Physiological and Pathological Mitochondrial Clearance Is Related to Pectoralis Major Muscle Pathogenesis in Broilers With Wooden Breast Syndrome

Wooden breast syndrome (WB) constitutes an emerging myopathy in the pectoralis major muscle (PM) of broiler chickens, characterized by myofiber hypertrophy and degeneration along with severe fibrosis. WB pathogenesis has been considered to involve hypoxia induced by rapid growth of the PM. In this study, we focused on mitochondrial morphology and dynamics in the myofibers, as these organelles are sensitive to damage by hypoxia, and examined the effects on WB pathogenesis. Specifically, the PMs of a flock of 35 broilers at 50 days of age were evaluated. First, the severity of disease in each bird was determined by measuring histopathological indices including the fibrotic area (FA) in the muscle and circularity of myofibers (CM). These values were 29.4 ± 9.6% and 0.70 ± 0.042, respectively, showing variety among the flock. Myofiber vacuolization was observed in all birds including numerous small- or large-rimmed vacuoles, with the former consisting of ultrastructurally autophagosome-like vacuoles engulfing degenerated mitochondria. The large-rimmed vacuoles frequently occurred in the PMs with more severe FA and CM, indicating a relationship between altered autophagy/mitophagy and WB severity. Next, the expression levels of hypoxia-adaptive and mitochondrial dynamics-related genes were analyzed, and their correlations with the histopathological indices were examined. The histopathological indices were negatively correlated with the expression of vascular endothelial growth factor A (VEGFA), indicating that less angiogenesis owing to weakened hypoxia-inducible factor signaling induces more severe WB pathology. In addition, the observed negative correlation with mitochondrial dynamics-related genes implied that WB pathology deteriorates concomitant with reduced mitochondrial dynamics. Furthermore, the expression of mitochondrial dynamics-related genes showed strong positive correlation with that of VEGFA and autophagy-/mitophagy-related genes. These results revealed that the PMs of broilers possess the mechanism of physiological clearance of mitochondria damaged by the hypoxia resulting from the continuous mitochondrial dynamics and autophagy/mitophagy accompanying rapid PM growth. In turn, the altered mitochondrial clearance induced by chronic hypoxia and the accumulation of damaged mitochondria likely underly the severe pathological features of WB.ArticleFrontiers in Pharmacology.11:579(2020)journal articl

Oocyte cumulus complex quality and oviduct transportation velocity in systemic autoimmune disease model mice

Oocyte transportation by the oviduct involves the interaction between ciliated epithelial cells and cumulus cells. To determine whether the quality of cumulus-oocyte complexes (COCs) changes the transportation property of COCs, we compared the transportation velocity of COCs (TVC) by the infundibulum ex vivo with various combinations of infundibula and COCs collected from different mice. We used young and aged C57BL/6N and MRL/MpJ, and MRL/MpJ-Fas (lpr/lpr) mice as the strains with intact female reproductive function and the systemic autoimmune disease model exhibiting oocyte pick-up dysfunction owing to the morphofunctional abnormality of ciliated epithelium, respectively. The TVC of aged MRL strains was less than that of aged C57BL/6N mice, suggesting that aging affects the transportation of COCs in MRL strains. The TVC of aged MRL/MpJ-Fas( lpr/lpr ) mice was the least among all examined combinations, whereas the TVC accelerated when the infundibulum or COCs were collected from other strains. These results indicate that the transportation property of COCs is determined not only by the ciliary function in the infundibulum but also by the properties of COCs

Altered ciliary morphofunction in the oviductal infundibulum of systemic autoimmune disease-prone MRL/MpJ-Fas(lpr/lpr) mice

According to our previous reports, impaired oocyte pickup was observed in the oviductal infundibulum of an autoimmune disease (AD) mouse model, suggesting a relationship between female infertility and AD. This study examines the relationship between AD and infundibulum morphofunction by focusing on the epithelial cilia. Healthy MRL/MpJ and AD-prone MRL/MpJ-Fas(lpr/lpr) mice were examined at 3 and 6 months of age, representing early and late disease stages, respectively. Oocyte pickup indices decreased with AD progression indicated by splenomegaly, autoantibody production and increased T cell counts of infundibulum mucosa in MRL/MpJ-Fas(lpr/lpr) mice. Ciliary beating frequency (CBF) and height in the infundibulum were faster and higher in MRL/MpJ-Fas(lpr/lpr) mice than in MRL/MpJ mice at the early AD stages, although the absolute CBF values were lower at the late AD stage. At the late stage, ciliary height did not differ between mouse lines but the morphological index of cilia beating direction indicated randomized patterns in MRL/MpJ-Fas(lpr/lpr) mice. The tracheal mucosa was also examined as a representative example of cilia morphology; its CBF decreased at the late AD stage in MRL/MpJ-Fas(lpr/lpr); however, there were no AD-related morphological changes. Our results demonstrate altered cilia motility in systemic and reproductive organs, with such morphological changes of the infundibulum likely impairing function, including oocyte pickup

Nutrition During the Early Rearing Period Affects the Incidence of Wooden Breasts in Broilers

This study aimed to evaluate the relationship between early nutrition and the incidence of wooden breasts (WB) in broilers. Sixteen male and twenty female neonatal ROSS 308 broiler chicks were divided equally into four flocks. From 0–12 days of age, starter diet H, composed of 22.4% crude protein (CP), 6.6% crude fat (CF), 1.25% lysine, 0.48% methionine, and ≥3,070 kcal/kg metabolizable energy (ME), was fed to two flocks, and starter diet L, composed of 19.9% CP, 2.5% CF, 1.04% lysine, 0.38% methionine, and ≥2,930 kcal/kg ME, was fed to the remaining two flocks. All the flocks were fed the same commercial finisher diet, composed of 20.3% CP, 7.5% CF, 1.18% lysine, 0.44% methionine, and ≥3,300 kcal/kg ME, from 12–47 days of age. The birds were weighed every 2–5 days, subjected to a wing-lift test, and histology was conducted on the pectoralis major muscle tissue samples from all the birds necropsied at 47 days of age. Significant differences in the mean body weight between groups H and L were observed during 6–16 days and 24–26 days of age in males and during 6–26 days of age in females. Regarding the score evaluation of the individual lesions reflecting wooden breast, the birds in which back-to-back wing contact was not possible had higher lesion scores than those in which back-to-back wing contact was possible. The absence of back-to-back wing contact appeared more frequently in flocks fed the starter diet L, particularly in males. These results indicate that inappropriate nutrition levels in the starter diet increase the incidence of WB. Therefore, avoiding early nutrition deficits is a cost-effective feeding strategy

Expression of Indian hedgehog signaling in murine oviductal infundibulum and its relationship with epithelial homeostasis

Homeostasis of the oviductal infundibulum epithelium is continuously regulated by signaling pathways under physiological and pathological conditions. Herein, we investigated the expression of hedgehog (Hh) signaling-related components in the murine oviductal infundibulum, which is known to maintain homeostasis in the adult epithelium. Additionally, using autoimmune disease-prone MRL/MpJ-Fas(lpr/lpr) (MRL/lpr) mice showing abnormal morphofunction of the ciliated epithelium of the infundibulum related to the oviductal inflammation, we examined the relationship between Hh signaling and pathology of the infundibulum. The expression and localization of Pax8, a marker for progenitor cells in the oviductal epithelium, and Foxj1, a marker for ciliogenesis, were examined in the infundibulum. The results showed that Pax8 was downregulated and Foxj1 was upregulated with aging, suggesting that homeostasis of the infundibulum epithelium of MRL/lpr mice was disturbed at 6 months of age. In all mice, the motile cilia of ciliated epithelial cells in the infundibulum harbored Hh signaling pathway-related molecules: patched (Ptch), smoothened (Smo), and epithelial cells harbor Gli. In contrast, Ptch, Smo, and Gli2 were significantly downregulated in the infundibulum of MRL/lpr mice at 6 months of age. The expression levels of Pax8 and Foxj1 were significantly positively correlated with those of Ptch1, Smo, and Gli2. Hh signaling is thought to be involved in homeostasis of the ciliated epithelium in the infundibulum. In MRL/lpr mice, which show exacerbated severe systemic autoimmune abnormalities, molecular alterations in Hh signaling-related components are considered to interact with local inflammation in the infundibulum, leading to disturbances in epithelial homeostasis and reproductive function

Nutrition During the Early Rearing Period Affects the Incidence of Wooden Breasts in Broilers

This study aimed to evaluate the relationship between early nutrition and the incidence of wooden breasts (WB) in broilers. Sixteen male and twenty female neonatal ROSS 308 broiler chicks were divided equally into four flocks. From 0–12 days of age, starter diet H, composed of 22.4% crude protein (CP), 6.6% crude fat (CF), 1.25% lysine, 0.48% methionine, and ≥3,070 kcal/kg metabolizable energy (ME), was fed to two flocks, and starter diet L, composed of 19.9% CP, 2.5% CF, 1.04% lysine, 0.38% methionine, and ≥2,930 kcal/kg ME, was fed to the remaining two flocks. All the flocks were fed the same commercial finisher diet, composed of 20.3% CP, 7.5% CF, 1.18% lysine, 0.44% methionine, and ≥3,300 kcal/kg ME, from 12–47 days of age. The birds were weighed every 2–5 days, subjected to a wing-lift test, and histology was conducted on the pectoralis major muscle tissue samples from all the birds necropsied at 47 days of age. Significant differences in the mean body weight between groups H and L were observed during 6–16 days and 24–26 days of age in males and during 6–26 days of age in females. Regarding the score evaluation of the individual lesions reflecting wooden breast, the birds in which back-to-back wing contact was not possible had higher lesion scores than those in which back-to-back wing contact was possible. The absence of back-to-back wing contact appeared more frequently in flocks fed the starter diet L, particularly in males. These results indicate that inappropriate nutrition levels in the starter diet increase the incidence of WB. Therefore, avoiding early nutrition deficits is a cost-effective feeding strategy

Compartmentalization of interleukin 36 subfamily according to inducible and constitutive expression in the kidneys of a murine autoimmune nephritis model

The interleukin (IL) 36 subfamily belongs to the IL-1 family and is comprised of agonists (IL-36 alpha, IL-36 beta, IL-36 gamma) and antagonists (IL-36Ra, IL-38). We previously reported IL-36 alpha overexpression in renal tubules of chronic nephritis mice. To understand the localization status and biological relationships among each member of the IL-36 subfamily in the kidneys, MRL/MpJ-Fas(lpr/lpr) mice were investigated as autoimmune nephritis models using pathology-based techniques. MRL/MpJ-Fas(lpr/lpr) mice exhibited disease onset from 3 months and severe nephritis at 6-7 months (early and late stages, respectively). Briefly, IL-36 gamma and IL-36Ra were constitutively expressed in murine kidneys, while the expression of IL-36 alpha, IL-36 beta, IL-36Ra, and IL-38 was induced in MRL/MpJ-Fas(lpr/lpr) mice. IL-36 alpha expression was significantly increased and localized to injured tubular epithelial cells (TECs). CD44(+)-activated parietal epithelial cells (PECs) also exhibited higher IL-36 alpha-positive rates, particularly in males. IL-36 beta and IL-38 are expressed in interstitial plasma cells. Quantitative indices for IL-36 alpha and IL-38 positively correlated with nephritis severity. Similar to IL-36 alpha, IL-36Ra localized to TECs and PECs at the late stage; however, MRL/MpJ-Fas(lpr/lpr) and healthy MRL/MpJ mice possessed IL-36Ra(+) smooth muscle cells in kidney arterial tunica media at both stages. IL-36 gamma was constitutively expressed in renal sympathetic axons regardless of strain and stage. IL-36 receptor gene was ubiquitously expressed in the kidneys and was induced proportional to disease severity. MRL/MpJ-Fas(lpr/lpr) mice kidneys possessed significantly upregulated IL-36 downstream candidates, including NF-kappa B- or MAPK-pathway organizing molecules. Thus, the IL-36 subfamily contributes to homeostasis and inflammation in the kidneys, and especially, an IL-36 alpha-dominant imbalance could strongly impact nephritis deterioration