In the early 1960s I applied 3H-thymidine autoradiography to the study of the cells constituting the neural tube, and found that its wall was composed solely of one kind of single-layered epithelial cell, which perform an elevator movement between the mitotic and DNA-synthetic zones in the wall in accord with the cell cycle. They were identified as multipotent stem cells of the central nervous sytem (CNS) to which I gave the name of matrix cells. ³H-thymidine autoradiography also revealed the chronology of development of these matrix cells: At first they proliferate only to expand the population (stage I), then switch to differentiate specific neuroblasts in given sequences (stage II), and finally change themselves into ependymoglioblasts, common progenitors of ependymal cells and neuroglia (stage III). Based on these findings, I proposed a monophyletic view of cytogenesis of the central nervous sytem. This matrix cell theory claiming the existence of multipotent stem cells has long been the target of severe criticism and not been accepted among neuro-embryologists for a long time. Recent findings by experimental and clinical neuroscientists on the importance of stem cells have renewed interest in the nature and biology of the multipotent neural stem cells. The present paper describes how the concept of the matrix cell (multipotent neural stem cells in vivo) emerged and what has come out from this view over the last 45 years, and how the basic concept of the matrix cell theory has recently been reconfirmed after a long period of controversy and neglect.

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The fibrillar collagen I gel induced the formation of numerous dendritic cell-like protrusions (cell spikes) from the cell body, whereas monomeric collagen I induced typical cell spreading with filopodia and lamellipodia in skin fibroblasts. Peripheral, not central stress fibers appeared upon adhesion to fibrillar collagen gel, whereas both types of fibers were evident upon adhesion to monomeric collagen. Microtubules and vimentin filaments were elongated inside stress fibers along the terminal tip of cell spikes. Spike formation was totally inhibited by nocodazole and severely delayed by cytochalasin D. This suggests that cell spike formation is dependent on microtubules rather than on F-actin. We then investigated the intracellular signaling responsible for cytoskeleton organization to identify the key factor that induces cell spike morphology. During cell spike formation, FAK and CAS were activated. More CAS was activated in cells on fibrillar collagen gel than on the monomeric form, whereas FAK was activated to the same level on either. At 90 min of culture, Rac1 was activated in cells on monomeric collagen I, whereas Cdc42, Rac1 and RhoA were activated in cells on fibrillar collagen gel. These results suggest that microtubule organization via CAS and small GTPases is important for the cell spike formation that is involved in collagen gel contraction and in wound retraction in skin.

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Circulating and nonadherent polymorphonuclear leukocytes (PMNs) become activated to attain adhesive state in an integrin-dependent manner by various stimuli, and perform a variety of microbicidal functions such as phagocytosis and superoxide production. We found that, in the absence of serum, a physiological concentration of hemopexin has a strong inhibitory action on Mg²⁺-dependent adhesion of PMA-activated PMNs to fibrinogen- and serum-coated surfaces. Under these conditions, Ca²⁺ had no effect on Mg²⁺-dependent adhesion or the adhesion-inhibitory activity of hemopexin. In contrast, PMNs suspended in serum containing sufficient amounts of hemopexin to inhibit adhesion showed marked adherence, which was inhibited by EGTA. Next, we prepared a small-molecule fraction of serum by ultrafiltration followed by boiling. PMA-activated PMNs was found to adhere in the presence of both hemopexin and the small-molecule fraction, and the adhesion was enhanced by exogenous Ca²⁺. EGTA abolished the effect of the small molecule fraction. The data suggest that serum contains adhesion-promoting factor(s) which allows PMNs to adhere despite the presence of hemopexin and that Ca²⁺ is required for adhesion-promoting activity. Further study of hemopexin may provide clues for new therapeutic strategies aimed at interfering with PMN adhesion to control inflammation and tissue injury.

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PVLA poly-(N-p-vinylbenzyl-O-β-D-galactopyranosyl-D-gluconamide) is a glycopolymer composed of hydrophilic carbohydrate side chain and hydrophobic styrene polymer. The hydrophilic carbohydrate residue of PVLA can be recognized as a ligand for hepatocytes asialoglycoprotein receptor (ASGP-R), which is abundant on the hepatocyte cell surface. Adhering to the PVLA coated dishes, hepatocytes try to form aggregates that have a long time survival and also cells in these aggregates exhibit better maintenance of specific hepatocyte functions. Stimulation of the cells with IGF-1 in this culture condition results in the formation of lower aggregates. In addition to the morphological influences of IGF-1 to these cells, we have also found that IGF-1 transmits growth stimulatory responses to hepatocytes on PVLA through both mitogen activated protein kinase (MAPK) pathway and β-catenin pathways. The phosphorylation of MAPK can take place within 5 min of stimulation with IGF-1 and within at least 10 ng/ml of IGF-1 concentration. Inhibition of MAPK activation by MEK-1 inhibitor PD98059 reduces IGF-1 induced MAPK phosphorylation, and also IGF-1 stimulated DNA synthesis. Similarly, the use of PI3-K inhibitor LY294002 also inhibits IGF-1 stimulated DNA synthesis. IGF-1 stimulation enhances the migration of β-catenin from the cytoskeleton and cell membrane to the cytoplasm which also is the reason behind formation of spheroids and lower aggregates. IGF-1 stimulation also shows increased translocalization of β-catenin to the nucleus that is essentially important to produce β-catenin responsive effects to the cells. These studies thus suggest that IGF-1 can stimulate the growth and survival of hepatocytes on PVLA through both MAPK and β-catenin signaling pathways, and that the activation of β-catenin signaling pathway produces the morphological changes of IGF-1 induced cells.

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