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Highlights • A distinctive miRNA signature between visceral and subcutaneous adipose tissues. • The novel role and function of miR-122 in adipose tissue. • The ratio of miR-122 between visceral and subcutaneous adipose tissues correlates with bariatric surgery outcome. Abstract The physiological roles and clinical impacts of the differences between visceral fat (VF) and subcutaneous fat (SF) are unclear. The present study aimed to compare the miRNA signatures between visceral fat (VF) and subcutaneous fat (SF) and study their influences on outcomes of bariatric surgery. To study the microRNA signatures of the VF and SF in obesity, we performed paired microRNA arrays of the adipose tissues from 20 bariatric surgery patients. The microRNA analysis identified miR-122 as the most significant signature between VF and SF. The tissue distribution, functions, and influences on adipogensis of miR-122 were analysed by Northern blotting, microRNA mimics and inhibitors, and whole-genome microarray analysis. The outcomes of body weight changes after bariatric surgery were analysed and correlated with the miR-122 abundances. Northern blotting confirmed that miR-122 was highly expressed in VF and SF. Bioinformatics analysis of the microarray revealed that proliferator activator receptor-γ (PPAR-γ) signalling was critically affected by miR-122. The modulation of PPAR-γ by miR-122 was confirmed in murine adipocytes and human adipose tissues. Furthermore, the differentiation of preadipocytes was significantly influenced by miR-122. In obese patients receiving bariatric surgery, the ratio of VF and SF miR-122 abundance correlated with 6-month and 1-year % excess body weight loss. Our findings indicate that miR-122 is highly expressed in adipose tissue. The abundance of miR-122 affects PPAR-γ signalling and adipocytes differentiation in vitro and human adipose tissues. Higher miR-122 in VF may be associated with greater body weight loss after bariatric surgery. [ABSTRACT FROM AUTHOR]

Background: Brown adipose tissue (BAT) contains both α- and β-adrenergic receptors. In the literature, the activity of α-adrenoreceptors is less documented, and their functions still remain puzzling. The present investigation has been undertaken to understand α-adrenoreceptors’ activity and their relation between uncoupling protein 1 (UCP1) mRNA expression and cyclic AMP (cAMP) generation in BAT. Methods: BAT precursor cells from young mice were grown in culture. Cells were exposed to norepinephrine (NE) and other agents. RNA was isolated after harvesting the cells, and northern blot was performed. Filters were exposed to film after hybridization with nick-translated complementary DNA probes, and results were evaluated by scanning. Amersham assay kit was used for cAMP measurement. Results: Treatment of prazosin and yohimbine separately with 1 μM of NE shows stimulation of UCP1 mRNA expression 106% and 154%, respectively, whereas with that of both drugs shows only 76%. cAMP generation occurs 282% with prazosin, 100% with yohimbine, and 382% with both drugs with 1 μM of NE, whereas it is 310%, 40%, and 358%, respectively, with 10 μM of NE. Conclusions: Stimulation of thermogenesis after treatment of prazosin and NE may be due to the inhibition of phosphodiesterase enzyme and with yohimbine and NE indicates the possibility of inhibition of the inhibitory effect of α2- and stimulation of α1-receptors. Increase of cAMP concentration with yohimbine and both drugs with NE are not correlated to UCP1 mRNA expression. This indicates that the relationship between cAMP elevation and stimulation of thermogenesis is not simple. This study clearly shows the interaction between β- and α-adrenoreceptor activities. [ABSTRACT FROM AUTHOR]

The article discusses research on the stable isotope fractionation in fish otoliths and muscles. It references a study by T. S. Elsdon at the University of Adelaide in South Australia, and his colleagues, published in the June 3, 2010 issue of "Marine Ecology Progress Series." The researchers studied the factors controlling the trophic fractionation of nitrogen and carbon stable isotopes in fish otoliths and muscles as well as whether fractionation was consistent in fishes with varying diets. They conclude that questions still remain regarding the fundamental assumptions of stable isotope analyses (SIA) as well as the right analytical techniques needed.

The article focuses on the ecto-5'-nucleotidase (CD73) which is greatly expressed on endothelial cells. It notes that CD73 functions to catalyze the terminal phosphohydrolysis of 5'-adenosine monophosphate and regulates the barrier function on endothelial cells. It mentions that CD73 modulates the vascular immune regulation under homeostatic conditions and under stress conditions performs a dysregulation.

The article reports on the disruption of the insulin⁄insulin-like growth factor-I signaling (IS) pathway and phenotypic analyses of the gene-disrupted animal. The gene-disruption by ins-7 RNAi knockdown of another insulin like gene, showed a significant influence on an adult lifespan, where it antagonized the lifespan extension induced. By gene-disruption, a clarification on the physiological function of ins-11, C.elegans insulin-like gene is provided by the study.

Background: The reconstruction of lost periodontal support including bone, ligament, and cementum is a major goal of therapy. Bone morphogenetic proteins (BMPs) have shown much potential in the regeneration of the periodontium. Limitations of BMP administration to periodontal lesions include need for high-dose bolus delivery, BMP transient biological activity, and low bioavailability of factors at the wound site. Gene transfer offers promise as an alternative treatment strategy to deliver BMPs to periodontal tissues.Methods: This study utilized ex vivo BMP-7 gene transfer to stimulate tissue engineering of alveolar bone wounds. Syngeneic dermal fibroblasts (SDFs) were transduced ex vivo with adenoviruses encoding either green fluorescent protein (Ad-GFP or control virus), BMP-7 (Ad-BMP-7), or an antagonist of BMP bioactivity, noggin (Ad-noggin). Transduced cells were seeded onto gelatin carriers and then transplanted to large mandibular alveolar bone defects in a rat wound repair model.Results: Ad-noggin treatment tended to inhibit osteogenesis as compared to the control-treated and Ad-BMP-7-treated specimens. The osseous lesions treated by Ad-BMP-7 gene delivery demonstrated rapid chrondrogenesis, with subsequent osteogenesis, cementogenesis and predictable bridging of the periodontal bone defects.Conclusion: These results demonstrate the first successful evidence of periodontal tissue engineering using ex vivo gene transfer of BMPs and offers a new approach for repairing periodontal defects. [ABSTRACT FROM AUTHOR]

Objective: The study was conducted to determine whether relaxin has a proliferative effect on amniotic epithelial cells and to show that this effect is caused by its stimulation of the insulin-like growth factor-II (IGF-II) gene.Study Design: Immunolocalization and Northern analysis were used to confirm the expression of IGF-II by the fetal cells in the membranes. Human amniotic epithelial (WISH) cells were treated with doses of IGF-II or human relaxin and their proliferative effects measured. The mechanism of the effect of relaxin on cellular proliferation was studied with the use of an IGF-II-blocking antibody and Northern analysis for IGF-II gene expression after treatment with relaxin. An in vivo correlate was sought by quantitation of relaxin gene expression in 10 fetal membranes from women with normally grown and large for gestational age infants.Results: The amniotic epithelial and cytotrophoblast cells of the fetal membranes expressed IGF-II, as did the amniotic epithelial-like (WISH) cell line. Treatment of WISH cells with IGF-II or relaxin caused a significant (P <.03) and dose-related increase in WISH cell proliferation over 5 days. The concurrent treatment with a blocking antibody to IGF-II significantly decreased the proliferative response to IGF-II (P <.002) and relaxin (P <.002). Treatment with relaxin caused a significant increase (P <.003) in the transcription of IGF-II in 24 hours. In fetal membranes, the levels of relaxin gene expression correlated with fetal membrane surface area (r = 0.76) and was significantly greater (P <.008) in the membranes from macrosomic infants (4020-4729 g) compared with those normally grown (2855-3830 g).Conclusion: IGF-II and relaxin both caused the proliferation of WISH cells. Concurrent treatment with an IGF-II-blocking antibody abrogated the proliferative effects of both hormones. Relaxin increased the transcription of IGF-II, and its expression levels in the fetal membranes correlated with the membrane surface area as well as neonatal birth weight. These data suggest that relaxin is a growth factor for the fetal membranes. [ABSTRACT FROM AUTHOR]

Objective: Transforming growth factor-beta1 is the prototype of a bimodal regulator of cell growth, which can either inhibit or stimulate the proliferation of smooth muscle cells. Part of transforming growth factor-beta1-mediated stimulation of growth is associated with the increased production of platelet-derived growth factor. The conversion of latent-to-active transforming growth factor-beta provides a pivotal mechanism for the regulation of the biologic activity of transforming growth factor-beta. We investigated the differential expression and production of the active form of transforming growth factor-beta1 in the myometrium and leiomyoma throughout the menstrual cycle. We also studied the mitogenic effects of transforming growth factor-beta1 and platelet derived growth factor on myometrial and leiomyoma cells in culture.Study Design: Myometrium and leiomyoma tissue pairs were obtained from 28 women who underwent hysterectomy. Total RNA from each tissue was extracted, and Northern blot analysis was performed for the detection of TGF-beta1 messenger RNA. Active and total transforming growth factor-beta1 protein was quantified with enzyme-linked immunosorbent assay. Cell proliferation of cultured human myometrial and leiomyoma cells that are treated with TGF-beta1 (0.01-1 ng/mL), anti-transforming growth factor-beta antibody (0.01-10 ng/mL), or platelet-derived growth factor (10 ng/mL) was assessed by the [(3)H]thymidine incorporation method.Results: Overall, the transforming growth factor-beta1 messenger RNA level in myometrial samples was 1.2-fold higher than in the leiomyoma samples (P <.05). Active transforming growth factor-beta1 protein levels in follicular and luteal phase myometrial and leiomyoma samples were significantly greater than the levels in samples from women with atrophic endometrium (P < 0.05). Transforming growth factor-beta1, at low concentrations (0.01 ng/mL), induced an increase in cell proliferation (2- to 3-fold; P <.05). When cells were treated with anti-transforming growth factor-beta antibody, there was a larger magnitude of increase observed (7- to 20-fold; P <.05). Platelet-derived growth factor (10 ng/mL) consistently increased the rate of cell proliferation both in myometrium and leiomyoma cells (5- to 6-fold; P <.05).Conclusion: Levels of active transforming growth factor-beta1 that were produced in follicular and luteal phases indicate a stimulatory role for ovarian hormones. The finding that transforming growth factor-beta1, only at low concentrations, stimulates cell proliferation mainly in leiomyoma cells is in agreement with the bimodal and dose-dependent effects of transforming growth factor-beta1 that is observed in smooth muscle cells of other tissues. The persistent and high rate of cell proliferation with platelet-derived growth factor suggests that the growth stimulatory effect of transforming growth factor-beta1 may be mediated through its up-regulatory effect on platelet-derived growth factor. [ABSTRACT FROM AUTHOR]

Background: A challenge in the reconstruction of periodontal structures is the targeted delivery of growth-promoting molecules to the tooth root surface. Polypeptide growth factors such as platelet-derived growth factor (PDGF) stimulate both cementogenesis and osteogenesis. Recent advances in gene therapy offer the advantage of delivering recombinant proteins to tissues for extended periods of time in vivo.Methods: Recombinant adenoviral vectors encoding for the PDGF-A gene were constructed to allow delivery of PDGF transgenes to cells. The recombinant adenoviruses were assembled using the viral backbone of Ad2/CMV/EGFP and replacing GFP (reporter gene encoding green fluorescent protein driven by the cytomegalovirus promoter [CMV] within adenovirus type 2) with the PDGF-A gene. Root lining cells (cloned cementoblasts) were transduced with Ad2/PDGF-A and evaluated for gene expression, DNA synthesis, and cell proliferation. PDGF-inducible genes, c-myc and osteopontin, were also evaluated following gene delivery of Ad2/PDGF-A.Results: The results revealed high level transduction of cementoblasts by gene transfer for 7 days as evidenced by flow cytometry and Northern blotting. Cementoblast DNA synthesis and subsequent proliferation were stimulated by Ad2/PDGF-A at levels equal to or greater than continuous rhPDGF-AA application. Strong message for the PDGF-A gene and protein as evidenced by Northern blotting and immunocytochemistry was noted. Furthermore, the potent induction of c-myc and osteopontin mRNA was found after PDGF gene delivery to cementoblasts.Conclusions: These findings demonstrate that gene delivery of platelet-derived growth factor stimulates cementoblast activity that is sustained above that of rhPDGF-AA application. The use of gene therapy as a mode of growth factor delivery offers a novel approach to periodontal tissue engineering. [ABSTRACT FROM AUTHOR]

Objective: We previously identified a protein, Brx, that interacted with estrogen receptor alpha. Sequence analysis determined that Brx is a novel member of the Dbl family of oncoproteins involved in signaling pathways that regulate cell growth. Because the Brx protein was found to be highly expressed in hormoneresponsive breast epithelium, the objective of this study was to determine whether Brx was expressed in both normal and neoplastic ovarian tissues.Study Design: A polyclonal antiserum directed against the Brx protein was used to perform immunolocalization on sections from 5 normal ovaries and 20 ovarian neoplasms. Chromosomal localization of the brx gene was accomplished by means of fluorescence in situ hybridization. Northern and Western blot analyses were performed on extracts prepared from human ovaries.Results: Brx protein was localized to the cytoplasm of granulosa cells from mature graafian follicles, the corpus luteum, and islands of hilar cells in normal ovaries. In tumors with low malignant potential and ovarian carcinomas the neoplastic epithelium stained strongly for Brx protein. Northern and Western blot analyses, respectively, confirmed expression of Brx messenger ribonucleic acid and protein in normal ovary. Finally, the brx gene was localized to 15q25.Conclusion: The proto-oncogene brx is expressed in specific normal human ovarian tissues and is also present in ovarian epithelial neoplasms. [ABSTRACT FROM AUTHOR]

Objective: This study was undertaken to determine which genes were up-regulated by acute distention in an amniotic epithelial cell line and in human fetal membranes.Study Design: WISH cells, a human amniotic epithelial cell line, were grown on silicone elastomer sheets coated with extracellular matrix and reproducibly distended by 40% in a novel device for 4 hours. Differential gene expression was analyzed by means of suppression subtractive hybridization. Expression of the identified genes was then quantitated by Northern blot analysis in fetal membrane explants after distention in the same device for 4 hours. The effect of distention on apoptosis of the cells and tissue samples was concomitantly studied by means of the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling method.Results: The genes for interleukin 8 and pre-B-cell colony-enhancing factor were found to be up-regulated in both the WISH cells and the distended fetal membranes. The apoptotic index values in both the cells and the tissue samples were unaffected by distention.Conclusions: Acute distention induces the up-regulation of interleukin 8 and pre-B-cell colony-enhancing factor in both WISH cells and human fetal membranes and does not cause apoptosis. [ABSTRACT FROM AUTHOR]

Objective: This study was undertaken to identify genes with expression up-regulated by acute distention in the human fetal membranes.Study Design: Fetal membrane explants were distended reproducibly in a novel device in vitro for 4 hours, and suppression subtractive hybridization was used to identify the candidate genes for up-regulation of expression in response to this stimulus. The up-regulation in response to distention was confirmed by quantitative Northern blot analysis both after a 4-hour in vitro distention and after labor in vivo.Results: Suppression subtractive hybridization identified 3 genes with expression up-regulated by acute distention: an interferon-stimulated gene encoding a 54-kd protein, the gene for huntingtin-interacting protein 2 (a ubiquitin-conjugating enzyme), and a novel transcript. Expression of each of the distention-responsive genes found to be up-regulated in vitro was also up-regulated in fetal membranes in association with labor.Conclusions: Suppression subtractive hybridization was successfully applied to a complex tissue, the human fetal membranes, and 3 novel distention-responsive genes were identified. Both acute in vitro distention and labor in vivo up-regulate expression of at least 3 genes in the human fetal membranes. [ABSTRACT FROM AUTHOR]

Objective: The purposes of this study were to determine (1) whether uterine tissues as well as the fetal placenta are involved in the development of prostaglandin-synthesizing capacity associated with impending labor in pregnant sheep and (2) whether the key enzyme of prostaglandin synthesis, prostaglandin H synthase 2, is differentially expressed in the different intrauterine tissues during late gestation and in association with labor.Study Design: Myometrium, endometrium, and fetal placenta were removed from ewes at 95 days' gestation, (n = 3), 101 to 110 days' gestation (n = 3), 111 to 120 days' gestation (n = 3), 121 to 130 days' gestation (n = 3), 131 to 140 days' gestation (n = 3), and 141 to 145 days' gestation (n = 4) and from ewes in spontaneous term labor at 143 to 147 days' gestation (n = 4). Expressions of prostaglandin H synthase 2 messenger ribonucleic acid and protein were determined by Northern blot and Western blot analyses. Prostaglandin H synthase 2 was localized in the fetal placenta by immunohistochemical means.Results: Levels of both prostaglandin H synthase 2 messenger ribonucleic acid and protein increased gradually from 115 days' gestation in the fetal placenta and from 131 days' gestation in the endometrium. A further and more significant increase in prostaglandin H synthase 2 concentration occurred in the placenta and endometrium during spontaneous term labor. In contrast, myometrial concentrations of prostaglandin H synthase 2 messenger ribonucleic acid and protein remained at steady basal levels during the course of pregnancy and increased only during labor. Prostaglandin H synthase 2 was localized in the trophoblast cells of the fetal placenta.Conclusions: Tissue-specific ontogenic expression of prostaglandin H synthase 2 was observed in myometrium, endometrium, and placenta during late ovine gestation and spontaneous term labor. Fetal placenta and endometrium showed increased expression of prostaglandin H synthase 2 messenger ribonucleic acid and protein during late ovine gestation, whereas myometrial prostaglandin H synthase 2 concentration remained low throughout late gestation. Prostaglandin H synthase 2 concentrations in the myometrium, endometrium, and placenta are all upwardly regulated during labor. [ABSTRACT FROM AUTHOR]

Objective: We sought to investigate the expression of cannabinoid receptors in human placenta and BeWo choriocarcinoma cells and study their role in the regulation of the serotonin transporter.Study Design: Expression of the 2 types of cannabinoid receptors (CB1 and CB2) in human placenta and BeWo cells was investigated by reverse transcriptase-polymerase chain reaction and Northern blot analysis. The involvement of the receptors in the regulation of the serotonin transporter expression was studied by using a cannabinoid receptor agonist (WIN 55212-2). BeWo cells were treated with the agonist in the presence or absence of forskolin, and the serotonin transporter activity was measured by assessing paroxetine-sensitive serotonin transport. Serotonin transporter density in cell membranes was monitored by measuring paroxetine-sensitive binding of RTI-55, a specific high-affinity ligand for the transporter. Agonist-induced changes in intracellular levels of cyclic adenosine monophosphate were also monitored.Results: Reverse transcriptase-polymerase chain reaction and Northern blot analysis demonstrated unequivocally that human placenta and BeWo cells express both types of cannabinoid receptors. Treatment of BeWo cells with the receptor agonist blocked the activity of the constitutive, as well as the forskolin-induced, serotonin transporter without affecting the serotonin transporter density. This effect is not mediated by alterations in intracellular cyclic adenosine monophosphate levels.Conclusion: The results show that cannabinoid receptors are expressed in human placenta and BeWo cells and play a role in the regulation of the serotonin transporter activity. Human placenta is therefore a direct target for cannabinoids, and marijuana use during pregnancy is likely to affect the placental clearance of serotonin through the serotonin transporter. [ABSTRACT FROM AUTHOR]

Objective: Our purpose was to determine differential effects of natural and synthetic glucocorticoids on cytochrome 17alpha-hydroxylase and cytochrome P-450 side-chain cleavage messenger ribonucleic acid in the sheep placenta and to determine whether feed-forward effects during labor are involved in further inducing placental cytochrome 17alpha-hydroxylase.Study Design: Sheep underwent placement of myometrial electromyogram electrodes while they were under general anesthesia at 117 days' gestation. At 125 days' gestation either saline solution (early control animals not in labor, n = 5), 0.48 mg betamethasone during 48 hours (n = 7), 0.48 mg dexamethasone during 48 hours (n = 7), or 55 mg cortisol during a maximum of 96 hours (n = 4) was directly administered intravenously to the fetus. Necropsies were performed at 127 to 129 days' gestation. We also studied 6 ewes in spontaneous term labor at 143-147 days' gestation, 6 term control animals not in labor at 140 to 147 days' gestation, and 6 sheep in which myometrial activity was inhibited by intravenous infusion to the ewe of the selective cyclooxygenase 2 inhibitor nimesulide 9 hours after the onset of labor beginning at 147 to 148 days' gestation. Total fetal placental ribonucleic acid was analyzed by Northern blot with complementary deoxyribonucleic acid probes for cytochrome 17alpha-hydroxylase, cytochrome P-450 side-chain cleavage, and 18S ribosomal ribonucleic acid to correct for loading.Results: Placental cytochrome 17alpha-hydroxylase messenger ribonucleic acid was detectable neither in term control animals not in labor nor in early control animals not in labor. Placental cytochrome 17alpha-hydroxylase messenger ribonucleic acid was induced in spontaneous term labor and all cortisol-infused sheep in labor with respect to term control animals not in labor and early control animals not in labor (P <.01). All betamethasone-infused sheep had myometrial contraction activity; however, only 4 of 7 had detectable placental cytochrome 17alpha-hydroxylase messenger ribonucleic acid. Placental cytochrome 17alpha-hydroxylase messenger ribonucleic acid was not detected in dexamethasone-infused sheep, even the 2 that had myometrial contractions. After reversal of the progression of spontaneous labor with nimesulide placental cytochrome 17alpha-hydroxylase messenger ribonucleic acid was significantly lower than the spontaneous term labor group (without nimesulide treatment). The placentas from all animals expressed cytochrome P-450 side-chain cleavage messenger ribonucleic acid, but no changes were associated with either gestational age studied (130 versus > 140 days' gestation) or glucocorticoid-induced premature labor and spontaneous term labor.Conclusions: (1) In sheep the expression of placental cytochrome 17alpha-hydroxylase is tightly associated with spontaneous term labor, and active synthesis of placental cytochrome 17alpha-hydroxylase is required during the progression of labor. (2) Cortisol is a more potent stimulator of placental cytochrome 17alpha-hydroxylase messenger ribonucleic acid in sheep than are synthetic glucocorticoids. (3) Betamethasone has a greater effect in inducing labor in sheep than does dexamethasone, possibly mediated through placental cytochrome 17alpha-hydroxylase. [ABSTRACT FROM AUTHOR]

Objective: Our purpose was to determine whether human ovarian surface epithelial cells express calcium-sensing receptors and whether changes in extracellular calcium modulate the proliferation of these cells.Study Design: Ovarian surface epithelial cells from normal patients and from ovarian tumors were tested for calcium-sensing receptor expression by Northern hybridization and immunoblot analysis. Functional responses to agonists of the calcium-sensing receptor were monitored by inositol triphosphate analysis and measurements of intracellular calcium release. The effect of extracellular calcium on proliferation was monitored by thymidine incorporation and growth curve analysis.Results: Increasing extracellular calcium above 0.8 mmol/L produces a marked proliferative response in normal ovarian surface epithelial cells. Both normal and transformed cells express calcium-sensing receptor messenger ribonucleic acid and protein; some tumor lines overexpress calcium-sensing receptor messenger ribonucleic acid. The calcium-sensing receptors in normal ovarian surface epithelial cells respond to the agonist gadolinium with increases in inositol triphosphate production and calcium release.Conclusion: Human ovarian surface epithelial cells are growth regulated by extracellular calcium, and calcium-sensing receptors may mediate this response. [ABSTRACT FROM AUTHOR]

Objective: The purpose of this study was to evaluate the regulation of lipoprotein lipase activity, protein mass, and messenger ribonucleic acid by estradiol.Study Design: Premenopausal women not taking exogenous sex steroids had transdermal 17 beta-estradiol and placebo patches placed in the gluteal region during the early follicular phase of the menstrual cycle. Adipose biopsies were performed from beneath the patches. Adipose tissue lipoprotein lipase activity was determined by a radiometric assay, protein mass was determined by enzyme-linked immunosorbent assay, and messenger ribonucleic acid level was determined by Northern analysis. Comparisons between the treated and placebo sides were analyzed by nonparametric statistics.Results: Adipose tissue from beneath the 17 beta-estradiol patch had significantly decreased lipoprotein lipase activity and extracellular protein mass than did adipose tissue from beneath the placebo patch. There was no difference in lipoprotein lipase messenger ribonucleic acid levels.Conclusion: Estrogen decreases lipoprotein lipase activity by a posttranscriptional modification of protein levels. A hypothesis of sex steroid regulation of body fat distribution is proposed. [ABSTRACT FROM AUTHOR]