Studies on the molecular evolution (affinity maturation) of antibodies: unexpected encounter

Antibodies are unique proteins in that they improve their affinity for antigens during antibody responses. This process termed “affinity maturation” is the result of random hypermutation of immunoglobulin genes in B cells followed by selection of the clones that acquired high-affinity antibodies. However, what mechanisms are responsible for the induction of hypermutation and the subsequent B cell selection in germinal centers has remained unknown in many respects. I would like to talk about our long-lasting studies on these immunologic mechanisms focusing on the functional role of follicular dendritic cell

Regulatory Role for Complement Receptors (CD21/CD35) in the Recombination Activating Gene Expression in Mouse Peripheral B Cells

A population of peripheral B cells have been shown to express recombination activating gene products, RAG-1 and RAG-2, which are considered to be involved in revising the B cell antigen receptor (BCR) in the periphery. BCR engagement has been reported to turn off RAG expression in peripheral B cells, whereas the same treatment has an opposite effect in immature B cells in the bone marrow. In contrast to receptor editing that is involved in the removal of autoreactivity in immature B cells, it has been shown that secondary V(D)J rearrangement in peripheral B cells, termed receptor revision, contributes to affinity maturation of antibodies. Here, we show that RAG-2 expression in murine splenic B cells was abrogated by the coligation of BCR with complement receptors (CD21/CD35) much more efficiently than by the engagement of BCR alone. On the other hand, the same coligation augmented proliferation of anti-CD40-stimulated B cells. Consistent with these observations, RAG-2 expression was lower in the draining lymph nodes of the quasi-monoclonal mice when they were immunized with a high-affinity antigen than with a low-affinity one. These findings suggest a crucial role for CD21/CD35 in directing the conservation or the revision of BCRs in peripheral B cells

Significance of Trans Fatty Acids in Cancer.

Trans fatty acids (TFAs) are a recent focus of health problems. TFA is a definitive risk factor for cardiovascular diseases and the death. TFA is also possible risk factor for Alzheimer's disease, obesity, diabetes, fatty liver, and ovulation infertility. The relationship between TFA and carcinogenic risk is controversial; however, TFA is reported to increase the risk of breast cancer and prostate cancer. Elaidic acid (EA), a trans form of oleic acid, enhances cancer cell growth, invasion, and anti-apoptotic survival. In animal models, EA promotes tumor growth and metastasis to the lung, liver, and peritoneum. EA induces sternness in cancer cells through transactivation of EFGR via SRC from GPR40/120 as receptors in EA-integrated cholesterol rafts. Activated EGFR relays the signals to activate canonical and non-canonical wnt pathways and to inactivate notchl pathways. EA also increases miR-494, which inhibits cell differentiation through decrease of target genes. Continuous EA feeding with dosage alteration increased cancer cell sternness. EA diminishes the efficiency of 5-fluorouracil by increasing residual cancer stem cells. These findings suggest that TFA is a relevant cancer promoting factor. The decision to remove TFA from foods made by the FDA might have an impact on cancer clinics

Generation of IgM and IgG1 monoclonal antibodies with identical variable regions: comparison of avidity

Generally, IgM antibodies (Abs) produced in a primary immune response show lower affinity for an inducing antigen (Ag) compared with the corresponding IgG Abs that are major switched isotypes formed in the secondary response. An IgM molecule is a pentamer with 10 Ag-binding sites that will contribute to an increase of avidity for an Ag. To estimate the contribution of the pentameric structure to the avidity of an IgM Ab, we generated IgM and IgG1 monoclonal Abs (mAbs) with identical V regions that are specific for 4-hydroxy-3-nitrophenylacetyl (NP) by in vitro class switching of B cells followed by the cell fusion with a mouse myeloma cell line. Compared with an anti-NP IgG1 mAb, the corresponding IgM mAb showed much higher avidity for NP-conjugated bovine serum albumin, which was drastically reduced after being dissociated into monomers

Effects of Nanosecond Laser Fabrication on Bioactivity of Pure Titanium

AbstractWe are developing surface modification techniques for dental implants with the aim of reducing the time required to realize good adhesion between bone and implant surfaces. A nanosecond Nd:YVO4 laser was used to modify the surfaces of commercially pure titanium (CP Ti) disks and their bioactivities were then evaluated. The surfaces of the CP Ti disks were covered by lines after laser treatment. This treatment created complex microasperities of titania with rutile and anatase crystal structures. This results in the formation of hydroxyapatite on surfaces immersed in 1.5-times concentrated simulated body fluid for 7 days, whereas no hydroxyapatite was observed on conventionally polished surfaces that were immersed under the same conditions. This indicates that laser treatment improves the bioactivity of CP Ti, which is a critical property for osseointegrated implants

Genetic manipulation of an exogenous non-immunoglobulin protein by gene conversion machinery in a chicken B cell line

During culture, a chicken B cell line DT40 spontaneously mutates immunoglobulin (Ig) genes by gene conversion, which involves activation-induced cytidine deaminase (AID)-dependent homologous recombination of the variable (V) region gene with upstream pseudo-V genes. To explore whether this mutation mechanism can target exogenous non-Ig genes, we generated DT40 lines that bears a gene conversion substrate comprising the green fluorescent protein (GFP) gene as a donor and the blue fluorescent protein (BFP) gene as an acceptor. A few percent of the initially BFP-expressing cells converted their fluorescence from blue to green after culture for 2–3 weeks when the substrate construct was integrated in the Ig light chain locus, but not in the ovalbumin locus. This was the result of AID-dependent and the GFP gene-templated gene conversion of the BFP gene, thereby leading to the introduction of various sizes of GFP-derived gene segment into the BFP gene. Thus, G/B construct may be used to visualize gene conversion events. After switching off AID expression in DT40 cells, the mutant clones were isolated stably and maintained with their mutations being fixed. Thus, the gene conversion machinery in DT40 cells will be a useful means to engineer non-Ig proteins by a type of DNA shuffling

Fabrication of small aspheric moulds using single point inclined axis grinding

Single point inclined axis grinding techniques, including the wheel setting, wheel-workpiece interference, error source determination and compensation approaches, were studied to fabricate small aspheric moulds of high profile accuracy. The interference of a cylindrical grinding wheel with the workpiece was analysed and the criteria for selection of wheel geometry for avoiding the interference was proposed. The grinding process was performed with compensation focused on two major error sources, wheel setting error and wheel wear. The grinding results showed that the compensation approach was efficient and the developed grinding process was capable to generate small aspheric concave surfaces on tungsten carbide material with a profile error of smaller than 200. nm in PV value after two to three compensation cycles