BACKGROUND:A short-peptide small molecule hydrogel (SMH) developed in the previous study has more obvious advantages than other hydrogels to improve local microenvironment, carry bioactive substances and interfere with stem cell signal transduction pathways. OBJECTIVE:To explore the effect of SMHs on bone marrow mesenchymal stem cells (BMSCs) proliferation, apoptosis and differentiation into myocardial cells. METHODS: (1) Passage 9 rat BMSCs in vitro were divided into control group and experimental group, followed by routine culture and culture in SMHs, respectively. At 7 days of culture, cell proliferation and apoptosis were detected. Cells in the two groups were exposed to anaerobic environment for 12 hours, and expression levels of Bcl-2, Bax and Caspase-3 in BMSCs were detected. (2) Passage 9 BMSCs were divided into four groups and then cultured in 5-azacytidine, SMHs, SMHs+5-azacytidine, and L-DMEM (normal control), respectively. After 4 weeks of induction, expression of CTnT, desmin and Cx-43 proteins was detected and expression levels of early cardiac transcription factors, NKX2.5 and GATA-4, were also measured. RESULTS AND CONCLUSION: (1) Compared with the control group, better proliferation and lower apoptosis of BMSCs were found in the experimental group. Under anaerobic conditions, the number of survival cells was reduced in both groups, but less apoptosis or necrosis was found in the experimental group than the control group (P < 0.05). Moreover, the level of Bcl-2 was higher in the experimental group than the control group (P < 0.01), while the levels of Bax and Caspases-3 protiens were lower in the experimental group than the control group (P < 0.01). (2) NKx2.5 and GATA-4 mRNA expression was found in both 5-azacytidine and SMHs+5-azacytidine groups, and moreover, the mRNA levels of early cardiac transcription factors were significantly higher in the SMHs+5-azacytidine group than in the 5-azacytidine group (P < 0.05). In the normal control group, cTnT expressed negatively, and desmin and Cx-43 expressed weakly. The expression of cTnT, desmin and Cx-43 proteins was higher in the SMHs+5-azacytidine group than in the 5-azacytidine and SMHs groups, while there was no significant difference between the latter two groups. To conclude, SMHs as a culture medium is conducive to the proliferation of BMSCs, reduces cell apoptosis, and promotes myocardial differentiation of BMSCs.

Objective: To clone Fd and light chain genes of monoclonal antibody HAb18 against human hepatoma and verify their accuracy and liability. Methods: Total RNA was extracted from hybridoma cell line secreting MAb HAb18, and Fd and light chain genes were amplified by RT-PCR. After PCR products were ligated into pMD18T vector, positive clones were screened and DNA sequences were tested and analysed by relative softwares. Then, light chain and Fd genes were sequential cloned into phage display vector pComb3. After recombinant vector was transformed into E.coli XL1-blue, recombinant vector was rescued by helper phage M13K07 and the specificity of phages to antigen was detected by indirect ELISA. Results: The size of amplified Fd and light chain genes was separately 665 bp and 668 bp. The results of sequence analysis showed that both VL and VH contained 2 characteristic cystines and CH1 was IgG1 classes and CL was ?. ELISA result identified that expressed Fab antibody could specially bind to corresponding antigen. Conclusion: Fd and light chain genes of MAb HAb18 were successfully cloned, which lay a good foundation for constructing a diversity of engineering antibody.

Objective To study CT and MRI features of intracranial chondrosarcomas. Methods CT and MRI of 3 cases with intracranial chondros arcomas proved by pathology were retrospectively analyzed.Results CT scans showed the tumors were lobular mass, per itumoral edema were not serious, intratumorous calcification and bone invasion w ere seen in 2 cases.MRI showed that tumor had low to intermediate signal intensi ty or hypointensity on T 1WI, high signal intensity on T 2WI. On contrast enha nced MRI,the center of lesions which was low signal intensity on T 2WI was no e nhancement, but there was observable enhancement at periphery of tumors.Conclusion Intracranical chondrosarcomas are often orgina ted from synchondrosis of the skull base,the lesions are often associated with c alcification, and bone invasion, the accurate dignosis should depend on microsco pic examination and immunohistochemical staining.

AIM:To investigate the kinetics of tumor necrosis factor alpha(TNF-?)in an animal model of chronic P.aeruginosa biomembrane associated with lung infection.METHODS:Rats were challenged with 0.1 mL of PAO579(1012 CFU/L)in alginate beads or 0.1 mL of planktonic PAO579(1012 CFU/L).After challenging for 3,7 and 14 d,bacteriological and pathological features,and TNF-? expression in lung tissue were observed.RESULTS:(1)CFU/lung in alginate beads group was significantly higher than that in planktonic bacteria group(P

OBJECTIVE To understand the effects of lasR/rhlR mutations on the biofilm formation of Pseudomonas aeruginosa in vitro.METHODS The biofilms of PAEO1 and lasR,rhlR,and lasR rhlR mutants were incubated in a polyvinyl chloride tube in 0.9% saline and studied by means of scanning electronic microscopy(SEM) after three days′ incubation.RESULTS After 3 day′s incubation,PAEO1 biofilm showed a well-developed structure,whereas thin and poor developed biofilms were seen in lasR/rhlR mutant groups.CONCLUSIONS lasR/rhlR Genes play probably an important role in the formation of P.aeruginosa biofilm in vitro.

OBJECTIVE To understand the importance of quorum sensing(QS)system in Pseudomonas aeruginosa(PAE)biofilm infection in lungs,the pathogenic effects of the wild-type P.aeruginosa PAEO1 were compared with QS double mutants PAEO1 lasR rhlR and PAEO1 lasⅠ rhⅡ in vivo.METHODS Rats were challenged intratracheally with alginate embedded PAE strain PAEO1 and the mutants of QS in the concentration of 1?109 colony-forming units per milliliter(CFU/ml).Two weeks post intratracheal challenge with P.aeruginosa,parameters were evaluated.RESULTS Two weeks after challenge,significantly milder microscopic and macroscopic lung pathology(P

Objective To explore the miRNA regulating the potential cancer-promoting gene CCL18 in cutaneous malignant melanoma.Methods Bioinformatics analysis was conducted by using online software miRanda and TargetScan,so as to predict the miRNA targeting CCL18 gene.Three kinds of C CL18 3'UTR dual-luciferase reporter vectors,including mutant 3'UTR vector (mutant 3'UTR group),wildtype 3'UTR vector (wild-type 3'UTR group) and empty vector (blank control group),as well as miRNA vectors carring selected miRNAs were constructed according to human gene sequence analysis,and then were used to co-transfect 293T cells.After 48-hour treatment,the cells were lysed for detection of luciferase activity.Real-time fluorescence-based quantitative PCR was performed to measure the expression of CCL 18 and selected miRNA in 14 fresh malignant melanoma tissue specimens and 14 paracancerous normal skin tissue specimens (control tissues),and their correlations were analyzed.Results Online software analysis showed that some miRNAs were identified to target the 3'UTR of CCL18 gene,including miR-183,miR-128 and miR-33a.Luciferase reporter vectors and miRNA vectors were constructed successfully.As luciferase activity assay showed,when miR-183 and miR-128 were bound to the CCL18 3'UTR,the luciferase activities were significantly higher in their mutant 3'UTR groups (11.63 ± 0.42;8.80 ± 0.49) than in their wild-type 3'UTR groups (4.86 ± 0.39;5.01 ± 0.54;both P ＜ 0.05) and blank control groups (2.41 ± 0.13;2.39 ± 0.05;both P ＜ 0.01),while there were no significant differences between miR-33a-hinding mutant 3'UTR group (6.41 ± 0.47) and miR-33a-binding wild-type 3'UTR group (6.16 ± 0.22,P ＞ 0.05).Real-time fluorescence-based quantitative PCR revealed higher mRNA expression of the CCL18 gene (3.52 ± 1.68),but lower expression of miR-183 (0.49 ± 0.32),miR-128 (0.30 ± 0.20) and miR-33a (0.46 ± 0.40) in the malignant melanoma tissues compared with the control tissues.The mRNA expression of the CCL18 gene was negatively correlated with the expression of miR-128 (rs =-0548,P ＜ 0.05),but showed no significant correlations with the expression of miR-183 and miR-33a (both P ＞ 0.05).Conclusion miR-128 may play a role in regulating the potential malignant melanoma-promoting gene CCL18.

OBJECTIVE To understand the effects of meropenem on the biofilms of Pseudomonas aeruginosa lasR/rhlR mutations in vitro.METHODS The biofilms of PAEO1 and its lasR,rhlR,and lasR rhlR mutants were incubated in a polyvinyl chloride tube in 0.9% saline for three days,then were immersed in meropenem solution of 26 ?g/ml for 24 hours,and examined by scanning electron microscopy(SEM).RESULTS After three days incubation,PAEO1 biofilms showed a well-developed structure,however,the biofilms of PAEO1 lasR,rhlR,and lasR rhlR mutants were thin and poor developed;after 24 hours in meropenem solution,PAEO1 biofilm become rareness,whereas PAEO1 lasR,rhlR,and lasR rhlR mutants biofilms were almost destructed and only small pieces left.CONCLUSIONS lasR/rhlR Genes play probably an important role in the formation of P.aeruginosa biofilm and in the resistance to meropenem.

AIM: To study the expression profile of QKI m RNA and protein in rat heart during pathological cardiac hypertrophy. ME THODS: A rat cardiac hypertrophy model was established using continuous norepinephrine (NE) perfusion. Real-time PCR and Western blotting were applied t o examine QKI mRNA and protein expression respectively in rat heart. RES ULTS: Both mRNA and protein of three QKI isoforms were detected in adult rat heart. QKI-5 mRNA and total QKI protein were remarkably decreased in NE-ind uced hypertrophic heart compared with those in control group (P

A mouse-anti-human monoclonal antibody was produced by using the membrane proteins of human lung carcinoma cell line A549 as the immunogen to generate monoclonal antibodies against lung carcinoma with the use of hybridoma techniques. McAb4E7 was prepared successfully. To identify its antigen, proteomic technologies such as two-dimenstional electrophoresis, western blotting and mass spectrometry were employed. The targeting antigen of McAb4E7 expressed positive in human lung cancer cell lines A549 and human hepatocarcinoma cell line HepG2, moreover, the expression of the antigen was stronger in A549 cells. Finally, we obtained one positive protein in A549 cell line that has strong affinity and specificity for McAb4E7, which was identified to be ATP synthase beta subunit. We identified ATP synthase beta subunit as the targeting antigen of lung carcinoma special monoclonal antibody McAb4E7.
Animals ; Antibodies, Monoclonal ; biosynthesis ; chemistry ; immunology ; Antibodies, Neoplasm ; immunology ; Antibody Specificity ; Antigens, Neoplasm ; genetics ; immunology ; Cell Line, Tumor ; Humans ; Lung Neoplasms ; immunology ; Membrane Proteins ; immunology ; Mice ; Mice, Inbred BALB C ; Mitochondrial Proton-Translocating ATPases ; immunology