Gene microarray analysis of peripheral blood cells in pulmonary arterial hypertension

The importance of genetic predisposition, inflammation, and autoimmune mechanisms in the development of pulmonary arterial hypertension (PAH) is becoming increasingly clear. We hypothesized that the analysis of gene expression profiles from peripheral blood mononuclear cells would distinguish patients with PAH from normal volunteers. We also hypothesized that a subset of genes would discriminate between patients with idiopathic PAH and pulmonary hypertension related to secondary causes. Mononuclear cells were isolated from 15 patients diagnosed with PAH and 6 normal control subjects. Microarray expression was performed, and the expression profiles were analyzed for consistent and predictive differences in gene expression. We identified a signature set of 106 genes that discriminated with high certainty (p < or = 0.002) between patients with PAH and normal individuals. The results of the microarray analysis were retrospectively and prospectively confirmed by quantitative polymerase chain reaction for 2 of the 106 genes. Supervised clustering analysis generated a list of differentially expressed genes between patients with idiopathic and secondary causes of pulmonary hypertension. Microarray expression profiling of peripheral blood cells can discriminate between patients with PAH and normal volunteers. These findings may have important implications toward diagnosis, screening, and pathogenesis of this disease.     

A heteroplasmic point mutation of mitochondrial tRNALeu(CUN) in non-lymphoid haemopoietic cell lineages from a patient with acquired idiopathic sideroblastic anaemia

Acquired idiopathic sideroblastic anaemia (AISA) has been proposed to be a disorder of mitochondrial DNA (mtDNA). The hallmark of mitochondrial iron overload may be attributable to a respiratory chain defect leading to impaired reduction of ferric iron (Fe^3 +) to ferrous iron (Fe^2 +), which is essential to the last step of mitochondrial haem biosynthesis. In a 71-year-old patient we identified a point mutation in one of the two mitochondrial transfer-RNAs coding for leucine (tRNA^leu(CUN)). The mutation involves a G → A transition in the anticodon loop, immediately adjacent to the anticodon triplet (mtDNA position 12301). The mutated guanine is highly conserved in a wide range of species. The mutation is heteroplasmic, i.e. there is a mixture of normal and mutated mitochondrial genomes (ratio c. 50:50). Heteroplasmy of mtDNA is not found in normal individuals, but is a typical feature of mitochondrial cytopathies. The point mutation was present in the patient's bone marrow and whole blood samples, in purified platelets, and in the granulocyte/erythrocyte pellet after mononuclear cell separation by density gradient centrifugation. The mutation was not found in T- and B-lymphocytes isolated by immunomagnetic bead separation. It was also absent from buccal mucosa cells and cultured skin fibroblasts. This pattern of involvement suggests that the mutation occurred in a self-renewing myeloid stem cell of the CFU-GEMM type.     

Pacemaker-related myocardial perfusion defects worsen during higher pacing rate and coronary flow augmentation.

BACKGROUND: Asynchronous activation resulting from right ventricular apical (RVA) pacing can adversely affect left ventricular function and myocardial perfusion despite normal coronary arteries. This situation makes detection of coronary heart disease in paced patients difficult. OBJECTIVES: The purpose of this study was to assess the distribution, extent, and severity of myocardial perfusion defects with RVA pacing at low and high rates and increased coronary blood flow with adenosine. METHODS: Fourteen patients with permanent RVA pacing and angiographically normal coronary arteries underwent myocardial perfusion single-photon emission computed tomography at rest at low and high pacing rates and with pacing at low rates with adenosine. Data were analyzed semi-quantitatively using a 20-segment scoring model and coded using a four-point scoring system. RESULTS: At rest, 23 (55%) of 42 coronary flow territories showed abnormal perfusion and 52 (19%) of 280 corresponding segments demonstrated abnormal perfusion; mean perfusion score was 0.22. After high-rate pacing, perfusion was abnormal in 31 (74%) of 42 flow territories and 122 (44%) of 280 segments; mean perfusion score was 0.67. Adenosine infusion resulted in 28 (67%) of 42 abnormal flow territories and 90 (32%) of 280 abnormal segments; mean perfusion score was 0.44. Perfusion defects were observed most often in close proximity to the origin of the pacing site. CONCLUSION: RVA pacing results in myocardial perfusion defects. The false-positive findings are present at rest and more obvious with high-rate pacing than during adenosine infusion. Detection of coronary artery disease should be performed with caution in RVA paced patients because of the high number of perfusion defects observed in the absence of coronary artery disease.     

16, 16-Dimethyl PGE2 fails to prevent ethanol-induced drop of gastric potential difference in man

Digestive Diseases and Sciences -     

Gene expression pattern in biomechanically stretched cardiomyocytes: evidence for a stretch-specific gene program

Biomechanical stress ie, attributable to pressure overload, leads to cardiac hypertrophy and may ultimately cause heart failure. Yet, it is still unclear how mechanical stress is sensed and transduced on the molecular level. To systematically elucidate the underlying signal transduction pathways, we analyzed the gene expression profile of stretched cardiomyocytes on a genome-wide scale in comparison with other inducers of hypertrophy such as pharmacological stimulation. Neonatal rat ventricular cardiomyocytes were either stretched biaxially or stimulated with phenylephrine (PE), both resulting in a similar degree of hypertrophy. Microarray analyses revealed 164 genes >2.0-fold up- and 21 genes <0.5-fold downregulated (P<0.01). Differential expression was confirmed by real-time polymerase chain reaction. Genes of the "fetal gene program" such as BNP were induced by both stretch (4.2x) and PE (2.9x). We also verified upregulation of known stretch-responsive genes, including HSP70 (20.9x) and c-myc (3.0x). Moreover, several genes were found to be preferentially induced by stretch, such as the cardioprotective cytokine GDF15 (24.8x) and heme oxygenase 1 (Hmox1, 10.8x; both confirmed on protein level). Neither PE nor endothelin-1 upregulated GDF15 and Hmox1, whereas angiotensin II significantly induced both genes. Conversely, the AT(1) receptor blocker irbesartan markedly blunted stretch-mediated GDF15 and Hmox1 upregulation, suggesting that the angiotensin receptor transduces the biomechanical induction of these genes. In conclusion, we report a comprehensive gene expression profile of cardiomyocytes subjected to biomechanical stress in comparison with pharmacologically induced hypertrophy. Our data imply that a stretch-specific gene program exists, which is mediated, at least in part, by angiotensin II-dependent signaling.     

IL-10 Synergizes with IL-4 and IL-13 in Inhibiting Lysosomal Enzyme Secretion by Human Monocytes and Lamina Propria Mononuclear Cells from Patients with Inflammatory Bowel Disease

Tissue injury and inflammation in inflammatorybowel disease (IBD) are associated with enhancedmonocytic lysosomal enzyme release. In this study,peripheral monocytes and lamina propria mononuclearcells (LPMNC) were isolated from IBD patients andnormal controls. Cells were stimulated withlipopolysaccharide after treatment with IL-13, IL-4, andIL-10, and enzyme secretion was assessed by using thecorresponding p-nitrophenyl glycosides as substrates.Molecular forms of cathepsin D were examined to describethe mode of enzyme release. IL-10 and IL-4 stronglydown-regulate enzyme secretion in IBD monocytes. IBD monocytes showed a diminished responsiveness tothe inhibitory effect of IL-13. Impaired monocyteresponse was not found with combinations of IL-13 andIL-10 or IL-4 and IL-10. LPMNC from involved IBD mucosa showed significantly higher enzyme secretioncompared with LPMNC from noninvolved IBD mucosa butresponded inefficiently to either IL-4, IL-13, or IL-10alone. However, combined treatment with IL-10 and IL-4 or IL-10 and IL-13 strongly suppressedenzyme release by these cells. Both the precursor andmature forms of cathepsin D were elevated in IBDpatients. While IL-13 reduced mainly the precursor form, the effect of IL-4 and IL-10 concerns both theprecursor and mature form of cathepsin D. Our resultsfavor the potent clinical utility of combined treatment,thus improving chances of developing effective treatments for human IBD.     

Endoscopic retrograde cholangiopancreatography, intraductal ultrasonography, and magnetic resonance cholangiopancreatography in bile duct strictures: a prospective comparison of imaging diagnostics with histopathological correlation

OBJECTIVES: A variety of imaging techniques are available to diagnose bile duct strictures; the most effective imaging technique, however, has not been established yet. In the present study, we compared the impact of endoscopic retrograde cholangiopancreatography (ERCP), intraductal ultrasonography (IDUS), and magnetic resonance cholangiopancreatography (MRCP) with regard to diagnosing bile duct strictures. METHODS: We prospectively examined 33 patients with jaundice due to bile duct strictures by ERCP plus IDUS and MRCP. The objectives were to assess diagnostic quality of imaging, complete presentation of the bile duct, and differentiation of malignant from benign lesions. Surgical and histopathological correlations, which were used as the gold standard, were available in all cases since all included patients underwent laparotomy. RESULTS: Diagnostic image quality for ERCP was 88% and 76% for MRCP (p > 0.05). Comparing ERCP and MRCP, complete presentation of the biliary tract was achieved in 94% and 82%, respectively (p > 0.05). ERCP and MRCP allowed correct differentiation of malignant from benign lesions in 76% and 58% (p= 0.057), respectively. By supplementing ERCP with IDUS, the accuracy of correct differentiation of malignant from benign lesions increased significantly to 88% (p= 0.0047). CONCLUSIONS: Comparing ERCP with MRCP, we found adequate presentation of bile duct strictures in high imaging quality for both techniques. ERCP supplemented by IDUS gives more reliable and precise information about differentiation of malignant and benign lesions than MRCP alone without additional imaging sequences.     

Sildenafil for long-term treatment of nonoperable chronic thromboembolic pulmonary hypertension

Only a small percentage of patients with chronic thromboembolic pulmonary hypertension are eligible for pulmonary thrombendarterectomy. We investigated the effects of oral sildenafil on hemodynamics and exercise capacity in 12 nonoperable chronic thromboembolic pulmonary hypertension patients. All patients were in disease progression despite sufficient long-term anticoagulation and the best supportive care and suffered from severe pulmonary hypertension (pulmonary vascular resistance index 1,935 +/- 228 dyn. s. cm-5. m2, cardiac index 2.0 l. min-1. m-2, 6-minute walking distance 312 +/- 30 m). After approximately 6 months of sildenafil treatment, pulmonary hemodynamics and exercise capacity improved significantly (pulmonary vascular resistance index 1,361 +/- 177 L. min-1. m2, p = 0.004, cardiac index 2.4 +/- 0.2 L. min-1. m-2, p = 0.009, 6-minute walking distance 366 +/- 28 m, p = 0.02). Therefore, oral sildenafil may offer a new option for medical treatment of this devastating disease.     

Transplantation of peripheral blood progenitor cells from HLA-identical sibling donors

Transplantation of peripheral blood progenitor cells (PBPCs) has largely replaced autologous bone marrow transplantation. The same might occur in the allogeneic setting if the favourable initial experience with allogeneic PBPCT is confirmed. We analysed all primary transplants utilizing unmodified PBPC from HLA-identical sibling donors reported to the European Group for Blood and Marrow Transplantation (EBMT) for 1994. 59 patients with a median age of 39 years received myeloablative therapy for acute myelogenous leukaemia (23 patients), acute lymphoblastic leukaemia (13), chronic myelogenous leukaemia (nine), lymphoma (seven), or other diagnoses (seven) mostly of advanced stages followed by transplantation of allogeneic PBPC. Three patients died soon after grafting, the others showed prompt haemopoietic recovery with median times to recover an absolute neutrophil count (ANC) above 0.5 and 1.0×10⁹/l of 15 (range 9–27) and 17 d (range 10–28) respectively. Time to platelet recovery above 20 or 50×10⁹/l was 16 (range 9–76) and 18 d (range 12–100) respectively. 27 patients (46%) developed no or mild acute graft-versus-host disease (GVHD). The incidence of moderate (grade II) disease was 27%; 24% of the patients developed severe acute GVHD (grades III or IV). 55% of patients who were alive 90 d after transplantation developed chronic GVHD, the probability to develop extensive chronic GVHD was 32% (95% confidence interval 22–42) with a median follow-up of 14 months. Overall and event-free survival (EFS) at 1 year were 54% (CI 48–60) and 50% (CI 43–57), respectively, the relapse incidence was 23% (CI 17–29). EFS was 67% (CI 55–79) in patients transplanted for acute leukaemias in first complete remission, chronic myelogenous leukaemia in first chronic phase, or severe aplastic anaemia. Transplantation of allogeneic PBPC resulted in prompt and durable engraftment. The incidence and severity of acute and chronic GVHD seemed comparable to that observed after allogeneic BMT. Overall and event-free survival in this cohort of patients, most of whom suffered from advanced leukaemia or lymphoma, is encouraging, suggesting that the high numbers of T lymphocytes and/or natural killer cells contained in a typical PBPC collection product exert a vigorous graft-versus-leukaemia effect. Further evaluation of allogeneic PBPCT is highly desirable.