Hematopoietic development is a complex process that involves a large number of growth factors and cytokines. Many cytokines are known to act on more mature, lineage-restricted cells of the hematopoietic system. However, no specific factors have yet been identified that induce the expansion of the most primitive hematopoietic cells without also inducing differentiation. To search for such factors, we isolated novel cell lines from the yolk sac in order to identify genes important in early hematopoietic and endothelial development. This approach led to the discovery of B219, a sequence that is expressed in at least four isoforms in very primitive hematopoietic cell populations and which may represent a novel hemopoietin receptor. The recently published receptor for the obesity (ob) gene product (leptin) is an isoform of B219 with a nearly identical ligand binding domain. B219/obr is expressed in the yolk sac, early fetal liver, enriched hematopoietic stem cells and in a variety of lymphohematopoietic cell lines. B219/obr is also expressed at high levels in adult reproductive organs. B219/obr maps to human chromosome 1p32, a region syntenic with the recently reported location of obr on murine chromosome 4 (ref. 5).

No abstract

Rationale: Idiopathic pulmonary fibrosis (IPF) is a disease of progressive lung fibrosis with a high mortality rate. In organ repair and remodeling, epigenetic events are important. MicroRNAs (miRNAs) regulate gene expression post-transcriptionally and can target epigenetic molecules important in DNA methylation. The miR-17z92 miRNA cluster is critical for lung development and lung epithelial cell homeostasis and is predicted to target fibrotic genes and DNA methyltransferase (DNMT)-1 expression. Objectives: We investigated the miR-17z92 cluster expression and its role in regulating DNA methylation events in IPF lung tissue. Methods: Expression and DNA methylation patterns of miR-17z92 were determined in human IPF lung tissue and fibroblasts and fibrotic mouse lung tissue. The relationship between the miR-17z92 cluster and DNMT-1 expression was examined in vitro. Using a murine model of pulmonary fibrosis, we examined the therapeutic potential of the demethylating agent, 59-aza-29-deoxycytidine. Measurements and Main Results: Compared with control samples, miR17z92 expression was reduced in lung biopsies and lung fibroblasts from patients with IPF, whereas DNMT-1 expression and methylation of the miR-17z92 promoter was increased. Several miRNAs from the miR-17z92 cluster targeted DNMT-1 expression resulting in a negative feedback loop. Similarly, miR-17z92 expression was reduced in the lungs of bleomycin-treated mice. Treatment with 59-aza-29-deoxycytidine in a murine bleomycin-induced pulmonary fibrosis model reduced fibrotic gene and DNMT-1 expression, enhanced miR-17z92 cluster expression, and attenuated pulmonary fibrosis. Conclusions: This study provides insight into the pathobiology of IPF and identifies a novel epigenetic feedback loop between miR-17z92 and DNMT-1 in lung fibrosis.Keywords: microRNA; miR-17z92; pulmonary fibrosis; DNA methylation; DNMT-1 Idiopathic pulmonary fibrosis (IPF) represents the most aggressive form of interstitial lung disease with a median survival of 3-5 years (1). Failure to resolve epithelial cell injury in the lung is critical to the pathogenesis of IPF (2-4). In addition, epithelialmesenchymal transition (EMT) (5), fibroblast proliferation and activation (6), and recruitment of inflammatory cells (7,8) all contribute to extracellular matrix accumulation in the lung (7). The current study focused on identifying the molecular mechanisms underlying the pathogenesis of IPF.Because changes in fibrotic gene expression (2, 9-11) and few genetic mutations have been identified in IPF (12, 13), we focused on microRNA (miRNA, miR) expression and epigenetic regulators in lung epithelial cells and fibroblasts. MiRNAs can either block translation or degrade target mRNAs (14,15). Notably, a single miRNA can regulate upward of 30 genes. MiRNAs can be encoded in intronic or exonic DNA regions and encoded in their own open reading frame and controlled by DNA promoter elements, such as DNA methylation by DNA methyltransferases (DNMTs) of CpG islands (15,16). Of the three DNMTs expressed in h...

The effects of dorsal hippocampal lesions on retention of classical trace conditioned responses were examined using the rabbit nictitating membrane preparation. Animals were trained to criteria and then lesioned either in the cortex or in the hippocampus and the cortex. Hippocampal damage had no effect on the retention of responses but produced significantly longer onset latencies. A control group of hippocampal animals acquired conditioned responses (CRs) at least as quickly as the prelesion subjects, and they also exhibited longer response onset latency. A second experiment evaluated the performance of hippocampal lesioned animals in classical trace conditioning with either a low-intensity periorbital shock or corneal air puff as the unconditioned stimulus (UCS). Hippocampal animals successfully acquired CRs under both conditions but exhibited an alteration of response onset which was dependent on the form of the UCS. Hippocampal animals displayed shorter response onset in the air-puff condition and longer response onset in the shock condition. Cortical animals timed responses consistently regardless of the UCS. These findings strongly suggest that the hippocampus modulates temporal characteristics of learned behavior.

The effects of hippocampal ablation on acquisition rates and temporal characteristics of classically conditioned nictitating membrane responses were examined in groups of rabbits trained with a 150-, 300-, or 600-ms interstimulus interval. Acquisition rates were accelerated in the 150- and 600-ms groups. No effect was present in the 300-ms group. Response onset latencies were also affected in the 150-ms group. These findings were interpreted to support the notion that the hippocampus modulates learned motor behavior by a neural model of the response to be executed.

The p38 mitogen-activated protein kinases (MAPK) play a crucial role in stress and inflammatory responses and are also involved in activation of the human immunodeficiency virus gene expression. We have isolated the murine cDNA clones encoding p38-␦ MAPK, and we have localized the p38-␦ gene to mouse chromosome 17A3-B and human chromosome 6p21.3. By using Northern and in situ hybridization, we have examined the expression of p38-␦ in the mouse adult tissues and embryos. p38-␦ was expressed primarily in the lung, testis, kidney, and gut epithelium in the adult tissues. Although p38-␦ was expressed predominantly in the developing gut and the septum transversum in the mouse embryo at 9.5 days, its expression began to be expanded to many specific tissues in the 12.5-day embryo. At 15.5 days, p38-␦ was expressed virtually in most developing epithelia in embryos, suggesting that p38-␦ is a developmentally regulated MAPK. Interestingly, p38-␦ and p38-␣ were similar serine/threonine kinases but differed in substrate specificity. Overall, p38-␦ resembles p38-␥, whereas p38-␤ resembles p38-␣. Moreover, p38-␦ is activated by environmental stress, extracellular stimulants, and MAPK kinase-3, -4, -6, and -7, suggesting that p38-␦ is a unique stress-responsive protein kinase.

The receptor for the gene product of the obesity gene, leptin, was recently reported to be expressed on murine and human hematopoietic progenitor cells. Therefore, we studied the expression of the leptin receptor, OB-R, in normal myeloid precursors, human leukemia cell lines, and primary leukemic cells using reverse-transcriptase polymerase chain reaction. In normal hematopoiesis, OB-R was expressed in CD34+ cells. Normal promyelocytes (CD34−33+ and CD34−13+) expressed only very low levels of the short, presumably nonsignaling isoform. Both the long and short isoforms of OB-R were expressed in 10 of 22 samples from patients with newly diagnosed primary or secondary acute myeloid leukemia (AML), with a higher incidence of the long isoform in primary AML (87.6% v28.6%; P = .01). The incidence of OB-R expression was higher in recurrent than in newly diagnosed AML (P < .001), and samples from four patients with refractory AML showed strong expression of both isoforms. Both OB-R isoforms were also expressed in newly diagnosed and recurrent acute promyelocytic leukemia cells but were essentially absent in samples of chronic or acute lymphocytic leukemia. In vitro growth of myeloid leukemic cell lines and of blasts from 14 primary AMLs demonstrated that recombinant human leptin alone induced low level proliferation, significantly (P < .05) increased proliferation induced by recombinant human granulocyte colony-stimulating factor, interleukin 3, and stem cell factor in a subset of AML and increased colony formation (P < .005). Also, leptin reduced apoptosis induced by cytokine withdrawal in MO7E and TF-1 cells. Serum leptin levels correlated only with body mass index (P < .001) and gender (P = .03). Results confirm the reported expression of leptin receptor in normal CD34+ cells and demonstrate the frequent expression of leptin receptors in AML blasts. While normal promyelocytes lack receptor expression, leukemic promyelocytes express both isoforms. We also demonstrate proliferative effects of leptin alone and in combination with other physiologic cytokines, and anti-apoptotic properties of leptin. These findings could have implications for the pathophysiology of AML.

No abstract