Journal: Endocrinology

Article Title: Thymic PTH Increases After Thyroparathyroidectomy in C57BL/KaLwRij Mice

doi: 10.1210/en.2017-03083

Figure Lengend Snippet: Thymus-derived PTH increases after TPTX. After serum PTH returned to a pre-TPTX level, TPTX and control mice were euthanized, and parathyroid and thyroid (Thyroid+Parathyroid), liver, and thymus tissues were removed. Harvested tissues were washed extensively in PBS and homogenized in T-PER buffer, and proteins were immunoprecipitated with goat anti-mouse PTH antibodies, including anti-mouse PTH(1-12) and anti-mouse PTH(53-84) or goat IgG as an isotype control. (A) PTH was detected in whole cell lysates (WCL) of the parathyroid+thyroid, after immunoprecipitation from the parathyroid+thyroid, and in the thymus (indicated by arrows). PTH protein was not detected in the liver. (B) Thymi from TPTX mice contained more PTH than thymi from nonoperated (control) mice. (C) Immunohistochemistry showing increased PTH protein production in the thymus after TPTX. Thymus sections were prepared and incubated with anti–PTH(1-12) antibody. The antibody was visualized with 3,3′-diaminobenzidine and counterstained with methyl green. The representative PTH signals are indicated by yellow arrows. (D) PTH-positive cells were enumerated in at least four sections from each group and plotted as mean ± standard error. Student t test was performed. ***P < 0.001. IP, immunoprecipitated; Iso, isotype control antibody; PT, parathyroid glands; PTH*, parathyroid hormone antibody.

Article Snippet: Tissue lysates were incubated with 1 μg goat anti-mouse PTH antibodies, including anti-mouse PTH(1-12) [catalog no. 20-2320; Research Resource Identifier: AB_2721076 ; Quidel, San Diego, CA] and anti-mouse PTH(53-84) (catalog no. 20-2310; Research Resource Identifier: AB_2721077 ; Quidel) or with 1 μg goat immunoglobulin G (IgG; catalog no. ab37373; Abcam, Cambridge, MA) as an isotype control for 30 minutes at 4°C on a rotary shaker.

Techniques: Derivative Assay, Immunoprecipitation, Immunohistochemistry, Incubation

Journal: Research and Practice in Thrombosis and Haemostasis

Article Title: Activation of coagulation FXI promotes endothelial inflammation and amplifies platelet activation in a nonhuman primate model of hyperlipidemia

doi: 10.1016/j.rpth.2023.102276

Figure Lengend Snippet: Levels of activated FXI were elevated in a model of hyperlipidemia. PPP was isolated from NHPs on a standard chow diet ( n = 6, lean) or NHPs on a high-fat diet ( n = 8, obese). Clotting times were measured by incubating PPP with an aPTT reagent followed by CaCl 2 to initiate clot formation (A) or by stimulating PPP with Dade Innovin Reagent to measure PT. Activated coagulation protease species, FXIa–AT and T-AT, were measured using a custom ELISA (C, D). Statistical analyses were conducted using a Mann–Whitney test or an unpaired t -test. Statistical significance is indicated by an asterisk for P ≤ .05. Data are shown as mean ± SEM. aPTT, activated partial thrombospondin time; AT, antithrombin; ELISA, enzyme-linked immunosorbent assay; F, factor; PPP, platelet-poor plasma; PT, prothrombin time; NHP, nonhuman primate.

Article Snippet: Plasma samples were analyzed for C-reactive protein using an enzyme-linked immunosorbent assay kit (ELISA, ALPCO) following the manufacturer’s instructions.

Techniques: Isolation, Coagulation, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: Research and Practice in Thrombosis and Haemostasis

Article Title: Activation of coagulation FXI promotes endothelial inflammation and amplifies platelet activation in a nonhuman primate model of hyperlipidemia

doi: 10.1016/j.rpth.2023.102276

Figure Lengend Snippet: FXI inhibition prolonged aPTT clotting times in a model of hyperlipidemia. PPP was isolated from NHPs on a high-fat diet ( n = 5) treated with a FXI function–blocking antibody. Clotting times were measured by incubating PPP with an aPTT reagent followed by CaCl 2 to initiate clot formation (A) or by stimulating PPP with Dade Innovin Reagent to measure PT (B). Activated coagulation protease species, FXIa–AT and T–AT, were measured using a custom ELISA (C, D). Statistical analyses were conducted using a Friedman test with a Dunn’s post-hoc test or repeated measures one-way ANOVA with a Dunnett’s post-hoc test. Statistical significance is indicated by an asterisk for P < .05. Data are shown as mean ± SEM. aPTT, activated partial thrombospondin time; AT, antithrombin; ELISA, enzyme-linked immunosorbent assay; F, factor; NHP, nonhuman primate; PPP, platelet-poor plasma; PT, prothrombin time.

Article Snippet: Plasma samples were analyzed for C-reactive protein using an enzyme-linked immunosorbent assay kit (ELISA, ALPCO) following the manufacturer’s instructions.

Techniques: Inhibition, Coagulation, Isolation, Blocking Assay, Enzyme-linked Immunosorbent Assay