Concanavalin A-mediated binding and sphering of human red blood cells by homologous monocytes.

Human red blood cells sensitized with concanavalin A became bound to homologous peripheral blood monocytes. Binding occured at a concentration of 10(5) molecules of tetrameric Con A per red blood cell (RBC) and increased with additional Con A. RBC binding began within 5 min and was maximal at 90 min. Phagocytosis of sensitized RBCs was minimal. RBC attachment was prevented by 0.01 M alpha-methyl-D-mannopyranoside, and, once the RBC-monocyte rosette was established, bound RBCs were largely removed with this specific saccharide inhibitor of Con A. RBCs attached to monocytes became spherocytic and osmotically fragile. The recognition of concanavalin A (Con A)-coated RBCs was not mediated through the monocyte IgG-Fc receptor. These studies demonstrate that, like IgG and C3b, Con A is capable of mediating the binding of human RBCs to human monocytes. Red cells so bound are damaged at the monocyte surface.

Genetic and physical interactions between Srp1p and nuclear pore complex proteins Nup1p and Nup2p.

Nup1p is a yeast nuclear pore complex protein (nucleoporin) required for nuclear protein import, mRNA export and maintenance of normal nuclear architecture. We have used a genetic approach to identify other proteins that interact functionally with Nup1p. Here we describe the isolation of seventeen mutants that confer a requirement for Nup1p in a background in which this protein is normally not essential. Some of the mutants require wild-type Nup1p, while others are viable in combination with specific nup1 alleles. Several of the mutants show nonallelic noncomplementation, suggesting that the products may be part of a hetero-oligomeric complex. One is allelic to srp1 which, although it was identified in an unrelated screen, was shown to encode a protein that is localized to the nuclear envelope (Yano, R., M. Oakes, M. Yamaghishi, J. A. Dodd, and M. Nomura. 1992. Mol. Cell. Biol. 12:5640-5651). We have used immunoprecipitation and fusion protein precipitation to show that Srp1p forms distinct complexes with both Nup1p and the related nucleoporin Nup2p, indicating that Srp1p is a component of the nuclear pore complex. The distant sequence similarity between Srp1p and the beta-catenin/desmoplakin family, coupled with the altered structure of the nuclear envelope in nup1 mutants, suggests that Srp1p may function in attachment of the nuclear pore complex to an underlying nuclear skeleton.

Yeast N1e3p/Nup170p is required for normal stoichiometry of FG nucleoporins within the nuclear pore complex.

The FG nucleoporins are a conserved family of proteins, some of which bind to the nuclear localization sequence receptor, karyopherin. Distinct members of this family are found in each region of the nuclear pore complex (NPC), spanning from the cytoplasmically disposed filaments to the distal end of the nuclear basket. Movement of karyopherin from one FG nucleoporin to the next may be required for translocation of substrates across the NPC. So far, nothing is known about how the FG nucleoporins are localized within the NPC. To identify proteins that interact functionally with one member of this family, the Saccharomyces cerevisiae protein Nup1p, we previously identified 16 complementation groups containing mutants that are lethal in the absence of NUP1 These mutants were referred to as nle (Nup-lethal) mutants. Mutants in the nle3/nlel7 complementation group are lethal in combination with amino-terminal nup1 truncation mutants, which we have previously shown to be defective for localization to the NPC. Here we show that NLE3 (which is allelic to NUP170) encodes a protein with similarity to the mammalian nucleoporin Nup155. We show that Nle3p coprecipitates with glutathione S-transferase fusions containing the amino-terminal domain of Nup1p. Furthermore, a deletion of Nle3p leads to changes in the stoichiometry of several of the XFXFG nucleoporins, including the loss of Nup1p and Nup2p. These results suggest that Nle3p plays a role in localizing specific FG nucleoporins within the NPC. The broad spectrum of synthetic phenotypes observed with the nle3delta mutant provides support for this model. We also identify a redundant yeast homolog that can partially substitute for Nle3p and show that together these proteins are required for viability.

Invading the yeast nucleus: a nuclear localization signal at the C terminus of Ty1 integrase is required for transposition in vivo.

Retrotransposon Ty1 faces a formidable cell barrier during transposition--the yeast nuclear membrane which remains intact throughout the cell cycle. We investigated the mechanism by which transposition intermediates are transported from the cytoplasm (the presumed site of Ty1 DNA synthesis) to the nucleus, where they are integrated into the genome. Ty1 integrase has a nuclear localization signal (NLS) at its C terminus. Both full-length integrase and a C-terminal fragment localize to the nucleus. C-terminal deletion mutants in Ty1 integrase were used to map the putative NLS to the last 74 amino acid residues of integrase. Mutations in basic segments within this region decreased retrotransposition at least 50-fold in vivo. Furthermore, these mutant integrase proteins failed to localize to the nucleus. Production of virus-like particles, reverse transcriptase activity, and complete in vitro Ty1 integration resembled wild-type levels, consistent with failure of the mutant integrases to enter the nucleus.

Adenotonsillectomy for upper airway obstruction carries increased risk in children with a history of prematurity.

To define better the clinical presentation and perioperative outcome in children undergoing adenotonsillectomy (T&A) for relief of upper airway obstruction (UAO), we reviewed the hospital records of 60 consecutive, otherwise normal children aged 12 years or younger. Seven patients with trisomy 21, neurologic impairments, or preoperative cor-pulmonale were excluded. Intraoperative and postoperative complications were experienced by 15 (34%) and 13 (25%), respectively, of the 53 children with preoperative UAO. The most severe complications comprised pulmonary edema and prolonged postoperative oxyhemoglobin desaturation. Multivariate logistic regression analysis found a history of prematurity and/or low birth weight to be the most significant risk factors related to the occurrence of complications. Twenty-eight % of the study population had a history of prematurity and they had approximately 85% of the perioperative complications seen in children with UAO undergoing T&A. Other significant risk factors included adenoidal facies and evidence of respiratory distress at the time of surgery. It appears that T&A poses significant risk for children with UAO who were born prematurely and have evidence of abnormal facial development or respiratory distress preoperatively.

Prognostic value of eustachian tube function in pediatric tympanoplasty.

Poor eustachian tube function and prevalence of infectious ear disease are thought to be the reasons for less successful outcome associated with tympanoplasty in children. Since both of these factors are related to age, identification of those patients who may benefit by delaying the surgery has been a concern to otolaryngologists. In an effort to investigate the role of eustachian tube function on the outcome, we tested the tubal function in 63 ears (56 children) undergoing tympanoplasty for central perforations. In 49 ears (78%), the graft took; of these, 33 had good middle-ear function, 8 developed persistent or recurrent otitis media, and 8 had severe retraction or atelectasis. There was a significant association (p less than 0.01) between outcome and preoperative tubal function as determined by combining the active and passive function parameters. However, the prognostic value of tubal function testing was low; predictive values for success and lack of success being 68% and 56%, respectively. Other factors, such as graft placement (medial or lateral) contralateral ear status, and child's age, were not associated with outcome. Consistent with other studies, good eustachian tube function was shown to predict good outcome, but poor tubal function was not helpful in predicting poor outcome.

Medical management of chronic suppurative otitis media without cholesteatoma in children.

Tympanomastoid surgery is considered standard management for chronic suppurative otitis media (CSOM) without cholesteatoma, which is unresponsive to ototopical/oral antimicrobial therapy. The following makes this sequence of management less attractive today: 1. potential ototoxicity of ototopical agents; 2. lack of oral antimicrobial agents effective against most common pathogens (e.g., Pseudomonas aeruginosa); 3. frequent occurrence in children who have tympanostomy tubes; and 4. failure of tympanomastoid surgery to eradicate the disease in all cases. We conducted a study in 36 pediatric patients with chronic suppurative otitis media, in which all received parenteral antimicrobial therapy and daily aural toilet (mean duration of treatment = 9.7 days). Thirty-two patients (89%) had resolution of their infection with medical therapy alone; four children required tympanomastoidectomy. Further investigation is needed to understand the etiology, pathogenesis, and most effective methods of management/prevention of CSOM in children.

Nasal surgery in children with cystic fibrosis: complications and risk management.

Children with cystic fibrosis (CF) are predisposed to pansinusitis and nasal polyposis and, therefore, require frequent surgery by otolaryngologists. These children are at risk to have complications following surgery, both locally, at the surgical site, as well as systemically from their underlying pulmonary disease. A 20-year retrospective study of children (average census 219 per year) revealed 39 children developed nasal polyps and these children required 85 nasal polypectomies. No major complications occurred and only three minor complications (fever, minor bleeding) were noted. Over 95% of our CF children were able to return home in less than 48 hours, and extensive hospital stays appeared not to be required.

The relationship between dental overbite and eustachian tube dysfunction.

OBJECTIVE: The purpose of this study was to investigate the association between deep dental overbite and eustachian tube dysfunction (ETD). DESIGN: Case-control study. SETTING: Tertiary care pediatric otolaryngology outpatient clinic at the Children's Hospital, Boston, Massachusetts. PATIENTS: 105 patients between the ages of 2 and 6 years. STUDY MEASUREMENTS: Dental overbite, overjet, and occlusal relationships were measured by an observer who was unaware of ETD status. ETD was defined as having ventilation tubes in place or having the recommendation for ventilation tube placement by an attending pediatric otolaryngologist. In addition, demographic information and medical and social histories were prospectively recorded. RESULTS: In a multivariate logistic regression model, children with deep bites were 2.8 times more likely to have ETD than those without deep bites (P = .03). Other independent risk factors for ETD identified in this model were family history of otitis media (OM) and age less than 3 years. CONCLUSIONS: Children with deep dental overbites are at a significantly increased risk for developing ETD.

Sensorineural hearing loss from quinolinic acid: a neurotoxin in middle ear effusions.

Quinolinic acid (QUIN) is an endogenous metabolite that exerts a neurotoxic effect by binding to specific neuronal receptors. Studies involving a broad spectrum of infectious and inflammatory central nervous system diseases have suggested a role for QUIN in causing neuronal injury. Since there is evidence for presence of the QUIN receptor in mammalian cochleas, QUIN was measured in middle ear effusions (MEEs). Gas chromatography/mass spectrometry detected QUIN in each of 65 diluted human MEEs, with a mean of 482 +/- 75 (SEM) nmol/L and a range from 15 to 2667 nmol/L. QUIN was also detected in each of 197 chinchilla MEEs from five different models of otitis media, with a mean of 10.6 +/- 1.3 (SEM) mumol/L and a range from 0.23 to 146.0 mumol/L (corrected for dilution). To determine whether QUIN causes sensorineural hearing loss (SNHL), QUIN solutions were placed on round window membranes (RWM) for 20 to 240 minutes, in 20 chinchillas. SNHL was detected by electrocochleography in QUIN-exposed animals, but not in saline controls. We conclude that QUIN is present in MEEs and that QUIN in the middle ear has the potential to cross the RWM and cause sensorineural hearing loss, possibly by binding to specific neuronal receptors in mammalian cochleas.

Causes of pediatric sensorineural hearing loss: yesterday and today.

OBJECTIVE: To ascertain the present common causes of sensorineural hearing loss (SNHL) in children and compare them with those of previous reports. DESIGN: A retrospective review of the medical records for all children with a diagnosis of SNHL seen from January 1, 1993, through September 30, 1996, at our institution. SETTING: A tertiary care children's hospital. PATIENTS: Three hundred one children, aged 1 week through 18 years, who presented for evaluation of SNHL. RESULTS: Of the 301 children, 68.1% had a definite or probable cause of their SNHL identified; 18.9%, 1 or more possible causes; and 31.9%, no obvious cause. A family history of SNHL or prematurity and/or complicated perinatal course was found in 28.6% of patients. Named syndromes, multiple congenital anomalies, meningitis, or prenatal maternal factors, including maternal prenatal substance abuse, were present in another 38.5%. However, syndromes commonly reported to be associated with SNHL, such as Waardenburg syndrome, were seen in less than 1% of patients. The average age at diagnosis was 3.02 years for the bilateral moderate or worse SNHL; for unilateral SNHL, the average age was 3.97 years. The most useful diagnostic study was computed tomographic scanning. CONCLUSIONS: Sensorineural hearing loss is fairly common in children. Extensive workups, often without clear direction, should be reconsidered based on the children with SNHL who otolaryngologists are now seeing. Infant screening programs, although identifying many children earlier, will also provide the opportunity to fine-tune the evaluation (ie, cytomegalovirus titers and/or cultures at birth), increasing the diagnostic yield.

Therapeutic implications in the treatment of aural Pseudomonas infections based on in vitro susceptibility patterns.

OBJECTIVE: To examine the in vitro susceptibility patterns of aural isolates of Pseudomonas aeruginosa and to identify changes over a 4-year period. DESIGN: Retrospective case series. SETTING: The outpatient department at Children's Hospital of Pittsburgh (Pa), a tertiary referral center. PATIENTS: Ambulatory children younger than 18 years from whose ears P aeruginosa was isolated. OUTCOME MEASURES: The in vitro susceptibility of aural isolates of P aeruginosa to ampicillin, cefotaxime, chloramphenicol, sulfisoxazole, ticarcillin, mezlocillin, gentamicin, tobramycin, cefazolin, tetracycline, piperacillin, nitrofurantoin, cephalexin hydrochloride, ceftriaxone, cefuroxime axetil, and sulfamethoxazole-trimethoprim. RESULTS AND CONCLUSIONS: No changes were found in the trends of the susceptibility patterns over the 4-year study period, with the exception of the semisynthetic penicillins, ticarcillin and mezlocillin. These two agents were found to be relatively ineffective against the strains of P aeruginosa isolated in 1989 (59% and 18% susceptibility, respectively). This finding is in contrast to their effectiveness over the remainder of the study period (96% and 90% susceptibility, respectively), which was excellent. These observations likely reflect a change in the breakpoints for the minimal inhibitory concentrations between these periods. The intravenous agent with the best susceptibility profile was piperacillin (96%). Of the aminoglycosides tested, 94% of the isolates were sensitive to tobramycin, as opposed to only 79% for gentamicin. This finding may have significance when one is empirically selecting ototopical therapy, since both tobramycin and gentamicin are available as topical preparations. Of the oral agents, the combination of sulfamethoxazole-trimethoprim was most effective (46%).

Connexin 26 studies in patients with sensorineural hearing loss.

OBJECTIVE: To determine the spectrum of connexin 26 (Cx26) mutations and their phenotypes in children with sensorineural hearing loss (SNHL) or mixed hearing loss (MHL). DESIGN: Children with SNHL or MHL were prospectively tested for mutations in the entire coding region of the Cx26 gene. PATIENTS: Children with SNHL or MHL with no obvious etiology for the hearing loss. RESULTS: Between December 1, 1998, and July 1, 2000, 107 patients with SNHL or MHL from 99 families underwent Cx26 testing. Most patients were aged 1 week to 16 years (61 boys and 46 girls). Thirty (30%) of 99 probands had Cx26 mutations: biallelic mutations were detected in 18 (9 homozygous and 9 compound heterozygous) and single mutations were detected in 12. Twelve previously reported mutations (35delG, 167delT, E47X, L90P, M34T, G12V, V37I, R143W, V84L, V153I, V27I, and 310del14) and 3 novel mutations (E129K, T8M, and N206S) were found. Hearing loss in patients with biallelic Cx26 mutations ranged from unilateral high frequency to bilateral profound. Four children, 2 with biallelic mutations, had temporal bone abnormalities. CONCLUSIONS: Connexin 26 mutations are common in children with SNHL, and it is likely that the homozygous and compound heterozygous mutations cause the SNHL. However, pathogenicity is less certain when only a single Cx26 mutation is present. Patients with biallelic Cx26 mutations had a slightly higher incidence of milder hearing loss than in previous studies. Children with SNHL or MHL should be tested for Cx26 mutations early in their evaluation.

Craniofacial, temporal bone, and audiologic abnormalities in the spectrum of hemifacial microsomia.

OBJECTIVES: To evaluate the clinical, audiologic, and temporal bone computed tomograpic findings in patients with hemifacial microsomia and to use the OMENS (each letter of the acronym indicates 1 of the following 5 dysmorphic manifestations: O, orbital asymmetry; M, mandibular hypoplasia; E, auricular deformity; N, nerve involvement; and S, soft tissue deficiency) grading system to assess possible correlations between the severity of dysmorphic features with the type of abnormalities in the temporal bone and with degree of hearing deficit. DESIGN: Retrospective study. SETTING: Tertiary care children's hospital. PATIENT: Forty patients with hemifacial microsomia. RESULT: Mandibular hypoplasia and auricular abnormalities were the most common clinical manifestations, present in 39 patients (97%) and 38 patients (95%), respectively. Conductive hearing loss was noted in 35 patients (86%) and sensorineural hearing loss in 4 patients (10%). Facial nerve weakness was present in 20 patients (50%). Twenty patients had unilateral aural atresia, 12 patients had unilateral aural stenosis, and 7 patients had bilateral anomalies. Moderate hypoplasia or atresia of the middle ear was noted in 36 patients (90%) and ossicles were malformed in 30 patients (75%). Hypoplasia of the oval window was the most common inner ear abnormality. CONCLUSIONS: Severity of craniofacial features (total OMENS score) significantly correlated with the degree of temporal bone abnormality, but no correlation was noted with the degree or type of hearing loss. We recommend the following: (1) use of the OMENS classification system for documentation and analysis of dysmorphic finding in hemifacial microsomia; (2) complete audiologic evaluation in all patients with hemifacial microsomia regardless of the type of craniofacial abnormalities; and (3) temporal bone computed tomography for further evaluation of hearing deficit.

Duration of intubation in children with acute epiglottitis.

Nasotracheal intubation has been demonstrated to be effective in supporting the airways of children with acute epiglottitis. Length of intubation and criteria used for extubation are still controversial. A 6-year retrospective review at Children's Hospital of Pittsburgh identified 100 cases of acute epiglottitis, which were initially managed with nasotracheal intubation. Extubation was based on direct laryngeal inspection performed in the operating room (1979-1981) and, more recently, in the intensive care unit (1982-1984). Length of intubation decreased from 63.8 hours in 1979 to 42.1 hours in 1984. The percent of children intubated longer than 48 hours decreased from 69% to 22% in the same time period. These data indicate that a shorter period of intubation is aided by daily laryngeal inspection in the ICU. We propose a staging system for acute epiglottitis to aid in the decision to safely extubate these children.

Head and neck manifestations of Beckwith-Wiedemann syndrome.

Beckwith-Wiedemann syndrome is a congenital disorder manifested by organomegaly, omphalocele, hypoglycemia, and macroglossia. We have found a significant number of these children to be at risk for upper airway obstruction during infancy or childhood. In this review of 13 children, 2 required tracheotomy during infancy for cor pulmonale caused by macroglossia. Seven of nine children older than 1 year required tonsillectomy and adenoidectomy to relieve upper airway obstruction. Although macroglossia can be a cause of airway obstruction in infants with Beckwith-Wiedemann syndrome, we have found that airway obstruction during childhood is related to tonsillar and adenoidal hypertrophy and not to macroglossia. Anterior tongue reduction is reserved for the correction of malocclusion, articulation errors, or cosmesis, whereas tonsillectomy and adenoidectomy may be curative of obstructive symptoms.

Surgical management of tracheal stenosis in an infant with multiple congenital anomalies: when is baby inoperable?

Congenital tracheal stenosis is a rare disorder that carries a significant morbidity and mortality. It is an even greater challenge when present in combination with other life-threatening congenital anomalies. We present an infant with multiple congenital anomalies, including trisomy 21, who was diagnosed with congenital tracheal stenosis at 3 months of age. At age 5 months, the infant underwent tracheoplasty with costal cartilage graft and pulmonary artery banding. Bronchoscopy 3 weeks later revealed a patent airway. However, the patient died 2 months postoperatively of cardiac complications. At autopsy, whole organ and histologic specimens revealed excellent incorporation of the graft with no evidence of re-stenosis. Our documentation of graft incorporation in this patient is evidence that long segments of stenotic trachea can be successfully treated by tracheoplasty utilizing rib cartilage grafts.

Tracheotomy in the preterm infant.

Over the last decade, prolonged survival of preterm infants (gestation less than or equal to 36 weeks) who require lengthy periods of mechanical ventilation has necessitated that many of these infants undergo tracheotomy. The complication rate for tracheotomy in these preterm infants has not been reported. We compared 83 full-term (FT) infants who underwent tracheotomy in their first year of life with 41 preterm infants. Twenty-three preterm infants had birth weight greater than or equal to 1,500 g (PT), and 18 of the preterm infants had gestational age less than or equal to 32 weeks and birth weight less than or equal to 1,500 g (PT-VLBW). Early complications (day 0 to 7) occurred in over 50% of the PT-VLBW compared to only 24% of the FT infants. Late complication rates were similar for all three groups. This higher early complication rate for PT-VLBW infants may be related to gestational age, low birth weight, and medical condition rather than surgical technique.

Treatment of chronic suppurative otitis media with topical ciprofloxacin.

To date, only ofloxacin has been approved by the US Food and Drug Administration for treatment of ears with a nonintact tympanic membrane. The purpose of this study was to determine the safety and efficacy of topical ciprofloxacin hydrochloride in the treatment of experimental chronic suppurative otitis media caused by Pseudomonas aeruginosa infection in cynomolgus monkeys. Forty adult cynomolgus monkeys were divided into 4 equal groups, and their ears were challenged with P aeruginosa, drained for 3 weeks, then treated twice daily for 4 weeks with 1 of 4 randomly assigned agents: 1) ciprofloxacin, 2) saline, 3) Cortisporin, or 4) vehicle. The animals were followed up with auditory brain stem response testing, culture, otoscopy, and histopathology. Both ciprofloxacin and Cortisporin treatment resulted in a significantly more rapid rate of clearance of P aeruginosa as compared to treatment with saline (100% versus 20%). Eradication was not associated with resolution of otorrhea after a 4-week period of treatment. There were no significant changes in auditory brain stem response wave latencies for any of the treatment groups. Histopathologic data revealed that there was no statistically significant difference in the amount of outer hair cell loss for the ciprofloxacin group as compared to the control ear and other treatment groups. We conclude, therefore, that topical ciprofloxacin is not ototoxic and is effective in sterilizing the otorrhea, but does not promote resolution of the drainage, in this animal model.

Tidal flow measurement in the decision to decannulate the pediatric patient.

Measurements of peak inspiratory flow obtained through the tracheostomy cannula (MIFT) during tidal breathing were compared to peak inspiratory flow measurements obtained through the mouth (MIFM) in 40 children to assess physiologic readiness to decannulate the tracheostomized pediatric patient. Ratio of peak flow MIFM/MIFT was 1.40 for 34 successfully decannulated children compared to 0.83 for 22 unsuccessful attempts (p less than 0.01). Tidal flow measurements are highly predictive (84%) in identifying children who are unlikely to be ready for decannulation. A schema is proposed to utilize tidal flow measurements as the first step in the decannulation process.