Undernutrition induced growth restriction in early life increases the risk of chronic disease in adulthood. While metabolic impairments have been observed, few studies have characterized the gut microbiome and gut-liver metabolome profiles of growth restricted animals during early-to-mid-life development. To induce growth restriction, mouse offspring were either born to gestational undernutrition (GUN) or suckled from postnatal undernutrition (PUN) dams fed a protein-restricted diet (8% protein) or control diet (CON; 20% protein) until weaning at postnatal age of 21 days (PN21). At PN21, all mice were fed the CON diet until adulthood (PN80). Livers were collected at PN21 and PN80, and fecal samples were collected weekly for gut microbiome and metabolome analyses. PUN mice exhibited the most alterations in gut microbiome and gut and liver metabolome compared to CON. These mice had altered fecal microbial Beta-Diversity (p=0.001) and exhibited higher proportions of Bifidobacteriales (Linear Mixed Model (LMM) p=7.158x10-6), Clostridiales (p=1.459x10-5), Erysipelotrichales (p=0.0003) and lower Bacteroidales (p=4.121x10-5). PUN liver and fecal metabolome had a reduced total bile acid pool (p<0.01), as well as lower Riboflavin (p=0.003), amino acids (i.e., methionine (p=0.0018), phenylalanine (p=0.0015), and tyrosine (p=0.0041)), and higher excreted total peptides (LMM p=0.0064), dipeptides (p=0.001), and tripeptides (p=0.0123) compared to CON. PUN also had abnormally reduced Epiandrosterone (p=0.02) at PN80. PUN liver and fecal metabolome varied in specific acylcarnitines: higher liver oleoyl-L-carnitine (p=0.0038) and palmitoylcarnitine (p=0.0096) vs. CON at PN21, which recovered at PN80. However, PUN also had higher fecal R-ButyrylCarnitine (p=0.0125) through adulthood, despite refeeding. Overall, protein restriction during lactation permanently alters the gut microbiome into adulthood. While the liver bile acids, amino acids, and acylcarnitines recovered, the fecal peptides and microbiome remained permanently altered into adulthood, indicating that inadequate protein intake in a specific time frame in early life can have an irreversible impact on the microbiome and fecal metabolome.

Low birth weight (LBW) offspring have a higher prevalence of long‐term cognitive impairments, possibly the result of altered blood flow control in the brain. Using a pig model, we tested the hypothesis that LBW progeny exhibit abnormal cerebral hemodynamics and vasomotor control during early developmental stages of growth. At birth, littermate pigs were identified as normal (NBW=1.7±0.1 kg) or low birth weight (LBW=1.2±0.2 kg). Measurements were obtained at 28 d (weaning) and 56 d of age (n=8/group, 4M/4F per group) : i) 28‐d old NBW (28‐NBW; body mass=10.0±0.4 kg); ii) 28‐d old LBW (28‐LBW; body mass=8.0±0.4 kg); iii) 56‐d old NBW (56‐NBW; body mass=23.0±1.0 kg); and iv) 56‐d old LBW (56‐LBW; body mass=17.0±0.8 kg). Cerebral blood flow velocity was measured using Transcranial Doppler ultrasound. Thereafter, pigs were euthanatized, their brains were harvested, weighed and their pial arteries were dissected for pressure myography experiments. Briefly, pial artery vasomotor control was studied in response to a Ca2+‐activated potassium (BK) channel α‐subunit agonist NS1619 (elicits vasodilation; 1e‐10‐1e‐4 M) and sympathetic co‐transmitter neuropeptide Y (NPY; potent vasoconstrictor; 1e‐10‐1e‐4 M). Brain mass and body mass were significantly lower in LBW vs. NBW pigs (p<0.01). Mean cerebral blood flow velocities were similar between LBW and NBW pigs (p≥0.20). Peak systolic velocities were greater (main effect of BW, p<0.01) and reductions in end diastolic velocities approached significance in LBW vs. NBW pigs (main effect of BW, p=0.08). Accordingly, the resistive index was greater in LBW vs. NBW pigs (main effect of BW, p<0.01). The maximum vasodilator response to NS1619 was not significantly different between LBW and NBW pigs (p=0.12). However, the overall magnitude of dilation (area under the curve) was lower in LBW vs. NBW pigs (main effect of BW, p<0.05). The maximum vasoconstrictor response to NPY was greater in LBW vs. NBW groups (main effect of BW, p<0.05). Similarly, the overall magnitude of constriction was greater in LBW vs. NBW pigs (main effect of BW, p<0.01). With respect to cerebral hemodynamics and vasomotor control, no main effects of age were observed (p≥0.50). Results from this study show that LBW pigs display alterations in cerebral hemodynamics as well as cerebral vasomotor control that persist for at least 56 days. Impaired BK channel function and enhanced vasoreactivity to sympathetic co‐transmitter NPY may underlie altered cerebrovascular hemodynamics in LBW pigs. Support or Funding Information SHRF Establishment grant (#4522) and NSERC

William Arthur Ward stated, “The mediocre teacher tells. The good teacher explains. The superior teacher demonstrates. The great teacher inspires.” Discovery experimentation is an inductive method that demonstrates and inspires by creating an interest in determining the underlining basis of a phenomenon. This experiential approach also fosters motivation and enhances learning. Starting with what the student knows augments this approach. By starting with what the student already knows, the student can consciously and explicitly link the subsequent new information with previous knowledge. Accordingly, we used a simple battery as an analogy for electrocardiogram vectors to introduce the theoretical physics of how the heart produces voltages that are detectable at the body surface. This extraordinarily complex physics was approached in a straightforward and inexpensive way while still providing an understanding of the fundamental concepts developed by Willem Einthoven in 1895.

Exercise is essential for reducing age‐related bone mineral loss by providing mechanical loading. Due to greater shear strain forces generated, downhill walking or running at slower speeds may provide similar stimuli to faster running, sprinting, or jumping on flat surfaces. Results from previous studies are mixed when comparing changes in morphological and mechanical properties from slower, downhill running to level or uphill running and sedentary controls. The purpose of this study was to determine if downhill running (DHR) could improve bone mineral content, calcium concentration, and total calcium in the limbs of mice. This study was approved by the MSU IACUC and FVB mice (Charles River) were housed according the Guide for the Care and Use of Laboratory Animals. Forty‐five days after birth (PN45), mice were allocated to either a sedentary or exercised group. Exercised mice began running on a treadmill set at a −16% gradient five times per week for 12 weeks in 30 minute bouts. After a two‐minute warm‐up speeds were increased from 10 m·min−1 to 17 m·min−1 for the remaining 28 minutes. At PN129, mice were euthanized, and fore‐ and hindlimbs were collected as these were the bones most likely to respond to exercise. Bones were cleaned, ether‐extracted, and ashed as a measure of bone mineral content. Ash was recorded and then microwave digested in nitric acid for calcium concentration analysis via atomic absorption spectrophotometry. Total calcium was calculated from multiplying calcium concentration by amount of ash for each sample. Due to the quantity of ash needed for digestion and the small size of mice, bones were pooled. Sedentary (n=7) and exercised (n=6) mouse data were found to be normally distributed and analyzed using the mixed model procedure in SAS 9.4 for effects of sex, exercise, and their interaction on ash, calcium concentration, and total calcium. There was no interaction between sex and exercise as well as no effect of sex nor exercise (P > 0.10) on any of the parameters measured (Table 1). Surprisingly, DHR in this study did not produce greater bone ash or total calcium when compared to sedentary controls. Though DHR can produce compressive strain and high shear strain rates, the slower speed may not be enough for bone to respond as it does to higher speeds or larger impacts. Additionally, these mice were analogous to human adults when they began training, and since younger bone responds to exercise better, this age may have limited the mice’s ability to accrue greater bone mineral. In this study, DHR failed to stimulate bone to accrue or even maintain mineral above sedentary values and therefore may not be an appropriate exercise recommendation to reduce age‐related mineral loss. Ash as a measure of bone mineral content, calcium concentration, and total calcium from the limbs of either downhill running (DHR) or sedentary (Sed) mice. Neither sex nor exercise had an effect on any of the values (P > 0.10). Sex Exercise Ash (mg) Calcium Concentration (μg·ml−1) Total Calcium (mg) Female DHR ...

INTRODUCTION Early life growth restriction increases susceptibility to non‐communicable adult‐onset diseases. However, there is minimal mechanistic rationale for elevated disease risk. The gut microbiome assists in regulating host physiological processes, with emerging evidence suggesting microbiome dysregulation leads to increased risk of chronic disease. Thus, the purpose of this investigation was to determine differences in the gut microbiome of growth restricted mice as compared to non‐restricted mice. METHODS A cross‐fostering, protein‐restricted (8% protein) nutritive model in FVB mice was used to induce growth restriction during gestation (GUN; N=10 male,15 female) or lactation from days 1–21 of life (PUN; N=11 male, 12 female) along with a control group (CON, 20% protein, N=12 male, 11 female). At 21 postnatal days of age (PN21) all mice were weaned to a non‐restricted diet (20% protein), isolating undernutrition to early life windows. Fecal samples were collected weekly across the lifespan (until PN80) to determine longitudinal programming effects of growth restriction on the gut microbiome. Cecum samples were collected at PN21 and PN80. Microbiome analysis of fecal and cecum samples were conducted by first extracting the DNA using Qiagen Powersoil DNA extraction kits and conducting PCR amplification of the bacterial 16S rRNA gene amplicon profiles through the Qiita bioinformatics platform. Phenotypic growth markers of all three diet groups (CON, GUN, PUN) were analyzed using repeated measures ANOVA. Metabolomics data on the same samples is currently being analyzed and will be integrated with the microbiome dataset. RESULTS The fecal microbiome was strongly separated by treatment group using the Bray‐Curtis distance measure (PERMANOVA p = 0.001). A random forest machine learning classification validated this separation showing an overall out‐of‐bag error rate of 18.6% across the three groups in the whole dataset. The PUN group was distinctly separated from GUN and CON, with a group error rate of only 15.0%. Differences in the microbiome were not evident through analysis at the Phylum level (Firmicutes/Bacteroidetes ratio) but was instead driven by differential abundances of specific rare taxa. Bifidobacterium was significantly elevated in the PUN group (Kruskal‐Wallis test p=0.00017), beginning as early as PN21 and remaining elevated in the PUN group through the lifespan (PN80). CONCLUSIONS A protein‐restricted nutritive model induces strong imprinting on the structure of the murine microbiome. Changes in the PUN group are primarily driven by differences in low abundance taxa. Understanding the mechanisms that lead to the microbiome dysbiosis shown here could lead to the development of therapeutic interventions to limit the development of a stunted or immature gut microbiota. Support or Funding Information MSU Department of Biochemistry and Molecular Biology; MSU Department of Kinesiology

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Introduction Growth-restriction (GR) reduces ribosome abundance and skeletal muscle mass in mice. A reduction in skeletal muscle mass increases the risk of frailty and is associated with high morbidity and mortality rates. As eccentric type exercise increases muscle mass, this investigation aimed to determine if eccentric loading of skeletal muscle via downhill running (DHR) increased muscle mass in GR mice. Methods Mice were growth-restricted either gestational undernutrition (GUN: n = 8 litters), postnatal undernutrition (PUN: n = 8 litters) or were not restricted (CON: n = 8 litters) via a validated cross-fostering nutritive model. On postnatal day (PN) 21, all mice were weaned to a healthy diet, isolating the period of GR to early life as seen in humans. At PN45, mice were assigned to either a downhill running (DHR, CON n = 4 litters, GUN n = 4 litters, PUN n = 4 litters) or sedentary (SED, CON n = 4 litters, GUN n = 4 litters, PUN n = 4 litters) group. DHR (16% decline: 18 m/min) was performed in 30-minute bouts, three times per week, for 12 weeks on a rodent treadmill. At PN129, the quadriceps femoris was dissected and evaluated for mass, myofiber size and type, and molecular markers of growth. Results Following training, CON-DHR mice having larger cells than CON-SED, GUN-SED, PUN-SED, and PUN-DHR mice (p < 0.05). The PUN group (as compared to CON) had reduced body mass (p < 0.001), UBF abundance (p = 0.012), phosphor-mTOR (p < 0.001), and quadriceps mass (p = 0.02). The GUN and PUN groups had increased MuRF1 abundance (p < 0.001) compared to CON (p < 0.001). Conclusions The blunted response to training suggests GR mice may have anabolic resistance when exposed to eccentric type exercise.