Daily Eicosapentaenoic Acid Infusion in IUGR Fetal Lambs Reduced Systemic Inflammation, Increased Muscle ADRß2 Content, and Improved Myoblast Function and Muscle Growth.

Intrauterine growth-restricted (IUGR) fetuses exhibit systemic inflammation that contributes to programmed deficits in myoblast function and muscle growth. Thus, we sought to determine if targeting fetal inflammation improves muscle growth outcomes. Heat stress-induced IUGR fetal lambs were infused with eicosapentaenoic acid (IUGR+EPA; n = 9) or saline (IUGR; n = 8) for 5 days during late gestation and compared to saline-infused controls (n = 11). Circulating eicosapentaenoic acid was 42% less (p < 0.05) for IUGR fetuses but was recovered in IUGR+EPA fetuses. The infusion did not improve placental function or fetal O2 but resolved the 67% greater (p < 0.05) circulating TNFα observed in IUGR fetuses. This improved myoblast function and muscle growth, as the 23% reduction (p < 0.05) in the ex vivo differentiation of IUGR myoblasts was resolved in IUGR+EPA myoblasts. Semitendinosus, longissimus dorsi, and flexor digitorum superficialis muscles were 24-39% lighter (p < 0.05) for IUGR but not for IUGR+EPA fetuses. Elevated (p < 0.05) IL6R and reduced (p < 0.05) ß2 adrenoceptor content in IUGR muscle indicated enhanced inflammatory sensitivity and diminished ß2 adrenergic sensitivity. Although IL6R remained elevated, ß2 adrenoceptor deficits were resolved in IUGR+EPA muscle, demonstrating a unique underlying mechanism for muscle dysregulation. These findings show that fetal inflammation contributes to IUGR muscle growth deficits and thus may be an effective target for intervention.

Daily injection of the ß2 adrenergic agonist clenbuterol improved poor muscle growth and body composition in lambs following heat stress-induced intrauterine growth restriction.

Background: Intrauterine growth restriction (IUGR) is associated with reduced ß2 adrenergic sensitivity, which contributes to poor postnatal muscle growth. The objective of this study was to determine if stimulating ß2 adrenergic activity postnatal would rescue deficits in muscle growth, body composition, and indicators of metabolic homeostasis in IUGR offspring. Methods: Time-mated ewes were housed at 40°C from day 40 to 95 of gestation to produce IUGR lambs. From birth, IUGR lambs received daily IM injections of 0.8 µg/kg clenbuterol HCl (IUGR+CLEN; n = 11) or saline placebo (IUGR; n = 12). Placebo-injected controls (n = 13) were born to pair-fed thermoneutral ewes. Biometrics were assessed weekly and body composition was estimated by ultrasound and bioelectrical impedance analysis (BIA). Lambs were necropsied at 60 days of age. Results: Bodyweights were lighter (p ≤ 0.05) for IUGR and IUGR+CLEN lambs than for controls at birth, day 30, and day 60. Average daily gain was less (p ≤ 0.05) for IUGR lambs than controls and was intermediate for IUGR+CLEN lambs. At day 58, BIA-estimated whole-body fat-free mass and ultrasound-estimated loin eye area were less (p ≤ 0.05) for IUGR but not IUGR+CLEN lambs than for controls. At necropsy, loin eye area and flexor digitorum superficialis muscles were smaller (p ≤ 0.05) for IUGR but not IUGR+CLEN lambs than for controls. Longissimus dorsi protein content was less (p ≤ 0.05) and fat-to-protein ratio was greater (p ≤ 0.05) for IUGR but not IUGR+CLEN lambs than for controls. Semitendinosus from IUGR lambs had less (p ≤ 0.05) ß2 adrenoreceptor content, fewer (p ≤ 0.05) proliferating myoblasts, tended to have fewer (p = 0.08) differentiated myoblasts, and had smaller (p ≤ 0.05) muscle fibers than controls. Proliferating myoblasts and fiber size were recovered (p ≤ 0.05) in IUGR+CLEN lambs compared to IUGR lambs, but ß2 adrenoreceptor content and differentiated myoblasts were not recovered. Semitendinosus lipid droplets were smaller (p ≤ 0.05) in size for IUGR lambs than for controls and were further reduced (p ≤ 0.05) in size for IUGR+CLEN lambs. Conclusion: These findings show that clenbuterol improved IUGR deficits in muscle growth and some metabolic parameters even without recovering the deficit in ß2 adrenoreceptor content. We conclude that IUGR muscle remained responsive to ß2 adrenergic stimulation postnatal, which may be a strategic target for improving muscle growth and body composition in IUGR-born offspring.

Primary myoblasts from intrauterine growth-restricted fetal sheep exhibit intrinsic dysfunction of proliferation and differentiation that coincides with enrichment of inflammatory cytokine signaling pathways.

Intrauterine growth restriction (IUGR) is linked to lifelong reductions in muscle mass due to intrinsic functional deficits in myoblasts, but the mechanisms underlying these deficits are not known. Our objective was to determine if the deficits were associated with changes in inflammatory and adrenergic regulation of IUGR myoblasts, as was previously observed in IUGR muscle. Primary myoblasts were isolated from IUGR fetal sheep produced by hyperthermia-induced placental insufficiency (PI-IUGR; n = 9) and their controls (n = 9) and from IUGR fetal sheep produced by maternofetal inflammation (MI-IUGR; n = 6) and their controls (n = 7). Proliferation rates were less (P < 0.05) for PI-IUGR myoblasts than their controls and were not affected by incubation with IL-6, TNF-α, norepinephrine, or insulin. IκB kinase inhibition reduced (P < 0.05) proliferation of control myoblasts modestly in basal media but substantially in TNF-α-added media and reduced (P < 0.05) PI-IUGR myoblast proliferation substantially in basal and TNF-α-added media. Proliferation was greater (P < 0.05) for MI-IUGR myoblasts than their controls and was not affected by incubation with TNF-α. Insulin increased (P < 0.05) proliferation in both MI-IUGR and control myoblasts. After 72-h differentiation, fewer (P < 0.05) PI-IUGR myoblasts were myogenin+ than controls in basal and IL-6 added media but not TNF-α-added media. Fewer (P < 0.05) PI-IUGR myoblasts were desmin+ than controls in basal media only. Incubation with norepinephrine did not affect myogenin+ or desmin+ percentages, but insulin increased (P < 0.05) both markers in control and PI-IUGR myoblasts. After 96-h differentiation, fewer (P < 0.05) MI-IUGR myoblasts were myogenin+ and desmin+ than controls regardless of media, although TNF-α reduced (P < 0.05) desmin+ myoblasts for both groups. Differentiated PI-IUGR myoblasts had greater (P < 0.05) TNFR1, ULK2, and TNF-α-stimulated TLR4 gene expression, and PI-IUGR semitendinosus muscle had greater (P < 0.05) TNFR1 and IL6 gene expression, greater (P < 0.05) c-Fos protein, and less (P < 0.05) IκBα protein. Differentiated MI-IUGR myoblasts had greater (P < 0.05) TNFR1 and IL6R gene expression, tended to have greater (P = 0.07) ULK2 gene expression, and had greater (P < 0.05) ß-catenin protein and TNF-α-stimulated phosphorylation of NFκB. We conclude that these enriched components of TNF-α/TNFR1/NFκB and other inflammatory pathways in IUGR myoblasts contribute to their dysfunction and help explain impaired muscle growth in the IUGR fetus.

Myoblasts are stems cells whose functional capacity can limit muscle growth. However, stressful intrauterine conditions cause these cells to be intrinsically dysfunctional. This restricts muscle growth capacity, leading to intrauterine growth restriction (IUGR) of the fetus, low birth weight, and less muscle mass after birth. Consequently, meat yield is reduced in IUGR-born food animals and glucose homeostasis is impaired in IUGR-born humans, which contributes to metabolic dysfunction. Intrinsic dysfunction of IUGR myoblasts has been previously observed, but the fetal programming changes (i.e., permanent changes in the development of cellular mechanisms that explains different functional outcomes) have not been identified. This study shows that one mechanism is the enhancement of signaling pathways for TNF-α and other inflammatory cytokines. These cytokines have roles in stress responses and regulation of muscle growth. Programmed enhancement of these pathways means that IUGR myoblasts are more responsive to even normal amounts of circulating cytokines. Unfortunately, the primary response of myoblasts to cytokines is slower differentiation (i.e., cellular transformation necessary for muscle growth). Programmed enhancement of this response directly impedes myoblast-dependent muscle growth, and the deficit is lifelong. However, identifying this mechanism is a fundamental step for developing strategies to improve muscle growth in low birth weight offspring.

Inflammatory Mediation of Heat Stress-Induced Growth Deficits in Livestock and Its Potential Role as a Target for Nutritional Interventions: A Review.

Heat stress is detrimental to well-being and growth performance in livestock, and systemic inflammation arising during chronic heat stress contributes to these poor outcomes. Sustained exposure of muscle and other tissues to inflammation can impair the cellular processes that facilitate muscle growth and intramuscular fat deposition, thus reducing carcass quality and yield. Climate change is expected to produce more frequent extreme heat events, increasing the potential impact of heat stress on sustainable livestock production. Feedlot animals are at particularly high risk for heat stress, as confinement limits their ability to seek cooling from the shade, water, or breeze. Economically practical options to circumvent heat stress in feedlot animals are limited, but understanding the mechanistic role of inflammation in heat stress outcomes may provide the basis for treatment strategies to improve well-being and performance. Feedlot animals receive formulated diets daily, which provides an opportunity to administer oral nutraceuticals and other bioactive products to mitigate heat stress-induced inflammation. In this review, we examine the complex associations between heat stress, systemic inflammation, and dysregulated muscle growth in meat animals. We also present evidence for potential nutraceutical and dietary moderators of inflammation and how they might improve the unique pathophysiology of heat stress.