Journal: Scientific Reports

Article Title: 5T4-specific chimeric antigen receptor modification promotes the immune efficacy of cytokine-induced killer cells against nasopharyngeal carcinoma stem cell-like cells

doi: 10.1038/s41598-017-04756-9

Figure Lengend Snippet: 5T4-28Z CAR gene modification promotes the anti-NPC activity of CIK cells in a 5T4-dependent manner. Target cells consisted of 5T4 low-expressing S18 and S26 cells and their spheroid cells with an intermediate or high level of 5T4 expression. Effector cells included 5T4-28Z-CIK, 5T4-28Z-CIK (NKG2D-), C-28Z-CIK and NT-CIK cells. ( a–d ) Briefly, 1 × 10 4 target cells per well were co-cultured with different effector cells at E/T ratios of 10/1, 5/1 and 2.5/1 for 4 h. The cytotoxicity of effector cells was examined with LDH release assays. Values in the line graphs represent the mean ± SD of three parallel wells. ( e ) Then, 2 × 10 4 target cells per well were co-incubated with 1 × 10 5 effector cells per well for 24 h. Effector cells cultured alone in medium served as the negative control. IFN-γ production of the effector cells was assessed using ELISAs. Values in the histogram represent the mean ± SD of three parallel wells. ( f ) In addition, 2 × 10 5 target cells per well were co-cultured with 1 × 10 6 effector cells per well for 5 h in the presence of 1 × Protein Transport Inhibitor Cocktail and anti-human CD107α-APC antibody or IgG isotype control. Effector cells cultured alone served as the negative control. Degranulation of effector cells was evaluated using FACS. Values presented in the histogram represent the mean ± SD of triplicate samples. ( g ) S26 cells or S26 spheroids labelled with CM-Dil were co-cultured with 5T4-28Z-CIK cells and monitored using an inverted fluorescence microscope with climate control. Images captured intermittently are displayed. The results are representative of at least three independent experiments. * indicates p < 0.05 (one-way ANOVA).

Article Snippet: Next, 50 μl of supernatant per well was collected to measure LDH release using a cytotoxicity LDH Assay Kit-WST ® (Dojindo) according to the manufacturer’s specifications.

Techniques: Modification, Activity Assay, Expressing, Cell Culture, Incubation, Negative Control, Control, Fluorescence, Microscopy

Journal: The Korean Journal of Internal Medicine

Article Title: Angiotensin III increases monocyte chemoattractant protein-1 expression in cultured human proximal tubular epithelial cells

doi: 10.3904/kjim.2016.31.1.116

Figure Lengend Snippet: Angiotensin III (Ang III)-stimulated monocyte chemoattractant protein-1 (MCP-1) production in HK-2 cells. Cells were incubated for the indicated times in the presence or absence of Ang III. MCP-1 protein in culture medium was quantified by enzyme-linked immunosorbent assay. Results are shown as mean ± SEM from six independent experiments. Lactate dehydrogenase (LDH) release from Ang II-, Ang III-, or losartan-treated cells. LDH release is expressed as percentage of maximal LDH release induced by 1% Triton X-100 for 48 hours. a p < 0.05 vs. 8 hours MCP-1 level, b p < 0.05 vs, control cells.

Article Snippet: HK-2 cell viability was assessed by measuring lactate dehydrogenase (LDH) release into the culture medium using a LDH cytotoxicity detection kit according to the manufacturer’s protocol (Takara Biomedical, Kyoto, Japan).

Techniques: Incubation, Enzyme-linked Immunosorbent Assay

Journal: The Korean Journal of Internal Medicine

Article Title: Angiotensin III increases monocyte chemoattractant protein-1 expression in cultured human proximal tubular epithelial cells

doi: 10.3904/kjim.2016.31.1.116

Figure Lengend Snippet: Lactate dehydrogenase (LDH) release from angiotensin (Ang) II-, Ang III-, or losartan-treated cells. LDH release is expressed as percentage of maximal LDH release induced by 1% Triton X-100 for 48 hours.

Article Snippet: HK-2 cell viability was assessed by measuring lactate dehydrogenase (LDH) release into the culture medium using a LDH cytotoxicity detection kit according to the manufacturer’s protocol (Takara Biomedical, Kyoto, Japan).

Techniques:

Journal: Biomedicines

Article Title: Self-Organization Provides Cell Fate Commitment in MSC Sheet Condensed Areas via ROCK-Dependent Mechanism

doi: 10.3390/biomedicines9091192

Figure Lengend Snippet: Cells from condensed areas of the cell sheet (CS) are characterized by an increased content of the cytochrome-C oxidase IV component (COX-IV) and higher lactate dehydrogenase (LDH) activity. ( A ) Western blotting of the mitochondrial ETC COX-IV subunit in lysates of ML, total CS and microdissected condensed or scattered areas. Graphs present a densitometry analysis of the COX-IV content normalized to β-actin (densitometry analysis). ( B ) LDH activity (LDHact) in the lysates of microdissected condensed areas and scattered areas. DNA amounts in the respective cell lysates were used for the normalization of LDH activity to the cell number. Control data was obtained from lysates of the A431 cell line that increased the proportion of anaerobic glycolysis and the THP-1 cell line with the major input of oxidative phosphorylation but not glycolysis. The band numbers indicate individual biological replicate samples. In the figure graphs and histograms, significant differences were marked by * ( p < 0.05) and ** ( p < 0.005).

Article Snippet: The enzymatic activity of lactate dehydrogenase (LDH) in condensed and scattered areas of CS was assayed in the samples obtained immediately after laser microdissection using a Lactate Dehydrogenase (LDH) Assay Kit (Fluorometric) (ab197000, Abcam, Cambridge, UK) according to the manufacturer’s instructions.

Techniques: Activity Assay, Western Blot