Transforming growth factor beta family expression at the bovine feto-maternal interface

<p>Abstract</p> <p>Background</p> <p>Endometrial remodelling is necessary for implantation in all mammalian species. The TGF beta super-family plays a crucial role in this event in humans and mice. However, the role of TGF beta super-family members during implantation is still unclear in ruminants. In the present study, the spacio-temporal expression of TGF beta super-family members including activin was explored in bovine trophoblasts and endometrial tissue during the peri-implantation period in order to elucidate whether it is essential for promoting cell proliferation at the implantation site.</p> <p>Methods</p> <p>Gene expression in the fetal membrane and endometrium of the gravid and non-gravid horn around Day 35 of gestation were analyzed with a custom-made oligo-microarray in cattle. The expression of activin and its related genes was also analyzed with quantitative RT-PCR. Activin-like activity in trophoblastic tissue and BT-1 cells was examined using a fibroblast cell proliferation test and Western blotting.</p> <p>Results</p> <p>The expression of various TGF beta super-family related genes including activin was detected in trophoblasts and the endometrium in cattle. The most intensive activin expression was found in the gravid horn endometrium, and rather intense expression was detected in the non-gravid trophoblastic tissue. Extracts from the fetal membrane including trophoblasts and purified activin both stimulated fibroblast proliferation effectively, and activin was immunologically detected in BT-1 cells, which have trophoblastic features.</p> <p>Conclusions</p> <p>Specific expression of the activin gene (gene name: inhibin beta A) was found in the gravid horn endometrium during peri-implantation. An activin-like molecule, which was derived from the endometrium and trophoblasts, stimulated the proliferation of fibroblast cells. These results suggested that as in other species, the activity of TGF beta super-family members including activin-like molecules plays a pivotal role in endometrial remodelling, which is an essential process in implantation and placentogenesis during the peri-implantation period in cattle.</p

Update on new aspects of the renin-angiotensin system in liver disease: clinical implications and new therapeutic options

A B S T R A C T The RAS (renin-angiotensin system) is now recognized as an important regulator of liver fibrosis and portal pressure. Liver injury stimulates the hepatic expression of components of the RAS, such as ACE (angiotensin-converting enzyme) and the AT 1 receptor [AngII (angiotensin II) type 1 receptor], which play an active role in promoting inflammation and deposition of extracellular matrix. In addition, the more recently recognized structural homologue of ACE, ACE2, is also up-regulated. ACE2 catalyses the conversion of AngII into Ang-(1-7) [angiotensin-(1-7)], and there is accumulating evidence that this &apos;alternative axis&apos; of the RAS has anti-fibrotic, vasodilatory and anti-proliferative effects, thus counterbalancing the effects of AngII in the liver. The RAS is also emerging as an important contributor to the pathophysiology of portal hypertension in cirrhosis. Although the intrahepatic circulation in cirrhosis is hypercontractile in response to AngII, resulting in increased hepatic resistance, the splanchnic vasculature is hyporesponsive, promoting the development of the hyperdynamic circulation that characterizes portal hypertension. Both liver fibrosis and portal hypertension represent important therapeutic challenges for the clinician, and there is accumulating evidence that RAS blockade may be beneficial in these circumstances. The present review outlines new aspects of the RAS and explores its role in the pathogenesis and treatment of liver fibrosis and portal hypertension

cDNA microarray analysis of bovine embryo gene expression profiles during the pre-implantation period

BACKGROUND: After fertilization, embryo development involves differentiation, as well as development of the fetal body and extra-embryonic tissues until the moment of implantation. During this period various cellular and molecular changes take place with a genetic origin, e.g. the elongation of embryonic tissues, cell-cell contact between the mother and the embryo and placentation. To identify genetic profiles and search for new candidate molecules involved during this period, embryonic gene expression was analyzed with a custom designed utero-placental complementary DNA (cDNA) microarray. METHODS: Bovine embryos on days 7, 14 and 21, extra-embryonic membranes on day 28 and fetuses on days 28 were collected to represent early embryo, elongating embryo, pre-implantation embryo, post-implantation extra-embryonic membrane and fetus, respectively. Gene expression at these different time points was analyzed using our cDNA microarray. Two clustering algorithms such as k-means and hierarchical clustering methods identified the expression patterns of differentially expressed genes across pre-implantation period. Novel candidate genes were confirmed by real-time RT-PCR. RESULTS: In total, 1,773 individual genes were analyzed by complete k-means clustering. Comparison of day 7 and day 14 revealed most genes increased during this period, and a small number of genes exhibiting altered expression decreased as gestation progressed. Clustering analysis demonstrated that trophoblast-cell-specific molecules such as placental lactogens (PLs), prolactin-related proteins (PRPs), interferon-tau, and adhesion molecules apparently all play pivotal roles in the preparation needed for implantation, since their expression was remarkably enhanced during the pre-implantation period. The hierarchical clustering analysis and RT-PCR data revealed new functional roles for certain known genes (dickkopf-1, NPM, etc) as well as novel candidate genes (AW464053, AW465434, AW462349, AW485575) related to already established trophoblast-specific genes such as PLs and PRPs. CONCLUSIONS: A large number of genes in extra-embryonic membrane increased up to implantation and these profiles provide information fundamental to an understanding of extra-embryonic membrane differentiation and development. Genes in significant expression suggest novel molecules in trophoblast differentiation

Mas-related G protein-coupled receptor type D antagonism improves portal hypertension in cirrhotic rats

Splanchnic vasodilatation contributes to the development and aggravation of portal hypertension (PHT). We previously demonstrated that in cirrhosis, angiotensin‐ mediates splanchnic vasodilatation through the Mas receptor (MasR). In this study, we investigated whether the recently characterized second receptor for angiotensin‐(1–7), Mas‐related G protein‐coupled receptor type D (MrgD), contributes to splanchnic vasodilatation in cirrhotic and noncirrhotic PHT. Splanchnic vascular hemodynamic and portal pressure were determined in two rat models of cirrhotic PHT and a rat model with noncirrhotic PHT, treated with either MrgD blocker D‐Pro(7)‐Ang‐(1‐7) (D‐Pro) or MasR blocker A779. Gene and protein expression of MrgD and MasR were measured in splanchnic vessels and livers of cirrhotic and healthy rats and in patients with cirrhosis and healthy subjects. Mesenteric resistance vessels isolated from cirrhotic rats were used in myographs to study their vasodilatory properties. MrgD was up‐regulated in cirrhotic splanchnic vessels but not in the liver. In cirrhotic rats, treatment with D‐Pro but not A779 completely restored splanchnic vascular resistance to a healthy level, resulting in a 33% reduction in portal pressure. Mesenteric vessels pretreated with D‐Pro but not with A779 failed to relax in response to acetylcholine. There was no splanchnic vascular MrgD or MasR up‐regulation in noncirrhotic PHT; thus, receptor blockers had no effect on splanchnic hemodynamics. Conclusion: MrgD plays a major role in the development of cirrhotic PHT and is a promising target for the development of novel therapies to treat PHT in cirrhosis. Moreover, neither MrgD nor MasR contributes to noncirrhotic PHT

The role of the vagal innervation of the stomach (abomasum and pylorus) and intestine (duodenum) in insulin and oxytocin release in sheep : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Animal Science at Massey University, Palmerston North, New Zealand

Although mechanisms regulating nutrient partitioning and milk synthesis are not fully understood in ruminants, recent studies in lactating monogastric animals have shown that the vagus nerve modulates secretion of various hormones that are implicated in the short- and long-term control of nutrient partitioning. Therefore, the overall aim of this study was to examine the role of the vagal innervation of the GI tract on insulin and oxytocin release and milk yield in sheep. In a series of experiments described in this thesis, the effect of vagotomy was studied in ewes and wethers by comparing the responses of vagotomized animals (i.e. abomasal, pyloric, duodenal and hepatic branches sectioned) with control (sham-operated) animals. Insulin release in response to a bolus injection of glucose was studied in lactating ewes (Chapters 2 and 4) or wethers (Chapters 3 and 5). The differences in responses were not significant in the experiments described in Chapters 3 and 4. However, the release of insulin from the pancreas in response to glucose injection was significantly (P < 0.05) suppressed in the vagotomized animals used in the experiments described in Chapters 2 and 5. Moreover in the experiments with wethers (Chapter 5), insulin secretion in response to glucose bolus injection was significantly (P < 0.05) higher when administered 2 h (i.e. fed state) following feeding than 22 h (i.e. fasting state). In addition, postprandial insulin concentrations were significantly (P < 0.05) lower in the vagotomized wethers than in the sham-operated wethers, but insulin secretion in the vagotomized wethers was apparently unaffected by plane of nutrition, despite significantly (P < 0.05) higher blood glucose levels in wethers on the HP intake. The insulin concentrations were, however, higher (P < 0.05) in the control group of wethers fed on the high plane (HP) of nutrition than those fed on the low plane (LP) of nutrition (Chapter 5). Insulin was released in response to the sight and/or ingestion of food, cephalic phase insulin release (CPIR), without any significant changes in blood glucose concentrations. However, the increase in insulin concentration was significantly (P < 0.05) suppressed in both vagotomized wethers and ewes in comparison with control animals (Chapters 5 and 6). Suckling increased plasma insulin concentrations in the sham-operated ewes but not in the vagotomized ewes (Chapter 6), although the difference in the concentrations between the two groups was not statistically significant (P < 0.09). Milk and fat yields were significantly (P < 0.05) reduced for one day and two days, respectively, in the vagotomized ewes compared with those of sham-operated controls, but was restored over the next 2-3 days (Chapter 2). Milk yield was not different between the two treatment groups in the second study (Chapter 4). Suckling-associated plasma oxytocin concentrations were significantly (P < 0.01) lower in the vagotomized ewes than in the sham-operated control ewes, although the difference was not statistically significant when corrected for baseline values (Chapter 4). In the next experiment (Chapter 6), oxytocin concentrations between the two treatment groups of ewes were not significantly different. However, in this experiment, suckling caused a significant (P < 0.05) increase in oxytocin concentrations from the baseline values in the sham-operated ewes fed and suckled simultaneously but not in the vagotomized ewes fed and suckled simultaneously. Vagotomy significantly (P < 0.05) increased digestibility of dry matter and nitrogen in wethers, although food intake was not different between the two treatment groups (Chapter 3). In conclusion, the findings in wethers (Chapter 5) agree with those in lactating ewes (Chapters 2 and 4) and, indicated that the effect of vagotomy on insulin release in response to glucose injection is more apparent over a short period (i.e. 2-4 h; Chapters 2 and 5) following feeding than after a longer period (i.e. 6-22 h; Chapters 4 and 5). This suggested that the pancreatic β-cells are more sensitive soon after feeding, because the vagal inputs reaching the β-cells from the GI tract are higher due to the recent consumption of food. The finding that post-prandial insulin concentrations in the vagotomized animals of HP group were significantly reduced, despite their significantly higher blood glucose levels, provide further evidence that the vagus nerve is a major determinant for sensitizing pancreatic β-cells. Furthermore the vagal innervation of the GI tract plays a major role in CPIR, and also appears to play an important role in insulin release during suckling in sheep. The concentrations of oxytocin measured in these experiments suggest that vagotomy interferes with oxytocin secretion although differences between vagotomized and sham operated ewes were often non significant. However, the data suggest that feeding stimulates OT secretion. It is possible that the failure to achieve consistent differences in milk ejection and hence removal in these studies may have been partly masked because of the anatomical features of the mammary gland of the ewe. Finally the activity of the vagus nerve influences the digestibility of dry matter and nitrogen in sheep

Development and progression of non-alcoholic fatty liver disease: the role of advanced glycation end products

Non-alcoholic fatty liver disease (NAFLD) affects up to 30% of the adult population and is now a major cause of liver disease-related premature illness and deaths in the world. Treatment is largely based on lifestyle modification, which is difficult to achieve in most patients. Progression of simple fatty liver or steatosis to its severe form non-alcoholic steatohepatitis (NASH) and liver fibrosis has been explained by a ‘two-hit hypothesis’. Whilst simple steatosis is considered the first hit, its transformation to NASH may be driven by a second hit. Of several factors that constitute the second hit, advanced glycation end products (AGEs), which are formed when reducing-sugars react with proteins or lipids, have been implicated as major candidates that drive steatosis to NASH via the receptor for AGEs (RAGE). Both endogenous and processed food-derived (exogenous) AGEs can activate RAGE, mainly present on Kupffer cells and hepatic stellate cells, thus propagating NAFLD progression. This review focuses on the pathophysiology of NAFLD with special emphasis on the role of food-derived AGEs in NAFLD progression to NASH and liver fibrosis. Moreover, the effect of dietary manipulation to reduce AGE content in food or the therapies targeting AGE/RAGE pathway on disease progression is also discussed

Portal pressure responses and angiotensin peptide production in rat liver are determined by relative activity of ACE and ACE2

Angiotensin converting enzyme (ACE) 2 activity and angiotensin-(1-7) [Ang-(1-7)] levels are increased in experimental cirrhosis; however, the pathways of hepatic Ang-(1-7) production have not been studied. This study investigated the role of ACE2, ACE, and neutral endopeptidase (NEP) in the hepatic formation of Ang-(1-7) from angiotensin I (Ang I) and Ang II and their effects on portal resistance. Ang I or Ang II were administered to rat bile duct ligated (BDL) and control livers alone and in combination with the ACE inhibitor lisinopril, the ACE and NEP inhibitor omapatrilat, or the ACE2 inhibitor MLN4760 (n = 5 per group). BDL markedly upregulated ACE, ACE2, and NEP. Ang-(1-7) was produced from Ang II in healthy and in BDL livers and was increased following ACE inhibition and decreased by ACE2 inhibition. In contrast, Ang-(1-7) production from Ang I was minimal and not affected by ACE or NEP inhibition. Surprisingly, ACE2 inhibition in BDLs dramatically increased Ang-(1-7) production from Ang I, an effect abolished by ACE2/NEP inhibition. Ang II and Ang I induced greater portal pressure increases in BDL livers than controls. The effects of Ang I were closely correlated with Ang II production and were strongly attenuated by both ACE and ACE/NEP inhibition. These findings show that the major substrate for hepatic production of Ang-(1-7) is Ang II and this is catalyzed by ACE2. Ang I is largely converted to Ang II by ACE, and net conversion of Ang I to Ang-(1-7) is small. NEP has the ability to generate large amounts of Ang-(1-7) in the BDL liver from Ang I only when ACE2 activity is greatly decreased or inhibited