The ACE2/Ang-(1-7)/Mas Axis Regulates the Development of Pancreatic Endocrine Cells in Mouse Embryos

Angiotensin-converting enzyme 2 (ACE2), its product Angiotensin-(1-7) [Ang-(1-7)], and Ang-(1-7) receptor Mas, have been shown to regulate organogenesis during embryonic development in various species. However, it is not known whether a local ACE2/Ang-(1-7)/Mas axis is present in the fetal pancreas. It is hypothesized that there is a local ACE2/Ang-(1-7)/Mas axis in the embryonic pancreas in mice that is involved in regulating islet cell development. To address this issue, the endogenous expression profile of axis constituents in embryonic mouse pancreata was examined. Involvement of the ACE2 axis in the regulation of pancreatic development was also examined. The present experiments showed in an in vivo animal model that endogenous expression levels of ACE2 and the Mas receptor were upregulated in mouse pancreata in late embryogenesis, peaking on embryonic day E16.5, when it reached 3 folds compared to that seen at E12.5. Consistently, endogenous expression of Ang-(1-7) also peaked at E16.5. Treatment with the ACE2 inhibitor DX600 did not alter islet development. However, prenatal treatment with A779, a Mas receptor antagonist, reduced the β-cell to α-cell ratio in neonatal islets, impaired islet insulin secretory function, and impaired the pups’ glucose tolerance. In ex vivo pancreas explant cultures, A779 again decreased the β-cell to α-cell ratio, apparently through its effects on β-cell proliferation (reduced proliferation shown with Ki67 staining), and also decreased Insulin and Ngn3 mRNA expression. Furthermore, treatment of explant cultures with Ang-(1-7) increased mRNA levels of Insulin and pancreatic progenitor marker Ngn3, as well as Nox4, the ROS generation enzyme; these stimulatory effects were attenuated by co-treatment with A779, suggesting that Ang-(1-7), via Mas receptor signaling, may promote differentiation of pancreatic progenitors into insulin-producing cells via modulation of reactive oxygen species. These data together suggest that a Mas receptor-mediated mechanism may stimulate pancreatic cell development.     

Fibroblast growth factor receptor signaling is essential for lens fiber cell differentiation

The vertebrate lens provides an excellent model to study the mechanisms that regulate terminal differentiation. Although fibroblast growth factors (FGFs) are thought to be important for lens cell differentiation, it is unclear which FGF receptors mediate these processes during different stages of lens development. Deletion of three FGF receptors (Fgfr1-3) early in lens development demonstrated that expression of only a single allele of Fgfr2 or Fgfr3 was sufficient for grossly normal lens development, while mice possessing only a single Fgfr1 allele developed cataracts and microphthalmia. Profound defects were observed in lenses lacking all three Fgfrs. These included lack of fiber cell elongation, abnormal proliferation in prospective lens fiber cells, reduced expression of the cell cycle inhibitors p27^kip1 and p57^kip2, increased apoptosis and aberrant or reduced expression of Prox1, Pax6, c-Maf, E-cadherin and α-, β- and γ-crystallins. Therefore, while signaling by FGF receptors is essential for lens fiber differentiation, different FGF receptors function redundantly.     

Down-regulating annexin gene GhAnn2 inhibits cotton fiber elongation and decreases Ca2+ influx at the cell apex

Cotton fiber is a single cell that differentiates from the ovule epidermis and undergoes synchronous elongation with high secretion and growth rate. Apart from economic importance, cotton fiber provides an excellent single-celled model for studying mechanisms of cell-growth. Annexins are Ca²⁺- and phospholipid-binding proteins that have been reported to be localized in multiple cellular compartments and involved in control of vesicle secretions. Although several annexins have been found to be highly expressed in elongating cotton fibers, their functional roles in fiber development remain unknown. Here, 14 annexin family members were identified from the fully sequenced diploid G. raimondii (D₅ genome), half of which were expressed in fibers of the cultivated tetraploid species G. hirsutum (cv. YZ1). Among them, GhAnn2 from the D genome of the tetraploid species displayed high expression level in elongating fiber. The expression of GhAnn2 could be induced by some phytohormones that play important roles in fiber elongation, such as IAA and GA₃. RNAi-mediated down-regulation of GhAnn2 inhibited fiber elongation and secondary cell wall synthesis, resulting in shorter and thinner mature fibers in the transgenic plants. Measurement with non-invasive scanning ion-selective electrode revealed that the rate of Ca²⁺ influx from extracellular to intracellular was decreased at the fiber cell apex of GhAnn2 silencing lines, in comparison to that in the wild type. These results indicate that GhAnn2 may regulate fiber development through modulating Ca²⁺ fluxes and signaling.     

Induction of a dwarf phenotype with IBH1 may enable increased production of plant‐made pharmaceuticals in plant factory conditions

Year‐round production in a contained, environmentally controlled ‘plant factory’ may provide a cost‐effective method to produce pharmaceuticals and other high‐value products. However, cost‐effective production may require substantial modification of the host plant phenotype; for example, using dwarf plants can enable the growth of more plants in a given volume by allowing more plants per shelf and enabling more shelves to be stacked vertically. We show here that the expression of the chimeric repressor for Arabidopsis AtIBH1 (P35S:AtIBH1SRDX) in transgenic tobacco plants (Nicotiana tabacum) induces a dwarf phenotype, with reduced cell size. We estimate that, in a given volume of cultivation space, we can grow five times more AtIBH1SRDX plants than wild‐type plants. Although, the AtIBH1SRDX plants also showed reduced biomass compared with wild‐type plants, they produced about four times more biomass per unit of cultivation volume. To test whether the dwarf phenotype affects the production of recombinant proteins, we expressed the genes for anti‐hepatitis B virus antibodies (anti‐HBs) in tobacco plants and found that the production of anti‐HBs per unit fresh weight did not significantly differ between wild‐type and AtIBH1SRDX plants. These data indicate that P35S:AtIBH1SRDX plants produced about fourfold more antibody per unit of cultivation volume, compared with wild type. Our results indicate that AtIBH1SRDX provides a useful tool for the modification of plant phenotype for cost‐effective production of high‐value products by stably transformed plants in plant factory conditions.     

Inhibition of the angiotensin-converting enzyme decreases skeletal muscle fibrosis in dystrophic mice by a diminution in the expression and activity of connective tissue growth factor (CTGF/CCN-2)

The renin-angiotensin system (RAS), through angiotensin II and the angiotensin-converting enzyme (ACE), is involved in the genesis and progression of fibrotic diseases characterized by the replacement of normal tissue by an accumulation of an extracellular matrix (ECM). Duchenne muscular dystrophy (DMD) presents fibrosis and a decrease in muscle strength produced by chronic damage. The mdx mouse is a murine model of DMD and develops the same characteristics as dystrophic patients when subjected to chronic exercise. The connective tissue growth factor (CTGF/CCN2) and transforming growth factor type beta (TGF-β), which are overexpressed in muscular dystrophies, play a major role in many progressive scarring conditions. We have tested the hypothesis that ACE inhibition decreases fibrosis in dystrophic skeletal muscle by treatment of mdx mice with the ACE inhibitor enalapril. Both sedentary and exercised mdx mice treated with enalapril showed improvement in gastrocnemius muscle strength explained by a reduction in both muscle damage and ECM accumulation. ACE inhibition decreased CTGF expression in sedentary or exercised mdx mice and diminished CTGF-induced pro-fibrotic activity in a model of CTGF overexpression by adenoviral infection. Enalapril did not have an effect on TGF-β1 expression or its signaling activity in sedentary or exercised dystrophic mice. Thus, ACE inhibition might improve muscle strength and decrease fibrosis by diminishing specifically CTGF expression and activity without affecting TGF-β1 signaling. Our data provide insights into the pathogenic events in dystrophic muscle. We propose ACE as a target for developing therapies for DMD and related diseases.