Bordetella bronchiseptica fimbrial protein-enhanced immunogenicity of a Mannheimia haemolytica leukotoxin fragment.

Leukotoxin produced by Mannheimia (Pasteurella) haemolytica is an important virulence factor in shipping fever pneumonia in feedlot cattle and is a critical protective antigen. In this study, the immune response to a chimeric protein generated by combining a gene fragment encoding neutralizing epitopes of M. haemolytica leukotoxin and a fimbrial protein gene (fim N) from Bordetella bronchiseptica was evaluated. The recombinant gene was cloned in a bacterial expression vector under the control of the tac promoter and expressed as a fusion protein with glutathione-S-transferase (GST) in Escherichia coli. Immunization of mice with the recombinant protein, GST-LTXFIM elicited a significantly stronger anti-leukotoxin antibody response than comparable immunizations with GST-LTX fusion proteins lacking FIM N. The GST-LTXFIM was also more stable than GST-LTX during storage at -80 degrees C, thus alleviating a stability problem inherent to leukotoxin. This chimeric protein may be a candidate for inclusion in new generation vaccines against shipping fever pneumonia.

Promoting the recruitment and retention of minority dental faculty: federal, state, and personal programs.

There are federal programs available for both minority students and faculty. There are also private, state, and foundation funds available for minority students. The most important factor for these programs to succeed is the understanding of peoples' needs. We have to create an environment in which people are judged on their ability. At the same time we have to create an environment that rewards diversity.

Principles of dentofacial orthopedics.

The differences in the response of patients to the same orthodontic treatment are, to a great extent, the result of variability in the direction and rate of craniofacial growth. Furthermore, there is currently little scientific evidence that the temporary improvement of skeletal relationships from orthopedic appliances will alter the craniofacial skeleton on a permanent basis. However, contemporary literature is beginning to show that certain appliances may be more effective than others at a specific point in the growth process. Timing of treatment in a patient is becoming of increased clinical importance. A review of the anatomy of palatal expansion indicates that expansion is much greater in the anterior portion of the palate, in both horizontal and vertical planes. Assessment of skeletal maturity for treatment timing and growth prediction is most commonly performed with the hand/wrist radiograph. A new method is presented which uses epiphyseal and diaphyseal widths and fusion of selected phalanges to determine the relative position of the individual on the pubertal growth curve. Skeletal maturity assessment is a traditional attempt to judge physiological development. The future of craniofacial growth assessment lies in the development of physiological measurements which are both replicable and valid and clinically feasible. The data from these studies provides information to allow better quantitative diagnosis and treatment as well as objective assessment of the treatment outcome.

X-ray diffraction of nitinol orthodontic arch wires.

Superelastic mechanical behavior of nitinol alloy orthodontic wires is thought to be the result of a stress induced crystallographic transformation from austenite to martensite. The purpose of the study was to compare the SE mechanical behavior of nitinol wires to stress-induced phase changes. Eight nitinol arch wires having rectangular cross-sections were strained from 0% to 10% in tension with a mechanical testing machine. Load/extension plots were subjectively ranked for SE behavior. X-ray diffraction patterns were collected with and without 6% strain. Without strain, nitinol wires were found to be predominantly austenite with some wires containing a small amount of martensite. When strained 6%, superelastic wires demonstrated a phase transformation from austenite to martensite. XRD patterns were ranked for percent transformation and 110 peak width. Product rankings of the degree of superelasticity were positively correlated with the rank of martensitic transformation (p < 0.05). Superelasticity ranks were negatively correlated with XRD peak width ranks (p < 0.01). Increased peak width indicates greater cold work. A range of superelastic mechanical behavior and martensitic transformation is exhibited by wires currently on the market. Cold work and heat treatments are important variables to be controlled during the manufacture of nitinol products.

Ferritin stimulation of a monokine inhibitor of lipopolysaccharide-augmented myelopoiesis is ferroxidase dependent.

Ferritin inhibition of myelopoiesis has been associated with intrinsic ferroxidase activity of heavy-chain ferritin and with production of a monokine inhibitor of lipopolysaccharide (LPS)-augmented monocytopoiesis. We report here that intrinsic ferroxidase activity of heavy-chain ferritin is required for stimulated production of the monokine inhibitor of LPS-augmented monocytopoiesis.

Comparative evaluation of ceramic bracket base designs.

Since the initial introduction of ceramic brackets, base designs have been modified to reduce tooth damage during debonding. The purpose of this study was to compare shear and tensile bond strengths and fracture sites of four second-generation ceramic brackets: Allure IV (A) (GAC International, Inc., Central Islip, N.Y.), Ceramaflex (C) (TP Orthodontics, Inc., LaPorte, Ind.), Intrigue (I) (Lancer Orthodontics, Carlsbad, Calif.), Transcend 2000 (T) (Unitek Corp., Monrovia, Calif.), and a foil-mesh base stainless steel bracket, DynaBond II (D) (Unitek Corp., Monrovia, Calif.). Twenty brackets of each type were bonded to 100 mandibular bovine incisor teeth with Concise bonding adhesive. The samples were thermocycled for 24 hours and the brackets were debonded with an Instron universal testing machine (Instron Corp., Canton, Mass.). A modified Transcend debonding instrument was used for tensile debonding, whereas a chisel was used for shear debonding. An analysis of variance was performed with a 0.05 level of confidence. Mean shear strengths (kg/cm2) necessary to debond were 174.0 (A), 71.0 (C), 189.0 (I), 228.0 (T), and 160.0 (D). Mean tensile strengths (kg/cm2) were 27.0 (A), 26.7 (C), 51.3 (I), 56.5 (T), and 48.6 (D). Fracture sites examined with a light microscope showed no enamel damage with any of the ceramic brackets. Intrigue was the only bracket to fracture and had 30% bracket fracture in the tensile mode and 20% bracket fracture in the shear mode. The percentage of fractures at the adhesive-bracket base interface for shear and tensile modes, respectively, were 80, 100 (A); 100, 90 (C); 10, 60 (I); 60, 90 (T); and 90, 80 (D).(ABSTRACT TRUNCATED AT 250 WORDS)

Prostaglandin E (PGE) and interleukin-1 beta (IL-1 beta) levels in gingival crevicular fluid during human orthodontic tooth movement.

The purpose of this study was to examine gingival crevicular fluid (GCF) levels of two potent bone resorbing mediators, prostaglandin E (PGE) and interleukin-1 beta (IL-1 beta), during human orthodontic tooth movement. The study included 10 patients, each having one treatment tooth undergoing orthodontic movement and a contralateral control tooth. The GCF was sampled at control sites and treatment (compression) sites before activation and a 1, 24, 48, and 168 hours. Prevention of plaque-induced inflammation allowed this study to focus on the dynamics of mechanically stimulated PGE and IL-1 beta GCF levels. The PGE and IL-1 beta levels were determined with radioimmunoassay. At 1 and 24 hours, mean GCF IL-1 beta levels were significantly elevated at treatment teeth (8.9 +/- 2.0 and 19.2 +/- 6.0 pg, respectively) compared with control teeth (2.0 +/- 1.1 pg, p = 0.0049, and 2.9 +/- 1.0 pg, p = 0.0209, respectively). The GCF levels of PGE for the treatment teeth were significantly higher at 24 and 48 hours (108.9 +/- 11.9 and 97.9 +/- 7.3 pg) than the control teeth (61.8 +/- 7.2 pg, p = 0.0071, and 70.8 +/- 7.4 pg, p = 0.0021, respectively). The GCF levels of PGE and IL-1 beta remained at baseline levels throughout the study for the control teeth, whereas significant elevations from baseline in GCF IL-1 beta (24 hours) and PGE levels (24 and 48 hours) were observed over time in the treatment teeth (p < or = 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Mandibular condyle position: comparison of articulator mountings and magnetic resonance imaging.

This study evaluated the reliability of jaw positions, the existence of distinct jaw positions, and condyle-disk-fossa relationships in a symptom-free population by using articulator mountings and magnetic resonance imaging (MRI). The subjects examined included 28 men, 22 to 35 years of age, all having Angle Class I molar relationships and no discernible TMJ dysfunction. Records taken included the following: an axiographic face-bow to locate retruded hinge axis position, an interocclusal registration of retruded position (RE), a series of interocclusal registrations for centric occlusion (CO), a leaf gauge-generated centric relation (CR), a series of interocclusal registrations for CR, and MRI. The mandibular position indicator of the SAM articulator (Great Lakes Orthodontics, Ltd., Tonawanda, N.Y.) was used to determine reliability and existence of distinct jaw positions. Magnetic resonance imaging also evaluated jaw positions and anatomic relationships. The results indicate: (1) The articulator analysis of CO and CR is statistically replicable. (2) A distinct jaw position could be demonstrated for CO that was separate from RE and CR. It was not possible to discriminate between RE and CR. (3) Condylar concentricity was observed in half of the sample and remained consistent in RE, CO, and CR. (4) Of the sample 13% demonstrated anteriorly displaced disks that were not influenced by posterior condyle placement. (5) The clinical concept of treating to CR as a preventive measure to improve disk-to-condyle relationships was not supported by this study.

Cytokine mediation of the suppressive effect of ferritin on colony-stimulating factor-1-dependent monocytopoiesis.

Peptide-specific IgG from a rabbit immunized with an alanine-lysine-proline-arginine ((ALA1)-tuftsin) containing 14-mer "ferritin" peptide neutralized rat liver ferritin inhibition of in vitro CSF-1-dependent monocytopoiesis. Antiferritin IgG similarly neutralized the inhibitory effect of ferritin but did not neutralize peptide inhibition of the in vitro myelopoietic response. No cross-reactivity between the respective antibodies and Ag was detected either by Western immunoblot or by competitive ELISA. Depletion of adherent cells before marrow cell culture significantly reduced the inhibitory effect of ferritin but did not influence peptide inhibition of CSF-1-stimulated colony formation. Adherent marrow cells and P388D1 cells treated with both CSF-1 and ferritin, but not either alone, produced inhibitory supernatant culture media that were neutralized by antipeptide but not antiferritin IgG. High resolution molecular sieve chromatography of the inhibitory adherent marrow cell and P388D1 supernatants resolved two peaks of 50 to 60 kDa and approximately 30 kDa in each. The inhibitory activity in all four peaks was neutralized by antipeptide but not antiferritin IgG. The ferritin/CSF inhibitors were not further characterized although identity with IL-6, IL-8, TNF-alpha, transforming growth factor-beta, and IFN-alpha/beta could be eliminated. The results indicate that ferritin inhibition of CSF-1-dependent monocytopoiesis is mediated by an endogenously produced inhibitor, or inhibitors, that shares antigenic similarity with the (ALA1)-tuftsin-containing 14-mer peptide and that adherent marrow cells, most likely monocytes or macrophages, produce the endogenous inhibitors in response to both CSF-1 and ferritin.

Isolation and functional studies on feline bone marrow derived macrophages.

In this report, we describe an in vitro culture method for feline bone marrow cells, which yields large numbers of quiescent macrophages after 14 days of culture. The bulk of the cultured cell population consists of macrophages as assessed by morphology, macrophage specific cytochemistry, and phagocytosis. The remaining cells were lymphocytes, bone marrow stromal cells, fibroblasts and occasional polymorphonuclear leukocytes. While resting cells produced no detectable interleukin 1, stimulation with lipopolysaccharide (LPS) induced the production of biologically active interleukin 1. After 6 h LPS stimulation, mRNA for tumor necrosis factor alpha and interleukin 1 beta was detectable. The absence of mRNA in unstimulated cells indicates cultured macrophages were not activated until stimulated by LPS or plastic adherence. This approach provides a useful means to measure potential modulatory effects by virus infections or other agents upon feline macrophage gene expression.

Opposing effects of tumor necrosis factor alpha and leukemia inhibitory factor in lipopolysaccharide-stimulated myelopoiesis.

Three myelopoietically active, lipopolysaccharide (LPS)-stimulated monokines, interleukin-1 alpha (IL-1 alpha), tumor necrosis factor alpha (TNF-alpha), and leukemia inhibitory factor (LIF), were tested for effect in an in vitro model for LPS-induced inflammatory murine monocytopoiesis. Neither cytokine stimulated clonal proliferation of marrow-derived progenitors; however, both IL-1 alpha and TNF-alpha enhanced macrophage colony-stimulating factor (M-CSF)-dependent colony formation. The additional progenitors stimulated by IL-1 alpha and TNF-alpha to form colonies in response to M-CSF were equivalent to the precommitment, transitional progenitors stimulated by M-CSF and bacterial LPS. In addition, the additional colonies elicited by IL-1 alpha and TNF-alpha were not additive in cultures containing both M-CSF and LPS, indicating these colonies arose from the same LPS-responsive, two-signal-dependent transitional progenitors. Leukemia inhibitory factor did not influence M-CSF-stimulated colony formation; however, LIF effected a dose-dependent inhibition of colony formation by transitional progenitors responding to combinations of M-CSF and LPS, IL-1 alpha, TNF-alpha, or an additional transitional cell costimulant, substance P. Neutralizing anti-murine TNF-alpha antibodies abrogated transitional cell colony formation stimulated by combinations of M-CSF and TNF-alpha, IL-1 alpha, LPS, or substance P but had no effect on colony formation stimulated solely by M-CSF. The results indicate that TNF-alpha may be an important positive stimulus for commitment of progenitors to the mononuclear phagocyte lineage and that TNF-alpha may be the endogenous regulator of the costimulatory effects of LPS, IL-1, and substance P. In addition, the results indicate that LIF may play an opposing negative regulatory role acting to inhibit LPS and TNF-alpha stimulation of the transitional progenitors.

Nanomolar cyclic adenosine and guanosine monophosphates stimulate macrophage colony-stimulating factor responsiveness by murine transitional progenitors.

Both 3':5' cyclic adenosine monophosphate (cAMP) and 3':5' cyclic guanosine monophosphate (cGMP) stimulated colony-stimulating factor 1 (CSF-1)-dependent colony formation by murine two-signal-dependent progenitors without influencing colony formation by committed CSF-1-responsive progenitors. The stimulatory effect was optimal at 10(-9) M and did not diminish with increasing concentrations of the cyclic nucleotides. The membrane-permeating analogs dibutyryl cAMP and 8-Br-cGMP similarly augmented colony formation by the transitional progenitors at 10(-9) M; however, with increasing concentration, enhancement diminished with eventual inhibition of total colony formation at micromolar concentrations. Stimulation by the two cyclic nucleotides was mutually incompatible. The results indicate that physiological levels of extracellular cyclic nucleotides may significantly influence myelopoiesis. Furthermore, the results introduce the interesting possibility that stimulation, unlike inhibition, may be initiated through an extracytoplasmic mechanism that does not require direct activation of cytoplasmic cyclic nucleotide-dependent protein kinases.

Characterization of a neutralizing monoclonal antibody to Pasteurella haemolytica leukotoxin.

Six hybridoma clones producing monoclonal antibodies (MAbs) reactive with Pasteurella haemolytica A1 leukotoxin were derived from mice immunized with leukotoxin excised from sodium dodecyl sulfate-polyacrylamide gels. Of the six MAbs, only one, Ltx-2, neutralized leukotoxin in a BL-3 cell cytotoxicity assay. MAb Ltx-2 blocked association of A1 leukotoxin to BL-3 cells, as measured by flow cytometric analysis. The epitope recognized by Ltx-2 was localized to the carboxyl half of the native protein, between residues 450 and 939, by Western immunoblot analysis of CNBr fragments. Further analysis with leukotoxin deletion proteins indicated either that the Ltx-2-reactive epitope was localized in the carboxyl portion of the leukotoxin between amino acids 768 and 939 or that this region influences MAb recognition of the epitope. MAb Ltx-2 was tested for neutralizing activity against leukotoxin produced by P. haemolytica serotypes 1 through 12. The MAb neutralized leukotoxin produced by all of the A biotype isolates (serotypes 1, 5, 6, 7, 8, 9, and 12), with the exception of serotype A2, but did not neutralize any T biotype leukotoxin tested (T3, T4, or T10). The results indicate that MAb Ltx-2 neutralizes leukotoxin by interfering with target cell association and that the MAb-specific epitope is either not present or not critical for function in the leukotoxin produced by P. haemolytica serotypes A2, T3, T4, and T10.

Effects of bisphosphate treatment and mechanical loading on bone modeling in the rat tibia.

Bisphosphonate treatment has been shown to decrease endosteal bone formation and increase periosteal bone apposition in the rat tibial diaphysis. This study tested the hypothesis that the increase in periosteal apposition is a compensatory attempt to maintain skeletal mass appropriate for the mechanical load. Twenty-four rats were divided into four groups. In two of the groups, one hindlimb in each rat was immobilized with a sling device to increase the mechanical load on the opposite limb. Daily injections of dichloromethylene bisphosphonate (Cl2MBP) were given at 10 mg/kg to one immobilized group and one mobile group. The other two groups were given daily injections of normal saline. Fluorescent bone labels were administered at two-week intervals. All rats were killed after ten weeks of treatment, and calcified tibial cross sections were prepared for fluorescence microscopy. Bone dimensions and periosteal and endosteal apposition rates were calculated. When compared with saline controls, Cl2MBP treatment decreased endosteal apposition rate in all tibias. Periosteal apposition rate was increased with Cl2MBP treatment in all tibias except the unloaded limb of immobilized rats. The Cl2MBP-induced increase in periosteal apposition rate was greatest in loaded limbs and was proportional to the relative amount of body weight supported.

Pulmonary immunity in calves following stimulation of the gut-associated lymphatic tissue by bacterial exotoxin.

Antibodies in serum and pulmonary lavage fluids were measured in calves following stimulation of the gut-associated lymphatic tissue (GALT) by inoculation of crude leukotoxin of Pasteurella haemolytica into the duodenum through a surgically placed catheter. Nine calves free of P. haemolytica were divided into two groups. Group 1 received an intraduodenal (ID) inoculation of leukotoxin and group 2 received an ID inoculation of phosphate buffered saline. Serum and pulmonary lavage fluids were collected weekly and assayed for antibodies specific to P. haemolytica including immunoglobulin (Ig)G, leukotoxin neutralizing antibodies (LNA), and IgA (lavage fluids only). The multiplicative increase (over baseline) in each class of antibody titer following ID inoculation of leukotoxin, the composite geometric mean increase of all antibodies together, and the composite number of the five antibody titers which increased at least fourfold were computed. Results showed that the geometric mean of each antibody titer and the two composite indices was higher in the GALT-primed groups than in the sham-primed group. The differences were statistically significant (p less than 0.05) for serum IgG and for the two composite indices. This experiment demonstrates for the first time that GALT stimulation by bacterial exotoxins results in increased pulmonary antibody levels in calves.

Differential effects of CD4+ and CD8+ cells in acute, systemic murine candidosis.

Monoclonal antibody (mAb) depletion was used to assess contributions of CD4+ and CD8+ cells in resistance to systemic murine Candida albicans infection. Depletion of CD8+ cells did not influence either survival or mean survival time (MST); however, depletion of CD4+ cells significantly enhanced both survival and MST. Combined depletion of both CD4+ and CD8+ cells significantly lengthened the MST but did not enhance survival. A protective influence of CD8+ cells could be deduced but, to be manifested, required depletion of an overshadowing immunopathologic CD4+ response.

A technique for measurement of intraoral lip pressures with lip bumper therapy.

One of the reported effects of functional appliance therapy is muscular adaptation achieved through the use of vestibular shields. To develop a method for measuring these effects, 10 children with Class I molar occlusion in the late mixed-dentition stage underwent lip bumper therapy for 8 months. Semiconductor pressure transducers mounted on Tru-Tain stents in the mandibular midline and left canine areas were used to measure lip pressures with the patient at rest and during five functional exercises. Resting and speaking lip pressures for the midline transducer showed significant increases 1 month after lip bumper placement and then decreased to near or below pretreatment levels, possibly an adaptive response. Left-side resting pressures also showed a gradual decrease. In contrast, left-side speaking pressures showed no significant changes over time. The swallowing pressures were not replicable because of excessive background fluctuations. The results of this study indicate that this method has potential for improving clinical diagnosis. The preliminary data also suggest an adaptive response of lip muscles that varies according to anatomic location.