The shaker 2 (sh2) and pirouette (pi) mouse mutants display severe inner ear dysfunction that involves both auditory and vestibular manifestation. Pathology of the stereocilia of hair cells has been found in both mutants. This study was designed to further our knowledge of the pathological characteristics of the inner ear sensory epithelia in both the sh2 and pi strains. Measurements of auditory brainstem responses indicated that both mutants were profoundly deaf. The morphological assays were specifically designed to characterize a pathological actin bundle that is found in both the inner hair cells and the vestibular hair cells in all five vestibular organs in these two mutants. Using light microscope analysis of phalloidin-stained specimens, these actin bundles could first be detected on postnatal day 3. As the cochleae matured, each inner hair cell and type I vestibular hair cell contained a bundle that spans from the region of the cuticular plate to the basal end of the cell, then extends along with cytoplasm and membrane, towards the basement membrane. Abnormal contact with the basement membrane was found in vestibular hair cells. Based on the shape of the cellular extension and the actin bundle that supports it, we propose to name these extensions cytocauds. The data suggest that the cytocauds in type I vestibular hair cells and inner hair cells are associated with a failure to differentiate and detach from the basement membrane. 相似文献
Tamm-Horsfall protein (THP) is the major glycoprotein component of urine, yet its biological role remains obscure. Recent reports have suggested that a concanavalin A (Con A)-binding fraction of THP from pregnancy urine can bind the cytokines tumour necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL-1). In order to investigate this claim in relation to THP from normal adult urine we raised monoclonal antibodies to THP and sought THP/TNF-alpha interactions in three separate assay systems. We found no evidence that THP binds to TNF-alpha under physiological conditions, but we observed that it exerts a weak, probably not physiologically relevant, but reproducible inhibitory effect on the toxicity of TNF-alpha for monolayers of L929 cells, even when the cells are pretreated with the THP, and washed before addition of the cytokine. Since our preparations of THP do not interact directly with TNF-alpha we postulated an interaction with the cells themselves, or with their extracellular matrix. The THP was found by ELISA, immunoblotting and immunohistology, to bind to as yet unidentified components of the extracellular matrix in a manner dependent on cations, pH and carbohydrates. These data, considered in the light of the published amino acid sequence and biochemical properties, suggest that THP is a member of a structural glycoprotein family known to modulate cell adhesion. 相似文献
A plasma HIV-1 RNA amplification assay (RNA assay), a quantitative peripheral blood mononuclear cell (PBMC) microculture (culture), and a PBMC HIV-1 DNA amplification assay (DNA assay) were compared for diagnosis of HIV-1 infection in infants receiving zidovudine in Pediatric AIDS Clinical Trials Group protocol 185; assays were performed for all 24 infected and 100 uninfected infants. HIV-1 infection was defined as >or=2 positive cultures or positive antibody to HIV-1 at >or=18 months. Cultures were performed at birth and 6 and 24 weeks of age; DNA and RNA assays were performed on cryopreserved specimens. The sensitivity of culture and DNA and RNA assays at birth was 20.8%, 10.5%, and 26.7%, respectively. At older ages, sensitivity typically exceeded 80%, remaining highest for the RNA assay (>85%). Assay specificity was >99%. Positive predictive values exceeded 93% for each assay at each age; negative predictive values were highest (>90%) for the RNA assay. At birth (P < 0.005) and age 6 weeks (P < 0.001), a significantly larger proportion of infected infants were identified by means of the RNA assay than by the other assays. The diagnostic performance of the RNA assay matched or exceeded that of culture and the DNA assay. Given that RNA assays require less blood volume and yield rapid results, our study adds to existing data suggesting that RNA assays may be used for early diagnosis of HIV-1 infection in infants. 相似文献
The occurrence of immune complexes in the serum and the level of the C3 breakdown product C3d in the plasma from patients with leprosy were studied by quantitative methods and the results were compared in various forms of the disease. These studies were performed on sixty-two samples from twenty-six patients. The serum 125I-C1q binding activity was found to be increased by more than 2 s.d., as compared to the normal values, in most of the sera from patients with erythema nodosum leprosum (ENL) (80%) and uncomplicated lepromatous leprosy (82%), but also in the sera from patients with tuberculoid leprosy (58%). In vitro studies suggested that immune complexes involving mycobacterial antigens were present in leprosy sera. An increased C3d level (greater than 2s.d.) was also found in most of the plasma from patients with ENL (70%), but rarely in the plasma from patients with uncomplicated lepromatous leprosy (18%) and never in tuberculoid leprosy patients' plasma. The absence of a significant correlation between the 125I-C1q binding activity and the C3d level in leprosy patients may suggest that extravascular immune complexes are involved in the complement activation occurring in ENL. The quantitation of C3d in plasma may be of some practical interest in the early diagnosis of ENL complications of leprosy. 相似文献
In a human in-vitro fertilization (IVF) programme, the effect of co-
culture of embryos with human fibroblasts was evaluated with respect to
pregnancy rate and embryo development. Patients were included in the study
after giving informed written consent. The IVF treatments were randomly
assigned by stratification of both age (<36 versus > or =36 years)
and previous IVF attempts (yes versus no). After fertilization was
established, the zygotes were transferred to a 4-well dish with or without
fibroblasts and cultured for 2 days. On the third day after ovum pick-up
(OPU), cell number and quality [5 (good) to 1 (poor)] of the embryos were
scored and a maximum of three embryos was transferred. Supernumerary
embryos of good quality were cryopreserved. The design of this study was a
group sequential trial with the objective of detecting differences between
pregnancy rates following IVF with conventional incubation or incubation in
co-culture with fibroblasts. This design included one evaluation at
half-way data collection. In the study, 148 patients had an OPU, of whom 77
were allocated to the co-culture group. There was no statistically
significant difference in pregnancy rate, cell number and embryo quality
between the two groups. The ongoing pregnancy rate per embryo transfer was
27% in co-culture and 30% in the conventional culture group. The
implantation rates per transferred embryo were 17 and 18% respectively.
Using a multivariate logistic regression model for the probability of
ongoing pregnancies, the odds ratio of co-culture, adjusted for age and
previous IVF attempts, was not statistically significant. In conclusion,
co-culture with human fibroblasts does not contribute to an improvement of
embryo quality nor to a higher pregnancy rate after IVF in an unselected
group of patients.
相似文献
Bovine conglutinin was used in a solid-phase assay for the detection of immune complexes. In a first step, the tested serum sample is incubated in polypropylene tubes coated with conglutinin to allow C3-coated immune complexes to bind to solid-phase conglutinin. In a second step, the conglutinin-bound complexes are detected using an enzyme-conjugated or radiolabelled anti-immunoglobulin antibody.
The conglutinin-binding (KgB) test does not suffer from the interference of DNA, heparin or endotoxins. Its limit of sensitivity for aggregated IgG is 3 μg/ml undiluted human serum. Immune complexes prepared in vitro using tetanus toxoid, or DNA, and corresponding antibodies in human sera could be detected at various antigen/antibody ratios and at antibody concentrations lower than 8 μg/ml. The KgB test allowed for the detection of immune complexes in sera from patients with systemic lupus erythematosus, rheumatoid arthritis, idiopathic vasculitis, leprosy and leukemia. These sera were also tested using the 125I-labelled Clq-binding activity (BA) test and the KgB test simultaneously, and a significant rank order correlation was observed. In patients with leukemia, a significant correlation was observed using three tests, KgB, 125I-labelled Clq BA and Raji-cell radioimmunoassay (RIA).
Therefore, the KgB test appears as a simple and reproducible method, utilizing a very stable reagent, with a sensitivity and specificity comparable to the other tests studied and allowing for clinical application.
The proteins of respiratory syncytial (RS) virus were analyzed by SDS-polyacrylamide gel electrophoresis. Eight virion structural proteins with molecular weights of 180,000, 89,000, 48,000, 42,000, 34,000, 28,000, 25,000, and 21,000 were identified. These proteins were given tentative designations of L (180,000), G (89,000), F1 (48,000), NP (42,000), P (34,000), M (28,000), Vp25 (25,000), and F2 (21,000). The 89,000-, 48,000-, and 21,000-dalton polypeptides were glycosylated and could be purified on lentil-lectin sepharose columns. All three glycoproteins could be immunoprecipitated from extracts of infected cells but not from uninfected cells, suggesting that they are viral specified. The host cell affected the apparent molecular weights of the largest and smallest glycosylated polypeptides possibly by differences in glycosylation. The 48,000- and 21,000-dalton glycopolypeptides were disulfide linked subunits of a 68,000-dalton glycoprotein that was seen on unreduced gels. The 68,000-dalton glycoprotein was thus similar to the fusion (F) protein of paramyxoviruses. Treatment of infected cultures with tunicamycin, a drug that blocks glycosylation, inhibited syncytial formation and resulted in over a 1000-fold reduction of extracellular infectious virus. Virions purified from tunicamycin-treated cells had reduced amounts of all three glycosylated proteins. No new forms of these proteins were conclusively identified, suggesting that unglycosylated forms of RS glycoproteins were not incorporated into virion membranes. 相似文献
PROBLEM: During normal pregnancy, major changes occur in the production of Th2/Th1 cytokines at the feto-maternal interface. Th2 cytokines such as interleukin-4 (IL-4) or interleukin-10 (IL-10) are predominantly produced locally in the uterine and placental tissues, whereas the production of Th1 cytokines such as tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ) are decreased. Because these modulations might be induced by the embryo, the current study was carried out to test the effect of rabbit blastocoelic fluid on the production of Th2/Th1 cytokines by lymphocytes, and to investiate the possible implication of transforming growth factor β2 (TGF-β2) prostaglandin E2 (PGE2) as modulators of the production of these cytokines. METHOD OF STUDY: Human peripheral blood lymphocytes (PBL) were cultured along with ConcanavalinA (Con A), and rabbit blastocoelic fluid was collected on day 12 of gestation (BF d-12). Concentrations of cytokines in culture media were determined by enzyme-linked immunoadsorbent assay (ELISA). RESULTS: Addition of BF d-12 in the culture medium induced a strong inhibition of IL-2, TNF-α, IL-10, and granulocyte-macrophage colony-stimulating factor (GM-CSF) production. However, an initial pretreatment of the lymphocytes with BF d-12, followed by a Con A stimulation, led to a marked increase in GM-CSF production, whereas IL-2, TNF-α, and IL-10 secretions were inhibited. It was also demonstrated, for the first time, that a pretreatment of the lymphocytes with TGF-β2 and PGE2 increased GM-CSF production to the same level reached after the addition of BF d-12. Furthermore, removal of TGF-β2 and PGE2 from BF d-12 by affinity chromatography reduced the effect of BF d-12 on GM-CSF production. CONCLUSIONS: Taken together, these findings suggest that the embryo, in modulating harmful and beneficial cytokine production locally, plays an active role in its protection against maternal immune cellular assault. These results also emphasize the importance of growth factors for successfully maintaining pregnancy. 相似文献
The impact of the insertion (I)/deletion (D) (I/D) polymorphism in the angiotensin 1-converting enzyme (ACE) gene on the extent of white matter myelin loss (ML) was investigated in four regions of the cerebral cortex in an autopsy-confirmed series of 93 patients with Alzheimer's disease (AD). The possible influence of APO E epsilon4 allele acting in concert with ACE D allele was assessed. The extent of ML did not differ between D/D, I/D and I/I genotype groups when data from all four brain regions were combined. However, separate analysis showed that the frontal and temporal cortex tended to be affected more severely by ML in patients with D/D genotype compared to those with I/D and I/I genotypes. Stratification according to APO E epsilon4 allele revealed a greater overall ML in patients bearing at least one copy of ACE D allele and one APO E epsilon4 allele, especially in individuals homozygous for both. The APO E epsilon4 allele may therefore act synergistically in patients with AD (and other subjects) bearing ACE D/D genotype to increase the risk of ML, perhaps through transient ischaemic episodes consequent upon poor cardiac output associated with coronary atherosclerosis in patients with the APO E epsilon4 allele. 相似文献