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31.
Delivery of a hammerhead ribozyme specifically down-regulates the production of fibrillin-1 by cultured dermal fibroblasts 总被引:4,自引:1,他引:4
Kilpatrick MW; Phylactou LA; Godfrey M; Wu CH; Wu GY; Tsipouras P 《Human molecular genetics》1996,5(12):1939-1944
The hammerhead ribozyme is a small catalytic RNA molecule. Potential
hammerhead ribozymes that possess a catalytic domain and flanking sequence
complementary to a target mRNA can cleave in trans at a putative cleavage
site within the target molecule. We have investigated the potential of
hammerhead ribozymes to down-regulate the product of the fibrillin-1 gene
(FBN1). Fibrillin is a 347 kDa glycoprotein that is a major constituent of
the elastin-associated microfibrils. Mutations in the FBN1 gene are
responsible for Marfan syndrome (MFS), a common systemic disorder of the
connective tissue. Many FBN1 mutations responsible for MFS appear to act in
a dominant-negative fashion, raising the possibility that reduction of the
amount of product from the mutant FBN1 allele might be a valid therapeutic
approach for MFS. A trans-acting hammerhead ribozyme (FBN1-RZ1) targeted to
the 5' end of the human FBN1 mRNA has been designed and synthesized, and
shown to cleave its target efficiently in vitro. FBN1-RZ1 cleavage is
magnesium dependent and efficient at both 37 and 50 degrees C. Delivery of
the FBN1-RZ1 ribozyme into cultured dermal fibroblasts, by receptor-
mediated endocytosis of a ribozyme-transferrin-polylysine complex,
specifically reduces both cellular FBN1 mRNA and the deposition of
fibrillin in the extracellular matrix. These results suggest that the use
of hammerhead ribozymes is a valid approach to the study of fibrillin gene
expression and possibly to the development of a therapeutic approach to
MFS.
相似文献
32.
Burwinkel B; Maichele AJ; Aagenaes O; Bakker HD; Lerner A; Shin YS; Strachan JA; Kilimann MW 《Human molecular genetics》1997,6(7):1109-1115
Glycogen storage disease due to phosphorylase kinase deficiency occurs in
several variants that differ in mode of inheritance and tissue-
specificity. This heterogeneity is suspected to be largely due to mutations
affecting different subunits and isoforms of phosphorylase kinase. The gene
of the ubiquitously expressed beta subunit, PHKB, was a candidate for
involvement in autosomally transmitted phosphorylase kinase deficiency of
liver and muscle. To identify such mutations, the complete PHKB coding
sequence was amplified by RT-PCR of RNA isolated from blood samples of
patients and analyzed by direct sequencing of PCR products. The
characterization of mutations was complemented by PCR of genomic DNA. In
one female and four male patients, we identified five independent nonsense
mutations (Y418ter; R428ter; Y974H+E975ter; Q656ter in two cases), one
single-base insertion in codon N421, one splice-site mutation affecting
exon 31, and a large deletion involving the loss of exon 8. Although these
severe translation-disrupting mutations occur in constitutively expressed
sequences of the only known beta subunit gene of phosphorylase kinase,
PHKB, they are associated with a surprisingly mild clinical phenotype,
affecting virtually only the liver, and relatively high residual enzyme
activity of approximately 10%.
相似文献
33.
Human ovarian granulosa cells and follicular fluid indices: the relationship to oocyte maturity and fertilization in vitro 总被引:1,自引:0,他引:1
The study investigates the correlation between oocyte maturity and
fertilization and a variety of hormonal parameters in follicular fluid and
ovarian granulosa cells. A methodology for purification of granulosa cells
from contaminating blood cells is also established. A total of 63
follicular aspirates were collected at oocyte retrieval from 30 women
superovulated using the long luteinizing hormone- releasing hormone (LHRH
analogue)/human menopausal gonadotrophin regimen. Oestradiol, progesterone,
testosterone and human chorionic gonadotrophin (HCG) were quantified in
follicular fluid and granulosa cells were immunostained for human chorionic
gonadotrophin. Immunopurification of granulosa cells from contaminating
blood cells was performed. HCG in follicular fluid was significantly high
in follicles yielding immature (grade 3) oocytes (P=0.002); there was no
correlation with fertilization. Aspirates from follicles containing mature
(grade 1) oocytes and oocytes that subsequently fertilized had
significantly more granulosa cells immunobound to HCG (P < 0.001,
P=0.02). Moreover, the immunomagnetic purification technique provided
>98% pure population of granulosa cells. The data demonstrate that HCG
in follicular fluid and on granulosa cells may help to predict oocyte
maturity and fertilization. Furthermore, immunomagnetic beads provide a
reliable procedure for the purification of ovarian granulosa cells.
相似文献
34.
35.
Intravenous gammaglobulin treatment of chronic idiopathic thrombocytopenic purpura 总被引:11,自引:0,他引:11
Bussel JB; Kimberly RP; Inman RD; Schulman I; Cunningham-Rundles C; Cheung N; Smithwick EM; O'Malley J; Barandun S; Hilgartner MW 《Blood》1983,62(2):480-486
High-dose intravenous gammaglobulin (IVIgG) was given to 12 children and adults with chronic idiopathic thrombocytopenic purpura (ITP) to avoid splenectomy or because they either failed to respond to or required maintenance with high doses of steroids and/or immunosuppressives. The average platelet count increase to initial therapy was 239,500/microliters (range 23,000-790,000). A concomitant IgG Fc receptor blockade, measured by IgG-sensitized 51Cr-labeled autologous erythrocytes, was seen in 11 of 11 patients tested, both splenectomized and not splenectomized, lasting 3-4 wk. Six or more months after treatment, 2 children are in remission, 2 children and 2 adults are stable requiring no therapy with platelet counts of approximately 50,000 and 30,000, respectively, 3 children require maintenance IVIgG therapy at 2-10-wk intervals, and 1 child and 2 adults have become refractory to further IVIgG. Splenectomy was not performed in 4 children. Two adults were able to discontinue daily prednisone. The 3 patients who became unresponsive to Swiss Red Cross gamma-globulin (IgSRK) therapy did so in conjunction with a markedly elevated platelet-associated IgG and IgM. Serum IgM increased an average of 103 mg/dl after the IVIgG infusions. No significant side effects were seen. 相似文献
36.
37.
Twenty patients, aged 4 months to 58 years, were evaluated for liver transplantation by duplex sonography, and 15 transplantations were completed; 42 postoperative examinations were performed. Sonographic findings were correlated with seven preoperative and five postoperative angiographic evaluations. Preoperative duplex US findings included tumors, portal vein occlusion, varices, biliary obstruction, and variant vascular anatomy. Postoperative findings included hepatic artery occlusion, portal vein occlusions (one with cavernous transformation), portal vein stenosis, biliary obstruction, intrahepatic and extrahepatic fluid collections, and air in the portal vein due to ischemic bowel. Use of angiography allowed confirmation of the vascular abnormalities and demonstrated evidence of rejection in patients with normal Doppler waveforms. Duplex sonography is a valuable portable technique for evaluating these patients and can be used in triage of patients requiring angiography. 相似文献
38.
Greenlee MW; Koessler M; Cornelissen FW; Mergner T 《Cerebral cortex (New York, N.Y. : 1991)》1997,7(3):253-267
Visual discrimination and short-term recognition memory for computer-
generated random patterns were explored in 23 patients with a postsurgical
lesion in one of the cortical hemispheres. Their results are compared with
those of 23 age-matched volunteers. In a same- different forced-choice
discrimination task, d' and log beta (measures of sensitivity and bias), as
well as reaction time (RT) were determined. All participants viewed
patterns defined either by luminance contrast or isoluminant red-green
color contrast, the amplitude of which was adjusted to be 10 times the
respective detection threshold level. Block patterns consisting of a 6 x 6
matrix of light and dark (red and green) checks were randomly configured on
each presentation. They were presented in pairs, randomly in two visual
quadrants for a duration of 200 msec. Three presentation conditions were
used: simultaneous presentation of reference and test stimulus, sequential
presentation with a short delay (interstimulus interval, ISI = 3 s), and
sequential presentation with a long delay (ISI = 6 s). The results indicate
that patients with a lesion in the occipitotemporal cortex, the superior
temporal cortex and the frontal cortex were significantly impaired on both
luminance-contrast and color-contrast pattern discrimination. Patients with
damage in the anterior inferotemporal cortex showed no overall impairment.
The results suggest that performance in visual discrimination and
recognition memory tasks rely on distributed neural processes with more
than one neocortical location.
相似文献
39.
40.
We examined differences in approaches to HIV-related infection control practices in two university teaching hospitals in the United States and Nigeria. Health care workers (n = 202 in Nigeria and 186 in the USA) responded to a previously validated measure of infection control practices. There were significant differences in the estimated probability of treating a person with HIV disease (higher in USA), and a greater probability of peer ridicule as a way of enforcing group norms on infection control in Nigeria. Peer enforcement of norms was significantly lower in the USA. In both countries, more precautions would be taken if it was known that the patient was HIV infected. Infection control practices were more likely to be followed in the USA compared with Nigeria if they were praised for this activity, if appropriate facilities (sharps containers, gloves, etc.) were nearby, and if they felt that infection control procedures were effective. These data point to the importance of normative social pressures in Nigeria and of knowing the patient is HIV infected in the USA and feeling that infection control procedures are effective ways of avoiding occupational HIV infection. The role of normative pressures and assumptions about HIV infection status as well as cues and availability of facilities for infection control appear to differ between these health care workers in Nigeria and the USA. 相似文献