Human cytomegalovirus persists in myeloid progenitors and is passed to the myeloid progeny in a latent form

CD34+ progenitor cells can harbour latent human cytomegalovirus (HCMV); however, the mechanisms of HCMV latency remain unclear. We have investigated the effects of the haematopoietic lineage restriction on the establishment and spread of the latent HCMV to progeny cells. In vitro-infected and latently-infected haematopoietic progenitor cells derived from HCMV seropositive donors were studied. The presence of HCMV DNA in bone marrow progenitor (BMP) cells was determined by single colony polymerase chain reaction and fluorescent in situ hybridization (FISH). The presence of CMV DNA was found to be restricted to myeloid progenitors and the percentage of HCMV-infected cells was lower in naturally-infected cells than in in vitro-infected cells. Erythroid differentiation resulted in an abortive infection with persistence of the viral nucleic acids in red cell precursors. In BMP cells from HCMV seronegative donors, HCMV DNA was localized in the nucleus. Bone marrow progenitors in the presence of granulocyte-macrophage colony stimulating factor (GMCSF) maintained HCMV DNA for extended periods of time. No viral production could be detected throughout the culture but the comparison of the numbers of latently-infected cells prior to and after the culture suggests that proliferation of haematopoietic progenitor cells may lead to the expansion of latently-infected cells.     

Genetic and epidemiological relationships among Campylobacter isolates from humans

84 Campylobacter jejuni isolates from Swedish patients with domestic infection were characterized with pulsed-field gel electrophoresis (PFGE), and the subtype information considered in relation to epidemiological data. Based on pattern combinations from restriction cleavage with SmaI and SalI, 52 different PFGE types were identified. Types with an average pattern similarity of at least 82% and 63% were assembled in groups and clusters, respectively. The 2 largest clusters included 71% of the isolates. The distribution in time varied between different groups and clusters, where some were isolated sporadically during the whole period and others appeared more concentrated in time. Types in 1 cluster were significantly more often isolated in summer than other types in the study. Isolates from children showed lower pattern similarity to other isolates than isolates from adults. Sets of type and time related cases, possibly representing small outbreaks, were identified when indistinguishable PFGE patterns were found in isolates from temporally related cases. Our results indicate that although a large number of genotypes may be found among C. jejuni strains infecting humans, a large proportion of these may be genetically related, and that different genotypes may appear during different seasons and infect individuals of different ages.     

The Y-box binding protein YB-1 is associated with progressive disease and mediates survival and drug resistance in multiple myeloma

Current knowledge about molecular mechanisms underlying disease progression and drug resistance in multiple myeloma (MM) is still limited. Here, we analyzed the potential pathogenetic role of the Y-box binding protein YB-1 in MM. YB-1 is a member of the cold-shock domain protein superfamily and involved in various cellular functions such as proliferation. Immunohistochemical analyses revealed that neither normal bone marrow (BM) plasma cells (PCs), premalignant PCs of patients with monoclonal gammopathy of unknown significance (MGUS), nor MM cells with a mature morphology showed expression of YB-1 in situ. In contrast, YB-1 was strongly expressed in situ in normal PC precursor blasts as well as in a MM subset and in vitro in all of the evaluated MM cell lines. The YB-1-expressing MM cells were characterized by an immature morphology and a highly proliferative phenotype as defined by Ki 67 expression. We observed that siRNA-mediated knockdown of YB-1 decreased proliferation and induced apoptosis in MM cells even in the presence of BM stromal cells. Furthermore, we found that overexpression of YB-1 mediated resistance toward doxorubicin-induced apoptosis in MM cells. Thus, YB-1 contributes to disease progression, survival, and drug resistance in MM and might therefore provide an attractive therapeutic target.     

Ligand-induced inactivation of type I iodothyronine 5'-deiodinase: protection by propylthiouracil in vivo and reversibility in vitro

The inactivation of the phenolic ring deiodinases by natural substrates and iodinated contrast agents such as iopanoic acid (IOP) is a principal mechanism regulating the peripheral metabolism of thyroid hormones. The present studies were designed to investigate the cellular and biochemical mechanisms mediating this effect on type I 5'-deiodinase (5'DI) in rat liver and kidney. We reasoned that if inactivation requires the direct interaction of ligand (e.g. IOP) with 5'DI, then this effect should be blocked by a prior injection of 6n-propyl-2-thiouracil (PTU), which binds to an essential sulfhydryl group at the enzyme's active site and inhibits catalytic cycling. Experiments were performed using liver and kidney microsomes prepared from male rats injected 2.5 or 6 h earlier with various combinations of PTU, IOP, and/or vehicle. In initial studies, conditions were defined whereby active 5'DI could be regenerated in vitro from PTU-injected animals. This involved the treatment of microsomes with 50 mM dithiothreitol, pH 7.0, before assay. After this reductive treatment, 5'DI activity in both liver and kidney microsomes from rats previously injected with PTU, or the combination of PTU followed 30 min later by IOP, was equivalent to that in vehicle-injected control animals, whereas activity was decreased 60% in microsomes from animals injected with only IOP. This demonstrates that PTU protects 5'DI against ligand-induced inactivation, suggesting that this process is initiated by substrate binding to the enzyme's active site. In other experiments, liver microsomes from IOP-injected animals were pretreated with 100 mM dithiothreitol, pH 9.0, then washed, and 5'DI activity was determined. Under these stringent reducing conditions, 5'DI activity was completely recovered in microsomes from rats injected with IOP 2.5 h earlier, but only partially recovered 6 h after the IOP injection. Thus, ligand-induced inactivation of 5'DI may involve initially a reversible oxidation of the enzyme, followed by its irreversible inactivation.     

Infection of hematopoietic progenitor cells by human cytomegalovirus.

The susceptibility of hematopoietic progenitor cells to infection by human cytomegalovirus (HCMV) was investigated using several strains of HCMV, including the recombinant strain RC256. RC256 is derived from the laboratory strain Towne and contains the Escherichia coli LacZ gene coding for beta-galactosidase (beta-gal) regulated by an early HCMV promoter. Expression of LacZ allowed the detection of HCMV in individual hematopoietic cells. Clonogeneic bone marrow (BM) progenitors, including CD34+ cells, could be infected with HCMV and would then form normal hematopoietic colonies. By polymerase chain reaction (PCR) amplification of DNA, HCMV could be detected in both erythroid and myeloid colonies. LacZ activity was observed predominantly in cells of myelomonocytic lineage. When cells derived from HCMV-infected progenitors were cocultivated with permissive human fibroblasts, infectious virus expressing LacZ was recovered. Although no characteristic HCMV cytopathology was observed in BM colonies, high virus to cell ratios resulted in a moderate inhibition of colony formation. Since infected hematopoietic progenitors can harbor HCMV for weeks and through several differentiation steps in culture, we postulate that in vivo these cells may serve as a reservoir of latent virus and contribute to HCMV dissemination.     

Surgery for fulminating colitis during pregnancy

Two cases of fulminating colitis presenting during pregnancy are described. In both cases, resectional surgery was performed. In the first case, cesarean section was combined with subtotal colectomy and ileostomy during the 32nd week of gestation. In the second case, cesarean section was performed during the 33rd week of gestation and proctocolectomy in the puerperium. In both cases, histopathologic examination showed colitis more consistent with Crohn's disease. It is concluded that if fulminating colitis appears during pregnancy it should be treated in the same manner as in the nonpregnant state.     

Chronic pancreatitis and extrapancreatic cancer A retrospective study among 181 patients with chronic pancreatitis

Summary The relationship between chronic pancreatitis (CP) and extrapancreatic cancer has been debated in the recent years. In prospective studies, it has been found that pancreatic cancer develops in 0–5% of patients with chronic pancreatitis. Many papers describe an increased relative risk for developing extrapancreatic cancer in patients suffering from chronic pancreatitis. In this study including 181 patients with CP, we found 14 patients with extrapancreatic cancer (three of these had two different types of cancer). No patient had pancreatic cancer. It was found that the respiratory airways and upper gastrointestinal tract were the dominating locations (five and four cases, respectively), but also genital and hemolymphopoietic cancers were represented (four and two cases, respectively). Two patients had metastatic cancer with unknown primary tumor. The patients with cancer tended to be older than those without cancer. The patients with CP had a 2.43 times greater risk of developing cancer than the general Danish population (age and sex standardized comparison). The relatively large number of cancers in the upper gastrointestinal tract and respiratory airways suggest that tobacco and alcohol may be responsible, as these organs have the highest exposure to these compounds, which are well known carcinogens.     

Circulating adhesion molecules in allergic and non-allergic asthma

Circulating forms of adhesion molecules (intercellular-adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin ) are related to the turnover of these molecules on the cell surface. In contrast to the other molecules, the levels of E-selectin probably exclusively reflect the activity of endothelial cells. The aim of this study was to compare levels of circulating adhesion molecules in patients with allergic (AA) and non-allergic asthma (NA) and to relate the levels of soluble adhesion molecules to methacholine responsiveness and lung function. The study comprised 19 patients with AA, 15 patients with NA and 17 healthy subjects. Soluble adhesion molecules, spirometry, methacholine responsiveness and peak flow variability was measured. The group of patients with AA had higher levels of sE-selectin than the reference group (P=0.046). Serum levels of sE-selectin correlated significantly with bronchial responsiveness (r=0.76) and peak flow variability (r=0.75) (P<0.01) in the NA but not in the AA group. All adhesion molecules in AA (P<0.05-<0.001), but only sE-selectin in NA (P<0.05), were correlated to airway conductance. sVCAM-1 was reduced by inhaled steroids (P<0.01). Our results indicate that endothelial cells are activated in asthma and that this activity has a bearing on airflow variability and bronchial responsiveness in NA.     

Controlling systolic blood pressure is difficult in patients with diabetic kidney disease exhibiting moderate-to-severe reductions in renal function

This study compared the use of antihypertensive treatment and blood pressure (BP) controls between patients with diabetic kidney disease (DK+) and patients with non-diabetic kidney disease (DK-) exhibiting moderate-to-severe chronic renal failure who did not need renal replacement therapy. A cross-sectional survey included all renal patients with s-creatinine at ?200 micromol/l attending regular control sessions at six renal units in Norway. Of the 351 patients included, 73 (20.8%) were DK+. The proportion reaching a BP goal of <130/80 mmHg was similar in DK+ and DK- (14.1% vs 13.6%, p = 0.92), while 38% and 39% achieved a BP of <140/90 mmHg, respectively. The systolic BP goal was more difficult to achieve than the diastolic BP goal in DK+ patients (35% vs 15%) despite a mean of three different types of drugs being used. Loop diuretics and beta-adrenergic-receptor antagonists were the most frequently prescribed drugs, and the use of angiotensin-converting enzyme inhibitors or angiotensin-II-receptor antagonists declined when renal function deteriorated, from 80% to 0% and from 66% to 20% in the DK+ and DK- groups, respectively (p = 0.001). Thus, despite the use of multiple antihypertensive drugs, controlling BP - especially the systolic BP - is difficult in high-risk patients with chronic renal failure caused by diabetic kidney disease.     

Periscope Proteins are variable-length regulators of bacterial cell surface interactions

Changes at the cell surface enable bacteria to survive in dynamic environments, such as diverse niches of the human host. Here, we reveal “Periscope Proteins” as a widespread mechanism of bacterial surface alteration mediated through protein length variation. Tandem arrays of highly similar folded domains can form an elongated rod-like structure; thus, variation in the number of domains determines how far an N-terminal host ligand binding domain projects from the cell surface. Supported by newly available long-read genome sequencing data, we propose that this class could contain over 50 distinct proteins, including those implicated in host colonization and biofilm formation by human pathogens. In large multidomain proteins, sequence divergence between adjacent domains appears to reduce interdomain misfolding. Periscope Proteins break this “rule,” suggesting that their length variability plays an important role in regulating bacterial interactions with host surfaces, other bacteria, and the immune system.

Bacteria encounter complex and dynamic environments, including within human hosts, and have thus evolved various mechanisms that enable a rapid response for survival within, and exploitation of, new conditions. In addition to classical control by regulation of gene expression, bacteria exploit mechanisms that give rise to random variation to facilitate adaptation [e.g., phase and antigenic variation (1)]. In Gram-positive and Gram-negative human pathogens, DNA inversions (2, 3), homologous recombination (4), DNA methylation (1), and promoter sequence polymorphisms (5) govern changes in bacterial surface components, including capsular polysaccharide and protein adhesins, which can impact bacterial survival and virulence in the host (1, 6). Many of these mechanisms are very well studied and widespread across bacteria.A less well-studied mechanism is length variation in bacterial surface proteins. Variability in the number of sequence repeats in the Rib domain (7)–containing proteins on the surface of Group B streptococci has been linked to pathogenicity and immune evasion (8). The repetitive regions of the Staphylococcus aureus surface protein G (SasG) (9) and Staphylococcus epidermidis SasG homolog, Aap (10), also demonstrate sequence repeat number variability. In SasG, this variability regulates ligand binding by other bacterial proteins in vitro (11) in a process that has been proposed to enable bacterial dissemination in the host. Variations in repeat number have also been noted in the biofilm forming proteins Esp from Enterococcus faecalis (12) and, more recently, CdrA from Pseudomonas aeruiginosa (13). High DNA sequence identity in the genes that encode these proteins is likely to facilitate intragenic recombination events that would lead to repeat number variation (14) and, in turn, to protein sequence repetition. However, such sequence repetition is usually highly disfavored in large multidomain proteins (15), so its existence in these bacterial surface proteins suggests that protein length variation provides an evolutionary benefit. SasG, Aap, and Rib contain N-terminal host ligand binding domains and C-terminal wall attachment motifs; thus our recent demonstration that the repetitive regions of both SasG (16) and Rib (17) form unusual highly elongated rods suggests that host-colonization domains will be projected differing distances from the bacterial surface.Here, we show that repeat number variation in predicted bacterial surface proteins is more widespread and we characterize a third rod-like repetitive region in the Streptococcus gordonii protein (Sgo_0707) formed by tandem array of Streptococcal High Identity Repeats in Tandem (SHIRT) domains. Thus, we propose a growing class of “Periscope Proteins,” in which long, highly similar DNA repeats facilitate expression of surface protein stalks of variable length. This mechanism could enable changes in response to selection pressures and confer key advantages to the organism that include evasion of the host immune system (8) and regulation of surface interactions (11) involved in biofilm formation and host colonization.