Ocular surface tissues are vulnerable to various stimuli from the external environment,including microbial invasion,mechanical damage,chemical stimulation,dust,UV light,and allergen stimulation.Conjunctival goblet cells play an important role in maintaining ocular surface self-homeostasis and health.These cells are the only unicellular glands in humans and mammals that are mainly distributed in the epithelium of the digestive tract,respiratory tract,and the ocular surface.Many factors affect goblet cell development and differentiation,especially the newly discovered molecule Spdef,which is a transcription factor found in the ETS family.The main function of conjunctival goblet cells is to secrete mucin to protect and lubricate the ocular surface;however,they also secrete a variety of other bioactive substances.Recent studies have demonstrated that goblet cells are involved in antigen presentation,regulation of dendritic cell phenotype differentiation,and induction of immune tolerance on the mucosa.Many ocular surface diseases,such as dry eye,will alter density and function of conjunctival goblet cells.Therefore,ophthalmologists should be concerned about the biological behavior of conjunctival goblet cells and the relationship between them and ocular surface diseases.

Objective: To obtain the 12-mer phage clones displaying the Hantaan virus mimic epitopes.Methods: HFRSV-positive sera were used as selective molecules for biopanning.A 12-mer phage peptide library was bio-panned for 5 rounds,and the result was confirmed by sandwich ELISA,competition ELISA and DNA sequencing.Results: After 5 rounds of effective screening,the results detected by sandwich ELISA and competition ELISA showed that the majority of the selected clones could react to positive sera in a dose-dependent manner,but could not bind to BSA and the control sera.Sequencing and alignment analyses indicated that amino acid sequences of 45 positive clones fell into 8 groups,and 7 of them exhibited putative motifs: LVXKR,LTXR,IXKP,LXPA,VGA,KXIR and EKXP.Four of the putative motifs had a homologous region within the structural proteins of HFRSV.Conclusion: The peptides displayed by the phage can mimic the epitopes of HFRSV antigens,which provides the potential for preparing more effective epitope-based vaccines and specific diagnostic reagents.

Objective: To clone,identify and express the Shigalike toxin 2B(Stx2B) subunit gene of EHEC O157∶H7.Methods: A pair of primers were designed based on the Stx2B subunit gene sequence of EHEC O157∶H7.The Stx2B gene was amplified from the EHEC O157∶H7 chromosome by PCR and cloned into the pMD18-T vector.Thereafter,the gene was cut from the pMD18-T vector and cloned into the prokaryotic expression plasmid pET-28a vector.Then the recombinant plasmids were transformed into E. coli BL21(DE3) and the transformed host strain induced by IPTG.The expression protein was detected by SDS-PAGE and Western-blot analyses.Results: The Stx2B gene was successfully cloned into pMD18-T and pET-28a vectors,and the expressed protein identified by SDS-PAGE and Western blot.The molecular mass(Mr) of the expressed product was about 7 500,and the expression rate about 40%. Conclusion: The Stx2B gene was successfully cloned and effectively expressed in the prokaryotic expression system.

Objective To observation the effect of treating wind-cold type periarthritis of shoulder with acupuncture with warmed needle. Methods Study the influence of acupuncture with warmed needle on clinical indexes as total therapeutic effect, pain, and range of activity of 60 patients with wind-cold type periarthritis of shoulder. Result The total effective rate was over 80%. Conclusion acupuncture with wanned needle is a good method in treating wind-cold type periarthritis of shoulder clinically.

OBJECTIVE To compare and analyze the drug resistance of Chryseobacterium meningosepticum which producing metallo-?-lactamase(MBL) and extended-spectrum-?-lactamases(ESBLs) from ICU patients′ and non ICU patients′ specimens of sputa so as to guide the rational application of antibiotics.METHODS Identified the strains with VITEK 32,MBL and ESBLs were also screened by double disk synergy;the antimicrobial sensitivity of clinical isolates was tested by VITEK GNS143 and the antimicrobial sensitivity was added and tested by K-B method.RESULTS As a result the rate of producing MBL of C.meningosepticum was 49.0% from ICU patients′ specimens of sputa which was higher than the rate of 13.8% from non ICU patients′;the rate of producing ESBLs of C.meningosepticum was 37.2% from ICU patients′ specimens of sputa which was higher than the rate of 30.6% from non ICU patients′;the rate of drug resistance to AMP/SUB,TZP,CEP,CIP,and LEV from ICU patients′ was higher than that from non ICU patients.CONCLUSIONS Why the high resistance rate of C.mengingosepticum in ICU patients′ specimens of sputa might be due to the high producing ?-lactamases(MBL and ESBLs).

The mrp gene of SS2 human strain Habb was truncated and cloned into a prokaryotic expression vector pGEX4T-2,and a fusion-expressed protein MRP-GST of 61 000 was obtained in E.coli.The GST was cut from MRP-GST with thrombin protease to gain the purified MRP of 35 000 which showed a strong reaction to the SS2 positive sera in Western blotting.BALB/c mice were immunitied intraperitoneally with purified MRP protein.Murine myeloma cells were fused with the splenocytes of the immunized mice after the third immunization.An indirect ELISA coated with purified MRP was used to screen hybridomas for production of specific antibody.The six McAb recognized MRP specially.According to the results of 71 standard strains (48 SS and 34 SS2)by Sandwich ELISA,the coincidence rate was 97.2%,but for 34 SS standard serotype was 100 %.The ELISA method has a potential value for clinical and epidemiological applicationgs.

Objective:To observe the influence of multi-disciplinary cooperation and optimization management mode on the negative situation and rehabilitation effect of patients with affective disorder after coronary heart disease and cerebrovascular disease.Methods:A total of 188 patients with emotional disorders caused by coronary heart disease and cerebrovascular disease from January 2017 to December 2018 to our hospital were selected. According to the random number method, 94 cases in the control group adopted the conventional management model, and 94 cases in the experimental group adopted the multidisciplinary collaborative optimization management model. The negative emotions [evaluated by Self-rating Anxiety Scale (SAS), Self-rating Depression Scale(SDS) ] scores before and after management, the coping style (Jalowies) scores, and quality of life [evaluated by World Health Organization Quality of Life-Brief (WHOQOL-BREF) ] scores were compared between the two groups. The clinical rehabilitation effect and satisfaction rate of related management between the two groups were compared.Results:There were no significant differences between the two groups before management of SAS scores, SDS scores, Jalowies scores, and WHOQOL-BREF quality of life scores ( P> 0.05). After management, the SAS scores and SDS scores of the two groups were significantly reduced, and the Jalowies scores and WHOQOL-BREF quality of life scores were significantly increased. After management, the SAS score and SDS score of the experimental group were 47.32±5.68, 49.93±6.49 and 54.95±6.59, 55.33±8.30 in the control group, the difference was significant between two groups ( t value was 8.503, 4.969, P<0.01). The Jalowies scores and WHOQOL-BREF scores of the experimental group were higher than those of the control group ( t value was -27.662--4.290, P<0.01). The total effective rate of rehabilitation in the experimental group was 98.94%(93/94), which was significantly higher than 90.43%(85/94) in the control group, the difference was statistically significant ( χ2 value was 6.760, P <0.05). The total satisfaction rate of the experimental group for management intervention was 100.00%(94/94), which was significantly higher than the total satisfaction rate of the control group, 91.49%(86/94), the difference was statistically significant ( χ2 value was 6.397, P <0.05). Conclusions:In the patients with affective disorder caused by coronary heart disease and cerebrovascular disease, multi-disciplinary cooperation and optimization management mode can significantly reduce the negative emotions of patients after management, improve the way of patients to deal with the disease, effectively improve the clinical rehabilitation effect and the quality of life of patients, improve the satisfaction rate of patients with management, shorten the distance between nurses and patients, and achieve ideal results.

Objective To study the expression and significance of miR-26a in intrahepatic cholangiocarcinoma.Methods The expression of miR-26a in 46 intrahepatic cholangiocarcinoma(ICC) tissues and peritumoral tissues was detected by quantitative real time polymerase chain reaction (qRT PCR).The intrahepatic eholangiocarcinoma cell line HCCC-9810 and RBE were transfected with miR 26a mimics and miR 26a inhibitors,respectively,by lipofectamine 2000.The growth curves were constructed by the CCK 8 method.The migration and invasion ability was demonstrated by wound healing and transwell assay.The cell cycle was analyzed by flow cytometry.The potential mechanism was illustrated by Western blotting.Results For the 46 ICC tissues and peritumoral tissues,miR 26a levels were significantly higher in the tumor tissues than in the peritumoral tissues (P＜0.05).Vascular invasion,TNM Ⅲ～Ⅳ stage and lymph node metastasis were significantly associated with high miR 26a expression levels (P＜0.05),but gender,age,tumor amounts,tumor encapsulation,tumor diameter and tumor differentiation showed no significant association (P＞0.05).Enhanced cell proliferation,migration and invasion ability,accelerated G0/G1 phase to S phase transition,activated AKT by PTEN suppression were observed in HCCC-9810 cells with up regulation of miR-26a.Conversely,cell proliferation,migration and invasion ability was inhibited,G0/G1 phase was blocked and AKT was restrained by PTEN increase wkh down regulation of miR-26a in RBE cells.PTEN mRNA in versely correlated with the miR-26a level (r=-0.8272,P＜0.01).Patients with a high miR-26a expression had significantly poorer overall survival (P＜0.05).A high miR 26a exprcssion,multiple tumors and lymph node metastasis were independent prognostic factors of overall survival (P＜0.01).Conclusion Overexpression of miR-26a in intrahepatic cholangiocarcinoma correlated with clinicopath ological features and overall survival,and it potentially promoted tumor proliferation and metastasis through the PTEN/AKT pathway.

Objective To examine the localization of neurokinin B receptor (NK3)\|like immunoreactivity (\|LI) in the central nervous system of the mouse. Methods An immunohistochemcial staining method was used. Results NK3 receptor\|LI was localized in somatic and dendritic profiles in the most parts and in neuropil in a few regions of the mouse central nervous system. A large number of neurons with NK3\|LI was seen in the anterior olfactory nuclei, accumbens nucleus, septal area, ventral pallidum, pallidum, caudate putamen, nucleus of the stria terminalis, anterior hypothalamic area, tuber cincreum area, lateral hypothalamic area, perifornical nucleus, supraoptic nucleus, arcuate nucleus, mammillar nuclei, substatia nigra, ventral tegmental area, retrorubral area, superior and inferior colliculus, periaqueductal gray, nucleus of the solitary tract, and superficial layers of the medullary and spinal dorsal horns. The superfical layers of the cerebral cortex, piriform cortex, dorsal hippocampus, amygdal complex, reticular formation of the brainstem contained some neurons with NK3 receptor\|LI. In the ventral hippocampus, median and intralaminar nuclei of the thalamus and interpeduncular nuclei, NKR\|LI was localized in neuropil. Conclusion\ Neurons with NK3 receptor\|LI were widely distributed in the central nervous system. It may be involved in many physiological functions in the central nervous system of the mouse.\;[

Objective: To construct a phage display library of human single-chain Fv antibodies against blood group Rh(D) substance. Methods: Combining phage display library techniques, isolated total RNA from B lymphoblastoid cell lines secreting anti-Rh(D) antibodies was used for the synthesis of the first strand of cDNA, V_ H and V_ L genes were amplified by 2nd PCR and linked together by splicing overlap extension (SOE) with the use of a (Gly_ 4Ser)_ 3 linker. The resulted scFv genes were then cloned into pCANTAB5E vectors and displayed on the phage. Phage clones were selected using intact red cells as a source of antigen. After 4 rounds of "binding-elution-enrichment", each clone was assayed for specificity by Dot ELISA. Results: A phage antibody library, with the sink size being 1.2?107, was obtained. The percentage of full-length scFv gene inserted into phage DNA was 0.80. Rescued by helper phage, a phage scFv library with titer of 3?108 pfu/ml was established. Specific phages with scFv were acquired after 4 rounds of panning, one clone exhibiting specific binding to Rh+ cell was identified by Dot ELISA. Conclusion: A strategy for construction phage antibody library by means of phage display technique was practicable, which would be useful in screening engineered antibodies against human Rh (D) blood group substances.