Objective: To investigate the molecular changes in bladder urothelial carcinoma via different pathways. Methods:Polymerase-chain reaction (PCR) or coamplification at low denaturation temperature-PCR and Sanger direct sequencing were per-formed to detect the status of fgfr3, p53, and h-ras gene mutations in 88 tissue samples of human bladder cancer and 10 normal control tissues. The relative mRNA expression levels of motility-related protein-1 (MRP-1)/CD9 and the relationship between genes and tumor recurrence were also determined. Logistic regression and relative analyses were conducted to compare the significance and interrelation of genes among tumor recurrences. Results:The mutation rate of p53 increased as pathological grades and stages increased. Recurrence rate was higher in patients with MT-p53 genotype than in patients with WT-p53 genotype. Conversely, the mutation rate of fgfr3 gene decreased as pathological grades and stages increased. Recurrence rate was also higher in patients with WT-fgfr3 genotype than in pa-tients with MT-fgfr3 genotype. In low-grade and early stage tumors, MT-fgfr3/WT-p53 was the most prevalent genotype;in high-grade and late stage tumors, WT-fgfr3/MT-p53 was the most prevalent genotype. The mutations of h-ras were mainly observed in low-grade tumors in early stages. Moreover, the relative mRNA levels of MRP-1/CD9 decreased as pathological grades and stages increased. The mRNA levels of MRP-1/CD9 were negatively correlated with p53 mutations and positively correlated with fgfr3 mutations. Logistic re-gression analysis results showed that patients with WT-fgfr3 genotypes exhibited 3.88 times higher relative risk of tumor recurrence than those with MT-fgfr3 genotypes;by contrast, patients with MT-p53 genotypes exhibited 4.53 times higher relative risk of tumor re-currence than those with WT-p53 genotypes. Conclusion:Fgfr3 and h-ras gene mutations may play important roles in tumorigenesis of low-grade and early stage bladder cancer. p53 gene mutation and mRNA levels of MRP-1/CD9 may be implicated in the tumorigenesis of high-grade tumors in late stage of bladder cancer. In general, the two variants of urothelial carcinoma exhibit distinct genetic defects. fgfr3 gene mutation revealed a pathway of favorable prognosis, and p53 gene mutation demonstrated a pathway associated with poor prognosis.

Micro-ribonucleic acids (miRNAs) are endogenous single-stranded small non-coding RNAs. miRNAs bind to a com-plementary site in the 3' untranslated region of their target mRNAs through canonical base pairing, which can direct the degradation or translational repression of these transcripts. Thus, miRNAs can effectively silence the protein expression of target genes post-transcrip-tionally. miRNAs may also regulate the expression of oncogenes and tumor suppressor genes and could be involved in almost all known hallmarks of cancinogenesis. In this paper, we discuss the following in detail:(1) biogenesis and main functions of cellular miR-NAs, (2) stability and detectability of exosomal miRNAs in biological fluids;and (3) feasibility of miRNAs as a potential new class of biomarkers derived from urinary exosome in the malignancy of urinary system. Finally, we summarize studies on urinary exosomal miRNAs as potential biomarkers of prostate, bladder, and kidney cancers.

By using of combined HRP retrograde tracing and immunocytoche mistry methods, the projection from the caudal part of the ventrolateral medulla (VLM) to the nucleus accumbens (Acb) was examined. When HRP was injected into the ventral and medial area of the caudal part of Acb, the labeled cell bodies were mostly found in the bilateral (predominantly ipsilateral) caudal part of the VLM. When HRP technique (HRP injected into the Acb)was combined with immunocytochemical method, many HRP-TH, HRP-NT and HRP-CCK double labeled cell bodies were found in the VLM. The number of the HRP-TH double labeled cell bodies were numerous, while HRP-NT and HRP-CCK doublelabeled cells were less. HRP-TH double labeled neurons were also found in the reticular formation between the solitary tract nucleus and VLM.

The projection from the nucleus of solitary tract (Sol) to the nucleus accumbens (Acb) was examined by using HRP retrograde and anterograde tracing combined with immunocytochemical double-labeling technique. The following results were obtained: (1) when WGA-HRP was injected into the caudal part of the Sol, the labeled fibers, terminals and cell bodies were found in the ventromedial area of the caudal Acb. After injecting HRP into the ventromedial area of the caudal Acb, labeled cell bodies and terminals were found in the ipsilateral and contralateral caudal part of the Sol,mainly in the commissure nucleus and the medial subnucleus of nucleus of solitary tract. (2) After injecting HRP into the Acb and combined with immunocytochemical method, many HRP-TH, HRP-NT, and HRP-CCK double-labeled cell bodies were found in the caudal part of the Sol. The number of the HRP-TH double-labeled cell bodies was most numerous, HRP-NT cells was next and HRP-CCK cells was even less.

The lymphonodus cervicothoracious and lumbaris of 19 Pekin ducks were studied using the histological methods. The lymph nodes are consisted of the capsule, loose areas, and dense areas. There are lymphatic tissue cords, connective tissue cords and intercord lymph sinus and lymphatic nodules also in the loose areas. The dense area are consisted of lymphatic nodules and mass of lymphoid cells. The lymphatic nodules are irregularly distributed and no germinal centers. The lymph sinuses may be classified into the central sinus and peripheral sinus. The latter may be subdivided into subcapsule sinus, paralymphonodular sinus and the intercord lymph sinus. There are reticular cells in the intercord lymph sinus. No true trabeculae are present but there are many blood vessels and nerves in the lymph nodules.

Objective To explore the heme oxygenase-1 (HO-1) content of carbon monoxide (CO) levels and clinical significance of Mycoplasma pneumoniae infection in children.Methods 219 cases of mycoplasma pneumonia from October 2013 to June 2015 in our hospital were divided into two groups,which were non-asthmatic group (156 cases) and wheezing group (63 cases),and the wheezing group were divided into two groups according to the presence or absence of hypoxemia and mild asthmatic group (39 cases) and severe asthmatic group (24 cases).HO-1 content in the blood was measured by enzyme-linked immunosorbent assay,using dual-wavelength spectrophotometric determination of hemoglobin oxygen percentage of broken (COHb％),analysis of the differences between the groups.Results Children with blood COHb content wheezing group were (2.59 ± 0.40)％ and HO-1 levels were (1 813.24 ± 28.34)ng/L and non-respite group were significantly increased,it had significant differences between the two groups (P＜ 0.05).Group of children with severe wheezing COHb content were (3.63 ± 0.45)％ and HO-1 levels were (2 594.34±23.94)ng/L and mild wheezing group were significantly increased,it had significant differences between the two groups (P＜0.05).Positive correlation between blood COHb content and HO-1 levels were significantly (r=0.733,P＜0.05).Conclusion Mycoplasma pneumoniae infection in children with wheezing blood COHb content and HO-1 levels were significantly increased,and with wheezing exacerbation and showed an increasing trend,suggesting that CO and HO-1 may be involved in Mycoplasma pneumoniae infection in children in the pathogenesis of wheezing,should lead to clinical attention.

Objective To investigate the effects of Chinese medicine HUOXUESHENGJI(promoting blood flow and tissue regeneration) on the oxidative stress in rat skeletal muscles after acute contusion.Methods Seventy two male SD rats were randomly and equally divided into three groups:(1)Chinese medicine HUOXUESHENGJI group(A),(2)physiological saline group(B),(3)natural healing group(C),and(4)blank control group(D).The muscle contusion was produced at the gastrocnemius by specific device;Local injection at the injured site with Chinese medicine HUOXUESHENGJI in group A;local injection with physiological saline instead of HUOXUESHENGJI in group B,and natural healing group served as control.The muscles were sampled at day 1,4,7 and 14 respectively after contusion,and then the MDA and the activities of antioxidant enzymes(CAT,SOD and GSH-px)in gastrocnemius were investigated.Results As compared with the group B and group C,SOD and GSH-px activities in group A were significantly increased after contusion(P0.05).The con-centrations of MDA in group A were lower than that in group B and group C at day 4 and day 7 after contusion(P

Objective To analyze the features of onset, clinical pathological characteristics about the primary bladder mucinous adenocarcinoma.Methods From January 1990 to June 2015, we retrospectively reviewed the data from 15 patients diagnosed as primary bladder mucinous adenocarcinoma, including 10 male patients and 5 female patients.Their mean age was 58 years old, ranged from 41 to 78 years old.Among the fifteen patients, the initial symptoms included hematuria in 13 cases, lower abdominal pain in 1 case and urinary irritation symptom in 1 case.The ultrasound and CT scan revealed bladder tumors, which the size ranged from 2 to 6 cm.The location of bladder tumors included front wall in 12 cases, trigone zone in 2 cases and top wall in 1 case.Nine cases was suspected as tumor from urachal remnants.Eleven patients underwent partial cystectomy, three patients accepted the radical cystectomy and one case accepted transurethral resection of bladder tumor (TURBT).Result Pathological diagnosis was bladder mucinous adenocarcinoma in all patients, including nine from urachal remnant and the others from urothelimn.The tumor exhibited the mushroom liked prominence, which was associated with surface ulceration and infiltrated into the depth of bladder.Meanwhile, it was covered with thick mucinous substances.The histologic examination revealed the presence of andenoid structure, composed by various degree of diferenfiated mucinous cells.In cases with adenocarcinomas from urachus, the residue of urachal tissue could be noticed.The bladder mucous was intact or ulcerated.No sign of metaplasia was observed.In the pathological diagnosis, the classification included grade Ⅲ in 3 cases, grade Ⅱ in 7 cases and grade Ⅰ in 5 cases.Ten persons reported the information of the follow up.Eight of them, whose tumor originated from urachus, accepted bladder-sparing surgery.One died from acute myocardial infarction after 23 months postoperatively.And one died from cerebral hemorrhage 45 months postoperatively.The others have been followed up from 8 to 65 months with no sign of recurrence.In two cases with urothelial carcinoma, one was found the new urothelial carcinoma 50 months after TURBT and one died from cancer metastasis 29 months after partial cystectomy.Conclusions Primary mucinous adenocarcinoma of the bladder possess the relatively high malignant tendency.The hematuria is the main initial symptoms.The histologic examination revealed the presence of different differentiated mucinous cells and formed the andenoid structure.The case with urachal remnant adenocarcinoma has the better prognosis than other types.

Objective To study the biomarker panel of superficial bladder transitional cell carcinoma(SBTCC)and analyze the biological pathway in tumorigenesis by Shotgun proteomics strategy.Methods Normal urothelium cells and cancer cells were harvested by laser capture microdissection from clinical specimen and the proteomic expression profile was identified by two-dimensional liquid chromatography tandem mass spectrometry.The isoelectric point,molecular weight,grand average of hydropathicity,transmembrane helices were analyzed by using proteomics tools.Gene ontology was used to comment the identified proteins.The pathway analysis was performed by ArrayTrack software,and visualized by GenMAPP.Results There were 440 and 218 proteins expressed in cancer cells and normal cells respectively,among them 388 proteins were differerntially expressed.All the database about identified proteins was deposited in an accessible form to researchers at http://www.Proteome-SBTCC.org.cn and http://www.Proteome-NHTE.org.cn.There were 267(68.8%)differentially expressed proteins which had GO biological process comments.The biological pathwavs of these proteins included MAPK signaling pathway,focal adhesion,oxidative phosphorylation,ECMreceptor interaction,etc.Conclusion Shotgun strategy proteomies database of normal transitional epithelium and SBTCC is successfully constructed.And the basis for the understanding of cell biology and discovery of biomarker panel for SBTCC iS provided.

Objective To construct a naive human Fab fragment phage display library,from which the anti-gp96 antibodies may be panned by the gp96 purified from the tissue of urothelial carcinoma in the urinary bladder and provide a basis to new therapy for the malignant tumors.Methods Peripheral blood lymphocytes were isolated from 800 ml of blood,which was obtained from four healthy blood donors.The heavy chain Fd and light chain cDNA synthesized from the total RNA of lympbocytes were amplified by PCR with variable regions 5' and 3' primers of heavy and light chain, and the amplification products were ligated into the phagemid vector pComb3, then the ligated sample was transformed into competent E.coli XL1-Blue by electroperation.The transformed cells were infected with VCSM13 helper phage to yield recombinant phage antibody Fabs.The phagemids abstracted from amplified E.coli were cut with endonucleases such as Sac Ⅰ,Xba Ⅰ,Xho Ⅰ and Spe Ⅰ, and both the phage antibody Fabs and phage-raids abstracted from amplified E.coil were amplified by PCR to monitor the insertion of the genes of light chain or heavy chain Fd fragment.The gp96 purified from the urothelial carcinoma tissue of the bladder by affinity chromatog-raphy on eoncanavalin-A sephnrose and DEAE-sephnrose ion exchange chromatography were utilized as antigens to process three rounds of panning to the original Fab antibody library.Results The quantity of total RNA and cDNA were qualified.By combination of light chain and heavy chain genes, an antibody library containing 6.6×106 clones was obtained, and both the cutting of enzymes and PCR showed that there were the genes of light chain or heavy chain Fd fragment in the phagemids.The gp96 protein was obtained from urothelial carcinoma tissue in the urinary bladder.After having been panned by gp96, the original antibody library gained enrichment by 68 times.Conclusion Utilizing the technology of phage surface display, specific antibody can be gained from the human naive Fab phage display library,which can be used for immunological therapy for tumors.