Cross-sectional follow-up of voice outcomes in children who have a history of airway reconstruction surgery.

OBJECTIVES: This study reports vocal function in a cross-section of children with subglottic stenosis. Each child had a history of laryngotracheal reconstruction and/or cricotracheal resection surgery. Vocal function was measured using laryngoscopy, acoustic analysis, perceptual evaluation and impact of voice on quality of life. DESIGN: All patients aged >5 years with history of laryngotracheal reconstruction and/or cricotracheal resection surgery at the Scottish National Complex Airways service were invited to participate. SETTING: Data were gathered in the Royal Hospital for Children in Glasgow in a single outpatient appointment. PARTICIPANTS: Twelve of 56 former patients (aged 5-27) provided a voice sample and eleven consented to awake laryngoscopy. All consented for detailed evaluation of their medical records. MAIN OUTCOME MEASURES: Acoustic analysis of fundamental frequency and pitch perturbation was conducted on sustained vowel [a]. Perceptual evaluation was conducted by 4 trained listeners on a series of spoken sentences. Impact on quality of life was measured using the paediatric voice-related quality of life questionnaire. Laryngeal function was descriptively evaluated. RESULTS: Four children had normal voice acoustically, perceptually and in relation to voice-related quality of life. One of these had vocal fold nodules unrelated to surgical history. Two other children had "near normal" vocal function, defined where most voice measurements fell within the normal range. CONCLUSIONS: Normal or "near normal" voice is a possible outcome for children who have had this surgery. Where there is an ongoing complex medical condition, voice outcome may be poorer.

Observations of the marginal band system of nucleated erythrocytes.

The marginal band (MB) of nucleated erythrocytes (thos of nonmammalian vertebrates) is a continuous peripheral bundle of microtubules normally obscured by hemoglobin. Treatment of these elliptical cells with modified microtubule polymerization media containing Triton X-100 yields a semilysed system in which MB, nucleus, and trans-MB material (TBM) are visible under phase contrast. The TBM apparently interconnects structural components, passing around opposite sides of the nucleus and suspending it in native position. In uranyl acetatestained whole whole mounts (goldfish) examined by transmission electron microscopy, the TBM appears as a network. MBs of semilysed cells are relatively planar initially, but twist subsequently into a range of "figure-8" shapes with one of the two possible mirror-image configurations predominant. Nuclei and MBs can be released using proteolytic enzymes, to which the TBM seems most rapidly vulnerable. MBs thus freed are birefringent, generally untwisted, and much more circular than they are in situ. As a working hypothesis, it is prosposed that the flattened, elliptical shape of nucleated erythrocytes is a result of TBM tension applied asymmetrically across an otherwise more circular MB, and that the firure-8 configuration occurs when there is extreme TBM shrinkage or contraction.

Association of centrioles with the marginal band of a molluscan erythrocyte.

Continuous circumferential bundles of microtubules, or marginal bands (MBs), are best known as a prominent structural feature of all nonmammalian vertebrate erythrocytes and mammalian blood platelets. Since their discovery in the late 19th century, MBs have been thought to play a cellular morphogenetic role, but no cytological clues to the mechanism of MB biogenesis have been reported. In previous work we have established the presence of MBs in serveral invertebrate blood cell types, including amebocytes and coelomocytes of certain Arthropod species and erythrocytes of a Sipunculan. We report here the occurrence of MBs in erythrocytes of the ark Anadara transversa (Mollusca) and four closely related species. The MBs of these arks have a striking structural feature; each is physically associated with a pair of centrioles. The centrioles are identified as such on the basis of morphological criteria: size, cylindrical shape, right-angle orientation, pairing, and 9-triplet ultrastructure. This intimate association between centrioles and MBs suggests that centrioles may be MB-organizing centers and invites comparative investigation of their possible role in vertebrate erythrocyte and platelet morphogenesis.

Centrioles as microtubule-organizing centers for marginal bands of molluscan erythrocytes.

The erythrocytes of blood clams (arcidae) are flattened, elliptical, and nucleated. They contain elliptical marginal bands (MBs) of microtubules, each physically associated with a pair of centrioles marginal bands (MBs) of microtubles, each physically associated with a pair of centrioles (Cohen, W., and I. Nemhauser, 1980, J. Cell Biol., 86:286-291). The MBs were found to be cold labile in living cells, disappearing within 1-2 h at 0 degrees C. After the cells had been rewarmed for 1-2 h, continuous MBs with associated centrioles were once again present. Time-course studies utilizing phase contrast, antitubulin immunofluorescence, and electron microscopy of cytoskeletons prepared during rewarming revealed structural evidence of centriole participation in MB reassembly. At the earliest stage of reassembly, a continuous MB was not present. Instead, relatively short and straight microtubules focused on a pointed centriolar "pole," and none were present elsewhere in the cytoskeleton. Thin continuous MBs then formed, still pointed in the centriolar region. Subsequently, the MBs regained ellipticity, with their thickness gradually increasing but not reaching that of controls even after several hours of rewarming. At these later time points, microtubules still radiated from the centrioles and joined the MBs some distance away. In the presence of 0.1 mM colchicines, MB reassembly was arrested at the pointed stage. Electron microscopic observations indicate that pericentriolar material is involved in microtubule nucleation in this system, rather than the centriolar triplets directly. The results suggest a model in which the centrioles and associated material nucleate assembly and growth of microtubules in diverging directions around the cell periphery. Microtubules of opposite polarity would then pass each other at the end of the cell distal to the centrioles, with continued elongation eventually closing the MB ellipse behind the centriole pair.

The cytoskeletal system of nucleated erythrocytes. III. Marginal band function in mature cells.

Marginal bands (MBs) of microtubules are believed to function during morphogenesis of nonmammalian vertebrate erythrocytes, but there has been little evidence favoring a continuing role in mature cells. To test MB function, we prepared dogfish erythrocytes with and without MBs at the same temperature by (a) stabilization of the normally cold-labile MB at 0 degree C by taxol, and (b) inhibition of MB reassembly at room temperature by nocodazole or colchicine. We then compared the responses of these cells to mechanical stress by fluxing them through capillary tubes. Before fluxing , cells with or without MBs had normal flattened elliptical shape. After fluxing , deformation was consistently observed in a much greater percentage of cells lacking MBs. The difference in percent deformation between the two cell types was highly significant. That the MB is an effector of cell shape was further documented in studies of the formation of singly or doubly pointed dogfish erythrocytes that appear during long-term incubation of normal cells at room temperature. On-slide perfusion experiments revealed that the pointed cells contain MBs of corresponding pointed morphology. Incubation of cells with and without MBs showed that they become pointed only when they contain MBs, indicating that the MB acts as a flexible frame which can deform and support the cell surface from within. To test this idea further, cells with and without MBs were exposed to hyperosmotic conditions. Many of the cells without MBs collapsed and shriveled , whereas those with MBs did not. The results support the view that the MB has a continuing function in mature erythrocytes, resisting deformation and/or rapidly returning deformed cells to an efficient equilibrium shape in the circulation.

The cytoskeletal system of nucleated erythrocytes. II. presence of a high molecular weight calmodulin-binding protein.

Calmodulin was detected in dogfish erythrocyte lysates by means of phosphodiesterase activation. Anucleate dogfish erythrocyte cytoskeletons bound calmodulin. Binding of calmodulin was calcium-dependent, concentration-dependent, and saturable. Cytoskeletons consisted of a marginal band of microtubules containing primarily tubulin, and trans-marginal band material containing actin and spectrinlike proteins. Dogfish erythrocyte ghosts and cytoskeletons were found to contain a calcium-dependent calmodulin-binding protein, CBP, by two independent techniques: (a) 125I-calmodulin binding to cytoskeletal proteins separated by SDS PAGE, and (b) in situ azidocalmodulin binding in whole anucleate ghosts and cytoskeletons. CBP, with an apparent molecular weight of 245,000, co-migrated with the upper band of human and dogfish erythrocyte spectrin. CBP was present in anucleate ghosts devoid of marginal bands and absent from isolated marginal bands. CBP therefore appears to be localized in the trans-marginal band material and not in the marginal band. Similarities between CBP and high molecular weight calmodulin-binding proteins from mammalian species are discussed.

The cytoskeletal system of nucleated erythrocytes. I. Composition and function of major elements.

We have studied the dogfish erythrocyte cytoskeletal system, which consists of a marginal band of microtubules (MB) and trans-marginal band material (TBM). The TBM appeared in whole mounts as a rough irregular network and in thin sections as a surface-delimiting layer completely enclosing nucleus and MB. In cells incubated at 0 degrees C for 30 min or more, the MB disappeared but the TBM remained. MB reassembly occurred with rewarming, and was inhibited by colchicine. Flattened elliptical erythrocyte morphology was retained even when MBs were absent. Total solubilization of MB and TBM at low pH, or dissolution of whole anucleate cytoskeletons, yielded components comigrating with actin, spectrin, and tubulin standards during gel electrophoresis. Mass-isolated MBs, exhibiting ribbonlike construction apparently maintained by cross-bridges, contained four polypeptides in the tubulin region of the gel. Only these four bands were noticeably increased in the soluble phase obtained from cells with 0 degrees C-disassembled MBs. The best isolated MB preparations contained tubulin but no components comigrating with high molecular weight microtubule-associated proteins, spectrin, or actin. Actin and spectrin therefore appear to be major TBM constituents, with tubulin localized in the MB. The results are interpreted in terms of an actin- and spectrin-containing subsurface cytoskeletal layer (TBM), related to that of mammalian erythrocytes, which maintains cell shape in the absence of MBs. Observations on abnormal pointed erythrocytes containing similarly pointed MBs indicate further that the MB can deform the TBM from within so as to alter cell shape. MBs may function in this manner during normal cellular morphogenesis and during blood flow in vivo.

Elevated 1,25-dihydroxyvitamin D plasma levels in normal human pregnancy and lactation.

Plasma 1,25-dihydroxyvitamin D levels are elevated in early pregnancy and continue to increase throughout pregnancy. They remain elevated postpartum in lactating women. The elevated levels probably represent a physiological response to increased calcium requirements.

Evidence for selective accumulation of intrathyroidal T lymphocytes in human autoimmune thyroid disease based on T cell receptor V gene usage.

We have investigated the T cell receptor V alpha and V beta gene family usage by T lymphocytes infiltrating affected thyroids in patients with autoimmune thyroid disease. We show that the intrathyroidal T lymphocytes from patients (n = 6) with autoimmune thyroid disease display a widespread usage of V beta gene families with an average of 14.4/19 V beta gene families similar to the peripheral T lymphocytes of the same patients. Because we recently reported that the utilization of V alpha gene families is markedly reduced within these mitogen-stimulated intrathyroidal T cell populations, as well as within intact tissue from similar patients (n = 4) (overall mean of 4.0/18 families detected), these results indicate that in thyroids of patients with autoimmune thyroid disease the lymphocytes are selectively accumulating based on their V alpha rather than V beta elements. This preferential hTcR V alpha and widespread V beta gene usage was not mimicked in most 7-d autologous mixed lymphocyte reactions using non-T cell stimulators (n = 6) or EB-virus immortalized autologous B cell lines (n = 3). Hence, the selective V gene utilization by intrathyroidal T cells is likely to be secondary to multiepitopic thyroidal autoantigens activating thyroid infiltrating T cells or to the presence of a superantigenlike thyroidal self-antigen, capable of determining a selective infiltration or activation of a variety of T lymphocytes on the basis of their V alpha gene usage.

Correlation of MR imaging findings with intraoperative findings after cervical spine trauma.

BACKGROUND AND PURPOSE: There are limited data correlating MR imaging and anatomic findings of ligamentous injury in cervical spine trauma. This study compares acute MR imaging with surgical observations of disk/ligamentous injury after blunt cervical trauma. MATERIALS AND METHODS: Consecutive patients with acute cervical spine trauma who underwent preoperative MR imaging and surgery from 1998 to 2001 were identified. MR imaging was obtained within 48 hours of injury for most patients. All scans included sagittal T1, T2 fat-saturated, and short tau inversion recovery sequences. At surgery, extent of injury at the operated level was recorded on a standardized form for either anterior or posterior structures or both depending upon the operative approach. MR examinations were separately evaluated by 2 readers blinded to the intraoperative findings. Radiologic and surgical findings were then correlated. RESULTS: Of 31 patients, an anterior surgical approach was chosen in 17 patients and a posterior approach in 13 patients. In one patient anterior and posterior approaches were utilized. Seventy-one percent of patients had spinal cord injury on MR imaging. MR imaging was highly sensitive for injury to disk (93%), posterior longitudinal ligament (93%), and interspinous soft tissues (100%), but it was less sensitive for injury to the anterior longitudinal ligament (71%) and ligamentum flavum (67%). For most ligamentous structures, there was limited agreement between specific MR imaging findings and injury at surgery. CONCLUSION: In acute cervical spine trauma, MR imaging has moderate to high sensitivity for injury to specific ligamentous structures but limited agreement between specific MR imaging findings and injury at surgery. MR imaging may overestimate the extent of disruptive injury when compared with intraoperative findings, with potential clinical consequences.

Labour characteristics and neonatal Erb's palsy.

OBJECTIVE: To determine risk factors for Erb's palsy, with a focus on graphic labour patterns. DESIGN: A case-control study. SETTING: New York City. SAMPLE: A total of 45 consecutive cases of Erb's palsy and 90 controls. METHODS: Pregnancies and labours of neonatal Erb's palsy cases were compared with 90 controls using univariate and multiple logistic regression analysis. MAIN OUTCOME MEASURES: Erb's palsy and shoulder dystocia. RESULTS: Mothers of children with Erb's palsy had a higher term body mass index and more gestational diabetes than those of controls. Even cases without diabetes had higher blood glucose values after a 50-g glucose challenge than did controls. Cases had a higher birthweight and a lower ratio of head-to-thoracic circumference than controls. Shoulder dystocia occurred in 67% of cases and in 2% of controls (P = 0.001). Only 46% of labours had a completely normal dilatation pattern. In a multiple logistic regression model, variables independently associated with brachial plexus injury were long deceleration phase of labour, long second stage, high birthweight, black race, and high neonatal or maternal body mass. CONCLUSIONS: Erb's palsy was frequently preceded by abnormal labour and shoulder dystocia; however, a substantial proportion of cases occurred after normal labour and delivery. Predictive models will be necessary to determine to what extent careful monitoring of the terminal portion of dilatation and of fetal descent and incorporation of maternal body mass and race (all independent risk factors in this study) will help identify fetuses at risk for brachial plexus palsy.

Inhibition of photosystem II by uncouplers at alkaline pH and its reversal by artificial electron donors.

When chloroplasts are aged for 5 min at pH 9.6, or are exposed to uncouplers at pH 8.5-9.0, electron flow from water to Hill acceptors is inhibited. Both treatments induce rapid millisecond dark decay of delayed light emission. 3-(3,4-Dichlorophenyl)-1,1-dimethylurea-sensitive electron transport through Photosystem II can be regenerated in both types of inhibited chloroplasts by the artificial electron donor, 1, 5-diphenylcarbohydrazide. Neither treatment inhibits electron flow through Photosystem I. Uncouplers at alkaline pH, when added in the light, are less effective in producing the inhibition than when added in the dark. These results are interpreted as indicating inhibition of the oxygen-evolving apparatus by alkaline intrathylakoid pH.

Hyperglycemia suppresses the adrenal medullary response to hypoxemia.

An in vivo microdialysis technique was used to study adrenal medullary function in 6 euglycemic and 6 hyperglycemic anesthetized Sprague-Dawley rats exposed to hypoxia. After stabilization of an adrenal dialysis probe, dialyzable adrenal epinephrine and norepinephrine were measured in response to 15 min of 7.8% oxygen breathing in both groups. In euglycemic rats, hypoxia increased epinephrine and norepinephrine by 650 and 320% above baseline, respectively. During hyperglycemia, (mean plasma glucose level 30.0 mM) epinephrine and norepinephrine rose only 119 and 104%, respectively. The catecholamine increase in the hyperglycemic rats was significantly attenuated in comparison to euglycemic controls (epinephrine, P = 0.0232; norepinephrine, P = 0.0079). These data demonstrate that acute hyperglycemia has the capacity to suppress the normal adrenal medullary response to hypoxemia.

High plasma norepinephrine concentrations at birth in infants of diabetic mothers.

Analysis of plasma norepinephrine (NE) concentrations in umbilical artery and vein from infants of diabetic and nondiabetic mothers revealed high plasma NE values in those of diabetic mothers. While birth weight and arterial plasma NE did not correlate in infants of nondiabetic mothers (r = 0.07, NS), birth weight and plasma NE were related inversely in infants of diabetic mothers (r = -0.73, P less than 0.05).

Medial Occipital Lobe Hyperperfusion Identified by Arterial Spin-Labeling: A Poor Prognostic Sign in Patients with Hypoxic-Ischemic Encephalopathy.

Hypoxic-ischemic encephalopathy carries an uncertain prognosis. We sought to retrospectively assess the prognostic value of arterial spin-labeling MR imaging in 22 adult patients diagnosed with hypoxic-ischemic encephalopathy. Quantitative CBF maps were generated from the M0 map, and arterial spin-labeling data on a per-voxel basis were regionally interrogated via visual inspection and ROI placement. Hyperperfusion was defined as regional increases in CBF of >20% (relative to global CBF) and/or >100 mL/100 g/min. Eleven of 22 patients had prominent bilateral medial occipital lobe hyperperfusion, all of whom died before hospital discharge. One patient who had nondistinct arterial spin-labeling hyperperfusion and restricted diffusion survived. Medial occipital lobe hyperperfusion is a distinctive pattern that merits prospective investigation in a cohort of patients with moderate hypoxic-ischemic encephalopathy to determine its predictive ability in patients with a higher likelihood of survival.

Detergent-based isolation of marginal bands of microtubules from nucleated erythrocytes.

We have developed a new, detergent-based method for the isolation of marginal bands (MBs) of microtubules from non-mammalian vertebrate erythrocytes. The critical step in MB isolation is selective removal of the "membrane skeleton" network (MS), within which the MB is enclosed. To test potential MS solubilizing agents systematically, we prepared dogfish (Mustelus canis) erythrocyte cytoskeletons in the presence of protease inhibitors and stored them at -20 degrees C in medium containing 50% glycerol and 10 microM taxol to stabilize the MB. Using this as a standard starting material, we found that low concentrations of sodium dodecyl sulfate (SDS) (0.025-0.1%) in the presence of Triton X-100 (0.1-0.4%) released both MBs and nuclei intact from cytoskeletons. Either detergent alone was ineffective. MB release from cytoskeletons was blocked by excess Triton X-100 and slowed by glycerol, and this was useful for stopping the release reaction during quantitative time-course studies. Most MBs (greater than 90%) were liberated from cytoskeletons in 5 to 30 min, depending upon detergent concentrations and other conditions, and they were sufficiently stable for mass isolation by differential centrifugation. Added standard proteins were not proteolyzed during MB release, nor was release blocked by protease inhibitors, indicating that endogenous proteases were not involved. As observed in thin sections and negatively stained whole mounts (transmission electron microscopy) and in critical-point dried preparations (scanning electron microscopy), the isolated MBs consisted principally of bundled microtubules, with some additional adhering material. SDS-polyacrylamide gel electrophoresis showed the isolated MBs to be composed primarily of four tubulin region polypeptides, with the same stoichiometry as in the whole cytoskeleton. As determined by immunofluorescence microscopy, isolated MBs bound antibody to both chicken brain and erythrocyte tau, in addition to anti-tubulin. Thus, proteins of the tau family may be involved in bundling of MB microtubules. Unlike previous MB isolation methods, that employed here is applicable to erythrocytes of diverse species, including the marine toad (Bufo marinus) and the chicken (Gallus domestica), both of which should be of value for comparative studies.

Adrenal secretion of catecholamines during hypoxemia in fetal lambs.

A new method for quantifying adrenal secretory function in chronically catheterized fetal lambs was developed. This preparation included insertion of a catheter distally into the fetal left renal vein and placement of a remotely operated, hydraulically controlled choker around the renal vein at its junction with the vena cava. With the choker open, adrenal venous blood flowed into the renal vein and then into the vena cava. With the choker closed, adrenal blood flowed into the catheter so that timed samples of adrenal venous effluent could be obtained. With this technique, left adrenal secretory rates of norepinephrine and epinephrine were determined across a spectrum of oxygen concentrations in five near-term fetal lambs. There was a rapid rise in norepinephrine secretion after induction of hypoxemia. Maximum secretory rates occurred at about 5 min, concomitantly with the lowest attained fetal arterial partial pressure of oxygen (PO2s). There was an inverse exponential relation between these catecholamine secretion rates and fetal arterial PO2 (P less than 0.001). Norepinephrine secretion appeared to increase in response to lesser degrees of hypoxemia than did epinephrine, although a longer time delay between stimulation and epinephrine response may have been a factor. Overall, norepinephrine secretion was greater than that of epinephrine. The ratios of norepinephrine to epinephrine in individual adrenal samples varied considerably and in some instances were less than one. These ratios did not correlate significantly with the degree of hypoxemia. During 25 min of a relatively steady state of hypoxemia, norepinephrine secretion declined markedly after about 5 min, although it remained above control levels throughout. Epinephrine secretion rose more gradually but then was relatively stable during the remaining period of hypoxemia.

Neurologic complications in long-standing nephropathic cystinosis.

The central nervous system has been considered to be uninvolved in nephropathic cystinosis. Survival into adulthood, following renal dialysis and transplantation, has brought attention to the sequelae of long-standing cystinosis. We examined 14 patients with cystinosis, 12 of whom had undergone renal transplantation. Two patients had neurologic symptoms. One patient had progressive bradykinesia, dementia, and spasticity with computed tomographic scan evidence of cerebral atrophy and multifocal mineralization in bilateral internal capsules and periventricular white matter. One patient had behavioral and, to a lesser extent, cognitive disturbance and computed tomographic scan evidence of marked, progressive cerebral atrophy. Although the remaining patients had normal results of neurologic examinations, 11 had roentgenographic evidence of generalized cerebral atrophy; 2 of these had abnormal electroencephalograms, 1 had borderline-deficient intellectual function, and 2 had computed tomographic scan evidence of multifocal, intracerebral mineralization. The patients with nervous system abnormalities were not distinguished by patterns of medication use, demographic or laboratory features, or the relative severity of cystinosis. Although the neurologic involvement in these patients suggests that cystinosis may eventually involve the central nervous system, the differential diagnosis must include other complications from renal failure, dialysis, and immunosuppression.

Central nervous system manifestations in human immunodeficiency virus infection without AIDS.

To characterize neurological and neuropsychological findings associated with human immunodeficiency virus type-I (HIV) infection, 77 seropositive homosexual or bisexual males with no or minor symptoms of HIV were compared prospectively to 44 HIV seronegative men by observers blinded to serological status of the subjects. Neurological symptoms and examination findings were not significantly different between seropositives and seronegatives except for cranial nerve findings, predominantly mild hearing impairment. Mean performance scores for a 15-test neuropsychological battery were within an unimpaired range for both groups, although for five tests, mean scores were significantly poorer in seropositives. After adjustment for vocabulary score, and demographic and psychosocial variables, the mean score of seropositives was significantly worse only for the Benton Visual Retention Test. Magnetic resonance (MR) images of brain were abnormal in 14 (27%) of 52 seropositives and one of 10 seronegatives (value was not significant). HIV was isolated from cerebrospinal fluid (CSF) in 31 (61%) of 51 seropositives. The only clinical or laboratory difference between CSF culture positives and negatives was a higher CSF immunoglobulin synthesis rate in the former subjects (medians of 10.3 versus 0.1 mg/day; p = 0.03). An additional 13 seropositive subjects had immunologic evidence of central nervous system HIV infection, defined by a serum-to-CSF HIV antibody ratio of less than 5.5. Intracranial abnormalities on MR imaging were associated with CSF immunologic responses to HIV. Nervous system involvement occurred in the vast majority of men with early HIV infection, but clinically significant impairment was uncommon.