Modulation of retinoic acid-induced differentiation of human leukemia (HL-60) cells by serum factors and sphinganine.

The human cell line HL-60 was used to investigate the role of protein kinase C in the regulation of retinoic acid-induced maturation of promyelocytic leukemia cells by growth and differentiation factors found in serum. Cells grown in serum-containing medium differentiated less than cells in serum-free medium due to several factors, including albumin binding of retinoic acid. Addition of an inhibitor (sphinganine) of protein kinase C, an enzyme that participates in cellular responses to many serum factors, facilitated the retinoic acid-induced differentiation. Cells treated with both retinoic acid and sphinganine produced more superoxide when stimulated by formylmethionylleucylphenylalanine; hence, this combination generated a more functional population of cells. The ability of sphinganine to promote retinoic acid-induced differentiation suggests that retinoic acid therapy might be improved by the concurrent use of a modulator of protein kinase C activity.

Differential effects of long-chain (sphingoid) bases on the monocytic differentiation of human leukemia (HL-60) cells induced by phorbol esters, 1 alpha, 25-dihydroxyvitamin D3, or ganglioside GM3.

Conditions were developed to prolong the ability of sphinganine, a potent inhibitor of protein kinase C, to block the phorbol ester-induced adherence of HL-60 cells beyond 24 h. The loss of inhibition after this time seen previously (A.H. Merrill, Jr., A.M. Sereni, V.L. Stevens, Y.A. Hannun, R.M. Bell, and J.M. Kinkade, Jr., J. Biol. Chem., 261: 12610-12615, 1986), which appeared to be due to metabolism of this long-chain base, was overcome by supplying sphinganine daily. After 4 days, phorbol myristate acetate-induced adherence was inhibited approximately 50% by sphinganine. Sphinganine significantly decreased the expression of nonspecific esterase induced by phorbol myristate acetate in the nonadherent cells, indicating that other aspects of maturation besides adherence were blocked. The effects of daily sphinganine treatments on the monocytic differentiation induced by 1 alpha-25-dihydroxyvitamin D3 or ganglioside GM3 were also investigated. The increases in nonspecific esterase expression, nitroblue tetrazolium reduction, and morphological maturation caused by either agent were unaffected by the long-chain base. In addition, the changes in several cell surface antigens caused by 1 alpha,25-dihydroxyvitamin D3 were unaltered by sphinganine. Although phorbol esters, 1 alpha,25-dihydroxyvitamin D3, and ganglioside GM3 all induce the maturation of HL-60 cells along the monocytic lineage, the finding that sphinganine only affected the differentiation initiated by phorbol esters, in which protein kinase C clearly is a major regulator, suggests that this enzyme does not play a major role in these other pathways of differentiation.

Assessment of input-output properties and control of neuroprosthetic hand grasp.

Three tests have been developed to evaluate rapidly and quantitatively the input-output properties and patient control of neuroprosthetic hand grasp. Each test utilizes a visual pursuit tracking task during which the subject controls the grasp force and grasp opening (position) of the hand. The first test characterizes the static input-output properties of the hand grasp, where the input is a slowly changing patient generated command signal and the outputs are grasp force and grasp opening. Nonlinearities and inappropriate slopes have been documented in these relationships, and in some instances the need for system returning has been indicated. For each subject larger grasp forces were produced when grasping larger objects, and for some subjects the shapes of the relationships also varied with object size. The second test quantifies the ability of the subject to control the hand grasp outputs while tracking steps and ramps. Neuroprosthesis users had rms errors two to three times larger when tracking steps versus ramps, and had rms errors four to five times larger than normals when tracking ramps. The third test provides an estimate of the frequency response of the hand grasp system dynamics, from input and output data collected during a random tracking task. Transfer functions were estimated by spectral analysis after removal of the static input-output nonlinearities measured in the first test. The dynamics had low-pass filter characteristics with 3 dB cutoff frequencies from 1.0 to 1.4 Hz. The tests developed in this study provide a rapid evaluation of both the system and the user. They provide information to 1) help interpret subject performance of functional tasks, 2) evaluate the efficacy of system features such as closed-loop control, and 3) screen the neuroprosthesis to indicate the need for retuning.