Characteristics of uroflowmetry patterns in men with detrusor underactivity revealed by artificial intelligence.

OBJECTIVES: To elucidate the characteristics of uroflowmetry (UFM) observed in men with detrusor underactivity (DU) using our developed artificial intelligence (AI) diagnostic algorithm to distinguish between DU and bladder outlet obstruction (BOO). METHODS: Subjective and objective parameters, including four UFM parameters (first peak flow rate, time to first peak, gradient to first peak, and the ratio of first peak flow rate to maximum flow rate [Qmax ]) selected by analyzing the judgment basis of the AI diagnostic system, were compared in 266 treatment-naive men with lower urinary tract symptoms (LUTS). Patients were divided into the DU (70; 26.32%) and non-DU (196; 73.68%) groups, and the UFM parameters for predicting the presence of DU were determined by multivariate analysis and receiver operating characteristic (ROC) curve analysis. Detrusor underactivity was defined as a bladder contractility index <100 and a BOO index <40. RESULTS: Most parameters on the first peak flow of UFM were significantly lower in the DU group. On multivariate analysis, lower first peak flow rate and lower ratio of first peak flow rate to Qmax were significant parameters to predict DU. In the ROC analysis, the ratio of the first peak flow rate to Qmax showed the highest area under the curve (0.848) and yielded sensitivities of 76% and specificities of 83% for DU diagnosis, with cutoff values of 0.8. CONCLUSIONS: Parameters on the first peak flow of UFM, especially the ratio of the first peak flow rate to Qmax , can diagnose DU with high accuracy in men with LUTS.

Development of an artificial intelligence diagnostic system for lower urinary tract dysfunction in men.

OBJECTIVES: To establish an artificial intelligence diagnostic system for lower urinary tract function in men with lower urinary tract symptoms using only uroflowmetry data and to evaluate its usefulness. METHODS: Uroflowmetry data of 256 treatment-naive men with detrusor underactivity, bladder outlet obstruction, or detrusor underactivity + bladder outlet obstruction were used for artificial intelligence learning and validation using neural networks. An optimal artificial intelligence diagnostic model was established using 10-fold stratified cross-validation and data augmentation. Correlations of bladder contractility index and bladder outlet obstruction index values for the artificial intelligence system and pressure flow study values were examined using Spearman's correlation coefficients. Additionally, diagnostic accuracy was compared between the established artificial intelligence system and trained urologists with uroflowmetry data of 25 additional patients by χ2 -tests. Detrusor underactivity was defined as bladder contractility index ≤100 and bladder outlet obstruction index ≤40, bladder outlet obstruction was defined as bladder contractility index >100 and bladder outlet obstruction index >40, and detrusor underactivity + bladder outlet obstruction was defined as bladder contractility index ≤100 and bladder outlet obstruction index >40. RESULTS: The artificial intelligence system's estimated bladder contractility index and bladder outlet obstruction index values showed significant positive correlations with pressure flow study values (bladder contractility index: r = 0.60, P < 0.001; bladder outlet obstruction index: r = 0.46, P < 0.001). The artificial intelligence system's detrusor underactivity diagnosis had a sensitivity and specificity of 79.7% and 88.7%, respectively, and those for bladder outlet obstruction diagnosis were 76.8% and 84.7%, respectively. The artificial intelligence system's average diagnostic accuracy was 84%, which was significantly higher than that of urologists (56%). CONCLUSIONS: Our artificial intelligence diagnostic system developed using the uroflowmetry waveform distinguished between detrusor underactivity and bladder outlet obstruction with high sensitivity and specificity in men with lower urinary tract symptoms.

Dietary aloe vera gel powder and extract inhibit azoxymethane- induced colorectal aberrant crypt foci in mice fed a high- fat diet.

Aloe vera gel exhibits protective effects against insulin resistance as well as lipid-lowering and anti-diabetic effects. The anti-diabetic compounds in this gel were identified as Aloe-sterols. Aloe vera gel extract (AVGE) containing Aloe-sterols has recently been produced using a new procedure. We previously reported that AVGE reduced large-sized intestinal polyps in Apc-deficient Min mice fed a high fat diet (HFD), suggesting that Aloe vera gel may protect against colorectal cancer. In the present study, we examined the effects of Aloe vera gel powder (AVGP) and AVGE on azoxymethane-induced colorectal preneoplastic aberrant crypt foci (ACF) in mice fed a HFD. Male C57BL/6J mice were given a normal diet (ND), HFD, HFD containing 0.5% carboxymethyl cellulose solution, which was used as a solvent for AVGE (HFDC), HFD containing 3% or 1% AVGP, and HFDC containing 0.0125% (H-) or 0.00375% (L-) AVGE. The number of ACF was significantly lower in mice given 3% AVGP and H-AVGE than in those given HFD or HFDC alone. Moreover, 3% AVGP, H-AVGE and L-AVGE significantly decreased the mean Ki-67 labeling index, assessed as a measure of cell proliferation in the colonic mucosa. In addition, hepatic phase II enzyme glutathione S-transferase mRNA levels were higher in the H-AVGE group than in the HFDC group. These results suggest that both AVGP and AVGE may have chemopreventive effects on colorectal carcinogenesis under the HFD condition. Furthermore, the concentration of Aloe-sterols was similar between 3% AVGP and H-AVGE, suggesting that Aloe-sterols were the main active ingredients in this experiment.

Reduction of intestinal polyp formation in min mice fed a high-fat diet with aloe vera gel extract.

Aloe vera gel supercritical CO2 extract (AVGE) has been shown to contain five phytosterols, reduce visceral fat accumulation, and influence the metabolism of glucose and lipids in animal model experiments. Recent epidemiologic studies have shown that obesity is an established risk factor for several cancers including colorectal cancer. Therefore, we examined the effects of AVGE on intestinal polyp formation in Apc-deficient Min mice fed a high-fat diet. Male Min mice were divided into normal diet (ND), high fat diet (HFD), low dose AVGE (HFD+LAVGE) and high dose AVGE (HFD+HAVGE) groups. The ND group received AIN-93G diet and the latter 3 groups were given modified high-fat AIN-93G diet (HFD) for 7 weeks. AVGE was suspended in 0.5% carboxymethyl cellulose (CMC) and administered orally to mice in HFD+LAVGE and HFD+HAVGE groups every day (except on Sunday) for 7 weeks at a dose of 3.75 and 12.5 mg/kg body weight, respectively. ND and HFD groups received 0.5% CMC alone. Between weeks 4 and 7, body weights in the HFD and HFD+LAVGE groups were reduced more than those in the ND group. However, body weights were not reduced in the HFD+HAVGE group. Mice were sacrificed at the end of the experiment and their intestines were scored for polyps. No significant differences were observed in either the incidence and multiplicity of intestinal polyps (≥0.5 mm in a diameter) among the three groups fed HFD. However, when intestinal polyps were categorized by their size into 0.5-1.4, 1.5-2.4, or ≥2.5 mm, the incidence and multiplicity of large polyps (≥2.5 mm) in the intestine in the HFD+HAVGE group were significantly lower than those in the HFD group. We measured plasma lipid (triglycerides and total cholesterol) and adipocytokine [interleukin-6 and high molecular weight (HMW) adiponectin] levels as possible indicators of mechanisms of inhibition. The results showed that HMW adiponectin levels in the HFD group were significantly lower than those in the ND group. However, the levels in the HFD+HAVGE group were significantly higher than those in the HFD group. These results indicate that HAVGE reduced large-sized intestinal polyps and ameliorated reduction in plasma HMW adiponectin levels in Min mice fed HFD.

Aloe vera gel extract attenuates ethanol-induced hepatic lipid accumulation by suppressing the expression of lipogenic genes in mice.

We have previously reported that Aloe vera gel had hypoglycemic activity and anti-obesity effects, although the effect on alcoholic fatty liver was unclear. We examined in this present study the effect of an Aloe vera gel extract (AVGE) on hepatic lipid metabolism by using an ethanol-induced transient fatty liver mouse model. Ethanol (3 g/kg of mouse weight) was orally administered to induce an accumulation of triglyceride (TG) and increase the mRNA expression of such lipogenic genes as sterol regulatory element-binding protein-1 (SREBP-1) and fatty acid synthase (FASN) in the liver. Although ethanol ingestion caused a 5.4-fold increase in liver TG, pre-treating with AVGE (1 mg/kg/d) for 1 week significantly suppressed this elevation of the ethanol-induced liver TG level. The expression of lipogenic genes was also lower in the AVGE pre-treatment group than in the control group. This inhibitory effect on the ethanol-induced accumulation of TG was attributed to a reduction in the expression of lipogenic genes that were increased by ethanol.

Administration of dried Aloe vera gel powder reduced body fat mass in diet-induced obesity (DIO) rats.

The aim of the present study was to investigate the anti-obesity effects of Aloe vera gel administration in male Sprague-Dawley (SD) rats with diet-induced obesity (DIO). SD rats at 7 wk of age were fed either a standard diet (10 kcal% fat) (StdD) or high-fat (60 kcal% fat) diet (HFD) during the experimental period. Four weeks after of HFD-feeding, DIO rats (11 wk of age) were orally administered with two doses of Aloe vera gel powder (20 and 200 mg/kg/d) for 90 d. Body weights (g) and body fat (%) of HFD fed rats were significantly higher than those of StdD-fed rats. Although a modest decrease of body weight (g) was observed with the administration of dried Aloe vera gel powder, both subcutaneous and visceral fat weight (g) and body fat (%) were reduced significantly in Aloe vera gel-treated rats. Serum lipid parameters elevated by HFD were also improved by the Aloe vera gel treatment. The oxygen consumption (VO(2)), an index of energy expenditure, was decreased in HFD-fed rats compared with that in StdD-fed rats. Administration of Aloe vera gel reversed the change in VO(2) in the HFD-fed rats. These results suggest that intake of Aloe vera gel reduced body fat accumulation, in part, by stimulation of energy expenditure. Aloe vera gel might be beneficial for the prevention and improvement of diet-induced obesity.

Oral ingestion of aloe vera phytosterols alters hepatic gene expression profiles and ameliorates obesity-associated metabolic disorders in zucker diabetic fatty rats.

We investigated the effects of the oral administration of lophenol (Lo) and cycloartanol (Cy), two kinds of antidiabetic phytosterol isolated from Aloe vera , on glucose and lipid metabolism in Zucker diabetic fatty (ZDF) rats. We demonstrated that the administrations of Lo and Cy suppressed random and fasting glucose levels and reduced visceral fat weights significantly. It was also observed that treatments with Lo and Cy decreased serum and hepatic lipid concentrations (triglyceride, nonesterified fatty acid, and total cholesterol). Additionally, Lo and Cy treatments resulted in a tendency for reduction in serum monocyte chemotactic protein-1 (MCP-1) level and an elevation in serum adiponectin level. Furthermore, the expression levels of hepatic genes encoding gluconeogenic enzymes (G6 Pase, PEPCK), lipogenic enzymes (ACC, FAS), and SREBP-1 were decreased significantly by the administrations of aloe sterols. In contrast, Lo and Cy administration increased mRNA levels of glycolysis enzyme (GK) in the liver. It was also observed that the hepatic ß-oxidation enzymes (ACO, CPT1) and PPARα expressions tended to increase in the livers of the Lo- and Cy-treated rats compared with those in ZDF-control rats. We therefore conclude that orally ingested aloe sterols altered the expressions of genes related to glucose and lipid metabolism, and ameliorated obesity-associated metabolic disorders in ZDF rats. These findings suggest that aloe sterols could be beneficial in preventing and improving metabolic disorders with obesity and diabetes in rats.

Safety evaluation of supercritical carbon dioxide extract of Aloe vera gel.

UNLABELLED: The gel of the Aloe vera plant has been used safely for oral and external applications. Previously, we found phytosterols derived from an extract of Aloe vera gel obtained with an organic solvent to have hypoglycemic and antiobesity effects. While developing of functional foods using Aloe vera gel, we produced an active Aloe vera gel extract (AVGE) using a supercritical carbon dioxide (CO2) extraction procedure. In this study, we tested the safety of AVGE in vitro and in vivo. In an acute oral toxicological test in which AVGE was administered to rats at a dose of 150 mg/kg body weight, there were no deaths or apparent abnormalities at necropsy. In a 90-d toxicity test in which rats were continuously administrered AVGE at 30 or 150 mg/kg, euthanized, and subjected to pathological examinations, no abnormalities attributable to the AVGE were found. AVGE was nonmutagenic in the Ames test and a chromosomal aberration test at concentrations of up to 5000 µg/plate and 1600 µg/plate, respectively, and in an in vivo bone marrow micronucleus test at up to 150 mg/kg/d. PRACTICAL APPLICATION: AVGE can be safely used as a functional food material.

Aloe vera phytosterols act as ligands for PPAR and improve the expression levels of PPAR target genes in the livers of mice with diet-induced obesity.

SUMMARY: Lophenol (Lo) and cycloartanol (Cy), minor phytosterols of Aloe vera gel, were previously identified as anti-diabetic compounds, and these compounds also reduced body fat in a type 2 diabetic model animal. In this study, we investigated the effects of Lo and Cy on peroxisome proliferator activated receptors (PPAR) using a luciferase reporter assay. DNA microarray and real-time quantitative RT-PCR (qPCR) analyses were also performed in a diet-induced obesity (DIO) mouse model. The Aloe phytosterols activated PPAR in a dose-dependent manner. The expression levels of many PPAR target genes were changed in the Aloe phytosterol group compared with those in the control high-fat diet (HFD) group. In particular, the expression levels of Fatp1, Acox1, Cpt1, and Hmgcs2 were significantly increased in the Aloe phytosterol group compared with those in the control HFD group; however, the expression level of ApoCIII was significantly decreased in the Aloe phytosterol group. We confirmed that Aloe phytosterols activate PPAR transcription in vitro. In addition, quantitative gene expression analysis in DIO mice suggested that Aloe phytosterols improve fatty acid metabolism in the liver.:

Administration of phytosterols isolated from Aloe vera gel reduce visceral fat mass and improve hyperglycemia in Zucker diabetic fatty (ZDF) rats.

SUMMARY: We examined the effects of lophenol (Lo) and cycloartanol (Cy), minor phytosterols of Aloe vera gel, in obese animal model of type II diabetes, Zucker diabetic fatty (ZDF) rats. Male ZDF rats were administered Lo and Cy at 25 µg/(kg day) daily for 44 days. Consecutive treatment of phytosterols suppressed the hyperglycemia, and random blood glucose levels after 35 days of treatment were 39.6 and 37.2% lower than the control, in Lo and Cy treatment groups, respectively. Consistent with the random blood glucose level, hemoglobin A1c (HbA1c) values of phytosterols treated rats were also lower than the control (Lo: 5.5 ± 0.8, Cy: 4.6 ± 0.7 vs. control: 7.2 ± 1.5). In the oral glucose tolerance test (OGTT) after 28 days of administration, the glucose intolerance was improved in phytosterols treatment groups. Additionally, the continuous administration of Lo and Cy also reduced the serum free fatty acid (FFA) and triglyceride (TG) levels except total cholesterol (T-Cho). Furthermore, the weights of total abdominal fat tissues were significantly lower than the control in ZDF rats with Lo (27.7%) and Cy (26.3%) treatment. These observations suggest that Aloe vera-derived phytosterols could reduce visceral fat accumulation, and would be useful for the improvement of hyperlipidemia and hyperglycemia.:

Isolation and structure of four new ceramides from the starfish Luidia maculata.

A new sphingosine-type ceramide LMCer-1-1 (1) and three new phytosphingosine-type ceramides, LMCer-2-1 (2), LMCer-2-6 (3), and LMCer-2-7 (4), were isolated from the anti-hyperglycemic active ceramide molecular species LMCer-1 and LMCer-2, obtained from the less polar fraction of the chloroform-methanol extract of the whole bodies of Luidia maculata. The structures of these ceramides were determined on the basis of chemical and spectroscopic evidence as: (2S,3R,4E,2'R)-2-(2-hydroxyhexadecanoylamino)-16-methyl-4-octadecene-1,3-diol (1), (2S,3S,4R,2'R)-2-(2-hydroxyhexadecanoylamino)-16-methyl-octadecane-1,3,4-triol (2), (2S,3S,4R,2'R)-2-(2-hydroxydocosanoylamino)-hexadecane-1,3,4-triol (3), and (2S,3S,4R,2'R)-2-(2-hydroxydocosanoylamino)-14-methyl-hexadecane-1,3,4-triol (4).

Identification of five phytosterols from Aloe vera gel as anti-diabetic compounds.

The genus Aloe in the family Liliaceae is a group of plants including Aloe vera (Aloe barbadensis MILLER) and Aloe arborescens (Aloe arborescens MILLER var. natalensis BERGER) that are empirically known to have various medical efficacies. In the present study, we evaluated the anti-hyperglycemic effect of Aloe vera gel and isolated a number of compounds from the gel. On the basis of spectroscopic data, these compounds were identified as lophenol, 24-methyl-lophenol, 24-ethyl-lophenol, cycloartanol, and 24-methylene-cycloartanol. These five phytosterols were evaluated for their anti-hyperglycemic effects in type 2 diabetic BKS.Cg-m(+/+)Lepr(db/J) (db/db) mice. In comparison with the hemoglobin A1c (HbA1c) levels of vehicle-treated mice, statistically significant decreases of 15 to 18% in HbA1c levels were observed in mice treated with 1 mug of the five phytosterols. Considering the ability to reduce blood glucose in vivo, there were no differences between the five phytosterols. Administration of beta-sitosterol did not reduce the blood glucose levels in db/db mice. After administration of the five phytosterols for 28 d, fasting blood glucose levels decreased to approximately 64%, 28%, 47%, 51%, and 55% of control levels, respectively. Severe diabetic mice treated with phytosterols derived from Aloe vera gel did not suffer weight reduction due to glucose loss in the urine. These findings suggest that Aloe vera gel and phytosterols derived from Aloe vera gel have a long-term blood glucose level control effect and would be useful for the treatment of type 2 diabetes mellitus.

Administration of macrophage colony-stimulating factor mobilized both CD11b+CD11c+ cells and NK1.1+ cells into peripheral blood.

We attempted the phenotypic characterization of peripheral blood (PB) cells after daily administration of macrophage colony-stimulating factor (M-CSF) in mice. The number of CD11b+ cells was increased by M-CSF treatment (2- and 5-day injections). Notably, CD11bbrightCD11cdim, CD11b+CD11c+ and CD11b+CD80+ cells were significantly increased by 2-day treatment of M-CSF. On the other hand, the number of NK1.1+ cells was not changed by the 2-day treatment, but it was significantly increased by the 5-day treatment. However, the numbers of CD3+ and NK1.1+CD3+ cells were not changed by M-CSF treatment. Then, mononuclear cells (MNCs) were separated from the PB of mice treated with saline or M-CSF, and they were incubated with GM-CSF + IL-4 or IL-2. Compared with the saline-treated one (S-MNCs), the MNCs of M-CSF-treated mice (M-MNCs) showed strong proliferation by the GM-CSF + IL-4 stimulation. The MNCs could stimulate proliferation of allo-T cells in the mixed lymphocyte reaction (MLR), especially the M-MNCs showed strong reaction. On the other hand, the stimulation by IL-2 induced strong cell growth of MNCs. And M-CSF treatment enhanced this response. Furthermore, the M-MNCs (stimulated by IL-2 in vitro) exhibited greater cytotoxicity against Yac-1 cells than the S-MNCs. In conclusion, we found that administration of M-CSF mobilized CD11b+, CD11b+CD11c+, CD11b+CD80+, and NK1.1+cells into PB. And the injection of M-CSF facilitates the generation of dendritic and natural killer cells from PB cells in vitro. These results suggest that the mobilized cells may provide for application of immunotherapy.

Cloning and characterization of a novel RING-B-box-coiled-coil protein with apoptotic function.

We have identified a novel RING-B-box-coiled-coil (RBCC) protein (MAIR for macrophage-derived apoptosis-inducing RBCC protein) that consists of an N-terminal RING finger, followed by a B-box zinc finger, a coiled-coil domain, and a B30.2 domain. MAIR mRNA was expressed widely in mouse tissues and was induced by macrophage colony-stimulating factor in murine peritoneal and bone marrow macrophages. MAIR protein initially showed a granular distribution predominantly in the cytoplasm. The addition of zinc to transfectants containing MAIR cDNA as part of a heavy metal-inducible vector caused apoptosis of the cells characterized by cell fragmentation; a reduction in mitochondrial membrane potential; activation of caspase-7, -8, and -9, but not caspase-3; and DNA degradation. We also found that the RING finger and coiled-coil domains were required for MAIR activity by analysis with deletion mutants.

Booster effect of interleukin-2 on natural killer 1.1+ cells stimulated by administration of macrophage colony-stimulating factor in mice.

The authors studied the combined effects of macrophage colony-stimulating factor (M-CSF) and interleukin (IL)-2 on the functions and antitumor activity of natural killer (NK) 1.1+ cells in vitro and in vivo. NK1.1+ cells were isolated from the spleen of mice treated with saline or M-CSF, and their functions (proliferation, production of IFN-gamma, and cytotoxicity) evaluated in vitro. Although the proliferation of and production by NK1.1+ cells was stimulated by the addition of IL-2, the cells from the M-CSF-treated mice responded better. Furthermore, the cytotoxicity against Yac-1 cells and B16 melanoma cells was stimulated by M-CSF administration and enhanced by the addition of IL-2 and IL-12. These results demonstrated that M-CSF treatment augmented the functions of NK1.1 cells, and IL-2 and IL-12 boosted these activities in vitro. The authors then examined the effects of co-administration of M-CSF and IL-2 in vivo. The clearance of B16 cells in lung was augmented by the administration of M-CSF but not IL-2. However, M-CSF + IL-2 treatment further enhanced the clearance activity. The anti-metastatic activity was also enhanced by the M-CSF + IL-2 treatment. Furthermore, the survival of B16-bearing mice was prolonged by M-CSF + IL-2. These results suggested that administration of IL-2 boosts the functions of NK1.1+ cells, which are augmented preliminarily by the administration of M-CSF.

Molecular cloning of a novel immunoglobulin superfamily gene preferentially expressed by brain and testis.

We have cloned and characterized a novel gene from both human and mouse that encodes a new member of the immunoglobulin superfamily. The gene is preferentially expressed in both brain and testis, and hence, termed BT-IgSF (brain- and testis-specific immunoglobulin superfamily). The predicted protein consists of V-type and C2-type immunoglobulin domains as well as a hydrophobic signal sequence, a single transmembrane region, and a cytoplasmic domain. Human BT-IgSF protein (431 amino acids) is 88% identical to the mouse protein (428 amino acids) and both show significant homology to coxsackie and adenovirus receptor (CAR) and endothelial cell-selective adhesion molecule (ESAM). We examined the expression of BT-IgSF with various cultured cells and found that the gene was expressed in both neurons and glial cells in vitro. Furthermore, the expression was preferentially detected in pyramidal cell layers of the dentate gyrus and hippocampus and in commissure fibers of the corpus callosum, in brain tissue sections examined. These findings suggest that BT-IgSF plays a role in the development or function of the central nervous system.

Effect of coadministration of M-CSF and IFN-alpha on NK1.1+ cells in mice.

The purpose of this study was to evaluate the effect of coadministration of macrophage colony-stimulating factor (M-CSF) and interferon-alpha (IFN-alpha) on NK1.1(+) cells in mice. Administration of M-CSF, but not IFN-alpha, increased the number of NK1.1(+) cells and CD11b(+) cells in spleen and blood. Coadministration of the two agents induced a greater increase in NK1.1(+) cells than did administration of M-CSF alone. Administration of M-CSF or IFN-alpha augmented the clearance activity of Yac-1 cells in lung, and coadministration of these agents further augmented this effect. The combination of M-CSF and IFN-alpha effectively reduced the formation of tumor nodules in lung and liver in an experimental metastasis model using B16 melanoma. The combination of M-CSF and IFN-alpha induced the increase and activation of NK1.1(+) cells more than either agent alone. These effects may contribute to the antimetastatic reaction by NK1.1(+) cells in vivo.

Leukemic cell-surface CD13/aminopeptidase N and resistance to apoptosis mediated by endothelial cells.

BACKGROUND: Attachment of leukemic cells to vascular endothelial cells induces the vascular endothelial cells to release endothelial cell-derived interleukin 8 (endothelial IL-8), which then induces leukemic cells to undergo apoptosis. NB4, a human promyelocytic leukemic cell line that expresses high levels of cell-surface CD13/aminopeptidase N, does not undergo endothelial IL-8-induced apoptosis. Consequently, we investigated the relationship between cell-surface aminopeptidase activity and endothelial IL-8 induction of apoptosis in various leukemic cell lines. METHODS: CD13/aminopeptidase N activity and IL-8-induced apoptosis were examined in leukemic cell lines. Endothelial IL-8-induced apoptosis was examined further in NB4 cells, K562 cells (human chronic myelogenous leukemic cells expressing low levels of CD13/aminopeptidase N), CD13/aminopeptidase N-transfected K562 (K562/CD13) cells that overexpress aminopeptidase, and mock-transfected K562 cells (vector only). These cells were also cocultured with a vascular endothelial cell layer to investigate the association between aminopeptidase activity and apoptosis in this system. All statistical tests were two-sided. RESULTS: Endothelial IL-8 induced apoptosis in K562 cells but not in K562/CD13 cells. A combination of an aminopeptidase inhibitor (such as bestatin) and endothelial IL-8 induced apoptosis in NB4 cells and K562/CD13 cells (2.88-fold difference [95% confidence interval [CI] = 1.82-fold to 3.94-fold], P =.004 for bestatin-treated NB4 cells and 4.31-fold difference [95% CI = 3.52-fold to 5.10-fold], P<.001 for bestatin-treated K562/CD13 cells). When aminopeptidase activity in NB4 cells was modulated by aminopeptidase inhibitors, a statistically significant correlation was found between aminopeptidase activity and the proportion of apoptotic cells induced by endothelial IL-8 (r = -.837, P<.001 by Pearson's correlation coefficient; r = -.697, P =.013 by Spearman's correlation analysis by ranks). K562/CD13 cells cocultured with vascular endothelial cells did not undergo apoptosis, but the addition of bestatin resulted in the induction of apoptosis in K562/CD13 cells (2.70-fold difference [95% CI = 1.77-fold to 3.63-fold], P<.001). Bestatin treatment increased the level of IL-8 mRNA in and the amount of IL-8 secreted by vascular endothelial cells. CONCLUSIONS: High levels of cell-surface CD13/aminopeptidase N appear to allow leukemic cells to resist endothelial IL-8-induced apoptosis. The combination of endothelial IL-8 and bestatin induce leukemic cells expressing high levels of CD13/aminopeptidase N to undergo apoptosis. Bestatin may be useful for treating patients with leukemia.

New human myelodysplastic cell line, TER-3: G-CSF specific downregulation of Ca2+/calmodulin-dependent protein kinase IV.

We have established a new hematopoietic cell line from a patient with myelodysplastic syndrome (MDS), which was refractory anemia with excess blasts (RAEB). This cell line, designated TER-3, depends on several cytokines for long-term survival and growth, and requires interleukin-3 (IL-3) for continuous growth. Cytochemical analysis revealed that TER-3 cells are weakly dianisidine positive and nonspecific esterase positive, but peroxidase negative. The surface marker profile shows that the TER-3 cells are strongly positive for myeloid, lymphoid, and megakaryocytic antigens such as CD15, CD19, and CD61, and negative for some common multilineage antigens such as CD13, CD33, and CD34. Thus, this cell line has a multilineage phenotype, suggesting that the transformation event occurred in multipotent stem cells. Dianisidine- and nonspecific esterase-positive TER-3 cells increase with granulocyte-colony stimulating factor (G-CSF) rather than with IL-3. These results suggest that the cell line is useful for understanding the mechanism underlying G-CSF-associated hematopoietic cell differentiation and activation in the patient with MDS.

Comparative study and effects of macrophage colony-stimulating factor (M-CSF) administration on NK1.1+ cells in mouse spleen and bone marrow.

We conducted a comparative study of NK1.1+ cells in spleen and bone marrow and the effects of administration of M-CSF on them. Administration of M-CSF to mice increased the number of NK1.1+ cells in spleen but not in bone marrow. The NK1.1+ cells in spleen (Spl-NK1.1) and bone marrow (BM-NK1.1) were purified by magnetic cell sorter. Their cell surface markers and functions were then examined. The percentage of Mac-1 antigen-positive cells (and F4/80 antigen-positive cells) was higher among BM-NK1.1 than Spl-NK1.1. Moreover, the administration of M-CSF increased the number of Mac-1 and F4/80 antigen-positive cells in both Spl-Nk1.1 and BM-NK1.1. The functions (cytolytic activity and IFN-gamma production) of Spl-NK1.1 and BM-NK1.1 were the same and were enhanced by the administration of M-CSF. But Spl-NK1.1 produced more IFN-gamma than BM-NK1.1 when M-CSF was administered. BM-NK1.1 showed a greater proliferative response to IL-2 than Spl-NK1.1. Administration of M-CSF augmented this response. BM-NK1.1 proliferated in response to IL-4 and IL-15, but Spl-NK1.1 responded only slightly. However, administration of M-CSF stimulated Spl-NK1.1 to respond to these cytokines. Both Spl-NK1.1 and BM-NK1.1 showed only a weak response to M-CSF in vitro. But the expression of c-fms antigen (M-CSFR) increased after the M-CSF injections in vivo. These results suggested that there are phenotypical and functional differences between Spl-NK1.1 and BM-NK1.1. The administration of M-CSF led to an accumulation of NK1.1+ cells which were mobilized from bone marrow in spleen.