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Neural stem cells differentiate from embryonic stem cells via formation of neural stem spheres under free-floating conditions in astrocyte-conditioned medium. Subsequent culture of neural stem spheres on an adhesive substrate with fibroblast growth factor-2 promotes the migration of neural stem cells onto the substrate, resulting in an increase in the number of cells. These embryonic stem cell-derived neural stem cells can be differentiated almost exclusively into astrocytes by withdrawing fibroblast growth factor-2 from the medium without any additional instructions.

We have developed a simple method to efficiently produce a large number of neural stem cells and neurons from mouse embryonic stem (ES) cells. When cultured in astrocyte-conditioned medium (ACM) with mitogens (FGF-2 and EGF) under free-floating conditions, colonies of undifferentiated ES cells give rise to neural stem spheres (NSSs), composed of plentiful neural stem cells. Subsequent culture of the NSSs on an adhesive substrate with mitogens results in the migration of neural stem cells onto the substrate. These cells can be expanded, preserved by freezing, and differentiated into functional neurons. Neural stem cells and neurons provided by this NSS method may be valuable as potential donor cells for neuronal transplantation and also as convenient alternatives to tissue-derived neural cells.

The cytotoxicities of lipoproteins of Mycoplasma fermentansand Mycoplasma salivariumto a lymphocytic cell line, MOLT‐4, and a monocytic cell line, HL‐60, was upregulated by ATP added extracellularly in a dose‐dependent manner. These lipoproteins induced ATP release and plasma membrane permeability increase in these cell lines. In addition, periodate‐oxidized ATP, an antagonist for P2X purinergic receptors, suppressed the cytotoxicity of the lipoproteins, suggesting the possibility that P2X receptors for ATP play crucial roles in the cytotoxicity. Activation of caspase‐3 induced by the lipoproteins, which was assessed by the cleavage of the synthetic substrate DEVD‐pNA and the endogenous substrate poly(ADP‐ribose) polymerase, was also upregulated and downregulated by extracellular ATP and periodate‐oxidized ATP, respectively. On the basis of these results, this study suggests that mycoplasmal lipoproteins induce the permeability increase in lymphocytes and monocytes, by which ATP is released, and the ATP regulates the cytotoxicities of the lipoproteins to the cells, possibly by interaction with ATP receptors such as P2X purinergic receptors.

Abstract. The Na+,K+-ATPase (= Na pump), which produces the concentration gradient of Na+and K+across the cell membrane, was studied in diabetic erythrocytes. The activity and number of Na pumps, which are functional and quantitative expression of the Na+, K+-ATPase in erythrocytes, were measured by ouabainsensitive 42K or 43K influx and by [3H]ouabain binding, respectively. The turnover rate of the pumps was calculated from the two measurements to evaluate in situ activity of the pump.The Na pump activity was found to be higher in the diabetic (193 ± 12 nmol K+/h · 109cells) than in the normal group (164 ± 6) (P< 0.05), though the affinities of the pump for extracellular K+were not different. The number of Na pumps and the Kdof the pumps for ouabain in the diabetic group were not significantly different from those in the normal group (354 ± 12 site/cell and 4.33 ± 0.20 nm). The turnover rate of the diabetic group tended to be higher than that of the normal group. The rate was about one third of the molecular activity reported for Na+,K+-ATPase under optimum conditions. These results indicate that the enzymatic properties as well as the number of Na pumps were not altered in diabetic erythrocytes despite the increased Na pump activity. Therefore the increased activity of the erythrocyte Na pump in diabetes mellitus suggests an increase of cation permeability associated with a possible disorder in the diabetic membrane.

Neuron-rich and glial nuclear preparations and liver nuclei were isolated from adult guinea pigs. These nuclei were incubated to carry out DNA-ligation and -synthesis reactions. Before and after incubation, the sizes of single-standed DNA and DNA-synthesis patterns in single strands were analysed by using alkaline sucrose-density-gradient centrifugation. Isolation of nuclei by cell-fractionation technique shortened chromatin DNA and decreased markedly the number-average molecular weight of DNA strands. Chromatin DNA in neuronal and glial nuclei was ligated at the nicks during incubation in a reaction mixture containing ATP, Mg2+, dithiothreitol and four deoxyribonucleotides. The number-average molecular weights were estimated to increase 1.1-and 2.1-fold in neuronal and glial nuclei respectively. DNA strands in liver nuclei were shortened during incubation, but elongated under conditions that inhibit deoxyribonuclease. Since the endogenous deoxyribounuclease activity was conspicuously higher in liver nuclei than in neuronal and glial nuclei, the shortening and elongation were thought to depend on the balance between DNA ligase and deoxyribonuclease reactions. DNA synthesis occurred at the gaps in chromatin DNA and about 50% of the total synthesized DNA was found in the shorter strands having 6 to 297 bases in all species of nuclei. Based on these results, it was concluded that in nuclei isolated from non-dividing cells (neurons) and slowly dividing cells (glial and liver cells) DNA-ligation and -synthesis reactions proceeded in parallel at the breaks in single-stranded DNA, which was produced mainly by endogenous deoxyribonuclease during isolation and incubation processes.

The ontogeny of the GLUT3 glucose transporter gene and protein expression was studied in rat brain. Northern blot analysis using total RNA from rat brains at different developmental stages revealed that the levels of GLUT3 mRNA were very low during the embryonic stage and increased towards the postnatal stage. Immunohistochemistry using a specific antibody showed that the expression of GLUT3 protein was barely detectable in the embryonic stage, but was clearly detected on the plasma membrane of neuronal cells from 10 days after birth to the adult. Expression of GLUT3 mRNA and protein in the cerebral neuronal cell cultures was also examined during the maturation of neurons. GLUT3 glucose transporter of primary neuronal cultured cerebral cortical neurons was only detected in mature neurons after they were cultured for 14 days. These results indicate that GLUT3 plays an important role in glucose homeostasis postnatally in neurons of the rat brain.

Abstract: Introduction: Postoperative alopecia is a relatively rare event, and therefore both patients and surgeons are puzzled once it develops even though it is said to improve spontaneously with time in most cases. We report a parieto-occipital pressure-induced alopecia firstly developed in a patient who had undergone repeated surgery for 10 years after a traffic accident. Case report: A 29-year-old male underwent segmental osteotomy at the upper and lower frontal edentulous areas for distraction osteogenesis. Throughout the operation, he was in the supine position with the hair covered with a paper cap and the head on a plastic vinyl chloride-covered soft foam horseshoe-shaped urethane sponge placed on the horseshoe-shaped headrest. About 2 weeks after the surgery, two patches of parieto-occipital alopecia were observed at the barber’s. It cured gradually during the follow-up visits. Conclusion: The pathophysiology of the condition is thought to be mostly pressure-induced ischemia of hair follicles, and then we have to try not to make the situation. Scalp massages and the head repositioning during the surgery should be one of the means of prevention.