bluuebunny/crossref_metadata_2025 huggingface dataset:
A non-glycosylated form of pulmonary surfactant protein A appears in rat amniotic fluid
Authors: Sakai K , Kohri T , Kweon MN , Kishino Y
Published: October 2003

Surfactant protein A (SP-A) is a family of glycoproteins that have a triplet with 26, 32 and 36 kDa under reducing conditions in rat lung. We wanted to evaluate the SP-A forms in amniotic fluid of pregnant rats compared to those found in rat lungs. By Western blot analysis, glycosylated SP-A, was not found in the amniotic fluid in contrast to the pulmonary surfactant triplet SP-A, which comprises a 26 kDa protein and its glycosylated 32 and 36 kDa forms. The SP-A concentration in amniotic fluid was barely detectable at 18 days of gestation (20 +/- 12 ng.ml-1), and then increased and reached 700 +/- 333 ng.ml-1 at the final gestational day 21, as determined by an enzyme-linked immunoabsorbent assay. Immunohistochemically, SP-A was found in some epithelial cells of larger respiratory bronchi, but not, or to a lesser degree, in smaller respiratory bronchi at gestational day 18. At 21 days of gestation, SP-A was detected in bronchial and bronchiolar nonciliated epithelial Clara cells, alveolar epithelial type II cells and some alveolar macrophages. The ratio of the 26, 32 and 36 kDa SP-A forms in bronchoalveolar, bronchobronchiolar and tracheal lavage fluids prepared from adult rats was 6:29:65, 84:5:11 and 100:0:0, respectively. These findings show the presence of a non-glycosylated SP-A in rat amniotic fluid. This may reflect the increased ratio of non-glycosylated SP-A to bronchoalveolar, bronchobronchiolar and tracheal lavage fluids, respectively.
Diagnostic value of direct examination of the protected specimen brush in ventilator-associated pneumonia
Authors: Marquette CH , Wallet F , Neviere R , Copin MC , Saulnier F , Drault JN , Mehdaoui H , Mathieu D , Ramon P
Published: October 2003

Interpretation of the protected specimen brush (PSB) technique is based on quantitative bacterial cultures (QC), which unfortunately requires at least 24 h. We prospectively compared the diagnostic value of direct examination (DE) and QC of PSB specimens in 75 patients with suspected pneumonia. We also determined the optimal technique for DE. QC was performed using the serial dilution technique. From the original suspension, two cytospin slides were obtained and stained by the May-Grunwald Giemsa (MGG) and the Gram method for DE. If the prescreening on the MGG-stained slide was positive, the morphology and the Gram staining of the organisms were assessed on the Gram-stained slide. Using the 10(3) colony forming units (cfu.ml-1) threshold for defining PSB as positive or negative, DE had a sensitivity of 85% and a specificity of 94%. In a parallel in vitro study, 18 pairs of PSB specimens were collected from respiratory secretions inoculated with S. aureus. From each pair, one brush was processed as described above and the other was smeared on a glass slide prior to performance of QC. Using direct smear instead of cytocentrifuged preparation, slightly but significantly affected QC. Direct examination of cytospin slides is highly predictive of quantitative bacterial culture results, and provides rapid information regarding the Gram-stain morphology of the causative organisms. It may therefore guide initial therapy.
Bronchoalveolar lavage during neutropenic episodes: diagnostic yield and cellular pattern
Authors: Cordonnier C , Escudier E , Verra F , Brochard L , Bernaudin JF , Fleury-Feith J
Published: October 2003

Few data are available concerning the relationship between alveolar and blood cell populations during neutropenia. We wanted to compare the value of pulmonary endoscopic procedures with lavage in neutropenic (polymorphonuclear (PMN) count < or = 1,000.mm-3) and non-neutropenic settings. We therefore, retrospectively, reviewed the results of 118 investigations for pneumonia in patients with malignant haematological diseases. All had bronchoalveolar lavage (BAL), and some had additional studies with protected bacteriological samples. Each BAL specimen was studied after cytocentrifugation by cytological examination for opportunistic infections, haemorrhage, virus, legionellae, and bacteriological cultures. The diagnostic yield of all endoscopic procedures (BAL, telescoping plugged catheter and protected specimen brush) was 53% in neutropenic (Group 1) and 61% in non-neutropenic (Group 2) patients. The aetiological pattern of pneumonia was nearly the same in the two groups, except for more alveolar proteinosis in Group 1 and more cytomegalovirus (CMV) in Group 2. The absolute number of alveolar cells recovered through BAL (total number, macrophages, lymphocytes and PMNs) was significantly lower in neutropenic patients. We conclude that: 1) neutropenic patients with pneumonia require the same investigative approach as non-neutropenic patients; 2) profound neutropenia may be concomitant with a decreased cellularity of alveoli, which may reflect the consequences of marrow aplasia on the pulmonary cell population and/or direct effect of chemotherapy on the lung.
Altered bactericidal activity against Staphylococcus aureus in tuberculous bronchoalveolar lavage fluids
Authors: Selvaraj P , Venkataprasad N , Vijayan VK , Narayanan PR
Published: October 2003

We wished to evaluate the pulmonary defence capacity against common bacterial infections in patients with pulmonary tuberculosis. Bactericidal activity against Staphylococcus aureus of bronchoalveolar lavage fluids (cell-free supernatants) of patients with active (n = 13) and inactive pulmonary tuberculosis (n = 8), and normal individuals (n = 6), were studied. The 2 and 4 h bactericidal activities were higher than the 0 h activity in lavage fluids of healthy subjects and patients with inactive pulmonary tuberculosis. Active tuberculous lavage fluids were equally competent in their bactericidal activity against S. aureus at 0 and 2 h, but a reduced S. aureus killing was seen at 4 h of incubation. Estimation of total phospholipid levels revealed no significant difference between the various lavage fluids. This reduced killing of S. aureus showed a relationship with the cellular components (lymphocytes and macrophages) of active tuberculous lavage fluids. A reduced killing was associated with no lymphocytic alveolitis, and an increased killing with lymphocytic alveolitis. This study suggests that alveolar lining material of patients with active pulmonary tuberculosis has less bactericidal activity against bacterial infections, such as S. aureus.
Lung cavitation associated with Pneumocystis carinii infection in the acquired immunodeficiency syndrome: a report of six cases and review of the literature
Authors: Ferre C , Baguena F , Podzamczer D , Sanchez C , Viladrich PF , Garau J , Gudiol F
Published: October 2003

Lung cavitation in patients with the acquired immune deficiency syndrome (AIDS) and Pneumocystis carinii pneumonia (PCP) has mainly been reported as single case studies. Among 160 episodes of PCP seen in a 1,000 bed teaching hospital and a 600 bed teaching hospital from 1985-1992, we found six cases presenting with lung cavitation and documented Pneumocystis carinii infection. In the cases we report, as well as in the cases reviewed, cavities appear either alone or within an area of pulmonary consolidation, a mass or a nodule. They may present with haemoptysis, show unusual locations, and, most importantly, may frequently be misdiagnosed by bronchoalveolar lavage.
Increased gamma/delta-positive T-cells in blood and bronchoalveolar lavage of patients with sarcoidosis and hypersensitivity pneumonitis
Authors: Raulf M , Liebers V , Steppert C , Baur X
Published: October 2003

A small population of T-cells does not express the conventional T-cell receptor (TCR), characterized by the alpha and beta polypeptide chains (alpha/beta TCR) but two polypeptides termed gamma and delta (gamma/delta TCR). Changes in gamma/delta TCR expression may be relevant as the cause or consequence of several diseases. Our study was undertaken to determine and compare the distribution of T-cells expressing gamma/delta TCR in blood and bronchoalveolar lavage (BAL) of patients with sarcoidosis, hypersensitivity pneumonitis (HP), idiopathic pulmonary fibrosis (IPF), and of healthy controls. In addition, the association between gamma/delta TCR of blood T-lymphocytes and accessory molecules was analysed. Using direct immunofluorescence with the anti-gamma/delta TCR and anti-CD3 monoclonal antibodies (MoAbs) followed by flow cytometric analysis, the blood of patients with pulmonary sarcoidosis, HP, IPF and of healthy controls was analysed. To reveal the association between gamma/delta TCR of blood T-lymphocytes and the accessory molecules, expression of CD4, CD8 and CD25 were determined. Calculating the percentage and the total number of CD3+ gamma/delta TCR cells in blood, the data indicated a significant increase of gamma/delta T-cells in individuals with pulmonary sarcoidosis and HP, compared to healthy controls and IPF patients. In sarcoidosis patients with elevated CD3+ gamma/delta TCR levels, significantly lower CD4/CD8 ratios were observed. In addition, our data demonstrate a correlation between the decrease of CD4+ cells in blood and the amplified appearance of gamma/delta TCR expression in sarcoidosis patients, but not in HP patients.(ABSTRACT TRUNCATED AT 250 WORDS)
Medical treatment of pulmonary hypertension in chronic lung disease
Authors: Weitzenblum E , Kessler R , Oswald M , Fraisse P
Published: October 2003

In chronic respiratory diseases, especially chronic obstructive pulmonary disease (COPD) pulmonary arterial hypertension is generally mild to moderate and the necessity for treating it can, therefore, be questioned. In fact, pulmonary hypertension, even when modest, may worsen markedly during acute episodes, exercise and sleep. These acute increases in mean pulmonary artery pressure (PAP) could contribute to the development of right heart failure. Therefore, the medical treatment of pulmonary hypertension is justified. There are, at the present time, no selective pulmonary vasodilators, with the exception of inhaled nitric oxide. Indeed, vasodilators appear less effective in COPD compared to primary pulmonary hypertension. Thus, there is, at present, no justification for the long-term use of vasodilators in COPD patients. Long-term oxygen therapy (LTOT) attenuates and sometimes reverses the progression of pulmonary hypertension, although the condition rarely returns to normal. We do not know whether the structural changes of the pulmonary vasculature in COPD patients are potentially reversible with LTOT. The longer the daily duration of LTOT the better are the haemodynamic results. At present, LTOT remains the best treatment for pulmonary hypertension in COPD patient. In the future, treatment of this condition in COPD patients could combine LTOT and specific vasodilators.
The epidemiology of occupational asthma
Authors: Beckett WS
Published: October 2003

With the accelerating pace of new knowledge about occupational asthma, systematic epidemiological approaches have yielded important new insights into its clinical characteristics. Such questions as "what proportion of all cases of asthma are caused by occupational exposures?" and "is occupational asthma incidence rising?" can now be answered. Surveillance programmes estimate the number of exposed individuals and the number of incident cases, allowing us to perceive the magnitude of the health problem. Although occupational asthma is a relatively common illness, very little is known about quantitative exposure-response relationships. Epidemiological techniques can quantify the importance of predisposing factors, such as atopy or cigarette smoking, in determining individual risk factors for occupational asthma. Still to be determined are the major, possibly genetic factors which make occupational asthma an "idiosyncratic" illness, affecting only a minority of those with equivalent exposures in the workplace. The goal of applying epidemiological techniques to the study of occupational asthma is ultimately to identify more effective means to prevent its occurrence.
Exposure to environmental tobacco smoke and adult non-neoplastic respiratory diseases
Authors: Tredaniel J , Boffetta P , Saracci R , Hirsch A
Published: October 2003

Exposure to environmental tobacco smoke (ETS) is widespread in European countries, the most serious exposures occurring at home and in the workplace. Epidemiological studies have, essentially, addressed the association between ETS exposure and respiratory health in children, and increased risk of lung cancer among adult nonsmokers. Relatively few studies have been reported on ETS and adult non-neoplastic respiratory diseases. On the basis of the available data, no definite conclusion (excluding the acute irritating effect of ETS on respiratory mucous membranes) can be drawn. Although biologically plausible, it remains controversial whether ETS exposure is associated with chronic respiratory symptoms and occurrence of chronic obstructive pulmonary disease, including asthma. Most of the studies that have used the most sensitive indicators of pulmonary function have suggested a negative impact of ETS exposure. However, if really present, the physiological significance of such small changes is unclear, and the relationship to long-term changes in lung function is not established. Moreover, the possibility of bias and confounding factors must be taken into account. Thus, there is a need for further epidemiological studies on ETS exposure and adult non-neoplastic respiratory disorders.
Serum tumour markers in lung cancer: history, biology and clinical applications
Authors: Ferrigno D , Buccheri G , Biggi A
Published: October 2003

The association of biological markers with cancer has been recognized for many decades. Current interest in markers for cancer arose in the mid 1960s, with the discoveries of alpha-fetoprotein and carcinoembryonic antigen. They were called oncofetal proteins, because of their presence in high concentrations during embryonic development, their virtual disappearance in the neonatal period, and their reappearance with cancers of specific cell types. Essentially, any molecular species may be produced in abnormal amounts or under abnormal circumstances by a tumour, and thereby become useful as a tumour marker. Several tumour markers have been studied in lung cancer. Unfortunately, none of these appear to be sufficiently sensitive and specific to be reliable for screening and diagnostic purposes. However, there is a body of evidence which proves that at least some of these substances may be useful in the evaluation of the course and prognosis of the disease. This review presents data concerning the most studied and interesting tumour markers in lung cancer.
Prevention of thermal and condensation errors in pneumotachographic recordings of the maximal forced expiratory manoeuvre
Authors: Miller MR , Sigsgaard T
Published: October 2003

When recording respiratory manoeuvres using a pneumotachograph (PT), it is customary to heat the pneumotachograph head to avoid condensation errors. It is technically demanding to heat the PT correctly, and this increases the complexity of the equipment. We have studied whether a simpler method can eradicate condensation errors and preserve thermal stability of the PT. We tested the accuracy of a PT by recording the discharge of 1,000 ml of air through the PT before and after a subject had blown through it, thus causing both condensation within the PT and a rise in its temperature. We then determined whether the thermal and condensation errors could be avoided by placing the PT, between blows, on a fan that was blowing air at a constant 1.65 l.s-1. With ambient temperature at 20 degrees C, a single blow through the PT was found to cause a subsequent 4% over-reading in volume, due to condensation and temperature changes in the PT. After five consecutive blows the error was 7%. Placing the PT on the fan for 15 s after a single blow abolished these errors, whereas a wait of 15 s did not. At an ambient temperature of 14 degrees C there was a 5% over-reading of volume after a single blow, and an over-reading of 9% after five blows. At an ambient temperature of 26 degrees C these errors were 3 and 5%, respectively. All of these errors were abolished by placing the PT on the fan for 15 s and 30 s, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Bochdalek hernia: a rare cause of pleural empyema
Authors: Steenhuis LH , Tjon Tham A RT , Smeenk FW
Published: October 2003

This case report describes pleural empyema, caused by an intrathoracic ruptured stomach, in an adult patient with Bochdalek hernia. The possible complications and difficulties in diagnosing Bochdalek hernia in the adult are discussed.
Following severe injury, hypocholesterolemia improves with convalescence but persists with organ failure or onset of infection
Authors: Dunham CMichael , Fealk Michael H , Sever Wilbur E
Published: October 2003

Introduction

Our primary objective was to determine the impact of traumatic injury, onset of infection, organ/metabolic dysfunction, and mortality on serum cholesterol.

Methods

During 676 surgical intensive care unit (SICU) days, 28 ventilated trauma patients underwent daily measurement of white blood cell (WBC) count and differential, cholesterol, arterial oxygen tension/fractional inspired oxygen, bilirubin, glucose, creatinine, and bicarbonate. With the onset of infection, WBC response was considered positive if the WBC count was 16.0 or greater, immature neutrophils were 10% or greater, or WBC count increased by 20%. Cholesterol response was considered positive if cholesterol decreased or failed to increase by 10%.

Results

Injury Severity Score was 30.6 ± 8.6 and there were 48 infections. Initial cholesterol was decreased (119 ± 44 mg/dl) compared with expected values from a database (201 ± 17 mg/dl; P < 0.0001). The 25 survivors had higher cholesterol at SICU discharge (143 ± 35 mg/dl) relative to admission (112 ± 37 mg/dl; P < 0.0001). In the three patients who died, the admission cholesterol was 175 ± 62 mg/dl and the cholesterol at death was 117 ± 27 mg/dl. The change in percentage of expected cholesterol (observed value divided by expected value) from admission to discharge was different for patients surviving (16 ± 19%) and dying (-29 ± 19%; P = 0.0005). With onset of infection, the WBC response was positive in 61% and cholesterol response was positive in 91% (P = 0.001). Percentage of expected cholesterol was decreased with each system dysfunction: arterial oxygen tension/fractional inspired oxygen < 350, creatinine > 2.0 mg/dl, glucose > 120 mg/dl, bilirubin > 2.5 mg/dl, and bicarbonate ≥ 28 or ≤ 23 (P < 0.01). Percentage of expected cholesterol decreased as the number of dysfunctions increased (P = 0.0001).

Conclusion

Hypocholesterolemia is seen following severe injury. Convalescing patients (ready for SICU discharge) have improved cholesterol levels, whereas dying patients appear to have progressive hypocholesterolemia. Decreasing or fixed cholesterol levels suggest the development of infection or organ/metabolic dysfunction. Cholesterol responses are more sensitive for the onset of infection than are WBC responses. Sequential cholesterol monitoring is recommended for patients with severe trauma.
CCL16/LEC powerfully triggers effector and antigen-presenting functions of macrophages and enhances T cell cytotoxicity
Authors: Cappello Paola , Caorsi Cristiana , Bosticardo Marita , De Angelis Stefania , Novelli Francesco , Forni Guido , Giovarelli Mirella
Published: October 2003

The huan CC chemokine CCL16, a liver-expressed chemokine, enhances the killing activity of mouse peritoneal macrophages by triggering their expression of tumor necrosis factor α (TNF-α) and Fas ligand. Macrophages also respond to CCL16 by enhancing their production of monocyte chemoattractant protein-1, regulated on activation, normal T cells expressed and secreted chemokines, and interleukin (IL)-1β, TNF-α, and IL-12. The effect of CCL16 is almost as strong as that of lipopolysaccharide and interferon-γ, two of the best macrophage activators. Moreover, CCL16-activated macrophages overexpress membrane CD80, CD86, and CD40 costimulatory molecules and extensively phagocytose tumor cell debris. On exposure to such debris, they activate a strong, tumor-specific, cytolytic response in virgin T cells. Furthermore, cytolytic T cells generated in the presence of CCL16 display a higher cytotoxicity and activate caspase-8 in tumor target cells. This ability to activate caspase-8 depends on their overexpression of TNF-α and Fas ligand induced by CCL16. These data reveal a new function for CCL16 in the immune-response scenario. CCL16 significantly enhances the effector and the antigen-presenting function of macrophages and augments T cell lytic activity.
The interaction of human dendritic cells with yeast and germ-tube forms ofCandida albicansleads to efficient fungal processing, dendritic cell maturation, and acquisition of a Th1 response-promoting function
Authors: Romagnoli G , Nisini R , Chiani P , Mariotti S , Teloni R , Cassone A , Torosantucci A
Published: October 2003

T helper cell type 1 (Th1) cell-mediated immunity plays a rical role in protection against the opportunistic pathogen Candida albicans. Virulence of the fungus is closely associated with its ability to form germ-tubes (GT), the early phase of the dimorphic transition from the commensal yeast (Y) to the more invasive hyphal (H) form. In this study, we examined the functional outcome of the interaction of Y or GT forms with human dendritic cells (DCs), professional antigen-presenting cells, which are pivotal for initiation and modulation of T cell responses. DCs phagocytosed and killed Y and GT cells with a comparable efficiency, becoming able to trigger strong proliferative responses by Candida-specific, autologous T cell clones. Both fungal forms induced DC maturation, as indicated by up-regulation of CD83, CD80, CD86, CD40, and major histocompatibility complex classes I and II surface antigens. Chemokine receptors were also modulated in Candida–DCs, which showed increased CCR7/CXCR4 and decreased CCR5 expression. Y- and GT-activated DCs differed in the pattern of cytokine expression. In particular, GT cells, in common with fully differentiated H cells, induced significantly more elevated levels of interleukin (IL)-10 than Y cells. Nevertheless, Y-, GT-, or H-pulsed DCs secreted comparable amounts of IL-12p70. In addition, irrespective of the fungal form triggering DC activation, Candida–DCs acquired the ability to prime naive T lymphocytes with a defined Th1 phenotype. Overall, our findings highlight the induction of substantially similar functional patterns in human DCs encountering the different forms of growth of C. albicans, both seemingly activating the Th1-type immunity which is characteristic of the healthy human subjects, naturally immunized and protected against the fungus.
Injury, sepsis, and the regulation of Toll-like receptor responses
Authors: Murphy Thomas J , Paterson Hugh M , Mannick John A , Lederer James A
Published: October 2003

Although we tend to think that the immune system has evolved to protect the host from invading pathogens and to discriminate between self and nonself, there must also be an element of the immune system that has evolved to control the response to tissue injury. Moreover, these potential immune-regulatory pathways controlling the injury response have likely coevolved in concert with self and nonself discriminatory immune-regulatory networks with a similar level of complexity. From a clinical perspective, severe injury upsets normal immune function and can predispose the injured patient to developing life-threatening infectious complications. This remains a significant health care problem that has driven decades of basic and clinical research aimed at defining the functional effects of injury on the immune system. This review and update on our ongoing research efforts addressing the immunological response to injury will highlight some of the most recent advances in our understanding of the impact that severe injury has on the innate and adaptive immune system focusing on phenotypic changes in innate immune cell responses to Toll-like receptor stimulation.
Expression and regulation of antimicrobial peptides in the gastrointestinal tract
Authors: Cunliffe R N , Mahida Y R
Published: October 2003

The gastrointestinal (GI) tract is exposed to a wide range of microorganisms. The expression of antimicrobial peptides has been demonstrated in different regions of the GI tract, predominantly in epithelial cells, which represent the first host cells with which the microorganisms have to interact for invasion. The intestinal epithelial monolayer is complex, consisting of different cell types, and most have a limited lifespan. Of the GI antimicrobial peptides, α- and β-defensins have been studied the most and are expressed by distinct types of epithelial cells. Enteric α-defensin expression is normally restricted to Paneth and intermediate cells in the small intestine. However, there are important differences between mice and humans in the processing of the precursor forms of enteric α-defensins. Parasite infection induces an increase in the number of enteric α-defensin-expressing Paneth and intermediate cells in the murine small intestine. In the chronically inflamed colonic mucosa, metaplastic Paneth cells (which are absent in the normal colon) also express enteric α-defensins. Epithelial expression of β-defensins may be constitutive or nducible by infectious and inflammatory stimuli. The production of some members of the β-defensin family appears to be restricted to distinct parts of the GI tract. Recent studies using genetically manipulated rodents have demonstrated the likely in vivo importance of enteric antimicrobial peptides in innate host defense against microorganisms. The ability of these peptides to act as chemoattractants for cells of the innate- and adaptive-immune system may also play an important role in perpetuating chronic inflammation in the GI tract.
Interferon-γ: an overview of signals, mechanisms and functions
Authors: Schroder Kate , Hertzog Paul J , Ravasi Timothy , Hume David A
Published: October 2003

Interferon-γ (IFN-γ) coordinates a diverse array of cellular programs through transcriptional regulation of immunologically relevant genes. This article reviews the current understanding of IFN-γ ligand, receptor, ignal transduction, and cellular effects with a focus on macrophage responses and to a lesser extent, responses from other cell types that influence macrophage function during infection. The current model for IFN-γ signal transduction is discussed, as well as signal regulation and factors conferring signal specificity. Cellular effects of IFN-γ are described, including up-regulation of pathogen recognition, antigen processing and presentation, the antiviral state, inhibition of cellular proliferation and effects on apoptosis, activation of microbicidal effector functions, immunomodulation, and leukocyte trafficking. In addition, integration of signaling and response with other cytokines and pathogen-associated molecular patterns, such as tumor necrosis factor-α, interleukin-4, type I IFNs, and lipopolysaccharide are discussed.
Activation of A2A adenosine receptors inhibits expression of α4/β1 integrin (very late antigen-4) on stimulated human neutrophils
Authors: Sullivan Gail W , Lee David D , Ross William G , DiVietro Jeffrey A , Lappas Courtney M , Lawrence Michael B , Linden Joel
Published: October 2003

The α4/β1 integrin very late antigen-4 (CD49d/CD29) is up-regulated on circulating neutrophils of septic patients. Although no individual agent mimics this effect of sepsis, we now report that following priming of human neutrophils with lipopolysaccharide or tumor necrosis factor α (TNF-α), addition of formyl-Met-Leu-Phe (fMLP) results in a “stimulated”, sepsis-like, four- to fivefold rise in CD49d expression. TNF/fMLP stimulation also produced a similar increase in CD49d-mediated adhesion of neutrophils to a vascular cell adhesion molecule-1 (VCAM-1)-coated surface. Adenosine is a naturally occurring, anti-inflammatory mediator released from injured or inflamed tissues. We observed that stimulated neutrophil CD49d expression was decreased by activation of A2A adenosine receptors (A2AAR) with the selective agonist 4-{3-[6-amino-9-(5-ethylcarbamoyl-3,4-dihydroxy-tetrahydro-furan-2-yl)-9H-purin-2-yl]-prop-2-ynyl}-cyclohexanecarboxylicacid methyl ester (ATL146e; EC50=6.4 nM). ATL146e (100 nM) also reduced the fraction of stimulated neutrophils that adhered to VCAM-1 from 38 ± 6% to 27 ± 5%. Inhibition of CD49d expression was equally inhibited by ATL146e, added before or after TNF priming, and was reversed by incubation with the A2AAR-selective antagonist 4-{2-[7-amino-2-(2-furyl) (1, 2, 4)triazolo(2,3-a) (1, 3, 5)triazin-5-yl-amino]ethyl}-phenol (ZM241385; 100 nM). A suboptimal ATL146e concentration (1 nM) combined with the type IV phosphodiesterase inhibitor rolipram (100 nM) synergistically decreased stimulated CD49d expression by >50%. The cyclic adenosine monophosphate (cAMP)-dependent kinase [protein kinase A (PKA)] inhibitor H-89 (10 μM) reversed the effect of ATL146e on stimulated CD49d expression. Other means of increasing cAMP in neutrophils also decreased stimulated CD49d expression. We conclude that adenosine binding to A2AAR counteracts stimulation of neutrophil CD49d integrin expression and neutrophil binding to VCAM-1 via a cAMP/PKA-mediated pathway.
Subset-specific, uniform activation among Vγ6/Vδ1+ γδ T cells elicited by inflammation
Authors: Roark Christina L , Aydintug M Kemal , Lewis Julie , Yin Xiang , Lahn Michael , Hahn Youn-Soo , Born Willi K , Tigelaar Robert E , O’Brien Rebecca L
Published: October 2003

The Vγ6/Vδ1+ cells, the second murine γδ T cell subset to arise in the thymus, express a nearly invariant T cell receptor (TCR), colonize select tissues, and expand preferentially in other tissues during inflammation. These cells are thought to help in regulating the inflammatory response. Until now, Vγ6/Vδ1+ cells have only been detectable indirectly, by expression of Vγ6-encoding mRNA. Here, we report that 17D1, a monoclonal antibody, which detects the related epidermis-associated Vγ5/Vδ1+ TCR, will also bind the Vγ6/Vδ1+ cells if their TCR is first complexed to an anti-Cδ antibody. Features of this special condition for recognition suggest the possibility that an alternate structure exists for the Vγ6/Vδ1 TCR, which is stabilized upon binding to the anti-Cδ antibody. Using the 17D1 antibody as means to track this γδ T cell subset by flow cytometry, we discovered that the response of Vγ6/Vδ1+ cells during inflammation often far exceeds that of other subsets and that the responding Vγ6/Vδ1+ cells display a strikingly uniform activation/memory phenotype compared with other γδ T cell subsets.
In vitro model for hematopoietic progenitor cell homing reveals endothelial heparan sulfate proteoglycans as direct adhesive ligands
Authors: Netelenbos Tanja , van den Born Jacob , Kessler Floortje L , Zweegman Sonja , Huijgens Peter C , Dräger Angelika M
Published: October 2003

Proteoglycans (PGs) play a dominant role within the bone marrow (BM), but their role in homing of transplanted hematopoietic progenitor cells (HPC) is unknown. In this study, the role of heparan sulfate (HS) PGs on BM endothelium as adhesive structures was investigated. HPC (primary CD34+ cells and cell line KG-1a) were able to bind fractionated heparin, which could be competed by highly sulfated heparin/HS-glycosaminoglycans (GAGs). Under flow conditions, HPC adhered to immobilized heparin after rolling over E-selectin. Rolling of KG-1a on BM endothelial cell (EC) line 4LHBMEC was completely E selectin-dependent. Addition of heparin/HS-GAGs, endothelial treatment with chlorate, or anti-HS all partially inhibited firm adhesion. Moreover, enzymatic removal of endothelial HS-GAGs reduced initial adhesion. Finally, HPC-bound PGs isolated from 4LHBMEC, which was largely inhibited by enzymatic HS-degradation. In summary, we identified sulfated structures on BM endothelium, most likely HSPGs, as a novel class of glycoconjugates involved in the multistep homing cascade of HPC.
Antiequine Chorionic Gonadotropin (eCG) Antibodies Generated in Goats Treated with eCG for the Induction of Ovulation Modulate the Luteinizing Hormone and Follicle-Stimulating Hormone Bioactivities of eCG Differently
Authors: Hervé Virginie , Roy François , Bertin Jean , Guillou Florian , Maurel Marie-Christine
Published: October 2003

In dairy goats, treatments associating a progestogen and the equine chorionic gonadotropin (eCG) are the easiest way to induce and synchronize estrus and ovulation and to permit artificial insemination (AI) and/or out of season breeding. From the first treatment, the injection of eCG induces, in some females, the production of anti-eCG antibodies (Abs) that will interfere with the effectiveness of subsequent treatments. These anti-eCG Abs delay the preovulatory LH surge and the ovulation time, leading to poor fertility of the treated females. In this study, by in vitro bioassays, we show that anti-eCG Abs can positively or negatively modulate the LH and/or FSH bioactivities of eCG. Moreover, the modulation level of eCG bioactivity does not depend on the anti-eCG Ab affinity for eCG, as shown by surface plasmon resonance technology. The specificity of anti-eCG Abs tested by competitive ELISA highlighted the importance of a glycan environment in the recognition mechanism, especially the sialic acids specific to eCG. The different effects of anti-eCG Abs on eCG bioactivities could be explained by two hypotheses. First, steric hindrance preventing the interaction of eCG with its receptors would explain the inhibitory effect of some anti-eCG Abs; second, a conformational change in eCG by anti-eCG Abs could induce inhibition or potentiation of eCG bioactivities. It is significant that these modulations of eCG bioactivities by anti-eCG Abs impact mainly on the FSH bioactivity of eCG, which is essential for ovarian stimulation and subsequent fertility after treatment and AI, and to a lesser extent on LH bioactivity.
The Volume of a Sexually Dimorphic Nucleus in the Ovine Medial Preoptic Area/Anterior Hypothalamus Varies with Sexual Partner Preference
Authors: Roselli Charles E. , Larkin Kay , Resko John A. , Stellflug John N. , Stormshak Fred
Published: October 2003

Sheep are one of the few animal models in which natural variations in male sexual preferences have been studied experimentally. Approximately 8% of rams exhibit sexual preferences for male partners (male-oriented rams) in contrast to most rams, which prefer female partners (female-oriented rams). We identified a cell group within the medial preoptic area/anterior hypothalamus of age-matched adult sheep that was significantly larger in adult rams than in ewes. This cell group was labeled the ovine sexually dimorphic nucleus (oSDN). In addition to a sex difference, we found that the volume of the oSDN was two times greater in female-oriented rams than in male-oriented rams. The dense cluster of neurons that comprise the oSDN express cytochrome P450 aromatase. Aromatase mRNA levels in the oSDN were significantly greater in female-oriented rams than in ewes, whereas male-oriented rams exhibited intermediate levels of expression. Because the medial preoptic area/anterior hypothalamus is known to control the expression of male sexual behaviors, these results suggest that naturally occurring variations in sexual partner preferences may be related to differences in brain anatomy and capacity for estrogen synthesis.
Activins Are Critical Modulators of Growth and Survival
Authors: Brown Chester W. , Li Liunan , Houston-Hawkins Dianne E. , Matzuk Martin M.
Published: October 2003

Activins βA and βB (encoded by Inhba and Inhbb genes, respectively) are related members of the TGF-β superfamily. Previously, we generated mice with an Inhba knock-in allele (InhbaBK) that directs the expression of activin βB protein in the spatiotemporal pattern of activin βA. These mice were small and had shortened life spans, both influenced by the dose of the hypomorphic InhbaBK allele. To understand the mechanism(s) underlying these abnormalities, we now examine growth plates, liver, and kidney and analyze IGF-I, GH, and major urinary proteins. Our studies show that activins modulate the biological effects of IGF-I without substantial effects on GH, and that activin signaling deficiency also has modest effects on hepatic and renal function. To assess the relative influences of activin βA and activin βB, we produced mice that express activin βB from the InhbaBK allele, and not from its endogenous Inhbb locus. InhbaBK/BK, Inhbb−/− mice have failure of eyelid fusion at birth and demonstrate more severe effects on somatic growth and survival than either of the corresponding single homozygous mutants, showing that somatic growth and life span are supported by both activins βA and βB, although activin βA plays a more substantial role.
Evidence for lymphatic transport of insulin by topically applied biphasic vesicles
Authors: King Martin J , Michel Deborah , Foldvari Marianna
Published: October 2003

The cutaneous delivery pathway through the lymphatics of a novel transdermal lipid-based delivery system (biphasic vesicles), which was previously shown to deliver sustained physiological levels of basal insulin in a pain-free manner across the skin, was evaluated in a diabetic rat model. Transdermal patches (one per rat) containing insulin in biphasic vesicles (1–10 mg recombinant human insulin dose) were applied to the shaved abdominal skin of streptozotocin-induced diabetic rats for 73 h. Blood glucose was monitored approximately every 2–10 h using a Lifescan glucose meter. Inguinal lymph node insulin levels were analysed by ELISA. Insulin in the lymph nodes increased in a dose- and time-dependent manner. Maximal transdermal insulin concentrations in the lymph nodes were observed with both 140 IU (5 mg: 43.0 + 18.0 μIU mg−1 (mean + s.e.m., n = 4)) and 280 IU (10 mg: 48.0 + 19.6 μIU mg−1 (mean + s.e.m., n = 4)) doses of recombinant insulin at t = 73 h. The level of insulin in the lymph nodes after subcutaneous injection of 1 mg insulin at the peak blood glucose response was 35.8 μIU mg−1 (n = 2), before falling to 0.35 μIU mg−1 by t = 48 h (n = 2). The lymphatics is involved in the transdermal insulin delivery by biphasic vesicles. This is the first report on the lymphatic transport of a protein after non-invasive topical application on the skin.
Wnt signaling is required at distinct stages of development for the induction of the posterior forebrain
Authors: Braun Michelle M. , Etheridge Alton , Bernard Amy , Robertson Christie P. , Roelink Henk
Published: October 2003

One of the earliest manifestations of anteroposterior pattering in the developing brain is the restricted expression of Six3 and Irx3 in the anterior and posterior forebrain, respectively. Consistent with the role of Wnts as posteriorizing agents in neural tissue, we found that Wnt signaling was sufficient to induce Irx3 and repress Six3 expression in forebrain explants. The position of the zona limitans intrathalamica (zli), a boundary-cell population that develops between the ventral (vT) and dorsal thalamus (dT), is predicted by the apposition of Six3 and Irx3 expression domains. The expression patterns of several inductive molecules are limited by the zli,including Wnt3, which is expressed posterior to the zli in the dT. Wnt3 and Wnt3a were sufficient to induce the dT marker Gbx2 exclusively in explants isolated posterior to the presumptive zli. Blocking the Wnt response allowed the induction of the vT-specific marker Dlx2 in prospective dT tissue. Misexpression of Six3 in the dT induced Dlx2expression and inhibited the expression of both Gbx2 and Wnt3. These results demonstrate a dual role for Wnt signaling in forebrain development. First, Wnts directed the initial expression of Irx3 and repression of Six3 in the forebrain, delineating posterior and anterior forebrain domains. Later, continued Wnt signaling resulted in the induction of dT specific markers, but only in tissues that expressed Irx3.
Genetic analysis of adventitious root formation with a novel series of temperature-sensitive mutants ofArabidopsis thaliana
Authors: Konishi Mineko , Sugiyama Munetaka
Published: October 2003

When cultured on media containing the plant growth regulator auxin,hypocotyl explants of Arabidopsis thaliana generate adventitious roots. As a first step to investigate the genetic basis of adventitious organogenesis in plants, we isolated nine temperature-sensitive mutants defective in various stages in the formation of adventitious roots: five root initiation defective(rid1 to rid5) mutants failed to initiate the formation of root primordia; in one root primordium defective (rpd1)mutant, the development of root primordia was arrested; three root growth defective (rgd1, rgd2, and rgd3) mutants were defective in root growth after the establishment of the root apical meristem. The temperature sensitivity of callus formation and lateral root formation revealed further distinctions between the isolated mutants. The rid1mutant was specifically defective in the reinitiation of cell proliferation from hypocotyl explants, while the rid2 mutant was also defective in the reinitiation of cell proliferation from root explants. These two mutants also exhibited abnormalities in the formation of the root apical meristem when lateral roots were induced at the restrictive temperature. The rgd1and rgd2 mutants were deficient in root and callus growth, whereas the rgd3 mutation specifically affected root growth. The rid5 mutant required higher auxin concentrations for rooting at the restrictive temperature, implying a deficiency in auxin signaling. The rid5 phenotype was found to result from a mutation in the MOR1/GEM1 gene encoding a microtubule-associated protein. These findings about the rid5 mutant suggest a possible function of the microtubule system in auxin response.
Neuronal defects in the hindbrain ofHoxa1, Hoxb1andHoxb2mutants reflect regulatory interactions among these Hox genes
Authors: Gavalas Anthony , Ruhrberg Christiana , Livet Jean , Henderson Christopher E. , Krumlauf Robb
Published: October 2003

Hox genes are instrumental in assigning segmental identity in the developing hindbrain. Auto-, cross- and para-regulatory interactions help establish and maintain their expression. To understand to what extent such regulatory interactions shape neuronal patterning in the hindbrain, we analysed neurogenesis, neuronal differentiation and motoneuron migration in Hoxa1, Hoxb1 and Hoxb2 mutant mice. This comparison revealed that neurogenesis and differentiation of specific neuronal subpopulations in r4 was impaired in a similar fashion in all three mutants,but with different degrees of severity. In the Hoxb1 mutants, neurons derived from the presumptive r4 territory were re-specified towards an r2-like identity. Motoneurons derived from that territory resembled trigeminal motoneurons in both their migration patterns and the expression of molecular markers. Both migrating motoneurons and the resident territory underwent changes consistent with a switch from an r4 to r2 identity. Abnormally migrating motoneurons initially formed ectopic nuclei that were subsequently cleared. Their survival could be prolonged through the introduction of a block in the apoptotic pathway. The Hoxa1 mutant phenotype is consistent with a partial misspecification of the presumptive r4 territory that results from partial Hoxb1 activation. The Hoxb2 mutant phenotype is a hypomorph of the Hoxb1 mutant phenotype, consistent with the overlapping roles of these genes in facial motoneuron specification. Therefore, we have delineated the functional requirements in hindbrain neuronal patterning that follow the establishment of the genetic regulatory hierarchy between Hoxa1, Hoxb1 and Hoxb2.
Mixer/Bon and FoxH1/Sur have overlapping and divergent roles in Nodal signaling and mesendoderm induction
Authors: Kunwar Prabhat S. , Zimmerman Steven , Bennett James T. , Chen Yu , Whitman Malcolm , Schier Alexander F.
Published: October 2003

Transcription factors belonging to the FoxH1 and Mixer families are required for facets of Nodal signaling during vertebrate mesendoderm induction. Here, we analyze whether zebrafish proteins related to FoxH1[Schmalspur (Sur)] and Mixer [Bonnie and clyde (Bon)] act within or downstream of the Nodal signaling pathway, test whether these two factors have additive or overlapping activities, and determine whether FoxH1/Sur and Mixer/Bon can account for all Nodal signaling during embryogenesis. We find that sur expression is independent of Nodal signaling and that bon is expressed in the absence of Nodal signaling but requires Nodal signaling and Sur for enhanced, maintained expression. These results and the association of FoxH1 and Mixer/Bon with phosphorylated Smad2 support a role for these factors as components of the Nodal signaling pathway. In contrast to the relatively mild defects observed in single mutants, loss of both bon and sur results in a severe phenotype characterized by absence of prechordal plate, cardiac mesoderm, endoderm and ventral neuroectoderm. Analysis of Nodal-regulated proteins reveals that Bon and Sur have both distinct and overlapping regulatory roles. Some genes are regulated by both Bon and Sur, and others by either Bon or Sur. Complete loss of Nodal signaling results in a more severe phenotype than loss of both Bon and Sur,indicating that additional Smad-associated transcription factors remain to be identified that act as components of the Nodal signaling pathway.
TheCaenorhabditis elegansnonmuscle myosin genesnmy-1andnmy-2function as redundant components of thelet-502/Rho-binding kinase andmel-11/myosin phosphatase pathway during embryonic morphogenesis
Authors: Piekny Alisa J. , Johnson Jacque-Lynne F. , Cham Gwendolyn D. , Mains Paul E.
Published: October 2003

Rho-binding kinase and the myosin phosphatase targeting subunit regulate nonmuscle contractile events in higher eukaryotes. Genetic evidence indicates that the C. elegans homologs regulate embryonic morphogenesis by controlling the actin-mediated epidermal cell shape changes that transform the spherical embryo into a long, thin worm. LET-502/Rho-binding kinase triggers elongation while MEL-11/myosin phosphatase targeting subunit inhibits this contractile event. We describe mutations in the nonmuscle myosin heavy chain gene nmy-1 that were isolated as suppressors of the mel-11hypercontraction phenotype. However, a nmy-1 null allele displays elongation defects less severe than mutations in let-502 or in the single nonmuscle myosin light chain gene mlc-4. This results because nmy-1 is partially redundant with another nonmuscle myosin heavy chain, nmy-2, which was previously known only for its role in anterior/posterior polarity and cytokinesis in the early embryo. At the onset of elongation, NMY-1 forms filamentous-like structures similar to actin, and LET-502 is interspersed with these structures, where it may trigger contraction. MEL-11, which inhibits elongation, is initially cytoplasmic. In response to LET-502 activity, MEL-11 becomes sequestered away from the contractile apparatus, to the plasma membrane, when elongation commences. Upon completion of morphogenesis, MEL-11 again appears in the cytoplasm where it may halt actin/myosin contraction.