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The approved interferon beta-1b (Betaseron®/Betaferon®) dose is 250 μg (8 MIU) administered subcutaneously (sc) every other day (eod). Clinical trial data suggest a dose–response effect for interferon beta in multiple sclerosis (MS) treatment and a maximum dose has yet to be established. The Interferon Dose Escalation Assessment of Safety (IDEAS) study evaluated the safety and tolerability of interferon beta-1b 500 μg (16 MIU) sc eod with structured dose escalation and adverse event (AE) management in 22 patients (20 interferon beta-1b-treated (SD) and two interferon beta-1b-naïve (ND)) with relapsing-remitting (RR) MS, secondary-progressive (SP) MS, or progressive relapsing MS. IDEAS comprised an eight-week dose escalation period and a 12-week maintenance period, with modification as clinically warranted. Autoinjectors were used for all injections ≥0.4 mL. Clinical laboratory values were monitored monthly. Baseline and exit assessments included the MS Functional Composite score, EDSS, and neutralizing antibody MxA assay. AEs were recorded at every injection. Dose escalation ranged from two to 12 weeks. Some 91% of patients (20/22) achieved the 500-μg dose, and of these 90% (18/20) completed the maintenance phase. There were no differences in response between ND and SD patients. Most common AEs were decreased general well-being, insomnia, and injection site reactions (mostly mild). The 500-mg dose of interferon beta-1b was well tolerated in the short-term with escalation and premedication in these patients, most of whom had previously been receiving 250 μg interferon beta-1b. [ABSTRACT FROM AUTHOR]

The Interferon Dose Escalation Assessment of Safety extension trial monitored neutralizing antibodies to interferon beta-1b in patients who currently or had previously received the double dose (500 μg) for up to 28 months. Fifteen patients entered the extension trial; five patients were neutralizing antibody-positive at the start of the trial. The present study demonstrates that when neutralizing antibodies develop in patients receiving higher doses of interferon beta-1b they tend to persist for a prolonged period, although neutralizing antibody titers tend to decrease over time and some patients may revert to neutralizing antibody-negative status. [ABSTRACT FROM AUTHOR]

Background: Given that HIV-protease inhibitors (HIV-PIs) are substrates/inhibitors of the multidrug transporter ABCB1, can induce ABCB1 expression, and are used in combination with doxorubicin for AIDS-Kaposi's Sarcoma (KS) treatment, the role that ABCB1 plays in mediating multidrug resistance (MDR) in a fully transformed KS cell line (SLK) was explored.Methods: The KS cells were exposed to both acute and chronic treatments of physiological concentrations of different HIV-PIs (indinavir, nelfinavir, atazanavir, ritonavir, or lopinavir), alone or together with doxorubicin. The ABCB1 mRNA and protein expression levels were then assessed by qRT-PCR and western blotting, flow cytometry, and immunofluorescence.Results: Chronic treatment of SLK cells with one of the five HIV-PIs alone or together resulted in increased resistance to doxorubicin. Co-treatment with one of the HIV-PIs in combination with doxorubicin resulted in a synergistic increase in resistance to doxorubicin, and the degree of resistance was found to correlate with the expression of ABCB1. The SLK cells were also revealed to be cross-resistant to the structurally unrelated drug paclitaxel.Conclusion: These studies suggest that ABCB1 is primarily responsible for mediating MDR in SLK cells selected with either HIV-PIs alone or in combination with doxorubicin. Therefore, the roles that ABCB1 and drug cocktails play in mediating MDR in KS in vivo should be evaluated. [ABSTRACT FROM AUTHOR]

The project was a master's level graduate program that prepares occupational therapists, physical therapists, speech and language pathologists, nurses and special educators for early intervention practice. The purpose of this descriptive report is to explain the conceptual framework for the graduate program and to identify themes in students' learning from the portion of the program focused on professional-family relationships-the family practicum. In the family practicum, each student spent 50 hours over 6 months with one family who had a young child with significant disabilities. The students wrote reflective journals during the practicum that revealed how they interpreted their experiences for application to their own practices. Through qualitative analysis of the students' journals, four themes emerged: (1) Acknowledge that parenting a child with a disability is a 24/7 job; (2) Recognize that internal and external resources are essential to family adaptation; (3) Respect parents as the experts on their child; and (4) Accept the family's values. These themes were validated by the families' evaluation of the practicum and provide evidence that students grew in their appreciation of and competence in relationship-centered early intervention. [ABSTRACT FROM AUTHOR]

Cyclosporin A (CsA) is an immunosuppressive drug commonly given to transplant patients. Its application is accompanied by severe side effects related to calcium, among them hypertension and nephrotoxicity. The Na+/Ca2+ exchanger (NCX) is a major calcium regulator expressed in the surface membrane of all excitable and many nonexcitable tissues. Three genes, NCX1, NCX2, and NCX3 code for Na+/Ca2+ exchange activity. NCX1 gene products are the most abundant. We have shown previously that exposure of NCX1-transfected HEK 293 cells to CsA, leads to concentration-dependent reduction of Na+/Ca2+ exchange activity and surface expression, without a reduction in total cell-expressed NCX1 protein. We show now that the effect of CsA on NCX1 protein expression is not restricted to transfected cells overexpressing the NCX1 protein but exhibited also in cells expressing endogenously the NCX1 protein (L6, H9c2, and primary smooth muscle cells). Exposure of NCX2- and NCX3-transfected cells to CsA results also in reduction of Na+/Ca2+ exchange activity and surface expression, though the sensitivity to the drug was lower than in NCX1-transfected cells. Studying the molecular mechanism of CsA–NCX interaction suggests that cyclophilin (Cyp) is involved in NCX1 protein expression and its modulation by CsA. Deletion of 426 amino acids from the large cytoplasmic loop of the protein retains the CsA-dependent downregulation of the truncated NCX1 suggesting that CsA–Cyp–NCX interaction involves the remaining protein domains. [ABSTRACT FROM AUTHOR]

SV40 vectors packaged in vitro (pseudovirions) are an efficient delivery system for plasmids up to 17.7 kb, with or without SV40 sequences. A truncated Pseudomonas exotoxin gene (PE38) was delivered into various human cells (HeLa, KB-3-1, human lymphoblastoids, and erythroleukemia cells), in vitro using pseudovirions. The number of viable cells was reduced significantly in the PE38-transduced cells. Human KB adenocarcinomas growing in mice were treated with intratumoral injection of PE38 packaged in vitro, and tumor size decreased significantly. Intraperitoneal treatments were as effective in reducing tumor size as intratumoral treatments. To check the viability of mock- or PE38-treated mice, every 4 days they were weighed, their blood was tested, and various tissues were screened for pathology. All parameters showed that the in vitro-packaged vectors, injected into tumors or intraperitoneally, caused no abnormalities in mice. The combined treatment of doxorubicin with in vitro-packaged PE38 reduced tumor size slightly more than each of the treatments separately. However, the combined treatment did not cause the weight loss seen with doxorubicin alone. These results indicate that SV40 in vitro packaging is an effective system for cancer gene delivery using two different routes of injection and in combination with chemotherapy.Cancer Gene Therapy (2006) 13, 648–657. doi:10.1038/sj.cgt.7700943; published online 24 February 2006 [ABSTRACT FROM AUTHOR]

Abstract: The Jezre’el Valley, Israel has inherent drainage problems; high water tables are common. Intensive irrigation has led to the development of salinity problems. A number of drainage system designs were installed in the same field to manage the prevailing causes of the salinity hazards. The shallow drainage system was designed to prevent damage to the winter crops and especially to enable early seeding of summer crops. The deep drainage system was designed to maintain a relatively low water table, thereby avoiding concentration of salts in the root zone. The combined drainage system with relief wells was designed to relief upward hydraulic pressure exerted by the semi-confined shallow aquifer. To study the mechanisms of salinization and the nature of the water table regime, soil salinity and water table monitoring systems were installed perpendicular to the parallel subsurface drains. The water table monitoring systems consisted of batteries of piezometers installed at different depths and located between the drainage lines. The most efficient method of drainage was found to be a combination of shallow and deep drains with relief wells wherever the semi-confined shallow aquifer was present. A noticeable improvement of the soil salinity, to below 4dS/m, was observed 2–3 years after the drainage system was installed. It was found that the water table should be more than 1m below the soil surface, in the spring season, to prevent soil salinization. [Copyright &y& Elsevier]

To explore the relationship between the ingestion of Agouti paca (AP) and human leptospirosis in Guyana, 19 febrile men who said they had hunted and eaten A. paca were screened for malaria, using bloodsmears, and for leptospirosis, using an enzyme immuno-assay that detects Leptospira-specific IgM. Those found positive for anti-Leptospira IgM were then evaluated further, with a microscopical agglutination test based on a limited panel of serovars from three pathogenic species of Leptospira. Although six of die 18 patients who provided suitable samples for the serology were found seropositive for acute leptospirosis, only three of the 19 patients were found smear-positive for malaria, A clinical-decision model, based on medical histories, the results of physical examinations, and the use of routine urine dipsticks, and enabling prediction of the serological results, was developed. This model, which had 83% sensitivity and 100% specificity for leptospirosis, indicated that, in the absence of serology, most febrile patients reporting AP ingestion could be correctly treated if each was checked for malaria using traditional bloodsmears. The smear-positives should be treated with antimalarial drugs whereas the smear-negatives should be treated for leptospirosis if they had any of the following: a skin rash; lymphadenopathy; abnormal urine sediment (proteinuria or haematuria); and/or no previous history of malaria. In the present study, the relative risk of leptospirosis among the patients who were smear-negative for malaria and fulfilled at least one of these four criteria was 13 (P=0.0007). In Guyana at least, leptospirosis appears to be common among men who hunt, prepare and ingest AP. Vaccines may be the best, practical form of protection among such men. [ABSTRACT FROM AUTHOR]

Reduced accumulation of cisplatin is the most consistent feature seen in cisplatin-resistant (CP-r) cells that are cross-resistant to other cytotoxic compounds, such as methotrexate. In this report, defective uptake of a broad range of compounds, including [(14)C]-carboplatin, [(3)H]MTX, [(3)H]folic acid (FA), [(125)I]epidermal growth factor, (59)Fe, [(3)H]glucose, and [(3)H]proline, as well as (73)As(5+) and (73)As(3+), was detected in CP-r human hepatoma and epidermal carcinoma cells that we have previously shown are defective in fluid-phase endocytosis. Downregulation of several small GTPases, such as rab5, rac1, and rhoA, which regulate endocytosis, was found in CP-r cells. However, expression of an early endosomal protein and clathrin heavy chain was not changed, suggesting that the defective endocytic pathway is clathrin independent. Reduced expression of the cell surface protein, folate-binding protein (FBP), which is a carrier for the uptake of MTX, was also observed in the CP-r cells by confocal immunofluorescence microscopy and Real-Time PCR. Reactivation of the silenced FBP gene in the CP-r cells by a DNA demethylation agent, 2-deoxy-5-aza-cytidine (DAC) demonstrates that hypermethylation occurred in the CP-r cells. The uptake of [(14)C]carboplatin, [(3)H]FA, and [(3)H]MTX increased in an early stage CP-r cell line (KB-CP1) after treatment with DAC. Both a defective endocytic pathway and DNA hypermethylation resulting in the downregulation of small regulatory GTPases and cell surface receptors contribute to the reduced accumulation of a broad range of compounds in CP-r cells. [ABSTRACT FROM AUTHOR]

Transthyretin is the major thyroxine-binding protein in the plasma of rodents, and the main thyroxine-binding protein in the cerebrospinal fluid of both rodents and humans. The choroid plexus synthesizes transthyretin and secretes it to the cerebrospinal fluid. Although it was suggested that transthyretin might play an important role in mediating thyroxine transfer from the blood into the brain across the choroid plexus–cerebrospinal fluid barrier, newer findings question this hypothesis. Because thyroid hormone passage across brain barriers is a precondition for its action in the CNS, and because brain is an important target of thyroid hormone action, we investigated the role of transthyretin in mediating thyroid hormone access to and distribution within the brain in a transthyretin-null mouse model system. In this report we describe the results derived from use of film autoradiography, a technique that yields definitive morphological results. Film autoradiograms were prepared at 3 and 19 h after intravenous injection of either high specific activity [125I]thyroxine or [125I]triiodothyronine. Image analyses were designed to demonstrate regional changes in hormone distribution, and to highlight alterations in iodothyronine delivery from ventricles to brain parenchyma. We find no qualitative or quantitative differences in these parameters between the transthyretin-null and the wild-type mouse brain after either [125I]thyroxine or [125I]triiodothyronine administration.The data presented here now provide definitive evidence that, under standard laboratory conditions, transthyretin is not required for thyroid hormone access to or distribution within the mouse brain. This study also provides the first map of iodothyronine distribution in the brain of the mouse. [Copyright &y& Elsevier]

The MDR1 (multidrug resistance) gene, transferred to hematopoietic cells, is expected to protect them from anticancer chemotherapy and may serve as a selectable marker, restoring gene expression in vivo. Appropriate selection strategies, however, need to be established. To investigate whether preselection ex vivo affects chemoresistance, murine bone marrow cells were retrovirally transduced with high-titer or, as a model for suboptimal gene expression, Iow-titer retroviruses and exposed to daunomycin or colchicine for 48-96 h. Selection significantly increased chemoresistance of clonogenic progenitor cells. In tissue culture, the entire target population was rendered highly drug resistant after MDR1 transfer with high-titer viruses. If transduction was performed under suboptimal conditions, drug selection increased the frequency of chemoresistant colonies up to 40% over the number of unselected cells. Colchicine and daunomycin were equally efficient in increasing drug resistance ex vivo, but colchicine-preselected cells rescued lethally irradiated mice under conditions where daunomycinselected bone marrow cells failed to do so. Hence, while hematopoietic cells can be protected by MDR1, the selection strategy is critical for repopulation of bone marrow with transduced cells. Preselection in culture before transplantation significantly increased P-gp expression and chemoresistance in vivo in mice reconstituted with transduced bone marrow cells. This study may help to facilitate the use of MDR1 as a selectable marker in gene therapy of the hematopoietic system. [ABSTRACT FROM AUTHOR]

X-linked severe combined immunodeficiency (XSCID) is a hereditary disorder characterized by severe T cell lymphopenia and abnormal B cell function. The disease is caused by mutations in IL2RG, the gene encoding the interleukin-2 receptor common γ chain (γc) shared by several interleukin receptors. A Harvey retroviral bicistronic vector containing an IL2RG cDNA and cDNA encoding the multidrug transporter (MDR1) was constructed to investigate the correction of XSCID. Translation of the MDR1 cDNA is achieved from an internal ribosome entry site (IRES). Mouse fibroblasts transfected or transduced with the vector expressed both membrane proteins as detected with specific monoclonal antibodies by fluorescence activated cell sorting. Two human XSCID B cell lines were transduced using a filter concentration method in combination with phosphate depletion. Significant expression of both proteins was detected by Western blot analysis. This construct might be particularly useful if high expression of γc is required, as might be achievable through in vivo selection for drug resistance of recipient lymphocytes. [ABSTRACT FROM AUTHOR]

The development of simultaneous resistance to multiple structurally unrelated drugs is a major impediment to cancer chemotherapy. Multidrug resistance in human KB carcinoma cells selected in colchicine, vinblastine, or Adriamycin is associated with amplification of specific DNA sequences (the multidrug resistance locus, mdrl). During colchicine selection resistance is initially accompanied by elevated expression of a 4.5-kilobase mdrl messenger RNA (mRNA) without amplification of the corresponding genomic sequences. During selection for inceased levels of resistance, expression of this mRNA is increased simultaneously with amplification of mdrl DNA. Increased expression and amplification of mdrl sequences were also found-in multidrug-resistant sublines of human leukemia and ovarian carcinoma cells. These results suggest that increased expression ofmdrl mRNA is a common mechanism for multidrug resistance in human cells. Activation of the mdrl gene by mutations or epigenetic changes may precede its amplification during the development of resistance. [ABSTRACT FROM AUTHOR]