Objective To develop a colloidal gold immunochromatographic test strip for rapid and accurate detection of Pseudomonas aeruginosa(P.aeruginosa,Pa).Methods After bioinformatics analysis of Pa outer membrane protein OprF,the gene sequence with abundant antigenic determinants and high intraspecific homology was chemically synthesized,and then connected to pET-28a(+)vector to construct the expression vector pET-28a-OprF,which was transformed into E.coli BL21(DE3)and induced by IPTG. The recombinant OprF protein was purified by Ni Sepharose~(TM)6 Fast Flow and used to immunize two female BALB/c mice for 3～4 times by multi-point subcutaneous injection in the back at the first immunization and intraperitoneal injection at subsequent immunizations. The monoclonal antibodies were screened by animal cell fusion technique,and the colloidal gold immunochromatographic test strip for rapid detection of Pa was prepared by using monoclonal antibody and double antibody sandwich immunochromatography technique. The specificity,sensitivity and stability of the test strip were evaluated.Results Two monoclonal antibodies,Pa-1# and Pa-2#,were obtained with the titer of 1∶409 600,and both of them recognized OprF specifically. The prepared colloidal gold immunochromatographic test strip showed a sensitivity of 1. 0×10~6CFU/mL and had no cross reaction with 9 common respiratory pathogens with a good stability.Conclusion The prepared colloidal gold immunochromatographic test strip can detect Pa rapidly within 15 min,with high specificity and good stability.

BACKGROUND:Mesenchymal stem cels have the specific chemotaxis to the inflammation and tumor tissue, but the effect of mesenchymal stem cels on the growth of cervical cancer cels becomes an urgent problem. OBJECTIVE:To investigate the effect of human umbilical cord mesenchymal stem cels on the proliferation of cervical cancer Hela cels. METHODS:Hela cels were co-cultured with human umbilical cord mesenchymal stem cels at different number or its conditioned medium at different concentrations for 3 days. Then, cel counting kit-8 was used to detect the proliferation of Hela cels. RESULTS AND CONCLUSION:When Hela cels were co-cultured with human umbilical cord mesenchymal stem cels at ratios of 1:3, 1:2, 1:1, 2:1, 4:1, 8:1, the relative proliferation inhibition rates were 67.12%, 47.18%, 31.15%, 27.61%, 15.55% and 15.95%, respectively. When the Hela cels were co-cultured with human umbilical cord mesenchymal stem cel conditioned medium at 5, 10, 20, 40, 60, 100 mg/L, the relative proliferation inhibition rates were 0.61%, 40.1%, 63.47%, 80.61%, 93.56%, 90.65%, respectively. These findings indicate that the proliferation of Hela cels can be inhibited by co-culture with human umbilical cord mesenchymal stem cels at a certain concentration-dependent manner.

Objective To analyze the clinical value of chemotherapy combined with endocine therapy after standard treatment failure for advanced metastatic breast cancer.Methods 30 metastatic breast cancer patients after standard treatment failure were analyzed.Etoposide (75-100 mg/d) wasused on days 1-10,followed by 11 days of rest combined with medroxyprogesterone 0.5 g,twice per day,or megestrol 160 mg/d for 21 days.Clinical effects and life quility were analysed.Results The median treatment line of this therapy was 6 (range 3-9).The clinical benefit rate is 16.7 ％ (5/30),and the median progression free survival (PFS) was 4.0 months (range 1.0-13.0 months).Conclusion The combination of chemotherapy (etoposide) and endocrine therapy (progesterone) is a choice of treatment after standard drug failure for advanced mastatic breast cancer patients.

Background To study the pathogenesis and management of diabetic retinopathy (DR) has an important clinical significance.With the development of biomedical photonics in recent years,photobiomodulation therapy has been paid more and more attention.However,the sudy on biological regulation of light to DR is rarely reported.Objective This study was to explore the photobiomodulating effects on the apoptosis of retinal neuronal cells induced by high glucose environment and tried to offer a basis for the management of DR.Methods The retinal neurons were isolated from Wistar rats using immunomagnetic beads and primarily cultured in Neurobasal,and the cells were identified by Nissl staining.The cells were divided into normal control group,high-glucose group and high-glucose+LED group.The glucose at the concentration of 25 mmol/L was added into medium for 48 hours in the high-glucose group,and the cells induced by high-glucose were irradiated in incubator by LED for consecutive 300 seconds per time in a 12-hour interval with the wavelength of 620 nm,maximal power of 1 W,central light radiation exposure of 6.67 mW/cm2 and spot diameter of 2.0 cm.The apoptosis rate of the cells was assayed by flow cytometry;the intracellular Ca2+ content was determined by laser scanning confocal microscope;the relative expression level of phosphorylated serine-threonine kinase (p-AKT) protein in the cells was detected by Western blot.Results The cells grew well 2-3 days after cultured with the polygon and oval shape,and nucleolus were visible.More neuronal processes were obtained in 5-7 days after culture.Nissl staining showed the blue violet color in cytoplasma of neurons.The proportion of neurons and glial cells was 91％.The apoptosis rates of the cells were (7.634±3.176)％,(33.642 ±9.315)％ and (23.914±6.375)％ in the normal control group,high-glucose group and high-glucose+LED group,respectively,and the apoptosis rates of high-glucose group and high-glucose + LED group were significantly higher than that in the normal control group,while the apoptosis rate in the high-glucose+LED group was lower than that in the high-glucose group (all at P＜0.01).The fluorescence of Ca2+ in the cytoplasma was strong in the highglucose group and weak in the normal control group.The fluorescence pixel values in the high-glucose group and highglucose+LED group were significantly higher than that in the normal control group,and that in the high-glucose+LED group was reduced in comparison with high-glucose group (all at P＜0.05).The expressing band of p-AKT protein was strong in the normal control group and weak in the high-glucose group.The relative expressing levels were 10.34± 3.18,2.16±0.46 and 7.15 ±1.72 in the normal control group,high-glucose group and high-glucose+LED group,and relative expression level of high-glucose+LED group was significatly lower than that in the high-glucose group (P＜ 0.05).Conclusions High-glucose environment inhibits PI3K/AKT pathway and calcium homeostasis of retinal neurons,which results in cell apoptosis.Low intensity of LED light irradiation activates the anti-apoptotic PI3K/AKT pathway and therefore reduces apoptosis induced by high glucose.

Objective To analyze the clinic characteristics, lifetime and prognostic factors of young female breast cancer patients. MethodsClinical data of 155 patients under 35 years of age with breast cancer were retrospectively reviewed and followed up.ResultsThe positive rate of hormone receptors was 61.6 % (77/125) in all cases who had been detected receptor status. The median survival time in hormone receptors positive and negative group were 119.0 and 51.3 months (P＜0.01), and 5-year survival rates were 68 % and 33 %, respectively. For patients who had been treated with adjuvant tamoxifen (47.1%), the median survival time was 182 months which longer than without tamoxifen (P ＜0.05). The median disease-free survival time and median survival time were 24 and 91 months in all cases. The overall 3-, 5- and 10-year survival rates were 79 %, 60 % and 51%, respectively. Multifactor analysis with the COX model indicated that tumor size, axillary metastatic status, tamoxifen treatment and overexpression of Her-2 were independent prognostic factors. While clinic stage and hormone receptors status might be referenced prognostic factors. ConclusionYoung women breast cancer patient may have good prognosis if multimodality treatment is conducted. Tumor size, axillary metastatic status, adjuvant endocrine therapy and overexpression of Her-2 are independent prognostic factors.

Objective To detect the expression of Toll-like receptor 9 (TLR9) in pancreatic cancer,and to explore its clinical significance. Methods The real-time PCR, Western blotting and immunohistochemistry were used to examine the expression of TLR9 in 30 samples of pancreatic cancer and the adjacent tissues, and 10 samples of normal pancreatic tissues. The relationships of TLR9 with clinicopathological parameters were analyzed. Results Compared with normal pancreatic tissues, the amplification value of TLR9 mRNA expression was 2.32 fold (1.41 ～ 3.22 ) in human pancreatic cancer, was 1.23 fold (1.18 ～ 1.28) in paracancerous tissues, respectively, and the difference was statistically significant (P ＜ 0.05 ). The percentage of positive cells expressing TLR9 protein in human pancreatic cancer tissues,paracancerous tissues and normal tissues were 73.3% (22/30), 33.3% (10/30) and 20.0% (2/10), and the relative expressions of TLR9 protein were 0.780 ±0. 026,0. 400 ±0.018,0. 173 ±0.043 ,respectively, and the difference was statistically significant( P ＜ 0.05 ). The highly expressed TLR9 was positively correlated with the degree of tumor differentiation, TNM stage and lymph node metastasis. Conclusions TLR9 was highly expressed in pancreatic cancer tissues. TLR9 expression plays a role in the canceration of pancreatic tissues.

Objective To detect the expression of Toll-like receptor 9 (TLR9) in pancreatic cancer and to study the effect of CPG ODN2216 on the biological behavior of pancreatic cell carcinoma, and to explore their clinical significance. Methods Immunohistochemical method was used to examine the expression of TLR9 protein in pancreatic cancer tissue and immunofluorescence staining was also performed to detect TLR9 protein expression in pancreatic carcinoma cells. In vitro cell adhesion, wound-healing scrape assay, transwell invasion assay and cell colony formation assay were performed to assess the effect of CPG ODN2216 on the invasive properties of Panc-1 cells. Results TLR9 were highly expressed in the pancreatic cancer tissue and pancreatic carcinoma cells. In vitro experiments as cell spreading assays, cell adhesion, colony formation assay and invasion assays showed the cell adhesion and cell motility properties of CPG ODN 2216 group to be apparently weakened compared with the control group. MTT assay showed cell proliferation ability in the CPG ODN group to be notably decreased, and CPG ODN2216 had inhibitive effects on the growth of panc-1 cells in a dose and time-dependent manner. Conclusions TLR9 gene was correlated with the invasive and metastatic potentials of pancreatic carcinoma. The used of CPG ODN2216 induced the inhibition of migration and invasion of the Panc-1 cell line.

Objective To analysis the relationships between bone markers, bone-specific alkaline phosphatase (BAP) and cross-linked telopeptide of type Ⅰ collage (ICTP), and bone metastasis of breast cancer.Methods A total of 217 patients' serum were collected.The 217 cases were divided into two groups:109 cases with bone metastasis, 108 cases without bone metastasis. Serum BAP and ICTP was measured by ELISA. The relationships between factors of bone metastasis and serum levels of BAP, ICTP were analyzed.Results The levels of serum BAP and ICTP in bone metastases group were significantly higher than those in non-bone metastasis group[BAP:24.8 μg/L(7.60-213.70 μg/L) vs 21.2 μg/L(7.3～68.8 μg/L),ICTP:7.0μg/L(1.4～32.4 μg/L) vs 4.1 μg/L(0.0～15.8 μg/L) (P=0.003,P=0.000)].The level of serum BAP and ICTP in patients with multiple bone metastasis was significantly higher than that in patients with single bone metastasis[BAP:32.3 μg/L(9.A～213.7 μg/L) vs 18.1 μg/L(7.6～60.0 μg/L),ICTP:7.6 μg/L(1.4～32.4 μg/L) vs 4.9 μg/L(1.8～10.5 μg/L),(P=0.001,P=0.010)].The sensibility of BAP and ICTP was 45.0 ％ (49/109)and 46.8 ％ (51/109),respectively.The specificity of ICTP and BAP was 83.3 ％ (90/108)and 84.3 ％ (91/108),respectively.Joint detection of BAP and ICTP had improved sensibility in the diagnosis of bone metastasis in breast cancer patients. Conclusion Joint detection of serum bone biochemical markers ICTP and BAP have a little values for diagnosing bone metastasis in breast cancer patients.

Objective To evaluate the correlation of the clinical effects and prognosis in patients receiving medical ovarian suppression (goserelin)combined with anastrozole treatment with premenopausal metastatic breast cancer.Methods 44 hormone dependent mastatic breast cancer patients were treated by goserelin,3.6mg hypodermic injection every 28 days and anastrozole 1 mg were administered orally,clinical effects and prognosis were analysed.Results The clinical benefit rates of goserelin combination with anastrozole in patients with metastatic breast cancer were 52.4 ％(23/44),and the median progression free survival (PFS)was 8.3(5.3-11.2)months.In the analysis of whether to accept chemotherapy,the PFS of the not received chemotherapy group was better than received chemotherapy group (16.9 months vs 5.8 months P=0.048).Conclusion The combination of goserelin and anastrozole is an effective endocrine therapy regiment for patients with premenopausal metastatic breast cancer.It can be recommended for the premenopausal and hormone dependent mastatic breast cancer patients.