Understanding reasons for asthma outpatient (non)-attendance and exploring the role of telephone and e-consulting in facilitating access to care: exploratory qualitative study.

OBJECTIVE: To understand factors influencing patients' decisions to attend for outpatient follow up consultations for asthma and to explore patients' attitudes to telephone and email consultations in facilitating access to asthma care. DESIGN: Exploratory qualitative study using in depth interviews. SETTING: Hospital outpatient clinic in West London. PARTICIPANTS: Nineteen patients with moderate to severe asthma (12 "attenders" and 7 "non-attenders"). RESULTS: Patients' main reasons for attending were the wish to improve control over asthma symptoms and a concern not to jeopardise the valued relationship with their doctor. Memory lapses, poor health, and disillusionment with the structure of outpatient care were important factors implicated in non-attendance. The patients were generally sceptical about the suggestion that greater opportunity for telephone consulting might improve access to care. They expressed concerns about the difficulties in effectively communicating through non-face to face media and were worried that clinicians would not be in a position to perform an adequate physical examination over the telephone. Email and text messaging were viewed as potentially useful for sending appointment reminders and sharing clinical information but were not considered to be acceptable alternatives to the face to face clinic encounter. CONCLUSIONS: Memory lapses, impaired mobility due to poor health, and frustration with outpatient clinic organisation resulting in long waiting times and discontinuity of care are factors that deter patients from attending for hospital asthma assessments. The idea of telephone review assessments was viewed with scepticism by most study subjects. Particular attention should be given to explaining to patients the benefits of telephone consultations, and to seeking their views as to whether they would like to try them out before replacing face to face consultations with them. Email and text messaging may have a role in issuing reminders about imminent appointments.

Ectomycorrhizal root development in wet Alder carr forests in response to desiccation and eutrophication.

Effects of desiccation and eutrophication on ectomycorrhizal (ECM) root development in wet Alder carr forests in The Netherlands were studied. In northwestern Europe, wet Alder carr forests are found mostly in peatlands and along streams, forming an important component of wetland ecosystems. The dominant tree species in wet Alder carr forests is Alnus glutinosa (L.) Gaertn. (Black alder), which associates with ectomycorrhizal fungi. During recent decades, wet Alder carr forests in Europe have declined because of desiccation and eutrophication, particularly in The Netherlands. In the present study, the number of root tips of A. glutinosa trees was highest in an undisturbed wet Alder carr forest in a peatland area. Eutrophication in the peatland area significantly inhibited ectomycorrhizal (ECM) root development of A. glutinosa. In the eutrophied forest, ECM root tips were observed only close to A. glutinosa trees growing on hummocks. The concentrations of nitrate and potassium in soil water of the eutrophied forest were significantly higher than in the undisturbed forest, while magnesium and iron concentrations and the pH were significantly lower. The number of ECM root tips of A. glutinosa in a desiccated forest along a stream was generally lower than in an undisturbed wet Alder carr forest on waterlogged soil in the same area. The sulphate concentration in soil water in the desiccated forest was significantly higher than in the forest on waterlogged soil. ECM root development of A. glutinosa may have been negatively affected by the chemical composition of the soil water.

Oncology thinking cap: scaffolded use of a simulation to learn clinical trial design.

BACKGROUND: Physicians often are called on to participate in and interpret clinical trials, but their training in this area may not provide them with the inquiry skills that are needed. Simulations have the potential to be a promising tool for helping medical students learn the skills involved in clinical trial design. However, simulations may be complex and require additional scaffolding to support learning. DESCRIPTION: The goal of this study was to teach aspects of cancer clinical trial design through the scaffolded use of a simulation, the Oncology Thinking Cap. The software-based scaffolding provided guidance in designing the trial. Subsequently, the simulation allowed students to run the designed trial, which produces detailed patient histories. This feedback then could be used to redesign the trial. EVALUATION: Twenty-four 4th-year medical students were asked to design a clinical trial in advance, on paper, to test a new anticancer drug. Student groups then designed and simulated running the clinical trial assisted by the software environment. Instructional effectiveness was measured using a pretest-posttest design that included having students (a) write a group research proposal and (b) individually critique a flawed proposal. At the group level (N = 6 groups), students demonstrated a 34% increase in the number of elements of a clinical trial that they included in their research proposals. At the individual level (N = 24), students improved by 48% in their critiques of flawed proposals. CONCLUSIONS: Scaffolding embedded in the simulator is a promising approach to helping students learn about clinical trial design.

Primary non-Hodgkin's lymphoma of bone.

Primary non-Hodgkin's lymphoma of bone (PLB) constitutes approximately 5% of all extranodal non-Hodgkin's lymphoma (NHL) and 7% of primary bone tumors. The peak incidence for PLB is in the fifth decade, with a slight preponderance of males over females. The presenting symptoms usually consist of localized bone pain and occasionally a palpable mass. Most patients with PLB have B-cell tumors with a diffuse mixed-cell or diffuse large cell histology. While most patients present with early-stage disease, it is not clear whether such patients benefit from combined-modality therapy (CMT) consisting of radiation therapy (RT) and chemotherapy (CT) compared with either RT or CT alone. However, there is strong evidence that CMT is beneficial in the treatment of localized NHL, and these results might be applicable to the therapy for PLB. Nevertheless, only a phase III randomized, controlled clinical trial will determine whether CMT is superior to either CT or RT alone.

Autologous human monocyte-derived dendritic cells genetically modified to express melanoma antigens elicit primary cytotoxic T cell responses in vitro: enhancement by cotransfection of genes encoding the Th1-biasing cytokines IL-12 and IFN-alpha.

DNA-based immunization strategies designed to elicit cellular antitumor immunity offer an attractive alternative to protein- or peptide-based approaches. In the present study we have evaluated the feasibility of DNA vaccination for the induction of CTL reactivity to five different melanoma Ags in vitro. Cultured, monocyte-derived dendritic cells (DC) were transiently transfected with plasmid DNA encoding human MART-1/Melan-A, pMel-17/gp100, tyrosinase, MAGE-1, or MAGE-3 by particle bombardment and used to stimulate autologous PBMC responder T cells. CTL reactivity to these previously identified melanoma Ags was reproducibly generated after two or three stimulations with genetically modified DC. Co-ordinate transfection of two melanoma Ag cDNAs into DC promoted CTL responders capable of recognizing epitopes from both gene products. Coinsertion of genes encoding the Th1-biasing cytokines IL-12 or IFN-alpha consistently enhanced the magnitude of the resulting Ag-specific CTL reactivity. Importantly, DC transfected with a single melanoma Ag cDNA were capable of stimulating Ag-specific CTL reactivity restricted by multiple host MHC alleles, some of which had not been previously identified. These results support the inherent strengths of gene-based vaccine approaches that do not require prior knowledge of responder MHC haplotypes or of relevant MHC-restricted peptide epitopes. Given previous observations of in situ tumor HLA allele-loss variants, DC gene vaccine strategies may elicit a greater diversity of host therapeutic immunity, thereby enhancing the clinical utility and success of such approaches.

Potentiation of E7 antisense RNA-induced antitumor immunity by co-delivery of IL-12 gene in HPV16 DNA-positive mouse tumor.

Down-regulation of oncogene expression by antisense-based gene therapy has been extensively studied, and in some cases, therapeutic effects have been demonstrated. We have previously shown that down-regulation of HPV16 E6 and E7 gene expression inhibited HPV DNA-positive C3 mouse tumor growth. Although not all of the tumor cells were transfected by pU6E7AS plasmid, complete tumor regression was achieved if the tumor size was small at the start of therapy in a syngeneic host. This suggests that some other antitumor mechanisms may be involved in addition to the direct down-regulation of HPV16 E7 oncogene expression by the antisense effect of E7AS. In the current study, we demonstrated that E7AS induces tumor cell apoptosis. More importantly, a strong antitumor immune response was elicited in the pU6E7AS-treated and tumor-regressed mice. There was no tumor growth after rechallenging the tumor-regressed mice with 1 million C3 cells. This E7AS-induced antitumor immune response was augmented by co-delivery of mIL-12 cytokine gene. The combination therapy strategy resulted in complete regression of 26 of 28 (93%) tumors. Only 12 of 31 (38%) tumors from the group treated with pU6E7AS alone and 14 of 28 (50%) tumors from the group treated with pCMVmIL-12 alone had completely regressed. Complete regression was also demonstrated in tumors located 1 cm from the treated tumors, which indicates that a systemic antitumor effect was induced by E7AS and mIL-12. Immunohistochemistry demonstrated that a significant amount of CD4+ and CD8+ cells infiltrated into tumors treated with pU6E7AS, pCMVmIL-12 and pU6E7AS+pCMVmIL-12. These data indicate that host immunity is an important factor for antisense-based gene therapy approach which can be further enhanced by combination with cytokine gene therapy.

Interferon-alpha gene therapy for cancer: retroviral transduction of fibroblasts and particle-mediated transfection of tumor cells are both effective strategies for gene delivery in murine tumor models.

Stable transfection of tumor cells with IFN-alpha genes has been shown to result in abrogation of tumor establishment and induction of antitumor immunity. However, strategies suitable for the clinical application of IFN-alpha gene therapy for cancer have not been reported. In this study, we investigated two gene delivery systems capable of mediating the local paracrine production of high levels of biologically active IFN-alpha in murine tumor models: retroviral transduction of fibroblasts and particle-mediated transfection of tumor cells. In spite of the antiproliferative effects of IFN-alpha, it was possible to obtain stable retroviral producer cell lines and transduce a variety of murine tumor cells including syngeneic fibroblasts to stably secrete 2000-5000 U (40-100 ng) murine IFN-alpha/10(6) cells/24 h. IFN-alpha transduction of tumor cells abrogated tumorigenicity in establishment models and induced antitumor immunity in several murine tumor model systems. Importantly, IFN-alpha gene delivery using retrovirally transduced syngeneic fibroblasts was capable of suppressing the establishment of the poorly immunogenic TS/A mouse mammary adenocarcinoma and induced antitumor immunity. Particle mediated transient transfection of tumor cells using the gene gun led to the production of up to 20,000 U IFN-alpha/10(6) cells during the first 24 h and proved to be equally effective in suppressing establishment of TS/A adenocarcinoma and inducing antitumor immunity. These results suggest that retroviral transduction of autologous fibroblasts can serve as an effective gene delivery method for IFN-alpha gene therapy of cancer. Particle-mediated transfection of freshly isolated tumor cells may represent a clinically attractive alternative approach for nonviral gene delivery. Both strategies circumvent the difficulties in routinely establishing primary tumor cell lines from the vast majority of human cancers.

Analysis of a hot spot for DNA insertion suggests a mechanism for Ig switch recombination.

We recently reported that transfected DNA inserts into the VDJ-Cmu intron much more frequently than into average DNA, and that insertion within this intron occurs preferentially into the switch region. To gain information about the mechanisms involved in DNA insertion, we sequenced the 5' and 3' junctions of typical transformants. Although the junction sequences did not indicate a preferred insertion motif within the switch region, our results suggest that joining of the transfected and chromosomal DNAs is facilitated by short regions of identity. Our analysis of the insertions into the non-switch part of the intron suggests that breakage of the chromosomal DNA occurs preferentially at sites that are flanked by short complementary sequences. This correlation suggests that the self-complementary DNA might form short stem-loops, which, in turn, are prone to enzymatic cleavage and thus facilitate the insertion of transfected DNA. A model is proposed in which this effect can account for both the higher than average frequency of insertion into the VDJ-Cmu intron and the preference for the switch region within this intron. An extension of this model is proposed to explain why the repetitive switch regions are the preferred breakage/rejoining sites for isotype switch rearrangements.

The Ig heavy chain switch region is a hotspot for insertion of transfected DNA.

The Ig heavy chain class switch usually occurs by breaking and rejoining DNA in the switch (S) regions, which consist of tandemly repeated sequences 5' of the constant region exons. Various studies have suggested that S DNA can also recombine with non-S sequences. To measure the frequency of such recombination events, the hybridoma cell line igm692, a deletion mutant that lacks the C mu 1 and C mu 2 exons and the 3' end of the S mu region, was transfected with a fragment bearing the C mu 1-2 exons, but no S mu DNA. Insertion of this fragment into the residual VDJ-C mu intron of igm692 can restore a functional mu gene, yielding a transformant that is detected as a plaque-forming cell (PFC). PFC comprise approximately 8 x 10(-7) of the surviving transfected cells. In 10 of 12 PFCs, the C mu 1-2 fragment inserted into the 2.5-kb residual S mu region, whereas insertion in two cases occurred in the 3.5-kb segment 5' of S mu. Using a PCR assay to measure the frequency of insertion of the transferred fragment elsewhere in the hybridoma genome, we found that approximately 9% of the surviving transfected cells had stably acquired the C mu 1-2 fragment. These results indicate that the S mu region is approximately 100-fold more recombinogenic than the average genomic site, and approximately 7-fold more recombinogenic than the non-S mu segment of the residual VDJ-C mu, i.e., the S mu region is a hotspot for insertion of transfected DNA.

Primary non-Hodgkin's lymphoma of bone. A clinicopathologic study.

BACKGROUND: This study relates our experience in the diagnosis and treatment of a rare clinical entity, non-Hodgkin's primary lymphoma of bone. METHODS: Seventeen cases of patients with primary lymphoma of bone diagnosed and treated at a single institution between 1975 and 1992 are reviewed. Ten patients received combined-modality therapy, consisting of an anthracycline-containing combination chemotherapy (CT) regimen, followed by adjuvant radiotherapy (RT) to the primary site of disease. Five patients were treated with CT alone; one patient received RT alone; and one patient was treated with CT after emergency RT for spinal cord compression. RESULTS: Thirteen patients presented with Stage I disease, two with Stage II; and two with Stage IV disease (multiple bony sites only). Thirteen patients had an intermediate-grade diffuse large cell lymphoma; two had an intermediate-grade mixed small and large cell lymphoma; and two had a high-grade lymphoma (one immunoblastic and one small non-cleaved cell lymphoma). The overall response rate was 94% (18% complete response, 58% partial response 1, and 18% partial response 2). Thirteen patients are alive and disease-free at a median of 29 months; 10 of these received CT+RT, and 3 received CT alone. Three patients have died; one of these received CT+RT and one CT alone, and one relapsed immediately after CT. One patient, who was initially treated with RT and then with CT+RT after relapse, was lost to follow-up 40 months from the start of treatment. CONCLUSIONS: Because experience in the literature suggests a 50% distant relapse rate in primary lymphoma of bone treated with RT alone, our policy is to treat all patients with combined-modality therapy (CT+RT). However, only a Phase III randomized, controlled clinical trial will determine whether CT+RT is superior to either modality alone.

Organic mood disorder, manic type, associated with hyponatremia: a case report.

OBJECTIVE: Mania and other mood changes are known to be associated with various organic etiologies. This article documents a case of organic mania associated with hyponatremia, a phenomenon which has not been previously reported, and reviews the pertinent medical literature. METHOD: The mental status of a patient with end stage renal disease and no previous psychiatric history who presented with mania and hyponatremia was carefully monitored while her serum sodium concentration was corrected. RESULTS: The manic symptoms of the patient presented resolved completely with correction of serum sodium concentration alone. CONCLUSIONS: The acute episode of hyponatremia experienced by this patient with chronic, end stage renal disease appears to have precipitated the symptoms of mania. This observation may shed some additional light on the pathophysiology of organic mania.

Automated enzyme immunoassay for lutropin with the Abbott IMx analyzer.

An automated enzyme immunoassay for human lutropin for use with the Abbott IMx analyzer is described. The assay provides results in approximately 40 min with a sensitivity of 0.25 int. units of LH per liter for up to 23 serum or plasma samples. Cross-reactivity with follitropin (2000 int. units/L) and thyrotropin (2 int. units/L) was negligible; it was 0.016% with human choriogonadotropin (1 X 10(6) int. units/L). There was no interference by high concentrations of bilirubin (0.5 g/L), hemoglobin (7.50 g/L), or triglycerides (13.5 g/L). Intra-, inter-, and total assay CVs were less than or equal to 3.75%, less than or equal to 7.1%, and less than or equal to 7.94%, respectively. Values obtained with the IMx correlated well (r = 0.98, n = 194) with values obtained with Diagnostic Products' LH Double Antibody RIA, and Serono's LH MAIAclone assay. This assay should be useful for small to medium-size laboratories involved in the clinical diagnosis of reproductive pathology.

Analyzing the same data in two ways: a demonstration model to illustrate the reporting and misreporting of clinical trials.

The methods used to analyze and interpret clinical trials of chemotherapy may have a major impact on the conclusions that are drawn in papers reporting them. To illustrate this problem, we constructed a hypothetical clinical trial in which patients with metastatic cancer were treated with chemotherapy. The following two articles provide reports of this trial, analyzed by methods that we would interpret as being of low and high quality, respectively. In the present report, we describe briefly the methodological differences that led to the opposite conclusions based on this single set of data. The errors of reporting and omissions of the first article (A) are similar to those that have been extracted from recent issues of the Journal of Clinical Oncology and other leading cancer journals, although they have not all appeared within a single report. This demonstration model illustrates problems in the reporting of clinical trials and suggests guidelines for improved reporting.

Isolation and structural mapping of a human c-src gene homologous to the transforming gene (v-src) of Rous sarcoma virus.

We have utilized a lambda Charon 4A human genomic library to isolate recombinant clones harboring a highly conserved c-src locus containing nucleotide sequences homologous to the transforming gene of Rous sarcoma virus (v-src). Four overlapping clones spanning 24 kilobases of cellular DNA were analyzed by restriction endonuclease mapping. Human c-src sequences homologous to the entire v-src region are present in a 20-kilobase region that contains 11 exons as determined by restriction mapping studies utilizing hybridization to labeled DNA probes representing various subregions of the v-src gene and by preliminary DNA sequencing analyses. A considerable degree of similarity exists between the organization of the human c-src gene and that of the corresponding chicken c-src gene with respect to exon size and number. However, the human c-src locus is larger than the corresponding chicken c-src locus, because many human c-src introns are larger than those of chicken c-src. alu family repetitive sequences are present within several human c-src introns. This locus represents a highly conserved human c-src locus that is detectable in human cellular DNAs from various sources including placenta, HeLa cells, and WI-38 cells.