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.     

Results of physical therapy for idiopathic clubfoot: a long-term follow-up study

Physical therapy without anesthesia or plaster casts was used to treat 338 cases of clubfoot (CF). Our technique is based on progressive sequential manipulations at birth. We first reduce the varus and later the equinus component of the CF. The gentle stretches used in this technique are complemented by active physiotherapy stimulating the muscles, and then a simple splint is suited to the foot to fix its degree of realignment. When used alone, this technique achieves 77% good and fair results. In resistant cases, complementary surgery was used. We obtained 96% good and fair results.     

Massive intratubular hemorrhage with herniations into renal veins: report of a case

A 24-year-old man had unilateral gross hematuria that required nephrectomy. Pathological examination revealed massive intratubular hemorrhage and frequent deposition of an amorphous and homogeneous material positive for periodic acid, Schiff stain in the corticomedullary junction. This substance had the characteristics of Tamm-Horsfall protein and frequently herniated into the lumen of thin-walled veins of arcuate size. There was no apparent cause for the bleeding. To the best of our knowledge this is the third reported case with these peculiar findings and no apparent cause. We discuss some hypotheses as to the etiopathogenesis of this rare and intriguing condition.     

Blood level of phosphatidylinositols as a criterion of diagnosis and control of the effectiveness of surgical treatment in gastric cancer

In patients with gastric cancer, the content of phosphatidylinositols in the blood was significantly decreased, in average 2-fold, as compared with their level in donors. After surgical treatment in 36 patients, these indices increased and achieved a maximum value in a year. In these patients, the general state improved, there were no disease recurrences. In 44 patients after the operation, the content of phosphatidylinositols remained low, or increased insignificantly as compared with their level before the operation. In this group, 16 patients died, 10 developed a disease recurrency.     

Primary adenocarcinoma of the appendix

Thirteen cases of primary appendicular adenocarcinoma are reported. This rare tumour usually presents as acute appendicitis. The correct diagnosis is rarely entertained before or during surgery. The main treatment choice lies between appendicectomy alone and appendicectomy followed by right hemicolectomy. There are anatomical reasons for advising the latter and survival figures from the literature tend to support this preference. The cases reported here indicate that in the period 1972-1984, in the North West Region, there was a slight preference for appendicectomy alone. The additional procedure of right hemicolectomy did not confer any clear-cut survival advantage.     

Cytochrome P450-dependent arachidonic acid metabolism in human kidney

Cytochrome P450-dependent arachidonic acid metabolism in human kidney cortex from several postmortem subjects has been characterized. Using HPLC and GC/MS, four cytochrome P450-arachidonic acid metabolites were tentatively but not unequivocally identified as epoxyeicosatrienoic acid (EET), dihydroxyeicosatrienoic acid (DHT) and 19- and 20-hydroxyeicosatetraenoic acids, suggesting the involvement of two major cytochrome P450 enzymes, epoxygenase and omega/omega-1 hydroxylases. This pattern of metabolism was similar to that found in rabbit and rat kidneys. The formation of these metabolites was dependent on the presence of NADPH and inhibited by IgG of NADPH-cytochrome P450 (c) reductase. Immunologic studies of renal cytochrome P450 epoxygenase demonstrated that antibodies prepared against human-purified hepatic cytochrome P450 epoxygenase recognized renal enzyme protein and inhibited the enzyme activity by 92%. In contrast, control immunoglobulin did not inhibit renal cytochrome P450 epoxygenase. Antibody inhibition of renal cytochrome P450 epoxygenase demonstrated a degree of conservation of both enzyme proteins between liver and kidney. Antibodies against lauric acid omega/omega-1 hydroxylases (P450 omega) inhibited the formation of omega/omega-1 hydroxylase products, 19- and 20-HETEs. Identical qualitative patterns of arachidonic acid metabolites were observed in all cortical microsomes studied. Interindividual variations were observed in the cytochrome P450-dependent arachidonic acid metabolism, and the activities ranged from 0.031 to 5.027 nmol arachidonic acid converted/mg protein/30 min. which is about a 150-fold difference. However, when the specific activities for total cytochrome P450-dependent arachidonic acid metabolism were calculated, two separate groups could be distinguished, high and low metabolizers of arachidonic acid.(ABSTRACT TRUNCATED AT 250 WORDS)     

Potentiation of cholinergic activity with pyridino[1,2-a]imidazo[5,4-b]indole: in vitro studies

1. Effects of a novel imidazoindole derivative on cholinergic function were studied in isolated tissue preparations. 2. The compound demonstrated a dose-dependent (10(-11)-10(-9) potentiation (20-60%) of acetylcholine induced tension in guinea pig ileal tissue. 3. Increases in the size of end-plate potentials and nerve evoked muscle twitches were observed in frog nerve-skeletal muscle preparations. 4. Cholinesterase activity was not inhibited. 5. The results suggest that the compound has actions at the post-synaptic muscarinic receptor complex in smooth muscle and causes pre-synaptic increases in ACh release at the neuromuscular junction.     

Absorption of a novel prodrug of L-dopa, L-3-(3-hydroxy-4-pivaloyloxyphenyl)alanine (NB-355). In vitro and in situ studies

Absorption mechanism and absorption site of a prodrug of L-DOPA, L-3-(3-hydroxy-4-pivaloyloxyphenyl)alanine (NB-355, 1) was investigated using rats. Prodrug 1 (0.5 mM) was taken up by intestinal tissue segments time-dependently in vitro at pH 6.0. However, the rate of uptake was less than that of L-dopa. Inhibitors of the amino acid active transport system (L-Phe, dinitrophenol, ouabain) had no effect on the uptake of prodrug 1. In the intestinal tissue segments, prodrug 1 was extensively hydrolyzed by diisopropylfluorophosphate-sensitive esterase(s). To characterize the absorption site, gastrointestinal tracts were ligated to make acute loops in situ and prodrug 1 or L-dopa was injected into the loops. L-dopa disappeared rapidly from the lumen of the jejunum. In contrast, prodrug 1 disappeared rapidly from the ileum rather than the duodenum or jejunum. From these results, it was suggested that prodrug 1 was slowly absorbed primarily from the lower small intestine.     

A novel method for culturing sweat gland epithelia: comparison of normal and cystic fibrosis tissues.

A new method is described to produce epithelial sheets by direct explantation of human sweat glands onto matrigel-coated millipore filters. The method is applicable to whole glands, separated coils or ducts and to normal and CF tissues. Electrogenic transport studies show that epithelia develop sodium transporting capability, even when explants are derived from secretory coils.     

Effects of hypothermia on testicular ischemia

The ischemic effects of prolonged testicular torsion have been well documented; however, prevention or arrest of the damaging effects of prolonged ischemia has been incompletely studied. Two groups of Sprague-Dawley rats were subjected to varying lengths of bilateral testicular ischemia. Group I underwent normothermic ischemia for two, four, and six hours. Likewise, Group II underwent similar time periods of ischemia, however, after thirty minutes of normothermic ischemia the scrotum of each animal was placed into an ice bath maintained at 4C. Two weeks postoperatively, bilateral orchiectomy was performed. Histology of the testes of the two groups was compared. Neither group revealed significant destruction of the germinal epithelium after two hours of ischemia. Group I revealed only 25% preservation of the germinal epithelium at four hours and only 8% preservation at six hours of ischemia. In contrast, Group II which received ice showed 90% preservation of germinal epithelium at four hours and 85% preservation at six hours of ischemia. We conclude that external ice application significantly preserves seminiferous tubules at four and six hours of ischemic injury in the rat testicle.