135 research outputs found

    Studies on the Cell Ni|Ni-Soap(s), K-Soap, Cu-Soap(s)|Cu

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    60-6

    Interaction of Cationic Surfactants with Polytungstate

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    346-34

    Effects of a second intron on recombinant MFG retroviral vector

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     The retroviral vectors based on an MFG-type backbone have superior expression characteristics, in part, due to the presence of the retroviral chimeric intron (MFG intron). We tested the hypothesis that inclusion of a second intron in MFG vectors may influence packaging and/or LTR-driven transgene expression. We constructed two MFG retroviral vectors, MFG/hFIXc and MFG/hFIXm2, containing human factor IX (hFIX) cDNA without and with a 0.3-kb hFIX intron, respectively. When tested with primary mouse myoblasts or HepG2 cells in culture for transient expression activity, pMFG/hFIXm2 plasmid produced two-to-three fold higher hFIX than pMFG/hFIXc. These vectors produced equivalent retroviral titers from packaging cells. In transduced cells, the splicing of the MFG intron in the retroviral transcripts occured at a similar efficiency; however, MFG/hFIXc virus gave two-fold higher hFIX expression than that of the MFG/hFIXm2 viral infection. Analyses of MFG/hFIXm2 virion RNA and transduced cell genomic DNA suggested that, although the hFIX intron containing viral RNA are packaged, these viruses fail to integrate their transgenes into the genome of transduced cells, suggesting a block at the reverse transcription and/or integration steps. Similar results were also obtained with the prototype vectors, LIXcSN and LIXm2SN, lacking the MFG intron. Together, these results suggest that a hFIX cDNA sequence in the retroviral vectors performs better over hFIX intron-containing minigene.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/42460/1/705-146-3-601_11460601.pd

    Bone marrow stromal cells as a genetic platformfor systemic delivery of therapeutic proteins in vivo : human factor IX model

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    Background Hemophilia B is an X-linked bleeding disorder that results from a deficiency in functional coagulation factor IX (hFIX). In patients lacking FIX, the intrinsic coagulation pathway is disrupted leading to a lifelong, debilitating and sometimes fatal disease. Methods We have developed an ex vivo gene therapy system using genetically modified bone marrow stromal cells (BMSCs) as a platform for sustained delivery of therapeutic proteins into the general circulation. This model exploits the ability of BMSCs to form localized ectopic ossicles when transplanted in vivo . BMSCs were transduced with MFG-hFIX, a retroviral construct directing the expression of hFIX. The biological activity of hFIX expressed by these cells was assessed in vitro and in vivo . Results Transduced cells produced biologically active hFIX in vitro with a specific activity of 90% and expressed hFIX at levels of ∼497 ng/10 6 cells/24 h and 322 ng/10 6 cells/24 h for human and porcine cells, respectively. The secretion of hFIX was confirmed by Western blot analysis of the conditioned medium using a hFIX-specific antibody. Transduced BMSCs (8 × 10 6 cells per animal) were transplanted within scaffolds into subcutaneous sites in immunocompromised mice. At 1 week post-implantation, serum samples contained hFIX at levels greater than 25 ng/ml. Circulating levels of hFIX gradually decreased to 11.5 ng/ml at 1 month post-implantation and declined to a stable level at 6.1 ng/ml at 4 months. Conclusions These findings demonstrate that genetically modified BMSCs can continuously secrete biologically active hFIX from self-contained ectopic ossicles in vivo , and thus represent a novel delivery system for releasing therapeutic proteins into the circulation. Copyright © 2002 John Wiley & Sons, Ltd.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/35232/1/292_ftp.pd

    5-Androstene-3β,7β,17β-triol (β-AET) Slows Thermal Injury Induced Osteopenia in Mice: Relation to Aging and Osteoporosis

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    5-androstene-3β,7β,17β-triol (β-AET), an active metabolite of dehydroepiandrosterone (DHEA), reversed glucocorticoid (GC)-induced suppression of IL-6, IL-8 and osteoprotegerin production by human osteoblast-like MG-63 cells and promoted osteoblast differentiation of human mesenchymal stem cells (MSCs). In a murine thermal injury model that includes glucocorticoid-induced osteopenia, β-AET significantly (p<0.05) preserved bone mineral content, restored whole body bone mineral content and endochondral growth, suggesting reversal of GC-mediated decreases in chondrocyte proliferation, maturation and osteogenesis in the growth plate. In men and women, levels of β-AET decline with age, consistent with a role for β-AET relevant to diseases associated with aging. β-AET, related compounds or synthetic derivatives may be part of effective therapeutic strategies to accelerate tissue regeneration and prevent or treat diseases associated with aging such as osteoporosis

    'I am on treatment since 5 months but I have not received any money': coverage, delays and implementation challenges of 'Direct Benefit Transfer' for tuberculosis patients - a mixed-methods study from South India.

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    Background: In March 2018, the Government of India launched a direct benefit transfer (DBT) scheme to provide nutritional support for all tuberculosis (TB) patients in line with END TB strategy. Here, the money (@INR 500 [~8 USD] per month) is deposited electronically into the bank accounts of beneficiaries. To avail the benefit, patients are to be notified in NIKSHAY (web-based notification portal of India's national TB programme) and provide bank account details. Once these details are entered into NIKSHAY, checked and approved by the TB programme officials, it is sent to the public financial management system (PFMS) portal for further processing and payment. Objectives: To assess the coverage and implementation barriers of DBT among TB patients notified during April-June 2018 and residing in Dakshina Kannada, a district in South India. Methods: This was a convergent mixed-methods study involving cohort analysis of patient data from NIKSHAY and thematic analysis of in-depth interviews of providers and patients. Results: Of 417 patients, 208 (49.9%) received approvals for payment by PFMS and 119 (28.7%) got paid by 1 December 2018 (censor date). Reasons for not receiving DBT included (i) not having a bank account especially among migrant labourers in urban areas, (ii) refusal to avail DBT by rich patients and those with confidentiality concerns, (iii) lack of knowledge and (iv) perception that money was too little to meet the needs. The median (IQR) delay from diagnosis to payment was 101 (67-173) days. Delays were related to the complexity of processes requiring multiple layers of approval and paper-based documentation which overburdened the staff, bulk processing once-a-month and technological challenges (poor connectivity and issues related to NIKSHAY and PFMS portals). Conclusion: DBT coverage was low and there were substantial delays. Implementation barriers need to be addressed urgently to improve uptake and efficiency. The TB programme has begun to take action
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