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We investigate the application of machine learning Agent Based Modelling (ABM) techniques to model and forecast various financial markets including Foreign Exchange and Equities, especially models that could reproduce the time-series properties of the financial variables. We model the economy by considering non-equilibrium economics. We adopt the features that are required for modelling non-equilibrium economics using ABMs and replicate the non-equilibrium nature of the financial markets by considering a set of bounded rational heterogeneous agents, with different strategies that are ranked according to their performance in the market. We consider markets where there are different agents interacting among themselves and forming some sort of patterns. For example, the patterns are equity prices or exchange rates. While the agents have been interacting in the artificial market, the generated patterns (price dynamics) they co-produce would match with the real financial time-series. In order to get the best fit to the real market, we need to search for the best set of artificial heterogeneous agents that represents the underlying market. Evolutionary computing techniques are used in order to search for a suitable set of agent configuration in the market. We verify the forecasting performance of the artificial markets by comparing that with the real financial market by conducting out-of-sample tests.

Background: Plasma apolipoprotein (apo) E, a sialoprotein, plays an important role in reverse cholesterol transport. Previously, we showed that chronic alcohol consumption impairs glycosylation of apo E in rat liver. Peritoneal macrophages are another significant apo E synthesis site. Objective: The main purpose of this study was to determine the effects of chronic alcohol feeding of rats on the synthesis, sialylation, and sialic acid content of macrophage apo E and its ability to bind to the HDL 3 molecule in vitro. Design: Rats were fed an alcoholic diet or an isoenergetic control diet for 8 wk, after which peritoneal macrophages isolated from them were cultured and analyzed for apo E metabolism. Results: Macrophages from alcohol-fed rats accumulated 33.3% more (P < 0.05) cholesterol than did those from control rats when incubated with acetylated LDL. These macrophages showed a 51-57% lower relative sialylation rate of apo E (P < 0.001) but no significant difference in relative protein synthetic rate. The sialic acid content of the intracellular and secreted forms of apo E was reduced by 41.8% (P < 0.001) and 50.3% (P < 0.001), respectively, with chronic alcohol treatment. Secretion of newly synthesized apo E was impaired by 53.7% (P < 0.001) and 26.1% (P < 0.001) in the absence and presence of HDL in the medium, respectively. Macrophages of alcohol-treated rats secreted apo E with 47.6-67.2% lower (P < 0.001) HDL 3 binding ability; binding ability was restored completely by resialylation of the desialylated apo E. Conclusion: In rats, an alcohol-mediated decrease in sialylation rate resulting in loss of sialic acid residues in apo E impairs the ability of apo E to bind to HDL and consequently in defective reverse cholesterol transport.

Clusterin is a N-glycosylated sialoglycoprotein present in rat brain cells. Clusterin, which elicits aggregation in a wide variety of cells, has been suggested to play an important role in synaptic remodeling through its cell adhesion property or lipid transport capacity in the brain. Sialic acid residues in clusterin may be responsible for its structural conformation, stability and functional ability. Maturation of clusterin is governed by the relative actions of sialyltransferases and sialidases that are present in brain microsomes, golgi bodies, cytosol and plasma membranes. We have earlier reported that chronic ethanol treatment in rats has a damaging effect on the hepatic glycosylation machinery. Others have reported increased hydrolysis of brain sialoconjugates in rats following chronic ethanol administration. Specificity of the effects of chronic ethanol treatment in the brain in relation to the glycosylation process, is still obscure. Therefore, in this investigation, we have studied the specific effects of chronic ethanol treatment on the glycosylation of rat brain clusterin and the causes that may lead to any possible defects in the glycosylation process. We have determined the effects of chronic ethanol treatment on (i) the incorporation of labeled leucine and N-acetylmannosamine into immunoprecipitable clusterin in whole brain homogenate, microsomes, golgi, cytosol, plasma membrane and synaptosomes, (ii) enzymatic activities of sialyltransferases in golgi and synaptosomes, and sialidase in brain cytosol and plasma membranes, and (iii) de novo synthetic rate of rat brain cytosolic sialidase. Our results showed that chronic ethanol treatment in rats resulted in (1) a decreased sialation index of brain clusterin by 47. 2% (p

A cellular carotenoid-binding protein was purified to homogeneity from beta-carotene-fed ferret liver utilizing the following steps: ammonium sulfate precipitation, ion exchange, gel filtration, and affinity chromatography. The final purification was 607-fold. [14C]beta-Carotene co-purified with the binding protein throughout the purification procedures. SDS-PAGE of the purified protein showed a single band with an apparent molecular mass of 67 kDa. Scatchard analysis of the specific binding of the purified protein to beta-carotene showed two classes of binding sites, a high affinity site with an apparent Kd of 56 x 10(-9) M and a low affinity site with a Kd of 32 x 10(-6) M. The Bmax for beta-carotene binding to the high affinity site was 1 mol/mol, while that for the low affinity site was 145 mol/mol. The absorption spectrum of the complex showed a 32-nm bathochromic shift in lambdamax with minor peaks at 460 and 516 nm. Except for alpha-carotene and cryptoxanthin, none of the model carotenoids or retinol competed with beta-carotene binding to the protein. Thus, a specific carotenoid-binding protein of 67 kDa has been characterized in mammalian liver with a high degree of specificity for binding only carotenoids with at least one unsubstituted beta-ionone ring.

In recent years, a number of studies have been reported that have clearly established that hepatic glycosylation machinery is affected by chronic ethanol treatment in rats. We have previously reported that chronic ethanol treatment in rats resulted in decreased glycosylation of transfenin and apolipoprotein E with concomitant decreases in enzymatic activities of Golgi galactosyltransferases and sialyltransferases. In all these studies investigators have invariably used the well-accepted dietary formulation of alcohol diet as proposed by Lieber and DeCarli. However, questions were raised whether the lower carbohydrate content in Lieber's alcohol diet may be responsible for observed effects of ethanol on hepatic glycosylation machinery. Therefore, to verify whether or not the crucial effects of chronic ethanol treatment on hepatic glycosylation machinery as observed in our studies, were truly caused by ethanol, we have extended our studies on protein glycosylation with the inclusion of a third dietary group that was compensated for carbohydrate content. In this investigation, rats were fed with three dietary regimen corresponding to control, ethanol, and carbohydrate compensated ethanol group and studies were done on (i) labeled leucine, galactose and N-acetylmannosamine incorporation into transferrin and apolipoprotein E, and (ii) hepatic galactosyltransferase and sialyttrans-ferase activities in Golgi rich fraction in rat. Our results clearly showed that regardless of the carbohydrate content, marked decreases in the incorporation of labeled sugars into transferrin and the enzymatic activities of galactosyltransferase and sialyltrans-ferase occurred in rats administered chronic ethanol. Thus, it is reasonable to conclude that it is not the carbohydrate content of the diet but ethanol per se, when administered chronically, greatly impairs the glycosylation machinery of rat liver and that the magnitudes of these effects are selectively specific with regard to the type of sugar or the glycosylation enzyme.

Carbohydrate residues of glycoproteins play important roles in their functions. We have previously shown that long-term ethanol treatment in rats alters the normal glycosylation pattern of plasma transferrin and apolipoprotein (apo) E. Glycosylation of proteins is a posttranslational process that is regulated by both glycosyltransferases and glycosidases, the resident enzymes of hepatic subcellular organelles. In this investigation using rat transferrin and apo E as model N- and O-glycosylated proteins, respectively, we have explored the effects of long-term ethanol treatment on the (1) incorporation of various labeled sugar precursors into these specific glycoproteins, (2) activities of mannosyltransferase, galactosyltransferase, and sialytransferases, and (3) hepatic synthetic rate of N-acetyl glucosamine (GlcNAc) alpha 2,6-sialyltransferase (2,6-ST). The relative ratio of labeled sugar to leucine incorporation (glycosylation index) showed a 43% (P < .01) decrease for relative mannosylation of transferrin molecule at both the microsomal and Golgi level in the ethanol group (AN) versus the control group (CN). For apo E, relative mannosylation was reduced by 48.9% (P < .01) and 46.9% (P < .01), respectively, at the microsomal and Golgi level in the AN versus CN. More importantly, relative sialation of transferrin was reduced by 86% (P < .001) in AN as compared with CN. Relative sialation of apo E was reduced by 35% (P < .01) in AN as compared with CN.(ABSTRACT TRUNCATED AT 250 WORDS)

Hepatic activities of rate limiting enzymes in fatty acid and cholesterol synthesis and cholesterol degradation were determined in lean and obese LA/N-cp rats. The hepatic activities of acetyl-CoA carboxylase and fatty acid synthetase, the key enzymes of fatty acid synthesis and 3-hydroxy-3-methylglutaryl coenzyme A reductase (the rate limiting enzyme in cholesterol synthesis), were increased 2-fold in the obese rats as compared with their lean littermates. In contrast, the activity of cholesterol 7α-hydroxylase, the rate limiting enzyme of cholesterol degradation to bile acids, was significantly decreased by 28% in the obese group as compared with the control group. Significantly, compared with the control group, the obese animals exhibited similar magnitudes of differences in the activities of the above enzymes even when they were pair-fed with the control animals. Thus these differences in the obese group are not due to hyperphagia but possibly to hypersecretion of the lipogenic hormone, insulin in this strain. These results indicate that the LA/N-cp obese rat has twice the capacity to synthesize body fat and cholesterol but has a reduced capacity to degrade the cholesterol, leading to increased accumulation of cholesterol and fat.

Transferrin is a N-glycosylated glycoprotein and plays an important role in iron transport from sites of absorption and storage to sites of utilization. Chronic ethanol alters the normal microheterogeneity pattern of transferrin as a consequence of changes in the sialic acid content. However the underlying basis of this change in sialic acid contents of transferrin in alcohol abuse remains unclear. We have undertaken this study in order to investigate the effects of chronic ethanol in rats with respect to the hepatic rate of (i) transferrin synthesis based on labeled leucine incorporation, (ii) the incorporation of labeled N-acetyl mannosamine (NAM) into sialic acid residues of transferrin, and (iii) roles of specific sialyltransferase and sialidase at hepatic subcellular level. The results showed no significant difference in the incorporation of labeled leucine into transferrin at all levels between the control and ethanol group, whereas the incorporation of NAM into transferrin was significantly decreased by 84% (p < 0.001) both at the whole cell and Golgi level. Thus, the incorporation of labeled NAM relative to the incorporation of labeled leucine into hepatic transferrin was significantly decreased by 86% (p < 0.001) in chronic ethanol-treated animals as compared with the controls both at the whole cell and Golgi levels. These data are further supported by our finding of concomitant decrease in the activity of β-galactoside α2,6-sialyltransferase by 58% (p < 0.01) in ethanol-treated rats as compared with control animals. In contrast, both the plasma membrane and plasma sialidase activities were increased by 95% (p < 0.01) and 85% (p < 0.01), respectively, in ethanol-fed rats as compared with their respective controls. We conclude that decreased activity of sialyltransferase and increased activity of sialidase sequentially at the plasma membrane and plasma compartment may be responsible for decreased incorporation of sialic acid residues in serum transferrin molecules after chronic ethanol treatment.

This article summarizes content proceedings of a symposium held at the 2004 Research Society on Alcoholism Scientific Annual Meeting in Vancouver, Canada. The chairs were Friedrich M. Wurst and Raye Litten. The presentations were (1) Introduction, by Raye Litten; (2) Direct Ethanol Metabolites--On the Threshold From Science to Routine Use, by Friedrich M. Wurst; (3) Sialic Acid Index of Plasma Apolipoprotein J (SIJ) as a Viable Marker for Chronic Alcohol Consumption, by Philippe Marmillot; (4) The Emergence of Ethyl Glucuronide (EtG) Testing as a Tool in Monitoring Healthcare Professionals, by Gregory E. Skipper; (5) Application of Biomarkers for Alcohol Use Disorders in Clinical Practice, by Tim Neumann; (6) Utility of Biomarkers in Assessing the Efficacy of Medications for Treating Alcoholism, by Marty Javors; and (7) Discussion, by Raye Litten.

Background: Shoulder joint instability results in great disability and it is possible to get good results of treatment in selected patients in whom intraarticular pathologies are diagnosed and treated appropriately. Aims: To investigate the pathology of unstable shoulder joint and study the effect of arthroscopic stabilization for the management of traumatic anterior glenohumeral instability and success rate of all arthroscopic Bankart repair. Materials and Methods: This prospective study was conducted in a tertiary care hospital of West Bengal in a time span of 1 year, after getting the approval of the Institutional Ethical Committee and informed consent of the patients. Patients aged between 20 and 40 years with post-traumatic recurrent anterior dislocation of the shoulder were included. Patients were operated arthroscopically and followed up. Results were analyzed per the assessment criteria of Rowe scale. Results: Ten percent had fair and 90% had poor Rowe score during preoperative period. Bankart lesion was present in 16 patients out of 20 patients in whom we performed arthroscopic Bankart repair with suture anchors. At last follow up, 13 patients were able to work above shoulder level normally; two patients had mild limitation of work. One patient had moderate type of restriction of activities. This patient had a history of trauma 3 months after operation. Conclusions: Overall 93.75% patients had excellent to good result in the present study. Based on this short series and minimum follow-up we recommended, this method of management for shoulder instability due to Bankart lesion.

Although plasma carbohydrate-deficient transferrin (CDT) is considered a viable biochemical marker for chronic alcohol consumption, it is valid only when an individual's daily alcohol consumption exceeds 60 g. In addition, it is less sensitive in women drinkers than in men drinkers. We have established that chronic alcohol consumption impairs the hepatic sialylation of a number of glycoproteins by specifically down-regulating Gal-beta-1,4GlcNAc alpha2,6-sialyltransferase mRNA. Significantly, we found that chronic ethanol consumption markedly inhibits hepatic sialylation of apolipoprotein J (Apo J), a 70-kDa N-glycosylated protein of plasma HDL. Because the sialic-acid index of Apo J (SIJ; moles of sialic acid per mole of Apo J protein) is approximately seven times more than that for transferrin (28 vs. 4), we have evaluated whether plasma SIJ would be an even more sensitive marker for chronic ethanol consumption than CDT in both rats and human subjects. The method involves immunoaffinity purification of plasma HDL-Apo J, followed by its sialic acid determination. We have found that chronic ethanol feeding resulted in loss of sialic acid residues of plasma HDL-Apo J in rats. This loss of sialic acid was positively correlated with both amount and duration of ethanol treatment. In human subjects, an intake of about 60 g of alcohol for 30 days led to almost 50% (P.01) depletion of sialic acid from plasma HDL-Apo J. Further, we established that there was a positive correlation of alteration in SIJ with alcohol consumption, detoxification, abstinence, and relapse in human alcohol-dependent patients (sensitivity, 90%-92%). In addition, plasma SIJ was decreased by 50%-57% (P.01) in both male and female alcohol-dependent subjects. We suggest that plasma SIJ can be used as a viable marker for early detection of chronic alcohol consumption in human beings.

The Silicon Tracking System (STS) is the central detector of the Compressed Baryonic Matter (CBM) experiment at future Facility for Anti-proton and Ion Research (FAIR) at Darmstadt. The task of the STS is to reconstruct trajectories of charged particles originating at relatively high multiplicities from the high rate beam-target interactions. The tracker comprises of 300 μm thick silicon double-sided micro-strip sensors. These sensors should be radiation hard in order to reconstruct charged particles up to a maximum radiation dose of 1 × 1014neqcm−2. Systematic characterization allows us to investigate the sensor response and perform quality assurance (QA) tests. In this paper, systematic characterization of prototype double-sided silicon micro-strip sensors will be discussed. This procedure includes visual, passive electrical, and radiation hardness test. Presented results include tests on three different prototypes of silicon micro-strip sensors.

In the Compressed Baryonic Matter (CBM) Experiment at FAIR, the Silicon Tracking System (STS) will perform track reconstruction and momentum determination of the charged particles created in interactions of heavy-ion beams with nuclear targets. The STS will consist of 8 tracking layers located at distances between 30 cm and 100 cm downstream of the target inside the 1 Tm magnetic dipole field. An ultra-low material budget is required to achieve momentum resolution of the order of Δp/p = 1%. Therefore, the front-end electronics is placed outside the physics aperture. The active volume of the STS is built from 300 μm thick double-sided silicon microstrip sensors mounted onto lightweight carbon fiber support ladders. The sensors will be read out through ultra-thin micro-cables with fast self-triggering electronics at the periphery of the stations where also other infrastructure such as cooling can be placed. In this paper, the development status of the detector system, highlighting the overview of the STS layout, tracking algorithm and performance simulations are presented.

We have previously shown that chronic ethanol treatment impairs the glycosylation of proteins in the rat liver. Changes in the microheterogeneity of transferrin, a N-sialoprotein under chronic alcohol consumption are well established. Apolipoprotein J, another N-glycoprotein, is a normal component of plasma high-density lipoproteins in the rat and human. Apo J is also highly sialylated and, thus, may be vulnerable to the deleterious actions of ethanol. Therefore, to understand the specific nature of alterations of Apo J sialylation as a consequence of chronic ethanol treatment, we have determined: (1) the sialylation index of Apo J (moles sialic acid per mole Apo J protein) in rats administered ethanol for 4, 6, and 8 weeks and a gradual withdrawal and a follow-up abstinence for 1, 2, and 4 weeks; and (2) enzymatic activities of hepatic sialyltransferase and plasma sialidase during the same periods of alcohol treatment and abstinence in rats. Although no significant differences in the Apo J sialylation index between rats of the control and ethanol groups were found at the 4th week of alcohol treatment, a highly significant loss of 24% (p < 0.001) and 44% (p < 0.001) was found after 6 and 8 weeks, respectively, of alcohol feeding of these animals. Furthermore, a significant recovery of 38% (p < 0.001), 78% (p < 0.001), 84% (p < 0.001) and 96% (p < 0.001) in the sialylation index of Apo J were found, respectively, during withdrawal and 1, 2, and 4 weeks of subsequent alcohol abstinence in these animals. These changes in the sialic acid content of Apo J were accompanied by a similar pattern of changes in the enzyme activities of hepatic sialyltransferase and plasma sialidase in animals undergoing chronic ethanol treatment, withdrawal, and abstinence periods. The analysis of the sialylation index of Apo J seems to be a simple and feasible method to use to evaluate the extent of ethanol exposure.

Gangliosides are sialo-glycosphingolipids that play important roles in the interaction of cells with their environment and are thus involved in the regulation of many cellular events. Sialic acid residues are important for the conformation of a glycomolecule, their structural stability and their functions. Although decreased brain ganglioside sialic acid has been previously reported as a result of chronic ethanol treatment in rats, no reports are available on the sialylation of specific gangliosides and/or the mechanism leading to depletion of their sialic add residues. Therefore, in this investigation, we have examined the effects of chronic ethanol treatment on (1) incorporation of [4,5- 3 H]N-acetylmannosamine (ManNAc) into specific rat brain gangliosides, GD 3 , GD 1a , GT 1a , and GT 1b ; and (2) enzymatic activities of brain sialyltransferase and sialidase at specific subcellular levels. The experiments were done in male Wistar rats pair-fed with either ethanol or control liquid diets for a period of 8 weeks. The rats were intracerebroventricularty injected with labeled ManNAc (30 μCi/rat) and killed after 90 min. Radioactivity was determined in respective ganglioside bands separated on a thin layer chromatography system. Specific activities of sialyltransferase and slalidase were assessed using GM 3 and GD 3 as substrates, respectively. The results showed significant decreases of 57.7% (p < 0.001) and 68.9% (p < 0.001), respectively, in the labeled ManNAc incorporation into GD 3 and GD 1a fractions in rats of the ethanol group, compared with rats of of the control group. No significant changes were noted in the incorporation of labeled ManNAc into GT 1a or GT 1b ganglioside fractions between the ethanol and control groups. Concomitantly, compared with control rats, a decrease of 18.9% (p < 0.05), 20.6% (p < 0.05), and 15.8% (p < 0.001) was found in the sialyltransferase activity, respectively, at the whole brain, and brain Golgi and synaptosomal levels. However, dramatic increases of 32.4% (p < 0.05), 105% (p < 0.001), and 150% (p < 0.001) in sialidase activity were found, respectively, at the whole brain and brain cytosol and synaptosomal fractions of rat treated chronically with ethanol. Thus, we conclude that the deleterious actions of ethanol on the sialylation of rat brain gangliosides is specific, and the reduced sialic acid label found in GD 3 and GD 1a in this study is mainly due to increased activity of brain sialidase. Furthermore, the study reaffirms our tenet that, regardless of whether it is the liver or the brain, glycosylation cascade is one of the main target of the deleterious attacks of ethanol.

We have previously shown in rats that chronic ethanol feeding significantly inhibits the incorporation of labeled leucine into Apo E secreted into the liver perfusate (p less than 0.01). Fish oil has been shown to counteract many of the adverse effects of ethanol. In order to explore whether this inhibitory effect of ethanol was due to the decreased synthesis and/or defective glycosylation of this glycoprotein, we have determined the effects of chronic ethanol and fish oil on the synthesis and glycosylation of Apo E in vivo. Four groups of male Wistar rats were pair-fed the following liquid diets for 8 weeks; (1) Ethanol Regular Fat, (2) Control Regular Fat, (3) Ethanol Fish Oil, and (4) Control Fish Oil. At the end, the rats were intraportally injected with a single dose of [U-14C]leucine (0.2 microCi/g body weight) and/or [2-3H]mannose (1 microCi/g body weight) and killed after 30 min. The incorporation of the labeled precursors into the immunoprecipitable Apo E was measured in the liver and its microsomal and Golgi fractions. The results showed marked decreases in mannose incorporation into total glycoproteins and specifically of Apo E in whole liver, microsomal, and Golgi fractions under ethanol treatment. In contrast, the leucine incorporation into liver Apo E increased 11% (p less than 0.048) by ethanol treatment. As a result, the [3H]mannose/[14C] leucine incorporation ratio also decreased 41% to 47% at the whole liver, microsomal, and Golgi fractions indicating a marked inhibition in glycosylation of Apo E in the ethanol group.(ABSTRACT TRUNCATED AT 250 WORDS)

Persistent excessive intake of ethanol can lead to liver cell injury and eventually to cirrhosis of the liver after prolonged abuse over many years. There are individual variations in susceptibility to alcohol toxicities. Hepatocellular necrosis results in a wide variety of clinical symptoms ranging from a relatively asymptomatic enlargement of the liver to massive fatty infiltration that ultimately leads to hepatic failure.1,2As the process of alcohol-mediated injury progresses, liver fibrosis and eventually cirrhosis ensue, resulting in total failure of the liver functions. The pathogenesis of alcohol-induced toxicity generally begins with specific episodes of hepatocellular injury accompanied by varying degrees of fatty liver.3As the alcohol abuse continues, this degenerative process manifests itself into liver cell dysfunction, chronic inflammation, and structural distortion leading to the proliferation of fibrous tissue, which ultimately culminates into the cirrhosis of the liver and death.

A soluble enzyme that catalyzes the oxidation of L-alpha-aminoadipate delta-semialdehyde to L-alpha-aminoadipic acid in the presence of NAD+ has been isolated and characterized from human liver. This enzyme L-alpha-aminoadipic delta-semialdehyde oxidoreductase has been found to be localized in the cytosol using subcellular fractionation and marker enzyme assays. The reaction product of this enzyme has been identified as L-alpha-aminoadipic acid by use of an amino acid analyzer and thin layer chromatography. The enzymatic reaction was irreversible and has a pH optimum of 8. The enzyme was stimulated by Mg2+, Cu2+ and Mn2+, and has a requirement of free -SH groups. The Km and Vmax values for its substrate L-alpha-aminoadipate delta-semialdehyde were shown to be 181 microM and 71.4 pmol.min-1.mg-1, respectively, and for its coenzyme NAD+ to be 454 microM and 142.9 pmol.min-1.mg-1, respectively. The characteristics of the oxidoreductase obtained from the human liver and Pseudomonas putida were compared.

We have previously shown that cultured human proximal tubular cells (PT) incubated with gentamicin contain numerous "myeloid bodies." This morphological change was accompanied by the storage of phosphatidylcholine and sphingomyelin. In order to delineate the biochemical mechanisms responsible for the accumulation of sphingomyelin in cells incubated with gentamicin, we pursued detailed studies on the activity of sphingomyelinase. Characterization studies on sphingomyelinase revealed that this enzyme has a bimodal pH optima in PT cells. Optimum activity was observed at pH 5.6 (designated as acid sphingomyelinase, A-SMase) and at pH 7.4 (designated as neutral sphingomyelinase, N-SMase). The activity of both the enzymes increased proportionately in control cells as a function of days of incubation. The activity of A-SMase was 16% lower in cells incubated with gentamicin as compared to control. The most striking observation was a gradual decline in the activity of N-SMase in cells incubated with gentamicin. Thus, following 21 days of incubation of cells with 0.3 mM gentamicin, the N-SMase was 2.7-fold lower than control cells. Mg2+ stimulated and Triton X-100 inhibited the activity of N-SMase. Whereas Mg2+ had no effects, Triton X-100 stimulated the activity of the A-SMase in PT cells. Moreover, A-SMase was relatively more heat-resistant than the N-SMase. The Km values for sphingomyelin using A-SMase in control cells and cells incubated with gentamicin were 0.07 X and 0.016 X 10(-7) M, respectively, whereas the Km values for sphingomyelin using N-SMase in control cells and cells incubated with gentamicin were 1.8 X and 1.5 X 10(-7) M, respectively. These findings suggest that gentamicin exerts a competitive inhibition of the A-SMase in PT cells. In contrast, gentamicin exerts a noncompetitive inhibition of the N-SMase in PT cells. Subcellular fractionation studies revealed that A-SMase was exclusively localized in the "lysosome-rich" fraction, whereas most, if not all, the N-SMase was localized in the microsomal fraction and "plasma-membrane"-rich fraction in cultured PT cells. Cells incubated with gentamicin for 21 days contained 25% lower activity of A-SMase associated with the lysosomal fraction as compared to control. In contrast, N-SMase activity in the microsomal and plasma membrane fraction was one-half as compared to control. We conclude that gentamicin-mediated decrease in sphingomyelinase activity may be responsible for the storage of sphingomyelin in cultured human PT cells.(ABSTRACT TRUNCATED AT 400 WORDS)