The counterion, a negatively charged amino acid residue that stabilizes a positive charge on the retinylidene chromophore, is essential for rhodopsin to receive visible light. The counterion in vertebrate rhodopsins, Glu113 in the third transmembrane helix, has an additional role as an intramolecular switch to activate G protein efficiently. Here we show on the basis of mutational analyses that Glu181 in the second extracellular loop acts as the counterion in invertebrate rhodopsins. Like invertebrate rhodopsins, UV-absorbing parapinopsin has a Glu181 counterion in its G protein-activating state. Its G protein activation efficiency is similar to that of the invertebrate rhodopsins, but significantly lower than that of bovine rhodopsin, with which it shares greater sequence identity. Thus an ancestral vertebrate rhodopsin probably acquired the Glu113 counterion, followed by structural optimization for efficient G protein activation during molecular evolution.

Sufficient cytotrophoblast (CT) invasion into the uterine wall and subsequent remodeling of maternal uterine vasculature is critical to establish uteroplacental circulation. The production of vascular endothelial growth factor (VEGF) family molecules is confirmed in placental cells including CTs, but it is not elucidated how the VEGF system in CTs is controlled by oxygen tension and how it is involved in the development of placental circulation. To address this, we explored the effect of oxygen tension on the expression of VEGF, placenta growth factor (PlGF), and their antagonist, soluble fms-like tyrosine kinase-1 (sFlt-1) using ELISA and real-time PCR in a primary CT cell culture. For comparison, the same was conducted in parallel using other cells comprising placenta, such as human umbilical vein endothelial cells (HUVECs) and villous fibroblasts (VFs). Reduced oxygen resulted in a pronounced increase in sFlt-1 mRNA amount and sFlt-1 release into the culture media in CTs, whereas this was not the case with HUVECs and VFs. Free (not bound to sFlt-1) VEGF was not detected in CT culture media regardless of oxygen concentration, even though VEGF expression was stimulated by reduced oxygen in CTs, which was similar to the stimulation in HUVECs and VFs. Free PlGF was also diminished in CT culture media by reduced oxygen. These results implicate that CTs possess a unique property to enhance sFlt-1 production under reduced oxygen, which could consequently antagonize angiogenic activity of VEGF and PlGF. The presented findings might provide a framework with which to understand the mechanism of uterine vascular remodeling and its perturbations as exemplified in preeclampsia.

Opn5 (neuropsin) belongs to an independent group separated from the other six groups in the phylogenetic tree of opsins, for which little information of absorption characteristics and molecular properties of the members is available. Here we show that the chicken Opn5 (cOpn5m) is a UV-sensitive bistable pigment that couples with Gi subtype of G protein. The recombinant expression of cOpn5m in HEK 293s cells followed by the addition of 11-cis-and all-trans-retinal produced UV light-absorbing and visible lightabsorbing forms, respectively. These forms were interconvertible by UV and visible light irradiations, respectively, indicating that cOpn5m is a bistable pigment. The absorption maxima of these forms were estimated to be 360 and 474 nm, respectively. The GTPγS binding assay clearly showed that the visible light-absorbing form having all-trans-retinal activates Gi type of G protein, whereas no Gt or Gq activation ability was observed. Immunohistochemical studies using an antibody against cOpn5m clearly showed that this pigment is localized within some types of amacrine cells and some cells in the ganglion cell layer of the retinas, the vast majority of cells in the pineal gland and serotonin-positive cells in the paraventricular organ. Because cOpn5m is the only UV-sensitive opsin among the opsins found so far in chicken, this study provides the molecular basis for UV reception in chicken.G protein-coupled receptor | nonvisual photoreception | phototransduction

Purpose:To investigate the utility of apparent diffusion coefficient (ADC) values for discriminating tumor in patients with prostate cancer from normal prostatic tissues in healthy adult men, and to identify correlations between ADC and histologic grade of prostate cancer. Materials and Methods:A total of 125 healthy male volunteers (mean age, 60 years; range, 50 -86 years) and 90 prostate cancer patients (mean age, 71 years; range, 51-88 years) underwent diffusion-weighted imaging (DWI) of the prostate with a single-shot echo-planar imaging sequence using b-factors of 0 and 800 sec/mm 2 . ADC was measured from two locations in the peripheral zone (PZ) and two locations in the central gland (CG) in normal subjects, and tumor locations of PZ or transition zone (TZ) in patients with prostate cancer. Results:Mean ADC values of tumor regions in both PZ (1.02 Ϯ 0.25 ϫ 10 Ϫ3 mm 2 /sec) and TZ (0.94 Ϯ 0.21 ϫ 10 Ϫ3 mm 2 /sec) were significantly lower than those in the corresponding normal regions (1.80 Ϯ 0.27 ϫ 10 Ϫ3 mm 2 /sec and 1.34 Ϯ 0.14 ϫ 10 Ϫ3 mm 2 /sec, respectively) (P Ͻ 0.0001 each). Furthermore, a significant negative correlation was identified between ADC in PZ cancer and tumor Gleason score ( ϭ Ϫ0.497, P Ͻ 0.0001). Conclusion:ADC values appear to provide acceptable diagnostic accuracy in both PZ and TZ.

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Interpretation of combined T2-weighted, dynamic contrast-enhanced, and DW MR image findings can yield reasonable diagnostic accuracy in both the PZ (80% [191 of 240 regions]) and the TZ (74% [59 of 80 regions]).

A non‐visual pigment melanopsin, which is localized in photosensitive retinal ganglion cells and is involved in the circadian photoentrainment and pupillary responses in mammals, is phylogenetically close to the visual pigments of invertebrates, such as insects and cephalopods. Recent studies suggested that melanopsin is a bistable pigment and drives a Gq‐mediated signal transduction cascade, like the invertebrate visual pigments. Because detailed electrophysiological properties are somewhat different between the visual cells and the photosensitive ganglion cells, we here expressed and purified the invertebrate visual pigment and melanopsin to comparatively investigate their Gq‐activation abilities. We successfully expressed and purified UV and blue light‐sensitive visual pigments of the honeybee as well as the amphioxus melanopsin. Although the purified UV‐sensitive pigment and the melanopsin lost their bistable nature during purification, reconstitution of the pigments in lipid vesicles resulted in return of the bistable nature. The light‐dependent Gq‐activation abilities among these reconstituted pigments are similar, suggesting that the electrophysiological differences do not depend on the Gq‐activation step but rather on the other signal transduction steps and/or on cell properties. Our findings are also important in that this is the first report describes a heterologous large‐scale expression of the Gq‐coupled invertebrate visual pigments in cultured cells.

Exquisitely regulated cytokine balance during early pregnancy is thought to be necessary for promoting survival of the fetal allograft. Our previous studies have demonstrated that membrane-bound human leukocyte antigen (mHLA-G) expressed on trophoblasts is one of the key factors in regulating cytokine balance by shifting the Th1/Th2 balance toward Th2 polarization, a favourable milieu for the maintenance of pregnancy. Given that trophoblasts secrete soluble HLA-G (sHLA-G), we examined its biological roles in comparison with mHLA-G. We cultured peripheral blood mononuclear cells (PBMC) with either the HLA-A and -B-deficient B lymphoblast cell line (721.221 cells) or the same cell line transfected with mHLA-G (721.221-G1 cells), in the presence or absence of recombinant sHLA-G. Cytokine concentrations in the culture media were determined by enzyme-linked immunosorbent assay. In contrast to mHLA-G protein, sHLA-G stimulated the release of tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma, whereas it reduced the release of interleukin (IL)-3, regardless of the presence of the presence of a stimulatory effect of the mHLA-G-expressing cells. Although mHLA-G reduced the release of IL-4, sHLA-G did not have any effect. Conversely, sHLA-G stimulated the release of IL-10 whereas mHLA-G was without effect. These results suggest that sHLA-G regulates the release of cytokines from PBMC chiefly by counterbalancing mHLA-G, and thereby may play a role in maintaining pregnancy.