Age-dependent response of murine female bone marrow cells to hyperbaric oxygen

Consequences of age on the effects of hyperbaric oxygen (HBO) on bone marrow (BM) derived stem cells and progenitors (SCPs) are largely unknown. We treated 2- and 18-month old C57BL/6 female mice by HBO. Hematopoietic stem cells and progenitors, enumerated as colony-forming units in culture, were doubled only in peripheral leukocytes and BM cells of young mice receiving HBO. In old mice colony-forming unit fibroblast numbers, a measure of mesenchymal stromal cells (MSCs) from BM, were high but unaffected by HBO. To further explore this finding, in BM-MSCs we quantified the transcripts of adipocyte early-differentiation genes peroxisome proliferator-activated receptor-γ, CCAAT/enhancer binding protein-β and fatty-acid binding protein 4; these transcripts were not affected by age or HBO. However, osteoblast gene transcripts runt-related transcription factor 2, osterix (OSX) and alkaline phosphatase (AP) were twofold to 20-fold more abundant in MSCs from old control mice relative to those of young control mice. HBO affected expression of osteoblast markers only in old MSCs (OSX gene expression was reduced by twofold and AP expression was increased threefold). Our data demonstrate the impact of aging on the response of BM SCPs to HBO and indicate the potentially different age-related benefit of HBO in wound healing and tissue remodeling

Structural Model of Porcine Factor Vil and Factor Villa Molecules Based on Scanning Transmission Electron Microscope (STEM) Images and STEM Mass Analysis

Abstract Porcine plasma factor VIII (fVIII) molecules are heterodimers composed of a 76,000-mol wt light chain (-A3-C1-C2) and a heavy chain ranging in molecular weight from 82,000 (Al-A2) to 166,000 (A,-A2-B). Proteolytic activation of fVIII by thrombin results in fVIlla heterotrimers lacking B domains (Al, A2, A3-Cl-C2). In this study, immunoaffinity purified fVIII was further fractionated by mono S or mono Q chromatography to prepare heterodimers containing a light chain and an Al-A2-B heavy chain (fVIII 166/76) or an Al-A2 heavy chain (fVIII 82/76). Mass analysis of scanning transmission electron microscopic (STE;M) images of fVIII 166/76 indicated that heterodimers (mass 237±20 kD) had irregularly globular core structures 10-12 nm across, and frequently displayed a diffuse, occasionally globular to ovoid satellite structure extending 5-14 nm from the core, and attached to it by a thin stalk. Factor VIII 82/76 molecules (mass 176±20 kD) had the same core structures as fVIII 166/76 molecules, but lacked the satellite structure. These findings indicate that Al-A2 domains of heavy chains and the light chains of the fVIII procofactor molecule are closely associated and constitute the globular core structure, whereas the B domainal portion of heavy chains comprises the peripheral satellite appendage. Factor VIII core structures commonly displayed a fingerlike projection near the origin of the B domainal stalk that was also a consistent feature of the free heavy chains (mass 128-162 kD) found in fVIII 166/76 preparations. Factor VIII light chain monomers (mass, 76±16 kD) were globular to cshaped particles 6-8 nm across. These chains commonly possessed a v-shaped projection originating from its middle region, that could also be observed at the periphery of fVIII core molecules. Factor VIIIa preparations contained heterotrimers (mass 162±13 kD) that had the same dimensions as fVIII core structures, lacked the B domainal appendage, and sometimes possessed the same core features as fVIII molecules. MolecuThis work was presented in part at the 12t

Prognostic value of discs large homolog 7 transcript levels in prostate cancer.

Hypoxia has been associated with malignant progression, metastasis and resistance to therapy. Hence, we studied expression of hypoxia-regulated genes in 100 prostate cancer (CaP) bulk tissues and 71 adjacent benign tissues. We found 24 transcripts significantly overexpressed (p ≤ 0.02). Importantly, higher transcript levels of disc large (drosophila) homolog-associated protein 5 (DLGAP5)/discs large homolog 7 (DLG7)/hepatoma up-regulated protein (HURP), hyaluronan-mediated motility receptor (HMMR) and cyclin B1 (CCNB1) were associated with higher Gleason score and more advanced systemic progression. Since the products of HMMR and CCNB1 have been identified recently as molecular markers of CaP progression, we postulated that DLG7 has prognostic value too. To test this hypothesis, we measured transcript levels for DLG7 in a 150-pair case-control cohort. The cases (progression to systemic disease within six years of surgery) and controls (no progression within eight years) were matched for clinical and pathologic prognostic variables, including grade, stage, and preoperative serum levels of PSA. The overall prognostic ability of DLG7, as tested in receiver operating characteristic analysis was of 0.74 (95% CI, 0.68 to 0.8). Overall, our data indicate that expression of DLG7, a hypoxia-controlled gene, holds prognostic potential in high-risk CaP; this also demonstrates that variation of oxygen tension may constitute a tool for identification of novel biomarkers for CaP

Transcript levels for CCNB1, DLG7, and HMMR measured in CaP and noncancerous prostate tissue.

<p>Panels on the left compare transcript levels in CaP bulk tissue (full symbols) with the levels measured in benign prostate tissue (open symbols) from men free of CaP (BP) and in benign prostate tissue (BPC) adjacent to CaP of combined Gleason score 6 (gs6). Panels on the right display transcript levels measured in non-neoplastic prostate epithelial cells isolated by laser capture microdissection (LCM) in benign tissues (open symbols): BP, benign prostatic hyperplasia (BPH) and BPC adjacent to CaP of the indicated Gleason score (gs). Full symbols in panels on the right denote transcript levels measured in LCM-isolated CaP cells: high-grade prostatic intraepithelial neoplasia (HGPIN), the cells isolated from areas of combined Gleason scores 6 through 8 and cells isolated from lymph node metastases (met). CCNB1, cyclin B1; DLG7, discs large homolog 7; HMMR, hyaluronan-mediated motility receptor.</p

Three hypoxia-controlled genes associated with Gleason score and prognosis.

<p>Among the hypoxia-regulated genes significantly overexpressed in CaP, cyclin B1 (CCNB1), DLGAP5 and hyaluronan-mediated motility receptor (HMMR) were associated with Gleason score and disease outcome.</p

Receiver operating characteristic (ROC) analysis for DLG7.

<p>The area under the curve was 0.74 (95% CI, 068–0.80). Inset: Scatter plot of the normalized expression values for cases (red) and controls (green).</p