Some properties of marrow derived adherent cells in tissue culture

It has previously been shown that monolayer cultures derived adherent cells (MDAC), apparently consisting of fibroblasts, macrophages, epithelioid cells, and fat cells, can support long-term stem cell proliferation in vitro. In the present study, the hematopoietic support capability of murine MDAC monolayers was confirmed and the cultured cells further characterized with respect to the following properties: esterase I activity, complement (C3) receptors, IgG (Fc) receptors, colony stimulating activity (csa) production, and collagen synthesis. The cultures were also examined immunohistochemically to localize fibronectin, laminin, and collagen synthesis and to identify the collagen subtypes synthesized. MDAC morphology was as described in previous studies, although fat cells were few in number. It was found that MDAC included some cells with esterase I activity and C3 receptors. Fc receptors were not, however, detected, nor did the cultures produce csa, indicating that mononuclear phagocytes were not present. MDAC synthesized collagen types I and III and also fibronectin. Staining for epithelial basement membrane proteins (collagen types IV and V and laminin) was negative. The results indicate that the vast majority of these cultured MDAC were fibroblasts.     

Expanding the mutational spectrum of LZTR1 in schwannomatosis

Schwannomatosis is characterized by the development of multiple non-vestibular, non-intradermal schwannomas. Constitutional inactivating variants in two genes, SMARCB1 and, very recently, LZTR1, have been reported. We performed exome sequencing of 13 schwannomatosis patients from 11 families without SMARCB1 deleterious variants. We identified four individuals with heterozygous loss-of-function variants in LZTR1. Sequencing of the germline of 60 additional patients identified 18 additional heterozygous variants in LZTR1. We identified LZTR1 variants in 43% and 30% of familial (three of the seven families) and sporadic patients, respectively. In addition, we tested LZTR1 protein immunostaining in 22 tumors from nine unrelated patients with and without LZTR1 deleterious variants. Tumors from individuals with LZTR1 variants lost the protein expression in at least a subset of tumor cells, consistent with a tumor suppressor mechanism. In conclusion, our study demonstrates that molecular analysis of LZTR1 may contribute to the molecular characterization of schwannomatosis patients, in addition to NF2 mutational analysis and the detection of chromosome 22 losses in tumor tissue. It will be especially useful in differentiating schwannomatosis from mosaic Neurofibromatosis type 2 (NF2). However, the role of LZTR1 in the pathogenesis of schwannomatosis needs further elucidation.     

Suppression of apoptosis in hematopoietic factor-dependent progenitor cell lines by expression of the FAC gene

Fanconi anemia (FA) is a genetically heterogeneous, inherited blood disorder characterized by bone marrow failure, congenital malformations, and a predisposition to leukemias. Because FA cells are hypersensitive to DNA cross-linking agents and have chromosomal instability, FA has been viewed as a disorder of DNA repair. However, the exact cellular defect in FA cells has not been identified. Sequence analysis of the gene defective in group C patients (FAC) has shown no significant homologies to other known genes. The FAC protein has been localized to the cytoplasm, indicating that FAC may either play an indirect role in DNA repair or is involved in a different cellular pathway. Recent evidence has indicated that FA cells may be predisposed to apoptosis, especially after treatment with DNA cross-linking agents. The demonstration that genes can suppress apoptosis has been accomplished by overexpression of such genes in growth factor-dependent cell lines that die by apoptosis after factor withdrawal. Using retroviral-mediated gene transfer, we present evidence that expression of FAC in the hematopoietic factor-dependent progenitor cell lines 32D and MO7e can suppress apoptosis induced by growth factor withdrawal. Flow cytometry and morphologic analysis of propidium iodide stained cells showed significantly lower levels of apoptosis in FAC-retroviral transduced cells after growth factor deprivation. Expression of FAC in both cell lines promoted increased viability rather than proliferation, which is consistent with other apoptosis-inhibiting genes such as Bcl- 2. These findings imply that FAC may act as a mediator of an apoptotic pathway initiated by growth factor withdrawal. Furthermore, the congenital malformations and hematologic abnormalities characterizing FA may be related to an increased predisposition of FA progenitor cells to undergo apoptosis, particularly in the absence of extracellular signals.     

Assessing the delivery of neutrophils to tissues in neutropenia

Studies of neutrophil kinetics in neutropenic individuals, as well as clinical observations of variability in the occurrence of infection among patients with neutropenia, have suggested that blood neutrophil counts may not uniformly reflect the effective delivery of neutrophils to extravascular tissues where the cells perform their principal host defense functions. To evaluate this possibility we developed a sensitive, reproducible method of measuring the extravascular delivery of neutrophils to a normal mucosal site of neutrophil turnover. This method is based upon the quantification of neutrophils recoverable from saline mouth wash specimens. Twenty-five mL specimens, obtained in a controlled manner from neutropenic patients and normal subjects, were centrifuged and the sediments resuspended in 1.0 mL Hank's buffer with 2 micrograms acridine orange, incubated at 37 degrees C for 15 minutes, and then examined in a hemocytometer chamber by fluorescence microscopy. Neutrophils could be clearly distinguished by their characteristic fluorescence and were counted. With this method as few as 1,500 neutrophils were detected reliably in mouth wash specimens. Mucosal neutrophil counts varied less than 10% with repeated sampling of individual subjects over 5-day periods and were consistently greater than 1.3 X 10(5)/specimen in non-neutropenic individuals. Although profound neutropenia was generally reflected by lower than normal oral mucosal neutrophil counts, these counts were significantly higher in individuals with chronic severe neutropenia (blood neutrophils less than 300/mm3) than in patients with acute neutropenia of comparable severity that had developed following chemotherapy. Also, in individuals recovering from profound neutropenia, neutrophils usually reappeared earlier in mouth wash specimens than in blood, and oral mucosal neutrophil counts attained recovery levels more rapidly than did blood counts. This phenomenon was particularly evident in an individual with cyclic neutropenia. Moreover, mucosal neutrophils could occasionally be detected in profoundly neutropenic patients when neutrophils were not present in blood samples. These findings indicate that mucosal neutrophil counts in individuals with neutropenia provide information about the delivery of neutrophils to tissues that may not be apparent from blood neutrophil counts alone.     

Evading and embracing normality: estrangement and ambivalence in the accounts of methamphetamine consumers

While much has been made of the governmentality evinced in drug policy, its effects on people who use drugs have received less attention. Scholars who have investigated these effects commonly focus on the views and experiences of individuals receiving treatment for their drug use, often reporting an explicit desire among individuals in treatment for a return to a normal, healthy life. Many authors trace this desire to the normalisation inherent in drug policy, and the governmentality involved in the delivery of drug treatment more directly. This article adds to these discussions by shifting focus from the experience of individuals in treatment to those out of treatment settings. In so doing, we aim to develop a more nuanced understanding of how heavy drug users negotiate power, governmentality and the modulations of health and illness in the course of everyday life. We ground our discussion in qualitative research conducted in Melbourne, Australia, with 31 current methamphetamine consumers. We argue that regular methamphetamine consumption involves a complex and ambivalent relationship with the ideas of health and normal life, imposing as well as reflecting a form of estrangement between its consumers and mainstream (or normal) society. This ambivalence has important implications for the delivery of health and social services among methamphetamine consumers, insofar as the restoration of normal health and the reintegration of former drug users into mainstream society are typical health service goals. We address some of these policy implications by way of conclusion.     

Anomalous origin of left coronary artery from the pulmonary artery with ventricular septal defect

Only two cases have been reported previously of the association of ventricular septal defect (VSD) with anomalous origin of the left coronary artery (ALCA) arising from the pulmonary artery. The purpose of this paper is to present two additional cases, to describe the pathophysiology, and to emphasize how the clinical course of this combination of defects differs from that of isolated ALCA. Patients with both of these anomalies present in infancy with manifestations only of a large left-right ventricular shunt and pulmonary hypertension. Initially the ALCA is well perfused from the high pressure in the pulmonary artery. In these instances in which the pulmonary artery pressure subsequently decreased because of spontaneous reduction in size of the VSD, the left coronary arterial system became less well perfused. Because of this decreased perfusion in association with the left ventricular myocardial stress initially caused by volume overload, myocardial ischemia and ultimately infarction occurred. Early identification and repair of the anatomic abnormality could prevent irreversible myocardial damage.     

Transseptal Left Heart Catheterization in Infants and Children

Transseptal left heart catheterization was performed in 80 infants and children with various forms of congenital heart disease. The majority had left heart obstructive lesions. Forty percent were under 5 years of age and less than 20 kg in weight. Uncomplicated cardiac perforation occurred in two patients. The technique is described in detail with emphasis on measures which increase the safety of the procedure for the patient. We conclude that this is a useful technique and in selected patients may be the preferred approach to the left heart.     

Amoxicillin for acute lower respiratory tract infection in primary care: subgroup analysis of potential high-risk groups

Background

Antibiotics are of limited overall clinical benefit for uncomplicated lower respiratory tract infection (LRTI) but there is uncertainty about their effectiveness for patients with features associated with higher levels of antibiotic prescribing.

Aim

To estimate the benefits and harms of antibiotics for acute LRTI among those producing coloured sputum, smokers, those with fever or prior comorbidities, and longer duration of prior illness.

Design and setting

Secondary analysis of a randomised controlled trial of antibiotic placebo for acute LRTI in primary care.

Method

Two thousand and sixty-one adults with acute LRTI, where pneumonia was not suspected clinically, were given amoxicillin or matching placebo. The duration of symptoms, rated moderately bad or worse (primary outcome), symptom severity on days 2–4 (0–6 scale), and the development of new or worsening symptoms were analysed in pre-specified subgroups of interest. Evidence of differential treatment effectiveness was assessed in prespecified subgroups by interaction terms.

Results

No subgroups were identified that were significantly more likely to benefit from antibiotics in terms of symptom duration or the development of new or worsening symptoms. Those with a history of significant comorbidities experienced a significantly greater reduction in symptom severity between days 2 and 4 (interaction term −0.28, P = 0.003; estimated effect of antibiotics among those with a past history −0.28 [95% confidence interval = −0.44 to −0.11], P = 0.001), equivalent to three people in 10 rating symptoms as a slight rather than a moderately bad problem. For subgroups not specified in advance antibiotics provided a modest reduction in symptom severity for non-smokers and for those with short prior illness duration (<7 days), and a modest reduction in symptom duration for those with short prior illness duration.

Conclusion

There is no clear evidence of clinically meaningful benefit from antibiotics in the studied high-risk groups of patients presenting in general practice with uncomplicated LRTIs where prescribing is highest. Any possible benefit must be balanced against the side-effects and longer-term effects on antibiotic resistance.