Journal: BMC Biology

Article Title: Involvement of Plasmodium falciparum protein kinase CK2 in the chromatin assembly pathway

doi: 10.1186/1741-7007-10-5

Figure Lengend Snippet: Reverse genetics of PfCK2β1 . 3D7 parasites transfected with pCAM-BSD-KOPfCK2β1 with or without pCHD-PfCK2β1 were analyzed by PCR and Southern blotting. (A) Locations of the primers used for PCR screening, and the restriction enzymes used to cut the genomic DNA to give a diagnostic pattern of bands for analysis by Southern blotting. One truncated copy possesses its promoter and initiation codon, but lacks the C-terminal cysteine pair, a stop codon and a 3'UTR, whereas the other copy possesses both cysteine pairs but lacks a promoter and an initiation codon, and has an artificial stop codon introduced. (B) PCR screening of DNA from untransfected 3D7 parasites, two separate pCAM-BSD-KOPfCK2β1-transfected lines (KOCK2β1 1 and KOCK2β1 2), and parasites transfected with both the knockout plasmid and the complementation plasmid (KOCK2β1 + comp). (1) Amplification of the wild-type locus; (2) amplification over the 5' integration boundary; (3) amplification of the insert in the pCAM-BSD-KOPfCK2β1 plasmid. Evidence of integration was seen only in the DNA from the doubly transfected parasite culture (KOCK2β1 + comp; lane 2, faint band seen at 598 bp). (C) The parasite DNA was digested using the restriction enzymes EcoR I and Cla I, and analyzed by Southern blotting, using BSD and PfCK2β1 sequences as probes. (1) Untransfected 3D7; (2) KOCK2β1 1; (3) KOCK2β1 2; (4) KOCK2β1 + complement. See text for details.

Article Snippet: Forty-eight hours later, the ring-stage parasites were transfected by electroporation with 100 μg of purified plasmid DNA in buffer (Cytomix; Gibco-BRL, Grand Island, New York, NY, USA) as described previously [ ].

Techniques: Transfection, Southern Blot, Diagnostic Assay, Knock-Out, Plasmid Preparation, Amplification

Journal: Scientific Reports

Article Title: Development of an acute lung injury model for drug testing

doi: 10.1038/s41598-025-02078-9

Figure Lengend Snippet: Exposure of the small airway epithelial cells (SAECs) in the model to cytomix (10 ng/ml each of tumor necrosis factor alpha [TNF-α], interferon gamma [IFN-γ], and interleukin-1 beta [IL-1β]). Confocal immunofluorescence microscopy images visualized using LASX 3D Viewer, of tight junctions (zonula occludens-1-AF647, red; E-cadherin-AF488, green) for untreated (control; left panel) and cytomix treated (right panel) and ( B ) showing the differentiated SAECs cell types including cilia (acetylated tubulin-AF647, magenta), goblet cells (MUC5AC-AF405, blue), club cells (SCGB3A2-AF555, red), and basal cells (cytokeratin 5-AF488, green) for the untreated (upper panel), cytomix treated SAECs (lower panel). ( C ) Transepithelial electrical resistance (Ω*cm 2 ) in untreated (control) and cytomix treated SAECs. Fold change in cytokine expression of SAECs following cytomix treatment compared to untreated control for ( D ) C–X–C motif chemokine ligand 10 (CXCL10), ( E ) C–C motif ligand 2 (CCL2), ( F ) C–C motif ligand 3 (CCL3), ( G ) interleukin-6 (IL-6), ( H ) IL-17, and ( I ) soluble receptor for advanced glycation end products (sRAGE) for 2 independent experiments with 4 replicates per experiment. Student’s t-test, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: To formulate Cytomix media, recombinant human TNF-α, IL-1β, and IFN-γ (Peprotech, Cranbury, NJ) were added to EMEM without FBS to a final concentration of 50 ng/ml each.

Techniques: Immunofluorescence, Microscopy, Control, Expressing

Journal: Scientific Reports

Article Title: Development of an acute lung injury model for drug testing

doi: 10.1038/s41598-025-02078-9

Figure Lengend Snippet: Drug testing using the small airway epithelial cell (SAEC)/human umbilical vein endothelial cell (HUVEC) model with cytomix as the inflammatory exposure. Measurement of ( A ) SAEC (epithelial) and ( B ) HUVECs (endothelial) expression of interleukin-6 (IL-6; pg/ml) in response to untreated (sham), cytomix (10 ng/ml each of tumor necrosis factor alpha [TNF-α], interferon gamma [IFN-γ], and interleukin-1 beta [IL-1β]), and cytomix plus the JAK1/2 inhibitor baricitinib (1 µM) for 24 h at 37 °C, 5% CO 2 . Independent experiments (n = 2), replicates per experiment (n = 6). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Confocal immunofluorescence microscopy images ( C ) showing basal cells (cytokeratin 5-AF488, green; left panels), club cells (SCGB3A2-AF555, red; right panels) and pSTAT3 (green) and STAT3 (red) expression in SAECs for the untreated (top panels), cytomix (middle panels), and cytomix plus baricitinib treated (lower panels) SAECs. Images were taken at the same laser intensity for untreated, cytomix and cytomix + baricitinib treated samples.

Article Snippet: To formulate Cytomix media, recombinant human TNF-α, IL-1β, and IFN-γ (Peprotech, Cranbury, NJ) were added to EMEM without FBS to a final concentration of 50 ng/ml each.

Techniques: Expressing, Immunofluorescence, Microscopy