Chronic heat stress induces renal fibrosis and mitochondrial dysfunction in laying hens

Abstract

Background: Heat stress in laying hens negatively affects egg production and shell quality by disrupting the homeostasis of plasma calcium and phosphorus levels. Although the kidney plays an important role in calcium and phosphorus homeostasis, evidence regarding the effect of heat stress on renal injury in laying hens is yet to be elucidated. Therefore, the aim of this study was to evaluate the effects of chronic heat stress on renal damage in hens during laying periods.

Methods: A total of 16 white-leghorn laying hens (32 weeks old) were randomly assigned to two groups (n = 8). One group was exposed to chronic heat stress (33 °C for 4 weeks), whereas the other group was maintained at 24 °C.

Results: Chronic heat exposure significantly increased plasma creatinine and decreased plasma albumin levels (P < 0.05). Heat exposure also increased renal fibrosis and the transcription levels of fibrosis-related genes (COLA1A1, αSMA, and TGF-β) in the kidney. These results suggest that renal failure and fibrosis were induced by chronic heat exposure in laying hens. In addition, chronic heat exposure decreased ATP levels and mitochondrial DNA copy number (mtDNA-CN) in renal tissue, suggesting that renal mitochondrial dysfunction occurs under conditions of heat stress. Damaged mitochondria leak mtDNAs into the cytosol and mtDNA leakage may activate the cyclic GMP-AMP synthase (cGAS) stimulator of interferon genes (STING) signaling pathway. Our results showed that chronic heat exposure activated the cGAS-STING pathway as indicated by increased expression of MDA5, STING, IRF7, MAVS, and NF-κB levels. Furthermore, the expression of pro-inflammatory cytokines (IL-12) and chemokines (CCL4 and CCL20) was upregulated in heat-stressed hens.

Conclusions: These results suggest that chronic heat exposure induces renal fibrosis and mitochondrial damage in laying hens. Mitochondrial damage by heat stress may activate the mtDNA-cGAS-STING signaling and cause subsequent inflammation, which contributes to the progression of renal fibrosis and dysfunction.

Keywords: Heat stress; Laying hens; Mitochondrial DNA; Renal fibrosis; cGAS-STING.

Fig. 1

Effect of chronic heat stress on renal function and histology in laying hens. a Plasma levels of blood urea nitrogen (BUN), creatine, and albumin in the control (n = 8) or heat stress for 4 weeks (n = 8) hen groups. Data are presented as the mean ± SEM. Statistical analysis was performed by a Student’s t-test. ^*P < 0.05 was considered statistically significant. b Representative histological images of Masson’s trichrome stained sections of the control and heat-stressed hen kidneys (upper). Morphometric analysis of the fractional cortical tubular area of Masson’s trichrome stained kidney images (lower). n = 8 for each group. Data are presented as the percentage of the total cortex and mean ± SEM. Statistical analysis was performed using a Student’s t-test. Bars = 100 µm. c The mRNA expression levels of COL1A1, COL1A2, αSMA, and TGFβ were measured by performing real-time PCR and normalized to 18S rRNA. n = 8 in each group. Statistical analysis was performed by a Student’s t-test

Fig. 2

Effect of chronic heat stress on the functionality of mitochondria in the kidney. a ATP levels in renal tissue. n = 8 in each group. Data are presented as the percentage of the total cortex and as the mean ± SEM. Statistical analysis was performed using a Student’s t-test. b mtDNA copy number of renal tissues in laying hens. mt/nucDNA = mtDNA relative to nuclear DNA (β-actin) copy number. ^*P < 0.05

Fig. 3

Effects of chronic heat stress on the expression of STING, NF-kB pathway, and related genes. a mRNA levels of MDA5, STING, IRF7, MAVS, and NF-kB, b mRNA levels of pro-inflammatory cytokines (IFN-β, IFN-γ, IL-1β, and IL-12) and chemokines (IL-8, CCL2, CCL4 and CCL20). The mRNA expression levels were quantified by real-time quantitative PCR and normalized to 18S rRNA. n = 8 in each group. Statistical analysis was performed using a Student’s t-test. ^*P < 0.05