Monolayer cultures of normal human bone cells contain multiple subpopulations of alkaline phosphatase positive cells

Summary Cytochemical staining of normal human bone cells in monolayer cultures for alkaline phosphatase (ALP) indicated that the cultures contained mixed-cell populations. Time course evaluations of the cytochemical staining revealed, in addition to the ALP-negative cell population, at least two subpopulations of ALP-positive human bone cells with different levels of ALP. A cytochemical method has been developed which separates the ALP-positive cells into high and intermediate ALP subpopulations. In this method, human bone cells were stained for ALP using an azo-dye method and incubating at 4°C for 10 and 30 minutes, respectively. We defined the cell population that stained positively for ALP at 10 minutes as strong ALP-positive cells, and both strong and intermediate cells were stained at 30 minutes. The intermediate cells were determined from the difference between the values at the two time points. The intra- and interassay variations of the assay, with the same investigator in blinded investigations, were both less than 10% and the interobserver variation was approximately 25%. Analysis of the distribution of ALP levels in cells with a laser densitometer confirmed the presence of at least three cell subpopulations. 1,25(OH)₂D₃ treatment increased the proportions of both ALP-positive cell populations, whereas TGF-beta treatment increased only the intermediate ALP-positive cell population. On the contrary, fluoride increased the proportion of the strong ALP cells, and IGF-1 had no effect on the proportions of either ALP-positive subpopulation. When the ALP-specific activity was compared with the percentage of each ALP-positive subpopulations for the cells treated with effectors, the ALP-specific activity correlated with the total ALP-positive and with the strong ALP-positive populations but not with the intermediate ALP-positive subpopulation. In summary, this study represents the first evidence that normal human bone cells in monolayer cultures contained at least two subpopulations of ALP-positive cells, and that bone cell effectors could have differential effects on each cell population.     

Early and midterm results of the arterial switch operation for transposition of the great arteries in Japan

A total of 267 infants and children who underwent the arterial switch operation for transposition of the great arteries in the 6 years before Jan. 1, 1988 in six main Japanese institutions were entered into this review. The current status of patients surviving more than 1 year after the operation were evaluated along with early operative results. One hundred forty-six patients had an intact ventricular septum, 103 had a significant ventricular septal defect, and 18 had so-called Taussig-Bing anomaly of the transposition type. Eighteen patients were less than 28 days of age, 73 were 1 to 5 months of age, and 176 were older than 6 months of age at the time of operation. The overall mortality rate was 35% in the first 3 years and 12% in the more recent 3 years. There was a significant difference between the overall mortality rate of primary and two-stage repair (22% versus 10%, p = 0.047) in patients with intact ventricular septum. The overall mortality rate in patients with type B or C coronary arteries of Yacoub, and Radley-Smith's classification was significantly higher than that of other types of arteries (86% versus 18%, p = 0.0001). A total of 156 patients survived more than 1 year after the operation, and 44 children (28%) were noted to have supravalvular pulmonary stenosis (greater than 20 mm Hg). This complication was more common in patients operated on in the newborn period. Trivial or mild aortic regurgitation was noted in 29 patients (19%) and was more common in patients with two-stage than with primary repair (24% versus 14%). Aortic regurgitation was significantly more prevalent in patients in whom coronary arteries were implanted into slits or U-shaped defects than in those whose arteries were implanted into punched-out holes made on the pulmonary root (28% versus 8%, p = 0.049). Normal sinus rhythm was present in 97% of 154 patients and left ventricular ejection fraction was within the normal range in 97% of 115 patients at catheterization 1 to 5 months after the operation.     

Effect of high NaCl intake on Na+ and K+ transport in the rabbit distal convoluted tubule

Morphological studies have demonstrated that a chronic increase in distal Na⁺ delivery causes hypertrophy of the distal convoluted tubule (DCT). To examine whether high NaCl-intake also causes functional changes in the well defined DCT, we measured transmural voltage (V _T), lumen-to-bath Na⁺ flux (J _Na(LB)), and net K⁺ secretion (J _K(net)) in DCTs obtained from control rabbits and those on high NaCl-intake diets. The lumen negativeV _T was significantly greater in the high NaCl group than in the control group. The net K⁺ secretion (pmol mm^–1 min^–1) was greater in the high NaCl-intake group (54.1±13.0 vs 14.7±5.6). The K⁺ permeabïlities in both luminal and basolateral DCT membranes, as assessed by the K⁺-induced transepithelial voltage deflection inhibitable with Ba²⁺, were increased in the experimental group. The lumen-to-bath²²Na flux (pmol mm^–1 min^–1) was also greater in the experimental group (726±119 vs 396±65). TheV _T component inhibitable with amiloride was also elevated in the high NaCl-intake group. Furthermore, Na⁺–K⁺-ATPase activity of the DCT was higher in the experimental than in the control group. We conclude that high NaCl intake increases both Na⁺ reabsorption and K⁺ secretion by the DCT. This phenomenon is associated with an increased Na⁺–K⁺-ATPase activity along with increased Na⁺ and K⁺ permeabilities of the luminal membrane, and an increase in the K⁺ permeability of the basolateral membrane. Cellular mechanisms underlying these functional changes remain to be established.     

Encoding-related brain activity during deep processing of verbal materials: a PET study

The recent advent of neuroimaging techniques provides an opportunity to examine brain regions related to a specific memory process such as episodic memory encoding. There is, however, a possibility that areas active during an assumed episodic memory encoding task, compared with a control task, involve not only areas directly relevant to episodic memory encoding processes but also areas associated with other cognitive processes for on-line information. We used positron emission tomography (PET) to differentiate these two kinds of regions. Normal volunteers were engaged in deep (semantic) or shallow (phonological) processing of new or repeated words during PET. Results showed that deep processing, compared with shallow processing, resulted in significantly better recognition performance and that this effect was associated with activation of various brain areas. Further analyses revealed that there were regions directly relevant to episodic memory encoding in the anterior part of the parahippocampal gyrus, inferior frontal gyrus, supramarginal gyrus, anterior cingulate gyrus, and medial frontal lobe in the left hemisphere. Our results demonstrated that several regions, including the medial temporal lobe, play a role in episodic memory encoding.     

Intracisternal injection of TRH antibody blocks gastric emptying stimulated by 2-deoxy-d-glucose in rats

We evaluated the effect of 2-deoxy-d-glucose (2-DG) on gastric emptying of a non nutrient solution in conscious rats using a Phenol red method. Intravenous injection of 2-deoxy-d-glucose dose-dependently increased the rate of gastric emptying. This stimulatory action of 2-DG was abolished by subdiaphragmatic vagotomy. Intracisternal injection of thyrotropin-releasing hormone (TRH) antibody blocked intracisternal TRH and intravenous 2-DG-induced enhancement of gastric empyting but not the stimulation of gastric emptying induced by intracisternal pancreatic polypeptide. The TRH antibody injected intraperitoneally had no effect. These results suggest that endogenous TRH in the brain is involved in vagal-dependent stimulation of gastric emptying by 2-DG.     

Overexpression of cyclin‐dependent kinase 4 protein in extramammary Paget's disease

Cyclin‐dependent kinase 4 and 6 (CDK4/6) plays an important role in cell cycle progression, and the CDK4/6–cyclin D1 complex controls the cell cycle transition from G₁ phase to S phase. CDK4 is enhanced in several types of cancers and CDK4/6 inhibitors attenuate the proliferation of several types of cancer in vitro/in vivo. The purpose of our study was to investigate the expression pattern of CDK4 and evaluate its clinical importance in extramammary Paget's disease (EMPD). Almost all EMPD tissues were positive for CDK4, and metastatic lesions had a similar immunostaining intensity to primary lesions. In addition, CDK4 protein levels were positively correlated with those of cyclin D1 protein. Taken together, CDK4 may assume a crucial role in EMPD progression.     

Mutations in the fifth immunoglobulin-like domain of kit are common and potentially sensitive to imatinib mesylate in feline mast cell tumours

The purpose of the current study was to investigate the mutation status of KIT in feline mast cell tumours (MCTs) and to examine the effects of tyrosine kinase inhibition on the phosphorylation of mutant kit in vitro and in clinical cases of cats. Sequence analysis of KIT identified mutations in 42/62 MCTs (67·7%). The vast majority of the mutations were distributed in exons 8 and 9, both of which encode the fifth immunoglobulin-like domain (IgD) of kit. All five types of kit with a mutation in the fifth IgD were then expressed in 293 cells and examined for phosphorylation status. The mutant kit proteins showed ligand-independent phosphorylation. The tyrosine kinase inhibitor imatinib mesylate suppressed the phosphorylation of these mutant kit proteins in transfectant cells. In a clinical study of 10 cats with MCTs, beneficial response to imatinib mesylate was observed in 7/8 cats that had a mutation in the fifth IgD of kit in tumour cells. Mutations in the fifth IgD of kit thus appear to be common and potentially sensitive to imatinib mesylate in feline MCTs. These data provide an in vivo model for paediatric mastocytosis where mutations in the fifth IgD of kit also occur.     

Delayed Gastric Emptying by Helicobacter pylori Lipopolysaccharide in Conscious Rats

The present study was carried out to investigatethe possibility that lipopolysaccharide deprived fromHelicobacter pylori may alter gastric motility. Toaddress the question, we examined the effect of H. pylori lipopolysaccharide on gastricemptying in conscious rats. Gastric emptying wasevaluated by the phenol red method. Time-course anddose-related effects of intraperitoneal administrationof H. pylori lipopolysaccharide were investigated.Intraperitoneal injection of H. pylorilipopolysaccharide significantly suppressed gastricemptying of a liquid meal in a dose-dependent manner.The inhibitory action of H. pylori lipopolysaccharide wasobserved 2, 4, 8, or 12 hr after the injection. Theseresults suggest for the first time that H. pylorilipopolysaccharide may suppress gastric emptying in along-lasting fashion. It is also suggested that H. pylorimay influence gastric function through its cell wallstructure named lipopolysaccharide.