Differential effect of phorbol esters and interleukin-3 on protein kinase C isoform content and kinase activity in the FDC-P1 cell line

FD/PMA is a subclone of the interleukin-3 (IL-3)-dependent, FDC-P1 cell line, which proliferates in response to either 12-O- tetradecanoylphorbol-13 acetate (PMA) or IL-3. While several endogenous substrates were phosphorylated in response to protein kinase C (PKC) activation in FDC-P1, phospholipid-dependent phosphorylation in the FD/PMA grown in PMA was not observed. Basal, phosphatidylserine- independent, and diolein-independent phosphorylation of cytosolic substrates with molecular weights of 17, 52, 57, and 105 Kd were enhanced in FD/PMA cells grown in PMA as compared with FDC-P1 cells cultured in IL-3. Phosphorylation of a 105-Kd substrate was enhanced in the particulate fraction of FD/PMA cells maintained in PMA. The 17-Kd substrate in FD/PMA cells comigrated with a substrate phosphorylated in a PKC-dependent manner in FDC-P1 cells. Phosphorylation of the 52- and 57-Kd substrates, but not of the 17-Kd substrate, was inhibited by H-7 and staurosporine. A portion of the PMA-induced cytosolic kinase activity coeluted with PKC on diethyl aminoethyl chromatography. While FD/PMA cells cultured in PMA contained negligible PKC-dependent phosphorylation of endogenous substrates or histone, alpha and epsilon PKC isoforms were detected by Western blot analysis. PKC phosphotransferase activity was observed in FD/PMA cells grown in PMA when peptides corresponding to residues 720 to 737 of PKC-epsilon or residues 4 to 14 of myelin basic protein were used as substrates. These data indicate that maintenance of FD/PMA cells in PMA stimulates proliferation and markedly alters PKC substrate specificity. Generation of at least two phospholipid-independent kinases occurs in PMA-treated cells.     

Correlation of drug sensitivity in vitro with clinical responses in childhood acute myeloid leukemia

Clonogenic cells from 41 children with newly diagnosed acute myeloid leukemia (AML) were tested in vitro for their sensitivity to cytarabine (Ara-C) and daunorubicin (DNR). The findings were then compared with the patients' responses to induction chemotherapy that uniformly included Ara-C and DNR. Light-density marrow cells were incubated with either or both drugs for one hour and cultured over leukocyte feeder layers; clusters and colonies were scored on days 7, 10, and 14. Only the percentage of cell kill in the presence of 1.8 mumol/L DNR was significantly associated with responses to induction therapy: median of 45% (range, 0% to 98%) for patients achieving complete remission v 16% (range, 4% to 23%) for nonresponders (P = .007). The relationship between clonogenic cell kill less than or equal to 23% and clinical responses was striking. Of the 11 evaluable patients with in vitro findings in this category, ten either failed induction therapy or relapsed within 1 year after attaining remission. Kaplan-Meier analysis of relapse-free survival times indicated longer durations of remission for patients whose blast cells showed increased sensitivity in vitro to Ara-C alone, DNR alone, or a combination of the two agents. Seven of 11 patients with cell kills of greater than or equal to 49% in the presence of 1.25 mumol/L Ara-C remain free of leukemia, compared with only one of 12 whose cells were less sensitive to the drug (P = .006). We conclude that the in vitro sensitivity of clonogenic leukemic progenitors to DNR and Ara-C correlates with treatment outcome in children with newly diagnosed AML.     

Sulfhydryl reagents induce altered spectrin self-association, skeletal instability, and increased thermal sensitivity of red cells

Incubation of erythrocytes with the sulfhydryl reagent N-ethyl- maleimide (NEM) results in altered spectrin self-association and formation of dimers on the membrane. Skeletons isolated from these cells exhibit marked skeletal instability. In addition, NEM treatment induces increased thermal sensitivity of both cells and purified spectrin. These effects were not produced in aerobically incubated glucose-6-phosphate dehydrogenase deficient cells and were therefore presumably not due to depletion of intracellular reduced glutathione. These effects were produced by another permeant sulfhydryl reagent, monobromobimane, but not by its membrane-impermeant derivative. We conclude that spectrin sulfhydryl groups play an important role in spectrin self-association and thermal stability.     

Telomeres in the mouse have large inter-chromosomal variations in the number of T2AG3 repeats

The ultra-long telomeres that have been observed in mice are not in accordance with the concept that critical telomere shortening is related to aging and immortalization. Here, we have used quantitative fluorescence in situ hybridization to estimate (T₂AG₃)_n lengths of individual telomeres in various mouse strains. Telomere lengths were very heterogeneous, but specific chromosomes of bone marrow cells and skin fibroblasts from individual mice had similar telomere lengths. We estimate that the shortest telomeres are around 10 kb in length, indicating that each mouse cell has a few telomeres with (T₂AG₃)_n lengths within the range of human telomeres. These short telomeres may be critical in limiting the replicative potential of murine cells.     

The Kheda malaria project: the case for environmental control

The Kheda project experience has shown that bio-environmentalcontrol of malaria is feasible, cost effective and ecologicallysound. It clearly brings out the need to consider health issuesat the planning stage of all developments. Bio-environmentalcontrol of malaria is suggested as the first line of attackfor the control of mosquitoes, malaria and other mosquito-bornediseases. Insecticides may be reserved for short-term use. Thiswill enable judicious and selective use of insecticides in solelyepidemic situations. The growing problem of resistance to insecticidesin mosquitoes as well as environmental pollution, can then besolved on a long term basis.