Kohtaro Takei

Kinetic simulation of signal transduction system in hippocampal long-term potentiation with dynamic modeling of protein phosphatase 2A

We modeled and analyzed a signal transduction system of long-term potentiation (LTP) in hippocampal post-synapse. Bhalla and Iyengar [Science 283(1999) 381] have developed a hippocampal LTP model. In the conventional model, the concentration of protein phosphatase 2A (PP2A) was fixed. However, it was reported that dynamic inactivation of PP2A was essential for LTP [J. Neurochem. 74 (2000) 807]. We introduced a dynamic modeling of PP2A; inactivation (phosphorylation) of PP2A by calcium/calmodulin-dependent protein kinase II (CaMKII) in the presence of calcium/calmodulin, self-activation (autodephosphorylation) of PP2A, and inactivation (dephosphorylation) of CaMKII by PP2A. This model includes complex feedback loops; both CaMKII and PP2A are autoactivated, while they inactivate each other. Moreover, we proposed an analysis strategy for model validation by applying the results of sensitivity analysis. In our system, calcineurin (CaN) played an essential role, rather than the activation of protein kinase C (PKC) as documented in the conventional model. From results of the analysis of our model, we found the following robustness as characteristics of bistability in our model: (1). PP2A reactions against calcium ion (Ca(2+)) perturbation; (2). PP2A inactivation against PP2A increase; (3). protein phosphatase 1 (PP1) activation against PF2A increase; and (4). PP2A reactions against PP2A initial concentration. These properties facilitated LTP induction in our system. We showed that another mechanism could introduce bistable behavior by adding dynamic reactions of PP2A.

Mosaic reconstruction of blood vessels in mouse neocortical tissue transplanted into the third ventricle of rat brain

Newborn mouse neocortical tissue was transplanted into the third ventricle of rat brain and the reconstruction and the origin of the blood vessels were investigated by using a monoclonal antibody against mouse endothelial surface antigen-1 (MESA-1). It was clearly demonstrated that some of the blood vessels in the graft originated in the donor mouse neocortical tissue. An India ink perfusion experiment revealed that the blood was supplied to the MESA-1-positive blood vessels. Furthermore, electron microscopic immunohistochemical studies demonstrated the existence of a mosaic reconstruction of blood vessels which consisted of mouse- and rat-derived vascular endothelial cells. It was concluded that the blood vessels originating in the donor tissue and those originating in the host tissue inoculate with each other in the grafted tissue.

Expression of Ia antigen on vascular endothelial cells in mouse cerebral tissue grafted into the third ventricle of rat brain

The mechanisms of the immunological rejection after xenogeneic neural transplantation were investigated with special reference to the expression of class II major histocompatibility complex (MHC) antigen (Ia antigen) on the grafted tissue. Tissue from a newborn mouse cerebral cortex was transplanted into the third ventricle of a 4-week-old rat brain. Infiltration of cytotoxic T-cells into the grafted tissue was investigated immunohistochemically by using a monoclonal antibody (OX-8). The infiltration began 8 days after transplantation and continued until about 4 weeks when the tissue was completely rejected. The expression of Ia antigen was also investigated immunohistochemically. The Ia antigen was first detected in the grafted tissue at 6 days after transplantation. The Ia antigen was considered to be expressed on the vascular endothelial cells judging from the staining patterns and the location of India ink which was perfused from the hosts left cardiac ventricle. The perfusion experiments with India ink also revealed that blood was supplied to the grafted tissue from 5 days after transplantation. These results suggest that the expression of Ia antigen on the vascular endothelial cells renders the grafted tissues competent to initiate and participate in the immune reaction. The results also raise a possibility that the expression of Ia antigen is triggered by blood supplied from the host brain. In addition, the results indicate that the Ia-positive blood vessels do not originate in the host brain but are intrinsic to the grafted tissue.

Chapter 13 Immunological rejection of grafted tissue in xenogeneic neural transplantation

Publisher Summary This chapter discusses the immunological rejection of grafted tissue in xenogeneic neural transplantation. It examines immunohistochemically the expression of Ia antigen, T-cell infiltration, and the reconstruction of blood vessels in mouse neocortical tissue grafted into the third ventricle of rat brain. The central nervous system (CNS) is an immunologically privileged site because of the existence of the blood–brain barrier, a poor lymphatic drainage system, and the lack of expression of major histocompatibility complex (MHC) antigen in the normal brain. However, the accumulating lines of evidence indicate that MHC-incompatible grafts are immunologically rejected in allogeneic or xenogeneic neural transplantation. The reconstruction of blood vessels in the grafted tissue is discussed in the chapter. The elucidation of the mechanisms of the rejection provides not only practical cues in successful grafting after cross species neural transplantation but also in the clarification of the etiology of the nervous diseases, which are associated with immune responses.

Major glycoproteins in carp CNS myelin: Homology to P0 protein with HNK-1/L2 carbohydrate epitope

We studied the myelin protein profiles of carp from a phylogenetic point of view. The carp central nerve myelin contained two reactive bands, 28 and 25 kDa, demonstrated with anti-bovine P0 antibody. Their molecular weights are slightly different from those of two positive bands found in carp peripheral myelin. The N-terminal amino acid sequences of these four positive bands were identical to one another and showed high homology with those of mammalian P0 protein, suggesting that carp central myelin contains the P0-like protein. Lectin binding analysis revealed that carbohydrate structure of the P0-like proteins in carp central myelin is similar to those in peripheral myelin of carp and other vertebrates. Further, the carp P0-like glycoproteins, like the P0 proteins of other vertebrates, reacted with antibodies that recognize the HNK-1/L2 carbohydrate epitope. We conclude that the major structural glycoprotein in central myelin of the carp is homologous to P0 protein in peripheral myelin of other higher classes of vertebrates.

Expression of a P0-like glycoprotein in central nervous system myelin of amphibians (Ambystoma mexicanus, Xenopus laevis and Rana catesbeiana)

1. The myelin protein profiles in the CNS and PNS of three species of amphibians were analyzed by biochemical and immunohistochemical methods. 2. The CNS myelin of the African clawed frog (Xenopus) and the Mexican salamander (axolotl) contained, in addition to proteolipid protein, a unique protein zero (P0)-like protein, whereas the adult bullfrog did not. 3. A strong expression of the P0-like protein in the bullfrog CNS myelin was found transiently at ontogenetically early phases including at the time of metamorphosis. 4. The CNS P0-like protein and the PNS P0 protein showed a difference in reactivity with lectins and anti-L2/HNK-1 antibodies, suggesting that the two proteins differ in some aspects of their carbohydrate structures.

Nuclear membrane antigen specific to nerve and muscle tissues.

A monoclonal antibody, 2F7, raised against a nuclear protein subfraction recognized the nuclear membrane of nervous and muscular tissues of guinea pig, rat and rabbit, but no other tissue was stained. In the nervous system, both neurons and glial cells were labelled. An electron microscopic immunohistochemical study demonstrated that 2F7 antibody labelled the inner surface of the nuclear membrane. Western blot analysis on the nuclear envelope fraction containing nuclear lamina revealed that this antibody reacted to two minor component proteins of 80 and 82KDa. These 2F7 antigens expressed preferentially in the nervous and muscular tissues were distinct from the major nuclear envelope proteins reported so far and might be related to neuronal or muscular tissue-specific functions.

Xenogeneic neural transplantation: role of vasculature and MHC antigen in immunological rejection.

The reconstruction of blood vessels in a graft is one of the important events inducing immunological rejection. First, we investigated the reconstruction of blood vessels in a graft by using a monoclonal antibody against mouse endothelial surface antigen 1. Secondly, the lymphocyte proliferative response to Ia antigen were examined in mixed lymphocyte cultures by using monoclonal antibodies against Ia antigen. The results show that the blood vessels originating in the host tissue inoculate with those originating in the donor graft tissue and that the expression of Ia antigen on the vascular endothelial cells plays an important role in the immunological rejection through sensitization of peripheral lymphocytes.