Adipose Tissue Of Adult Rats Research Articles

EFFECTS OF PROGENITOR CELL CONDITIONED MEDIA ON THE AMOUNT OF BRAIN CORTEX NEURONS IN A RAT MODEL OF TRAUMATIC BRAIN INJURY

Aim. The purpose of the study was to examine beneficial effect of conditioned media (CM) of progenitor cells of different origin (neurogenic progenitor cells, or NPCs, and adipose-derived mononuclear cells, or AMCs) as a source of mesenchymal multipotent stromal cells (MMSCs) on brain cortex neurons in rats with traumatic brain injury (TBI). Methods. TBI was reproduced in outbred sexually mature male rats by developing the model of free-falling load (50 g) with damage to the left hemisphere of the brain. The rats were injected 3 times with an interval of every other day intraperitoneally with NPCs CM and AMCs CM (dose 0.8 mg by total protein) that were obtained from cell cultures of fetal rat brain and adult rat adipose tissue. On the 5th day after TBI, the morphologic study of brain tissue was performed. Results. The number of neurons in the cortex of rats on the 5th day after TBI in damaged hemisphere as well as in contralateral hemisphere compared to control group decreased by half. Three i.p. injections of NPCs CM or AMCs CM increased the number of neurons in the cortex in both hemispheres in rats of corresponding groups compared to the rats with TBI without additional treatment. Conclusion. Obtained results indicate that CM obtained from NPCs and AMCs have noticeable neuroprotective effect on the damaged neurons and might be considered as an additional mode to treatment of TBI.

Oligodendroglia Generated From Adult Rat Adipose Tissue by Direct Cell Conversion.

Obtaining oligodendroglial cells from dispensable tissues would be of great interest for autologous or immunocompatible cell replacement therapy in demyelinating diseases, as well as for studying myelin-related pathologies or testing therapeutic approaches in culture. We evaluated the feasibility of generating oligodendrocyte precursor cells (OPCs) from adult rat adipose tissue by expressing genes encoding transcription factors involved in oligodendroglial development. Adipose-derived mesenchymal cells were lentivirally transduced with tetracycline-inducible Sox10, Olig2, Zfp536, and/or Nkx6.1 transgenes. Immunostaining with the OPC-specific O4 monoclonal antibody was used to mark oligodendroglial induction. O4- and myelin-associated glycoprotein (MAG)-positive cells emerged after 3 weeks when using the Sox10 + Olig2 + Zfp536 combination, followed in the ensuing weeks by GFAP-, O1 antigen-, p75NTR (low-affinity NGF receptor)-, and myelin proteins-positive cells. The O4+ cell population progressively expanded, eventually constituting more than 70% of cells in culture by 5 months. Sox10 transgene expression was essential for generating O4+ cells but was insufficient for inducing a full oligodendroglial phenotype. Converted cells required continuous transgene expression to maintain their glial phenotype. Some vestigial characteristics of mesenchymal cells were maintained after conversion. Growth factor withdrawal and triiodothyronine (T3) supplementation generated mature oligodendroglial phenotypes, while FBS supplementation produced GFAP+- and p75NTR+-rich cultures. Converted cells also showed functional characteristics of neural-derived OPCs, such as the expression of AMPA, NMDA, kainate, and dopaminergic receptors, as well as similar metabolic responses to differentiation-inducing drugs. When co-cultured with rat dorsal root ganglion neurons, the converted cells differentiated and ensheathed multiple axons. We propose that functional oligodendroglia can be efficiently generated from adult rat mesenchymal cells by direct phenotypic conversion.

Gender-specific effects of intrauterine growth restriction on the adipose tissue of adult rats: a proteomic approach.

BackgroundIntrauterine growth restriction (IUGR) may program metabolic alterations affecting physiological functions and lead to diseases in later life. The adipose tissue is an important organ influencing energy homeostasis. The present study was aimed at exploring the consequences of IUGR on the retroperitoneal adipose tissue of adult male and female rats, using a proteomic approach.Methods and ResultsPregnant Wistar rats were fed with balanced chow, either ad libitum (control group) or restricted to 50 % of control intake (restricted group) during the whole gestation. The offspring were weaned to ad libitum chow and studied at 4 months of age. Retroperitoneal fat was analyzed by two-dimensional gel electrophoresis followed by mass spectrometry.Both male and female restricted groups had low body weight at birth and at weaning but normal body weight at adulthood. The restricted males had normal fat pads weight and serum glucose levels, with a trend to hyperinsulinemia. The restricted females had increased fat pads weight with normal glucose and insulin levels.The restricted males showed up-regulated levels of proteasome subunit α type 3, branched-chain-amino-acid aminotransferase, elongation 1- alpha 1, fatty acid synthase levels, cytosolic malate dehydrogenase and ATP synthase subunit alpha. These alterations point to increased proteolysis and lipogenesis rates and favoring of ATP generation.The restricted females showed down-regulated levels of L-lactate dehydrogenase perilipin-1, mitochondrial branched-chain alpha-keto acid dehydrogenase E1, and transketolase. These findings suggest impairment of glycemic control, stimulation of lipolysis and inhibition of proteolysis, pentose phosphate pathway and lipogenesis rates.In both genders, several proteins involved in oxidative stress and inflammation were affected, in a pattern compatible with impairment of these responses.ConclusionsThe proteomic analysis of adipose tissue showed that, although IUGR affected pathways of substrate and energy metabolism in both males and females, important gender differences were evident. While IUGR males displayed alterations pointing to a predisposition to later development of obesity, the alterations observed in IUGR females pointed to a metabolic status of established obesity, in agreement with their increased fat pads mass.Electronic supplementary materialThe online version of this article (doi:10.1186/s12953-015-0088-z) contains supplementary material, which is available to authorized users.

Effects of a Moderate Physical Training on the Leptin Synthesis by Adipose Tissue of Adult Rats Submitted to a Perinatal Low-protein Diet

The aim of the study was to verify if moderate physical training affects leptin content in visceral and subcutaneous adipose tissue of adult rats subjected to a low-protein diet during the perinatal period. Male Wistar rats were divided into 2 groups according to their mother's diet during gestation and lactation: control (17% casein, C, n=12) and low-protein (8% casein, LP, n=12). On postnatal day 60, half of each group was submitted to moderate physical training (8 wks, 5 d · wk - 1, 60 min · d - 1, at 70% of VO2max, T) or not. After the physical training period, visceral and subcutaneous adipose tissues were removed. Leptin content was evaluated by western blotting. Starting from the fifth week on, T pups showed a reduction in the body weight. Similarly, LP+T offspring showed a lower body weight starting from the sixth week on. Western blotting analysis showed that leptin content in the visceral tissue was higher in the LP rats (p<0.01) and it was reversed in LP+T. No difference was found in the subcutaneous tissue. Moderate physical training attenuated the effects of a perinatal low-protein diet on the leptin content in visceral adipose tissue in adult offspring.

The Interactions Between Rat-Adipose-Derived Stromal Cells, Recombinant Human Bone Morphogenetic Protein-2, and Beta-Tricalcium Phosphate Play an Important Role in Bone Tissue Engineering

Cells, scaffolds, and growth factors are the three main factors for creating a stem-cell-based tissue-engineered construct, but the interactions between three factors are not very clear. We hereby explored the interactions between rat-adipose-derived stromal cells (rASCs), recombinant human bone morphogenetic protein-2 (rhBMP-2), and beta-tricalcium phosphate (beta-TCP) to provide evidence for their application in bone tissue engineering by evaluating the protein adsorption of beta-TCP, the cell attachment, alkaline phosphatase (ALP) activity/protein, osteocalcin (OCN) content, mineral formation, calcium content, phosphonium content, cell vitality, gene expression, and implantation in the backs of severe combined immunodeficient mice of rhBMP-2 preinducing rASCs seeded onto beta-TCP. The results showed that beta-TCP could adsorb the proteins from the media. The attachment, proliferation, and osteogenic properties of rASCs were supported by beta-TCP, as revealed using scanning electron microscopy. Compared with rASCs cultured on the culture plate, rASCs cultured on beta-TCP had significantly higher ALP activity/protein, OCN content, and mineral formation. These values for rASCs cultured on beta-TCP with rhBMP-2 increased most significantly. The rhBMP-2 significantly increased the calcium content, phosphonium content, and ALP, type I collagen, and OCN mRNA levels of rASCs cultured on beta-TCP. The methylthiazol tetrazolium method revealed that the vitality of rASCs cultured on beta-TCP with or without rhBMP-2 for 4, 7, and 28 days in vitro was insignificantly different. After 8 and 12 weeks of implantation, each group displayed increased bone formation over the 12-week period. The percentage of the new bone formed areas for beta-TCP/rhBMP-2 and beta-TCP was not significantly different. This value for rASCs/beta-TCP construct was significantly higher than that for beta-TCP group, but the maximal and robust bone formation was presented in rASCs/beta-TCP with rhBMP-2. The results implied that stem cells existed in adult rat adipose tissue. beta-TCP could adsorb rhBMP-2 from the media and had osteoinductivity when alone implanted in the back of severe combined immunodeficient mice. beta-TCP was also sufficient to trigger the differentiation of rASCs toward an osteoblastic phenotype without the addition of osteogenic factor. The rhBMP-2 could better sufficiently induce osteogenic differentiation of rASCs seeded onto beta-TCP. The rASCs and rhBMP-2 could promote the dissolution of beta-TCP to provide Ca2+ and PO4(3-) needed for bone formation. The interactions between the three factors could provide an optimizing microenvironment for osteogenic differentiation of rASCs, and this might be essential for sufficient and timely bone formation in vivo. This study may provide insight into the clinical repair of bone defect with ASCs+beta-TCP+rhBMP-2 construct.

Neonatal leptin exposure induces altered expression of specific PPARα transcripts in adipose tissue of adult rats

Neonatal leptin exposure induces altered expression of specific PPARα transcripts in adipose tissue of adult rats

Covalent modification of porous implants using extracellular matrix proteins to accelerate neovascularization

Healing associated with many polymeric biomedical implants commonly involves the formation of an avascular fibrous capsule. The lack of either formation or persistence of blood vessels in formed fibrous capsules, as well as a lack of new blood vessels within porous polymeric implants, often results in poor performance of the implant. The current study evaluated the use of extracellular matrix protein modification of a commonly used biomedical implant material, expanded polytetrafluoroethylene (ePTFE), as a mechanism to increase the neovascularization both within these porous implants and in tissue that forms in the peri-implant area. Discs of ePTFE were covalently modified with different extracellular matrix proteins including collagen type IV, fibronectin, and laminin type I. Discs were implanted into the adipose tissue of adult rats, and following a 5-week implant phase, histologic analysis of peri-implant tissue angiogenesis and implant neovascularization was performed. Striking differences were observed in angiogenic and neovascularization responses to matrix-modified ePTFE when compared with control, untreated ePTFE. Fibronectin treatment resulted in an extensive inflammatory response but, relative to the degree of inflammation, limited evidence of tissue angiogenesis or polymer neovascularization. Collagen type IV treatment groups exhibited a significant increase in angiogenesis in the peri-implant tissue with minimal evidence of implant neovascularization. In contrast to all other implant modifications, laminin type 1-treated ePTFE samples stimulated an extensive peri-implant tissue angiogenic response and a coordinate neovascularization of the porous interstices of the biomaterial.

TH-, NPY-, SP-, and CGRP-immunoreactive nerves in interscapular brown adipose tissue of adult rats acclimated at different temperatures: an immunohistochemical study.

Interscapular brown adipose tissue (IBAT), a site of nonshivering thermogenesis in mammals, is neurally controlled. The co-existence of sympathetic and peptidergic innervation has been demonstrated in different brown adipose depots. We studied the morphological profile of IBAT innervation and tested by immunohistochemical methods whether cold and warm stimulation are accompanied by modifications in the density of parenchymal noradrenergic nerve fibers. We also studied the immunoreactivity of afferent fibers--which contain calcitonin gene-related peptide (CGRP) and substance P (SP)--in different functional conditions. IBAT was obtained from adult rats (6 weeks old) acclimated at different temperatures (4 degrees, 20 degrees, and 28 degrees C). Tissue activity was evaluated by studying the immunolocalization of uncoupling protein (UCP-1), a specific marker of brown adipose tissue. Noradrenergic and peptidergic innervation were seen to arise from morphologically different nerves. Fibers staining for tyrosine hydroxylase (TH) were thin, unmyelinated hilar nerves, and CGRP- and SP-positive fibers were in thick nerves containing both myelinated and unmyelinated fibers. Under cold stimulation, noradrenergic neurons produce greater amounts of TH, and their axons branch, resulting in increased parenchymal nerve fibers density. Neuropeptide Y (NPY) probably co-localizes with TH in noradrenergic neurons, but only in the perivascular nerve fiber network. The parenchymal distribution of NPY to interlobular arterioles and capillaries suggests that this peptide must have other functions besides that of innervating arteriovenous anastomoses, as hypothesized by other researchers. The different distribution of CGRP and SP suggests the existence of different sensory neuronal populations. The detection of CGRP at the parenchymal level is in line with the hypothesis of a trophic action of this peptide.

Consumption of a Low Protein Diet Increases Norepinephrine Turnover in Brown Adipose Tissue of Adult Rats

Adult rats were fed diets containing 5 or 15% casein to determine effects of low protein intake on efficiency of energy retention and on brown adipose tissue metabolism. Efficiency of energy retention of rats fed the 5% casein diet for 4 or 8 weeks was approximately 40% lower than was efficiency of energy retention of rats fed the 15% casein diet. Rats fed the 5% casein diet also exhibited an enhanced norepinephrine (NE)-stimulated metabolic rate and an accelerated NE turnover in their brown adipose tissue relative to responses of rats fed the 15% casein diet. Additionally, NE turnover in white adipose tissue, but not heart, was accelerated in rats fed the 5% casein diet. The reduced efficiency of energy retention in rats fed the low protein diet may be linked to accelerated NE turnover and heat production by their brown adipose tissue.

Continuity of intracellular channels with extracellular space in adipose tissue and liver: demonstrated with tannic acid and lanthanum.

Tannic acid was used to demonstrate continuity of intracellular channels with extracellular space in white adipose tissue of adult rats, brown adipose tissue of suckling rats, and liver of diabetic rats. Electron-opaque material resulting from treatment of glutaraldehyde-fixed tissue with tannic acid was found in extracellular space, invaginations of cell surfaces, vesicles, and intracellular channels. Electron-opaque material was present in channels that surrounded lipid droplets in both white and brown adipocytes and in hepatocytes. The small distance between the lumen of marked channels and lipid droplets in adipocytes indicates that a monolayered structure, perhaps a leaflet of membrane lining the channel. Similar findings were obtained in brown adipose tissue using lanthanum instead of tannic acid to mark intracellular channels continuous with extracellular space. Since endoplasmic reticulum is the primary site of triacylglycerol synthesis in adipocytes, marked channels near lipid droplets may be elements of endoplasmic reticulum. Some of the channels marked with tannic acid in hepatocytes contained lipoprotein particles, whereas others were located, in relation to mitochondria and lipid droplets, in the same sites as endoplasmic reticulum in untreated tissue. This indicates that some of the channels marked with tannic acid in hepatocytes are endoplasmic reticulum. Presence of electron-opaque material in intracellular channels and vesicles, but not in cytoplasm, of treated tissue indicates the channels and vesicles were open to extracellular space during treatment with tannic acid or lanthanum and, furthermore, that their membranes were continuous with plasma membrane.

Increased cellular proliferation in adipose tissue of adult rats fed a high-fat diet.

The feeding of a high-fat diet to adult rats was shown to increase the incorporation of [3H]thymidine into DNA of the adipocyte and stromal fractions. After only 2 days on a high-fat diet there was a marked increase in the incorporation of label. When a 2-week period was interposed between [3H]thymidine administration and determination of DNA specific activity, the greatest increase in incorporation of label was found after 1 week on the diet, when incorporation increased 6-fold or more in both adipocytes and stroma and subsequently decreased to stabilize at a level two or three times that of chow-fed rats in the adipocyte fraction. Rats labeled when young and later placed on a high-fat diet showed a decrease in DNA specific activity in both adipocytes and stroma, confirming that cellular proliferation had occurred in both fractions. The specific activities of both stromal and adipocyte DNA were very similar at all time points studied. An attempt to increase the difference in specific activities by waiting many weeks after [3H]thymidine injection before isolating DNA was not successful. This may be because the total amount of DNA in the stromal and adipocyte fractions increases in parallel on the diet. The significance of these findings in terms of the normal turnover of adipose tissue DNA and the responsiveness to diet is discussed.

Isolation of fat cell precursors from adult rat adipose tissue

The possible existence of adipocyte precursors in adult rat adipose tissue was investigated. Cells were isolated from the stromal fraction of adipose tissue and were grown in culture. Skin fibroblasts were used as controls. The stromal fraction cells were initially fusiform and proliferated; in culture, they accumulated lipid inclusions, became rounder and acquired an eccentric nucleus. In contrast, the skin fibroblasts from the same rat grown under identical culture conditions, did not exhibit any appreciable lipid accumulation. The doubling time for both the stromal fraction cells and skin fibroblasts was 40-60 h. At confluency, the stromal fraction cells contained 5-7 times more glyceride-glycerol than skin fibroblasts. Thus, adipose tissue of adult rats contains cells with the potential to proliferate and acquire morphological characteristics similar to those of adipocytes.

Electron microscopic localization of adenylate cyclase activity of white and brown adipose tissue of the rat and chicken.

Brown adipose tissue of newborn rats and chicken embryos and white adipose tissue of adult rats were studied. Adenylate cyclase (EC 4.6.1.1.) activity stimulated by 0.1 mmol/l noradrenaline was demonstrated using an electron microscopic histochemical method. The reaction product was visualized as a cobalt salt in the plasmalemma of the adipocytes. The adipocytes of the brown adipose tissue of the newborn rats showed most intense reaction in the outer surfaces of their plasmalemma. Alloxan totally inhibited the enzymatic reaction. The histochemical reaction used in the present study probably demonstrated the hormonal receptor sites in the plasmalemmas of the adipocytes which are stimulated by noradrenaline.

Effect of fasting on the esterfied α-glycerol ethers of the neutral lipids of heart and adipose tissue of rats

The effect of prolonged fasting on the lipids of cardiac muscle and adipose tissue of adult rats has been studied with special emphasis on the acylated α-glycerol ethers of the neutral lipids. Adipose tissue was found to be an excellent source of neutral lipid glycerol ethers as well as plasmalogens, the latter being present in amounts previously observed only in brain. In the neutral lipids of adipose tissue 75–80 per cent of the total glycerol ethers were present as ether diesters. The remaining 20–25 per cent of the glycerol ethers consisted of ether monoesters and α-glycerol ethers in similar amounts. In contrast, the triacyl glycerides represented 97 per cent of the total glycerides of adipose tissue. Fasting has little effect on the total glycerol ether content of adipose tissue and cardiac muscle, but results in marked changes in the triacyl glyceride content of these tissues. The concentration of neutral glycerides was found to increase in cardiac muscle only while the stores of triacyl glycerides in adipose had not been depleted. Once the neutral glycerides of adipose tissue started to decrease precipitously, the endogenous glycerides of cardiac muscle declined, suggesting that the non-esterified fatty acid content of the plasma was no longer sufficient to meet the energy needs of the cardiac muscle cell.