Lymphocyte subpopulations in middle ear effusions: flow cytometry analysis.

OBJECTIVE: The aim of this study was to identify lymphocyte subpopulations in middle ear effusions, peripheral blood, and adenoids in children suffering from otitis media with effusion. SETTING: Tertiary referral center. PATIENTS: Thirty-three children (55 ears) undergoing myringotomy for otitis media with effusion. METHODS: CD3, CD4, CD8, CD19, and natural killer cell populations were investigated in middle ear effusion, peripheral blood, and adenoids using a three-color monoclonal antibody and flow cytometry method for quantitative estimation. RESULTS: T cells (CD3) are dominating lymphocytes in middle ear effusion. Among T lymphocytes, the majority are those of the helper type (CD4). The dominating isoform among CD4 lymphocytes are memory cells (CD4CD45RO); among CD8 lymphocytes, naive cells (CD8CD45RA). The percentage of CD4 cells, CD8 cells, and the CD4/CD8 ratio was significantly higher in middle ear effusions than in blood. The percentage of memory CD4 lymphocytes and naive CD8 lymphocytes was significantly lower in the middle ear effusion. Lymphocyte subsets were compared between 22 pairs of effusions from each patient. The percentage of each type of cell did not differ significantly. CONCLUSION: The results of this study indicate local regulation of the lymphocyte profile in middle ear effusions and the same phase of immune response in two ears of the same patient.     

The role of quinone reductase (NQO1) and quinone chemistry in quercetin cytotoxicity.

The effects of quercetin on viability and proliferation of Chinese Hamster Ovary (CHO) cells and CHO cells overexpressing human quinone reductase (CHO+NQO1) were studied to investigate the involvement of the pro-oxidant quinone chemistry of quercetin. The toxicity of menadione was significantly reduced in CHO+NQO1 cells compared to wild-type CHO cells, validating the NQO1-overexpression in the CHO+NQO1 transfectant. Quercetin inhibited the proliferation of wild-type CHO and CHO+NQO1 cells to a similar extent without affecting cell viability, indicating that NQO1 enrichment of CHO cells did not provide increased protection. On the other hand, inhibition of NQO1 in both types of cells by dicoumarol significantly potentiated the inhibitory effect of quercetin on cell proliferation, revealing the role of NQO1 in cellular protection against quercetin. Altogether, these results can be explained by the hypothesis that both wild-type CHO and CHO+NQO1 cells contain sufficient NQO1 activity for optimal protection against the pro-oxidant effect of quercetin on cell proliferation. The results also point at a cellular NQO1 threshold for optimal protection against quercetin. This NQO1 threshold seems to be in the range of NQO1 activities already present in various tissues.     

CD80 and CD86 expression on LPS-stimulated monocytes and the effect of CD80 and CD86 blockade on IL-4 and IFN-gamma production in nonatopic bronchial asthma

CD80 and CD86 seem to play an important role in the allergen-induced secretion of interleukin (IL)-5 and IL-13. Up to now, the expressions of CD80 (B7.1) and CD86 (B7.2) on monocytes and the kinetics of the expression of these molecules on lipopolysaccharide-stimulated monocytes in nonatopic asthma have not been defined. Using monoclonal antibodies, we have compared the expressions of CD80 (B7.1) and CD86 (B7.2) on the monocytes of healthy persons and nonatopic asthmatic patients. We have also assessed the effect of CD80 and CD86 inactivation on IL-4 and interferon (IFN)-gamma production in nonatopic asthmatics and healthy subjects. We found that a low expression of CD80 (1.64 +/- 0.65 vs. 3.53 +/- 1.43%) and a moderate expression of CD86 (41.25 +/- 13.4 vs. 49.46 +/- 11.49%) on the studied monocytes were characteristic for asthma. In nonatopic asthma patients inactivation of CD80 or CD86 blockade significantly reduced IFN-gamma production by T lymphocytes (p < 0.02; p < 0.03). In both the studied groups, anti-CD80 antibodies did not diminish T lymphocyte production of IL-4. However, anti-CD86 antibodies significantly (p < 0.04) reduced the IL-4 concentration in culture supernatants. Our results confirm that both the CD80 and CD86 molecules play an important role in the maintenance and amplification of the inflammatory process. It suggests that in the inflammatory process that occurs in nonatopic bronchial asthma, Th1 as well as Th2 lymphocytes are equally important.     

Human heat shock protein 60 (409-424) fragment is recognized by serum antibodies of patients with acute coronary syndromes.

Acute coronary syndromes (ACS), including unstable angina (UA) and acute myocardial infarction (MI), are clinical manifestations of a progressive atherosclerotic process. Antibodies (Ab) to heat shock proteins (hsp) have been reported to be associated with atherosclerosis. Blood samples from 35 patients with ACS and 20 healthy volunteers were tested for Ab to human hsp60 by an enzyme-linked immunosorbent assay (ELISA). Levels of specific serum Ab against hsp60 were significantly elevated in patients with ACS when compared to clinically healthy subjects. To determine the antigenic determinants recognized by these Ab, antibody binding to seven peptides, selected from the hydrophilic and acrophilic regions of the human hsp60 molecule, was assessed. Despite the individual variation in the immune response among patients, one immunodominant region was revealed corresponding to the hsp60 (409-424) peptide. The identification of this epitope may be important for understanding the function of this protein in the atherosclerotic process.     

TGFbeta1 enhances formation of cellular Abeta/apoE deposits in vascular myocytes

Brain injury increases the risk of Alzheimer's disease (AD) through unknown mechanisms. We studied deposition of amyloid-beta protein (Abeta) in cells exposed to transforming growth factor beta1 (TGFbeta1), a cytokine that regulates cell metabolism during brain injury, and apolipoproteinE (apoE), the major lipid transporter in the brain. The studies were conducted by using brain vascular smooth muscle cells that are engaged in beta-amyloidosis in vivo and produce Abeta in cell culture. We found that cell treatment with TGFbeta1 together with apoE4 strongly increased the amount of cellular Abeta. The intracellular Abeta co-localized with apoE but not with TGFbeta, similarly as in vascular beta-amyloid. Some cellular Abeta/apoE deposits increased in size and persisted in culture even after the TGFbeta1 and apoE4 were removed. The appearance of cellular deposits of Abeta was associated with increased production of the amyloid-beta precursor protein and cellular retention of its mature form. The results suggest that the concomitant presence of apoE and TGFbeta1 can trigger vascular beta-amyloidosis by inducing intracellular formation of stable Abeta/apoE deposits.     

Influence of isopropylvaleramide and allylisopropylacetamide on transformation of C3H/10T? cells induced by benzo[a]pyrene derivatives

We examined the effect of aliphatic amides isopropylvaleramide(IVA) and allylisopropylacetamide (AIA) on the oncogenic transformationof C3H/10T? cells induced by benzo[a]pyrene (B[a]P) or its proximateand ultimate metabolites (?)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene(B[a]P-7,8-diol) and (+)-7ß,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene(B[a]P-diol-epoxide), respectively. IVA and AIA given priorto, simultaneously with, or for 24 h intervals beginning upto 48 h after removal of carcinogens significantly suppressedtransformation induced by B[a]P or the 7,8-diol metabolite.Both modifiers were most effective when added for 24 h immediatelyfollowing carcinogen exposure. IVA and AIA were also very potentinhibitors of B[a]P-diol-epoxide transformation; however theywere most effective when added for 24 h simultaneously withthe B[a]P-diol-epoxide. No significant difference in B[a]P-diol-epoxidebinding to DNA in C3H/10T? cells was observed during 1 or 24h exposure to this carcinogen in the presence or absence ofIVA or AIA. Neither modifier affected X-ray transformation whenadded for 24 h immediately following X-irradiation of C3H/10?cells.These results suggest that AIA and especially IVA might be importanttools in studies directed at non-metabolic aspects of B[a]Pcarcinogenesis.     

Amphiphilic star-like macromolecules as novel carriers for topical delivery of nonsteroidal anti-inflammatory drugs

The objective of this study was to evaluate amphiphilic star-like macromolecules (ASMs) as a topical drug delivery system. Indomethacin, piroxicam, and ketoprofen were individually encapsulated into the ASMs using coprecipitation. The effects of the ASMs on percutaneous permeation of nonsteroidal anti-inflammatory drugs (NSAIDs) across full thickness, hairless mouse skin were evaluated in vitro using modified Franz diffusion cells. In addition, solubility and in vitro release experiments were performed to characterize ASMs behavior in aqueous media. Poly(ethylene glycol) (PEG) and Pluronic P-85 were used as polymer controls to compare the role of PEG and amphiphilic behavior in the ASMs. In vitro release experiments indicated that ASMs can delay drug release (P⋖05), whereas solubility measurements showed that ASMs can increase NSAIDs aqueous solubility (P⋖05). Percutaneous permeation studies revealed that ASMs decreased both flux and Q₂₄ of drugs compared with the control (P⋖10). Skin pretreatment studies with ASM-containing solution before drug application demonstrated that pretreatment similarly influenced NSAID percutaneous permeation. In conclusion, ASMs likely slow drug permeation through 2 mechanisms, delayed drug diffusion from its core and skin dehydration by its shell. Thus, ASMs may be useful for delayed dermal delivery or prevention of compound permeation through the skin (eg, sunscreens, N,N-diethyl-m-toluamide [DEET]) from aqueous formulations.     

Investigation of Structural,Physical, and Attenuation Parameters of Glass: TeO2-Bi2O3-B2O3-TiO2-RE2O3 (RE: La,Ce, Sm,Er, and Yb), and Applications Thereof

A novel series of glass, consisting of B₂O₃, Bi₂O₃, TeO₂, and TiO₂ (BBTT) containing rare earth oxide RE₂O₃, where RE is La, Ce, Sm, Er, and Yb, was prepared. We investigated the structural, optical, and gamma attenuation properties of the resultant glass. The optical energy bands, the linear refractive indices, the molar refractions, the metallization criteria, and the optical basicity were all determined for the prepared glass. Furthermore, physical parameters such as the density, the molar volume, the oxygen molar volume, and the oxygen packing density of the prepared glass, were computed. Both the values of density and optical energy of the prepared glass increased in the order of La₂O₃, Ce₂O₃, Sm₂O₃, Er₂O₃, and then Yb₂O₃. In addition, the glass doped with Yb₂O₃ had the lowest refractive index, electronic polarizability, and optical basicity values compared with the other prepared glass. The structures of the prepared glass were investigated by the deconvolution of infrared spectroscopy, which determined that TeO₄, TeO₃, BO₄, BO₃, BiO₆, and TiO₄ units had formed. Furthermore, the structural changes in glass are related to the ratio of the intensity of TeO₄/TeO₃, depending on the type of rare earth. It is also clarified that the resultant glass samples are good attenuators against low-energy radiation, especially those that modified by Yb₂O₃, which exhibited superior shielding efficiency at energies of 622, 1170, and 1330 keV. The optical and gamma ray spectroscopy results of the prepared glass show that it is a good candidate for nonlinear optical fibers, laser solid material, and optical shielding protection.