Towards a Large-Scale Assessment of the Relationship between Biological and Chronological Aging: The INSPIRE Mouse Cohort.

Aging is the major risk factor for the development of chronic diseases. After decades of research focused on extending lifespan, current efforts seek primarily to promote healthy aging. Recent advances suggest that biological processes linked to aging are more reliable than chronological age to account for an individual's functional status, i.e. frail or robust. It is becoming increasingly apparent that biological aging may be detectable as a progressive loss of resilience much earlier than the appearance of clinical signs of frailty. In this context, the INSPIRE program was built to identify the mechanisms of accelerated aging and the early biological signs predicting frailty and pathological aging. To address this issue, we designed a cohort of outbred Swiss mice (1576 male and female mice) in which we will continuously monitor spontaneous and voluntary physical activity from 6 to 24 months of age under either normal or high fat/high sucrose diet. At different age points (6, 12, 18, 24 months), multiorgan functional phenotyping will be carried out to identify early signs of organ dysfunction and generate a large biological fluids/feces/organs biobank (100,000 samples). A comprehensive correlation between functional and biological phenotypes will be assessed to determine: 1) the early signs of biological aging and their relationship with chronological age; 2) the role of dietary and exercise interventions on accelerating or decelerating the rate of biological aging; and 3) novel targets for the promotion of healthy aging. All the functional and omics data, as well as the biobank generated in the framework of the INSPIRE cohort will be available to the aging scientific community. The present article describes the scientific background and the strategies employed for the design of the INSPIRE Mouse cohort.

Ectopic adenine nucleotide translocase activity controls extracellular ADP levels and regulates the F1-ATPase-mediated HDL endocytosis pathway on hepatocytes.

Ecto-F1-ATPase is a complex related to mitochondrial ATP synthase which has been identified as a plasma membrane receptor for apolipoprotein A-I (apoA-I), the major protein of high-density lipoprotein (HDL), and has been shown to contribute to HDL endocytosis in several cell types. On hepatocytes, apoA-I binding to ecto-F1-ATPase stimulates extracellular ATP hydrolysis into ADP, which subsequently activates a P2Y13-mediated HDL endocytosis pathway. Interestingly, other mitochondrial proteins have been found to be expressed at the plasma membrane of several cell types. Among these, adenine nucleotide translocase (ANT) is an ADP/ATP carrier but its role in controlling extracellular ADP levels and F1-ATPase-mediated HDL endocytosis has never been investigated. Here we confirmed the presence of ANT at the plasma membrane of human hepatocytes. We then showed that ecto-ANT activity increases or reduces extracellular ADP level, depending on the extracellular ADP/ATP ratio. Interestingly, ecto-ANT co-localized with ecto-F1-ATPase at the hepatocyte plasma membrane and pharmacological inhibition of ecto-ANT activity increased extracellular ADP level when ecto-F1-ATPase was activated by apoA-I. This increase in the bioavailability of extracellular ADP accordingly translated into an increase of HDL endocytosis on human hepatocytes. This study thus uncovered a new location and function of ANT for which activity at the cell surface of hepatocytes modulates the concentration of extracellular ADP and regulates HDL endocytosis.

Key role of PI3Kγ in monocyte chemotactic protein-1-mediated amplification of PDGF-induced aortic smooth muscle cell migration.

BACKGROUND AND PURPOSE: Vascular smooth muscle cell (SMC) migration within the arterial wall is a crucial event in atherogenesis and restenosis. Monocyte chemotactic protein-1/CC-chemokine receptor 2 (MCP-1/CCR2) signalling is involved in SMC migration processes but the molecular mechanisms have not been well characterized. We investigated the role of PI3Kγ in SMC migration induced by MCP-1. EXPERIMENTAL APPROACHES: A pharmacological PI3Kγ inhibitor, adenovirus encoding inactive forms of PI3Kγ and genetic deletion of PI3Kγ were used to investigate PI3Kγ functions in the MCP-1 and platelet-derived growth factor (PDGF) signalling pathway and migration process in primary aortic SMC. KEY RESULTS: The γ isoform of PI3K was shown to be the major signalling molecule mediating PKB phosphorylation in MCP-1-stimulated SMC. Using a PI3Kγ inhibitor and an adenovirus encoding a dominant negative form of PI3Kγ, we demonstrated that PI3Kγ is essential for SMC migration triggered by MCP-1. PDGF receptor stimulation induced MCP-1 mRNA and protein accumulation in SMCs. Blockade of the MCP-1/CCR2 pathway or pharmacological inhibition of PI3Kγ reduced PDGF-stimulated aortic SMC migration by 50%. Thus PDGF promotes an autocrine loop involving MCP-1/CCR2 signalling which is required for PDGF-mediated SMC migration. Furthermore, SMCs isolated from PI3Kγ-deficient mice (PI3Kγ(-/-)), or mice expressing an inactive PI3Kγ (PI3Kγ(KD/KD)), migrated less than control cells in response to MCP-1 and PDGF. CONCLUSIONS AND IMPLICATIONS: PI3Kγ is essential for MCP-1-stimulated aortic SMC migration and amplifies cell migration induced by PDGF by an autocrine/paracrine loop involving MCP-1 secretion and CCR2 activation. PI3Kγ is a promising target for the treatment of aortic fibroproliferative pathologies.

Cell surface adenylate kinase activity regulates the F(1)-ATPase/P2Y (13)-mediated HDL endocytosis pathway on human hepatocytes.

We have previously demonstrated on human hepatocytes that apolipoprotein A-I binding to an ecto-F(1)-ATPase stimulates the production of extracellular ADP that activates a P2Y(13)-mediated high-density lipoprotein (HDL) endocytosis pathway. Therefore, we investigated the mechanisms controlling the extracellular ATP/ADP level in hepatic cell lines and primary cultures to determine their impact on HDL endocytosis. Here we show that addition of ADP to the cell culture medium induced extracellular ATP production that was due to adenylate kinase [see text] and nucleoside diphosphokinase [see text] activities, but not to ATP synthase activity. We further observed that in vitro modulation of both ecto-NDPK and AK activities could regulate the ADP-dependent HDL endocytosis. But interestingly, only AK appeared to naturally participate in the pathway by consuming the ADP generated by the ecto-F(1)-ATPase. Thus controlling the extracellular ADP level is a potential target for reverse cholesterol transport regulation.

The nucleotide receptor P2Y13 is a key regulator of hepatic high-density lipoprotein (HDL) endocytosis.

Cell surface receptors for high-density lipoprotein (HDL) on hepatocytes are major partners in the regulation of cholesterol homeostasis. We recently identified a cell surface ATP synthase as a high-affinity receptor for HDL apolipoprotein A-I (apoA-I) on human hepatocytes. Stimulation of this ectopic ATP synthase by apoA-I triggered a low-affinity-receptor-dependent HDL endocytosis by a mechanism strictly related to the generation of ADP. This suggests that nucleotide G-protein-coupled receptors of the P2Y family are molecular components in this pathway. Only P2Y1 and P2Y13 are present on the membrane of hepatocytes. Using both a pharmacological approach and small interference RNA, we identified P2Y13 as the main partner in hepatic HDL endocytosis, in cultured cells as well as in situ in perfused mouse livers. We also found a new important action of the antithrombotic agent AR-C69931MX as a strong activator of P2Y13-mediated HDL endocytosis.

New insight on the molecular mechanisms of high-density lipoprotein cellular interactions.

High-density lipoprotein (HDL) cholesterol is an independent negative risk factor for coronary artery disease and thus represents today the only protective factor against atherosclerosis. The protective effect of HDL is mostly attributed to its central function in reverse cholesterol transport (RCT), a process whereby excess cell cholesterol is taken up and processed in HDL particles, and is later delivered to the liver for further metabolism and bile excretion. This process relies on specific interactions between HDL particles and cells, both peripheral (cholesterol efflux) and hepatic (cholesterol disposal) cells, and on the maturation of HDL particles within the vascular compartment. The plasma level of HDL cholesterol will thus result also from the complex interplay with cellular partners. Among them, some contribute to HDL formation - for instance ATP binding cassette AI protein - while others are mostly involved in HDL catabolism, the scavenger receptor-class B type I or the recently described membrane-bound ATP synthase/hydrolase. The last decade has seen major breakthroughs in the identification and elucidation of the role of cellular partners of HDL metabolism, and in their transcriptional regulations, opening up new perspectives in the modulation of HDL cholesterol.

Characterization of two high-density lipoprotein binding sites on porcine hepatocyte plasma membranes: contribution of scavenger receptor class B type I (SR-BI) to the low-affinity component.

Two HDL(3) high- and low-affinity binding sites are present on the human hepatoma cell line (HepG(2)). Recently, we have suggested that the high-affinity binding sites might modulate the endocytosis of HDL through the low-affinity binding sites [Guendouzi, K. (1998) Biochemistry 37, 14974-14980], highlighting the physiological importance of this family of HDL high-affinity binding sites. The present data demonstrate the presence of HDL(3) high-affinity (K(d) = 0.37 microg/mL, B(max) = 260 ng/mg of protein) and low-affinity (K(d) = 86.2 microg/mL, B(max) = 14 300 ng/mg of protein) binding sites on purified porcine hepatocyte plasma membranes. By contrast, free apoA-I was strictly specific to the high-affinity sites (K(d) = 0.2 microg/mL and B(max) = 72 ng/mg of protein). Competition experiments between (125)I-labeled HDL(3) and either LDL, oxidized LDL, or anti-SR-BI IgG as competitors show that SR-BI is mostly responsible (70% displacement) for the binding of HDL(3) to the low-affinity binding sites. By contrast, the same competition experiments using (125)I-labeled free apoA-I clearly excluded SR-BI as the high-affinity binding receptor. We conclude that the binding of HDL onto hepatocyte plasma membranes involves: (1) two low-affinity binding receptors, one being SR-BI; (2) one family of high-affinity binding sites unrelated to SR-BI.

World Health Organization--basic radiographic system (WHO-BRS) unit: experimental clinical evaluation.

The basic radiographic system (BRS) is a simplified version of a standard radiographic unit designed by the World Health Organization and the Pan American Health Organization. The design of the BRS includes the requirement that it should be able to perform routine radiographic examinations with a minimum of operator training. This report describes the performance of a BRS manufactured by Siemens. The physical parameters of this BRS were very good and operation should require a minimum of training. A small number of BRS films were compared with films obtained using the standard equipment in the department. The BRS films were generally regarded as satisfactory. Radiation exposure to patients is equal to, and in some cases less than, exposures with conventional X-ray units.

Empty right iliac fossa.

In the diagnosis of intra-abdominal disease, absence of gas or fecal shadows from the right iliac fossa is an important but nonspecific finding on plain roentgenograms. We found the empty right iliac fossa most commonly associated with acute appendicitis, Crohn's disease, and carcinoma of the cecum. Retrospective analysis of 137 confirmed cases of disease in the right lower quadrant yielded 21 (15.4%) with an empty right iliac fossa. In an additional 21 cases, an empty right iliac fossa was prospectively identified, and 13 of these patients (61.9%) had confirmed disease in a particular area.

Charcoal-containing empyema complicating treatment for overdose.

A 25-year-old man was treated with gastric lavage and activated charcoal for alcohol and drug overdose; immediately following the initial treatment he developed tension pneumothorax successfully managed with a chest tube. The patient presented four weeks later with a charcoal-containing empyema, an unreported condition heretofore.

Thickening of the posterior wall of the bronchus intermedius in right lung pneumonia.

A recent paper in the literature described thickening of the posterior wall of the bronchus intermedius (PWBI) in patients with congestive heart failure, and/or enlarged lymph nodes and neoplastic infiltrations. At this institution we have seen two recent cases of right upper lobe viral pneumonia demonstrating abnormal thickening of the posterior wall of the bronchus intermedius on lateral chest radiographs. A retrospective survey of our museum cases supported the hypothesis that pneumonia of the right lung, involving the right upper, middle, lower or any combination of these lobes may show thickening of the posterior wall of the bronchus intermedius.

Diagnostic considerations in urinary bladder wall calcification.

Though a relatively uncommon finding in general radiologic practice, urinary bladder wall calcification has relatively few etiologies. A series of 19 patients with radiographically visible bladder wall calcification encompassing most of the known causes is presented and other reported causes are discussed. Eight patients had schistosomiasis, six had primary carcinoma of the bladder, two had encrustation cystitis, and one each had amyloidosis, cyclophosphamide-induced cystitis, and tuberculosis. While a correct diagnosis is often not possible solely on the basis of the appearance of the calcification, such a diagnosis can usually be obtained expediently from a combination of history, clinical examination, appropriate laboratory studies, and radiographic evaluation of the bladder calcification and remaining urinary tract. Cystoscopy with biopsy of involved tissues is almost necessary, however, for confirmation and to rule out bladder neoplasia.

Malignant fistulae of the gastrointestinal tract.

Internal malignant fistulae of the gastrointestinal tract involving two or more loops of different segments of the bowel are rare. The colon is frequently one of the participating loops. Observations on four such cases indicate that the fistulous tract connecting the two bowel loops is usually linear, somewhat wide, and does not produce significant approximation of the bowel loops involved. These radiographic features help to differentiate such fistulae from the benign type.

Cholangiographic findings in agenesis of the gallbladder.

A case of agenesis of the gallbladder and cystic duct and the cholangiographic findings are presented. We believe this to be the first case documented by intravenous cholangiography.