The virion-associated Gag-Pol is decreased in chimeric Moloney murine leukemia viruses in which the readthrough region is replaced by the frameshift region of the human immunodeficiency virus type 1

The human immunodeficiency virus type 1 (HIV-1) requires a programmed -1 translational frameshift event to synthesize the precursor of its enzymes, Gag-Pol, when ribosomes from the infected cells translate the full-length viral messenger RNA. Translation of the same RNA according to conventional translational rules produces Gag, the precursor of the structural proteins of the virus. The efficiency of the frameshift controls the ratio of Gag-Pol to Gag, which is critical for viral infectivity. The Moloney murine leukemia virus (MoMuLV) uses a different strategy, the programmed readthrough of a stop codon, to synthesize Gag-Pol. In this study, we investigated whether different forms of the HIV-1 frameshift region can functionally replace the readthrough signal in MoMuLV. Chimeric proviral DNAs were obtained by inserting into the MoMuLV genome the HIV-1 frameshift region encompassing the slippery sequence where the frameshift occurs, followed by the frameshift stimulatory signal. The inserted signal was either a simple stem-loop, previously considered as the stimulatory signal, or a longer bulged helix, now shown to be the complete stimulatory signal, or a mutated version of the complete signal with a three-nucleotide deletion. Although the three chimeric viruses can propagate essentially as the wild-type virus in NIH 3T3 cells, single-round infectivity assays revealed that the infectivity of the chimeric virions is about three to fivefold lower than that of the wild-type virions, depending upon the nature of the frameshift signal. It was also observed that the Gag-Pol to Gag ratio was decreased about two to threefold in chimeric virions. Comparison of the readthrough efficiency of MoMuLV to the HIV-1 frameshift efficiency, by monitoring the expression of a luciferase reporter in cultured cells, revealed that the frameshift efficiencies were only 30-60% of the readthrough efficiency. Altogether, these observations indicate that replacement of the readthrough region of MoMuLV with the frameshift region of HIV-1 results in virions that are replication competent, although less infectious than wild-type MoMuLV. This type of chimera could provide an interesting tool for in vivo studies of novel drugs targeted against the HIV-1 frameshift event.     

Combining immunoreactive trypsinogen and pancreatitis-associated protein assays, a method of newborn screening for cystic fibrosis that avoids DNA analysis

OBJECTIVES: To evaluate the performance of a strategy in which, after immunoreactive trypsinogen (IRT) determination, genetic analysis is replaced by a biological test, the pancreatitis-associated protein (PAP) enzyme-linked immunosorbent assay (ELISA). STUDY DESIGN: The French newborn screening program includes cystic fibrosis (CF) screening by the IRT/CFTR mutation strategy. PAP was assayed on screening cards, in parallel with IRT, in all newborns from 5 French regions (n = 204,749). Analysis of PAP values in CF and non-CF newborns with elevated IRT allowed direct comparison between the current strategy and the proposed IRT/PAP strategy. RESULTS: A protocol in which newborns with IRT >50 ng/mL and PAP >1.8 ng/mL and those with IRT >100 ng/mL and PAP >1.0 ng/mL are directly recalled for sweat testing would have the same performance as the IRT/CFTR mutation strategy. CONCLUSIONS: The IRT/PAP strategy is an alternative for CF newborn screening, which avoids the drawbacks of genetic analysis and is cheaper and easier to implement than the current IRT/CFTR mutation strategy.     

Inhibition of chronic rejection and development of tolerogenic T cells after ICOS-ICOSL and CD40-CD40L co-stimulation blockade

BACKGROUND.: Blockade of the CD40-CD40L pathway results in long-term allograft survival but does not prevent chronic rejection. ICOS-ICOSL are members of the CD28-B7 family that play an important role in T-cell activation. METHODS.: The authors analyzed the effect of single or combined treatment with an anti-ICOS monoclonal antibody and CD40 immunoglobulin (Ig) on acute and chronic rejection of heart allografts in rats. RESULTS.: Treatment with anti-ICOS resulted in a modest but significant prolongation of allograft survival. Treatment with CD40Ig resulted in long-term graft survival but the cardiac grafts developed chronic rejection lesions. Combined CD40Ig+anti-ICOS treatment led to indefinite graft survival in all recipients and a significant decrease of chronic rejection lesions compared with CD40Ig alone. Importantly, four of the seven CD40Ig+anti-ICOS-treated recipients showed a complete absence of chronic rejection lesions, whereas all of the CD40Ig-treated recipients showed chronic rejection. The CD40Ig+anti-ICOS group also showed significant decreased graft infiltration, decreased antidonor cytotoxic T-lymphocyte activity, and decreased alloantibodies compared with the CD40Ig-treated group. Adoptive transfer of splenocytes indefinitely prolonged allograft survival, whereas those depleted of T cells did not, suggesting the development of T-regulatory mechanisms. CONCLUSIONS.: These data indicate that the chronic rejection mechanisms that are CD40-CD40L independent are ICOS-ICOSL dependent. These results were obtained with conservation of cognate immune responses and development of tolerogenic T cells.     

Reduction of macrophage infiltration and chemoattractant gene expression changes in white adipose tissue of morbidly obese subjects after surgery-induced weight loss

In human obesity, the stroma vascular fraction (SVF) of white adipose tissue (WAT) is enriched in macrophages. These cells may contribute to low-grade inflammation and to its metabolic complications. Little is known about the effect of weight loss on macrophages and genes involved in macrophage attraction. We examined subcutaneous WAT (scWAT) of 7 lean and 17 morbidly obese subjects before and 3 months after bypass surgery. Immunomorphological changes of the number of scWAT-infiltrating macrophages were evaluated, along with concomitant changes in expression of SVF-overexpressed genes. The number of scWAT-infiltrating macrophages before surgery was higher in obese than in lean subjects (HAM56+/CD68+; 22.6 +/- 4.3 vs. 1.4 +/- 0.6%, P < 0.001). Typical "crowns" of macrophages were observed around adipocytes. Drastic weight loss resulted in a significant decrease in macrophage number (-11.63 +/- 2.3%, P < 0.001), and remaining macrophages stained positive for the anti-inflammatory protein interleukin 10. Genes involved in macrophage attraction (monocyte chemotactic protein [MCP]-1, plasminogen activator urokinase receptor [PLAUR], and colony-stimulating factor [CSF]-3) and hypoxia (hypoxia-inducible factor-1alpha [HIF-1alpha]), expression of which increases in obesity and decreases after surgery, were predominantly expressed in the SVF. We show that improvement of the inflammatory profile after weight loss is related to a reduced number of macrophages in scWAT. MCP-1, PLAUR, CSF-3, and HIF-1alpha may play roles in the attraction of macrophages in scWAT.     

Epidemic of Trichophyton tonsurans tinea capitis in a nursery school in the Southern suburbs of Paris

INTRODUCTION: In March 2001 a school and family survey was conducted in a nursery school in the Southern suburbs of Paris, during an epidemic of Trichophyton tondsurans tinea. PATIENTS AND METHODS: One hundred twenty-nine children aged 3 to 6 were examined as well as 15 adults working in the school. A survey of the contaminated children or asymptomatic carriers was performed. All the children and adults concerned were treated at the same time, without eviction from school. RESULTS: T. tonsurans was detected in 10 cases of tinea (7.7 p.cent of the persons examined), in 18 cases of cutaneous lesions (13.9 p.cent) and in 25 asymptomatic carriers on the scalp (19.4 p.cent). The majority of the positive cases came from the same school class as the original case: 23 of the 26 children (88 p.cent), with 6 tinea, 14 asymptomatic carriers and 13 cutaneous lesions. Only one of the 15 adults exhibited a T. tonsurans cutaneous lesion. Among the 13 families studied, 2 had several members involved, the first being that of the original case (3 tinea and 2 asymptomatic carriers). DISCUSSION: Several important points are underlined by this study: 1) the high contagiousness of T. tonsurans; 2) the detection of 2 mechanisms of indirect contamination (rag doll mascot in the class and the family hair-clipper); 3) the one-year time lapse between the arrival of the contaminating child in the class and the survey, explaining the extent of the contamination; 4) the underestimation of the epidemic due to the lack of mycological examinations; 5) the identification of several dermatophytes in the same school:M. canis, T soudanense and T. tonsurans, and 7) the futility of eviction from school when all the children can be treated.     

The embryonic origins of hematopoietic stem cells: a tale of hemangioblast and hemogenic endothelium

The developmental origin of hematopoietic stem cells has been for decades the subject of great interest. Once thought to emerge from the yolk sac, hematopoietic stem cells have now been shown to originate from the embryonic aorta. Increasing evidence suggests that hematopoietic stem cells are produced from an endothelial intermediate designated by the authors as hemangioblast or hemogenic endothelium. Recently, the allantois in the avian embryo and the placenta in the mouse embryo were shown to be a site of hematopoietic cell production/expansion and thus appear to play a critical role in the formation of the hematopoietic system. In this review we shall give an overview of the data obtained from human, mouse and avian models on the cellular origins of the hematopoietic system and discuss some aspects of the molecular mechanisms controlling hematopoietic cell production.     

Mitochondrial diseases: molecular mechanisms, clinical presentations and diagnosis investigations

Mitochondrial diseases are relatively common inherited metabolic diseases due to mitochondrial respiratory chain dysfunction. Their clinical presentation is extremely diverse, multisystemic or confined to a single tissue, sporadic or transmitted, by maternal or mendelian inheritance. The diagnosis of mitochondrial disorders is difficult. It is based upon several types of clues both clinical (family history, type of symptoms but also their association in syndromic presentation,...) and biological (alteration of the lactate metabolism, brain imaging, morphological alterations especially of muscle tissue). The diagnosis relies upon the demonstration of a defect of the respiratory chain activities and/or upon the identification of the underlying genetic alteration. Molecular diagnosis remains quite difficult and up to-date concerns essentially mitochondrial DNA mutations. On one hand, clinical and biological presentations as well as enzymatic defects lack specificity. On the other hand, candidate genes are very numerous and part of them are probably still unknown.     

The nurse as an action tool in care for the aged

This study approaches nursing care as related to the aged. The studied situation involved health care needs of hospitalized persons, using the following central question: which is the meaning of nurses' actions when attending hospitalized aged patients without expectation of recovery and when technology is no longer that important? We aimed to reflect about hospitalized elders' needs in nursing reality. Comprehensive Sociology was used as a theoretical-methodological framework. The study was carried out at an Intensive Care Service of a Municipal Hospital in the city of Rio de Janeiro-Brazil. The subjects were nurses who attend hospitalized aged persons without any expectation of recovery, who were approached through a phenomenological interview. Through a comprehensive analysis, we identified care by being together, providing at the same time physical comfort and well-being to cope with the situation as typical of nursing actions. This study indicates some contributions for nursing care, assistance, teaching and research, aimed at strengthening nurses' attitude as an action tool in care for aged patients.