Kineret

The following reagents were used in the study: highly purified LPS (LPS E. coli, Serotype O111:B4,TLRgrade™, Enzo Life Sciences, Inc., Farmingdale, NY) synthetic bacterial lipoprotein, Pam3CSK4 (TLR1/2 ligand), a synthetic diacylated lipoprotein FSL-1 (TLR2/6 ligand), all from InvivoGen (San Diego, CA); ZVAD (Z-Tyr-Val-Ala-Dl-Asp-Fluoromethylketone, Bachem Americas, Inc., Torrance, CA), Kineret (Amgen, Thousand Oaks, CA), human recombinant IL-1β (rIL-1β, Endotoxin level <0.1 ng/µg, Life Technologies, Grand Island, NY), and dynasore (Enzo).

The soluble TNF receptor, Etanercept (Enbrel; Wyeth Europa), which is known to fully inhibit the biological function of murine TNF (Fei et al., 2011 (link); Ali et al., 2015b (link)), was used in the experiment as anti-TNF treatment. Subcutaneous (s.c.) injections of Etanercept (0.2 mg/mouse in 0.1 ml of PBS) were performed. The administration began 2 days prior to exposing the knee joints with Lpl1(+sp), and continued on the day the knee joints where i.a. injected with Lpl1(+sp), and were followed by every other day until the termination date. PBS was s.c. injected as control treatment on days when Etanercept was not administered.
To block the function of IL-1 in the murine model, the IL-1 receptor antagonist Anakinra (Kineret; Amgen) was used as anti-IL-1 treatment (Sgroi et al., 2011 (link); Ali et al., 2015a (link)). Anakinra (1 mg/mouse in 0.1 ml of PBS) was s.c. administered daily, starting 2 days prior to exposing the knee joints with Lpl1(+sp), and continued until the termination date.
Phosphate-buffered saline was administered s.c. daily in the control treatment group, starting 2 days prior to exposing the knee joints with Lpl1(+sp), and continued until termination day.

IL-1Ra (anakinra, trade name Kineret^®) was a gift from Amgen Inc. (Thousand Oaks, CA). Our study involved daily subcutaneous injections of two groups of animals: (a) a control mdx group, dosed with 0.9% NaCl (n = 6), and (b) a drug-treated mdx group, which received IL-1Ra at 25 mg/kg/day in a 50-μL volume of 0.9% NaCl (n = 10). Mice were randomized on the basis of body mass and were treated for 45 days, beginning at 5 weeks of age ending when they were 12 weeks old. IL-1Ra and the vehicle were injected subcutaneously, 7 days a week.