Journal: Journal of Virology

Article Title: Stabilization of the human cytomegalovirus UL136p33 reactivation determinant overcomes the requirement for UL135 for replication in hematopoietic cells

doi: 10.1128/jvi.00148-23

Figure Lengend Snippet: Stabilizing UL136p33 rescues viral replication in the absence of UL135 in CD34+ HPCs. (A) CD34+ HPCs were infected with WT UL136myc, UL136mycΔ33kDa, UL136mycΔK→R, and ΔUL135STOP/UL136mycΔK→R at an MOI of 2. At 24 hpi, CD34+/GFP+ (infected cells) were sorted and seeded into long-term bone marrow culture. After 10 days in culture, parallel populations of either mechanically lysed cells or live cells were plated onto fibroblast monolayers in cytokine-rich media. 14 days later, GFP+ wells were scored, and the frequency of infectious centers was determined by extreme limiting dilution analysis. The mechanically lysed population defines the quantity of virus present prior to reactivation (pre-reactivation; white bar). The live-cell population defines the quantity of virus present after reactivation (reactivation; gray bar). The frequency was normalized to WT UL136myc pre-reactivation, and the average of three independent experiments is shown. Statistical significance was calculated using a two-way ANOVA with Tukey’s multiple comparison tests. *, P < 0.05; **, P < 0.01; ***, P < 0.001). (B) Total DNA was isolated from CD34+ HPCs infected with WT UL136myc, UL136mycΔ33kDa, UL136mycΔK→R, and ΔUL135STOP/UL136mycΔK→R at an MOI of 2 at 10 dpi. The number of viral genomes relative to the level of RNaseP expression was quantified by qPCR using β2.7kb RNA gene- and RNaseP-specific primers. Two biological replicates from two independent cell donors are shown.

Article Snippet: Briefly, CD34 + HPCs were infected at an MOI of 2 for 20 h after which a pure population (>97%) of infected (GFP + ) CD34 + cells were isolated via FACS (FACSAria, BD Biosciences Immunocytometry Systems, San Jose, CA, USA) using a phycoerythrin-conjugated CD34-specific antibody (BD Biosciences) and propidium iodide to exclude dead cells.

Techniques: Infection, Virus, Comparison, Isolation, Expressing

Journal: Oncology Reports

Article Title: Local recurrence of small cell lung cancer following radiofrequency ablation is induced by HIF-1α expression in the transition zone

doi: 10.3892/or.2015.4541

Figure Lengend Snippet: Angiogenesis potential is higher in the TZ than that in the RZ of the RUL and RLL, but has no effect on tumor proliferation. (A-C) Immunostaining showed CD34-positive staining in the TZ (A), RZ in RUL (B) and RZ in RLL (C) of the RFA group and RFA + YC-1 group. (D) Based on the semi-quantitative analysis of the CD34-positive stained area, it was noted that the MVD value of the TZ was higher than that of the RZ in the RUL and RLL at day 1, 7 and 14 following RFA treatment, but when treated with YC-1, MVD values of the TZ, RZ in RUL and RLL were decreased. * P<0.01, RFA group vs. RFA + YC-1 group in TZ, RZ in RUL and RZ in RLL; ** P<0.05, RFA group: TZ vs. RZ in RUL; *** P<0.05, RFA group: TZ vs. RZ in RLL. (E) The effect of PTK/ZK on perilesional outgrowth of established micrometastases evaluated 1, 7 and 14 days. After RFA treatment, tumor growth was expressed as the PRA in the TZ and RZ. In the RFA and sham groups, treatment with PTK/ZK reduced tumor growth in the RZ of the RUL and RLL. ※ p>0.05, RFA group vs. RFA + PTK/ZK group; ▲ p<0.05, RFA group vs. RFA + PTK/ZK group or sham group vs. PTK/ZK group; ★ P<0.01, RFA group vs. RFA + PTK/ZK group or sham group vs. PTK/ZK group.

Article Snippet: Sections were then incubated with a mouse anti-human NSE, HIF-1α and CD34 primary antibody (NSE, 1:50; HIF-1α, 1:1,000; CD34, 1:200 dilution; Wuhan Boster Biological Engineering Technology Co. Ltd.) overnight at 4°C.

Techniques: Immunostaining, Staining

Journal: Molecular Medicine Reports

Article Title: COX-2 inhibition in the endothelium induces glucose metabolism normalization and impairs tumor progression

doi: 10.3892/mmr.2017.8270

Figure Lengend Snippet: Characterization of normal and tumor endothelial cells. (A) In the immunofluorescence assay, ECs were stained for factor VIII (green) and CD34 (red). The merged image shows yellow and blue areas. (B) qRT-PCR of mRNA expression levels of VEGF in NECs and TECs. The values are expressed relative to NECs (n=3). (C) qRT-PCR of mRNA expression and WB of protein expression levels of PFKFB3 in NECs and TECs. The values are expressed relative to NECs (n=3). (D) Glucose levels in the medium of TECs relative to NECs (n=3). (E) Lactate levels in the medium of TECs relative to NECs (n=3). (F) Glycolytic flux in TECs relative to NECs (n=3). All data are shown as the means ± SEM. *P<0.05. ECs, endothelial cells; TECs, tumor endothelial cells; VEGF, vascular endothelial growth factor; PFKFB3, fructose-2,6-bisphosphatase; NECs, normal endothelial cells.

Article Snippet: The abovementioned cells were fixed with ice-cold 4% paraformaldehyde for 20 min at 37°C, blocked with normal serum for 20 min at room temperature and then incubated overnight in the dark at 4°C with specific antibodies against mouse monoclonal CD34 (ab 8158; Abcam, Cambridge, UK) and mouse monoclonal factor VIII (9035; InvitrogenTM; Thermo Fisher Scientific, Inc.).

Techniques: Immunofluorescence, Staining, Quantitative RT-PCR, Expressing