Hyperopic refractive errors as a prognostic factor in intermittent exotropia surgery.

UNLABELLED: Purpose: To evaluate and compare surgical outcomes with respect to refractive errors in strabismus surgery for the treatment of intermittent exotropia (IXT). METHODS: The medical records of patients with IXT who were treated by one surgeon from January 2005 and June 2011 were reviewed. Three hundred and thirty-three IXT patients were included and divided into three groups according to preoperative refractive error: IXT with hyperopia (group I), IXT with emmetropia (group II), and IXT with myopia (group III). The surgical outcomes with respect to sensory and motor criteria were compared among the three groups. RESULTS: The surgical success rates according to motor criteria and sensory and motor criteria combined were higher in groups I (29 patients) and III (124 patients) than in group II (180 patients) at postoperative 3 and 6 months and at the last follow-up. Stereopsis was significantly better in groups II and III than in group I preoperatively (P=0.002 by one-way analysis of variance test); however, the difference was not significant postoperatively. Twenty patients in group I (69.0%) were prescribed undercorrected hyperopic spectacles postoperatively, while only 22 patients in group III (17.7%) were prescribed spectacles with more myopic power than their refractive errors. CONCLUSION: In the surgical treatment of IXT, hyperopia was not an indicator of poor prognosis. Taking into consideration the age effect, follow-up period after IXT surgery, and stereopsis improvement, hyperopic refractive error is rather a good prognostic factor.

Results of conservative management for consecutive esotropia after intermittent exotropia surgery.

PURPOSE: To examine the clinical course of consecutive esotropia (ET) using conservative management, after intermittent exotropia (IXT) surgery. METHODS: This study included 149 out of 151 consecutive patients with ET after IXT surgery, who were managed conservatively. The clinical course of consecutive ET was examined and the patients were classified into two groups based on the duration of esodeviation: (1) >3 weeks (persistent ET group, n=56) and (2) <3 weeks (transient ET group, n=93). Patient characteristics and treatment outcomes, including the recurrence of exotropia and stereopsis, were compared between the two groups. RESULTS: All patients with ET were managed with full-time alternate occlusion and/or with a Fresnel prism. In 149 patients out of 151 consecutive patients, 82% of ET disappeared at 12-month follow-up and all at the last follow-up visit (31.4±23.5 months). At the final visit, a recurrence of exotropia of >10 prism dioptres was significantly less frequent in the persistent ET group than in the transient ET group (25% vs 62%, respectively; P=0.01). However, stereopsis outcome was not significantly different between the two groups, and stereopsis change was not affected by age. CONCLUSIONS: By using conservative management only, persistent consecutive ET after IXT surgery disappeared in most cases by the 1-year follow-up visit after surgery. Recurrence of exotropia was significantly less frequent in patients with persistent ET, yet the sensory outcome was not affected by the duration of consecutive ET or age.

Recognition and phagocytosis of multiple periodontopathogenic bacteria by anti-Porphyromonas gingivalis heat-shock protein 60 antisera.

The present study has been performed to evaluate Porphyromonas gingivalis heat shock protein (HSP) 60 as a candidate vaccine to protect against multiple putative periodontopathic bacteria. Mouse anti-P. gingivalis HSP antisera demonstrated the elevated IgG antibody titers against the multiple bacteria tested and cross-reacted with heat-induced bacterial proteins of the target bacteria. The antisera also demonstrated a significantly higher opsonophagocytosis function against all the target bacteria than the control sera (P<0.01). We concluded that P. gingivalis HSP 60 could potentially be developed as a vaccine against multiple periodontopathic bacteria.

Epitope mapping of Porphyromonas gingivalis heat-shock protein and human heat-shock protein in human atherosclerosis.

To identify T- and/or cross-reactive B-cell epitopes of P. gingivalis and human heat-shock protein (HSP)60 in atherosclerosis patients, we synthesized 104 overlapping synthetic peptides spanning whole molecules of P. gingivalis HSP60 and human HSP60, respectively. T-cell epitopes of P. gingivalis HSP were identified with the use of previously established P. gingivalis HSP-reactive T-cell lines. B-cell epitopes of P. gingivalis HSP60 and human HSP60 were identified by the use of patients' sera. Anti-P. gingivalis, anti-P. gingivalis HSP60, or anti-human HSP60 IgG antibody titers were higher in the atherosclerosis patients compared with the healthy subjects. Five immunodominant peptides of P. gingivalis HSP60, identified as T-cell epitopes, were also found to be B-cell epitopes. Moreover, 6 cross-reactive B-cell epitopes of human HSP60 were identified. It was concluded that P. gingivalis HSP60 might be involved in the immunoregulatory process of atherosclerosis, with common T- and/or B-cell epitope specificities and with cross-reactivity with human HSP60.

Identification of T-cell epitopes of Porphyromonas gingivalis heat-shock-protein 60 in periodontitis.

The heat shock proteins (hsp) of bacterial species are considered to be involved in regulating the autoimmune mechanism in human diseases due to the considerable homology of their sequences with human hsp. To elucidate how stress proteins contribute to the immunopathogenesis of periodontitis, mononuclear cells from gingival connective tissue of 10 periodontitis patients were simulated with Porphyromonas gingivalis hsp60. T-cell lines reactive to P. gingivalis hsp60 were established from each patient to define T-cell epitope specificities. Anti-P. gingivalis IgG antibody titres were elevated in all patients. We could establish P. gingivalis hsp-reactive T-cell lines from gingival mononuclear cells that were mixtures of CD4+ and CD8+ cells. Of 108 overlapping synthetic peptides spanning the whole P. gingivalis hsp60 molecule, 10 peptides with epitope specificities for T-cells were identified, and were identical to those reported be B-cell epitopes in periodontitis.

Fusobacterium nucleatum impairs serum binding to Porphyromonas gingivalis biofilm.

Mouse immune sera obtained by immunization with Fusobacterium nucleatum and then Porphyromonas gingivalis demonstrated an impaired binding capacity to P. gingivalis-biofilm and lower avidity to P. gingivalis when compared with sera obtained from mice immunized with P. gingivalis alone.

P2Y(11), a purinergic receptor acting via cAMP, mediates secretion by pancreatic duct epithelial cells.

Pancreatic duct epithelial cells (PDEC) mediate the exocrine secretion of fluid and electrolytes. We previously reported that ATP and UTP interact with P2Y(2) receptors on nontransformed canine PDEC to increase intracellular free Ca2+ concentration ([Ca2+](i)) and stimulate Ca2+-activated Cl- and K+ channels. We now report that ATP interacts with additional purinergic receptors to increase cAMP and activate Cl- channels. ATP, 2-methylthio-ATP, and ATP-gamma-S stimulated a 4- to 10-fold cAMP increase with EC(50) of 10-100 microM. Neither UTP nor adenosine stimulated a cAMP increase, excluding a role for P2Y(2) or P1 receptors. Although UTP stimulated an (125)I(-) efflux that was fully inhibited by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl) ester (BAPTA-AM), ATP stimulated a partially resistant efflux, suggesting activation of additional Cl- conductances through P2Y(2)-independent and Ca2+-independent pathways. In Ussing chambers, increased cAMP stimulated a much larger short-circuit current (I(sc)) increase from basolaterally permeabilized PDEC monolayers than increased [Ca2+](i). Luminal ATP and UTP and serosal UTP stimulated a small Ca2+-type I(sc) increase, whereas serosal ATP stimulated a large cAMP-type I(sc) response. Serosal ATP effect was inhibited by P2 receptor blockers and unaffected by BAPTA-AM, supporting ATP activation of Cl- conductances through P2 receptors and a Ca2+-independent pathway. RT-PCR confirmed the presence of P2Y(11) receptor mRNA, the only P2Y receptor acting via cAMP.

Activation of a heterologously expressed octopamine receptor coupled only to adenylyl cyclase produces all the features of presynaptic facilitation in aplysia sensory neurons.

Short-term behavioral sensitization of the gill-withdrawal reflex after tail stimuli in Aplysia leads to an enhancement of the connections between sensory and motor neurons of this reflex. Both behavioral sensitization and enhancement of the connection between sensory and motor neurons are importantly mediated by serotonin. Serotonin activates two types of receptors in the sensory neurons, one of which is coupled to the cAMP/protein kinase A (PKA) pathway and the other to the inositol triphosphate/protein kinase C (PKC) pathway. Here we describe a genetic approach to assessing the isolated contribution of the PKA pathway to short-term facilitation. We have cloned from Aplysia an octopamine receptor gene, Ap oa(1), that couples selectively to the cAMP/PKA pathway. We have ectopically expressed this receptor in Aplysia sensory neurons of the pleural ganglia, where it is not normally expressed. Activation of this receptor by octopamine stimulates all four presynaptic events involved in short-term synaptic facilitation that are normally produced by serotonin: (i) membrane depolarization; (ii) increased membrane excitability; (iii) increased spike duration; and (iv) presynaptic facilitation. These results indicate that the cAMP/PKA pathway alone is sufficient to produce all the features of presynaptic facilitation.

Spectrum of opportunistic infections and malignancies in patients with human immunodeficiency virus infection in South Korea.

To determine the frequency and types of major opportunistic diseases in patients with HIV infection in South Korea, we reviewed the medical records of 173 HIV-infected patients. The patients were seen from 1985 to 1998 at a referral hospital for AIDS in South Korea. Most patients (85%) were male, and 107 (62%) were infected by heterosexual contacts. CD4+ lymphocyte counts at presentation were <200/microL in 27% of the patients. Tuberculosis was the most frequent opportunistic infection (25% of patients), followed by candidiasis (21%), herpes zoster (20%), Pneumocystis carinii pneumonia (10%), cytomegalovirus disease (9.8%). There were no cases of toxoplasmosis. Kaposi's sarcoma developed in 3 patients (1.7%), and non-Hodgkin's lymphoma, in 2 (1.2%). Eleven patients (6.4%) developed peripheral neuropathy, and 8 (4.6%) had HIV encephalopathy. Tuberculosis was the single most important HIV-related infection in South Korean patients.

Cloning and functional characterization of a Caenorhabditis elegans muscarinic acetylcholine receptor.

A cDNA clone encoding a muscarinic acetylcholine receptor (mAChR) has been isolated from the nematode Caenorhabditis elegans. The nematode mAChR, consisted of 585 amino acids, displays a high degree of amino acid sequence homology to other invertebrate and vertebrate mAChRs. Excluding a highly variable middle portion of the third intracellular loop, the C. elegans mAChR shares about 51% amino acid sequence identity with a Drosophila mAChR and 42-44% identity with human m1-m5 mAChR subtypes. Comparison of the cDNA sequence with the corresponding genomic sequence reveals that the C. elegans mAChR gene contains ten introns, eight of them in the coding region. Pharmacological profiles of the C. elegans mAChR expressed in Chinese hamster ovary (CHO) cells were shown to be similar to those of mammalian counterparts, indicating that ligand binding domains of the receptor have been conserved during evolution. When this cloned receptor was expressed in Xenopus oocytes, acetylcholine evoked a transient Cl- current. Furthermore, activation of the receptor with oxotremorine, acetylcholine or carbachol resulted in the stimulation of phosphatidylinositol metabolism in CHO cells, suggesting that the receptor is coupled to phospholipase C activation.

Interactions of Escherichia coli primary replicative helicase DnaB protein with nucleotide cofactors.

Interactions between the Escherichia coli primary replicative helicase DnaB protein and nucleotide cofactors have been studied using several fluorescent nucleotide analogs and unmodified nucleotides. The thermodynamically rigorous fluorescent titration technique has been used to obtain true binding isotherms, independently of the assumptions of any relationships between the observed quenching of protein fluorescence and the degree of nucleotide binding. Fluorescence titrations using several MANT derivatives of nucleoside diphosphates (MANT-ADP, 3',2'-O-(N-methylantraniloyl)adenosine-5'-diphosphate; MANT-GDP, 3',2'-O(N-methylantraniloyl)guanosine-5'-diphosphate; MANT-CDP, 3',2'-O-(N-methylantraniloyl)cytidine-5'-diphosphate; MANT-UDP, 3',2'-O-(N-methylantraniloyl)uridine-5'-diphosphate) have shown that the DnaB helicase has a preference for purine nucleotides. Binding of all modified nucleotides is characterized by similar negative cooperativity, indicating that negative cooperative interactions are base-independent. Thermodynamic parameters for the interactions of the unmodified nucleotides (ADP, GDP, CDP, and UDP) and inorganic phosphate (P(i)) have been obtained by using the competition titration approach. To analyze multiple ligand binding to a finite circular lattice, for a general case in which each lattice binding site can exist in different multiple states, we developed a matrix method approach to derive analytical expressions for the partition function and the average degree of binding for such cases. Application of the theory to competition titrations has allowed us to extract the intrinsic binding constants and cooperativity parameters for all unmodified ligands. This is the first quantitative estimate of affinities and the mechanisms of binding of different unmodified nucleotides and inorganic phosphate for a hexameric helicase. The intrinsic affinities of all of the studied ATP analogs are lower than the intrinsic affinities of the corresponding ADP analogs. The implications of these results for the mechanism of helicase action are discussed.

Binding of Escherichia coli primary replicative helicase DnaB protein to single-stranded DNA. Long-range allosteric conformational changes within the protein hexamer.

Quantitative analyses of the interactions of the Escherichia coli primary replicative helicase DnaB protein with single-stranded DNA have been performed using the thermodynamically rigorous fluorescence titration technique. This approach allowed us to obtain absolute stoichiometries of the formed complexes and interaction parameters, without any assumptions about the relationship between the observed signal change and the degree of binding. The analysis of the DnaB helicase interactions with nonfluorescent, unmodified nucleic acids has been performed, using a novel spectroscopic Macromolecular Competition Titration (MCT) method developed in the accompanying paper [Jezewska, M. J., & Bujalowski, W. (1996) Biochemistry 35, 2117-2128]. In the presence of the ATP nonhydrolyzable analog AMP-PNP, the DnaB helicase binds polymer DNA with a site-size of 20 +/- 3 nucleotides per protein hexamer. This site-size is independent of the type of nucleic acid base as well as the salt concentration and type of salt. Direct thermodynamic studies of the polynucleotide and oligomer binding to the DnaB hexamer, as well as the competition studies, show that independently of the type of nucleic acid base, as well as salt concentration and type of salt in solution, the helicase has only a single, strong binding site for DNA. Only this site is used when the protein interacts with polymer DNA. Moreover, UV photo-cross-linking experiments with oligonucleotides of different lengths, dT(pT)19, dT(pT)55, and dT(pT)69, suggest that primarily a single subunit of the DnaB helicase hexamer is in contact with the DNA. In interactions with polymer nucleic acids, the DnaB protein shows preferential intrinsic affinity for poly(dA), characterized in our standard conditions (pH 8.1, 10 degrees C, 100 mM NaCl, 5 mM MgCl2) by the intrinsic binding constant K = 6 +/- 2 x 10(6) M-1. These affinities are comparable to the affinities of the single-strand binding proteins in the corresponding solution conditions and strongly suggest that the helicase is capable of binding DNA without additional facilitating factors. Both the intrinsic affinity and the cooperativity are salt dependent. The formation of the DnaB-DNA complex is accompanied by the net release of approximately 2 ions, while another net release of approximately 2 ions accompanies the cooperative interactions. The data indicate an anion effect on the studied interactions and suggests that the released ions most probably originate from both the protein and the nucleic acid. The presence of a single, strong binding site on the hexamer, built of six chemically identical subunits, the very low site-size of the large helicase-DNA complex, and the involvement of a single subunit in contact with the nucleic acid indicate the presence of long-range allosteric interactions in the DnaB helicase which encompass the entire DnaB hexamer. Our sedimentation velocity measurements of the DnaB protein-(AMP-PNP)-5'-fluorescein-(dT)20 ternary complex show that the sedimentation coefficient of the complex is S20,W = 12.3 +/- 0.3, compared with S20,W = 10.5 +/- 0.3 of the free enzyme, indicating large changes in the hydrodynamic properties of the enzyme in the complex. These results provide direct evidence that the DnaB hexamer undergoes dramatic conformational changes which include all six subunits of the enzyme in the ternary complex. Moreover, sedimentation velocity studies of the ternary complex provide direct evidence that the hexamer is the species which binds ss nucleic acid. The significance of these results for a mechanistic model of the functioning of the DnaB helicase in DNA replication is discussed.

Effect of ethidium binding and superhelix density on the apparent supercoiling free energy and torsion constant of pBR322 DNA.

The value of the twist energy parameter (ET) of pBR322 is determined near zero superhelix density from topoisomer distributions created under various conditions. The resulting value, ET = 1155 +/- 65, at 37 degrees C is essentially unaffected by adding 10 mM Mg2+, or by changing the kind of Topo I from chicken-red-cell to calf-thymus. This value significantly exceeds that (ET = 950 +/- 80) measured for p30 delta DNA under identical conditions by the same method in the preceding paper. Decreasing the temperature from 37 to 21 degrees C yields a slightly larger value, ET = 1340 +/- 130, but the statistical significance of the increase is marginal. Attempts to determine reliable ET values for pBR322 at higher superhelix densities by ethidium binding were frustrated by the fact that good fits of the equilibrium dialysis results could not be achieved using a single value of ET. Moreover, the curves of apparent ET versus binding ratio r vary considerably from one preparation to another, and for a given preparation vary with time after cell lysis up to about seven weeks, after which they settle in to nearly reproducible behavior. The apparent ET values obtained from competitive dialysis experiments are typically rather low (ET approximately 700) for small r and nearly native superhelix density, and rise up to 1300 to 1500 with increasing binding ratio (up to r = 0.055) and decreasing negative superhelix density.(ABSTRACT TRUNCATED AT 250 WORDS)

Dynamics and structures of DNA: long-range effects of a 16 base-pair (CG)8 sequence on secondary structure.

The effects of inserting 16 base pair (bp) of alternating CG [(CG)8] near the middle of a much longer restriction fragment (1097 bp) are investigated by measuring various properties that are sensitive to secondary and tertiary structure. Results for this fragment are compared with those for a control fragment (1089 bp) with the identical sequence except at the insert. Another fragment (1382 bp), which contains a 296-bp extension at the 5'-end of the 1089-bp control fragment, is also used as a secondary control in some experiments. When the 1097-bp (CG)8 insert fragment is compared with the control fragments in 0.1 M NaCl buffer, the (CG)8 insert is found to induce disproportionately large relative changes in the molar ellipticity at 273 nm ([theta]273), the torsion constant (alpha) measured by fluorescence polarization anisotropy, the optical melting profile, and the susceptibility to S1 nuclease. Estimates of the minimum distance over which the (CG)8 insert alters the secondary structure range from 330 to 550 bp. With increasing NaCl concentration, the 1097-bp insert fragment undergoes a structural transition between 2.0 and 2.5 M that is manifested in the apparent diffusion coefficient (Dplat) from dynamic light scattering at large scattering vector. This transition, which is not exhibited by the control DNAs, is presumed to involve formation of Z-helix at the insert. However, the observed decrease in (Dplat) is attributed to an increase in bending rigidity, which perforce must be globally distributed far beyond the (CG)8 insert per se. In 4.25 M NaCl (but not in 0.1 M NaCl), the addition of 1 ethidium dye per 300 bp induces an extensive structural transition in the 1097 bp (CG)8 insert fragment. This transition, which also is not exhibited by the control DNAs, significantly decreases the bending rigidity, doubles [theta]273, and takes place on a time scale of a few days. Removal of ethidium and salt by dialysis vs 0.1 M NaCl buffer restores the original properties of the 1097-bp (CG)8 insert fragment. The present results are consistent with a (fluctuating, long-range) description of the secondary structure in which a given short sequence transiently fluctuates among two or more distinct secondary structures that extend over much larger domains of variable position and size, and whose relative stabilities depend on distant as well as close-lying base pairs.

Renal angiomyolipoma associated with neurofibromatosis and primary carcinoid of mesentery.

A patient with von Recklinghausen's disease manifested by dermal neurofibromatosis and cafe-au-lait spots presented with complaints of malaise, weight loss, lower extremity weakness, and a palpable left lower abdominal quadrant mass. Evaluation revealed a lumbar neurofibroma, a localized primary carcinoid tumor of the mesentery, and a left renal angiomyolipoma. Although an association between neurofibromatosis and carcinoid has been previously reported, we believe this is the first report documenting the association of all three entities.