New fiber formation in the interstitial spaces of rat skeletal muscle during postnatal growth.

The purpose of this study was to determine whether fiber hyperplasia occurs in the rat plantaris muscle during postnatal weeks 3-20. Total muscle fiber number, obtained via the nitric acid digestion method, increased by 28% during the early postnatal rapid growth phase (3-10 weeks), whereas the number of branched fibers was consistently low. Whole-muscle mitotic activity and amino acid uptake levels showed an inverse relationship to the increase in total fiber number. The expression of MyoD mRNA (RT-PCR) levels decreased from 3 to 20 weeks of age, as did the detection of anti-BrdU- and MyoD-positive cells in histological sections. Immunohistochemical staining patterns for MyoD, myogenin, or developmental myosin heavy chain on sections stained for laminin (identification of the basal lamina) and electron micrographs clearly indicate that de novo fiber formation occurred in the interstitial spaces. Myogenic cells in the interstitial spaces were negative for the reliable specific satellite cell marker M-cadherin. In contrast, CD34 (an established marker for hematopoietic stem cells)-positive cells were located only in the interstitial spaces, and their frequency and location were similar to those of MyoD- and/or myogenin-positive cells. These findings are consistent with fiber hyperplasia occurring in the interstitial spaces of the rat plantaris muscle during the rapid postnatal growth phase. Furthermore, these data suggest that the new fibers may be formed from myogenic cells in the interstitial spaces of skeletal muscle and may express CD34 that is distinct from satellite cells.     

Perturbed skeletal muscle insulin signaling in the adult female intrauterine growth-restricted rat

To determine the molecular mechanism(s) linking fetal adaptations in intrauterine growth restriction (IUGR) to adult maladaptations of type 2 diabetes mellitus, we investigated the effect of prenatal seminutrient restriction, modified by early postnatal ad libitum access to nutrients (CM/SP) or seminutrient restriction (SM/SP), vs. early postnatal seminutrient restriction alone (SM/CP) or control nutrition (CM/CP) on the skeletal muscle postreceptor insulin-signaling pathway in the adult offspring. The altered in utero hormonal/metabolic milieu was associated with no change in basal total IRS-1, p85, and p110beta subunits of PI 3-kinase, PKCtheta, and PKCzeta concentrations but an increase in basal IRS-2 (P < 0.05) only in the CM/SP group and an increase in basal phospho (p)-PDK-1 (P < 0.05), p-Akt (P < 0.05), and p-PKCzeta (P < 0.05) concentrations in the CM/SP and SM/SP groups. Insulin-stimulated increases in p-PDK-1 (P < 0.05) and p-Akt (P < 0.0007), with no increase in p-PKCzeta, were seen in both CM/SP and SM/SP groups. SHP2 (P < 0.03) and PTP1B (P < 0.03) increased only in SM/SP with no change in PTEN in CM/SP and SM/SP groups. Aberrations in kinase and phosphatase moieties in the adult IUGR offspring were initiated in utero but further sculpted by the early postnatal nutritional state. Although the CM/SP group demonstrated enhanced kinase activation, the SM/SP group revealed an added increase in phosphatase concentrations with the net result of heightened basal insulin sensitivity in both groups. The inability to further respond to exogenous insulin was due to the key molecular distal roadblock consisting of resistance to phosphorylate and activate PKCzeta necessary for GLUT4 translocation. This protective adaptation may become maladaptive and serve as a forerunner for gestational and type 2 diabetes mellitus.     

Expression of fibroblast growth factor family during postnatal skeletal muscle hypertrophy

The potential role of the fibroblast growth factor (FGF) familyduring stretch-induced postnatal skeletal muscle hypertrophy wasanalyzed by using an avian wing-weighting model. After 2 or 11 days ofweighted stretch, anterior latissimus dorsi (ALD) muscles were, onaverage, 34 (P < 0.01) and 85%(P < 0.01) larger, respectively, than unweighted ALD control muscles. By using quantitative RT-PCR, FGF-1 mRNA expression was found to be significantly decreased in ALDmuscles stretched for 2 or 11 days. In contrast, FGF-4 and FGF-10 mRNAexpression was significantly increased 2 days after initiation ofstretch. FGF-2, FGF-10, fibroblast growth factor receptor 1, andFREK mRNA expression was significantly increased at 11 days poststretch. Increases in FGF-2 and FGF-4 protein could bedetected throughout the myofiber periphery after 11 days of stretch. Ona cellular level, FGF-2 and FGF-4 proteins were differentiallylocalized. This differential expression pattern and proteinlocalization of the FGF family in response to stretch-induced hypertrophy suggest distinct roles for individual FGFs during thepostnatal hypertrophy process.

     

Satellite cell proliferation and differentiation during postnatal growth of porcine skeletal muscle

Age-related changes in satellitecell proliferation and differentiation during rapid growth of porcineskeletal muscle were examined. Satellite cells were isolated fromhindlimb muscles of pigs at 1, 7, 14, and 21 wk of age (4 animals/agegroup). Satellite cells were separated from cellular debris by usingPercoll gradient centrifugation and were adsorbed to glass coverslipsfor fluorescent immunostaining. Positive staining for neural celladhesion molecule (NCAM) distinguished satellite cells from nonmyogeniccells. The proportion of NCAM-positive cells (satellite cells) inisolates decreased from 1 to 7 wk of age. Greater than 77% ofNCAM-positive cells were proliferating cell nuclear antigen positive atall ages studied. Myogenin-positive satellite cells decreased from 30%at 1 wk to 14% at 7 wk of age and remained at constant levels thereafter. These data indicate that a high percentage of satellite cells remain proliferative during rapid postnatal muscle growth. Thereduced proportion of myogenin-positive cells during growth may reflecta decrease in the proportion of differentiating satellite cells oraccelerated incorporation of myogenin-positive cells into myofibers.

     

1, 0 lattice spacing of skeletal muscle during postnatal growth in mice

1, 0 lattice spacing of extensor digitorum longus muscles of mice (ICR) during growth (2-12 weeks) were measured in situ by X-ray diffraction method. 1, 0 lattice spacing of EDL was 35.4 +/- 0.27 (mean +/- SD, N = 33) nm. No significant difference between spacings in EDL's of each aged mice was observed.     

Comparison of growth and development of the exocrine pancreas in pigs and rats during the immediate postnatal period

This study compared pancreatic tissue growth and functional changes during the first 3 postnatal days in piglets and rat pups. In piglets the absolute weight and the relative weight per unit body weight of the pancreas increased by 97 and 70%, respectively, while in rat pups the same parameters decreased by 33 and 48%, respectively, during this period. The specific activity of pancreatic amylase rose by 336% while that of trypsin, chymotrypsin and lipase remained at newborn level in piglets. In rat pups the specific activities of all enzymes measured declined by 61 to 92% during the first 3 postnatal days. The rate of postnatal pancreatic growth in the two species coincide with the levels of epidermal growth factor and insulin-like growth factors in maternal milk as reported in the literature, suggesting that milk-borne growth factors may stimulate pancreatic development in newborn animals.     

Myocardial macronutrient transporter adaptations in the adult pregestational female intrauterine and postnatal growth-restricted offspring

Associations between exponential childhood growth superimposed on low birth weight and adult onset cardiovascular disease with glucose intolerance/type 2 diabetes mellitus exist in epidemiological investigations. To determine the metabolic adaptations that guard against myocardial failure on subsequent exposure to hypoxia, we compared with controls (CON), the effect of intrauterine (IUGR), postnatal (PNGR), and intrauterine and postnatal (IPGR) calorie and growth restriction (n = 6/group) on myocardial macronutrient transporter (fatty acid and glucose) -mediated uptake in pregestational young female adult rat offspring. A higher myocardial FAT/CD36 protein expression in IUGR, PNGR, and IPGR, with higher FATP1 in IUGR, FATP6 in PNGR, FABP-c in PNGR and IPGR, and no change in GLUT4 of all groups was observed. These adaptive macronutrient transporter protein changes were associated with no change in myocardial [(3)H]bromopalmitate accumulation but a diminution in 2-deoxy-[(14)C]glucose uptake. Examination of the sarcolemmal subfraction revealed higher basal concentrations of FAT/CD36 in PNGR and FATP1 and GLUT4 in IUGR, PNGR, and IPGR vs. CON. Exogenous insulin uniformly further enhanced sarcolemmal association of these macronutrient transporter proteins above that of basal, with the exception of insulin resistance of FATP1 and GLUT4 in IUGR and FAT/CD36 in PNGR. The basal sarcolemmal macronutrient transporter adaptations proved protective against subsequent chronic hypoxic exposure (7 days) only in IUGR and PNGR, with notable deterioration in IPGR and CON of the echocardiographic ejection fraction. We conclude that the IUGR and PNGR pregestational adult female offspring displayed a resistance to insulin-induced translocation of FATP1, GLUT4, or FAT/CD36 to the myocardial sarcolemma due to preexistent higher basal concentrations. This basal adaptation of myocardial macronutrient transporters ensured adequate fatty acid uptake, thereby proving protective against chronic hypoxia-induced myocardial compromise.     

Mice lacking skeletal muscle actin show reduced muscle strength and growth deficits and die during the neonatal period

All four of the muscle actins (skeletal, cardiac, vascular, and enteric) in higher vertebrates show distinct expression patterns and display highly conserved amino acid sequences. While it is hypothesized that each of the muscle isoactins is specifically adapted to its respective tissue and that the minor variations among them have developmental and/or physiological relevance, the exact functional and developmental significance of these proteins remains largely unknown. In order to begin to assess these issues, we disrupted the skeletal actin gene by homologous recombination. All mice lacking skeletal actin die in the early neonatal period (day 1 to 9). These null animals appear normal at birth and can breathe, walk, and suckle, but within 4 days, they show a markedly lower body weight than normal littermates and many develop scoliosis. Null mice show a loss of glycogen and reduced brown fat that is consistent with malnutrition leading to death. Newborn skeletal muscles from null mice are similar to those of wild-type mice in size, fiber type, and ultrastructural organization. At birth, both hemizygous and homozygous null animals show an increase in cardiac and vascular actin mRNA in skeletal muscle, with no skeletal actin mRNA present in null mice. Adult hemizygous animals show an increased level of skeletal actin mRNA in hind limb muscle but no overt phenotype. Extensor digitorum longus (EDL) muscle isolated from skeletal-actin-deficient mice at day 2 to 3 showed a marked reduction in force production compared to that of control littermates, and EDL muscle from hemizygous animals displayed an intermediate force generation. Thus, while increases in cardiac and vascular smooth-muscle actin can partially compensate for the lack of skeletal actin in null mice, this is not sufficient to support adequate skeletal muscle growth and/or function.     

Development of different types of muscle fiber in the postnatal ontogeny of guinea pigs

As demonstrates estimation of myosin ATPase and SDG activity, the guinea pig is already born with differentiated muscle fibers (MF), and the first histochemical differences between them take place in the uterine 10 days before birth. Tonic oxidative fibers of the first type, arranging hexagonally, develop especially quickly at early stages of postnatal ontogenesis. Their relative contents up to the end of the observations (185 days) do not change, and area of their transversal section increases but slightly in comparison to the phasic fibers. The main age changes of the muscle tissue are connected with formation and rearrangement of the phasic fibers. The most intensive reconstructions of the phasic fibers coincide with the period of game activity and sex maturation. In mixed muscles the part of the glycolytic fibers increase during the postnatal ontogenesis. In the process of ontogenesis the soleus muscle fully consists of oxidative fibers. The definitive level of the MF development is established after the guinea pigs have reached their sex maturation. Comparing the results of the given investigation with the previous data on development of MF in rats, it is possible to conclude that term and premature animals have various rates in development of the muscle system, however, main stages of myogenesis coincide, though they are connected with various phases of ontogenesis.     

Ontogenetic and lung development in Tupaia belangeri during the early postnatal period

The Belanger's tree shrew (Tupaia belangeri) has an unusual reproductive strategy. The animals are born in altricial condition and remain in the nest for the first four weeks of life, nursed only once in 48 h. This is highly demanding for the constitution of the neonates. Despite their immaturity in the external appearance at birth, newborn tree shrews have to deal with the absence of the mother. We asked if the lung structure of the neonates match the high physiological requirements of this “absentee system”. To examine the lung development of nest young tree shrews, histological and ultrastructural investigations were performed. Newborn tree shrews are at the transition stage between the saccular and the alveolar stage of lung development. In addition to small saccules, the lung has alveoli and associated structures already at birth and thus appears more mature compared with typical altricial species. The results of the present study reveal that despite their immaturity in the external appearance newborn tree shrews are relatively mature in terms of lung development. This can be interpreted as a prerequisite for thermoregulatory abilities, necessary in neonate tree shrews to cope with the restricted nature of maternal care.     

Early postnatal growth of skeletal muscle blood vessels of the rat

The development of blood vessels during the first three postnatal weeks was studied in the ventral stripe of the spinotrapezius muscle of the rat by use of India ink-gelatine injections, and electron microscopy. The number of terminal arterioles and collecting venules remained unchanged postnatally in the observed area. A remarkable proximodistal gradient of vascular development was apparent: while the basic structure of the hilar vessels remained unchanged in the time studied, the intramuscular arteries and veins matured gradually. More peripherally, gradual maturation of terminal and precapillary arterioles was observed. The capillary endothelium and the pericytes showed immature features, and remained unchanged during the time studied. An intense rebuilding activity was found in the endothelial cells of the growing venules, expressed by various forms of gaps, covered by an intact basal lamina and pericytes. Numerous mast cells and macrophages were found along all vessels. Intramuscular lymphatics were not present prior to the first postnatal week.     

Diversification of the myofibrillar M-band in rat skeletal muscle during postnatal development

Summary The fine structure of the M-band in soleus (SOL) and extensor digitorum longus (EDL) muscles in newborn and four-week-old rats was studied using electron-microscopic techniques. In newborn rats, all myotubes and fibres in both muscles had an identical myofibrillar appearance. A five-line M-band pattern was seen in longitudinal sections and distinct M-bridges in cross-sections. The Z-discs were of medium width. On the other hand, in four-week-old rats, different muscle fibre types were observed on the basis of their myofibrillar pattern. In SOL two fibre types were distinguished in longitudinal sections. One had a four-line M-band pattern and very broad Z-discs, whereas the other type had five lines in the M-band and broad Z-discs. In EDL, three different myofibrillar patterns were observed. The M-bands were composed of three, four or five lines. Fibres had either thin, broad or medium Z-disc widths, respectively. In cross-sections of the SOL muscle one group of fibres showed indistinct M-bridges, whereas distinct M-bridges were seen in the other fibres and in all observed EDL muscle fibres. We conclude that initially there seems to be a single intrinsic program for M-band genesis; this program becomes modified upon the induction of functionally differentiated fibres.     

Vascular endothelial growth factor and placenta growth factor in intrauterine growth-restricted fetuses and neonates

The angiogenic factors vascular endothelial growth factor (VEGF) and placenta growth factor (PlGF) are respectively up- and downregulated by hypoxia. We aimed to study circulating levels of the above factors in intrauterine growth restriction (IUGR) and to correlate their levels with the customized centiles of the infants. The study included 25 IUGR and 25 appropriate for gestational age (AGA) full-term, singleton infants and their mothers. Maternal (MS), fetal (UC), and neonatal day 1 (N1) and 4 (N4) blood was examined. MS and N1 PlGF, as well as UC VEGF levels correlated with the customized centiles of the infants (r= 0.39, P=.007, r=0.34, P=.01, and r= -0.41, P=.004, resp). Furthermore, UC, N1, and N4 VEGF levels were higher in girls (r=0.36, P=.01, r=0.33, P=.02, and r=0.41, P=.005 resp). In conclusion, positive and negative correlations of examined factors with the customized centiles of the infant could rely on placental function and intrauterine oxygen concentrations-both being usually lower in IUGR cases-while higher VEGF levels in girls should possibly be attributed to the stimulating action of estrogens.     

Beta-adrenergic receptor-adenylate cyclase alterations during the postnatal development of skeletal muscle

The postnatal development of mammalian skeletal muscle is associated with an increased capacity for glycogenolysis. In the present study rabbit skeletal muscle underwent a 7-fold increase in glycogen synthase and glycogen phosphorylase activity over the postnatal period of 0--8 weeks. An enriched fraction of sarcolemma was prepared from neonatal and adult muscle to examine the development of the beta-adrenergic receptor-adenylate cyclase system. Adult membranes possessed a 2-fold greater Na+K+(Mg2+)-ATPase activity and a 6--8 fold greater sodium fluoride- and epinephrine-stimulated adenylate cyclase activity. The activation ratio (effector activity/basal activity) increased 2--3 fold for epinephrine and sodium fluoride in adult sarcolemma. The activation by catecholamines conformed to the physiological beta 2 type response with isoproterenol (1.8 . 10(-8) M) > epinephrine (1.1 . 10(-7) M) > norinephrine (3.2 . 10(-6) M). In contrast, binding studies employing (-)-[3H]dihydroalprenolol showed little difference between neonatal and adult membranes with respect to (1) number of binding sites, (2) equilibrium dissociation constant and (3) displacement of (-)-[3H]dihydroalprenolol by catecholamine agonists. Protein and lipid components of the sarcolemma were also modified during development. Neonatal membranes possessed two glycopeptides of Mr 80000 and 86000, whereas in the adult only a single Mr 113000 species was evident. The total lipid phosphorus and phospholipid composition was unchanged during development. The content of linoleic acid increased approx. 3-fold during development in the phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine phospholipids. The cholesterol content of adult membranes was decreased by 29% compared to neonatal membranes.     

Formation of dolichol-linked sugar intermediates during the postnatal development of skeletal muscle

The postnatal development of skeletal muscle is characterized by changes in membrane function associated with N-linked glycoproteins. In the present study, early reactions involved in the synthesis of the dolichol-linked core oligosaccharide were examined in neonatal and adult rabbit skeletal muscle sarcoplasmic reticulum membranes. The initial rate of N-acetylglucosamine incorporation in the presence of exogenous dolichol phosphate was similar between neonate and adult (3.5-4.1 pmol of GlcNAc/min/mg). The Km values for UDP-GlcNAc and exogenous dolichol phosphate were similar. Tunicamycin (0.04-0.08 micrograms/ml) inhibited N-acetylglucosamine incorporation by 50%. UDP-GlcNAc pyrophosphatase activity was greater in neonatal membranes than adult (840 versus 350 pmol of GlcNAc-1-P/min/mg), explaining, in part, the greater enhancement of neonatal GlcNAc incorporation by pyrophosphatase inhibitors. Nucleotide-sugar pyrophosphatase inhibitors (alpha, beta-methylene ATP and dimercaptopropanol) increased the capacity of neonatal activity 4-fold and adult enzyme 2-fold. Analysis of dolichol-linked products by mild acid hydrolysis however, revealed that neonate had higher capacity for N,N'-diacetylchitobiosyl(pyro)phosphoryldolichol synthesis than adult. Mannosyltransferase and glucosyltransferase were elevated 6- and 5-fold in neonate compared to adult membranes. Neonate exhibited 4-fold greater GDP-Man pyrophosphatase activity than adult (500 versus 125 pmol of Man-1-P/min/mg). The Km for GDP-Man increased in the presence of exogenous dolichol phosphate. Increasing concentrations of exogenous dolichol phosphate did not equalize neonate and adult mannosyltransferase activity, indicating that the decline in activity during development was not due to a decrease in a pool of dolichol phosphate accessible to mannosyltransferase. Glucosyltransferase for the synthesis of glucosylphosphoryldolichol was also elevated 5-fold in neonatal compared to adult sarcoplasmic reticulum (7 versus 1.4 pmol of Glc/min/mg). In a previous study, it was reported that glycoprotein sialyltransferase activity decreased by a factor of 6.5 during the postnatal maturation and that total membrane hexose content of sarcoplasmic reticulum decreased by a factor of 8. Together, these results suggest that the postnatal development of skeletal muscle is characterized by coordinated changes in the expression of enzymes involved in both the "early" and "late" reactions of N-linked oligosaccharide biosynthesis.     

β-adrenegic receptor-adenylate cyclase alterations during the postnatal development of skeletal muscle

The postnatal development of mammalian skeletal muscle is associated with an increased capacity for glycogenolysis. In the present study rabbit skeletal muscle underwent a 7-fold increase in glycogen synthase and glycogen phosphorylase activity over the postnatal period of 0–8 weeks. An enriched fraction of sarcolemma was prepared from neonatal and adult muscle to examine the development of the β-adrenergic receptor-adenylate cyclase system. Adult membranes possessed a 2-fold greater Na⁺K⁺(Mg²⁺)-ATPase activity and a 6–8-fold greater sodium fluoride- and epinephrine-stimulated adenylate cyclase activity. The activation ratio (effector activity/basal activity) increased 2–3-fold for epinephrine and sodium fluoride in adult sarcolemma. The activation by catecholamines conformed to the physiological β₂ type response with isoproterenol (1.8 · 10^?8 M) > epinephrine (1.1 · 10^?7 M) > norinephrine (3.2 · 10^?6 M). In contrast, binding studies employing (?)-[³H] dihydroalprenolol showed little difference between neonatal and adult membranes with respect to (1) number of binding sites, (2) equilibrium dissociation constant and (3) displacement of (?)-[³H]dihydroalprenolol by catecholamine agonists.Protein and lipid components of the sarcolemma were also modified during development. Neonatal membranes possessed two glycopeptides of M_r 80 000 and 86 000, whereas in the adult only a single M_r 133 000 species was evident. The total lipid phosphorus and phospholipid composition was unchanged during development. The content of linoleic acid increased approx. 3-fold during development in the phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine phospholipids. The cholesterol content of adult membranes was decreased by 29% compared to neonatal membranes.