Megaloblastic anemia associated with erythroid hypoplasia

Megaloblastic Anemia Associated with Erythroid Hypoplasia

JOHN

F. PEZZIMENTI,

JOHN

LINDENBAUM,

New York,

New

M.D.* M.D.

York

From the Medical Services, Harlem Hospital and Columbia-Presbyterian Medical Center, and the Department of Medicine, Columbia University, College of Physicians and Surgeons, New York, New York 10032. Requests for reprints should be addressed lo Dr. John Lindenbaum. Medical Service, Harlem Hospital Center. 135th Street and Lenox Avenue, New York, New York 10037. Manuscript received June 1, 1972; revised July 21, 1972. *Present address: 30 Hospital Avenue, Danbury, Connecticut 06610.

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Four patients are described in whom moderate to marked erythroid hypoplasia of the bone marrow was associated with frank megaloblastic changes in red and white cell precursors. Two of these patients were folate deficieut, secondary to alcoholism or pregnancy, and two had pernicious anemia. Serum lactic dehydrogenase (LDH) concentrations were lower than would be anticipated in patients with severe megaloblastic anemias. The erythroid marrow converted from hypoplastic to hyperplastic during folic acid or vitamin 812 therapy in each case although the reticulocyte responses were delayed or suboptimal. Each of the patients appeared malnourished, two had impaired renal function, and one was hypothyroid. In the marrow of the pregnant patient, a combination of granulocytic hyperplasia and erythroid hypoplasia appeared to be present. A review of the bone marrows of 125 consecutive patients with megaloblastic anemias seen at two hospitals revealed no other cases with erythroid hypoplasia. Few similar cases have been previously documented in the literature. The cause of the transient erythroid aplasia was not determined. Intense erythroid hyperplasia of the bone marrow is recognized as a characteristic feature of the megaloblastic anemias and is most striking in severely anemic patients [l-3]. The increase in marrow erythroid precursors in patients with vitamin B12 or folate deficiency is not associated with reticulocytosis in the peripheral blood and has been attributed to ineffective erythropoiesis; the ultimate fate of the majority of the nucleated red cells is thought to be intramedullary destruction [3,4]. We have recently encountered four severely anemic patients who initially presented problems in differential diagnosis in that frank megaloblastic changes in the bone marrow were associated with moderate to marked erythroid hypoplasia. We here present these cases in detail, record their responses to vitamin 812 and folate therapy, review the relevant previous literature and consider possible explanations for the findings observed. MATERIALS

AND METHODS

Peripheral blood smears and marrow aspirates were stained with Wright-Giemsa stains and 1,000 cell differential counts performed. Adequate marrow specimens rich in spicules were obtained from each patient. In addition, a marrow biopsy specimen was obtained before therapy in one (Case 1). Fixed smears of marrow spicules were stained with Prussian blue for evaluation of hemosiderin stores [5]. Serum folate, vitamin 812. lysozyme and lactic dehydrogenase (LDH) concentrations were measured using published methods [e-91.

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TABLE

I

Summary

of Laboratory

Data, Bone Marrow

Bone marrow cellularity M:E ratio* Megakaryocytes Iron stores

Serum folate (ng/ml) Serum vitamin BIz (pg/mI)

Serum iron (pg/lOO ml) Serum haptoglobin Serum lysozyme (pg/mI) Serum LDH (lU/ml) Serum bilirubin (mg/100 ml) Serum albumin (g/100 ml) Associated conditions

(TABLE

14 0.0 8,300 68,000 Macrocytic hypersegmented neutrophils Slight decrease 2O:l Adequate

4+ Occasional ringed sideroblasts 23.3 96 175 Decrease

4+

8 320 2.6 2.7 Alcoholism, malnutrition, urinary obstruction, liver disease, thyroxine-binding globulin deficiency

32t 359 0.6 2.4 Hypertension, malnutrition, chronic renal disease, congestive heart failure

3.2 83 Not done Absent

I)

An 83 year old white man (T.S.) was brought 1. to HHC following a syncopal episode. He had noted fatigability and exertional dyspnea for one year. He was unable to give an adequate history but admitted to the heavy use of alcohol. Physical examination revealed a blue-eyed elderly man who was afebrile, very pale, cachectic, lethargic and moderately dehydrated. Positive physical findings included multiple ecchymoses and petechiae, an enlarged prostate, generalized muscle weakness, very poor memory and absent vibratory sense throughout the lower extremities. Pancytopenia was present on admission (Table I). Other Case

HYPOPLASIA-PEZZIMENTI,

9 0.1-0.4 3,600 Decrease Macrocytic hypersegmented neutrophils Normal 21:l Adequate 2+ to 3+

Normal Range

Case4 16 0.6 3,800 32,000 Macrocytic hypersegmented neutrophils Increase 1O:l Adequate 3+

1.0

1.0 110

330 Absent 21 265 0.3 2.6 Alcoholism, malnutrition, gastrointestinal bleeding, organic brain syndrome

was 27 pg/ml

LINDENBAUM

Conditions

Case3

9 0.1 3,900 37) 000 Macrocytic hypersegmented neutrophils Slight decrease 26:l Decrease

The four patients described were admitted to the Harlem Hospital Center (HHC) or the Columbia-Presbyterian Medical Center (CPMC) during a 13 month period in 1970-1971. To determine whether other patients with megaloblastic anemia associated with erythroid hypoplasia were encountered in the recent past at these hospitals, the records and bone marrow slides of 67 consecutive cases of megaloblastic anemia seen at CPMC and 58 cases at HHC over the past 36 months were reviewed. REPORTS

ERYTHROID

Findings and Associated

NOTE: Hematologic values are those present on admission. * 1,000 cell count. i_Serum lysozyme level after one month of vitamin BIZ therapy dysfunction).

CASE

AND

Case2

Case1 Hematocrit (%) Reticulocyte count (%) White blood cell count/mm3 Platelet count/mm3 Peripheral blood smear

ANEMIA

(persistent

447 250 Decrease 7 532 1.7 3.3 Pregnancy, malnutrition, hypothyroidism

elevation

attributed

6-30 200-1,oao 70-160

... 5-7 90-200 0.15-1.0 3.5-5.0

to chronic

...

renal

laboratory data on admission included blood urea nitrogen 55 mg/lOO ml, creatinine 1.8 mg/lOO ml, total bilirubin 2.6 mg/lOO ml, direct-reacting bilirubin 1.2 mg/lOO ml, serum glutamic oxaloacetic transaminase (SGO’T) 105 units and prothrombin time 17.7 seconds (control 12.6 seconds). A urine culture was positive for Escherichia coli. Blocking antibodies to intrinsic factor were present in the serum. Bone marrow examination on admission showed frankly megaloblastic red and white cell precursors with striking red cell hypoplasia (Figure 1A). Identical findings were present on a bone marrow biopsy specimen obtained 24 hours after admission. During the first hospital week, the patient was treated with several 1 mg injections of vitamin 8,~. 15 mg of folic acid daily, hydration and catheter drainage of the bladder. Two days after admission severe epistaxis occurred. The hematocrit was 10 per cent, white blood cell count 2,100/mm3 and platelet count 6.000/mm3. He was given transfusions to raise the hematocrit level to 15 per cent. After five days of vitamin BY2 therapy, the reticulocyte count was 0.1 per cent, the white blood cell count 15,510/mm3 and the platelet count 41,000/mm3. The reticulocyte count began to

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rise on the sixth day, reaching a peak of 8.8 per cent on the eighth day; the platelet count became normal by the seventh day, increased to 834,000 on the 12th day and subsequently returned to normal. The hematocrit level rose to 27 per cent by the 19th day. Roentgenograms of the stomach and the small intestine and a xylose tolerance test were normal. Schilling tests showed 0.1 per cent excretion in 48 hours without intrinsic factor and 11.0 per cent with intrinsic factor (normal, greater than 10 per cent). The blood urea nitrogen and creatinine levels fell to normal but rose again after removal of the Foley catheter. After two and a half months the hematocrit stabilized at 33 per cent, with 0.6 to 1.8 per cent reticulocytes and low serum iron and iron-binding capacity. A bone marrow aspirate now showed moderate normoblastic erythroid hyperplasia (Figure 1B) with increased iron stores and no ringed sideroblasts. The blood urea nitrogen level fell to 22 mg/lOO ml after reinstitution of catheter drainage. Four months after admission he underwent transurethral prostatic resection. Postoperatively an Escherichia coli septicemia developed and the patient died. At autopsy benign prostatic hypertrophy with bilateral hydronephrosis, chronic pyelonephritis and a bilateral basal pneumonia were the principal findings.

Case 2. A 76 year old black woman (L.L.) with a history of 25 years of hypertension was admitted to Harlem Hospital Center because of 10 months of orthopnea, exertional dyspnea and progressive weakness. She appeared poorly nourished, cachectic, pale and in acute respiratory distress. Mild hypertension, cardiomegaly, gallop rhythm, venous distention, bilateral pulmonary rales and moderate anasarca were present. Vibratory and position sense were normal. Hematologic values obtained on admission are shown in Table I. The blood urea nitrogen was 56 creatinine 3.2 mg/lOO ml. mg/lOO ml and serum Blocking antibodies to intrinsic factor were present in

LINDENBAUM

the serum. The patient was treated for acute pulmonary edema with diuretics and digitalis, and was given a transfusion of 3 units of packed red blood cells. Her congestive heart failure responded to management, but she remained azotemic and anemic. On the ninth hospital day the hematocrit value was 21 per cent, reticulocyte count 0.0 per cent, white blood cell count 3,630/mm3 and platelet count 41 ,000/mm3. The serum folate concentration had risen to 5.7 ng/ml but the vitamin B12 level remained low. A bone marrow aspirate at this time was megaloblastic in the white and red cell series and slightly hypocellular, with decreased red cell precursors (Table I). Therapy with 1 pg of vitamin B12 given intramuscularly daily was begun with rapid return of the white blood cell and platelet counts to normal (Figure 2). A bone marrow aspirate obtained on the fourth day of therapy showed increased cellularity and mild erythroid hyperplasia with transitional megaloblastic changes, but after two weeks the hematocrit level had fallen to 19 per cent and the reticulocyte count had only increased to 4.4 per cent. A bone marrow specimen at this time showed normoblastic erythroid hyperplasia. The daily dose of intramuscular vitamin 81s was increased to 5 pg. The reticulocyte count reached a peak of 5.7 per cent (Figure 2) on the 18th day of vitamin 812 therapy. A marrow aspirate obtained after 12 days of therapy at the 5 pg dosage level revealed marked erythroid hyperplasia with increased iron stores. The hematocrit level failed to rise during the first six weeks of vitamin Bt2 therapy, despite a further increase in daily dosage to 1 mg. The anemia was attributed to persistent azotemia and mild hemolysis of unknown cause associated with continued 3 to 6 per cent reticulocytosis, an elevated plasma hemoglobin level, decreased haptoglobins, negative Coombs’ test and normal red cell glucose-6-phosphate dehydrogenase activity. Roentgenograms of the stomach and small intestine and a serum xylose tolerance test were normal. Histamine-fast achlorhydria was present. The Schilling test was not performed due to azotemia. Figure 1. Case 1. A, representative area from bone marrow aspirate obtained on admission. Giant bands and metamyelocytes are present but there is a marked paucity of red cell precursors. Wright-Giemsa stain; original magnification X 1,000 X 1. 6, representative area from bone marrow obtained after aspirate two and a halt months of vitamin 812 therapy. Moderate normoblastic erythroid hyperplasia is present. Wright-Giemsa stain, original magnification X 1,000 x 1.

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I Platelets

ANEMIA

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ERYTHROID

HYPOPLASIA-PEZZIMENTI,

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40

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2. Case 2. Response to treatment with increasing doses of intramuscular vitamin B12.

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A 74 year old black female chronic alcoholic 3. (E.S.) was admitted to HHC in a semicomatose state. She had noted weakness, anorexia and epigastric pain of several weeks duration. She appeared thin, pale, poorly nourished, debilitated and stuporous. Positive physical findings included a smooth, depapillated tongue and evidence of mild congestive failure. Pancytopenia was present on admission (Table I). A nasogastric aspirate obtained after admission reacted positively with Topfer’s reagent and for occult blood. Tomograms of the anterior mediastinum were normal as were tests of thyroid function and roentgenograms of the gastrointestinal tract. Blocking antibodies to intrinsic factor were not present in the serum. During the first week of hospitalization the patient vomited blood on several occasions and passed numerous tarry stools. Endoscopy revealed bleeding from erosive gastritis and esophagitis, and she was given multiple blood transfusions. The reticulocyte count remained very low (0.1 to 0.4 per cent), and platelets continued to appear diminished on smear. A bone marrow aspirate on the 11 th hospital day when the hematocrit level was 28 per cent revealed classic megaloblastic changes in the white and red cell precursors with markedly decreased numbers of the latter. Spontaneous, minimal elevations of the reticulocyte count in the 1 to 2 per cent range, as well as an increase in the number of platelets to 200,000/mm3, were noted after the 18th hospital day. Oral folic acid therapy, 5 mg, was begun on the 22nd day with a subsequent rise in the reticulocyte count to 5.1 per cent a week later. The hematocrit level remained in the 25 to 30 per cent range until the seventh week when it began to rise, ultimately reaching 40 per cent by the time of discharge. A repeat bone marrow specimen obtained during the fifth week in the hospital showed normal numbers of normoblastic red blood cell precursors and increased iron stores. Paranoid ideation, Case

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present since admission, became less frequently expressed but at discharge the patient was thought to have a chronic organic brain syndrome. Case 4. A 29 year old white woman (A.R.) was admitted to CPMC for evaluation of an incompetent certix at approximately 33 weeks’ gestation. In 1964 she had a partial thyroidectomy for toxic diffuse nodular goiter. Her first pregnancy, two years prior to admission, had been uncomplicated, with a normal hemoglobin level at term. During the fourth month of the current pregnancy the hemoglobin level was 11.0 g/100 ml. During the latter four months of the pregnancy she had experienced nausea, frequent vomiting, glossalgia, fatigue and an 11 pound weight loss. Physical examination showed marked pallor and a lower abdominal mass consistent with a 33 week gravid uterus. Pancytopenia was present on admission (Table I). A few myelocytes and promyelocytes were seen on peripheral smear. The blood urea nitrogen was 5 mg/ 100 ml, serum protein-bound iodine 2.0 pg/lOO ml and thyroxine by column chromatography 1.2 pg/lOO ml. On the day of admission the patient was given 2 units of packed red blood cells and taken to surgery for repair of an incompetent cervix. On the following day a bone marrow aspirate (Figure 3) was megaloblastic with decreased red cell precursors. The patient was given a transfusion of 2 more units of packed red blood cells and oral folic acid. After five days the reticulocyte count rose to 4.9 per cent and the platelet count to 119,00O/mm 3. A bone marrow aspirate on the sixth day showed normoblastic erythroid hyperplasia. On the seventh day the hemoglobin level was 9.6 g/100 ml; two days later the patient went into premature labor and delivered a healthy 1,900 g infant. After an uneventful postpartum period the patient was discharged with a hemoglobin level of 10.6 g/100 ml.

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3. Case 4. Bone marrow aspirate obtained before folic acid therapy. Marked granulocytic hyperplasia with a high M:E ratio was present. WrightGiemsa stain; original magnification X 1,200 X 1. Figure

REVIEW OF OTHER CASES OF MEGALOBLASTIC ANEMIA

A review of 125 consecutive cases of megaloblastic anemia seen at the two hospitals during the three year period in which these four patients were admitted revealed no other patient with erythroid hypoplasia. Thus the incidence of red cell hypoplasia in this series of megaloblastic anemias was 4 in 129 (3.1 per cent). Serum LDH concentrations were available in the records of 30 patients with megaloblastic anemia associated with erythroid hyperplasia seen during the past 18 months and are plotted in Figure 4. The LDH values in our four patients, indicated by encircled symbols, are shown for comparison. There was a significant correlation between severity of anemia and LDH concentration in the 30 patients with classic megaloblastic anemias (r = -0.680, p CO.001). The mean LDH concentration in the 30 patients was 1,972 f 1,508 I U/ml and in our four patients 369 f 115 I U/ml (p KO.001).

LINDENBAUM

markedly decreased numbers of red cell precursors, resulting in an increase in the M: E ratio rather than the characteristic decrease usually seen. The red cell precursors that were present in each case were frankly megaloblastic. That the findings appear to reflect an actual decrease in red cell precursors rather than an unusually intense expansion of the intramedullary leukocytic compartment is supported by a number of observations. Excellent marrow specimens, rich in spicules, were obtained in each patient and the overall cellularity of the marrows (confirmed by biopsy in Case 1) was not increased in the first three patients. In addition the serum LDH concentration was inappropriately low considering the severity of the anemia in each patient (Figure 4). The inverse relationship between hematocrit level and serum LDH observed in our larger series of patients with megaloblastic anemias confirms that previously reported by several groups [lo-141. The source of the LDH elevation is most likely the cytoplasm of marrow megaloblasts, and the increase of enzyme levels with increasing severity of anemia has been attributed to the associated greater intensity of erythroid hyperplasia and intramedullary cell death [15,16]. In Cases 2 and 3 (Table I) lack of the usual elevation of the serum bilirubin level seen in megaloblastic anemias may be taken as further evidence of a decreased marrow erythroid turnover (in Case 1, the presence of alcoholic liver

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Hyperplasia of red and white cell precursors (and often megakaryocytes) is a typical feature in the bone marrow of patients with megaloblastic anemias, presumably reflecting ineffective hematopoiesis in each of the three cell series [l-3]. In the four patients we describe, active megaloblastic leukopoiesis was observed in combination with

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Serum LDH concentrations in 30 consecutively admitted patients with megaloblastic anemias associated with erythroid hyperplasia (uncircled symbols) and in the four patients reported here (circled symbols). In the latter four patients the serum LDH levels are not as high as those of control patients with comparable degrees of anemia.

Figure

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disease on admission may have been responsible for the increase in serum bilirubin). Elevations of serum lysozyme levels in patients with untreated megaloblastic anemia are thought to reflect increased granulocyte turnover [17-l 91. The serum lysozyme levels in our four patients were normal or slightly elevated, not beyond the range previously reported in patients with more typical megaloblastic states [17]. Thus the serum LDH, bilirubin and lysozyme concentrations, as well as the delayed and suboptimal reticulocyte responses to therapy, appear to provide indirect evidence consistent with the morphologic impression of a reduction in erythroid activity unusual for severely anemic patients with vitamin 812 or folic acid deficiency. Case 4 may differ somewhat from the first three in that over-all hypercellularity of the marrow was present and it was not as difficult to find erythroid precursors as in the others. The serum LDH, although inappropriately low (Figure 4), was the highest of the group. This case may represent a combination of mild erythroid hypoplasia and unusual leukocytic hyperplasia. The latter may have been an exaggeration of the tendency to granulocytic hyperplasia often associated with normal pregnancy [20]. The possibility that prior transfusion in some of our patients may have contributed to the depression in erythroid precursors should be considered. Three patients (Cases 2, 3 and 4) had received packed red blood cells which elevated the hematocrit levels to 21, 28 and 22 per cent, respectively. Mason and Leave11 [21] reported the effects on marrow morphology of massive transfusion to hematocrit levels of 46 to 51 per cent in three patients with untreated pernicious anemia. There was an increase in the M:E ratio in each patient (from 1.4:l to 2.3:1, from 0.9:1 to 1.6:1 and from 2.2:1 to 5.2: 1). Thus an elevation of the hematocrit to much higher levels than was achieved with transfusion in our cases did not appear to result in marked erythroid hypoplasia (Table I) such as was seen in our patients. A review of the literature has revealed few previously reported cases similar to ours. Levine and Hamstra [22] reported two cases of megaloblastic anemia of pregnancy in which the initial diagnosis was acute leukemia. These cases resembled our Case 4 in that the bone marrow appeared markedly hypercellular, and a combination of erythroblastopenia and granulocytic hyperplasia appeared to be present. A third case of megaloblastic anemia of pregnancy and erythroid atrophy was recently reported [23], but in this case a decreased general cellularity of the marrow was present suggestive of an aplastic anemia. Generalized mar-

December

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row hypoplasia without selective erythroblastopenia has rarely been reported in the past as a complication of macrocytic anemia of pregnancy [24] or after prolonged therapy of pernicious anemia with massive doses of folic acid [25]. Although a careful search of the literature has thus uncovered only three cases of megaloblastic anemia with red cell hypoplasia, our observation of four such cases during a three year period suggests that this complication may not be as rare as the paucity of published reports would suggest. Erythroblastopenia was a reversible event in each of our patients. Reticulocyte responses occurred during therapy with vitamin BIP or folic acid (although delayed or suboptimal), and were associated with appropriate erythroid hyperplasia on subsequent marrow examinations. It is therefore possible that the erythroid atrophy was in some way a result of vitamin B12 or folate depletion. In animals, experimental folic acid deficiency is frequently associated with panhypoplasia of the bone marrow [26,27]. In addition in some species certain doses of folic acid antagonists cause marrow hyperplasia and larger doses result in hypoplasia [28]. It is possible that our patients may have had unusually severe or prolonged vitamin depletion, but there was no evidence by history to support this view. In view of the nearly universal association of deficiency of these vitamins in man with red cell hyperactivity, other possible causes of the transient erythroblastopenia should be considered. Selective erythroid aplasia or hypoplasia in the absence of megaloblastic anemia has been described in association with a variety of conditions, including malignant tumors, autoimmune disorders, hemolytic anemias, therapy with various drugs, acute infections and as a congenital disorder or a preleukemic state [3,29-321. None of these conditions appears to have been present in any of our patients. Acute or chronic renal insufficiency has been reported as a cause of selective erythroid hypoplasia [33,34]. It is a rare complication of chronic renal disease and apparently is only seen with severe azotemia [34]. Two of our patients were mildly azotemic. Red cell aplasia has frequently been reported as an acute or chronic complication of severe protein malnutrition in childhood [35-381. Experimental protein starvation in animals results in erythroid aplasia [36]. The red cell hypoplasia of kwashiorkor has been attributed to decreased erythropoietin production, possibly a consequence of reduction in oxygen requirements [37]. Red cell hypoplasia responsive to riboflavin therapy has also been reported in malnourished patients and in experimental human riboflavin deficiency [38].

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In view of these observations, it is of interest that all our patients had lost weight and appeared malnourished on admission to the hospital. Three were described as “cachectic” (Cases 1, 2 and 3), two were long-standing chronic alcoholics (Cases 1 and 3), three had significant hypoalbuminemia, and the fourth was hypothyroid. Although erythropoietin levels were not measured in our patients, it is possible that protein depletion and reduced metabolic demands resulted in decreased erythro-

LINDENBAIJM

poietin production and resultant erythroid aplasia or that deficiencies of other nutrients (such as riboflavin) played a role in the selective depression of red cell precursors. ACKNOWLEDGMENT

We thank Mrs. Therese Bosland and Mrs. Janifer Huckaby for expert technical assistance, and Dr. Medhi Faranghi for the serum lysozyme measurements.

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