Microscopic and histochemical studies on the Auer bodies in leukemic cells

(1) Seventeen cases of acute leukemia with Auer bodies have been reported.Studies were carried out on 7 cases of acute monocytic leukemia and 3 cases ofsubacute myelogenous leukemia.

(2) Histochemical studies showed the Auer bodies to be oxidase, peroxidase,and periodic acid-Schiff positive; sudanophilic, slightly metachromatic and to givepositive tests for acetal lipids and ribonucleic acid.

(3) The Auer bodies were negative for acid and alkaline phosphatase, lipase,glycogen, desoxyribonucleic acid and were non-birefringent.

(4) A change in the chemical nature of the Auer body from an acid conditionto a more neutral state was noted. This change corresponded with the changesof the normal cytoplasmic granulation of the myelocytes and monocytes duringmaturation.

(5) The effects of cellular movements, trauma, and temperature changes uponthe Auer bodies were studied.

(6) Several leukemic cells, containing Auer bodies, were studied during theprocess of mitosis.

(7) A theory as to the formation and disintegration of the Auer bodies hasbeen presented.

Note: ACKNOWLEDGMENTSThe author wishes to express his thanks to Dr. B. K. Wiseman, Dr. B. C. Houghton, Dr. R. A. Knouff,and Dr. E. R. Hayes for their help and suggestions, and to Dr. J. D. Thomas, Dr.J. F. Gamble, Dr. R. J.Rohn, Dr. H. Schiro, Mrs. Jo Myers, M.Sc., Miss Georgia Gwinner, M.T., Mrs. Susan Ragsdale, M.T.and Mrs. Jeanne Marie Willison, M.T., for their help in securing the experimental material.

     

Granulomatous lesions in the bone marrow in infectious mononucleosis; a comparison of the changes in the bone marrow in infectious mononucleosis with those in brucellosis,tuberculosis, sarcoidosis and lymphatic leukemia

1. The findings in the blood and in aspirated bone marrow in 23 cases of infectious mononucleosis have been described.

2. Unequivocal evidence of involvement of the bone marrow has been found in70 per cent of the cases.

3. Evidence of granulomatous inflammation of the marrow was found in 48 percent of the cases.

4. Epithelioid cells were found in the films of bone marrow in 48 per cent of thecases. These cells appear morphologically identical with those seen in imprints oflymph nodes from infectious mononucleosis and sarcoidosis and with the epithelioid cells seen in films of the marrow in brucellosis, sarcoidosis and tuberculosis.

5. The granulomatous lesions of infectious mononucleosis seem most similar tothose of brucellosis, but they also resemble the small granulomatous lesions ofsarcoidosis and tuberculosis.

6. Lymphocytosis of the marrow as well as of the blood was demonstrated in allcases. Evidence of formation of lymphocytes in the marrow was presented, and thealtered lymphocytes of infectious mononucleosis were found in films of the marrow.The degree of lymphocytosis of the marrow in infectious mononucleosis was shownto be less than that in lymphatic leukemia. Lymphocytosis of the marrow was notfound in brucellosis, sarcoidosis or tuberculosis. The lymphocytic reaction demonstrable in the marrow in infectious mononucleosis is believed to be of value in differential diagnosis.

Note: ACKNOWLEDGMENTSWe wish to acknowledge the generous cooperation of Dr. Ruth E. Boynton and the Staff of theStudent’s Health Service of the University of Minnesota throughout the course of this year long studyof infectious mononucleosis. We are indebted to Dr. T. Edward Bell and Dr. James Cardy for performingthe sternal aspirations. The photomicrographs were made by Mr. Henry Morris.

     

Eosinophilic leukemia; report of a case with autopsy confirmation; review of the literature

1. The data in a case of fatal leukemia with predominant eosinophilia in theperipheral blood and bone marrow are presented; we believe that this case was oneof eosinophilic leukemia.

2. During the period of observation, these eosinophils showed progesssiveimmaturity as the symptoms became more severe. Eventually this "left shift"became so marked that a large proportion of the cells were terminally myeloblastsin both the blood and the bone marrow.

3. Autopsy revealed invasion of many of the tissues and organs with thesemature and immature eosinophil granulocytes and with myeloblasts.

Note: ACKNOWLEDGMENTOur thanks for helpful criticism are particularly extended to Dr. Charles Doan and to Dr. WilliamDameshek.

     

Is leukemia a disease of the reticulo-endothelial system?

1. Evidence is given from case study of reticulo-endothelial disease supportingthe concept that monocytic leukemia is one form of reticulo-endotheliosis.

2. On the basis of varied types of cell reactions seen in monocytic leukemia, it issuggested that all forms of leukemia may be hematologic varieties of reticulo-endotheliosis.

     

Tuberculous splenomegaly with the hypersplenism syndrome; a case report

A case of tuberculous splenomegaly with leukopenia and anemia followingmiliary tuberculosis has been presented. Splenectomy was required after streptomycin failed to control the cytopenias, progressive emaciation, and splenic infection. However, following what appeared to be six weeks of marked improvement,the patient developed a fulminating tuberculous meningitis and died.

Note: ACKNOWLEDGMENTThe authors are indebted to Dr. Byrd S. Leavell for his suggestions in the preparation of this paper.

     

THE TREATMENT OF LYMPHOBLASTIC LEUKEMIA WITH CRUDE MYELOKENTRIC ACID

Eight cases of blastic lymphoid leukemia have been treated with myelokentricacid in crude form, because hypothetically in blastic lymphoid leukemia there isa deficiency of this material. The crude myelokentric acid was used because it wasmore easily obtained than partially purified material. Purification of biologicallyactive materials by methods of extraction and precipitation necessarily results in aconsiderable loss of material. Thirteen partial remissions occurred following theadministration of crude myelokentric acid. Seven of the 8 patients have died, and5 necropsies were performed.

The necropsy material adds further weight to the belief that the remissions wereinduced by the myelokentric acid in that in all 5 necropsies there was a definitealteration in the histologic morphology as contrasted with the findings in thenecropsies of the controls.

It seems inadvisable, however, to treat a large number of patients with this material because it is crude, it is relatively unavailable, and no standard dose has yetbeen devised.

Note: We wish to t hank Dr. D. L. Turner and Dr. W. A. Hause for valuable assistance in this work.

     

Autohemagglutinins and hemolysins with hemoglobinuria and acute hemolytic anemia,in an illness resembling infectious mononucleosis

A case is reported of a young man with acute hemolytic anemia and hemoglobinuria who presented: an initial blood picture consistent with infectious mononucleosis, associated with a heterophile agglutination test positive in high dilution;auto-hemagglutinins, active in the cold, at room temperature and at 37 Centigrade;a hemolysin active at 37 C. after chilling, requiring the presence of a thermolabilecomponent of serum for hemolysis; a positive Donath-Landsteiner test but no evidence of syphilis. In addition there was clubbing of the digits with certain otherroentgenologic changes in the bones; absence of any other etiologic factors knownto be concerned with such anemia; uneventful improvement under massive transfusion therapy, with apparent recovery from his hematologic disorder whenstudied two years later.

Note: ACKNOWLEDGMENTOur thanks are due to Dr. Maxwell Finland and Dr. Philip F. Wagley for suggestions in preparingthis report. We are especially indebted to Dr. T. Hale Ham for his patient and critical advice and encouragement.

     

MONOCYTIC LEUKEMIA A CASE REPORT ILLUSTRATING VARIATIONS IN THE CLINICAL PICTURE

Monocytic leukemia is an entity that has engendered a variety of divergentclinical and histologic opinions. A case is presented in which prolonged and intensive clinical and laboratory observations demonstrate the erratic course of thisdisease and illustrate the erroneous conclusions that may be derived from inconstant, momentary observations of this dynamic process. Study of the "naturalhistory" of monocytic leukemia yields observations which tend to reconcile manyof the hitherto conflicting opinions regarding this disease.

The origin and characteristics of the monocytic cell are discussed, and its probable derivation from mesenchyma is emphasized.

     

Folic acid antagonists in the treatment of acute and subacute leukemia

Forty cases of acute and subacute leukemia were treated with one or more ofvarious folic acid antagonists, usually aminopterin. Thirty-two cases were treatedfor at least one week, remissions occurring in 10. The remissions were of a temporary nature and variable in duration. As of June 1, 1949, two patients werestill alive and in good condition, seven and one-half and thirteen months, respectively, after onset of therapy.

Clinical, hematologic, and to lesser extent, marrow remissions were obtainedmost commonly in the lymphocytic type, none in the 4 monocytic cases. The subacute cases responded far better than did those of the acute, fulminating variety.

The exact mechanism of action of the folic acid antagonists is not known, although one may speculate that anti-PGA, an anti-growth factor which resemblesPGA so closely, is readily accepted by the primitive white cell with resultantcell death.

The margin between a therapeutic or effective dose and one causing a toxic reaction is a narrow one. A "toxic" reaction may in fact be an indication of a therapeutic response.

The pattern of therapy was (1) administration of the drug, usually by parenteralinjection until a toxic or a pronounced hematologic reaction occurred, at whichpoint (2) the drug was discontinued. With subsidence of the toxic reaction, (3)a maintenance dosage was then given, usually in the form of oral medications,0.5 mg. daily for adults and 0.25 mg. for children.

Transfusions and antibiotics were administered as indicated. Frequent transfusions were usually very helpful at the time of the initial reaction when anemiawas ordinarily severe. Penicillin and other antibiotics were used for supportivetherapy. Thrombocytopenia and hemorrhage were exceedingly difficult to control.

The observed remissions appeared to be directly attributable to the action ofthe drug. Although temporary, they indicate that acute leukemia is not necessarily completely irreversible. Thus, there is hope that more potent anti-growthfactors may someday be discovered which will be of value in ultimate control ofthe disease.

Note: ACKNOWLEDGMENTSWe are pleased to acknowledge the continued cooperation and generosity of various members of theStaff of Lederle Laboratories. We wish particularly to mention the help of the late Doctor YellapragadaSubbaRow, who was greatly interested in some of our first remissions obtained in adults. We also wish tothank the staffs of the U. S. Veterans Hospital at West Roxbury (Dr. Thomas Warthin, Physician-in-Chief) and of the Murphy General Hospital for their cooperation. We are greatly indebted for their helpto the medical and nursing staffs of the Boston Floating Hospital (Dr. James H. Baty, Physician-in-Chief) and of the New England Center Hospital (Dr. Samuel Proger, Physician-in-Chief).

     

Studies on the histamine content of blood,with special reference to leukemia,leukemoid reactions and leukocytoses

The literature concerning blood histamine has been reviewed, including theevidence that the incorporation of histamine within the myeloid leukocytes maybe one of the metabolic functions of myelopoiesis. Data are presented on the histamine content of blood in normal subjects and in subjects with acute leukemia,chronic myelocytic leukemia, erythremia, and with leukocytoses or leukemoidreactions of other etiologies. The changing relationships of total blood histamineto unit myeloid cell histamine in subjects receiving irradiation therapy for chronicmyelocytic leukemia are also presented.

The data indicate that total blood histamine in chronic myelocytic leukemiais usually very high in comparison to normal, that the values in erythremia showa definite but less marked tendency in the same direction, while the values inleukocytoses of other etiology are, with few exceptions, normal or low. The bloodhistamine per million myeloid cells is, on the average, about twice normal in subjects with chronic myelocytic leukemia, within the normal range or a little lowin erythremia, and very low on the average in the group of leukocytoses studied.The suggestion is tentatively made that an aberration in this metabolic process(histamine incorporation within myeloid leukocytes) may be an inherent component of chronic myelocytic leukemia, whereas in physiologic leukocytoses andleukemoid reactions this is not the case.

Note: ACKNOWLEDGMENTSThe authors wish to express their grateful appreciation to Mr. John Fuschetti for valuable technicalassistance in performing the histamine determinations. The cooperation and aid of Dr. Gurth E. Carpenter, Dr. Henry H. Henstell, Dr. Myron Prinzmetal and of many members of the staff of WadsworthGeneral Hospital, Veterans Administration, Los Angeles were invaluable in obtaining blood specimensfrom suitable clinical material, and to them the authors wish to express their sincere thanks.

     

"Monocytic" Cells of Normal Blood, Schilling and Naegeli Leukemia, and Leukemic Reticuloendotheliosis in Slide Chambers

1. The study of living blood cells in slide chambers by phase microscopyand cinemicrography adds useful morphologic, developmental and physiologicdata complementary to other methods for studying blood cells.

2. Normal monocytes were characterized by flattening in fresh serum withminimal ameboid motility, by ectoplasmic veils waving in the medium, andby transformation to fully developed macrophages in 5-7 days.

3. Cells from four patients with acute monocytic leukemia (Schilling type)showed only slight differences in morphology, development and function fromnormal monocytes in the slide chamber.

4. Monocytoid cells from four patients with myelomonocytic leukemia(Naegeli type), in the slide chamber, resembled those from two patients withleukemic reticuloendotheliosis in morphology, development and function, butdiffered from normal monocytes and cells of acute monocytic (Schilling)leukemia. The monocytoid cells were labeled "reticulum" cells with the implication that they were related to the primitive hematopoietic reticulum cell.

5. The "reticulum" cell in the slide chamber differed from the monocyte inhaving a coarse nuclear chromatin pattern with clumping of chromatin,larger numbers of varying sized cytoplasmic granules, less ectoplasm, development of characteristic star-shaped macrophage forms without ectoplasmicveils, and in exhibiting a considerable ameboid motility as opposed to thesluggish veil waving of the monocyte.

6. Speculation as to the significance of the "reticulum" cell has been presented with the suggestion that this cell precedes the blast cell in hematopoiesis. In Naegeli leukemia, unknown factors may cause alterations in the typeof cell released from the marrow. There may also be shifts in emphasis betweenmyeloblast and pre-myeloblast proliferation which become reflected in thecells appearing in the peripheral blood.

Submitted on March 19, 1962 Accepted on May 28, 1962     

Studies on Charcot-Leyden crystals

1. Synthetic detergents of the anionic, cationic and nonionic types result inthe rapid and constant formation of Charcot-Leyden crystals from eosinophils.

2. Charcot-Leyden crystals have a negative crystalline birefringence and formpenetration twins.

3. The changes taking place in the eosinophil in the formation of Charcot-Leyden crystals under the influence of wetting agents, utilizing phase and polarizing microscopy, are described.

4. In the formation of Charcot-Leyden crystals with wetting agents, the nucleusof the eosinophil lyses with no appreciable effect on the granules.

5. In the formation of Charcot-Leyden crystals with a wetting agent, there isno change in the lipoid cortex of the eosinophil as demonstrated by staining withsudan black B.

6. Charcot-Leyden crystals undergo changes on standing that affect theirsolubilities.

7. The staining reactions and solubilities of Charcot-Leyden crystals aredescribed.

8. Oxyhemoglobin crystals constantly form from red cells on exposure to Aerosol MA; on two occasions, tyrosine crystals formed from the blood of a patientwith chronic myelogenous leukemia.

9. Evidence is offered that Charcot-Leyden crystals are crystalline proteinsderived only from the nucleus of the eosinophil.

Note: ACKNOWLEDGMENTSI wish to thank Dr. William B. Ober, Dr. J. J. Englefried, and Lt. E. E. Ozburn, MSC, USN, forassistance in this work.

     

STUDIES ON THE PHYSIOLOGY OF THE WHITE BLOOD CELL: THE GLYCOGEN CONTENT OF LEUKOCYTES IN LEUKEMIA AND POLYCYTHEMIA

The technic of determining glycogen in isolated white blood cells was appliedto the study of the different types of leukemia and of polycythemia, in order toobtain information on the physiology of the white blood cell. From this study itis concluded that the granulated leukocyte is the only carrier of glycogen in wholeblood. The "reducing substances" in lymphocytes and blast cells are not consideredas true glycogen.

The glycogen content of wet white blood cells in the rabbit amounts to about1 per cent. In the human being a range of from 0.17 to 0.67 per cent was calculated.In disease higher percentages occur, in polycythemia up to 1.64 per cent and inglycogen storage disease up to 3.05 per cent.

The glycogen concentration of normal white blood cells is within the same rangeas that of the striated muscle.

Note: I acknowledge with gratitude my indebtedness to Dr. William Dameshek for giving me the opportunity of analyzing the blood of some of the patients studied. Miss M. H. Campbell, Miss H. A. Clark,and Miss L. M. Garofalo have aided in carrying out many of the blood counts.

     

Refined liver extract in tropical macrocytic anemia

1. A series of 45 cases of T.M.A. treated with "refined" liver extract is reported.

2. "Refined" liver extract was found to be effective in 39 cases.

3. It was found that 2 or 3 ml. of refined liver extract (Examen N.P.) was sufficient to produce an optimum response.

4. As judged from therapeutic observations, it is suggested that in the majorityof cases of T.M.A. the deficiency is similar to that in Addisonian pernicious anemia, though the mode of production of the deficiency may not be the same.

Note: ACKNOWLEDGMENTOur thanks are due to Dr. R. Row, Hon. Director, P.G. Singhance Hindu Hospital, Bombay, whereall the cases were treated, for permission to publish these reports, and the firms concerned for the generousgift of liver extracts.

     

Application of Hematopoietic Cell Grafts to the Treatment of Leukemias and Allied Diseases. A Critical Review

A brief summary has been given of the present knowledge concerningtransfusion of hematopoietic cells in laboratory animals and also in man.

A survey has been made of the possible application of grafts of hematopoietic cells in the "preventive" treatment of experimental leukemia withemphasis on irradiation-induced and spontaneous leukemias.

Numerous articles have been devoted to describing attempts at curingexperimental leukemias, especially the transplantable varieties, by irradiationfollowed by transfusion of hematopoietic cells. The results vary from oneleukemia to the other. Most authors admit the rarity of eradicating leukemiaby irradiation with lethal or supralethal doses followed by transfusion of isologous hematopoietic cells. It has been shown that eradication can eventually be obtained when irradiation is followed by transfusion of homologoushematopoietic cells. This, most probably, is due to an immune reaction ofthose cells against the leukemic cells. Furthermore, even when the leukemiacan be eradicated, the animals usually die from the secondary syndrome.

A detailed analysis is given of the reported literature dealing with attemptsat treating human leukemia by irradiation followed by transfusion of autologous, isologous or homologous bone marrow.

Submitted on January 27, 1959 Accepted on January 11, 1960     

THE ACTION OF PTEROYL GLUTAMIC ACID AND NATURAL SOURCES OF FOLIC ACID ON BLOOD DYSCRASIAS INDUCED BY SULFONAMIDE DRUGS

Sulfanilamide, sulfathiazole, and sulfadiazine have been fed at 1 per cent levelsin highly purified diets and their effect on growth, mortality, and blood dyscrasiascompared with that of sulfasuxidine.

The soluble drugs produce conditions which are similar to those produced bysulfasuxidine. The growth depression is alleviated in large measure in the case ofsulfanilamide and to a lesser extent for sulfathiazole and sulfadiazine by folic acid.liver extract powder, and dried yeast extract as well as by para-aminobenzoic acid,

The blood dyscrasias due to sulfanilamide, sulfathiazole, and sulfadiazine aresevere leukopenia, granulocytopenia, and mild-to-severe anemia. These are uniformly prevented or the severity greatly curtailed by feeding folic acid, liver extract powder, or dried yeast extract. PABA has a lesser effect in the amounts fed.

Liver extract powder seems to have a beneficial effect on growth and mortalitywhich is not shown by the other supplements. Both free and conjugated folic acid(as yeast extract and in liver extract powder 1:20) are active in combating thedyscrasias.

Evidence from in vitro experiments with Str. haemolyticus (B Lancefield) indicates that neither folic acid, liver extract powder, nor dried yeast extract in ratiosto sulfonamide which are effective in preventing the blood dyscrasias will inhibitor block the bacteriostatic action of the sulfonamide drugs in vitro.

It is suggested that the action of folic acid, liver powder, and yeast extract is notwholly explained by alleviating a folic acid deficiency caused by intestinal bacteriostasis due to the drugs, but by an increased demand of the animals for folicacid in the presence of certain sulfonamides.

Note: We are indebted to Harold Buskirk, Alice Bergdahl, Hallie Ferguson, E. M. Stapert, F. La Plante,and Evon Saggio for valuable assistance in carrying out the experimental work reported here.

     

A review of lesions in the optic fundus in various diseases of the blood

1. The frequency of retinopathy as seen on an active hematologic servicein a two year period is reported. The report is based on the findings in 50patients who had retinopathy associated with a blood disorder.

2. The occurrence of retinopathy in the course of leukemia did not significantly alter the mortality or duration of illness.

3. Anemia, thrombocytopenia, septicemia and immaturity of the leukocytes appear to be the most important factors in the production of retinopathy in hematologic diseases.

4. The various retinal lesions are illustrated in photographs. Perivascularsheathing was seen only in chronic granulocytic leukemia. Otherwise, retinopathy was not specific for any particular blood disorder.

Submitted on August 18, 1958 Accepted on October 12, 1958     

SERUM PROTEIN CHANGES IN MYELOGENOUS AND LYMPHOCYTIC LEUKEMIAS AND HODGKIN'S DISEASE

1. Using a methyl alcohol fractionation technic, the albumin, globulin, andtotal protein levels were determined in a series of normal adults and compared withcases of myelogenous and lymphocytic leukemias and Hodgkin’s disease.

2. Statistically significant decreases in albumin and increases in globulin werefound in the cases of Hodgkin’s disease and myelogenous leukemia, but withoutsignificant changes in total protein. Globulin levels above the highest normal valuewere found in 23 per cent of the former and 33 per cent of the latter group.

3. No apparent relationship was noted between the levels of the serum proteinfractions and (1) the hemoglobin level, (2) the erythrocyte count, (3) the peripheral white blood cell picture, and (4) the bone marrow smears.

Note: The authors wish to express their appreciation to Professor Ovid O. Meyer, Department of Medicine,for making available the clinical material in this study and for valuable suggestions. The authors arealso indebted to Professor J. A. E. Eyster, Department of Physiology, for suggestions as to statisticaltreatment of the data.

     

Chronic neutropenia; favorable response following splenectomy

The case of a patient with chronic neutropenia without splenomegaly, but responding favorably to splenectomy is reported. The surgical procedure appeared tobe indicated by the following: (1) exclusion of the extrinsic causes of neutropenia;(2) failure of response to the agents commonly employed to stimulate granulopoiesis; (3) demonstration of granulopoiesis in the sternal marrow; (4) increasein the circulating neutrophils following the parenteral administration of epinephrine; (5) the presence of coexisting diabetes with the potential hazard ofinfection.

The implication of the spleen as the main factor in the causation of the neutropenia in this case seems well established, although the specific mechanism is notapparent. There was no evidence of abnormal phagocytosis in the microscopicexamination of the spleen.

Note: ACKNOWLEDGMENTWe wish to express appreciation to Captain Earl F. Evans, Medical Corps, U. S. Navy, for his assistance in the management of this case.

     

Malic and Lactic Dehydrogenase Isozymes of Normal and Leukemic Leukocytes Separated on Glass Bead Columns

Five LDH and two MDH isozyme bands were obtained with acrylamidegel electrophoresis of leukocyte extracts. Normal lymphocytes showed a hightotal H-LDH (heart type) activity (67 per cent) with 25 per cent in LDH-1and only 3.5 per cent in LDH-5. Lymphocytes from chronic lymphatic leukemia(CLL) and lymphosarcoma leukemia (LSA-LL) had less LDH-1, and moreLDH-3 and LDH-4, than normal lymphocytes. The H-LDH fell to 60.5 per centin CLL and 56 per cent in LSA-LL. PMN leukocytes had low H-LDH activity(38.8 per cent) with 3.3 per cent in LDH-1 and 25.8 per cent in LDH-5. Inmyelogenous leukemia, myeloblasts had the most LDH-1 and H-LDH, whilemature PMN had the least. PMN leukocytes isolated from CLL, LSA-LL, andmyelogenous leukemia had LDH patterns like the normal. Monocytes fromacute monocytic leukemia were low in LDH-1 and LDH-5, but had a high totalenzyme content. They evidently were rich in LDH-2, 3, and 4.

Lymphocytes had less MDH-1 (60 per cent) than PMN leukocytes (78 percent). In CLL, lymphocyte MDH-2 increased. In myelogenous leukemia,myeloblasts had the most MDH-2 and mature PMN the least. Monocytesfrom monocytic leukemia contained a little more MDH-2 than PMN leukocytes.

In general, white cell immaturity and/or ability to divide was associatedwith high levels of LDH-1, total H-LDH, and MDH-2.

Submitted on May 5, 1966 Accepted on June 26, 1966