The Kinetics of Cell Proliferation in Cultures of Human Peripheral Blood

MacKinney, Archie A.; Stohlman, Frederick; Brecher, George

doi:10.1182/blood.v19.3.349.349

Cited by 267 publications

(32 citation statements)

References 2 publications

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“…It has been cautiously suggested that the transformation of the small lymphocyte is an antigen-antibody-like reaction to PHA (Mackinney et al, 1962;Elves and Wilkinson, 1963).…”

Section: Discussionmentioning

confidence: 99%

The Mitogenic Property of Phytohaemagglutinin: Studies on Human Leucocytes

Elves

Israëls

1963

Br J Haematol

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t PHYTOHAEMAGGLUTININ (PHA) has been used widely in the last 3 years for stimulating division ofperipheral leucocytes. It is extracted from beans of the genus Phaseolus (Dorset and Henley, 1916; Goddard and Mendel, 1929;Li and Osgood, 1949; Rigas and Osgood, 1955)~ and it was at first used for tissue culture as a means of separating viable leucocytes from peripheral blood. Later it was realized that it was responsible for the production of the mitoses observed in these cultures (Nowell, 1960).The changes in the cells under the influence of PHA have been well studied and the transformation of the small lymphocyte to the large blast-hke cell is now accepted (Cooper, Barkhan and Hale, 1961; Mackinney, Stohlmann and Brecher, 1962;Carstairs, 1962; Wdhson, 1962, 1963)~ but the reasons for these changes have not yet become clear.W e have performed a number of experiments which we hope will throw some light upon this problem. METHODSThe technique of culture employed in this work is essentially that described by Hungerford, Donnelly, Nowell and Beck (1959)~ variously modified to suit the demands of each experiment. Dextran (Dextraven-molecular weight about 120,000) was used to effect the separation of leucocytes from the blood. Unless otherwise stated, the PHA used was the PHA (P) prepared by Difco Ltd. (There are two forms of PHA at present obtainable; both are prepared by the method of Rigas and Osgood (1955). PHA (M) is a relatively crude bean extract which has been shown to consist of a protein fraction and a mucopolysaccharide fraction; PHA (P) has been further purified so that it consists almost entirely of the protein fraction alone.)The blood from which the leucocytes were obtained was drawn by venepuncture from normal healthy subjects. RESULTSThe first question was the possible association of the agglutinating and mitogenic properties of PHA. T o study this the following experiment was performed:PHA was lluted in 32 ml. of T.C.199 (see below), to give a concentration simdar to the fmal concentration present in the usual cell cultures (i.e. 2 vl./ml.). T.C.199 is a synthetic culture medium containing amino acids, glucose, vitamins and various coenzymes, and is made up in a balanced salt solution. The PHA medium was divided, under sterile conditions, into four equal parts, and to two of the parts was added 8 ml. of red blood cells, obtained by centrifugation, from the donors of the white cells to be cultured. After thorough mixing of the contents the containers of these two parts were left at 4' C. for 40 minutes. The red cells 406 * Medical Research Council Scholar. t Postal address: Royal Infirmary, Manchester 13.

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“…It has been cautiously suggested that the transformation of the small lymphocyte is an antigen-antibody-like reaction to PHA (Mackinney et al, 1962;Elves and Wilkinson, 1963).…”

Section: Discussionmentioning

confidence: 99%

The Mitogenic Property of Phytohaemagglutinin: Studies on Human Leucocytes

Elves

Israëls

1963

Br J Haematol

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“…Small lymphocytes from peripheral blood have been cultured, and with the aid of phytohemagglutinin (Hungerford et al, 1959;Nowell, 1960) have been shown to enlarge and undergo mitosis in the course of 24-30 hr. (Carstairs, 1962;Porter and Cooper, 1962~;Mackinney et al, 1962;Johnson and Roberts, 1963). Carstairs ( 1962) incubated blood with colloidal iron first of all, and then centrifuged to remove the phagocytic cells, including the monocytes, which had become heavier after ingesting the iron.…”

Section: The Lymphocyte-production Pathwaymentioning

confidence: 99%

Further Problems of Lymphocyte Production

Yoffey

1964

Annals of the New York Academy of Sciences

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“…A few days after phytohemagglutinin (PHA)$, or other mitogens, are added to cultures of peripheral blood lymphocytes of patients affected by chronic lymphatic leukemia (CLL) DNA synthesis and cell division occurs. However, compared to the response of normal lymphocytes, these events are impaired (Havemann & Rubin, 1968 ;MacKinney, Stohlman & Brecher, 1962;Nowell, 1960;Oppenheim, Whang & Frei, 1965;Quaglino & Cowling, 1964;Robbins, 1964;Rubin, Havemann & Dameshek, 1969;Schrek & Rabinowitz, 1963).…”

Section: Introductionmentioning

confidence: 99%

Kinetics of Phytohemagglutinin Stimulated Lymphocytes in Chronic Lymphocytic Leukemia

1976

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The DNA synthesis pattern and several kinetic parameters of in vitro PHA stimulated normal and CLL lymphocytes were determined. The DNA synthesis peak of CLL lymphocytes occurred 2-3 days later than that of normal lymphocytes. The generation time, estimated by the labeled mitoses method, was found to be 28 hr and 20 hr for CLL and normal lymphocytes respectively. This difference was mainly due to longer S and GI periods. It was also shown that both CLL and normal lymphocytes divide several times. These data were confirmed by the chromatid labeling pattern and by the halving of the grains and the double labeling techniques. By combining continuous and pulse labeling the growth fraction of CLL Iymphocytes was found to be progressively increasing, because of the recruitment of new cells in cycle, from the third day of culture. Therefore the delayed peak of DNA synthesis of CLL lymphocytes was caused by a longer cell cycle and by a longer pre-replicative phase.

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The Kinetics of Cell Proliferation in Cultures of Human Peripheral Blood

Cited by 267 publications

References 2 publications

The Mitogenic Property of Phytohaemagglutinin: Studies on Human Leucocytes

The Mitogenic Property of Phytohaemagglutinin: Studies on Human Leucocytes

Further Problems of Lymphocyte Production

Kinetics of Phytohemagglutinin Stimulated Lymphocytes in Chronic Lymphocytic Leukemia

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