Impact of fetal exposure to mycotoxins on longissimus muscle fiber hypertrophy and miRNA profile.

BACKGROUND: Longissimus muscle samples were collected from lambs exposed in utero to mycotoxins [E-, endophyte-free tall fescue seed without ergot alkaloids (negative control) or E + , endophyte-infected tall fescue seed containing ergot alkaloids] during mid-gestation (MID; E + /E-) or late-gestation (LATE; E-/E +) harvested at two developmental stages (FETAL, gestational d133) or (MAT, near maturity, 250 d of age; n = 3/treatment/developmental stage). Muscle samples were examined to determine the impact of in utero mycotoxin exposure on skeletal muscle fiber hypertrophy and the miRNA profile at FETAL and MAT. RESULTS: Longissimus weight was greater (P < 0.05) in E + /E- lambs compared to E-/E + lambs at MAT; however, FETAL longissimus weight did not differ (P > 0.10) between fescue treatments. Type I fiber cross sectional area was larger (P < 0.10) for E + /E- than E-/E + at MAT but did not differ (P > 0.10) between fescue treatments at FETAL. Type II fiber area was larger (P < 0.05) at MAT in E + /E- compared to E-/E + but did not differ (P < 0.05) between fescue treatments at FETAL. Cross-sectional Type I and Type II longissimus muscle fiber area increased (P < 0.05) from FETAL to MAT by 6.86-fold and 10.83-fold, respectively. The ratio of Type II:Type I muscle fibers was lower (P = 0.04) at MAT compared to FETAL. There were 120 miRNA differentially expressed (q < 0.05) between FETAL and MAT. Maternal fescue treatment did not alter (q > 0.05) expression of miRNAs in the longissimus muscle. miR-133, -29a, -22-3p, and -410-3p were identified as highly significant with a log2 fold change > 4. In vitro satellite cell cultures showed that selected miRNAs (miR-22-3p, 29a, 27a, and 133a) are differentially regulated during proliferation and differentiation indicating a role of miRNA in muscle hypertrophy. CONCLUSIONS: Exposure to mycotoxins did not alter fiber type but had long-term impacts on postnatal muscle hypertrophy and cross-sectional area. The miRNA profile of the longissimus was not altered by Maternal mycotoxin exposure at FETAL or MAT. Developmental age altered the miRNA transcriptome and mRNA expression of known genes related to muscle growth. These results indicate that Maternal exposure to E + fescue seed during LATE gestation can alter postnatal muscle hypertrophy in sheep; however, these changes are not regulated by the miRNA transcriptome of the longissimus muscle.

Use of AgomiR and AntagomiR technologies to alter satellite cell proliferation in vitro, miRNA expression, and muscle fiber hypertrophy in intrauterine growth-restricted lambs.

Introduction: microRNAs (miRNAs) are small non-coding RNAs that work at the posttranscriptional level to repress gene expression. Several miRNAs are preferentially expressed in skeletal muscle and participate in myogenesis. This research was conducted to alter endogenous miRNA expression in skeletal muscle to promote muscle hypertrophy. Methods: Two experiments were conducted using mimic/agomiR or antagomir technologies to alter miRNA expression and examine changes in myoblast proliferation in vitro (experiment 1) and muscle hypertrophy in vivo (experiment 2). In vitro experiments found that antagomiR-22-3p and mimic-127 increased myoblast proliferation compared to other miRNA treatments or controls. These miRNA treatments, antagomiR-22-3p (ANT22) and agomiR-127 (AGO127), were then used for intramuscular injections in longissimus muscle. Results and discussion: The use of antagomiR or mimic/agomiR treatments down-regulated or up-regulated, respectively, miRNA expression for that miRNA of interest. Expression of predicted target KIF3B mRNA for miR-127 was up-regulated and ACVR2a mRNA was up-regulated for miR-22-3p. ANT22 injection also up-regulated the major regulator of protein synthesis (mTOR). Proteomic analyses identified 11 proteins for AGO127 and 9 proteins for ANT22 that were differentially expressed. Muscle fiber type and cross-sectional area were altered for ANT22 treatments to transition fibers to a more oxidative state. The use of agomiR and antagomir technologies allows us to alter miRNA expression in vitro and in vivo to enhance myoblast proliferation and alter muscle fiber hypertrophy in IUGR lambs during early postnatal growth.