Journal: Diabetes

Article Title: UBASH3A Mediates Risk for Type 1 Diabetes Through Inhibition of T-Cell Receptor–Induced NF-κB Signaling

doi: 10.2337/db16-1023

Figure Lengend Snippet: IL-2 production in Jurkat cells and in Jurkat-derived clones deficient in or overexpressing UBASH3A. A and B: ELISA measurements of IL-2 in the culture supernatants of Jurkat cells and of UBASH3A−/− (A) and UBASH3A-overexpressing (B) clones after 24-h stimulation with either anti-CD3 alone or anti-CD3 plus anti-CD28. The dashed lines indicate the ELISA detection limit (i.e., 7.8 pg/mL), and data points below the dashed lines represent values extrapolated from the IL-2 ELISA standard curve. The data are pooled from four ELISA experiments. C: Relative mRNA levels of IL2 in Jurkat cells and in UBASH3A−/− 2.1D6 and 2.1F7 cells. The cells were stimulated for 6 h as described in research design and methods. The data are pooled from two quantitative PCR experiments. Each data point in A–C represents an individual measurement of one sample. Numbers of samples (n) are shown. The mean and SEM values are indicated by solid lines and error bars, respectively. Unpaired two-tailed Student t tests were performed to assess statistical significance: *P = 0.02, ***0.0001 ≤ P < 0.0002, ****P < 0.0001.

Article Snippet: IL-2 ELISA Cells were stimulated as described above for 24 h, and the Human IL-2 ELISA Max Deluxe kit (BioLegend) was used to measure IL-2 production in the culture supernatants.

Techniques: Derivative Assay, Clone Assay, Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction, Two Tailed Test

Journal: Nutrition & Metabolism

Article Title: High fat diet and PCSK9 knockout modulates lipid profile of the liver and changes the expression of lipid homeostasis related genes

doi: 10.1186/s12986-023-00738-z

Figure Lengend Snippet: Hepatic mRNA and protein expression of Pcsk9 and Anxa2 genes after 20 weeks on HFD. The hepatic expression was assessed at mRNA and protein levels by qPCR/ELISA for PCSK9 ( A, B ) and qPCR/Western blot for ANXA2 ( C, D ). The mRNA level was calculated with 2 − ΔΔ Ct method. The relative protein level on the blots was determined by ImageJ, using actin as an internal control. Values are reported as mean ± SD; n = 5 in each group. The p value was determined by unpaired T-test. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: PCSK9 secretion was measured from the conditioned medium using a LEGEND MAX™ Human PCSK9 ELISA Kit (BioLegend, USA).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Western Blot, Control

Journal: Nutrition & Metabolism

Article Title: High fat diet and PCSK9 knockout modulates lipid profile of the liver and changes the expression of lipid homeostasis related genes

doi: 10.1186/s12986-023-00738-z

Figure Lengend Snippet: Lipid accumulation in the livers of PCSK9 −/− mice on control diet. Lipid composition of the liver was examined by HPLC–MS/MS. The concentration of TG, DG and CER subtypes ( A, F, K ), their total amount ( B, G, L ), the proportion of saturated and monounsaturated TGs, DGs and CERs to total amount ( C, D, H, I, M, N ) and the ratio of monounsaturated and saturated lipids were determined ( E, J, O ). Values are reported as mean ± SD; n = 5 in each group. The p value was determined by multiple T-test ( A, F, K ) or unpaired T-test (all others). * p < 0.05

Article Snippet: PCSK9 secretion was measured from the conditioned medium using a LEGEND MAX™ Human PCSK9 ELISA Kit (BioLegend, USA).

Techniques: Control, Tandem Mass Spectroscopy, Concentration Assay

Journal: Nutrition & Metabolism

Article Title: High fat diet and PCSK9 knockout modulates lipid profile of the liver and changes the expression of lipid homeostasis related genes

doi: 10.1186/s12986-023-00738-z

Figure Lengend Snippet: Hepatic gene expression of Pcsk9 and Anxa2 genes in PCSK9 −/− mice on control diet. The hepatic expression was assessed at mRNA and protein levels by qPCR/ELISA for PCSK9 ( A, B ) and qPCR/Western blot for (ANXA2; C, D ). The mRNA level was calculated with 2 − ΔΔ Ct method. The relative protein level on the blots was determined by ImageJ, using actin as an internal control. Values are reported as mean ± SD; n = 5 in each group. The p value was determined by multiple T-test ( A and B ). * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: PCSK9 secretion was measured from the conditioned medium using a LEGEND MAX™ Human PCSK9 ELISA Kit (BioLegend, USA).

Techniques: Gene Expression, Control, Expressing, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Nutrition & Metabolism

Article Title: High fat diet and PCSK9 knockout modulates lipid profile of the liver and changes the expression of lipid homeostasis related genes

doi: 10.1186/s12986-023-00738-z

Figure Lengend Snippet: Hepatic gene expression of Cd36 and Ldlr genes in PCSK9 −/− mice. Hepatic expression at mRNA level was measured by qPCR and calculated with 2 − ΔΔ Ct method ( A, C ). The relative protein level on the blots was determined by ImageJ, using actin as an internal control ( B, D ). Values are reported as mean ± SD; n = 5 in each group. The p value was determined by multiple T-test ( A and B ). * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: PCSK9 secretion was measured from the conditioned medium using a LEGEND MAX™ Human PCSK9 ELISA Kit (BioLegend, USA).

Techniques: Gene Expression, Expressing, Control

Journal: Nutrition & Metabolism

Article Title: High fat diet and PCSK9 knockout modulates lipid profile of the liver and changes the expression of lipid homeostasis related genes

doi: 10.1186/s12986-023-00738-z

Figure Lengend Snippet: Co-localization of PCSK9 with LDLR, CD36 and ANXA2. HepG2 cells were examined for PCSK9 secretion 16 and 40 h after serum depletion ( A , n = 3). Cellular localization of PCSK9, LDLR, CD36 and ANXA2 proteins was examined by laser scanning confocal microscopy ( B ). Co-localization rate was used to quantify the degree of co-localization ( C , n = 5). The scale bar has a width of 5 µm. Values are reported as mean ± SD. The p value was determined by unpaired T-test ( A ) or one-way ANOVA combined with Tukey’s multiple comparisons test ( C ). * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: PCSK9 secretion was measured from the conditioned medium using a LEGEND MAX™ Human PCSK9 ELISA Kit (BioLegend, USA).

Techniques: Serum Depletion, Confocal Microscopy

Journal: Nutrition & Metabolism

Article Title: High fat diet and PCSK9 knockout modulates lipid profile of the liver and changes the expression of lipid homeostasis related genes

doi: 10.1186/s12986-023-00738-z

Figure Lengend Snippet: Overview figure summarizing the content of the manuscript. The figure represents changes in PCSK9, LDLR, ANXA2 and CD36 levels after 20 weeks HFD and in PCSK9 gene knockout mice. The quantity of the colour coded proteins is proportional with the amount found under experimental conditions. Changes in different lipid levels are listed in the tables

Article Snippet: PCSK9 secretion was measured from the conditioned medium using a LEGEND MAX™ Human PCSK9 ELISA Kit (BioLegend, USA).

Techniques: Gene Knockout