Biomedical subjects

I Fabian

Publications and source records attributed to I Fabian.

At least 19 recordsLinked to original sources

PubMed2006

Moxifloxacin enhances antiproliferative and apoptotic effects of etoposide but inhibits its proinflammatory effects in THP-1 and Jurkat cells.

Etoposide (VP-16) is a topoisomerase II (topo II) inhibitor chemotherapeutic agent. Studies indicate that VP-16 enhances proinflammatory cytokines secretion from tumour cells, including IL-8, a chemokine associated with proangiogenic effects. Fluoroquinolones inhibit topo II activity in eukaryotic cells by a mechanism different from that of VP-16. The fluoroquinolone moxifloxacin (MXF) has pronounced anti-inflammatory effects in vitro and in vivo. We studied the effects of MXF and VP-16 on purified human topo II activity and further analysed their combined activity on proliferation, apoptosis and caspase-3 activity in THP-1 and Jurkat cells. Moxifloxacin alone slightly inhibited the activity of human topo II; however, in combination with VP-16 it led to a 73% reduction in enzyme activity. VP-16 inhibited cell proliferation in a time and dose-dependent manner. The addition of moxifloxacin for 72 h to low-dose VP-16 doubled its cytotoxic effect in THP-1 and Jurkat cells (1.8- and 2.6-fold decrease in cell proliferation, respectively) (P<0.004). Moxifloxacin given alone did not induce apoptosis but enhanced VP-16-induced apoptosis in THP-1 and Jurkat cells (1.8- and two-fold increase in annexin V positive cells and caspase-3 activity, respectively) (P<0.04). VP-16 induced the release of IL-8 in a time and dose-dependent manner from THP-1 cells. Moxifloxacin completely blocked the enhanced release of IL-8 induced by 0.5 and 1 microg ml(-1) VP-16, and decreased IL-8 release from cells incubated for 72 h with 3 microg ml(-1) VP-16 (P<0.001). VP-16 enhanced the release of IL-1beta and TNF-alpha from THP-1 cells, whereas the addition of MXF prevented the enhanced cytokine secretion (P<0.001). We conclude that MXF significantly enhances VP-16 cytotoxicity in tumour-derived cells while preventing VP-16-induced proinflammatory cytokine release. This unique combination may have clinical benefits and cytotoxic drug 'sparing effect' and should be further studied in vivo.

Anti-Infective Agents↗

PubMed2001

Immunomodulatory effects of moxifloxacin in comparison to ciprofloxacin and G-CSF in a murine model of cyclophosphamide-induced leukopenia.

We analyzed the effect of the two quinolones moxifloxacin and ciprofloxacin on the repopulation of hematopoietic organs and on the production of cytokines by various organs of cyclophosphamide (CP)-induced leukopenic mice. The effect was compared to that of G-CSF. Cyclophosphamide injection induced a severe leukopenia, with nadir at day 4 post-injection. All the quinolone and G-CSF-treated animals showed WBC>500/microL at the nadir, compared to 50% of saline-treated mice. Cyclophosphamide induced a marked decrease in the number of myeloid progenitors (CFU-C) in bone marrow (BM) and spleen. Quinolone or G-CSF treatment resulted in a 1.4-4.3-fold increase in CFU-C numbers in the BM; no enhancement was observed in the spleen. Treatment with CP resulted in enhanced colony-stimulating activity (CSA) in bone shaft and spleen and decreased activity in bladder and lung. Treatment of CP-injected mice with quinolones significantly enhanced CSA in the bone shaft, spleen, lung and bladder on different days. In normal mice the highest levels of GM-CSF and IL-6 were observed in lung-conditioned medium (compared to bone shaft, spleen and bladder). Injection of CP resulted in a 22.5- and 93-fold decrease in GM-CSF and IL-6 levels, respectively, in lung-conditioned medium, while treatment with quinolones resulted in 2-4-fold increase in GM-CSF with no effect on IL-6 production. G-CSF treatment had no enhancing effect on GM-CSF nor on IL-6 production. We conclude that moxifloxacin and ciprofloxacin administered to CP-injected mice revert some of the immune suppressive effects of cyclophosphamide.

Animals↗

PubMed2001

Effect of simvastatin alone and in combination with cytosine arabinoside on the proliferation of myeloid leukemia cell lines.

BACKGROUND: Cholesterol biosynthesis is regulated by the activity of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. Cholesterol and its derivatives are required in high concentrations by neoplastic proliferating cells for both DNA synthesis and cell growth. Thus, inhibition of HMG-CoA reductase could effect cell cycle progression and proliferation. Therefore, we examined the effect of an HMG-CoA reductase inhibitor (simvastatin) alone and in combination with cytosine arabinoside (ARA-C) on the proliferation of two AML cell lines. METHODS: AML blasts derived from two cell lines (HL-60 and AML-2) were incubated with increasing concentrations of either simvastatin alone or simvastatin alone for 24 hours with ARA-C added thereafter. The effect of the drugs on cell proliferation in liquid culture (3H thymidine uptake) and on clonogenic assay was analyzed. RESULTS: We found that the number of proliferating AML blasts (suspension cultures) and colony formations (agar cultures) of both cell lines declined significantly after incubation with simvastatin. Preincubation of both cell lines with simvastatin by the addition of increasing concentrations of ARA-C produced a degree of growth inhibition that was significantly greater than that of the individual compounds. This antigrowth interaction was additive rather than synergistic. CONCLUSIONS: We conclude that simvastatin has a major antiproliferative effect on AML blasts in vitro. Also, the combination of simvastatin and ARA-C significantly enhanced the antiproliferative effect of each drug. These findings may open new avenues in both the laboratory and clinical research of the treatment of leukemia.

Anticholesteremic Agents↗

PubMed2000

Ultrasound imaging-guided noninvasive ultrasound thrombolysis: preclinical results.

BACKGROUND: Catheter-based therapeutic ultrasound thrombolysis was recently shown to be effective and safe. The purpose of this work was to study the safety and efficacy of external high-intensity focused ultrasound thrombolysis guided by ultrasound imaging in experimental settings. METHODS AND RESULTS: A therapeutic transducer was constructed from an acoustic lens and integrated with an ultrasound imaging transducer. In vitro clots were inserted into bovine arterial segments and sonicated under real-time ultrasound imaging guidance in a water tank. With pulsed-wave (PW) ultrasound, the total sonication time correlated with thrombolysis efficiency (r(2)=0.7666). A thrombolysis efficiency of 91% was achieved with optimal PW parameters (1:25 duty cycle, 200-micros pulse length) at an intensity (I(spta)) of >35+/-5 W/cm(2). Ultrasound imaging during sonication showed the cavitation field as a spherical cloud of echo-dense material. Within <2 minutes, the vessel lumen evidenced neither residual clot nor damage to the arterial wall. On serial filtration, 93+/-1% of the lysed clot became subcapillary in size (<8 microm). In vitro safety studies, however, showed arterial damage when an I(spta) of 45 W/cm(2) was used for periods of >/=300 seconds. CONCLUSIONS: External high-intensity focused ultrasound thrombolysis using optimal PW parameters for periods of </=300 seconds appears to be a safe and effective method to induce thrombolysis. The procedure can be guided by ultrasound imaging, thereby allowing the monitoring of therapy.

Animals↗

PubMed2000

Activation of c-K-ras mutations in human gastrointestinal tumors.

BACKGROUND & AIMS: Ras genes are the most frequently detected oncogenes in human malignancies. Data regarding the frequency of c-K-ras mutations in esophageal, gastric, and small bowel tumors are limited and controversial. METHODS: DNA was extracted from 262 formalin-fixed, paraffin-embedded sections of gastrointestinal samples and tumors, including Barrett's esophagus, esophageal squamous cell carcinomas and adenocarcinomas, and small and large bowel adenomas and adenocarcinomas. The presence of c-K-ras codon 12 mutations was determined using a nonradioactive polymerase chain reaction-based restriction fragment length polymorphism assay. RESULTS: c-K-ras mutations were detected in 1 of 39 (2%) patients with Barrett's esophagus, 1 of 21 (5%) adenocarcinomas, 0 of 27 squamous cell carcinomas of the esophagus, and 1 of 32 (3%) gastric adenocarcinomas. It was also present in 8 of 20 (40%) and 10 of 28 (36%) small bowel adenomas and adenocarcinomas, respectively. Similar numbers were observed in 10 of 25 (40%) large bowel adenomas and 11 of 30 adenocarcinomas (37%). Mutations were not associated with age, gender, histology, grade, stage, location, or mortality. CONCLUSIONS: The frequency of codon 12 c-K-ras mutations in small and large bowel tumors is approximately 10-fold higher than that of tumors in the upper gastrointestinal tract.

Adenocarcinoma↗

PubMed1999

Alteration of actin organization by jaspamide inhibits ruffling, but not phagocytosis or oxidative burst, in HL-60 cells and human monocytes.

Jaspamide, a naturally occurring cyclic peptide isolated from the marine sponge Hemiastrella minor, has fungicidal and growth-inhibiting activities. Exposure of promyelocytic HL-60 cells and human monocytes to jaspamide induces a dramatic reorganization of actin from a typical fibrous network to focal aggregates. HL-60 cells exposed to 5 x 10(-8) mol/L or 10(-7) mol/L jaspamide exhibited a reduced proliferation rate. In addition, 10(-7) mol/L jaspamide induced maturation of HL-60 cells as indicated by the appearance of a lobulated nucleus in 55% +/- 5% of the cells and immunophenotypic maturation of the leukemia cells (upregulation of CD16 and CD14 B antigens). Further characterization has shown that F-actin is aggregated both in HL-60 cells and in human monocytes exposed to 10(-7) mol/L jaspamide. Well-spread cultured human monocytes contracted and adopted round shapes after treatment with jaspamide. Moreover, a dose-dependent increase in both total actin and de novo synthesized portions of the soluble actin was observed in jaspamide-treated HL-60 cells. Jaspamide treatment inhibits ruffling and intracellular movement in HL-60 cells and monocytes, but does not affect phagocytic activity or respiratory burst activity. The consequential effects of jaspamide-induced actin reorganization on ruffling, versus its negligible effect on phagocytosis and oxidative burst, may shed light on molecular mechanisms of actin involvement in these processes. Jaspamide disrupts the actin cytoskeleton of normal and malignant mammalian cells with no significant effect on phagocytic activity and may, therefore, be considered as a novel therapeutic agent.

Actins↗

PubMed1999

Inhibition by ascorbic acid of apoptosis induced by oxidative stress in HL-60 myeloid leukemia cells.

The human myeloid leukemia cell line HL-60 transports the oxidized form of ascorbic acid, dehydroascorbic acid (DHA), and accumulates reduced ascorbic acid. We studied the effect of ascorbic acid loading on apoptosis induced by serum- and glucose-free culture and by oxidative stress induced by H2O2. Uptake accumulation studies indicated that incubation of HL-60 cells with DHA resulted in the accumulation of intracellular ascorbic acid which decreased with time when cells were incubated in DHA-free medium. Exposure of HL-60 cells to increasing concentrations of H2O2 resulted in dose-dependent intracellular accumulation of peroxides, as determined by the use of the oxidation-sensitive fluorescent probe 2',7'-dichlorofluorescin-diacetate (DCFH-DA), which was accompanied by a decrease in intracellular ascorbic acid and an increase in apoptosis. A dramatic decrease in intracellular ascorbic acid was noted when preloaded HL-60 cells were exposed to 150 microM H2O2 (the concentration dropped from 5.2 +/- 0.6 mM to 3.6 +/- 0.1 mM in cells preincubated with 150 microM DHA). A dose-dependent protective effect of DHA was observed. Ascorbic acid loading also provided strong protection from apoptosis associated with serum- and glucose-free culture. Flow cytometry studies showed that exposure of HL-60 cells to 150 microM H2O2 resulted in decreased Bcl-2 expression that was associated with enhanced apoptosis (up to 33.6 +/- 2.6%). No significant variation of Bcl-2 expression was measured following exposure of HL-60 cells, loaded with ascorbic acid, to 150 microM H2O2 and only a slight increase (up to 10.1 +/- 3.1%) in apoptosis. These findings indicate that ascorbic acid can inhibit apoptosis induced by oxidative stress in HL-60 cells.

Antioxidants↗

PubMed1999

Overexpression of cyclin D1 occurs in both squamous carcinomas and adenocarcinomas of the esophagus and in adenocarcinomas of the stomach.

Increased expression of the cyclin D1 gene frequently occurs in human squamous carcinomas of the esophagus. However, the expression of cyclin D1 has not been previously examined in detail in adenocarcinomas of the esophagus or stomach. Therefore, we examined, in parallel, the expression of cyclin D1 in both squamous and adenocarcinomas of the esophagus and in adenocarcinomas of the stomach. The level of expression of the cyclin D1 protein was assessed by immunohistochemistry in 39 esophageal and 34 gastric carcinomas and correlated with clinical and pathology parameters. Within the esophagus, 71% of the squamous carcinomas and 64% of the adenocarcinomas were positive for increased cyclin D1 nuclear staining. For adenocarcinomas of the stomach, the overall positive rate was 47%; in the gastric cardia, the rate was 44%, and in other regions of the stomach, it was 50%. In esophageal and gastric adenocarcinomas of the intestinal type, increased expression of cyclin D1 was seen in 70% of the samples, whereas with the diffuse type only 13% were positive (P < .01). Tumors from patients older than the median age of 67 years were more frequently positive than tumors from patients younger than 67 years (74% v 42%, respectively) (P < .01). Positive staining was also seen more frequently in well and moderately differentiated tumors than in poorly differentiated tumors (74% v 49%, respectively) (P < .05). Cytoplasmic staining for cyclin D1 was noted in 22% of the tumors, of various types. Therefore, increased expression of cyclin D1 frequently occurs in both adenocarcinomas and squamous carcinomas of the esophagus, and in adenocarcinomas of the stomach. The increased expression in adenocarcinomas is especially frequent in the intestinal-type lesions.

Adenocarcinoma↗

PubMed1999

Ascorbic acid inhibits apoptosis induced by X irradiation in HL60 myeloid leukemia cells.

Exposure of cells to ionizing radiation can cause apoptosis. Since antioxidants have been shown to protect against radiation-induced apoptosis, in this study we have evaluated the putative protective effect of ascorbate against radiation-induced apoptosis as well as the production of peroxides in the cells. HL60 cells transport the oxidized form of ascorbic acid, dehydroascorbic acid (DHA), and accumulate reduced ascorbate. Exposure of the cells to 5-40 Gy X radiation resulted in induction of apoptosis. Preincubation of the cells with DHA reduced the level of apoptosis after exposure to 5-20 Gy. Exposure of the cells to 5 or 20 Gy X radiation did not affect the intracellular concentration of peroxides, while phorbol myristate acetate (PMA), which is known to induce production of H(2)O(2) in cells (and served as a control), resulted in an increase in peroxides and a decrease in intracellular ascorbate. Irradiation of the cells with 1-3 Gy resulted in up-regulation of expression of BCL2 without affecting the level of apoptosis. At higher doses of radiation, enhanced BCL2 expression did not prevent radiation-induced apoptosis. Loading of the cells with ascorbate prior to their exposure to 1-3 Gy X radiation did not affect the enhanced BCL2 expression observed in the irradiated cells. At higher doses of radiation, ascorbate decreased apoptosis and restored the level of BCL2 in the cells. Exposure of the cells to 3-20 Gy X radiation enhanced the cell surface expression of TNFRSF6 (formerly known as Fas/APO-1) antigen and enhanced anti-TNFRSF6 antibody-induced apoptosis of the cells. Ascorbate loading did not affect expression of TNFRSF6 and did not overcome the anti-TNFRSF6 antibody-induced apoptosis. In conclusion, our data demonstrate that exposure of HL60 cells to radiation enhanced BCL2 and TNFRSF6 expression. Ascorbate did not affect BCL2 or TNFRSF6 expression. We therefore conclude that it protects HL60 cells against radiation-induced apoptosis, although the mechanisms of protection must still be elucidated.

Apoptosis↗

PubMed1999

Abnormalities in the expression of cell cycle-related proteins in tumors of the small bowel.

Tumors of the small bowel are quite rare for unknown reasons, although they resemble colorectal tumors in many respects. The purpose of this study was to determine whether abnormalities in the expression of several cell cycle control genes are of importance in small bowel tumorigenesis by comparing a series of samples of normal mucosa, adenomatous polyps, and adenocarcinomas. The levels of cyclin D1, cyclin E, p16, p21, p27, and p53 proteins were determined by immunohistochemistry in samples of normal small bowel (n = 16), small bowel adenomas (n = 20), and small bowel adenocarcinomas (n = 24). Normal small bowel mucosa expressed p27 protein, but not the other cell cycle-related proteins. About 20% of the tumors displayed a decrease in the expression of this protein. The most frequent alteration in the tumors was an increase in the p16 protein. Increased expression of p53 was associated with tumor progression because it was overexpressed in 45% of the adenomas and 65% of the adenocarcinomas (P<0.05). Advanced age and increased detection of cyclin D1 and p53 were associated with a decreased 3-year survival (P<0.05). Cell cycle abnormalities are early and important events in the multistep process of small bowel tumorigenesis, thus resembling colorectal carcinogenesis. As in colon cancer, deregulated expression of G1 proteins may perturb cell cycle control in benign adenomas of the small bowel and thereby enhance tumor progression. Increased expression of cell cycle inhibitors in tumors may serve as a defense mechanism for tumor progression.

Adenocarcinoma↗

PubMed1997

Human umbilical cord blood myeloid progenitor cells are relatively chemoresistant: a potential model for autologous transplantations in HIV-infected newborns.

Vertical transmission from mother to child occurs in 15-39% of women infected with the human immunodeficiency virus (HIV). Stem cell transplantation has recently been suggested as a potential therapy for patients with HIV infection. We have examined the possible advantages of human cord blood (HUCB) stem cells over bone marrow (BM) stem cells in the treatment of HIV-infected newborns. HUCB myeloid progenitors were found to be statistically more resistant to interferon-alpha (IFN-alpha), cytarabine (ARA-C), and eilatin than BM myeloid progenitor cells grown with IL-3 (P < 0.05). HUCB treated with IFN-alpha, ARA-C, and eilatin demonstrated a significantly higher capacity for self-renewal manifested by delta assay following 7 days in liquid culture. We, therefore, suggest that HUCB purged by anti-HIV drugs may be a source for autologous transplantation in HIV-infected newborns.

Antiviral Agents↗

PubMed1997

The putative role of arylsulfatase in interleukin-2-mediated cytotoxicity and interleukin-7-mediated bactericidal activity of natural killer cells.

We have examined the cytolytic and bactericidal activity of resting and cytokine-stimulated natural-killer (NK) cells against K562 and Daudi cell lines and Escherichia coli, respectively. Unstimulated NK cells showed considerable cytolytic activity against K562 (64 +/- 4%) and relatively low activity against the Daudi cell lines (22 +/- 9%). Pretreatment of NK cells with the arylsulfatase (AS) type-II-specific inhibitor NaH2PO4 reduced cytotoxicity towards K562 and Daudi 1.3- and 2.9-fold (p < 0.05; n = 12), respectively, indicating that AS participates in NK-mediated cytotoxicity. Interleukin-2 (IL-2) (200 units/ml) caused a 1.3- and 3.5-fold (p < 0.5; n = 12) enhancement of NK cytotoxic activity against K562 and Daudi, respectively. Pretreatment of these cells with the AS type-II-specific inhibitor NaH2PO4 reduced cytotoxicity 1.1-fold towards K562 (p < 0.05; n = 12) and 1.2-fold towards Daudi (p > 0.05; n = 12) indicating that AS does not participate in IL-2-mediated NK cytolytic activity against these cell lines. IL-7 (3 units/ml) did not cause any enhancement of NK cytolytic activity. Unstimulated NK cells showed considerable bactericidal activity against E. coli (23 +/- 4%). Incubation of resting NK cells with NaH2PO4 reduced the bactericidal effect only by 1.09-fold (p > 0.05; n = 12), indicating that AS does not mediate this effect. IL-2 (200 units/ml) and IL-7 (3 units/ml) enhanced the bactericidal activity 1.5- and 2.2-fold, respectively (p < 0.05; n = 12). This effect was not influenced by incubation of IL-2-stimulated cells with NaH2PO4, indicating that AS does not participate in the IL-2-mediated NK bactericidal effect. IL-2 seems to exert its stimulatory effect upon NK-mediated bactericidal activity by a different, non-AS-dependent mechanism. However, incubation of IL-7-stimulated NK cells with NaH2PO4 reduced the NK bactericidal effect by 1.2-fold (p < 0.05; n = 12), indicating that AS may have a role in this reaction. These data can be further confirmed by detection of AS through degranulation of NK cells, showing that IL-2 induced only mild degranulation of resting and f-MLP-stimulated NK cells (26 +/- 1% vs 22 +/- 2% and 31 +/- 2% vs 29 +/- 2, respectively) (p > 0.05; n = 8). In contrast, IL-7 showed significant enhancement of AS release in resting or f-MLP-induced NK cells (36 +/- 3% vs 22 +/- 2% and 49 +/- 3% vs 29 +/- 2%, respectively) (p < 0.05; n = 8).

Arylsulfatases↗

PubMed1997

Enhanced hematopoiesis in sublethally irradiated mice treated with various quinolones.

We compared the effect of 6 quinolones on growth of murine bone marrow (BM) progenitor cells in vitro, and their in vivo effect on repopulation of BM and on survival of sublethally irradiated mice. The addition of clinically attainable concentrations of ciprofloxacin, sparfloxacin or clinafloxacin, in concert with pokeweed mitogen (PWM) to murine spleen cells, resulted in a significant enhancement in colony stimulating activity. A 1.5-1.8 fold increase in the number of myeloid progenitors (CFU-C) was observed in the presence of quinolone-PWM spleen conditioned medium (SCM) (prepared with the above-mentioned quinolones) compared with control cultures exposed to PWM-SCM only. Three other quinolones showed either no stimulatory-effect (fleroxacin, norfloxacin) or had an inhibitory effect (ofloxacin) on CFU-C growth. The stimulatory quinolones share in common a cyclopropyl moiety at position N1 of the quinolone ring. This moiety is lacking in the other 3 quinolones. The secretion of interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) by murine spleen cells exposed to quinolone-PWM-SCM was significantly enhanced with all 6 quinolones. However, this effect was associated with a parallel increase in CFU-C only with ciprofloxacin (10 micrograms/mL), sparfloxacin (1 microgram/mL) and clinafloxacin (0.05 microgram/mL). The in vivo activity was assessed in sublethally irradiated mice (650 rad) treated with quinolones for 5 d. The number of CFU-C in BM and the number of peripheral white blood cells (WBC) 8 d post-irradiation was significantly enhanced in mice treated with ciprofloxacin (45 mg/kg/d), sparfloxacin (22.5 mg/kg/d) and clinafloxacin (11.25 mg/kg/d) compared to saline treated animals (p < or = 0.05). Clinafloxacin at higher dosage (45 mg/kg/d) resulted in a decrease in myeloid progenitors in BM. A similar increase in progenitors and WBC was observed in animals treated with high doses, above clinical relevance, of ofloxacin, and norfloxacin (90 mg/kg/d), and with fleroxacin (45 and 90 mg/kg/d). Quinolone-treated animals, at the above-cited doses, showed enhanced survival on d18 compared to saline treated animals. The only exception was the higher mortality of clinafloxacin-treated mice. The above observations imply that certain quinolones, sharing specific molecular structure, are potential immunomodulators at clinically relevant concentrations. These compounds should be further studied in neutropenic patients and BM or peripheral blood progenitor cell recipients.

Animals↗

PubMed1996

Granulocyte-macrophage colony-stimulating factor dependent monocyte-mediated cytotoxicity post-autologous bone marrow transplantation.

We investigated the in vitro antitumor activity of monocytes derived from autologous bone marrow transplanted (ABMT) patients treated in vivo with granulocyte-macrophage colony-stimulating factor (GM-CSF). Thirty-four patients (17 female, 17 male), median age 42 (range 3-57) years, were enrolled in the study. Fourteen patients were diagnosed with non-Hodgkin's lymphoma (NHL), eight with Hodgkin's disease (HD), nine with breast cancer and three with neuroblastoma. Six patients who did not receive GM-CSF post-ABMT served as controls. We assessed cytotoxicity, antibody-dependent cellular cytotoxicity (ADCC), expression of the activation antigen CD16, and cytokine production by an enriched population of monocytes (> 90% CD+14) pre-, during and post-GM-CSF administration. Within the group of patients receiving treatment, ADCC was significantly higher during in vivo GM-CSF administration than post-therapy (P < 0.05) and in 50% of these patients, ADCC increased during in vivo GM-CSF administration over pretreatment values. In addition, in vivo GM-CSF administration caused the monocytes to secrete elevated levels of tumor necrosis factor-alpha (TNF-alpha) and GM-CSF (P < 0.05). We conclude that GM-CSF augments monocyte-mediated cytotoxicity post-ABMT, and therefore may have a role in controlling minimal residual disease post-transplant.

Adolescent↗

PubMed1996

Synergistic effects of interleukin-11 with other growth factors on the expansion of hematopoietic progenitors from normal individuals and chronic myeloid leukemia patients resistant to treatment with cytosine arabinoside or eilatin.

Interleukin-11 (IL-11) is a novel cytokine that has been shown to stimulate human hematopoietic progenitors including the CD34+ CD33- DR- early progenitors. IL-11 has little effect on its own but it synergizes with other hematopoietic growth factors. We investigated the recovery of human myeloid progenitors incubated with IL-11 alone or in combination with other cytokines, including stem cell factor (SCF), interleukin-3 (IL-3) and granulocyte macrophage colony-stimulating factor (GM-CSF) following their in vitro treatment with ARA-C (10(-9) M) or Eilatin (10(-7) M). IL-11 in combination with IL-3 and GM-CSF markedly increased CFU-C colony growth pre- and post-ARA-C or Eilatin incubation from CML and normal individual bone marrow (BM) cells. Similarly, IL-11 alone or in combination with other cytokines increased cell recovery following 7-day suspension culture. A decrease in BCR/ABL fusion product was observed (by FISH analysis) after incubation of BM cells from CML patients in liquid culture for 7 days with 10(-9) M ARA-C or 10(-7) M Eilatin in the presence of IL-11 alone or in combination with other cytokines. These results indicate that following cytoreductive therapy IL-11 may enhance to a greater extent the growth of normal myeloid progenitors than the malignant clone and may, therefore, be of clinical importance for CML patients treated with chemotherapeutic agents.

Animals↗

PubMed1996

Automated monitoring of apoptosis in suspension cell cultures.

Cell death by apoptosis is often accompanied by extensive DNA cleavage at internucleosomal linker sites. Thus, the foremost techniques for estimating apoptosis are based on biochemical, electrophoretic, or flow cytometry analysis of nuclear DNA. However, apoptosis is also associated with a chain of morphologic changes in the nuclear and cytoplasmic structures that are easily recognizable using light microscopy. We suggest that changes in morphology of cells undergoing apoptosis might cause characteristic changes in their optical properties. It follows that continuous measuring of the OD of cells undergoing apoptosis would enable the study of the kinetics of cell death. We recently described an automated microculture kinetic (MiCK) assay that provides multiple OD measurements in nondisrupted cell cultures. In the present study the MiCK assay was employed to follow OD changes in HL-60 and OCI/AML-3 myelogenous leukemia cells and murine thymocytes exposed to ethanol, hydrogen peroxide, etoposide, cisplatin, doxorubicin, or hyperthermia; i.e., divers stimuli known to induce cell death via apoptosis or necrosis. The MiCK assay revealed prominent differences between the optical properties of the cells undergoing the two different modes of death. Plotting the OD data accumulated during the assay period against time betrayed characteristic patterns of either "apoptotic" or "necrotic" OD curves. The best fit slope of the indicative of apoptosis steep rising component of the apoptotic curve, correlated directly with the percentage of morphologically apoptotic cells in the culture. Criteria for graphical estimate of apoptosis were suggested and used to study apoptosis induced in HL-60 cells by the chemotherapy compounds etoposide and cisplatin. The MiCK assay demonstrated markedly varying time courses of the cell apoptotic response to these two drugs. In both cases, however, the time of graphically predicted peaks of apoptosis correlated with the time of morphologically and electrophoretically recognized peaks of apoptosis. Adaptability of the MiCK assay to a 96-well microplate format opens the way for large-scale studying of cell apoptotic response to various stimuli. An important technical advantage of the automated MiCK assay of apoptosis is that it does not require additional laboratory procedures after microcultures are initiated.

Animals↗

PubMed1995

Brain sites involved in mu-opioid receptor-mediated actions: a 2-deoxyglucose study.

Brain regions that may be functionally involved in the neuropharmacological actions of mu-opioid agonists have been examined in conscious rats using the quantitative [14C]2-deoxyglucose autoradiographic technique. At 0.5 microgram and 1 microgram intracerebroventricularly the highly selective mu-opioid receptor agonist D-Ala2, MePhe4, Gly-ol5-enkephalin effected statistically significant increases as well as statistically significant decreases in regional glucose utilization: in limbic structures, such as hippocampal formation, medial amygdala and lateral septum, glucose utilization was most prominently increased after D-Ala2, MePhe4, Gly-ol5-enkephalin; glucose utilization was further increased in the lateral habenular nucleus, the hypothalamus, ventromedial nucleus and dorsal raphe; whereas decreases were found in the mamillary body and anterior thalamus. Glucose utilization in structures associated with somatosensory and nociceptive processing was increased in the central gray of the midbrain and decreased in the nucleus gelatinosus. Only increases in glucose utilization were produced by D-Ala2; MePhe4, Gly-ol5-enkephalin in brain regions involved in motor control, including the globus pallidus, the substantia nigra, pars reticulata, the nucleus ruber and the cerebellum, and brain regions involved in visual processing--the visual cortex and superior colliculus deep layer. It is concluded that this pattern of regional changes underlies the mu-opioid receptor-mediated antinociceptive-, epileptogenic-, memory- and mood-modulating actions of mu-opioid agonists.

Amino Acid Sequence↗

PubMed1995

Histamine enhances granulocyte-macrophage colony-stimulating factor and interleukin-6 production by human peripheral blood mononuclear cells.

The effect of histamine on the production of cytokines by subpopulations of mononuclear cells was studied. A 3.5-fold increase in the number of myeloid colony-forming units (CFU-C) was observed when bone marrow cells were cultured in the presence of conditioned medium prepared from nonadherent mononuclear cells cultured with 10(-4) M histamine (CM-histamine) compared with phosphate-buffered saline (CM-PBS). Using ELISA and radioimmunoassay kits, histamine was found to enhance the production of GM-CSF (9.6-fold) and IL-6 (8.2-fold) by mononuclear cells but not by nonadherent cells or large granular lymphocytes. Anti-GM-CSF and anti-IL-6 antibodies markedly blocked cytokine activity in CM-PBS, whereas the blocking effect in CM-histamine was moderate, indicating enhanced GM-CSF and IL-6 activity in CM-histamine. No GM-CSF or IL-6 levels could be detected in CM-histamine or CM-PBS prepared from CD3+, CD4+, or CD8+ lymphocytes. Preincubation of CM-histamine with H1 and H2 receptor antagonists resulted in complete blocking of the histamine-enhanced colony-stimulating activity. We conclude that histamine is able to activate human mononuclear cells to generate cytokines such as GM-CSF and IL-6 via H1 and H2 receptors.

Antibodies↗