Chloroquine Downregulation of Intestinal Autophagy to Alleviate Biological Stress in Early-Weaned Piglets

Abstract

Early weaning stress impairs the development of gastrointestinal barrier function, causing immune system dysfunctions, reduction in feed intake, and growth retardation. Autophagy was hypothesized to be a key underlying cellular process in these dysfunctions. We conjectured that rapamycin (RAPA) and chloroquine (CQ), as two autophagy-modifying agents, regulate the autophagy process and may produce deleterious or beneficial effects on intestinal health and growth. To explore the effect of autophagy on early weaning stress in piglets, 18 early-weaned piglets were assigned to three treatments (each treatment of six piglets) and treated with an equal volume of RAPA, CQ, or saline. The degree of autophagy and serum concentrations of immunoglobulins and cytokines, as well as intestinal morphology and tight junction protein expression, were evaluated. Compared with the control treatment, RAPA-treated piglets exhibited activated autophagy and had decreased final body weight (BW) and average daily gain (ADG) (p < 0.05), impaired intestinal morphology and tight junction function, and higher inflammatory responses. The CQ-treated piglets showed higher final BW, ADG, jejuna and ileal villus height, and lower autophagy and inflammation, compared with control piglets (p < 0.05). Throughout the experiment, CQ treatment was beneficial to alleviate early weaning stress and intestinal and immune system dysfunction.

Chloroquine Improves Deoxynivalenol-Induced Inflammatory Response and Intestinal Mucosal Damage in Piglets

“…Western Blotting Analysis. Relative protein levels for Sequestosome 1 (P62/SQSTM1, P62), LC3, and β-actin in the jejunum were determined followed the methods of Simeng [23]. The primary antibodies used in the present study were as follows: anti-P62 (#23214), anti-LC3B, and anti-β-actin (#4970).…”

“…Permeability Related Indicators. We followed the methods of Simeng Liao et al [23] 2.7. Intestinal Morphology.…”

“…Intestinal Morphology. Intestinal morphology analyses were followed by the methods of [23]. Crypt depth, villous height, intestine wall thickness, goblet cell, and lymphocyte measured and counted with computer-assisted microscopy (Micro metrics TM; Nikon ECLIPSE E200, Tokyo, Japan).…”

“…Quantitative real-time PCR (qRT-PCR) was followed by the methods of [23]. In brief, total RNA was extracted using the Trizol reagent (Invitrogen, Thermo Fisher Scientific, USA).…”

“…The cDNA library was constructed using the TransScript One-Step gDNA Removal and cDNA Synthesis SuperMix kit (Transgen, China), then diluted as instructions. Amplification conditions were performed as follows: (1) denaturation for 10 min at 95°C; (2) 40 PCR cycles of denaturation for 15 s at 95°C, annealing for 15 s at 56-64°C and extension for 45 s at 56-64°C[23].…”