Effect of calcium dobesilate on vitreous hemorrhage in patients with PDR after pan retinal photocoagulation / 国际眼科杂志(Guoji Yanke Zazhi)

·AIM: To investigate the effect of calcium dobesilate on vitreous hemorrhage in patients with proliferative diabetic retinopathy ( PDR ) after pan retinal photocoagulation (PRP). ·METHODS:Totally 62 patients (30 cases with binocular lesions, 32 cases with monocular lesions, a total of 92 eyes) with PDR who were treated in our hospital from January 2015 to July 2017 were selected as the subjects. They were divided into the control group ( treated with pan retinal photocoagulation, n = 30, 17 cases with monocular lesions, 13 cases with binocular lesions, a total of 43 eyes ) and the study group ( treated with calcium dobesilate on the basis of treatment for the control group, n=32, 15 cases with monocular lesions, 17 cases with binocular lesions, a total of 49 eyes ). The recovery of visual acuity, blood rheology ( plasma viscosity, hematocrit, erythrocyte deformation index) and the incidence of complications such as vitreous hemorrhage in the two groups after surgery were observed. ·RESULTS: There was no significant difference between the two groups in the rate of excellent and good visual acuity, plasma viscosity, hematocrit or erythrocyte deformability index before treatment ( P>0. 05 ). After treatment, the rate of excellent and good visual acuity in the study group was significantly higher than that in the control group (P<0. 05). After treatment, the plasma viscosity and hematocrit decreased significantly while the erythrocyte deformability index significantly increased only in the study group, and changes of above -mentioned indexes in the study group were more obvious than those in the control group after treatment (P<0. 05). The incidence rate of vitreous hemorrhage and total incidence rate of complications in the study group were significantly lower than those in the control group ( P<0. 05). ·CONCLUSION: The application of calcium dobesilate in patients with PDR after pan retinal photocoagulation can effectively improve the recovery of visual acuity and reduce the incidence of complications such as vitreous hemorrhage. The mechanism may be related to effectively improving the hemodynamics.

Effect of miR-101 on biological characteristics of colorectal cancer cell line SW620 / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of miR-101 on biological behaviors of colorectal cancer cell line SW620.</p><p><b>METHODS</b>CCK-8 method, colony formation assay, cell cycle analysis and apoptosis analysis were applied to assess the effects of miR-101 on cell proliferation, invasion and apoptosis of SW620 cells.</p><p><b>RESULTS</b>Over-expression of miR-101 in SW620 cells significantly suppressed the cell proliferation and attenuated the colony-forming ability of the cells. Flow cytometry showed that over-expression of miR-101 in SW620 cells caused obvious cell cycle arrest in G2/M and 1/ phases, and significantly increased the cell apoptosis rate.</p><p><b>CONCLUSION</b>Over-expression of miR-101 can inhibit the proliferation, cause cell cycle arrest and promote apoptosis of colorectal cancer SW620 cells.</p>

Long-term culture and identification of CD133⁺ hematopoietic progenitor cells from human umbilical cord blood / 南方医科大学学报

<p><b>OBJECTIVE</b>To isolate CD133(+) hematopoietic progenitor cells from human umbilical cord blood and optimize the culture condition for maintaining their stem cell characteristics.</p><p><b>METHODS</b>CD133(+) hematopoietic progenitor cells were isolated from human umbilical cord blood using magnetic cell sorting system, and the cells were detected by flow cytometry. Four methods were used for culturing cells. After 8 weeks' culture, cytomorphology, flow cytometry, immunocytochemistry and immunofluorescence assay were used to identify the characteristics of the stem cells.</p><p><b>RESULTS</b>Over 80% of CD133(+) hematopoietic progenitor cells were isolated from human umbilical cord blood using magnetic cell sorting system. The cells were effectively expanded using optimized serum-free medium after 8 weeks of cell culture, whereas the cells in other media differentiated into adherent cells in a poor state.</p><p><b>CONCLUSION</b>The optimized serum-free medium allows effective expansion of CD133(+) hematopoietic progenitor cells that maintain stem cell characteristics after a long-term culture.</p>

Construction of has-miR-335 lentiviral vector and verification of the target gene of miR-335 / 南方医科大学学报

<p><b>OBJECTIVE</b>To construct a lentiviral vector of miR-335 gene and verify the target gene of miR-335.</p><p><b>METHODS</b>The precursor sequence of miR-335 gene was amplified from the genomic DNA by PCR and cloned into the lentiviral vector PLVTHM labeled with GFP. Real-time quantitative RT-PCR was used to detect miR-335 and RASA1 expression in different colorectal cancer cell lines. A recombinant vector psiCHECK-2-RASA1 containing RASA1 3'UTR was constructed followed by site-directed mutagenesis of RASA1 3'UTR to establish the vector psiCHECK-2-RASA1-Mut. Co-transfection of hsa-mir-335 or a NC with these recombined vectors in HEK293A and SW480 cells was performed, and dual-luciferase reporter assay was utilized to examine the changes in luciferase activities. The recombinant PLVTHM-miR335 plasmid was packaged into mature lentivirus by 293FT cells and used to infect SW620 cells. Flow cytometry was employed for sorting the GFP+ cells. The expression of miR-335 and RASA1 were determined by qRT-PCR, and Western blotting was used to detect the expression of RASA1 protein in SW620 cell lines.</p><p><b>RESULTS</b>The recombinant lentiviral vector PLVTHM-miR335, psiCHECK-2-RASA1 and the mutation expression vector psiCHECK-2-RASA1-Mut were successfully constructed. Dual-luciferase reporter assay showed that miR-335 decreased luciferase activity in cells co-transfected with psiCHECK-2-RASA1. The expression of miR-335 in SW620 cells infected with the lentivirus PLVTHM-miR335 was significantly increased, but the expression of RASA1 showed only slight changes. Overexpression of miR-335 suppressed the expression of RASA1 protein in SW620 cells.</p><p><b>CONCLUSION</b>We have successfully constructed the lentiviral vector containing mir-335 gene and a SW620 cell line with miR-335 overexpression. MiR-335 can suppress RASA1 gene expression by targeting the specific sequence of RASA1 3'UTR.</p>

Selection and identification of specific-binding peptides for cancer stem cell surface marker CD133 / 南方医科大学学报

<p><b>OBJECTIVE</b>To select the peptides that specifically bind human cancer stem cell surface marker CD133 from the Ph.D.-7>(TM) phage peptide library.</p><p><b>METHODS</b>With a biotinylated extracellular fragment of human cancer stem cell surface marker CD133 as the target protein, the CD133 high-affinity peptides were screened from the phage peptide library by liquid phase panning. The clones with high-binding force with human CD133 were then identified by sandwich ELISA and their single-stranded DNA was extracted to test the specificity by competitive ELISA. The amino acid sequences of the selected peptides derived from the phage DNA sequences were synthesized after sequence alignment analysis, and their capacity of binding with colorectal carcinoma cells was assessed by immunofluorescence technique.</p><p><b>RESULTS</b>After 4 rounds of liquid phase selection, the phages capable of specific binding with human CD133 were effectively enriched, with an enrichment ratio of 388 times compared to that at the fourth and first rounds. Thirteen out of the 20 clones from the fourth round of panning were identified as positive clones, among which 11 had identical amino acid sequence of TISWPPR, and 2 had the sequence of STTKLAL, and the former sequence showed a stronger binding specificity to CD133.</p><p><b>CONCLUSION</b>We have successfully obtained a peptide that specifically binds human CD133 from the Ph.D.-7(TM) phage peptide library, demonstrating the feasibility of screening small molecule high-affinity polypeptides from phage peptide library by liquid-phase panning.</p>