Metamorphosis of the Drosophila visceral musculature and its role in intestinal morphogenesis and stem cell formation.

The visceral musculature of the Drosophila intestine plays important roles in digestion as well as development. Detailed studies investigating the embryonic development of the visceral muscle exist; comparatively little is known about postembryonic development and metamorphosis of this tissue. In this study we have combined the use of specific markers with electron microscopy to follow the formation of the adult visceral musculature and its involvement in gut development during metamorphosis. Unlike the adult somatic musculature, which is derived from a pool of undifferentiated myoblasts, the visceral musculature of the adult is a direct descendant of the larval fibers, as shown by activating a lineage tracing construct in the larval muscle and obtaining labeled visceral fibers in the adult. However, visceral muscles undergo a phase of remodeling that coincides with the metamorphosis of the intestinal epithelium. During the first day following puparium formation, both circular and longitudinal syncytial fibers dedifferentiate, losing their myofibrils and extracellular matrix, and dissociating into mononuclear cells ("secondary myoblasts"). Towards the end of the second day, this process is reversed, and between 48 and 72h after puparium formation, a structurally fully differentiated adult muscle layer has formed. We could not obtain evidence that cells apart from the dedifferentiated larval visceral muscle contributed to the adult muscle, nor does it appear that the number of adult fibers (or nuclei per fiber) is increased over that of the larva by proliferation. In contrast to the musculature, the intestinal epithelium is completely renewed during metamorphosis. The adult midgut epithelium rapidly expands over the larval layer during the first few hours after puparium formation; in case of the hindgut, replacement takes longer, and proceeds by the gradual caudad extension of a proliferating growth zone, the hindgut proliferation zone (HPZ). The subsequent elongation of the hindgut and midgut, as well as the establishment of a population of intestinal stem cells active in the adult midgut and hindgut, requires the presence of the visceral muscle layer, based on the finding that ablation of this layer causes a severe disruption of both processes.

Understanding evolutionary variation in basal metabolic rate: An analysis in subterranean rodents.

Understanding how evolutionary variation in energetic metabolism arises is central to several theories in animal biology. Basal metabolic rate (BMR) -i.e., the minimum rate of energy necessary to maintain thermal homeostasis in endotherms- is a highly informative measure to increase our understanding, because it is determined under highly standardized conditions. In this study we evaluate the relationship between taxa- and mass-independent (residual) BMR and ten environmental factors for 34 subterranean rodent species. Both conventional and phylogenetically informed analyses indicate that ambient temperature is the major determinant of residual BMR, with both variables inversely correlated. By contrast, other environmental factors that have been shown to affect residual BMR in endotherms, such as habitat productivity and rainfall, were not significant predictors of residual BMR in this group of species. Then, the results for subterranean rodents appear to support a central prediction of the obligatory heat model (OHM), which is a mechanistic model aimed to explain the evolution of residual BMR. Specifically, OHM proposes that during the colonization of colder environments, individuals with greater masses of metabolically expensive tissues (and thus with greater BMR) are favored by natural selection due to the link between greater masses of metabolically expensive tissues and physiological capacities. This way, natural selection should establishes a negative correlation between ambient temperature and both internal organ size and residual BMR.

Portal-drained viscera heat production in Iberian pigs fed betaine- and conjugated linoleic acid-supplemented diets.

BACKGROUND: Betaine and conjugated linoleic acid (CLA) may alter growth and body composition in pigs, although their mode of action is not well understood. Portal-drained viscera (PDV) have a disproportionate influence with respect to their masses, and this may affect the productivity of more profitable tissues. The objective of this study was to determine if the use of betaine and/or CLA in the diet affects PDV heat production. RESULTS: Postprandial portal blood flow (PBF) was greater (19.0%, P = 0.004) for control compared with the other three diets. The lowest (P < 0.001) value for postprandial PDV O2 consumption corresponded to betaine + CLA followed by betaine and CLA diets (32.7, 25.4 and 17.7% respectively with respect to control diet). Postprandial PDV heat production was greater (26.4%, P < 0.001) for control with respect to the other three diets, with the minimum value corresponding to betaine + CLA (34.1% lower than control). CONCLUSION: Supplementation with betaine and/or CLA reduced the PBF, O2 consumption and therefore PDV heat production with respect to control diet. This effect was more pronounced when betaine and CLA were supplemented together, potentially increasing the energy availability for other body tissues. © 2016 Society of Chemical Industry.

Comparative effect of cane syrup and natural honey on abdominal viscera of growing male and female rats.

The high intake of refined sugars, mainly fructose has been implicated in the epidemiology of metabolic diseases in adults and children. With an aim to determine whether honey can substitute refined sugars without adverse effect, the long-term effects of natural honey and cane syrup have been compared on visceral morphology in growing rats fed from neonatal age. Honey increased the caecum and pancreas weights in male rats, which could enhance enzymatic activities of pancreas and digestive functions by intestinal microflora of caecum. Unlike honey, cane syrup caused fatty degenerations in the liver of both male and female rats. Honey enhanced intestinal villi growth, and did not cause pathology in the rodents' abdominal viscera, suggesting potential nutritional benefit as substitution for refined sugars in animal feed.

Developmental study of tripeptidyl peptidase I activity in the mouse central nervous system and peripheral organs.

Tripeptidyl peptidase I (TPPI) - a lysosomal serine protease - is encoded by the CLN2 gene, mutations that cause late-infantile neuronal ceroid lipofuscinosis (LINCL) connected with profound neuronal loss, severe clinical symptoms and early death at puberty. Developmental studies of TPPI activity levels and distribution have been done in the human and rat central nervous systems (CNS) and visceral organs. Similar studies have not been performed in mouse. In this paper, we follow up on the developmental changes in the enzyme activity and localization pattern in the CNS and visceral organs of mouse over the main periods of life - embryonic, neonate, suckling, infantile, juvenile, adult and aged - using biochemical assays and enzyme histochemistry. In the studied peripheral organs (liver, kidney, spleen, pancreas and lung) TPPI is present at birth but further its pattern is not consistent in different organs over different life periods. TPPI activity starts to be expressed in the brain at the 10th embryonic day but in most neuronal types it appears at the early infantile period, increases during infancy, reaches high activity levels in the juvenile period and is highest in adult and aged animals. Thus, in mice TPPI activity becomes crucial for the neuronal functions later in development (juvenile period) than in humans and does not decrease with aging. These results are essential as a basis for comparison between normal and pathological TPPI patterns in mice. They can be valuable in view of the use of animal models for studying LINCL and other neurodegenerative disorders.

Association of residual feed intake with growth and slaughtering performance, blood metabolism, and body composition in growing lambs.

The aim of this study was to determine the association of residual feed intake (RFI) with growth performance, blood metabolic parameters, and body composition factors in growing lambs. Individual body weight (BW) and dry matter intake (DMI) were determined in 137 male Hu lambs that were given a pellet feed four times a day for 50 d. RFI did not show a correlation with metabolic BW (MBW) or average daily gain (ADG), but it showed a positive correlation with DMI and feed conversation ratio (FCR). Organ weight and intestine length had a large influence on RFI in lambs. The low-RFI lambs have smaller rumen and longer duodenum indicating the less feed intake and more sufficient absorption rate of low-RFI lambs. The smaller organs like liver, lung and kidney in low-RFI lambs may be related to lower energy consumption and slower metabolic rate. The observed bigger testis was in low-RFI lambs was another cause of the improved feed efficiency. Finally, the plasma concentrations of thyroxine (T4) and adrenocorticotropic hormone (ACTH) were lower in the ELow-RFI group than in the EHigh-RFI group. This study provides new insight into the biological processes underlying variations in feed efficiency in growing lambs.

Isolation and characterization of a Japanese flounder clonal line, reversed, which exhibits reversal of metamorphic left-right asymmetry.

We have isolated a clonal line reversed (rev) of homozygous Japanese flounder through gynogenesis. The homozygous offspring gynogenetically produced from rev exhibited reversal of organization of the metamorphic L/R asymmetry such as the direction of eye-migration at a high frequency (20-30%). The molecular analysis using a left-specific marker pitx2 revealed that the embryonic L/R axis was ambiguously established: in more than half of rev embryos, pitx2 was expressed bilaterally in the lateral plate mesoderm (LPM). Previous studies in other animals demonstrated that ectopic pitx2 expression in the LPM could cause laterality defects of the visceral organs. Likewise, our results using rev imply that bilateral pitx2 expression could lead to randomization of the visceral organs. Coincidence of ectopic pitx2 expression and reversal of the direction of eye-migration in the population of rev offspring suggests that the rev locus is critical in specification of both the metamorphic and the visceral L/R asymmetries. However, reversal of the sidedness of the orientation of the visceral organs was not always accompanied by reversal of the direction of metamorphic eye-migration, suggesting that different mechanisms should be involved downstream of the rev locus in directing these two phases of asymmetric morphogenesis in the Japanese flounder.

Neurotrophins in murine viscera: a dynamic pattern from birth to adulthood.

There is growing evidence that target-derived neurotrophins regulate the function of visceral neurons after birth. However, the postnatal profile of neurotrophin supply from internal organs is poorly described. In this study, we compared neurotrophin concentrations in lysates of murine peripheral target tissues (lung, heart, liver, colon, spleen, thymus, kidney and urinary bladder) at different time points after birth. In most organs, there was a decrease of neurotrophin concentrations in the first weeks after birth. In contrast, there were characteristic increases of specific neurotrophins during adolescence or adulthood. These increases were found for nerve growth factor (NGF) in the heart, thymus, kidney and liver, for brain-derived neurotrophic factor (BDNF) in the lung, and for neurotrophin-3 (NT-3) in the colon. In conclusion, we show that neurotrophins display a very differential and dynamic profile in internal organs after birth.

Elevated C-reactive protein: another component of the atherothrombotic profile of abdominal obesity.

Recent studies have suggested that elevated plasma C-reactive protein (CRP) levels are associated with the features of insulin resistance syndrome. In the present study, we have examined the contribution of body composition measured by hydrostatic weighing and of abdominal adipose tissue (AT) accumulation assessed by computed tomography to the variation in plasma CRP levels associated with atherogenic dyslipidemia of the insulin resistance syndrome in a sample of 159 men, aged 22 to 63 years, covering a wide range of adiposity (body mass index values from 21 to 41 kg/m(2)). Plasma CRP levels showed positive and significant correlations with body fat mass (r=0.41, P<0.0001), waist girth (r=0.37, P<0.0001), and visceral AT accumulation measured by computed tomography at L4 to L5 (r=0.28, P<0.0003). Although CRP levels were associated with plasma insulin levels measured in the fasting state and after a 75-g oral glucose load, no significant correlations were found with plasma lipoprotein levels. Finally, comparison of body fatness, of abdominal fat accumulation, and of the features of the insulin resistance syndrome across quintiles of CRP revealed major differences in body fatness and in indices of abdominal AT accumulation between the lowest and the highest CRP quintiles, whereas no significant differences were found for variables of the plasma lipoprotein-lipid profile. These results suggest that obesity and abdominal AT accumulation are the critical correlates of elevated plasma CRP levels found in men with atherogenic dyslipidemia of the insulin resistance syndrome.

Effect of fetal undernutrition and postnatal overfeeding on rat adipose tissue and organ growth at early stages of postnatal development.

Intrauterine and perinatal life are critical periods for programming of cardiometabolic diseases. However, their relative role remains controversial. We aimed to assess, at weaning, sex-dependent alterations induced by fetal or postnatal nutritional interventions on key organs for metabolic and cardiovascular control. Fetal undernutrition was induced by dam food restriction (50 % from mid-gestation to delivery) returning to ad libitum throughout lactation (Maternal Undernutrition, MUN, 12 pups/litter). Postnatal overfeeding (POF) was induced by litter size reduction from normally fed dams (4 pups/litter). Compared to control, female and male MUN offspring exhibited: 1) low birth weight and accelerated growth, reaching similar weight and tibial length by weaning, 2) increased glycemia, liver and white fat weights; 3) increased ventricular weight and tendency to reduced kidney weight (males only). Female and male POF offspring showed: 1) accelerated growth; 2) increased glycemia, liver and white fat weights; 3) unchanged heart and kidney weights. In conclusion, postnatal accelerated growth, with or without fetal undernutrition, induces early alterations relevant for metabolic disease programming, while fetal undernutrition is required for heart abnormalities. The progression of cardiac alterations and their role on hypertension development needs to be evaluated. The similarities between sexes in pre-pubertal rats suggest a role of sex-hormones in female protection against programming.

Molecular profiling of behavioural development: differential expression of mRNAs for inositol 1,4,5-trisphosphate 3-kinase isoforms in naive and experienced honeybees (Apis mellifera).

In seeking genetic factors that may control the extended behavioural maturation of adult honeybees we found that inositol 1,4,5-trisphosphate (IP(3)) 3-kinase, a key enzyme in the IP(3)-mediated signalling cascade, is differentially expressed in brains of naive, newly emerged bees and experienced foragers. DNA sequencing yielded a contig of 21.5 kb spanning the honeybee IP(3)K locus and a 3' flanking gene similar to a transcription factor NFR-kappa-B. The IP(3)K locus gives rise to three differentially expressed major transcripts produced by alternative splicing that encode proteins with identical, highly conserved C-termini and distinct, non-conserved N-terminal domains. The type A transcript is dominant in the adult brain and its level of expression increases threefold during the first 4 days of adult development. The type B message is expressed in brains of naive bees, but is also found in the thorax and abdomen, whereas transcript C is expressed largely in non-neural tissues and in the antenna. In contrast to type A message, the brain levels of transcript B decrease during the first 4 days of adult life. Our data are evaluated in the context of the contrasting behavioural phenotypes of immature and experienced worker honeybees.

Real time PCR quantification of GFRalpha-2 alternatively spliced isoforms in murine brain and peripheral tissues.

The neurotrophic factor neurturin (NTN) is structurally related to the glial-derived neurotrophic factor (GDNF) and has been shown to prevent the degeneration of dopaminergic neurons both in vitro and in vivo. The preferred receptor for NTN is the GDNF family receptor alpha 2 (GFRalpha-2). To date, three protein-coding alternatively spliced GFRalpha-2 isoforms (GFRalpha-2a, GFRalpha-2b, GFRalpha-2c) have been identified in mammalian tissues. An accurate quantification of the expression levels is necessary when determining the contributions of these isoforms to NTN signaling in tissues. In this report, sequence independent real time RT-PCR is used to determine the expression levels of GFRalpha-2 isoforms at different developmental stages of the murine embryos, and in various adult tissues. In the adult murine brain, GFRalpha-2a was found to be the most abundant, GFRalpha-2c was slightly less and GFRalpha-2b was 10-fold lower. The testis did not appear to express significant levels of GFRalpha-2a, 2b or 2c, compared to the brain. A novel finding in this study is that in some tissues, including the adult brain, the expression levels of GFRalpha-2, as quantified by the amplification of the 3' sequences encoding the putative glycosyl-phosphatidylinositol anchor signal sequence, were significantly higher than the combined levels of GFRalpha-2a, GFRalpha-2b and GFRalpha-2c. This indicates the existence of yet to be identified forms of GFRalpha-2 in some tissues that may be of physiological significance.

Tissue restoration through regenerative biology and medicine.

The cost of tissue damage due to degenerative disease and injury is enormous in terms of health care costs, lost economic productivity, diminished quality of life and premature death. Advances in cell, developmental and molecular biology, and the discovery of regeneration-competent cells in many non-regenerating mammalian tissues, have given impetus to systematic investigations that will enable us to regenerate these tissues by cell transplantation or the pharmaceutical induction of regeneration from the body's own tissues. A significant avenue of research is the identification of the soluble and insoluble signals and their transduction pathways that govern the proliferation and differentiation of regeneration-competent cells, and the signals that inhibit their activity after injury. The most direct experimental strategy to identify the chemical and physical signals that promote regeneration and the factors that inhibit it is to make genomic and proteomic comparisons, using bioinformatic analyses, of regeneration-competent vs regeneration-deficient tissues. Once identified, the molecular promoters of regeneration can be used in clinical treatments. How far can we go in our quest for regeneration? We will probably be able to induce the regeneration of some tissues, such as skin or even spinal cord, within a few years. The regeneration of others, such as heart, lung, kidney or appendages, may be more complex and difficult, but we should not view them as impossible. They will just take a little longer.

In vivo and in vitro development of visceral adipose tissue in a nonhuman primate (Papio species).

We determined the development of the omental fat depot in a cross-sectional study of 242 baboons from birth to mature adulthood. The triglyceride content of the omentum increased during preweaning (birth to 4 months) and adolescence (2 to 5 years) and was associated with an increase in both fat cell number and size. Between weaning and 2 years of age omentum triglyceride mass decreased as a result of decreasing fat cell size, but fat cell number remained constant. After adolescence and up to 13 years of age, omental triglyceride mass and fat cell volume were stable, but fat cell number increased slightly in female baboons. We determined the in vitro potential of omental stromal vascular (S-V) cells from baboons at different stages of development to differentiate in a serum-free medium. Both the proportion of omental S-V cells that accumulated cytoplasmic lipid droplets and the induction of glycerol phosphate dehydrogenase (GPDH) activity were increased to the greatest degree in the presence of 1-methyl-3-isobutylxanthine, 2.0 nmol/L triiodothyronine (T3), 0.85 mumol/L insulin, and 1.0 mumol/L cortisol. Omental S-V cells from preweaning and adolescent baboons had a greater differentiation rate, GPDH activity, and triglyceride accumulation compared with cells from postweaned infants and mature adults. In summary, most of the growth of the baboon omentum occurs during the preweaning and pubertal periods of life, and omental S-V cells isolated from animals during these periods retain a greater potential to differentiate in vitro.

Systemic treatment with epidermal growth factor causes organ growth concomitant with reduced circulating levels of IGF-I and IGFBP-3: time-dependent changes in female rats.

Systemic treatment with epidermal growth factor (EGF) in neonatal rats reduces circulating levels of insulin-like growth factor I (IGF-I) and causes somatic growth retardation. In this study, we investigated the effects of EGF treatment on the IGF system and on visceral organ growth and longitudinal growth in mature rats. We treated female Wistar rats for 0 (n = 16), 1 (n = 8), 2 (n = 8), 3 (n = 8), or 4 (n = 8) weeks with subcutaneous EGF (150 microg/kg/day). The animals were weighed once a week. At sacrifice, various viscera were removed and weighed. Blood and serum samples obtained at sacrifice were analysed for growth hormone (GH), IGF-I, IGF binding proteins (IGFBPs) and various routine parameters. EGF treatment increased the total body weight. All parts of the gastrointestinal tract, the liver, the pancreas, the spleen, the bladder, the suprarenal glands and the ovaries increased proportionately more in weight than the increase in total body weight; the heart and the kidneys increased proportionately in weight whereas the weight of the perirenal fat was reduced. There were no changes in tail length but the mean length of the tibia was slightly increased in the group treated for 4 weeks with EGF. Circulating GH was unchanged but IGF-I and IGFBP-3 were reduced approximately 25 and 45%, respectively, in all EGF treated groups. There were no changes in the hepatic content of IGF-I and IGFBPs. In conclusion, systemic EGF treatment causes visceral growth concomitant with reduced circulating levels of IGF-I and IGFBP-3 in mature female rats.

Rates of protein synthesis in brain and other organs.

We previously found a decrease in protein synthesis in brain during development, which was much greater as measured in brain slices than in brain in vivo. In the present work such changes in brain were compared to those in other organs. With measurement of incorporation of flooding doses of [14C]valine into proteins of organs, the highest synthesis rate in the adult animal in vivo was found in liver (2.2%) followed by kidney (1.8%), spleen (1.6%), lung (1.0%), heart (0.7%), brain (0.6%) and muscle (0.5%). In immature animals the synthesis rate was highest in spleen (2.6%) followed by liver (2.4%), kidney (1.7%), lung (1.6%), brain (1.5%), heart (1.1%), and muscle (0.9%). Protein synthesis in slices from each tissue proceeded at lower rates than in vivo, especially in adults. The tissue affected the most by the preparation of the slices was muscle.

Lactational ethanol exposure: brain enzymes and [3H]spiroperidol binding.

Long-Evans lactating rats were fed 27% calories as ethanol in a liquid diet to determine whether alcohol received through the milk would alter normal brain development in the offspring. On days 16, 21 and 30, brains of the female offspring were removed, corpus striatum dissected and assayed for choline acetyltransferase activity, glutamic acid decarboxylase activity and [3H]spiroperidol binding activity. At day 16, there were no differences among the three treatment groups for the enzyme activities assayed. At day 21, glutamic acid decarboxylase activity in the pairfed group was higher than in ET and CT groups. Choline acetyltransferase activity in PF group was higher when compared to ad libitum controls and [3H]spiroperidol binding was not affected. At 30 days of age, animals exposed to ethanol had higher choline acetyltransferase activity and [3H]spiroperidol binding activity when compared to pairfed and ad libitum controls; and higher glutamic acid decarboxylase activity when compared to ad libitum controls. Data from the present study suggest that ethanol exposure during the brain growth spurt has a toxic effect on the late development of dopaminergic, cholinergic and GABAergic systems in the corpus striatum. These results may be related to the clinical symptoms of hyperactivity and problems with motor control in children exposed to alcohol during the third trimester and during lactation.

Mesenchymal stem cells from rat visceral fat exhibit multipotential differentiation in vitro.

Human subcutaneous fat-derived stem cells were recently shown to have the potential to differentiate in vitro into a variety of cell types, including adipocytes, osteoblasts, chondrocytes, and myoblasts (Zuk et al., Tissue Eng. 2001;7:211-228). Subcutaneous adipose tissue may therefore prove to be an easily acquired and abundant source of stem cells. Presently it is unclear whether mammals such as rats (which possess small or nonexistent subcutaneous fat pads) contain mesenchymal stem cells within the visceral fat of the abdominal cavity, or whether the visceral fat of any species contains stem cells. In this study we isolated and expanded a pool of mesenchymal cells from visceral fat of adult Sprague-Dawley rats and induced their differentiation in vitro into adipocytes, osteoblasts, neural cells, and chondrocytes. The differentiated phenotypes were verified by morphology as well as detection and expression of tissue-specific protein and mRNA. We conclude that despite well-documented differences in the metabolic and biochemical properties among anatomically distinct depots of fat, the visceral fat of rats contains adult mesenchymal stem cells with developmental potential similar to those isolated from subcutaneous fat in humans. Therefore, animals such as rats provide both a source of fat-derived stem cells and an immunocompetent, autologous host animal in which to investigate the capacity of the fat-derived cells to differentiate and form tissues in vivo.

Strain or deformation rate dependent finite growth in soft tissues.

The particular portion of the mechanical loading history of a tissue that serves as stimulus for growth or remodeling of that tissue is presently unknown. A kinematic basis for the implementation of strain or rate of deformation as a growth stimulus is presented. It is shown here that a recently proposed continuum theory for finite volumetric growth in soft tissue may be extended to include strain and rate of deformation as growth stimuli. The basis of the presentation is the recognition of two different time scales in the remodeling process, one on the order of seconds associated with the loading, and one on the order of weeks or months associated with the growth.

Fukutin expression in mouse non-muscle somatic organs: its relationship to the hypoglycosylation of alpha-dystroglycan in Fukuyama-type congenital muscular dystrophy.

Recent studies suggest that hypoglycosylation of alpha-dystroglycan (alpha-DG) may play an essential role in the pathogenesis of Fukuyama-type congenital muscular dystrophy (FCMD), which is caused by defects in the fukutin gene and characterized by dystrophic changes in the skeletal muscles and dysplastic lesions in the central nervous system. alpha-DG is expressed in many organs in addition to muscle and brain, although these organs are not affected in FCMD. To elucidate whether or not fukutin protein is involved in the glycosylation of alpha-DG in non-muscle somatic organs, we examined the distribution pattern of fukutin in developing and adult mouse tissues. The fukutin antisera labeled the acinar cells of the pancreas, the renal glomerular and tubular cells, and the epithelium of the bronchi, salivary gland, alimentary tract and skin in both fetal and adult mice. This distribution pattern was also confirmed by in situ hybridization. Antisera against alpha-DG and beta-DG labeled the same cellular populations in each organ, especially along the cell surface membrane. We also examined the glycosylation status of alpha-DG in autopsied FCMD cases (n = 5) and found evidence of hypoglycosylation in the kidney, lung, skin and intestine. These results suggest that fukutin protein is involved in the glycosylation process of alpha-DG in non-muscle somatic organs both during development and in the adult. It is unclear why muscle and brain symptoms predominate in FCMD, however re-evaluation of the functions of alpha-DG and fukutin protein in non-muscle somatic organs may provide valuable insight.