All 13 lipids, including two cardiolipins, one phosphatidylcholine, three phosphatidylethanolamines, four phosphatidylglycerols and three triglycerides, were identified in a crystalline bovine heart cytochrome c oxidase (CcO) preparation. The chain lengths and unsaturated bond positions of the fatty acid moieties determined by mass spectrometry suggest that each lipid head group identifies its specific binding site within CcOs. The X-ray structure demonstrates that the flexibility of the fatty acid tails facilitates their effective space-filling functions and that the four phospholipids stabilize the CcO dimer. Binding of dicyclohexylcarbodiimide to the O 2 transfer pathway of CcO causes two palmitate tails of phosphatidylglycerols to block the pathway, suggesting that the palmitates control the O 2 transfer process.The phosphatidylglycerol with vaccenate (cis-D 11 -octadecenoate) was found in CcOs of bovine and Paracoccus denitrificans, the ancestor of mitochondrion, indicating that the vaccenate is conserved in bovine CcO in spite of the abundance of oleate (cis-D 9 -octadecenoate). The X-ray structure indicates that the protein moiety selects cis-vaccenate near the O 2 transfer pathway against trans-vaccenate. These results suggest that vaccenate plays a critical role in the O 2 transfer mechanism.

No abstract

Bronchial asthma (BA) is a common chronic inflammatory disease characterized by hyperresponsive airways, excess mucus production, eosinophil activation, and the production of IgE. The complement system plays an immunoregulatory role at the interface of innate and acquired immunities. Recent studies have provided evidence that C3, C3a receptor, and C5 are linked to airway hyperresponsiveness. To determine whether genetic variations in the genes of the complement system affect susceptibility to BA, we screened single nucleotide polymorphisms (SNPs) in C3, C5, the C3a receptor gene (C3AR1), and the C5a receptor gene (C5R1) and performed association studies in the Japanese population. The results of this SNP case-control study suggested an association between 4896C/T in the C3 gene and atopic childhood BA (P = 0.0078) as well as adult BA (P = 0.010). When patient data were stratified according to elevated total IgE levels, 4896C/T was more closely associated with adult BA (P = 0.0016). A patient-only association study suggested that severity of childhood BA was associated with 1526G/A of the C3AR1 gene (P = 0.0057). We identified a high-risk haplotype of the C3 gene for childhood (P = 0.0021) and adult BA (P = 0.0058) and a low-risk haplotype for adult BA (P = 0.00011). We also identified a haplotype of the C5 gene that was protective against childhood BA (P = 1.4 x 10(-6)) and adult BA (P = 0.00063). These results suggest that the C3 and C5 pathways of the complement system play important roles in the pathogenesis of BA and that polymorphisms of these genes affect susceptibility to BA.

Human embryo implantation is a critical multistep process consisting of embryo apposition/adhesion, followed by penetration and invasion. Through embryo penetration, the endometrial epithelial cell barrier is disrupted and remodeled by an unknown mechanism. We have previously developed an in vitro model for human embryo implantation employing the human choriocarcinoma cell line JAR and the human endometrial adenocarcinoma cell line Ishikawa. Using this model we have shown that stimulation with ovarian steroid hormones (17␤-estradiol and progesterone, E2P4) and suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, enhances the attachment and adhesion of JAR spheroids to Ishikawa. In the present study we showed that the attachment and adhesion of JAR spheroids and treatment with E2P4 or SAHA individually induce the epithelial-mesenchymal transition (EMT) in Ishikawa cells. This was evident by up-regulation of N-cadherin and vimentin, a mesenchymal cell marker, and concomitant down-regulation of E-cadherin in Ishikawa cells. Stimulation with E2P4 or SAHA accelerated Ishikawa cell motility, increased JAR spheroid outgrowth, and enhanced the unique redistribution of N-cadherin, which was most prominent in proximity to the adhered spheroids. Moreover, an N-cadherin functional blocking antibody attenuated all events but not JAR spheroid adhesion. These results collectively provide evidence suggesting that E2P4-and implanting embryo-induced EMT of endometrial epithelial cells may play a pivotal role in the subsequent processes of human embryo implantation with functional control of N-cadherin.The first critical step in establishing a human pregnancy is implantation of an embryo onto the endometrial epithelial cells (EECs).2 In human, differentiation of the EECs is regulated across the menstrual cycle by changing levels of ovarian steroids. This ensures that an embryo-acceptable endometrial lining is present within 7-11 days after ovulation, the period termed the "implantation window" (1-3). Human embryonic implantation is a complex, multistep process, during which the embryo first comes into apposition with, and then later adheres onto the EEC layer. This is followed by embryo penetration through the EEC with invasion into the endometrial stromal cell (ESC) layer.Although several regulatory mediators of implantation have been identified in mice, the one or more mechanisms governing human implantation are largely still unknown. Ethical concerns and technical limitations pose significant obstacles to elucidating these processes in humans (4). Whereas mouse implantation models have enabled detailed molecular analyses, the human implantation site has remained inaccessible in vivo. To address the limitations of existing models, we adapted an in vitro implantation assay using human EECs and simulated model embryos (5). This model has been employed by several investigators studying early events in implantation (4 -7). For example, using this model, we investigated the effect of suberoylanilide hydroxamic aci...

BackgroundEndometrial stem/progenitor cells contribute to the cyclical regeneration of human endometrium throughout a woman's reproductive life. Although the candidate cell populations have been extensively studied, no consensus exists regarding which endometrial population represents the stem/progenitor cell fraction in terms of in vivo stem cell activity. We have previously reported that human endometrial side population cells (ESP), but not endometrial main population cells (EMP), exhibit stem cell-like properties, including in vivo reconstitution of endometrium-like tissues when xenotransplanted into immunodeficient mice. The reconstitution efficiency, however, was low presumably because ESP cells alone could not provide a sufficient microenvironment (niche) to support their stem cell activity. The objective of this study was to establish a novel in vivo endometrial stem cell assay employing cell tracking and tissue reconstitution systems and to examine the stem cell properties of ESP through use of this assay.Methodology/Principal FindingsESP and EMP cells isolated from whole endometrial cells were infected with lentivirus to express tandem Tomato (TdTom), a red fluorescent protein. They were mixed with unlabeled whole endometrial cells and then transplanted under the kidney capsule of ovariectomized immunodeficient mice. These mice were treated with estradiol and progesterone for eight weeks and nephrectomized. All of the grafts reconstituted endometrium-like tissues under the kidney capsules. Immunofluorescence revealed that TdTom-positive cells were significantly more abundant in the glandular, stromal, and endothelial cells of the reconstituted endometrium in mice transplanted with TdTom-labeled ESP cells than those with TdTom-labeled EMP cells.Conclusions/SignificanceWe have established a novel in vivo endometrial stem cell assay in which multi-potential differentiation can be identified through cell tracking during in vivo endometrial tissue reconstitution. Using this assay, we demonstrated that ESP cells differentiated into multiple endometrial lineages in the niche provided by whole endometrial cells, indicating that ESP cells are genuine endometrial stem/progenitor cells.