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- Ager, S, et al.
(författare)
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Retroviral display of antibody fragments; interdomain spacing strongly influences vector infectivity
- 1996
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Ingår i: Human Gene Therapy. - : Mary Ann Liebert Inc. - 1043-0342 .- 1557-7422. ; 7:17, s. 2157-2164
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Tidskriftsartikel (refereegranskat)abstract
- Five different single-chain antibody fragments (scFv) against human cell-surface antigens were displayed on murine ecotropic retroviral vectors by fusing them to the Moloney SU envelope glycoprotein. The spacing between the scFv and the SU glycoprotein was varied by fusing the scFv to residue +7 or to residue +1 of Moloney SU and by inserting linker sequences of different lengths between the domains. All of the chimeric envelopes were efficiently incorporated into vector particles and could bind to human cells through their displayed antibody fragments, but did not infect them. The spacing between the scFvs and the SU glycoproteins had no significant effect on the efficiency of envelope expression or viral incorporation and did not affect the binding properties of the chimeric envelopes, nor did it influence the efficiency of targeted gene delivery to human cells by scFv-displaying vectors. However, on murine fibroblasts the infectivity of vectors incorporating the chimeric envelopes was strongly influenced by the length of the interdomain spacer. The titers were very low when the single-chain antibodies were fused through a tripeptide linker to SU residue +7 and were greatly enhanced (up to 10(5)-fold) when they were fused to SU residue +1 through a heptapeptide linker. These results point to the importance of steric interactions between the domains of chimeric envelope glycoproteins and may have implications for retroviral vector design for human gene therapy.
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| 2. |
- J L Sussman, M Harel, M L Raves, K Giles, R B G Ravelli, L G M Peng, G J Kleywegt, D M Quinn, H K Nair, and I Silman
(författare)
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Studies on the 3-D structure of Torpedo Acetylcholinesterase
- 1996
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Ingår i: 1996 U.S. Army Medical Defense Bioscience Review. - : U.S. Army Medical Research Institute of Chemical Defense, Baltimore, MD. ; , s. 13-22
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Bokkapitel (refereegranskat)
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| 6. |
- Heyerdahl, H, et al.
(författare)
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Pharmacokinetic studies on 5-aminolevulinic acid-induced protoporphyrin IX accumulation in tumours and normal tissues
- 1997
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Ingår i: Cancer Letters. - 1872-7980. ; 112:2, s. 225-231
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Tidskriftsartikel (refereegranskat)abstract
- Laser-induced fluorescence (LIF) for in vivo point monitoring and fluorescence microscopy incorporating a CCD camera were used to study the fluorescence distribution of 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) in tumours. Fluorescence in a chemically induced adenocarcinoma in the liver of rats and in an aggressive basal cell carcinoma in a patient were studied after intravenous injection of ALA at a dose of 30 mg/kg body weight. The LIF technique demonstrated slightly more ALA-induced PpIX fluorescence in the tumour than in the surrounding normal liver and abdominal muscle of rats. The visible parts of the human basal cell carcinoma exhibited strong ALA-induced fluorescence, while this fluorescence was much weaker in the necrotic areas of the tumour and in the surrounding normal skin. (C) 1997 Elsevier Science ireland Ltd.
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| 7. |
- Liu, Y X, et al.
(författare)
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Prolactin delays gonadotrophin-induced ovulation and down-regulates expression of plasminogen-activator system in ovary.
- 1997
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Ingår i: Human Reproduction. - 0268-1161 .- 1460-2350. ; 12:12, s. 2748-55
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Tidskriftsartikel (refereegranskat)abstract
- This study was conducted to determine whether prolactin (PRL) suppresses gonadotrophin-induced ovulation and disturbs the co-ordinated gene expression of tissue type plasminogen activator (tPA) and plasminogen activator inhibitor type-1 (PAI-1) in rat ovary. Immature female rats were injected with 10 IU pregnant mare's serum gonadotrophin to stimulate follicle growth, and 48 h received different doses of prolactin followed by 7 IU human chorionic gonadotrophin (HCG). The oviducts were examined for the presence of ova, and the amounts of tPA and PAI-1 mRNA present in the ovary were measured at various times after the hormone treatment. PRL had no significant effect on ovarian weight but caused a dose-dependent decrease in ovulation number. In the control animals receiving HCG alone, 13.3 +/- 1.3 (mean +/- SEM) ova/oviduct were found; while in animals receiving HCG plus 50, 100 or 200 microg PRL, the ovulation number was dose-dependently suppressed by 53.6, 66.9 and 76% respectively at 18 h after treatment. PRL suppression of HCG-induced ovulation was time-dependent. By 24 h after treatment, the number of ova in the oviducts in HCG- and HCG plus PRL-treated groups was not significantly different. PRL also suppressed HCG-induced tPA gene expression in a dose- and time-dependent manner. At all time points examined, tPA mRNA content of whole ovaries and granulosa cells (GC) in PRL-treated groups was lower than in the HCG-treated controls. The activities of PAI-1 in ovarian extracellular fluid (OEF) and PAI-1 mRNA in the theca-interstitial cells (TI) in the PRL-treated groups were higher than in the HCG-treated controls. The highest stimulation by PRL of PAI-1 activity in OEF and of PAI-1 mRNA in TI was observed at 9 h and 6 h after HCG treatment respectively. The localization of tPA and PAI-1 antigens in the ovaries was consistent with changes in the mRNA and activity levels. These data suggest that PRL temporarily delays, but does not completely inhibit, HCG-induced ovulation, which may be caused by a suppression of PA-mediated proteolysis.
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| 8. |
- Liu, Y X, et al.
(författare)
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Prolactin regulation of tissue type plasminogen activator and plasminogen activator inhibitor type-I gene expression in eCG-primed rat granulosa cells in culture.
- 1998
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Ingår i: Biology of Reproduction. - 0006-3363 .- 1529-7268. ; 59:2, s. 409-16
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Tidskriftsartikel (refereegranskat)abstract
- The present study was designed to investigate the effect of prolactin (PRL) on plasminogen activator inhibitor-I (PAI-I) and tissue type plasminogen activator (tPA) gene expression in eCG-primed granulosa cells in vitro. At 46 h after the hormone treatment, ovaries were removed, and granulosa cells were prepared for culture. Cells were incubated for various times in serum-free medium in the presence or absence of LH and PRL alone or in combination. tPA and PAI-I activities in the media were assayed by fibrin overlay and reverse fibrin autograph, respectively. Cytoplasmic RNA from granulosa cells was prepared using the NP-40 method and was assayed for PAI-I and tPA mRNA levels. We demonstrated the following. 1) PRL increased PAI-I mRNA production in cultured granulosa cells. Inclusion of LH with PRL had a synergistic effect on increasing PAI-I mRNA levels. After 48-h culture, 3-fold increases in PAI-I mRNA levels were seen with LH in combination with PRL as compared with PRL alone. The synergistic increase in PAI-I mRNA levels occurred in a dose- and time-dependent manner. 2) The increase in PAI-I mRNA synthesis by PRL alone, or by PRL in combination with LH, was well correlated with the changes in PAI-I activity and antigen levels in the conditioned media. 3) PRL in the culture also dramatically decreased LH-induced tPA mRNA and activity in a dose- and time-dependent fashion. The decrease in the tPA activity by PRL was also correlated with an increase in the amount of PA-PAI-I complexes in the cell-conditioned media. 4) In situ hybridization of tPA and PAI-I mRNAs in the cultured granulosa cells also showed that PRL was capable of enhancing PAI-I mRNA while diminishing tPA mRNA production induced by LH. This suggests that the dose- and time-dependent decrease in the gonadotropin-induced tPA activity in the culture by the presence of PRL may be due to decreasing tPA mRNA synthesis on one hand and to neutralization of the tPA activity by the increased PAI-I activity on the other.
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