The role of fatty acids in the regulation of brush-border Ca2+ transport.

The role of the fatty-acid composition of the intestinal brush-border membrane in the control of transmembrane Ca2+ transport was examined by in vitro acylation of endogenous phospholipids. The initial rate of Ca2+ uptake into brush-border membrane vesicles was stimulated 1.6-fold by incubation with 50 microM linoleoyl CoA, which was the most effective CoA ester examined. Oleoyl CoA was also active, but stearoyl CoA, palmitoyl CoA and arachidonyl CoA displayed no activity. The effect of linoleoyl CoA was specific for Ca2+ transport; sodium-dependent phosphate uptake was slightly inhibited and the activity of alkaline phosphatase, a brush-border enzyme, was unaffected. Incubation of brush-border vesicles with either stimulatory (oleate) or nonstimulatory (stearate) CoA esters resulted in the incorporation of fatty acid into the four major phospholipid classes, suggesting a fatty-acid specificity of the Ca2+ transport phenomenon. These results are consistent with the hypothesis that fatty acids are important elements in the control of brush-border Ca2+ transport.

Discovery and spectroscopy of the young jovian planet 51 Eri b with the Gemini Planet Imager.

Directly detecting thermal emission from young extrasolar planets allows measurement of their atmospheric compositions and luminosities, which are influenced by their formation mechanisms. Using the Gemini Planet Imager, we discovered a planet orbiting the ~20-million-year-old star 51 Eridani at a projected separation of 13 astronomical units. Near-infrared observations show a spectrum with strong methane and water-vapor absorption. Modeling of the spectra and photometry yields a luminosity (normalized by the luminosity of the Sun) of 1.6 to 4.0 × 10(-6) and an effective temperature of 600 to 750 kelvin. For this age and luminosity, "hot-start" formation models indicate a mass twice that of Jupiter. This planet also has a sufficiently low luminosity to be consistent with the "cold-start" core-accretion process that may have formed Jupiter.

Interferon gamma expression in human nasal polyps.

One feature among nasal polyps (NPs) is the predominance of lymphocytes and eosinophils. We hypothesize that elevated levels of interferon gamma (IFN-gamma) activate lymphocytes and eosinophils within the NP microenvironment. Nasal polyps were evaluated for distribution and levels of IFN-gamma in specimens from 27 patients with nasal polyposis and 4 controls. Immunohistochemical study revealed IFN-gamma staining of eosinophils, glandular cells, and epithelium (27 of 27 patients). ELISA analysis indicated elevated IFN-gamma levels in total NP tissues (25.6 +/- 7.23 pg/mg total protein [TP]) compared with controls (16.27 +/- 6.54 pg/mg TP). Three subpopulations were identified based on IFN-gamma levels: low IFN-gamma group (10.7 +/- 5.51 pg/mg TP); medium IFN-gamma group (25.70 +/- 5.90 pg/mg TP); and high IFN-gamma group (52.58 +/- 10.29 pg/mg TP). The latter levels were approximately 3.5 times the control levels (P<0.0025). Patients with previous polypectomy surgery showed higher levels of IFN-gamma compared with controls (P<0.0423). A trend was found with increased IFN-gamma levels and allergy, asthma, and topical steroid use.

Otoacoustic emissions in full-term newborns at risk for hearing loss.

Distortion product otoacoustic emissions (DPOEs) and click-evoked otoacoustic emissions (CEOEs) characteristics of the normal newborn population have been previously reported in the literature. There is little information about DPOE evaluations in the newborn population at risk for hearing loss. The authors now report the DPOE and/or CEOE data from six full-term subjects at risk for hearing loss or with highly suspected hearing loss. These subjects were less than 1 year of age and at risk for hearing loss secondary to a history of hereditary hearing loss, meningitis, hyperbilirubinemia, and ototoxic drug exposure. Audiometric evaluation included auditory brainstem responses (ABR), behavioral observation audiometry, and tympanometry. The CEOEs and DPOEs were found to be decreased or absent in the subjects with suspected hearing loss secondary to cochlear pathology; they were found to be normal in a subject with a suspected central hearing loss. This study's data suggest that otoacoustic emissions when combined with ABR can provide a frequency-specific evaluation of cochlear function and help determine the anatomic site of a pathologic lesion.

Transforming growth factor-beta expression in otitis media with effusion.

OBJECTIVE: To characterize the existence and role of transforming growth factor-beta (TGF-beta) in otitis media with effusion (OME). STUDY DESIGN: Retrospective. METHODS: The levels of two major TGF-beta isoforms, TGF-beta1 and TGF-beta2, in the middle ear effusions (MEEs) of 44 children were evaluated using enzyme-linked immunospecific assays (ELISAs). Forty-eight MEEs were separated into three clinically relevant groups (i.e., serous, mucoid, and purulent), and TGF-beta levels were correlated with clinical parameters of disease for these MEEs. RESULTS: Both TGF-beta1 and TGF-beta2 were present in the samples. Mean levels of TGF-beta1 (920.36 +/- 437.75 pg/mg total protein) were generally 100-fold greater than those of TGF-beta2 (9.65 +/- 11.19 pg/mg total protein). TGF-beta1 levels were elevated in association with a history of previous tympanostomy tube placements (TTPs) (P = .029) and mucoid effusions (P = .042). TGF-beta2 levels were elevated in association with a history of previous TTPs (P = .100) and chronic (i.e., serous or mucoid) effusions (P = .003). CONCLUSIONS: TGF-beta1 is present in the MEEs of children with OME. Furthermore, TGF-beta1 and TGF-beta2 levels were elevated differentially in the presence of chronic disease indicators in OME, suggesting that these isoforms may have differing roles in the inflammatory processes that characterize OME.

Distortion-product and click-evoked otoacoustic emissions in healthy newborns.

Distortion-product otoacoustic emissions (DPOEs) are believed to provide frequency-specific information about cochlear function. The DPOEs have been reported in the adult population but have not been reported previously in the neonatal population. We recorded DPOEs from a group of healthy full-term newborn human subjects (35 ears) to establish the characteristics of these emissions in the newborn population. To our knowledge, this is the first study of DPOEs in newborns. The "DPOE audiograms" from the newborns tested revealed characteristics qualitatively similar to those seen in adults with normal hearing. This study demonstrates the feasibility of DPOE measurements among newborns and provides a normal baseline for this age group, thus fulfilling a necessary step toward the development of an objective, noninvasive frequency-specific test of cochlear function. Click-evoked otoacoustic emissions were also recorded from the newborn population and compared with click-evoked otoacoustic emissions from adults. The spectrum of the click-evoked emissions was variable and individualistic, similar to findings previously reported in adult subjects. The click-evoked otoacoustic emissions of the newborns had a higher overall level and contained stronger high-frequency (4.5 to 6 kHz) spectral components than those of the adults. We also found that the low-frequency components of the click stimulus spectrum were attenuated in the neonatal ears exhibiting a high-pass slope below about 2.5 kHz, whereas the stimulus spectrum was nearly flat in this frequency region in adult ears.

Topical corticosteroid treatment of anosmia associated with nasal and sinus disease.

OBJECTIVE: To establish the efficacy of topical corticosteroid nasal spray treatment of severe olfactory loss associated with severe nasal and sinus disease. DESIGN: Efficacy before and after open-label trial of topical corticosteroid nasal spray used exclusively in the head-down-forward position. SETTING: Taste and smell clinic of a university teaching hospital and research facility. PATIENTS: Taste and smell clinic patients with anosmia or severe hyposmia associated with paranasal sinus disease and nasal polyposis including 39 of 45 patients recruited from 1988 to 1994 who completed the topical corticosteroid treatment course and returned for subsequent testing. INTERVENTION: At least 8 weeks of treatment with flunisolide (Nasalide), 2 sprays in each nostril twice a day, with concurrent antibiotic treatment of any bacterial infection. MAIN OUTCOME MEASURES: Subjective olfactory symptoms, objective olfactory function tests, and otolaryngological evaluation (including endoscopic examination). RESULTS: Olfactory scores significantly improved following treatment (P < .001); signs of nasal and sinus disease significantly decreased (P < .001); and 26 (66%) of the patients reported a subjective improvement in their sense of smell. CONCLUSION: Topical corticosteroid nasal spray administered in a head-down-forward position is an effective treatment of severe olfactory loss associated with severe nasal and sinus disease.

Distortion product otoacoustic emissions in normal and impaired adult ears.

Distortion product otoacoustic emissions (DPOEs) were recorded in a group of normally hearing subjects (29 ears) and a group of subjects whose conditions were diagnosed as sensorineural hearing loss (23 ears) to study any correlation that might exist between DPOE characteristics and hearing impairment of different configurations. Three different DPOE paradigms have been used to investigate the influence of different test parameters on the DPOE data for normal and hearing-impaired ears. All normally hearing ears demonstrated detectable DPOEs, provided that the primary tone level was above a certain value. Hearing-impaired ears produced substantially reduced DPOEs compared with normally hearing subjects when the primary frequencies f1 and f2 corresponded to the region of hearing loss. Our data also suggested that, in general, more than one f2/f1 ratio is needed to examine any particular frequency region. The DPOEs provide frequency-specific information about cochlear function, which after further development, may form a basis for a noninvasive, objective method of evaluating cochlear function.

Interleukin-8 expression in human nasal polyps.

The cytokine interleukin 8 (IL-8) has been shown to be a potent mediator of leukocyte recruitment and neovascularization in inflammatory and neoplastic diseases. In this study we hypothesize that IL-8 produced in the nasal polyp microenvironment is responsible for the leukocyte recruitment seen in nasal polyposis. To test this hypothesis we evaluated nasal polyps for distribution and content of IL-8 antigen with immunohistochemical techniques and radioimmunoassay to determine tissue levels of IL-8. The immunohistochemical results demonstrated that IL-8 antigen staining occurred predominantly within inflammatory cells and epithelium. IL-8 was detected in all nasal polyp tissue homogenates (a mean value of 1767 +/- 1633 pg/mg total protein (TP) with a range of 134 to 3668 pg/mg TP vs control specimens with a mean value of 77 pg/mg TP with a range of 0.09 to 255 pg/mg TP). These data demonstrate the presence and distribution and levels of IL-8 antigen in nasal polyps in vivo, supporting our hypothesis that local production of IL-8 could be an important factor in the sustained recruitment of leukocytes in nasal polyposis. Thus IL-8 likely plays a significant role in the pathogenesis of this disease process and therefore is a potential target for therapeutic intervention.

Growth-related oncogene-alpha expression in human nasal polyps.

The cytokine growth-related oncogene-alpha (GRO-alpha) is a potent mediator of leukocyte recruitment and proliferation in inflammatory diseases. We hypothesized that GRO-alpha is produced in the inflammatory nasal polyp microenvironment. Evaluation of nasal polyps from 27 patients for distribution and content of GRO-alpha antigen, by use of immunohistochemical techniques and ELISA, revealed its presence in all 27 tissue samples. It was found predominantly within the eosinophils and neutrophils, with tissue levels ranging from 34 pg/mg total protein (TP) to 1746 pg/mg TP, with a mean value of 631 +/-98 pg/mg TP. Control tissues contained between 82 pg/mg TP and 316 pg/mg TP (mean 176+/-38 pg/mg TP). These results were statistically significant (P<0.03). Clinical correlations and statistical comparisons were calculated. These data suggest that GRO-alpha may be an important factor in the recruitment and activation of leukocytes in nasal polyposis, making it a potential target for therapeutic intervention.

Characterization of the eosinophil chemokine RANTES in nasal polyps.

Previous studies have demonstrated that the cytokine RANTES (Regulated And Normal T cell Expressed and Secreted) has been shown to be a potent mediator of eosinophil chemotaxis in vitro and of leukocyte recruitment. Because eosinophils are the hallmark cells in nasal polyposis, we hypothesize that RANTES is locally produced within the nasal polyp microenvironment and is responsible for the eosinophil recruitment seen in nasal polyposis. To begin to test this hypothesis, we evaluated nasal polyps from 17 patients and 3 control specimens for distribution and content of RANTES using immunohistochemical techniques and enzyme-linked immunosorbent assay technology. Our immunohistochemical studies demonstrated that in nasal polyposis, RANTES antigen staining occurred predominantly within eosinophils and epithelial cells. To quantify the relative levels of RANTES in normal and nasal polyp specimens, tissue homogenates were prepared, quantified, and normalized to protein levels. We detected RANTES in all 17 nasal polyp tissue homogenates (566 +/- 16 pg/mg total protein). The RANTES levels in nasal polyp homogenates were nearly 40-fold higher than the RANTES levels in normal tissue (15.7 +/- 28.2 pg/mg total protein). Thus, it appears that increased expression of RANTES by eosinophils and epithelial cells within the nasal polyp microenvironment promotes eosinophil recruitment and activation within nasal polyps. We hypothesize that RANTES induces increased recruitment and activation of eosinophils, presumably contributing to the increased tissue changes associated with nasal polyposis.

Soft tissue response in the rabbit larynx following implantation of LactoSorb (PLA/PGA copolymer) prosthesis for medialization laryngoplasty.

This project is designed to provide initial data regarding the use of polylactic acid/polyglycolic acid (PLA/PGA) copolymer ("LactoSorb" [Walter Lorenz Corp]), an alloplastic, resorbable material, as a prosthesis in an animal model of vocal fold medialization. Fifteen New Zealand white rabbits were utilized for left medialization laryngoplasty with LactoSorb implants after undergoing left recurrent laryngeal nerve section. At 1, 3, 6, and 9 months, the rabbits were sacrificed and their larynges were evaluated both grossly and histologically for tissue response to, and resorption characteristics of the implant, tissue cellularity, maintenance of vocal fold medialization, and airway patency. Additionally, 4 rabbits were used as controls, implanted with silicone rubber medialization implants, and sacrificed at 9 months for comparison. One rabbit underwent no surgery and was likewise used as a control. Grossly, no airway obstruction was noted, and no extrusions of the implants occurred. The LactoSorb implant maintained medialization in each group of sacrificed rabbits. Histologic findings revealed a very discrete, fibrous capsule around the implant in the 1- and 3-month rabbits, and the LactoSorb was still grossly visible. At 6 months, the thin fibrous capsule partially remained; at 9 months, the capsule was no longer evident, and the implant was no longer grossly visible. Endoscopic findings at the time of sacrifice in those rabbits implanted with silicone rubber included grossly patent airways with maintenance of medialization. In the rabbits implanted with silicone rubber, the histologic findings are similar to those described elsewhere. LactoSorb, because of its intermittent resorption rate, could offer an ideal alternative to currently utilized temporary, or resorbable, materials, and as such will hopefully prove an invaluable tool in the laryngologist's treatment planning and surgical repair of the patient with a paralyzed vocal fold.

Identification of a novel hepatic calcium binding protein.

Calcium binding activity in the 100,000 X g supernatant of bovine liver has been isolated by a procedure involving DEAE cellulose and Sephadex G-100 chromatography. In addition to calmodulin, two new high affinity calcium binding proteins have been identified. On gel filtration chromatography these proteins migrate with apparent molecular weights of 83,700 and 51,400; whereas by sodium dodecyl sulfate polyacrylamide gel electrophoresis, the two proteins migrate identically with Mr 63,000. In the presence of millimolar Mg2+, both proteins bind up to one mol Ca2+/mol protein. Half-maximal binding occurs at approximately 0.1 microM Ca2+. Amino acid compositional analysis reveals that both proteins are acidic, and contain about 40% glx and asx. Peptide mapping procedures suggest that these proteins may be highly homologous or multiple forms of a single protein. The results show the existence of calcium binding protein(s) other than calmodulin in hepatic cytosol.

Eosinophil expression of transforming growth factor-beta and its receptors in nasal polyposis: role of the cytokines in this disease process.

PURPOSE: Nasal polyposis (NP) is characterized by an increase in inflammatory processes including fibrosis. Because transforming growth factor beta (TGF-beta) has been proven to induce fibrosis, we hypothesize that TGF-beta and its receptors are present in NP and influence polyp development. MATERIALS AND METHODS: To test this hypothesis, we evaluated distribution (immunohistochemistry) of TGF-beta isoforms (TGF-beta 1, TGF-beta 2, and TGF-beta 3) and its receptors [(TGF-beta(RI) & TGF-beta(RII)] in NP from 36 NP patients and in five normal sinus tissue specimens obtained from septoplasty/inferior turbinectomy. Tissue levels of TGF-beta 1 and TGF-beta 2 levels were determined by enzyme-linked immunosorbent assay (ELISA) and protein content was determined by Bio Rad assay (Bio Rad, Richmond, CA). All tissue levels of TGF-beta were normalized and expressed as pg of TGF-beta per mg of total protein (pg/mg TP). RESULTS: Immunohistochemical studies showed eosinophils as the major cells positively staining for TGF-beta 1, TGF-beta 2, TGF-beta 3, TGF-beta(RI), and TGF-beta. In fibrotic sections, increased staining of eosinophils, fibroblast, and mononuclear cells was found for all three isoforms and both receptors. Evaluation of tissue levels indicated mean levels of TGF-beta 1 in the NP were 11.64 +/- 22.12 pg/mg TP versus normal control mean 44.36 +/- 22.12 pg/mg TP.TGF-beta 2 mean levels were 11.46 +/- 23.73 pg/mg TP versus normal control mean of 2.03 +/- 1.13 pg/mg TP. NP showed decreased expression of TGF-beta 1 and enhanced expression of TGF-beta 2 isoforms with presence of their receptors. Higher levels of TGF-beta 2 correlated with an increase in previous polypectomies perhaps indicative of severity of disease (P < or = .0001). CONCLUSION: Our studies show the presence of the TGF-beta isoforms and receptors in NP tissue. The results support our hypothesis that the eosinophil continues to be a pivotal inflammatory cell in NP, a differential regulation may govern the activity of TGF-beta in NP, and hence, the TGF-beta family of cytokines and receptors likely play a key role in controlling NP formation.

Identification of novel calcium binding proteins of heart and brain 100,000 x g supernatant.

The chelex competitive calcium binding assay has been used to assay the calcium binding activity of the 100,000 X g supernatant of bovine heart and brain. Chromatography of brain 100,000 X g supernatant on diethylamino-ethyl (DEAE) cellulose reveals the presence of two peaks of calcium binding activity, peak I eluting at about 0.05 M NaCl and peak II at about 0.18 M NaCl. Chromatography of peak I on Sephadex G-150 resolves a major and a minor peak of calcium binding activity, at Mr 40,000 and Mr 150,000. Chromatography of peak II (0.18 M NaCl) on Sepharose 6B produces two peaks of calcium binding activity, a broad peak of calcium binding activity composed of two molecular weight species of Mr 230,000 and Mr 420,000, and a sharp peak of calcium binding activity with Mr 75,000. Chromatography of the 100,000 X g supernatant of bovine heart on DEAE Cellulose reveals two peaks of calcium binding activity. Chromatography of the lower ionic strength peak on Sephadex G-150 resolved major and minor peaks of calcium binding activity at Mr 65,000 and 150,000, respectively. The results of this study suggest the presence of several calcium binding proteins, other than calmodulin, in these tissues.

Evidence that the erythrocyte calcium pump catalyzes a Ca2+:nH+ exchange.

Treatment of whole erythrocytes with 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene (DIDS) results in inhibition of ATP and phosphate-dependent Ca2+ transport in subsequently prepared inside-out vesicles (IOV). Accumulation of phosphate into IOV in the presence of Ca2+ is virtually abolished by prior DIDS treatment, consistent with the presumed inhibition of the band III anion-exchange protein by this agent. No inhibition of Ca2+-activatable ATP hydrolysis is observed following DIDS treatment when open membranes are used to prevent development of ion gradients. This indicates that DIDS does not affect the inherent ATPase activity of the calcium pump (Waisman, D. M., Smallwood, J., Lafreniere, D., and Rasmussen, H. (1982) FEBS Lett. 145, 337-340). In IOV prepared from untreated cells, ATP-dependent Ca2+ uptake is stimulated by phosphate, sulfate, or chloride. Rates of Ca2+ uptake into DIDS-IOV are not increased by these anions. Lipid-permeable organic acids such as acetate, however, do promote Ca2+ transport in DIDS-IOV. Lipophilic anions incapable of transporting protons into the vesicle interior (nitrate and thiocyanate) support sustained uptake only when the protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone is also added. These results support a model of the (Ca2+-Mg2+)-ATPase as a pump exchanging Ca2+ for protons, not transporting Ca2+ alone. Band III protein appears to promote Ca2+ transport in the presence of phosphate, sulfate, or chloride by exchanging external anion for the accumulating OH- (or HCO3-) produced by the calcium pump.

A product from human decidua inhibits prostaglandin production by human amnion.

Human decidua was obtained from nonlaboring women after elective cesarean section. Decidua was incubated with media alone for 20 hours and this media (decidual conditioned media) was then incubated with amnion cells in monolayer culture and amnion rings. A 90% decrease in PGE2 production by amnion cells in monolayer culture was demonstrated in the presence of decidual conditioned media when compared to controls. In short term incubations with fresh amnion, decidual conditioned media decreased the production of PGE2 in amnion by greater than 25% of the control rate of production in 17 of 21 experiments. These observations suggest that human decidua produces a factor capable of inhibiting prostaglandin production by amnion.

Do blood and meconium affect the detection of endotoxin in amniotic fluid with the limulus amebocyte gel clot assay?

The limulus amebocyte lysate (LAL), assay is the most sensitive technique for the detection of endotoxin in biological fluids. Because endotoxin is a component of gram-negative bacteria, the assay has been employed in the detection of gram-negative bacterial contamination of biological fluids. The LAL assay is rapid, inexpensive, easy to perform, and requires little laboratory expertise. When used in conjunction with the gram stain examination of amniotic fluid, it improves the detection of intra-amniotic infection before the availability of culture results. However, the usefulness of the LAL assay in the detection of endotoxin in other body fluids is limited by the presence of an inhibitor to the gelation of the assay. The studies reported in this communication were undertaken to establish if amniotic fluid contains such an inhibitor. Sterile amniotic fluid (AF) samples obtained from 93 patients by transabdominal amniocentesis before labor were used to determine the ED 50 dose of endotoxin necessary for a positive LAL result. The ED 50 dose of endotoxin required for gelation was significantly higher when AF--rather than pyrogen-free saline--was used as the diluent, implying that inhibitors are in fact present (ED 50 = 58.3 pgm/ml). The presence of blood or meconium in the AF did not enhance inhibition significantly: ED 50 doses were 58.3 pgm/ml and 56.2 pgm/ml, respectively. This is not significantly different from the ED 50 of clear amniotic fluid.