AICAr, a Widely Used AMPK Activator with Important AMPK-Independent Effects: A Systematic Review

Unless otherwise indicated, for immunocytochemical studies, TA and IC muscles and lumbar spinal cords were fixed by immersion in 4% paraformaldehyde in 0.1 M PB, pH 7.4, either for 2 h (in the case of TA and IC muscles) or overnight (in the case of spinal cords), and cryoprotected. Tissue samples were embedded in Tissue Freezing Medium (Triangle Biomedical Sciences) and frozen. Longitudinal (for TA and IC muscles) or transverse (for spinal cord) serial cryostat sections (16-μm thick) were obtained and stored at –80 °C. Immunocytochemical analysis of muscle fiber type composition was performed on unfixed TA muscles. For this, muscles were embedded in tragacanth gum, snap-frozen in liquid N2-cooled isopentane, and sectioned transversely (10-μm thick) on a cryostat.

• We found that knockout of Nrf2 limited the ability of AICAR to reduce the severity of PALI in mice (Figures 7A–E).
• Thus, the phosphorylation of a specific site can be semiquantitatively compared between different samples by normalizing the ion intensities of the phosphopeptides of interest to those of their cognate nonphosphopeptides, which serve as an internal standard.
• This is in contrast to the absence of changes in the cross-sectional area of myofibers reported in the dystrophic muscle of mdx mice following chronic treatment with AICAR 51.

At necropsy in the animals treated with HFD, a large number of fatty deposits in the abdominal cavity was revealed, as well as hydronephrosis of one of the kidneys. In the animals treated with HFD, there was a decrease in the mass of the liver, adrenal glands, and pancreas, as well as a significant increase in the mass of adipose tissue surrounding the epididymis. The introduction of AICAR, both alone and in combination with Methotrexate, reduced the body weight and body weight gain relative to animals on HFD, starting from the ninth week of the study. The feed intake in HFD-fed AICAR-treated animals was slightly higher than HFD-fed animals without treatment, indicating some normalization of this indicator.

5. AICAR Inhibits the Growth of Prostate Cancer Cells Through Activating an Ampk/Mtor-Dependent Pathway

It will be interesting https://killingtondistillery.com/supporting-athletes-in-overcoming-physical-and/ to explore if AICAR-induced MUC1-CT instability is correlated to MUC1-CT ubiquitination in the future. Ubiquitous expression of AMPKα1-, β1- and γ1-subunits in many tissues makes the α1β1γ1 complex a reference for AMPK assays to identify AMPK activators. However, given the unique functions and/or subcellular (or tissue)-specific distribution of the distinct AMPK complex,3, 4, 5 referencing screening to the α1β1γ1 complex may present a limited range of the physiology of AMPK. In 2003, Campas et al. reported that AICAr activates AMPK and induces apoptosis in primary samples of B-cell chronic lymphocytic leukemia (CLL) in vitro 11.

The normalized data were then consolidated, and the Python software was utilized for pattern recognition. Following unit variance (UV) scaling preprocessing, in-depth data analysis was performed. In order to ensure accurate results for the detection of AICAR in urine samples, it was necessary to validate the method for quantification purposes. The validation parameters were the selectivity, carry-over, linearity range, precision (intra/inter-day), accuracy, LOD, LOQ, matrix effect, recovery, and stability. QC samples were prepared by spiking human urine with working solutions and stock solutions reaching concentrations at LLOQ (10 ng mL−1), LQC (100 ng mL−1), MQC (1000 ng mL−1) and HQC (5000 ng mL−1) levels.

Thermal stability assay

For the quantification of invasiveness, crystal violet was dissolved by 50% methanol and the absorbance was determined at 570 nm using a microplate reader (BioTek Instruments, Inc.). T cell survival was analyzed by flow cytometric staining for Annexin V (BD Bioscience) and 7-AAD (BD Bioscience) at different conditions. For in vitro activation, cells from lymph nodes were activated with PMA/Ionomycin at indicated time points and collected for analysis. Surface and intracellular staining were performed as what we previously described 38, 39. The following antibodies were used for surface staining, which included anti-CD4 (clone RM4-5), anti-CD8 (clone 53-6.7), anti-CD69 (clone H1.2F3), anti-CD25 (clone PC61), anti-CD71 (clone RI7217). For intracellular staining of cytokines, LN cells with different treatments were stimulated with PMA/Ionomycin and golgiplug (BD Bioscience) for 5 hours.

Cell culture and cell lines

Tubes were then centrifuged on an SW60 Ti rotor (Beckman) at 35,000 rpm for 1 h at 4 °C, and the top fractions (about 200 μl) were collected as cytosolic fraction. For hepatic ischemia, surgery was performed as described previously72 with some modifications. Mice were anesthetized with pentobarbital sodium solution (100 mg/kg) by intraperitoneal injection and were placed on a heating plate set at 37 °C. The mouse abdomen was gently opened (from the mid-abdomen to the xiphoid) to expose the liver, and its intestine was covered by moistened gauze to prevent from drying. The portal vein, hepatic artery, and bile duct were then cross-clamped by an atraumatic clip at a place just above the branching to the right lateral lobe to subject the median and the left lateral lobes to ischemia. Mice that underwent the same surgery but did not undergo cross-clamping on the blood vessel and bile duct were considered as shams.

Recently, AMPK has been identified as an important factor regulating muscle fiber contractile gene expression and endurance (Narkar et al. 2008). Treatment with AMPK agonist AICAR increased running stamina by 45% in sedentary animals (Narkar et al. 2008) and enhanced spatial memory function in young mice (Kobilo et al. 2011). As AICAR has very low permeability across the blood–brain barrier (Marangos et al. 1990), effects on stamina and cognition are likely indirect. The best way to assess the role of AMPK in the effects of AICAr in vivo could be provided by AMPK knockout mice. As shown in Table 1, data obtained by transgenic mice models revealed that AICAr-mediated effects on glucose uptake in skeletal muscle cells 32,33,34,35, lipogenesis and fatty acid oxidation in the liver 36, and decreased fat synthesis in adipose cells were AMPK-dependent 37. However, a decrease in gluconeogenesis and an inhibition of oxidative phosphorylation (OXPHOS) can be observed in response to AICAr in mice lacking both AMPKα1 and α2 isoforms 38,39,40.