[Esophageal disease: is cross sectional imaging contributive?]. / Pathologie de l'oesophage. L'imagerie en coupes a-t-elle un intérêt?

For a long time, esophagography and endoscopy were the major diagnostic tests used for evaluation of the esophagus. Now, the development of computed tomography, endosonography and MR imaging has permitted more comprehensive evaluation of esophageal diseases. Cross sectional imaging is essential to evaluate the relationship between esophageal lesions and adjacent mediastinal structures and to evaluate the thickness of the esophageal wall.

Transcriptional profiling of human osteoblast differentiation.

Osteoblast differentiation is a key aspect of bone formation and remodeling. To further our understanding of the differentiation process, we have developed a collection of conditionally immortalized adult human osteoblast cell lines representing discrete stages of differentiation. To evaluate changes in gene expression associated with differentiation, polyA((+)) RNA from pre-osteoblasts, early and late osteoblasts, and pre-osteocytes was subjected to gene chip analysis using the Affymetrix Hu6800 chip in conjunction with an Affymetrix custom chip enriched in bone and cartilage cDNAs. Overall, the expression of 47 genes was found to change threefold or more on both chips between the pre-osteoblastic and pre-osteocytic stages of differentiation. Many of the observed differences, including down-regulation of collagen type I and collagen-processing enzymes, reflect expected patterns and support the relevance of our results. Other changes have not been reported and offer new insight into the osteoblast differentiation process. Thus, we observed regulation of factors controlling cell cycle and proliferation, reflecting decreased proliferation, and increased apoptosis in pre-osteocytic cells. Elements maintaining the cytoskeleton, extracellular matrix, and cell-cell adhesion also changed with differentiation reflecting profound alterations in cell architecture associated with the differentiation process. We also saw dramatic down-regulation of several components of complement and other immune response factors that may be involved in recruitment and differentiation of osteoclasts. The decrease in this group of genes may provide a mechanism for controlling bone remodeling of newly formed bone. Our screen also identified several signaling proteins that may control osteoblast differentiation. These include an orphan nuclear receptor DAX1 and a small ras-related GTPase associated with diabetes, both of which increased with increasing differentiation, as well as a high mobility group-box transcription factor, SOX4, that was down-regulated during differentiation. In summary, our study provides a comprehensive transcriptional profile of human osteoblast differentiation and identifies several genes of potential importance in controlling differentiation of osteoblasts.

Hormonal control of insulin-like growth factor I gene transcription in human osteoblasts: dual actions of cAMP-dependent protein kinase on CCAAT/enhancer-binding protein delta.

Insulin-like growth factor-I (IGF-I) is essential for somatic growth and promotes bone cell replication and differentiation. IGF-I production by rat osteoblasts is stimulated by activation of cAMP-dependent protein kinase (PKA). In this report, we define two interacting PKA-regulated pathways that control IGF-I gene transcription in cultured human osteoblasts. Stimulation of cAMP led to a 12-fold increase in IGF-I mRNA and enhanced IGF-I promoter activity through a DNA response element termed HS3D and the transcription factor CCAAT/enhancer-binding protein delta (C/EBPdelta). Under basal conditions, C/EBPdelta was found in osteoblast nuclei but was transcriptionally silent. Treatment with the PKA inhibitor H-89 caused redistribution of C/EBPdelta to the cytoplasm. After hormone treatment, the catalytic subunit of PKA accumulated in osteoblast nuclei. Inhibition of active PKA with targeted nuclear expression of PKA inhibitor had no effect on the subcellular location of C/EBPdelta but prevented hormone-induced IGF-I gene activation, while cytoplasmic PKA inhibitor additionally caused the removal of C/EBPdelta from the nucleus. These results show that IGF-I gene expression is controlled in human osteoblasts by two PKA-dependent pathways. Cytoplasmic PKA mediates nuclear localization of C/EBPdelta under basal conditions, and nuclear PKA stimulates its transcriptional activity upon hormone treatment. Both mechanisms are indirect, since PKA failed to phosphorylate human C/EBPdelta in vitro.

Regulated nuclear-cytoplasmic localization of CCAAT/enhancer-binding protein delta in osteoblasts.

Insulin-like growth factor I (IGF-I) plays a central role in skeletal growth by promoting bone cell replication and differentiation. Prostaglandin E2 (PGE2) and parathyroid hormone enhance cAMP production in cultured rat osteoblasts and stimulate IGF-I expression through a transcriptional mechanism mediated by cAMP-dependent protein kinase (PKA). We previously showed that PGE2 activated the transcription factor CCAAT/enhancer-binding protein delta (C/EBPdelta) in osteoblasts and induced its binding to a DNA element within the IGF-I promoter. We report here that a PKA-dependent pathway stimulates nuclear translocation of C/EBPdelta. Under basal conditions, C/EBPdelta was cytoplasmic but rapidly accumulated in the nucleus after PGE2 treatment (t(1/2) < 30 min). Nuclear translocation occurred without concurrent protein synthesis and was maintained in the presence of hormone. Nuclear localization required PKA and was blocked by a dominant-interfering regulatory subunit of the enzyme, even though C/EBPdelta was not a PKA substrate. Upon removal of hormonal stimulus, C/EBPdelta quickly exited the nucleus (t(1/2) < 12 min) through a pathway blocked by leptomycin B. Mutagenesis studies indicated that the basic domain of C/EBPdelta was necessary for nuclear localization and that the leucine zipper region permitted full nuclear accumulation. We thus define a pathway for PKA-mediated activation of C/EBPdelta through its regulated nuclear import.

Stimulation of exocytosis without a calcium signal.

More than 30 years ago, Douglas (Douglas & Rubin, 1961; Douglas, 1968) proposed that intracellular Ca2+ controls stimulus-secretion coupling in endocrine cells, and Katz & Miledi (1967; Katz, 1969) proposed that intracellular Ca2+ ions control the rapid release of neurotransmitters from synapses. These related hypotheses have been amply confirmed in subsequent years and for students of excitable cells, they dominate our teaching and research. Calcium controls regulated exocytosis. On the other hand, many studies of epithelial and blood cell biology emphasize Ca2+-independent regulation of secretion of mucin, exocytotic delivery of transporters and degranulation. The evidence seems good. Are these contrasting conclusions somehow mistaken, or are the dominant factors controlling exocytosis actually different in different cell types? In this essay, we try to reconcile these ideas and consider classes of questions to ask and hypotheses to test in seeking a more integrated understanding of excitation-secretion coupling. Our review is conceptual and narrowly selective of a few examples rather than referring to a broader range of useful studies in the extensive literature. The examples are taken from mammals and are documented principally by citing other reviews and two of our own studies. The evidence shows that protein phosphorylation by kinases potentiates Ca2+-dependent exocytosis and often suffices to induce exocytosis by itself. Apparently, protein phosphorylation is the physiological trigger in a significant number of examples of regulated exocytosis. We conclude that although sharing many common properties, secretory processes in different cells are specialized and distinct from each other.

CCAAT/enhancer-binding protein delta is a critical regulator of insulin-like growth factor-I gene transcription in osteoblasts.

Insulin-like growth factor-I (IGF-I) plays a major role in promoting skeletal growth by stimulating bone cell replication and differentiation. Prostaglandin E2 and other agents that induce cAMP production enhance IGF-I gene transcription in cultured rat osteoblasts through a DNA element termed HS3D, located in the proximal part of the major rat IGF-I promoter. We previously determined that CCAAT/enhancer-binding protein delta (C/EBPdelta) is the key cAMP-stimulated regulator of IGF-I transcription in these cells and showed that it transactivates the rat IGF-I promoter through the HS3D site. We now have defined the physical-chemical properties and functional consequences of the interactions between C/EBPdelta and HS3D. C/EBPdelta, expressed in COS-7 cells or purified as a recombinant protein from Escherichia coli, bound to HS3D with an affinity at least equivalent to that of the albumin D-site, a known high affinity C/EBP binding sequence, and both DNA elements competed equally for C/EBPdelta. C/EBPdelta bound to HS3D as a dimer, with protein-DNA contact points located on guanine residues on both DNA strands within and just adjacent to the core C/EBP half-site, GCAAT, as determined by methylation interference footprinting. C/EBPdelta also formed protein-protein dimers in the absence of interactions with its DNA binding site, as indicated by results of glutaraldehyde cross-linking studies. As established by competition gel-mobility shift experiments, the conserved HS3D sequence from rat, human, and chicken also bound C/EBPdelta with similar affinity. We also found that prostaglandin E2-induced expression of reporter genes containing human IGF-I promoter 1 or four tandem copies of the human HS3D element fused to a minimal promoter and show that these effects were enhanced by a co-transfected C/EBPdelta expression plasmid. Taken together, our results provide evidence that C/EBPdelta is a critical activator of IGF-I gene transcription in osteoblasts and potentially in other cell types and species.

Effects of a cross-training exercise program in persons with osteoarthritis of the knee a randomized controlled trial.

This study was designed to evaluate, by means of a randomized controlled trial, the effects of a physical activity program incorporating aerobic, strength, and stretching exercises in individuals with osteoarthritis of the knee. We randomly assigned 137 volunteers ages >/=50 to an experimental group or a control group. The experimental group participated in three 1-hour sessions of supervised exercises per week over a 3-month period. The control participants were instructed to continue their usual daily activities, and they attended 1-hour education sessions twice a month. The effectiveness of the program was evaluated using repeated measurements of parameters related to self-reported health status, physical capacity, and joint tenderness.After 3 months, significantly greater improvements were observed in the experimental group than the control group in terms of: arthritis pain (p = 0.02), ability to walk and bend (p = 0.03), aerobic capacity (p < 0.0001), hamstring and low back flexibility (p = 0.003), quadriceps and hamstring strength (p <0.01), and the perception of changes relating to osteoarthritis of the knee and general condition (p < 0.0001). However, no significant differences were observed between the groups in isokinetic strength of the quadriceps (all p's >== 0.05), joint tenderness (p = 0.18), and health perception (p = 0.7). The overall results suggest that this program is effective for older persons with osteoarthritis of the knee and that it could contribute to maintaining their independence and improving their quality of life.

Computed tomography of superior mesenteric vein thrombosis following appendectomy.

During a 5-year period, superior mesenteric vein (SMV) thrombosis was detected with computed tomography (CT) in six patients shortly after an appendectomy. No sign of SMV was present at appendectomy, and a period of more than 2 weeks free of clinical symptoms had elapsed between the appendectomy and the onset of the SMV thrombosis. In four cases, the appendicitis was complicated. These patients had nonspecific signs and symptoms, although two of them had elevation of blood hepatic enzyme levels. In all cases, postcontrast CT demonstrated enlargement of the SMV, with well-defined enhancement of the vascular wall and an intraluminal clot. In one case, CT showed extension of the thrombus to the portal vein with the presence of low-attenuation areas in the liver, consistent with hepatic infarcts. Two patients had predisposing diseases: idiopathic hypersplenism in one case and chronic hepatic disease in the other. SMV thrombosis is a possible complication of appendicitis, and early appendectomy in appendicitis can prevent this complication. Moreover, as in any abdominal surgery, early appendectomy may be complicated by thrombosis of the SMV, thus creating problems of postoperative diagnosis. The complication is more frequent when the initial operation is performed under difficult conditions (peritonitis), or when the patient presents with a coagulopathy. CT is useful in the diagnosis of SMV thrombosis, thus leading to early management with anticoagulant therapy, with a view to avoiding complications such as intestinal ischemia, portal vein thrombosis, and hepatic infarction.

Protein kinase C as a signal for exocytosis.

We have studied signaling mechanisms that stimulate exocytosis and luteinizing hormone secretion in isolated male rat pituitary gonadotropes. As judged by reverse hemolytic plaque assays, phorbol-12-myristate-13-acetate (PMA) stimulates as many gonadotropes to secrete as does gonadotropin-releasing hormone (GnRH). However, PMA and GnRH use different signaling pathways. The secretagogue action of GnRH is not very sensitive to bisindolylmaleimide I, an inhibitor of protein kinase C, but is blocked by loading cells with a calcium chelator, 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. The secretagogue action of PMA is blocked by bisindolylmaleimide I and is not very sensitive to the intracellular calcium chelator. GnRH induces intracellular calcium elevations, whereas PMA does not. As judged by amperometric measurements of quantal catecholamine secretion from dopamine- or serotonin-loaded gonadotropes, the secretagogue action of PMA develops more slowly (in several minutes) than that of GnRH. We conclude that exocytosis of secretory vesicles can be stimulated independently either by calcium elevations or by activation of protein kinase C.

Functional heterogeneity of pituitary gonadotropes in response to a variety of neuromodulators.

Agents previously implicated in control of the hypothalamo-pituitary-gonadal axis were screened for their ability to regulate male rat gonadotropes directly. GnRH-evoked gonadotropin release is accompanied by oscillations of intracellular Ca2+ concentration ([Ca2+]i) and of an outward K+ current that is activated by Ca2+. Substances that caused current responses similar to those with GnRH were hypothesized to evoke secretion. Endothelin-1, oxytocin, neurotensin, pituitary adenylate cyclase-activating polypeptide, and serotonin raised [Ca2+]i and evoked LH release as assayed by the reverse hemolytic plaque assay. These agents affected only subpopulations of gonadotropes establishing functional heterogeneity of pituitary gonadotropes. One neuromodulator (ATP) evoked ionic current in all gonadotropes but the current was different than that evoked by GnRH. Many other substances, including galanin and neuropeptide Y, caused no changes in currents and were considered not to affect [Ca2+]i and not to be secretagogues for gonadotropes.