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Biomedical subjects

B C Millar

Publications and source records attributed to B C Millar.

At least 109 records · Page 6Linked to original sources
PubMed1990

Evidence that multiple myeloma may be regulated by homeostatic control mechanisms: correlation of changes in the number of clonogenic myeloma cells in vitro with clinical response.

Myeloma colonies (MY-CFUc) could be grown in vitro for 6 months (median time) after a group of 12 myeloma patients had reached complete remission (CR). In a second group of 25 patients MY-CFUc increased in 17/25 and GM-CFUc in 20/25 patients after cyclophosphamide even though 24/25 patients had a partial response to VAMP and one was in CR. These data suggest that cell killing by cyclophosphamide stimulates residual tumour cells into proliferation and adds further support to the idea that myeloma is under some degree of homeostatic control which may be analogous to that in normal bone marrow. Although lymphoplasmacytoid myeloma cells may be more drug resistant than plasmacytoid myeloma cells in vitro, it was not possible to conclude that the emergence of lymphoplasmacytoid cells at relapse was indicative of resistance to further treatment.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1990

Colony stimulating activity in the serum of patients with multiple myeloma is enhanced by interleukin 3: a possible role for interleukin 3 after high dose melphalan and autologous bone marrow transplantation for multiple myeloma.

Sera from 36/37 multiple myeloma patients and 19/21 sera from patients with other solid or liquid tumours had granulocyte-macrophage colony stimulating activity (CSA) towards normal human donor bone marrow whereas 1/16 sera from normal donors had this activity. Unlike human rhGM-CSF and GM-CSF from 5637 (human bladder cell line) conditioned medium which is heat stable, CSA from serum is heat labile (56 degrees C/30 min). In multiple myeloma patients, CSA was detectable more than 2 years after treatment with 'high dose melphalan. Although multiple myeloma patients, at relapse, have sufficient CSA in their serum to produce maximal stimulation of GM-CFUc from normal donor bone marrow in vitro, their own GM population responds poorly. The results suggest that the failure of patients own bone marrow to respond to endogenous CSA may be due to damage to the stem cells of the marrow or the failure of precursor cells to respond to CSA. Addition of rhIL-3 to myelomatous serum increased the number of GM-CFUc from both normal and myelomatous bone marrow but did not stimulate the growth of MY-CFUc significantly. The results suggest that rhIL-3 may assist bone marrow recovery in multiple myeloma patients after intensive chemotherapy.

Bone Marrow↗
PubMed1990

Interleukin-6 is a cofactor for the growth of myeloid cells from human bone marrow aspirates but does not affect the clonogenicity of myeloma cells in vitro.

Several groups have claimed that IL-6 is a growth factor for human myeloma cells in vitro. Bone marrow aspirates from 30 patients at different stages of treatment with VAMP/high dose melphalan, were examined for myeloma colony formation (MY-CFUc) using a clonogenic assay in vitro. Myeloma cells from 16/30 patients produced MY-CFUc in our assay system, which uses heavily irradiated HL60 cells as an underlay in soft agar. These heavily irradiated cells were shown to be essential for the inhibition of granulocyte-macrophage colonies (GM-CFUc). The addition of recombinant human IL-6 (10 ng/plate) reduced the number of bone marrow samples which produced MY-CFUc from 16 to six. Furthermore, the addition of antibody to IL-6 (1 microgram/plate) failed to inhibit MY-CFUc from 6/7 samples. Conditioned medium from human peripheral blood mononuclear cells (PBMC-CM) contains approximately 2 ng/ml IL-6 and can be used to stimulate the growth and maintenance of the B9 murine IL-6 dependent hybridoma cell line. Recombinant human IL-6 supported the growth of B9 cells in a clonogenic assay and growth was inhibited by anti-IL-6 in the presence of rhIL-6 or PBMC-CM. Mononuclear cells from a second group of myeloma patients were cultured in soft agar in a mixture of PBMC-CM and fresh growth medium. Nine of the 10 samples produced myeloid colonies which consisted of granulocytes, monocytes and macrophages and the number of colonies was reduced by at least 50% in 6/8 samples when anti-IL-6 was added to the cultures. In no instance were MY-CFUc produced. Also, conditioned medium from the bladder carcinoma cell line 5637, which is used routinely as a source of granulocyte-macrophage colony stimulating factor (GM-CSF), contains approximately 4 ng/ml IL-6. Although rhIL-6 failed to stimulate GM-CFUc in the absence of other growth factors, addition of anti-IL-6 to cultures containing a suboptimal amount of 5637-CM reduced the number of colonies by 50%. These data provide evidence that IL-6 is a cofactor for the growth of myeloid precursors but does not affect the proliferation of human myeloma cells in vitro.

Adult↗
PubMed1989

The negative inotropic effect of neuropeptide Y on the ventricular cardiomyocyte.

The effect of neuropeptide Y (NPY) on cell contractions of ventricular myocytes isolated from the adult rat heart was investigated. Maximum changes in cell length (dL) during stimulated (0.5 Hz) contractions were determined in presence of the phosphodiesterase inhibitor Ro 20-1724 (0.5 mM) and adenosine deaminase (5 U/ml). Under these basal conditions NPY (10(-6) M) reduced dL by 39% of control. Isoproterenol (10(-6) M) increased dL by 105% of control; the EC50 was 2 x 10(-9) M. NPY reduced the increase in dL achieved by isoproterenol in a dose dependent manner. The IC50 value was 1 x 10(-9) M and NPY (10(-6) M) produced complete inhibition. In the absence of the phosphodiesterase inhibitor the IC50 was 4 x 10(-9) M. The EC50 of isoproterenol and IC50 of NPY producing accumulation of cAMP in myocytes (Millar et al. 1988) exceeded the respective values of dL by one order of magnitude. Prior treatment of the myocytes with pertussis toxin abolished the potency of NPY to antagonize the increase in dL by isoproterenol while not interfering with the response to the beta-agonist. These results demonstrate a negative inotropic effect of NPY on the ventricular myocardial cell. Complete abolition of the effect of NPY by pertussis toxin indicate that this effect is mediated by a sarcolemmal receptor for NPY linked to adenylate cyclase via an inhibitory guanine nucleotide binding protein.

4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone↗
PubMed1989

In vitro studies of ways to overcome resistance to VAMP--high dose melphalan in the treatment of multiple myeloma.

Myeloma colonies (MY-CFUc) from 7/24 patients undergoing treatment with VAMP (vincristine, adriamycin and methyl prednisolone) and high dose melphalan (HDM) were melphalan-resistant. It was not possible to conclude that VAMP induced melphalan resistance in MY-CFUc, but that resistance is endogenous in some myeloma cell populations. In 12/13 of the same patients of whom four had MY-CFUc which were melphalan resistant, the sensitivity of MY-CFUc and GM-CFUc to busulphan was similar. Thus resistance of MY-CFUc to melphalan did not confer resistance to busulphan. MY-CFUc from 1/7 of a second group of patients were adriamycin-resistant. This resistance was removed when the cells were treated with a combination of verapamil (3 micrograms/ml) and adriamycin. Verapamil also enhanced the toxicity of adriamycin to MY-CFUc from two patients where there was no evidence for adriamycin resistance. In these three patients the sensitivity of both MY-CFUc and GM-CFUc was similar after treatment with verapamil. Verapamil did not affect the uptake or efflux of 3H-daunorubicin in sensitive and resistant RPMI-8226 cells (myeloma) and peripheral blood mononuclear cells from a normal donor; neither did it affect the binding of 3H-daunorubicin to nucleic acid. It is concluded that verapamil may be a useful adjuvant to VAMP chemotherapy and that busulphan may provide an alternative to melphalan in patients whose myeloma cells are melphalan resistant.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1989

High-dose chemotherapy and autologous bone marrow transplantation for myeloma.

In three studies we have attempted to increase the complete remission rate in a series of patients with multiple myeloma under the age of 69. A CR rate of 27% was seen in 63 patients treated with intravenous high-dose melphalan 140 m/m2 with or without the addition of high-dose methyl prednisolone 1 g/m2 for 5 days (in 22 patients). In a third study a CR rate of 50% was seen in 50 patients treated in a programme in which vincristine, adriamycin and methyl prednisolone was first given and patients then received high-dose melphalan 140-200 mg/m2 with an autologous bone marrow transplant where possible. Median remission duration in the first two studies was 19 months with a median survival of 5 years. A definition of complete remission in myeloma is proposed.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1988

Increase in clonogenic tumour cells in bone marrow of patients with multiple myeloma treated with vincristine, doxorubicin, and methylprednisolone.

The relation between clonogenic myeloma cells, assayed in vitro, and clinical status was studied in 20 patients with multiple myeloma before and after VAMP therapy (vincristine, doxorubicin, and methylprednisolone). 14 patients showed an improvement in clinical status, as judged by a fall in myeloma protein and a decrease in plasmacytoid myeloma cell infiltration in the bone marrow. However, in 11 of these 14, there was an increase in the number of clonogenic myeloma cells. This suggests that the treatment changed the myeloma cell population in the residual bone marrow. To be effective, subsequent chemotherapy should be aimed at destroying this population of myeloma cells.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1988

The antiadrenergic effect of neuropeptide Y on the ventricular cardiomyocyte.

The effect of neuropeptide Y (NPY) on adenylate cyclase activity was examined in ventricular myocytes isolated from the adult rat heart. In the presence of the phosphodiesterase inhibitor Ro 20-1724 (0.5 mM) and adenosine deaminase (5 U/ml), these intact cells accumulate cyclic AMP when stimulated by isoproterenol. NPY (10(-9) to 10(-6) M) reduced the degree of cAMP accumulation achieved by 10(-7) M isoproterenol in a dose dependend manner by 10 to maximally 48%. The IC50 value was 3 x 10(-8) M NPY. A maximal concentration (10(-6) M) of N6-phenylisopropyladenosine (PIA) decreased cAMP levels by 39%, i.e. to a similar extent. Prior treatment of the myocytes with pertussis toxin (1 microgram/ml for 6 h) increased the mean stimulated values in the presence of isoproterenol (10(-7) M) by a factor 4.1. In such cells, NPY and PIA were ineffective in antagonizing the stimulation of cAMP production by isoproterenol. These results indicate that the ventricular myocyte has receptors for NPY, similar to the A1 adenosine-receptor in that they are linked to the adenylate cyclase by an inhibitory guanylate binding protein.

Adenylate Cyclase Toxin↗
PubMed1988

A simple method for culturing myeloma cells from human bone marrow aspirates and peripheral blood in vitro.

A double layer agar technique has been developed to grow myeloma colonies (MY-CFUc) from human bone marrow aspirates and peripheral blood. Heavily irradiated HL60 cells (5 x 10(5)/plate) are added to an agar underlay in growth medium containing 0.5% agar. Mononuclear cells from the test bone marrow or blood are overlayered in either 0.2 ml HL60-conditioned medium (HL60-CM) or in 0.5 ml growth medium containing 0.23% agar, and the cultures are incubated at 37 degrees C in an atmosphere of 5% CO2, 10% O2 and 85% N2. Colonies (greater than 50 cells) form between 2 and 3 weeks. Using this method 60/68 samples of bone marrow and 7/12 samples of blood from 54 patients have produced colonies in soft agar and in liquid on an agar underlay. The cells which form these colonies are of two distinct sizes, the larger cells being plasmacytoid and the smaller lymphoid. The two cell types are usually, but not always, present in separate colonies. Both plasmacytoid and lymphoid cells carry the isotype of the respective patient's myeloma protein and the plasma cell marker (HAN PC1). This technique has enabled us to culture myeloma cells from patients with as few as 2% plasma cells in the bone marrow but it does not permit the growth of normal B, T or granulocyte-macrophage colonies (GM-CFUc). The drug sensitivity of myeloma cells (MY-CFUc) compared with normal haemopoietic cells (GM-CFUc) can be measured using dose-response curves in individual patients. Furthermore, this method can detect resistant subpopulations within a given myeloma sample.

Bone Marrow↗
PubMed1987

Lymphotoxic activity of methyl prednisolone in vitro--I. Comparative toxicity of methyl prednisolone in human cell lines of B and T origin.

The cytotoxic activity of methyl prednisolone was compared in EB-3(B), NALM-6(B), CCRF-CEM(T) and RPMI-8226 (plasma cell) cell lines derived from human lymphoid malignancies. Whereas EB-3 cells were steroid-sensitive, NALM-6 cells were partially sensitive and CCRF-CEM and RPMI-8226 were steroid resistant at concentrations of methyl prednisolone up to 10(-4) M. A high concentration of methyl prednisolone, 2.5 X 10(-3) M was toxic to all cell lines. Steroid-sensitivity did not correlate with the incorporation of [3H] dexamethasone and could not be mimicked by flurbiprofen, a non-steroidal anti-inflammatory agent. Both theophylline and di-butyryl cAMP were toxic towards NALM-6, EB-3 and CCRF-CEM cells; however, this toxicity was reversible and did not reflect the cells' sensitivities towards methyl prednisolone. Furthermore, elevated levels of cAMP in theophylline-treated cells, were not demonstrable in cells treated with methyl prednisolone at toxic or non-toxic concentrations of the steroid. Steroid-sensitive EB-3 cells exposed to 10(-5) M methyl prednisolone, produced a soluble factor which was toxic CCRF-CEM cells.

B-Lymphocytes↗
PubMed1987

Comparison of melphalan toxicity in human lymphocytic cells and Chinese hamster cells in vitro: the relationship between DNA-DNA cross-link formation and clonogenic survival.

Human CCRF-CEM ('T' cell), EB3p ('B' cell) and RPMI-8226 (myeloma cell) lymphocytic cell lines were an order of magnitude more sensitive to melphalan (MEL) than Chinese hamster, V-79-753B, cells even though the amount of [14C]MEL they incorporated was less than 50% of that incorporated into the rodent cells: the D0 values were 0.16, 0.20 and 0.30 microgram/ml respectively compared with 1.6 micrograms/ml. Furthermore, MEL sensitivity was not related to the total thiol content of the cells. DNA-DNA cross-linking was not detectable in lymphocytic cells using the alkaline elution technique at doses of MEL used for clonogenic survival, whereas in Chinese hamster cells both parameters were assessable within the same dose range. At high concentrations of MEL there was a direct relationship between DNA-DNA cross-linking and drug dose in each lymphocytic cell line. Changes in the amounts of DNA-DNA cross-linking, at different MEL concentrations, increased directly with the sensitivity of the cells, viz. CCRF-CEM greater than EB3p greater than RPMI-8226. Calculated survival values for doses of MEL which produced measurable DNA-DNA cross-linking showed that there was a similar relationship between these parameters for the three lymphocytic cell lines which was different from that for Chinese hamster cells. It is concluded that the contribution of DNA-DNA cross-links in determining cell survival after MEL treatment is both quantitatively and possibly qualitatively different in human and rodent cells and that DNA-DNA cross-linking cannot be used as an indicator of MEL sensitivity in human lymphocytic cells unless parallel clonogenic survival studies are also undertaken.

Animals↗
PubMed1986

Comparative studies of total cross-linking, cell survival and cell cycle perturbations in Chinese hamster cells treated with alkylating agents in vitro.

The toxicities of 4-hydroperoxycyclophosphamide (4-OOH CY), phosphoramide mustard (PM), melphalan (MEL) and busulphan (BU) have been compared in Chinese hamster cells, V-79-753B. The initial total amount of cross-linking was a determining factor for the clonogenic survival of cells treated with MEL or PM. Although 4-OOH CY generated cross-links in this cell line, this damage did not account for the toxicity of the compound. There was no evidence for cross-link formation in cells treated with BU, even at a dose of the drug (1000 micrograms/ml) that was too toxic to measure clonogenic survival. Comparison for the four compounds at equitoxic doses showed that both PM and MEL caused the arrest of the cell cycle at G2 which persisted after drug removal. This was accompanied by a decline in the population growth rate and a decrease in total cell count. In contrast, both BU and 4-OOH CY caused a temporary arrest of the cell cycle G2, 24 hr after drug removal. However, the cell cycle distribution returned the control values within 3-4 days after treatment. Both BU and 4-OOH CY showed little effect on the initial growth rate of the cells. It is concluded that the initial amount of cross-links contributes to the toxicity of PM and MEL. However, it is unlikely that the generation of cross-links is of major importance for the toxicity of either 4-OOH CY or BU.

Alkylating Agents↗
PubMed1985

Mesna does not reduce cisplatin induced nephrotoxicity in the rat.

Although mesna afforded protection against the cytotoxicity of cisplatin in Chinese hamster cells, line V-79-753B, in vitro, there was no evidence for protection against nephrotoxicity when this drug combination was examined in the rat. It seems likely that cisplatin-induced nephrotoxicity is mediated by intracellular events in kidney cells which cannot be inhibited by mesna possibly due to its presence within cells in vivo as the stable and unreactive disulphide. On the basis of these data it is unlikely that combinations of mesna and cisplatin will be of therapeutic benefit in man.

Animals↗
PubMed1985

Studies on the relationship between the radiation resistance and glutathione content of human and rodent cells after treatment with dexamethasone in vitro.

A 20 h pre-treatment of human cells from normal (foetal lung) or malignant origin (glioma, lines U118 MG and U251 MG and bladder carcinoma, line EJ) with dexamethasone failed to increase their radiation resistance in vitro despite a 2-fold increase in the GSH content of a glioma cell line, U251 MG, and a small but significant increase in the GSH content of EJ bladder carcinoma cells. In contrast, there was a correlation between an increase in radiation resistance and an elevated GSH content of rodent cells (Chinese hamster lung, line V-79-379A; ovary, line CHO; rat hepatoma, line HTC, and mouse neuroblastoma, line NB413A) after a similar pre-treatment. The results suggest that enhancement of radiation resistance cannot be directly ascribed to an elevated GSH content in steroid-treated cells. On the basis of these data it is unlikely that the efficacy of radiotherapy will be diminished amongst patients receiving concomitant treatment with dexamethasone. However, in vivo testing is required to confirm these findings.

Animals↗
PubMed1985

Polyfunctional radiosensitizers. VII. Radiosensitization by conformationally-restricted isomers of a nitroxyl biradical in vitro.

Bothtrans-N,N'-bis(2,2,6,6-tetramethyl-1-oxyl-4-piperidinyl)-1, 2-diaminocyclopropane[Ro31-2269] and its cis isomer [Ro 31-2778] selectively sensitized hypoxic Chinese hamster cells, line V-79-753B, to radiation by decreasing both the D0 value and extrapolation number, whereas a related dibasic monoradical Ro 31-2655 decreased D0 alone. Although sensitization was maximal after a 1-hr cell-drug contact time, cells continued to accumulate both Ro 31-2269 and Ro 31-2778 when this contact time was increased up to 3 hr. There was no evidence for competition between either biradical and 2,2,6,6-tetramethyl-4-piperidinol-N-oxyl (TMPN) at equimolar concentration or biradical and 0.82 microM oxygen when cells were equilibrated with the biradicals for 3 hr prior to irradiation in the presence of mixtures of either oxygen and biradical, TMPN and biradical, or TMPN alone. Furthermore, when cells were equilibrated with an equimolar radical concentration of the trans isomer Ro 31-2269 and TMPN for 1 hr prior to irradiation in the presence of the mixture, there was no appreciable effect on sensitization of the slope of the hypoxic cell survival curve, but shoulder modification was reduced. When cells were equilibrated with the trans isomer Ro 21-2269 prior to irradiation in combination with 2.92 microM oxygen, cell survival was similar to that seen for cells irradiated with this concentration of oxygen alone. Examination of the plasma membrane from cells equilibrated with the trans biradical Ro 31-2269 showed that the drug accumulated in the membrane when compared with the concentration found in whole cells. Experiments with the conformationally-unrestricted biradical bis(2,2,6,6-tetramethyl-1-oxy-4-piperidinyl) succinate [Ro 03-6061] showed that when cells were equilibrated with the compound for 1 hr prior to irradiation in hypoxia in the presence of a mixture containing an equimolar radical concentration of TMPN, there was an increase in both the slope and the extrapolation number compared with values for hypoxic cells irradiated in the presence of this biradical alone. Furthermore, when cells which had been equilibrated with Ro 03-6061 were washed free of the drug, there was a residual decrease in both the D0 and extrapolation number of the hypoxic cell survival curve for at least 3 hr after removal of the compound. The results are discussed in terms of a model to account for sensitization by these compounds.

Animals↗