Reduced T cell response to beta-lactoglobulin by conjugation with acidic oligosaccharides.

We have previously reported that the conjugation of beta-lactoglobulin (beta-LG) with alginic acid oligosaccharide (ALGO) and phosphoryl oligosaccharides reduced the immunogenicity of beta-LG. In addition, those conjugates showed higher thermal stability and improved emulsifying properties than those of native beta-LG. We examine in this study the effect of conjugation on the T cell response. Our results demonstrate that the T cell response was reduced when mice were immunized with the conjugates. The findings obtained from an experiment using overlapping synthetic peptides show that novel epitopes were not generated by conjugation. One of the mechanisms for the reduced T cell response to the conjugates was found to be the reduced susceptibility of the conjugates to processing enzymes for antigen presentation. We further clarify that the beta-LG-ALGO conjugate modulated the immune response to Th1 dominance. We consider that this property of the beta-LG-ALGO conjugate would be effective for preventing food allergy as well as by its reduced immunogenicity. Our observations indicate that the method used in this study could be applied to various protein allergens to achieve reduced allergenicity with multiple improvements in their properties.

Degeneration of skeletal muscle fibers in the rat administrated carbon tetrachloride: similar histological findings of the muscle in a 64-year-old patient of LC with muscle cramp.

It is well known that painful muscle cramps occur frequently in patients with advanced liver cirrhosis (LC). Although many studies discuss the pathophysiological causes of these muscle cramps in various conditions, the results are not clear as far as the cause associated to LC is concerned. In order to investigate the cause of muscle cramps in LC, we examined the histological findings of skeletal muscle fibers in LC rat model and in a patient with LC. LC (n=9) was induced in rats by chronic carbon tetrachloride administration. The histological findings of skeletal muscle tissues from the lower leg in LC rats and those of the upper arm in a patient with LC were compared. The degenerated muscle fibers and centronucleus in LC rats were similar to the opaque fibers frequently observed in myotonic dystrophy with severe muscle rigidity in patients with LC. In conclusion, results of this study suggest that one of the causes for muscle cramps in patients with LC is due to skeletal muscle fiber degeneration. Therefore, histological observation of skeletal muscle fibers should be considered in the treatment of painful muscle cramps.

Reduced immunogenicity of beta-lactoglobulin by conjugation with acidic oligosaccharides.

Bovine beta-lactoglobulin (beta-LG) was conjugated with the acidic oligosaccharides, alginic acid oligosaccharide (ALGO) and phosphoryl oligosaccharides (POs) by the Maillard reaction to reduce the immunogenicity of beta-LG. The molar ratios of beta-LG to ALGO and POs in the conjugates were 1:6 and 1:8. The carbohydrate-binding sites in the beta-LG-ALGO conjugate were partially identified to be (60)Lys, (77)Lys, (100)Lys, (138)Lys, and (141)Lys. The isoelectric point of each conjugate was lower than that of beta-LG. CD spectra indicated that the secondary structure of beta-LG was almost maintained after conjugation. The results of fluorescence studies indicated that the conformation around Trp had not changed in each conjugate and that the surface of each conjugate was covered with a saccharide chain. Structural analyses with monoclonal antibodies indicated that the conformation around (8)Lys-(19)Trp (beta-sheet, random coil, short helix) in the conjugates had changed, whereas the native structure was maintained around (15)Val-(29)Ile (beta-sheet) and (125)Thr-(135)Lys (alpha-helix). The beta-LG-ALGO and beta-LG-POs conjugates maintained 77 and 70% of the retinol binding activity of beta-LG. Conjugation with ALGO and POs substantially enhanced the thermal stability of beta-LG. The anti-beta-LG antibody response was markedly reduced after immunization with both conjugates in BALB/c, C57BL/6, and C3H/He mice. B cell epitopes of beta-LG and the conjugate recognized in these mice were determined with 15-mer multipin peptides, and the linear epitope profiles of the conjugates were found to be similar to those of beta-LG, whereas the antibody response to each epitope was dramatically reduced. In particular, effective reduction of the antibody response was observed in the vicinity of the carbohydrate-binding sites. Conjugation of beta-LG with these acidic oligosaccharides was effective in reducing the immunogenicity of beta-LG. The conjugates obtained in this study are edible, so they would be very useful for food application.

Influence of knee alignment on quadriceps cross-sectional area.

Previous studies of methods for stimulating the individual muscles composing the quadriceps femoris have not considered the structural features of a subject's knee joint. In this study, we compared the ratios of the individual muscles composing the quadriceps between subjects with different knee alignments using magnetic resonance (MR) imaging. A total of 18 healthy males were examined: 6 normal knees (age, 23.0+/-0.6 yr; femorotibial angle (FTA), 176.8+/-0.4 degrees), 6 genu varum (age, 21.8+/-2.9 yr; FTA, 181.7+/-2.6 degrees) and 6 genu valgum (age, 21.0+/-1.6 yr; FTA, 172.3+/-1.5 degrees). The cross-sectional areas (CSAs) of quadriceps muscles were obtained by MR imaging of the entire left thigh. The CSAs of the vastus lateralis (VL), rectus femoris (RF), vastus medialis (VM) and vastus intermedius (VI) muscles were obtained by MR imaging of the entire left thigh in a supine position. The VM/VL ratio was also obtained by dividing the CSA of the VM by that of the VL and compared among the three groups of subjects with different knee alignments. The genu varum group showed a significantly higher %CSA of VM in the CSA of the quadriceps (VM/Quad) (49.0+/-2.6%) than values for the other two groups. The genu valgum group showed significantly higher values of RF/Quad (15.2+/-2.1%) and VL/Quad (40.6+/-4.0%) than the other groups. The VM/VL ratio was significantly higher in the genu varum than in values for the other two groups. This difference in CSA, in respect to knee alignment, may be considered when devising muscle training programs.

Resorbability and solubility of zinc-containing tricalcium phosphate.

Using zinc-containing tricalcium phosphate (ZnTCP) as the zinc carrier for zinc-releasing calcium phosphate ceramic implants promoted bone formation around the implants. Because no quantitative information was available on the equilibrium solubility and resorbability of ZnTCP, in vitro equilibrium solubility and in vivo resorbability of ZnTCP were determined and compared quantitatively in this study. The solubility of ZnTCP decreased with increasing zinc content. The negative logarithm of the solubility product (K(sp)) of ZnTCP was expressed as pK(sp) = 28.686 + 1.7414C - 0.42239C(2) + 0.063911C(3) - 0.0051037C(4) + 0.0001595C(5) in air, where C is the zinc content in ZnTCP (mol %). The solubility of ZnTCP containing a nontoxic level of zinc (<0.63 wt %) decreased to 52-92% of the solubility of pure tricalcium phosphate (TCP) in the pH range 5.0-7.4. However, the in vivo resorbed volume of ZnTCP containing the same amount of zinc was much lower than that expected from the in vitro solubility, becoming as low as 26-20% of that of TCP. Cellular resorption of TCP is substantially a process of dissolution in a fluid with an acidic pH that is maintained by the activities of cells. Therefore, the reduction of the resorbability of ZnTCP could be attributable principally to its lowered cellular activation property relative to that associated with pure TCP.