Movement of the epiglottis in mammals

In contrast to adult humans, the epiglottis of other mammals and infant humans is situated close to the soft palate. It has been argued that this posture is maintained during swallowing, with food passing laterally around an intact airway. To test this supposition, the movement of the epiglottis in two contrasting mammalian species, pigs and ferrets, was studied by placing radiopaque markers on the epiglottis and soft palate. Swallowing was observed with videofluoroscopy while the animals were feeding on hard and soft foods, liquids, and food mixed with barium sulfate. Analysis of the images showed that bolus formation and downward movement of the epiglottis away from the soft palate were unvarying phenomena in both animals for all tested foods. The duration of the epiglottic movement was approximately 0.3 s for liquids and slightly longer for solids. Because swallowing never occurred past an upright epiglottis, the results of this study do not support the hypothesis that adult animals maintain a patent airway during swallowing. Instead, the epiglottis in nonhuman mammals downfolds similarly to that of adult humans during swallowing. © 1996 Wiley-Liss, Inc.     

Osteoprotegerin, a crucial regulator of bone metabolism, also regulates B cell development and function

Osteoprotegerin (OPG) is a CD40-regulated gene in B cells and dendritic cells (DCs). We investigated the role of OPG in the immune system by generating opg(-/-) mice. Like its role as a regulator of bone metabolism, OPG also influences processes in the immune system, notably in B cell development. Ex vivo, opg(-/-) pro-B cells have enhanced proliferation to IL-7, and in opg(-/-) spleen, there is an accumulation of type 1 transitional B cells. Furthermore, opg(-/-) bone marrow-derived DCs are more effective in stimulating allogeneic T cells than control DCs. When challenged with a T-dependent Ag, opg(-/-) mice had a compromised ability to sustain an IgG3 Ag-specific response. Thus, in the immune system, OPG regulates B cell maturation and development of efficient Ab responses.     

Xenopus embryonic poly(A) binding protein 2 (ePABP2) defines a new family of cytoplasmic poly(A) binding proteins expressed during the early stages of vertebrate development

We describe a new RNA binding protein from Xenopus we have named ePABP2 (embryonic poly(A) binding protein type II). Based on amino acid similarity, ePABP2 is closely related to the ubiquitously expressed nuclear PABP2 protein that directs the elongation of mRNA poly(A) tails during pre-mRNA processing. However, in contrast to known PABP2 proteins, Xenopus ePABP2 is a cytoplasmic protein that is predominantly expressed during the early stages of Xenopus development and in adult ovarian tissue. Biochemical experiments indicate ePABP2 binds poly(A) with specificity and that this binding requires the RRM domain. Mouse and human ePABP2 proteins were also identified and mouse ePABP2 expression is also confined to the earliest stages of mouse development and adult ovarian tissue. We propose that Xenopus ePABP2 is the founding member of a new class of poly(A) binding proteins expressed in vertebrate embryos. Possible roles for this protein in regulating mRNA function in early vertebrate development are discussed.     

Bayesian inferences in the Cox model for order-restricted hypotheses

In studying the relationship between an ordered categorical predictor and an event time, it is standard practice to include dichotomous indicators of the different levels of the predictor in a Cox model. One can then use a multiple degree-of-freedom score or partial likelihood ratio test for hypothesis testing. Often, interest focuses on comparing the null hypothesis of no difference to an order-restricted alternative, such as a monotone increase across levels of a predictor. This article proposes a Bayesian approach for addressing hypotheses of this type. We reparameterize the Cox model in terms of a cumulative product of parameters having conjugate prior densities, consisting of mixtures of point masses at one, and truncated gamma densities. Due to the structure of the model, posterior computation can proceed via a simple and efficient Gibbs sampling algorithm. Posterior probabilities for the global null hypothesis and subhypotheses, comparing the hazards for specific groups, can be calculated directly from the output of a single Gibbs chain. The approach allows for level sets across which a predictor has no effect. Generalizations to multiple predictors are described, and the method is applied to a study of emergency medical treatment for stroke.     

Differential roles of CC chemokine ligand 2/monocyte chemotactic protein-1 and CCR2 in the development of T1 immunity

CCR2 and its major ligand, chemokine ligand 2 (CCL2)/monocyte chemotactic protein-1, have been found to influence T1/T2 immune response polarization. Our objective was to directly compare the roles of CCR2 and CCL2 in T1/T2 immune response polarization using a model of pulmonary Cryptococcus neoformans infection. Either deletion of CCR2 or treatment of wild-type mice with CCL2 neutralizing Ab produced significant and comparable reductions in macrophage and T cell recruitment into the lungs following infection. Both CCL2 neutralization and CCR2 deficiency resulted in significantly diminished IFN-gamma production, and increased IL-4 and IL-5 production by lung leukocytes (T1 to T2 switch), but only CCR2 deficiency promoted pulmonary eotaxin production and eosinophilia. In the lung-associated lymph nodes (LALN), CCL2-neutralized mice developed Ag-specific IFN-gamma-producing cells, while CCR2 knockout mice did not. LALN from CCR2 knockout mice also had fewer MHCII(+)CD11c(+) and MHCII(+)CD11b(+) cells, and produced significantly less IL-12p70 and TNF-alpha when stimulated with heat-killed yeast than LALN from wild-type or CCL2-neutralized mice, consistent with a defect in APC trafficking in CCR2 knockout mice. Neutralization of CCL2 in CCR2 knockout mice did not alter immune response development, demonstrating that the high levels of CCL2 in these mice did not play a role in T2 polarization. Therefore, CCR2 (but not CCL2) is required for afferent T1 development in the lymph nodes. In the absence of CCL2, T1 cells polarize in the LALN, but do not traffic from the lymph nodes to the lungs, resulting in a pulmonary T2 response.     

A novel disorder caused by defective biosynthesis of N-linked oligosaccharides due to glucosidase I deficiency

Glucosidase I is an important enzyme in N-linked glycoprotein processing, removing specifically distal alpha-1,2-linked glucose from the Glc3Man9GlcNAc2 precursor after its en bloc transfer from dolichyl diphosphate to a nascent polypeptide chain in the endoplasmic reticulum. We have identified a glucosidase I defect in a neonate with severe generalized hypotonia and dysmorphic features. The clinical course was progressive and was characterized by the occurrence of hepatomegaly, hypoventilation, feeding problems, seizures, and fatal outcome at age 74 d. The accumulation of the tetrasaccharide Glc(alpha1-2)Glc(alpha1-3)Glc(alpha1-3)Man in the patient's urine indicated a glycosylation disorder. Enzymological studies on liver tissue and cultured skin fibroblasts revealed a severe glucosidase I deficiency. The residual activity was <3% of that of controls. Glucosidase I activities in cultured skin fibroblasts from both parents were found to be 50% of those of controls. Tissues from the patient subjected to SDS-PAGE followed by immunoblotting revealed strongly decreased amounts of glucosidase I protein in the homogenate of the liver, and a less-severe decrease in cultured skin fibroblasts. Molecular studies showed that the patient was a compound heterozygote for two missense mutations in the glucosidase I gene: (1) one allele harbored a G-->C transition at nucleotide (nt) 1587, resulting in the substitution of Arg at position 486 by Thr (R486T), and (2) on the other allele a T-->C transition at nt 2085 resulted in the substitution of Phe at position 652 by Leu (F652L). The mother was heterozygous for the G-->C transition, whereas the father was heterozygous for the T-->C transition. These base changes were not seen in 100 control DNA samples. A causal relationship between the alpha-glucosidase I deficiency and the disease is postulated.     

Effects of intraoral splint wear on proteoglycans in the temporomandibular joint disc

Intraoral splints are a common dental treatment for dysfunctions of the temporomandibular joint (TMJ), but their effects on the structures of the joint, specifically the disc, have not been well investigated. This study examined proteoglycans (PGs) of the TMJ disc of the miniature pig and tested for alterations resulting from intraoral splint wear. Sixteen female pigs were divided into three groups: control (C), control splint (CS), and protrusive splint (PS). Splinted groups received chrome-cobalt ramp splints which were worn continuously for 2 months. PG content within various disc locations was determined by colorimeteric assay. PG synthesis and type were examined by labeling with (35)S-sulfate and SDS-PAGE analysis. Average water content of the disc was 77.1%, which places it at the high end of the normal range for collagenous biomaterials (60-80%). PGs migrating to the positions typical of aggrecan, biglycan, and decorin on SDS-PAGE were present in all locations of all groups. The highest content and synthesis of PGs were always found in the intermediate band of the disc regardless of group (P < 0.05), supporting the notion that this band encounters heavy compressive loading during function. The joints of animals from both splinted groups showed a high frequency of gross pathology. Biglycan synthesis was increased in both splinted groups (P < 0.05). Newly synthesized biglycan had a shorter migration distance in the intermediate bands of the CS group, suggesting increased hydrodynamic size. These findings suggest that intraoral splint wear may cause disc damage or remodeling.     

Strain in the braincase and its sutures during function

The skull is distinguished from other parts of the skeleton by its composite construction. The sutures between bony elements provide for interstitial growth of the cranium, but at the same time they alter the transmission of stress and strain through the skull. Strain gages were bonded to the frontal and parietal bones of miniature pigs and across the interfrontal, interparietal and coronal sutures. Strains were recorded 1) during natural mastication in conjunction with electromyographic activity from the jaw muscles and 2) during stimulation of various cranial muscles in anesthetized animals. Vault sutures exhibited vastly higher strains than did the adjoining bones. Further, bone strain primarily reflected torsion of the braincase set up by asymmetrical muscle contraction; the tensile axis alternated between +45 degrees and -45 degrees depending on which diagonal masseter/temporalis pair was most active. However, suture strains were not related to overall torsion but instead were responses to local muscle actions. Only the coronal suture showed significant strain (tension) during jaw opening; this was caused by the contraction of neck muscles. All sutures showed strain during jaw closing, but polarity depended on the pattern of muscle usage. For example, masseter contraction tensed the coronal suture and the anterior part of the interfrontal suture, whereas the temporalis caused compression in these locations. Peak tensile strains were larger than peak compressive strains. Histology suggested that the skull is bent at the sutures, with the ectocranial surface tensed and the endocranial surface predominantly compressed. Collectively, these results indicate that skulls with patent sutures should be analyzed as complexes of independent parts rather than solid structures.     

Characterization of terminal NeuNAcalpha2-3Galbeta1-4GlcNAc sequence in lipooligosaccharides of Neisseria meningitidis

Group B and C Neisseria meningitidis are the major cause of meningococcal disease in the United States and in Europe. N . meningitidis lipooligosaccharide (LOS), a major surface antigen, can be divided into 12 immunotypes of which L1 through L8 were found among Group B and C organisms. Groups B and C but not Group A may sialylate their LOSs with N-acetylneuraminic acid (NeuNAc) at the nonreducing end because they synthesize CMP-NeuNAc. Using sialic acid-galactose binding lectins as probes in an ELISA format, six of the eight LOS immunotypes (L2, L3, L4, L5, L7, and L8) in Groups B and C bound specifically to Maackia amurensis leukoagglutinin (MAL), which recognizes NeuNAcalpha2- 3Galbeta1-4GlcNAc/Glc sequence, but not to Sambucus nigra agglutinin, which binds NeuNAcalpha2-6Gal sequence. The combination of SDS-PAGE and MAL-blot analyses revealed that these six LOSs contained only the NeuNAcalpha2-3Galbeta1-4GlcNAc trisaccharide sequence in their 4.1 kDa LOS components, which have a common terminal lacto-N-neotetraose (LNnT, Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc) structure when nonsialylated as shown by previous studies. The LOS-lectin binding was abolished when the LOSs were treated with Newcastle disease viral neuraminidase which cleaves alpha2-->3 linked sialic acid. Methylation analysis of a representative LOS (L2) confirmed that NeuNAc is 2-->3 linked to Gal. Thus, these LOSs structurally mimic certain glycolipids, i.e., paragloboside (LNnT-ceramide) and sialylparagloboside and some glycoproteins in having LNnT and N-acetyllactosamine sequences, respectively, with or without alpha2-->3 linked NeuNAc. The molecular mimicry of the LOSs may play a role in the pathogenesis of N.meningitidis by assisting the organism to evade host immune defenses in man.