Journal: BMC Microbiology

Article Title: Role of Mycobacterium tuberculosis pknD in the Pathogenesis of central nervous system tuberculosis

doi: 10.1186/1471-2180-12-7

Figure Lengend Snippet: Invasion and survival of M. tuberculosis pknD mutant in host-derived cells . A . BALB/c mice were infected with M. tuberculosis CDC1551 or pknD mutant, and sacrificed at days 1 and 49 after infection. The mutant for M. tuberculosis pknD was significantly attenuated (P = 0.004) in mouse brain, but not lung tissue, 49 days after infection. No defect was observed in the lungs at either time point. Bacterial burden is represented as log 10 CFU/organ for all animal experiments. B . Invasion of host-cell monolayers by wild-type CDC1551, wild-type intergenic transposon control, pknD transposon mutant (pknD:Tn), and pknD genetic complement (pknD:Comp) was examined and normalized to the wild-type control. Invasion assays were performed in brain microvascular endothelial cells (HBMEC), epithelial A549 cells, and umbilical vein endothelia (HUVEC). No difference in invasion was observed in A549 cells (P = 0.31) or HUVEC (P = 0.41). A significant reduction in invasive capacity, however, was observed in the CNS-derived HBMEC (P = 0.02). This defect was restored by genetic complementation with the native pknD/pstS2 operon. N.S. = not significantly different. C . Intracellular survival of each of the above M. tuberculosis strains was examined in HBMEC at days 1, 3, 5, and 7 after infection. The pknD:Tn mutant demonstrated an invasion and intracellular survival defect in HBMEC relative to wild-type over the course of the seven day infection. D . Survival was also examined by infection of activated J774 macrophages. No corresponding survival defect for the pknD:Tn mutant was observed in these cells during the seven day infection. A mutant for the gene Rv0442c , known to be attenuated in the macrophage model, is included as a control. All CFU counts are represented as mean ± standard deviation.

Article Snippet: Primary human brain microvascular endothelial cells and HUVEC were kind gifts from Dr. Kwang Sik Kim, Department of Pediatrics, Johns Hopkins University School of Medicine.

Techniques: Mutagenesis, Derivative Assay, Infection, Control, Standard Deviation

Journal: Journal of Neuroinflammation

Article Title: Visualizing neuroinflammation with fluorescence and luminescent lanthanide-based in situ hybridization

doi: 10.1186/s12974-019-1451-2

Figure Lengend Snippet: Toll-like receptor 4 (TLR4) mRNA staining in BV2 microglia cells and J774 macrophage cells (biotinylated TLR4 cRNA probe conjugated to streptavidin (SA)-Alexafluor555) following 0 h and 4 h of LPS stimulation (0.5 μg/ml) compared to cellular autofluorescence. a DAPI (blue) and BV2 cell autofluorescence or TLR4 mRNA expression (orange) following LPS-induced inflammation. b DAPI (blue) and J774 cell autofluorescence or TLR4 mRNA expression (orange) following LPS-induced inflammation. c Brightness intensities of LPS-treated BV2 cells represented in arbitrary fluorescence units. d Brightness intensities of LPS-treated J774 cells represented in arbitrary fluorescence units. 40x magnification, scale bar = 20 μm

Article Snippet: The murine BV2 microglia cells were a generous gift from Prof. Gilles Guillemin (Macquarie University, Sydney, Australia), and J774 cells were a generous gift from Dr. Nisha Schwarz (SAHMRI, Adelaide, Australia).

Techniques: Staining, Expressing, Fluorescence

Journal: Journal of Neuroinflammation

Article Title: Visualizing neuroinflammation with fluorescence and luminescent lanthanide-based in situ hybridization

doi: 10.1186/s12974-019-1451-2

Figure Lengend Snippet: Luminescent lanthanide-based ISH staining in BV2 cells. a The detection of DAPI (blue; top panel) with epifluorescent microscopy and autofluorescence (red; bottom panel) with time-gated microscopy. b The detection of DAPI (blue; top panel) with epifluorescent microscopy and non-specific SA-Eu 3+ binding (red; bottom panel) by time-gated microscopy. c – e The detection of DAPI (blue; top panel) with epifluorescent microscopy and the detection of TLR4 mRNA (red) by luminescent lanthanide staining and time-gated microscopy at c 0 h, d 4 h, and e 24 h of LPS-induced inflammation. f TLR4 mRNA was significantly increased from 0 to 4 h following LPS stimulation and subsequently returned to near baseline levels of expression by 24 h. A filter with 343-nm excitation/441-nm emission was used for epifluorescent microscopy (DAPI filter); 340-nm excitation/615-nm emission was used for SA-BHHTEGST-Eu 3+ time-gated microscopy (100x magnification; scale bars = 100 μm)

Article Snippet: The murine BV2 microglia cells were a generous gift from Prof. Gilles Guillemin (Macquarie University, Sydney, Australia), and J774 cells were a generous gift from Dr. Nisha Schwarz (SAHMRI, Adelaide, Australia).

Techniques: Staining, Microscopy, Binding Assay, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Hypoxia Inhibits Cell Cycle Progression and Cell Proliferation in Brain Microvascular Endothelial Cells via the miR-212-3p/MCM2 Axis

doi: 10.3390/ijms24032788

Figure Lengend Snippet: Functional analysis of differentially expressed genes (DEGs) in bEnd.3 cells under 24 h hypoxia. ( A ) Volcano map of differentially expressed genes between the control and hypoxia conditions. ( B ) Heatmap of the top upregulated and downregulated genes. ( C ) Gene ontology (GO) term analysis of differentially expressed genes. ( D ) Top 20 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. ( E ) Top 10 upregulated pathways from gene set enrichment analysis (GSEA). ( F ) Top 10 downregulated pathways from GSEA.

Article Snippet: The mouse brain microvascular endothelial bEnd.3 cell line was obtained from iCell (Shanghai, China).

Techniques: Functional Assay