Disulfide bond formation between dimeric immunoglobulin A and the polymeric immunoglobulin receptor during hepatic transcytosis

The polymeric immunoglobulin receptor on rat hepatocytes binds dimeric IgA on the sinusoidal surface and mediates its transport to the canaliculus, where the complex of dimeric IgA and secretory component, the cleaved extracellular domain of polymeric immunoglobulin receptor, is secreted into bile. This process is unique in that disulfide bonds are formed between dimeric IgA and polymeric immunoglobulin receptor during transcytosis, permanently preventing their dissociation. Here we present three lines of evidence that disulfide bonding between dimeric IgA and polymeric immunoglobulin receptor occurs predominantly in a late transcytotic compartment and that hepatic transcytosis can proceed in the absence of disulfide bond formation. First, throughout the course of transcytosis the percentage of intracellular dimeric IgA disulfide bonded to polymeric immunoglobulin receptor is less than half that in bile, suggesting that disulfide bond formation is a late event in transcytosis. Second, dimeric IgA that recycles from early endocytotic compartments into the circulation is mostly noncovalently bound to secretory component. Finally, the rate of transcytosis of dimeric IgA and its appearance in bile are not affected when disulfide bond formation with polymeric immunoglobulin receptor is inhibited by blocking of free thiol groups on dimeric IgA with iodoacetamide. These results are consistent with other findings in the literature and indicate that the main physiological role of disulfide bond formation between dimeric IgA and polymeric immunoglobulin receptor is not to facilitate transcytosis but, rather, to stabilize the dimeric IgA-secretory component complex after its release into external secretions such as bile and intestinal secretions.     

Experimental Taenia solium cysticercosis in pigs: characteristics of the infection and antibody response

The biology of aging is reviewed from the perspective of a medical geneticist. This was the perspective of the late Sam Goldstein, and this article is, therefore, dedicated to his memory. Aging can be defined as the set of phenotypes that escape the force of natural selection. These phenotypes can be modulated by mutation or polymorphism at numerous genetic loci. Given the remarkable genetic and environmental heterogeneity that characterizes our species, it is understandable that there should be considerable variation in patterns of aging. A genetic approach involving the mapping and positional cloning of major loci could provide basic understanding of the mechanisms underlying such variability. Prototypic examples being investigated by the author and his colleagues are the Werner syndrome and dementias of the Alzheimer type. The biochemical genetic analysis of these and other disorders could lead to a new style of medicine based upon preventive approaches tailored to the needs of individuals. Such interventions should ideally involve pediatricians.     

Encapsulated hemoglobin: current issues and future goals

The promise of encapsulation systems for the sequestration of hemoglobin has been the long-held belief that encapsulation more closely mimics nature's strategy for circulating hemoglobin, and could alleviate hemoglobin based toxicities and increase circulation persistence. Various polymers have been proposed to deliver hemoglobin. One approach toward the encapsulation of hemoglobin has been to employ biodegradable, biocompatible vehicles such as phospholipid vesicles, or liposomes. The majority of encapsulation work with hemoglobin over recent years has focused on liposome encapsulated hemoglobin with demonstrations of efficacy and safety in total and partial isovolemic and hypovolemic exchange models, hemodynamics, circulation persistence and organ biodistribution, processing methods, long term storage through freeze-drying, and serum changes and histopathological consequences following administration in small animals. The data collected thus far indicate that encapsulation of hemoglobin does significantly alter many of the traditionally observed effects following the administration of cell free hemoglobin solutions. Liposome encapsulated hemoglobin circulates for 20-24 hours in small animals and principally distributes to the liver and spleen. The significant accumulation of liposome encapsulated hemoglobin in these organs poses new questions for short and long-term effects on the reticuloendothelial system and macrophage function which are currently being addressed. In addition, transient hemodynamic and serum changes have been observed following the administration of liposome encapsulated hemoglobin. Many of these are similar to the effects observed following administration of liposomes without intravesicular hemoglobin and are dictated by liposome parameters such as surface charge and character, size, and lipid composition. Finally, fundamental large scale production issues such as encapsulation efficiency and particle size distribution must be optimized to facilitate the commercial development of encapsulated hemoglobin. These issues are discussed in the historical context of encapsulated hemoglobin and basic liposome research, current research status, and future challenges for the development of encapsulated hemoglobin as an artificial oxygen carrying fluid.     

Inter- and intraspecies polymorphisms in the cholecystokinin-B/gastrin receptor alter drug efficacy

The brain cholecystokinin-B/gastrin receptor (CCK-BR) is a major target for drug development because of its postulated role in modulating anxiety, memory, and the perception of pain. Drug discovery efforts have resulted in the identification of small synthetic molecules that can selectively activate this receptor subtype. These drugs include the peptide-derived compound PD135,158 as well as the nonpeptide benzodiazepine-based ligand, L-740,093 (S enantiomer). We now report that the maximal level of receptor-mediated second messenger signaling that can be achieved by these compounds (drug efficacy) markedly differs among species homologs of the CCK-BR. Further analysis reveals that the observed differences in drug efficacy are in large part explained by single or double aliphatic amino acid substitutions between respective species homologs. This interspecies variability in ligand efficacy introduces the possibility of species differences in receptor-mediated function, an important consideration when selecting animal models for preclinical drug testing. The finding that even single amino acid substitutions can significantly affect drug efficacy prompted us to examine ligand-induced signaling by a known naturally occurring human CCK-BR variant (glutamic acid replaced by lysine in position 288; 288E --> K). When examined using the 288E --> K receptor, the efficacies of both PD135,158 and L-740, 093 (S) were markedly increased compared with values obtained with the wild-type human protein. These observations suggest that functional variability resulting from human receptor polymorphisms may contribute to interindividual differences in drug effects.     

Two year followup of anterior and vertical atlantoaxial subluxation in ankylosing spondylitis

OBJECTIVE: To describe the clinical and radiological 2-yr followup of 22 patients with anterior atlantoaxial subluxation (AAS) from a prospective cohort of patients with ankylosing spondylitis. METHODS: The 2-yr assessment included a structured questionnaire for rheumatologic and neurologic complaints and lateral cervical radiographs in maximal flexion view. Initial and 2-yr radiographs were assessed blind to patient data. The course of anterior AAS was classified as unchanged (< 1 mm), progression (> or = 1 mm) or regression (> or = 1 mm) at 2 yrs compared with baseline. Vertical AAS was classified using the Sakaguchi-Kauppi method. Magnification factor was corrected using the ratio of C3 width. RESULTS: Anterior AAS was detected in 22 patients at baseline examination. Two patients also had vertical AAS; 86% were male. Mean age was 33 +/- 9 yrs and mean disease duration was 12 +/- 7 yrs. At followup, one patient had died of acquired immunodeficiency syndrome, 3 could not be reached, and 2 had undergone surgical fusion due to severe myelopathy and now showed complete neurological recovery. Of the remaining 16 patients, 7 (32%) showed progression and 9 (41%) showed no change in the C1-odontoid distance. Vertical AAS developed in one patient. After the 2-yr assessment, 3 additional patients had surgical fusion because of notable progression of AAS, despite absence of neurological signs. CONCLUSION: Anterior AAS progressed in a number of these patients in the 2 yrs following its detection, and with or without neurological signs, surgical management was thought appropriate in a considerable number of them.     

Study of portal hypertension in children with special reference to sclerotherapy

Spinal cord injuries are rare in children, in face of their higher mobility comparing to adults. The high cervical and the thoracic segments of the spine are more frequently affected. In the last 10 years we had 90 cases of spinal injuries in our service being 12 with neurologic deficient (8 male and 4 female) and four of them without radiographic abnormality, even in the dynamics studies. The authors emphasise the possibility of occurrence of neurologic deficit in children after trauma, even without any radiographic abnormality.     

SAP97 is associated with the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor GluR1 subunit

Rapid glutamatergic synaptic transmission is mediated by ionotropic glutamate receptors and depends on their precise localization at postsynaptic membranes opposing the presynaptic neurotransmitter release sites. Postsynaptic localization of N-methyl-D-aspartate-type glutamate receptors may be mediated by the synapse-associated proteins (SAPs) SAP90, SAP102, and chapsyn-110. SAPs contain three PDZ domains that can interact with the C termini of proteins such as N-methyl-D-aspartate receptor subunits that carry a serine or threonine at the -2 position and a valine, isoleucine, or leucine at the very C terminus (position 0). We now show that SAP97, a SAP whose function at the synapse has been unclear, is associated with alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-type glutamate receptors. AMPA receptors are probably tetramers and are formed by two or more of the four AMPA receptor subunits GluR1-4. GluR1 possesses a C-terminal consensus sequence for interactions with PDZ domains of SAPs. SAP97 was present in AMPA receptor complexes immunoprecipitated from detergent extracts of rat brain. After treatment of rat brain membrane fractions with the cross-linker dithiobis(succinimidylpropionate) and solubilization with sodium dodecylsulfate, SAP97 was associated with GluR1 but not GluR2 or GluR3. In vitro experiments with recombinant proteins indicate that SAP97 specifically associates with the C terminus of GluR1 but not other AMPA receptor subunits. Our findings suggest that SAP97 may be involved in localizing AMPA receptors at postsynaptic sites through its interaction with the GluR1 subunit.     

Identification of 1,1'-bi(4-anilino)naphthalene-5,5'-disulfonic acid binding sequences in alpha-crystallin

The hydrophobic binding sites in alpha-crystallin were evaluated using fluorescent probes 1,1'-bi(4-anilino)naphthalenesulfonic acid (bis-ANS), 8-anilino-1-naphthalene sulfonate (ANS), and 1-azidonaphthalene 5-sulfonate (1,5-AZNS). The photolysis of bis-ANS-alpha-crystallin complex resulted in incorporation of the probe to both alphaA- and alphaB-subunits. Prior binding of denatured alcohol dehydrogenase to alpha-crystallin significantly decreased the photoincorporation of bis-ANS to alpha-crystallin. Localization of bis-ANS incorporated into alphaA-crystallin resulted in the identification of residues QSLFR and HFSPEDLTVK as the fluorophore binding regions. In alphaB-crystallin, sequences DRFSVNLNVK and VLGDVIEVHGK were found to be the bis-ANS binding regions. Of the bis-ANS binding sequences, HFSPEDLTVK of alphaA-crystallin and DRFSVNLNVK and VLGDVIEVHGK of alphaB-crystallin were earlier identified as part of the sequences involved in their interaction with target proteins during the molecular chaperone-like action. The hydrophobic probe, 1,5-AZNS, also interacted with both subunits of alpha-crystallin. Localization of 1,5-AZNS binding site in alphaB-crystallin lead to the identification of HFSPEEK sequence as the interacting site in this subunit of alpha-crystallin. Glycated alpha-crystallin displayed decreased ANS fluorescence and loss of chaperone-like function, suggesting the involvement of glycation site as well as ANS binding site in chaperone-like activity display.