluate the malignant extent and progression of bladder transitional cell carcinoma. PKC probably takes part in the activation process of NF-?B.

Objective To investigate the expression level of cyclooxygenase-2 (COX-2) and its effects on cell proliferation and apoptosis in bladder cancer. Methods The immunohistochemical method was used to determine the expression of COX-2 and Ki67 in 54 cases of bladder cancer, 29 cases of adjacent tumor tissue and 10 cases of normal bladder specimen respectively, and the apoptotic levels of the obove three tissues were assessed by counting the number of the cells labeled by TUNEL method. Results The positive rate of COX-2, the mean labelling index of Ki67 and the cellular apoptosis index in the cancer tissues were significantly higher than those in adjacent tumor tissues and normal bladder specimens(P

Objective The calcium oxalate stone is the most common type of the kidney stones.By building the rat renal calcium oxalate stone model, preliminary study the function of CaSR-Claudin14 regulating pathways on renal calcium oxalate stone for-mation model in rats. Methods 30 Male S-D rats were randomly divided into control group (n=15) and model group (n=15). Adult male S-D rats were given ethylene glycol and ammonium chloride to induce urolithiasis.Application of full automatic biochemical analyzer to test rat renal function and the changes of urine biochemical index.Immunohistochemistry was used to detect the expression of CaSR protein;RT-PCR was used to detect the Claudin-14 mRNA expression;Western Blotting was used to detect the expression of CaSR and Claudin-14 protein respectively. Results By observing model group has large stones crystallization under light microsco-py;model group rats 24 h urine calcium are significantly higher than control group([9.66 ±1.10]mmol vs [3.26 ±0.60]mmol, P<0.01); and model group 24 h urine volume are significantly higher than control group ([21.27 ±1.08]mL vs [13.2 ±0.55]mL, P<0.01 ); and urinary PH has no significant difference between the groups( P >0.05 ) .Expression of Claudin-14 mRNA in the model group is significantly higher than normal control group([0.150 ± 0.004] vs [0.047 ±0.008], P<0.01); Expression of Claudin-14 protein in the model group is significantly higher than normal control group([1.526 ±0.089] vs 0, P<0.01).Expression of CaSR protein in the model group is significantly higher than normal control group([6.697 ±0.051] vs [5.016 ±0.053], P<0.05). Conclusion CaSR can raise the expression of Claudin-14, increase re-nal tubular urinary calcium excretion to promote renal calcium oxalate stone formation.

Objective To discuss the types of ICC and characteristics of spontaneous Ca2+ waves of different types of ICC in the bladder of guinea pig. Methods Frozen-sections were made from the bladder of guinea pig and ICC were cultured in vitro. Cells were stained by indirect immunofluorescent method and detected by Laser scanning confocal microscope. The ICC cultured in vitro were divided randomly into 4 group: dimmer ICC,monomer ICC,dimmer ICC treated with 2-APB group and monomer ICC treated with 2-APB group according to the cell morphology and disrupted with 2-APB.The calcium concentration of ICC cultured in vitro were marked with Fluo-4 AM and disrupted by 2-aminoethoxydipheylbrate (2-APB, 100 μmol/L) in dimmer ICC treated with 2-APB group and monomer ICC treated with 2-APB group. The calcium oscillation function of ICC was observed under Laser scanning confocal microscope. Results For the monomer ICC and dimmer ICC in frozen sections and cultured in vitro,there were increased frequency (P＜0.01) and amplitude (P＜0.05) of spontaneous Ca2+ waves in dimmer ICC compared with the monomer monomer ICC. But after the cells disrupted by 2-APB after 15 min,There were decreased frequency (P＜0.01) and amplitude (P＜0.01) of spontaneous Ca2+ waves in the dimmer ICC treated with 2-ABP group compared with the dimmer ICC. The changes(P＞0.05) of spontaneous Ca2+ waves was not statistical significance in monomer ICC treated with 2-ABP group compared with monomer ICC group. Conclusions The bladder of guinea pig may exist 2 different types of ICC, dimmer ICC and monomer ICC. The excitability of spontaneous Ca2+ waves of dimmer ICC could be higher than in monomer ICC. The special structure of dimmer ICC may contribute to the formation of high spontaneous Ca2+ waves.

Objective To observe the effects of green fluorescence protein mediated by lentivirus in bladder, and to de?termine the amount of virus obtained good transfection effects. Methods lentivirus carring GFP gene was perfused using transurethral approach into bladder of guinea pigs. Samples of bladder, liver, kidney and lungs were collected for frozen sec?tions after feeding seven days. The distribution of green fluorescence was observed using laser confocal microscopy. Re?sults The titer of lentivirus was 4 × 108. GFP was found under the mucosa when the amount of lentivirus transurethral was 30μL. GFP was distributed widely in muscle layer using 40μL lentivirus. GFP was detected even stronger in muscle layer when the amount was 50μL. GFP was found in muscle layer when 25μL lentivirus was injected intravenously. GFP was not found in other tissues than in bladder via transurethral perfusion. There was higher GFP in liver, lungs and other organs than in bladder via intravenous injection. Conclusion Lentivirus can mediate GFP transfecting bladder of guinea pig successful?ly and escape the distribution of GFP all over the body intravenously, which will bring new research direction and method for clinical treatment of diseases in bladder.

Objective To study the antimalarial activity of naphthoquine phosphate combined with artemisinine against Plasmodium knowlesi in rhesus monkey.Methods Monkeys were randomly divided into 9 groups(3/group).The monkeys in groups A and B were treated i.g.once daily for 3 days with 6 or 10 mg/kg of naphthoquine phosphate respectively.Those in groups C and D were treated i.g.twice for the 1st day and once for the 2nd and 3rd day with 31.6 or 100 mg/kg of artemisinine respectively.In groups E, F and G, they were treated i.g.only once with the combination of naphthoquine phosphate 10 mg/kg and artemisinine 10, 20 or 25 mg/kg respectively.Groups H and I served as controls which were treated i.g.only once with 10 mg/kg of naphthoquine phosphate and 30 mg/kg of artemisinine respectively.Parasitemia was examined beginning 24 h after drug administration.The observation lasted 105 days when no more parasite was found.Results At 24 h after drug administration, the parasite reduction rate in all groups was higher than 90%.The parasite clearance time for groups E, F and G was(56.0?16.0),(53.3?4.6), and(56.0?8.0) h respectively, more rapid than that of Group H(69.3?4.6) h.There were 1, 3, 3, 2, 2, and 3 monkeys in groups A, B, D, E, F, and G respectively which were cured.No monkeys were cured in groups C, H and I.Conclusion The combination of naphthoquine phosphate and artemisinine is superior to the single component and the optimum proportion in the combination is 1:2.5 in treating P.knowlesi infection in monkeys.

AIM: To determine the expression of c-kit mRNA and protein in the bladders in guinea pigs with diabetic cystopathy (DCP) and to explore the correlation and mechanisms between c-kit expression and DCP. METHODS: Sixty guinea pigs were divided randomly into control group (n=20) and experimental group (n=40). The guinea pigs in experimental group were injected with streptozotocin (STZ) to induce diabetes mellitus. After fed for 10 weeks, the animals in both groups were tested with urodynamics, and the guinea pigs in experimental group were divided into the subgroups of DCP and the diabetic no-cystopathy (NDCP) group according to the results of urodynamics. mRNA expression of c-kit was detected by reverse transcription polymerase chain reaction (RT-PCR) and protein expression of c-kit was tested and analyzed by laser scanning confocal microscope. RESULTS: Decreased expression of c-kit mRNA was observed in DCP group compared to control and the NDCP group. The ratio of c-kit mRNA and GAPDH was 5.66±0.54 in controls (P<0.05), 5.54±1.28 in NDCP group (P<0.05) and 4.65 ±0.47 in DCP group. c-kit protein expression significantly declined in DCP group. The mean value of fluorescence intensity was 856.52± 53.03 in control group, 844.67± 59.24 in NDCP group and 548.69± 48.51 in DCP (P<0.01).CONCLUSION: The declined expression of c-kit) gene at transcription and translation levels destroys the SCF/c-kit signal pathway, leading to the dysfunction of Cajal-like) cells in DCP guinea pig, so the abnormal expression of c-kit gene is involved in the pathogenesis of DCP.