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Metabolism and ageing are intimately linked. Compared with ad libitum feeding, dietary restriction consistently extends lifespan and delays age-related diseases in evolutionarily diverse organisms (1,2). Similar conditions of nutrient limitation [...]

Capacitative [Ca.sup.2+] entry (CCE) is [Ca.sup.2+] entering after stimulation of inositol 1,4,5-trisphosphate (IP3) formation and initiation of [Ca.sup.2+] store depletion. One hallmark of CCE is that it can also be triggered merely by store depletion, as occurs after inhibition of internal [Ca.sup.2+] pumps with thapsigargin. Evidence has accumulated in support of a role of transient receptor potential (Trp) proteins as structural subunits of a class of [Ca.sup.2+]-permeable cation channels activated by agonists that stimulate IP3 formation - very likely through a direct interaction between the IP3 receptor and a Trp subunit of the [Ca.sup.2+] entry channel. The role of Trp's in [Ca.sup.2+] entry triggered by store depletion alone is less clear. Only a few of the cloned Trp's appear to enhance this type of [Ca.sup.2+] entry, and when they do, the effect requires special conditions to be observed, which native CCE does not. Here we report the full-length cDNA of mouse trp2, the homologue of the human trp2 pseudogene. Mouse Trp2 is shown to be readily activated not only after stimulation with an agonist but also by store depletion in the absence of an agonist. In contrast to other Trp proteins, Trp2-mediated [Ca.sup.2+] entry activated by store depletion is seen under the same conditions that reveal endogenous store depletion-activated [Ca.sup.2+] entry, i.e., classical CCE. The findings support the general hypothesis that Trp proteins are subunits of store- and receptor-operated [Ca.sup.2+] channels.

Mammalian genes related to the Drosophila trp gene encode proteins that form the subunits of the channels required for agonist-activated capacitative Ca2+ entry (CCE) into mammalian cells. In COS cells, the expression of two full-length cDNA's encoding the human trp homologs, Htrp1 and Htrp3, increase CCE by 75% and 230%, respectively. Ca2+ influx due to endogenous COS cell CCE channels is more sensitive to suppression by La3+ and Ni2+ than that mediated by Htrp3. The expression of the genes in the antisense orientation inhibits CCE.

Experimental studies of the primary structure and ligand-binding behavior of a human opsin-related gene encoding a retinaldehyde binding protein from bovine retinal pigment epithelium (RPE) reveal that RPE-retinal G protein coupled receptor (RGR) binds to 11-cis- and all-trans-retinal and the amino acid sequence of the protein is 86% similar to bovine RGR sequence. The 14.8 kb gene is present as seven exons with a structure different from that of visual pigment genes. The all-trans-retinal binding property of RGR indicates that it catalyzes chromophore isomerization.

[Figure: see text]. [ABSTRACT FROM AUTHOR]

Abstract Background Transgenic animals have become valuable tools for both research and applied purposes. The current method of gene transfer, microinjection, which is widely used in transgenic mouse production, has only had limited success in producing transgenic animals of larger or higher species. Here, we report a linker based sperm-mediated gene transfer method (LB-SMGT) that greatly improves the production efficiency of large transgenic animals. Results The linker protein, a monoclonal antibody (mAb C), is reactive to a surface antigen on sperm of all tested species including pig, mouse, chicken, cow, goat, sheep, and human. mAb C is a basic protein that binds to DNA through ionic interaction allowing exogenous DNA to be linked specifically to sperm. After fertilization of the egg, the DNA is shown to be successfully integrated into the genome of viable pig and mouse offspring with germ-line transfer to the F1 generation at a highly efficient rate: 37.5% of pigs and 33% of mice. The integration is demonstrated again by FISH analysis and F2 transmission in pigs. Furthermore, expression of the transgene is demonstrated in 61% (35/57) of transgenic pigs (F0 generation). Conclusions Our data suggests that LB-SMGT could be used to generate transgenic animals efficiently in many different species.

Interleukin 6 (IL6) and tumor necrosis factor contribute to the development of colitis-associated cancer (CAC). We investigated these signaling pathways and the involvement of G protein subunit alpha i1 (GNAI1), GNAI2, and GNAI3 in the development of CAC in mice and humans. B6;129 wild-type (control) or mice with disruption of Gnai1 , Gnai2 , and/or Gnai3 or conditional disruption of Gnai2 in CD11c+ or epithelial cells were given dextran sulfate sodium (DSS) to induce colitis followed by azoxymethane (AOM) to induce carcinogenesis; some mice were given an antibody against IL6. Feces were collected from mice, and the compositions of microbiomes were analyzed by polymerase chain reactions. Dendritic cells (DCs) and myeloid-derived suppressor cells (MDSCs) isolated from spleen and colon tissues were analyzed by flow cytometry. We performed immunoprecipitation and immunoblot analyses of colon tumor tissues, MDSCs, and mouse embryonic fibroblasts to study the expression levels of GNAI1, GNAI2, and GNAI3 and the interactions of GNAI1 and GNAI3 with proteins in the IL6 signaling pathway. We analyzed the expression of Gnai2 messenger RNA by CD11c+ cells in the colonic lamina propria by PrimeFlow, expression of IL6 in DCs by flow cytometry, and secretion of cytokines in sera and colon tissues by enzyme-linked immunosorbent assay. We obtained colon tumor and matched nontumor tissues from 83 patients with colorectal cancer having surgery in China and 35 patients with CAC in the United States. Mouse and human colon tissues were analyzed by histology, immunoblot, immunohistochemistry, and/or RNA-sequencing analyses. GNAI1 and GNAI3 (GNAI1;3) double-knockout (DKO) mice developed more severe colitis after administration of DSS and significantly more colonic tumors than control mice after administration of AOM plus DSS. Development of increased tumors in DKO mice was not associated with changes in fecal microbiomes but was associated with activation of nuclear factor (NF) κB and signal transducer and activator of transcription (STAT) 3; increased levels of GNAI2, nitric oxide synthase 2, and IL6; increased numbers of CD4+ DCs and MDSCs; and decreased numbers of CD8+ DCs. IL6 was mainly produced by CD4+/CD11b+, but not CD8+, DCs in DKO mice. Injection of DKO mice with a blocking antibody against IL6 reduced the expansion of MDSCs and the number of tumors that developed after CAC induction. Incubation of MDSCs or mouse embryonic fibroblasts with IL6 induced activation of either NF-κB by a JAK2-TRAF6-TAK1-CHUK/IKKB signaling pathway or STAT3 by JAK2. This activation resulted in expression of GNAI2, IL6 signal transducer (IL6ST, also called GP130) and nitric oxide synthase 2, and expansion of MDSCs; the expression levels of these proteins and expansion of MDSCs were further increased by the absence of GNAI1;3 in cells and mice. Conditional disruption of Gnai2 in CD11c+ cells of DKO mice prevented activation of NF-κB and STAT3 and changes in numbers of DCs and MDSCs. Colon tumor tissues from patients with CAC had reduced levels of GNAI1 and GNAI3 and increased levels of GNAI2 compared with normal tissues. Further analysis of a public human colorectal tumor DNA microarray database (GSE39582) showed that low Gani1 and Gnai3 messenger RNA expression and high Gnai2 messenger RNA expression were significantly associated with decreased relapse-free survival. GNAI1;3 suppresses DSS-plus-AOM–induced colon tumor development in mice, whereas expression of GNAI2 in CD11c+ cells and IL6 in CD4+/CD11b+ DCs appears to promote these effects. Strategies to induce GNAI1;3, or block GNAI2 and IL6, might be developed for the prevention or therapy of CAC in patients. [ABSTRACT FROM AUTHOR]

Lack of muscarinic regulation of [Ca.sup.2+] channels in [G.sub.i2][Alpha] gene knockout mouse hearts. Am J Physiol Heart Circ Physiol 280: H1989-H1995, 2001.--The purpose of the present study was to examine the role of [G.sub.i2][Alpha] in [Ca.sup.2+] channel regulation using [G.sub.i2][Alpha] gene knockout mouse ventricular myocytes. The whole cell voltage-clamp technique was used to study the effects of the muscarinic agonist carbachol (CCh) and the [Beta]-adrenergic agonist isoproterenol (Iso) on cardiac L-type [Ca.sup.2+] currents in both 129Sv wild-type (WT) and [G.sub.i2][Alpha] gene knockout ([G.sub.i2][Alpha]-/-) mice. Perfusion with CCh significantly inhibited the [Ca.sup.2+] current in WT cells, and this effect was reversed by adding atropine to the CCh-containing solution. In contrast, CCh did not affect [Ca.sup.2+] currents in [G.sub.i2][Alpha]-/- ventricular myocytes. Addition of CCh to Iso-containing solutions attenuated the Iso-stimulated [Ca.sup.2+] current in WT cardiomyocytes but not in [G.sub.i2][Alpha]-/- cells. These findings demonstrate that, whereas the Iso-[G.sub.s][Alpha] signal pathway is intact in [G.sub.i2][Alpha] gene knockout mouse hearts, these cells lack the inhibitory regulation of [Ca.sup.2+] channels by CCh. Therefore, [G.sub.i2][Alpha] is necessary for the muscarinic regulation of [Ca.sup.2+] channels in the mouse heart. Further studies are needed to delineate the possible interaction of [G.sub.i] and other cell signaling proteins and to clarify the level of interaction of G protein-coupled regulation of L-type [Ca.sup.2+] current in the heart. [Ca.sup.2+] current regulation; gene inactivation; signal transduction

Homologues of Drosophilia transient receptor potential (TRP) have been proposed to be unitary subunits of plasma membrane ion channels that are activated as a consequence of active or passive depletion of [Ca.sup.2+] stores. In agreement with this hypothesis, cells expressing TRPs display novel [Ca.sup.2+]-permeable cation channels that can be activated by the inositol 1,4,5-trisphosphate receptor (IP3R) protein. Expression of TRPs alters cells in many ways, including up-regulation of IP3Rs not coded for by TRP genes, and proof that TRP forms channels of these and other cells is still missing. Here, we document physical interaction of TRP and IP3R by coimmunoprecipitation and glutathione S-transferase-pulldown experiments and identify two regions of IP3R, F2q and F2g, that interact with one region of TRP, C7. These interacting regions were expressed in cells with an unmodified complement of TRPs and IP3Rs to study their effect on agonist- as well as store depletion-induced [Ca.sup.2+] entry and to test for a role of their respective binding partners in [Ca.sup.2+] entry. C7 and an F2q-containing fragment of IP3R decreased both forms of [Ca.sup.2+] entry. In contrast, F2g enhanced the two forms of [Ca.sup.2+] entry. We conclude that store depletion-activated [Ca.sup.2+] entry occurs through channels that have TRPs as one of their normal structural components, and that these channels are directly activated by IP3Rs. IP3Rs, therefore, have the dual role of releasing [Ca.sup.2+] from stores and activating [Ca.sup.2+] influx in response to either increasing IP3 or decreasing luminal [Ca.sup.2+].

Summary We discovered recently that the central metabolite α-ketoglutarate (α-KG) extends the lifespan of C. elegans through inhibition of ATP synthase and TOR signaling. Here we find, unexpectedly, that ( R )-2-hydroxyglutarate (( R )-2HG), an oncometabolite that interferes with various α-KG-mediated processes, similarly extends worm lifespan. ( R )-2HG accumulates in human cancers carrying neomorphic mutations in the isocitrate dehydrogenase (IDH) 1 and 2 genes. We show that, like α-KG, both ( R )-2HG and ( S )-2HG bind and inhibit ATP synthase and inhibit mTOR signaling. These effects are mirrored in IDH1 mutant cells, suggesting a growth-suppressive function of ( R )-2HG. Consistently, inhibition of ATP synthase by 2-HG or α-KG in glioblastoma cells is sufficient for growth arrest and tumor cell killing under conditions of glucose limitation, e.g., when ketone bodies (instead of glucose) are supplied for energy. These findings inform therapeutic strategies and open avenues for investigating the roles of 2-HG and metabolites in biology and disease. [ABSTRACT FROM AUTHOR]

The mechanic performance of ejecting pneumatic fire extinguisher is determined by the following parameters: air speed at funnel exit, outfire range and ejecting stability; and the second and third parameters depend on the fire extinguishing agent pellet's velocity on funnel exit and the amount of fire extinguishing agent pellets detained in the funnel when extinguisher is working. In this research, the CFD-DEM coupling model was adopted to simulate the working process of ejecting pneumatic fire extinguisher, and the type of 6MF-30 pneumatic extinguisher was used to test the reliability of the coupling model. The funnel throat angle X1, the adding position of fire extinguishing agent X2, and the funnel length X3 were selected as the influencing factors, and total 23 experiments were conducted by the simulation model under the quadratic orthogonal rotation design. The virtual tunnel was designed according to the tunnel equipped to 6MF-30 pneumatic extinguisher. The virtual fire extinguishing agent pellets were found on the EDEM interface, coupled to the original CFD by Lagrangian model. The pretest results showed that this model had strong convergence which was proved by its stable airflow performance when running at the time of 0.2-0.25 s, and stable pellet number when running at 0.34 s. The air velocity at exit calculated by the CFD-DEM coupling model was similar to the theoretical value. It could be found that the number of aggregating virtual pellets detained in the region between the air tunnel and the fire extinguishing agent's channel was consistent with the result of research by other scholars. The simulating experiment data were exported for analyzing. The average velocity in axial direction on each point at funnel exit was read by Fluent, and after subtracting the average value 197.550, the deviation data of the 23 experiments were got and selected as the outlet wind velocity (index 71). The max virtual pellet velocities in axial direction monitored by EDEM within 0-0.4 s of all experiments were selected as the spouting velocity (index Y2). The vertical coordinates of every virtual pellet at 0.37, 0.38, 0.39, 0.4 s in all experiments were read by EDEM, and 'countif' function was used to calculate the number of virtual pellet located in the tunnel (as assessment index Y3) when working. After the significant test and the variance analysis, the regression equation building was done by using SAS9.1. The result showed that the fire extinguishing agent pellet's velocity on funnel exit and the amount of fire extinguishing agent pellets detained in the funnel were significantly correlated to the three factors (P<0.0001, P<0.001 respectively), but the air speed at funnel exit had no correlation with these factors. By using response surface method combined with weighting method, the optimal aggregative index could be obtained under the condition that the funnel throat angle was 20°, the adding position of fire extinguishing agent was 140 mm and the funnel length was 700 mm. Under this condition, the efficiency of ejecting pneumatic fire extinguisher was mostly improved. The virtual test designed by the optimized parameters showed that the fire extinguishing agent pellet's velocity on funnel exit increased to 41.24 m/s, and the number of fire extinguishing agent pellets detained in the funnel decreased to 209. The verify test of the physical prototype indicated that the optimized parameters could improve the working effect of ejecting pneumatic fire extinguisher, which was manifested in extending the outfire range by 2.15 m, reducing the possibility of reburning and shortening the outfire time by 1.7 s. [ABSTRACT FROM AUTHOR]

ABSTRACT The p. Arg116 His mutation in the heat shock transcription factor-4 ( HSF4) has been associated with age-related cataracts, but it is also seen in 2% of the normal population, indicating either reduced penetrance or that the normal subjects were not old enough to express the phenotype. Based on the proximity of p. Arg116 His to two known mutations in the DNA-binding domain of HSF4, namely, p. Leu114 Pro and p. Arg119 Cys, which segregate with childhood lamellar cataract, we tested the possibility that this phenotype may have been missed by the ophthalmologist and/or that it did not spread to the visual axis so as to affect vision significantly. Here, we demonstrate via BAC (bacterial artificial chromosome) transgenesis that p. Arg116 His recreates the childhood lamellar cataract in mice suggesting that incomplete penetrance associated with early cataracts may not be an absence but a limitation of the detection of the phenotype. [ABSTRACT FROM AUTHOR]

The prereceptor activation of glucocorticoid production in adipose tissue by NADPH-dependent 11 hydroxysteroid dehydrogenase type 1 (11β-HSD1) has emerged as a potential mechanism in the pathogenesis of visceral obesity and metabolic syndrome. Hexose-6-phosphate dehydrogenase (H6PDH) is an endoplasmic reticulum lumen-resident enzyme that generates cofactor NADPH and thus mediates 11β-HSD1 activity. To determine the role of adipose H6PDH in the prereceptor modulation of 11β- HSD1 and metabolic phenotypes, we generated a transgenic (Tg) mouse model overexpressing H6PDH under the control of the enhancerpromoter region of the adipocyte fatty acid-binding protein (aP2) gene (aP2/H6PDH Tg mice). Transgenic aP2/H6PDH mice exhibited relatively high expression of H6PDH and elevated corticosterone production with induction of 11β-HSD1 activity in adipose tissue. This increase in corticosterone production in aP2-H6PDH Tg mice resulted in mild abdominal fat accumulation with induction of C/EBP mRNA expression and slight weight gain. Transgenic aP2/H6PDH mice also exhibited fasting hyperglycemia and glucose intolerance with insulin resistance. In addition, the aP2/H6PDH Tg mice have elevated circulating free fatty acid levels with a concomitant increased adipose lipolytic action associated with elevated HSL mRNA and Ser660 HSL phosphorylation within abdominal fat. These results suggest that increased H6PDH expression specifically in adipose tissue is sufficient to cause intra-adipose glucocorticoid production and adverse metabolic phenotypes. These findings suggest that the aP2/H6PDH Tg mice may provide a favorable model for studying the potential impact of H6PDH in the pathogenesis of human metabolic syndrome [ABSTRACT FROM AUTHOR]

A portable pneumatic extinguisher is an effective device that has been widely used for forest and grassland fire extinguishing in China. In order to enhance the effective range and rate of discharge of portable pneumatic extinguishers, a new method to weaken air velocity attenuation by increasing the flow rate using an air ejector was proposed and investigated in this study. This research belongs to the category of subsonic air ejector. First, the turbulivity of air jet "c" has been updated to the value of 0.106 by experiments. It is necessary to use this factor for the calculation of air velocity where 2.5 m downstream from the centrifugal fan which defined as u2.5. Experimental and the CFD (Computational Fluid Dynamics) methods are applied to investigate the influence on the performance of the air ejector. Three parameters were characterized: converging angle of entraining chamber θ; diameter of the mixing chamber Dm and the nozzle position (NXP) S. Up to 240 different models were established and meshed by Gambit 2.3, and then simulated and calculated by Fluent 6.3 with the turbulence model of RNG k-epsilon. Consequently, 240 different results containing the flow rate of nozzle outlet cross-section mp, the entrainment flow rate me, the flow rate of the mixing chamber outlet cross-section mc, and the air velocity of a mixing chamber outlet cross-section uc were acquired. Based on this data, the response surfaces for u2.5 and entrainment ratio that are used to investigate the interaction between Dm, θ and S to u2.5 and entrainment ratio were established. The results indicated that the parameters Dm and θ have a great influence on u2.5 and entrainment ratio. It was also demonstrated that the interaction between Dm and θ is significant. However, parameter S gave a relatively delicate influence on u2.5 and entrainment ratio, and also the interaction with the other parameters was weak. In addition, to find out the mechanism of interaction between Dm and θ, three groups of different pressure fields have been investigated and compared as well. It indicates that a high value of θ (when θ < 36.25°) could create a relatively significant negative pressure zone in the mixing chamber, which requires a larger Dm to provide sufficient air input, and as a consequence u2.5 and entrainment ratio increase accordingly. Because of the negative pressure zone nearby the nozzle outlet, the effective power of the centrifugal fan and engine are both increased. The optimum value of θ is 36.25°, and when Dm equals to 144 mm, u2.5 could reach the maximum value. The maximum u2.5, which was gained by experiments using an ejecting pneumatic extinguisher, was 36.4 m/s. Meanwhile, the value of u2.5 could keep adding up as Dm continued to increase, but the range of Dm has been limited by the design and practical applicability of the portable pneumatic extinguisher. [ABSTRACT FROM AUTHOR]

Purpose: Ocular Albinism type 1 (OA1) is a disease caused by mutations in the OA1 gene and characterized by the presence of macromelanosomes in the retinal pigment epithelium (RPE) as well as abnormal crossing of the optic axons at the optic chiasm. We showed in our previous studies in mice that Oa1 activates specifically Gαi3 in its signaling pathway and thus, hypothesized that a constitutively active Gαi3 in the RPE of Oa1−/− mice might keep on the Oa1 signaling cascade and prevent the formation of macromelanosomes. To test this hypothesis, we have generated transgenic mice that carry the constitutively active Gαi3 (Q204L) protein in the RPE of Oa1−/− mice and are now reporting the effects that the transgene produced on the Oa1−/− RPE phenotype. Methods: Transgenic mice carrying RPE-specific expression of the constitutively active Gαi3 (Q204L) were generated by injecting fertilized eggs of Oa1−/− females with a lentivirus containing the Gαi3 (Q204L) cDNA. PCR, Southern blots, Western blots and confocal microscopy were used to confirm the presence of the transgene in the RPE of positive transgenic mice. Morphometrical analyses were performed using electron microscopy to compare the size and number of melanosomes per RPE area in putative Oa1−/−, Gαi3 (Q204L) transgenic mice with those of wild-type NCrl and Oa1−/− mice. Results: We found a correlation between the presence of the constitutively active Gαi3 (Q204L) transgene and the rescue of the normal phenotype of RPE melanosomes in Oa1−/−, Gαi3 (Q204L) mice. These mice have higher density of melanosomes per RPE area and a larger number of small melanosomes than Oa1−/− mice, and their RPE phenotype is similar to that of wild-type mice. Conclusions: Our results show that a constitutively active Gαi3 protein can by-pass the lack of Oa1 protein in Oa1−/− mice and consequently rescue the RPE melanosomal phenotype. [ABSTRACT FROM AUTHOR]

Background: Ocular albinism type 1, an X-linked disease characterized by the presence of enlarged melanosomes in the retinal pigment epithelium (RPE) and abnormal crossing of axons at the optic chiasm, is caused by mutations in the OA1 gene. The protein product of this gene is a G-protein-coupled receptor (GPCR) localized in RPE melanosomes. The Oa1-/- mouse model of ocular albinism reproduces the human disease. Oa1 has been shown to immunoprecipitate with the Gαi subunit of heterotrimeric G proteins from human skin melanocytes. However, the Gαi subfamily has three highly homologous members, Gαi1, Gαi2 and Gαi3 and it is possible that one or more of them partners with Oa1. We had previously shown by in-vivo studies that Gαi3-/- and Oa1-/- mice have similar RPE phenotype and decussation patterns. In this paper we analyze the specificity of the Oa1-Gαi interaction. Methodology: By using the genetic mouse models Gαi1-/-, Gαi2-/-, Gαi3-/- and the double knockout Gαi1-/-, Gαi3-/- that lack functional Gαi1, Gαi2, Gαi3, or both Gαi1 and Gαi3 proteins, respectively, we show that Gαi3 is critical for the maintenance of a normal melanosomal phenotype and that its absence is associated with changes in melanosomal size and density. GSTpull- down and immunoprecipitation assays conclusively demonstrate that Gαi3 is the only Gαi that binds to Oa1. Western blots show that Gαi3 expression is barely detectable in the Oa1-/- RPE, strongly supporting a previously unsuspected role for Gαi3 in melanosomal biogenesis. Conclusion: Our results identify the Oa1 transducer Gai3 as the first downstream component in the Oa1 signaling pathway. [ABSTRACT FROM AUTHOR]

Presents information on a study which examined the role of G[sub i2] alpha protein in the regulation of cardiac calcium channels. Methodology; Results of the study; Discussion.

Proposes that the transient receptor potential (trp) gene homologs are subunits of capacitative calcium entry (CCE) channels in mammals. Functional expression; Molecular diversity of the trp family; Inhibition of endogenous CCE by trp antisense sequence; Primary structure; Tissue expression; Model of typology.

Excessive glucocorticoids (GCs) production in adipose tissue promotes the development of visceral obesity and the metabolic syndrome. 11β-Hydroxysteroid dehydrogenase type 1 (11 β-HSD1) is critical for controlling intracellular GC production in adipose tissue, and this process is also regulated by hexose-6-phosphate dehydrogenase (H6PDH) that provides cofactor NADPH for 11β-HSD1 amplifying local GC production. To better understand the integrated molecular and physiological role of adipose H6PDH, we created a tissue specific knockout of H6PDH gene mouse model in adipocytes (H6PDHAcKO mice) and characterized the metabolic related phenotypes. H6PDHAcKO mice had decreased levels of intra-adipose corticosterone with reduction of 11β-HSD1 activity in response to H6PDH deletion in adipose tissue. Moreover, H6PDHAcKO mice also exhibited reduced fasting blood glucose levels and increased glucose tolerance with increased insulin sensitivity. In addition, plasma FFA levels were decreased with a concomitant decrease in the expression of lipase ATGL and HSL in the abdominal adipose tissue. These results indicate that inactivation of H6PDH expression in adipose tissue is sufficient to cause intra-adipose GC inactivation leading to the favorable pattern of metabolic phenotype. These data suggest that H6PDHAcKO mice may provide a good model for study the potential contributions of fat-specific H6PDH inhibition in the metabolic phenotype and other pathological disorders in vivo. Disclosure: J. Wang: None. X. Wang: None. K. Lutfy: None. T. Friedman: None. M. Jiang: None. Y. Liu: None. Funding: National Institute of Diabetes and Digestive and Kidney Diseases (SC1DK104821) [ABSTRACT FROM AUTHOR]

Summary: The placenta provides the interface for gas and nutrient exchange between the mother and the fetus. Despite its critical function in sustaining pregnancy, the stem/progenitor cell hierarchy and molecular mechanisms responsible for the development of the placental exchange interface are poorly understood. We identified an Epcamhi labyrinth trophoblast progenitor (LaTP) in mouse placenta that at a clonal level generates all labyrinth trophoblast subtypes, syncytiotrophoblasts I and II, and sinusoidal trophoblast giant cells. Moreover, we discovered that hepatocyte growth factor/c-Met signaling is required for sustaining proliferation of LaTP during midgestation. Loss of trophoblast c-Met also disrupted terminal differentiation and polarization of syncytiotrophoblasts, leading to intrauterine fetal growth restriction, fetal liver hypocellularity, and demise. Identification of this c-Met-dependent multipotent LaTP provides a landmark in the poorly defined placental stem/progenitor cell hierarchy and may help us understand pregnancy complications caused by a defective placental exchange. [ABSTRACT FROM AUTHOR]

Abstract: Cytomegalic adrenal hypoplasia congenita (AHC) is an X-linked disease caused by mutations in DAX1-encoding gene NR0B1, previously thought to function primarily in steroidogenesis. We sought to determine the expression pattern for Dax1 along with known network partners in early embryogenesis and to determine a steroidogenic capacity for the embryo prior to the establishment of the urogenital ridge at embryonic day 9 (E9). Here, we report that murine Dax1 is a unique marker in early embryonic development, distinguishing the extraembryonic (proximal) endoderm from the remainder of the developing embryo. We showed that Wilms tumor 1, steroidogenic factor 1, and estrogen receptor β were expressed throughout the embryo, but the progesterone, estrogen α and androgen receptors, cytochrome P450 (Cyp11a1) and Nur77 were not observed in any of the embryonic layers. Lack of Cyp11A1 expression at this stage confirmed an absence of inherent steroidogenic capacity for the early embryo. The role of Nr0b1 in embryonic stem (ES) cells was investigated using siRNA knockdown, resulting in differentiation toward endoderm-like fate. Nr0b1 conditional knockout in ES cells led to differentiation, confirming our knockdown results. Our investigations suggest that Nr0b1 functions in a novel role in the maintenance of a relatively undifferentiated state. Our results further suggest that the failure of conventional murine Nr0b1 knockout attempts may be due to disregulated differentiation. [Copyright &y& Elsevier]

Go, a heterotrimeric G-protein, is enriched in brain and neuronal growth cones. Although several reports suggest that Go may be involved in modulation of neuronal differentiation, the precise role of Go is not clear. To investigate the function of Go in neuronal differentiation, we determined the effect of Goα, the α subunit of Go, on the expression of Cav2.2, the pore-forming unit of N-type calcium channels, at the transcription level. Treatment with cyclic AMP (cAMP), which triggers neurite outgrowth in neuroblastoma F11 cells, increased the mRNA level and the promoter activity of the Cav2.2 gene. Overexpression of Goα inhibited neurite extension in F11 cells and simultaneously repressed the stimulatory effect of cAMP on the Cav2.2 gene expression to the basal level. Targeted mutation of the Goα gene also increased the level of Cav2.2 in the brain. These results suggest that Go may regulate neuronal differentiation through modulation of gene expression of target genes such as N-type calcium channels. [Copyright &y& Elsevier]