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Vitamin E deficiency in Type II congenital dyserythropoietic anemia
Volume 84 Number 6
a peak response from baseline of 1.41 + 0.41 N moles per milligram creatine (M _+ range). Urinary cyclic AMP response to PTH was equally reduced regardless of age, birth weight, or presence of hypocalcemia. Phosphaturic response to PTH was difficult to demonstrate in premature infants and on the first day of life. A definite serum calcemic response occurred with a rise of 1.22 + 0.70 rag. per 100 ml. (M-• range) 12 hours postinfusion. Three premature infants demonstrated a 20-fold greater response to glucagon (30/~g per kilogram S.C. than to the PTH (5 U per kilogram per hour) infusion, with peak delta responses of 24.4 • 3.9 vs. 1.30 • 0.40 N moles per milligram creatine (M • range) respectively. Our studies suggest that in prematures the cyclic AMP response of the proximal renal tubule to PTH is substantially less developed than is that of the liver to glucagon. Cyclic AMP excretion does not reflect the hypoparathyroid state of premature infants. The reduced renal response to PTH may contribute to hypocalcemia but other factors must be involved. COMMENT: The relationship between the renal cyclic AMP response to PTH and the phosphaturic response was not studied. Urinary cyclic AMP excretion of hypocalcemic newborn infants did not differ from that of normocalcemic infants, suggesting that the hypocalcemia was not caused by hypoparathyroidism or that cyclic AMP excretion in premature infants is not regulated by parathormone.
19. Thymus transplant in lymphopenic immune deficiency (Nezelof's syndrome) D. G. Tubergen, Ann Arbor, Mich. A 5-year-old girl presented with a history of autoimmune hemolytic anemia, thrombocytopenic purpura, juvenile rheumatoid arthritis, and chronic respiratory infection. A brother died at age 18 months with anemia, thrombocytopenia and progressive pulmonary disease. He was lymphopenic and at autopsy was found to have a hypoplastic thymus with absent Hassall's corpuscles. Laboratory studies on our patient demonstrated normal immunoglobulin levels with antibody activity against bacterial and viral antigens. Lymphopenia, 400 to 1,500/ cmm, was persistent and there was no reaction to delayed hypersensitivity skin tests. In vitro response to phytohemagglutinin (PHA) was very low. Challenge with dinitrochlorobenzene two weeks after sensitization was associated with erythema and induration. No circulating lymphotoxins could be demonstrated. Transfer fa~ztor was administered once with no change in clinical condition or in skin test reactivity over a 3 month period. Transplantation of thymic tissue from a 17-week-old fetus was associated with a rise in in vitro lymphocyte response to PHA to normal levels in two days. There was no increase in circulating lymphocyte count. The PHA response was transient with decreased responsiveness evident by two weeks after transplant; by eight weeks the PHA response was only twice background. The transplant had no apparent effect on her clinical course. There was no evidence of graft vs. host disease. The rapid change in PHA responsiveness following transplantation is most compatible with a humoral effect of the thymus gland, The transient nature of the response suggests graft failure which may have had an immunologic basis. COMMENT: Dr. Lauren Pachman commented on the importance of this patient in confirming that transfer factor did not
Abstracts
915
work in thymic dysplasia and in documenting a return of PHA responsiveness following thymic transplant.
20. Correction of cyclic neutropenia L. M. Paehman, Chicago, IlL, D. W. Golde,* San Francisco, Calif., R. Barron,* and A. D. Schwartz, i2hicago, Ill. This severely neutropenic child had, since birth, cyclic episodes of aphthous ulcers, otitis media, cervical adenitis, and pneumonia. Serial total absolute granulocyte (PMN) counts documented a cyclic neutropenia--range 0 to 1,500 PMNs--occurring at 3 week intervals. Assays of PMN, thymic cell, complement, and antibody function were normal. An examination of bone marrow at the cycle's nadir demonstrated a maturation arrest at the myelocyte stage, in vitro bone marrow culture with liquid and semisolid media documented a maturation arrest at the myelocyte stage o f PMNs, with a marked increase in proliferative capacity. The patient received epinephrine, typhoid vaccine, and infusions of normal plasma without increase in PMN number. Normal donors were given typhoid vaccine and plasmaphoresed 60 minutes later. Their response to the typhoid vaccine was observed: local inflammation, rise in PMNs, and increase in antibody titer. On two separate occasions, post-typhoid plasma (10 c.c. per kilo) was given intravenously to the patient 10 days before his next anticipated PMN peak. On both occasions there was an increase in absolute PMNs within 24 hours, lasting 8 to 10 days. Sequential bone marrow studies showed progressive depletion in myelocytes with a shift to metamyelocytes and mature PMNs, accompanied by clinical improvement. The patient appears to have a defect in PMN maturation which is transiently corrected following infusion of plasma obtained from donors responsive to typhoid vaccine. COMMENT: Although the patient is able to produce the PMNstimulating factor, the conditions which regulate the production or release of the "factor" have not been defined. The physiologic characteristics and chemical identification of the "factor" are presently under investigation. It is not known which portion of the plasma contains the "factor"; it is not present in significant amounts in fresh frozen plasma.
21. VitaminE deficiency in Type H congenital dyserythropoietic anemia S. O.'Regan,* D. K. Melhorn,* A. J. Newman,* and R. C. Graham,* Cleveland, Ohio (Introduced by S. Gross, Cleveland, Ohio) An 8-year-old girl with Type II congenital dyserythropoietic anemia, characterized by morphologic abnormalities of erythroid precursors, marked nuclear and cytoplasmic membrane abnormalities shown by electron microscopy, immunologic alterations, hyperbilirubinemia, and chronic anemia, was found to be vitamin E deficient. Nutritional history and vitamin E absorption studies indicated that neither dietary lack nor intestinal malabsorption was the cause of the deficiency. Striking changes in the patient's hematologic status following "administration of vitamin E included rise in hemoglobin, decrease in bilirubin and reticulocyte count, and a marked increase in red cell survival. Erythrocyte phospholipids, decreased
916
Abstracts
The Journal of Pediatrics June 1974
while the patient was vitamin E deficient, returned toward normal levels during therapy. The most notable increase was seen in the phosphatidyl ethanolamine fraction. However, hematologic improvement was not complete, immunologic abnormalities persisted, and morphologic aberrations in erythrocyte precursors actually increased during vitamin E therapy. It is therefore concluded that vitamin E deficiency played a role in the production of the child's hematologic disorder, and may have been a result of the increased utilization of the vitamin in stabilization of defective cellular membranes.
cies of children there is demonstrable anti-tumor cell activity by lymphocytes which is countered by a patient serum factor capable of blocking it. COMMENT: It was asked if blocking antibodies were present; one serum was found to possess blocking antibody. In response to the question concerning the usefulness of this test to monitor therapy, it was aswered that follow-up was not yet of sufficient duration; several patients in the group had died. The question was raised whether 5tCr release from the target cells was related to toxicity; Dr. Ertel answered that with increased cell destruction there was increased 51Cr release. Dr. Howard asked if this reaction was specific; did lymphocytes from normal people kill tumor cells as well? Dr. Ertel replied that normal cells were capable of cytotoxicity but at a lower level.
22. L ymphocyte-mediated cytotoxicity in children's tumors I. J. Ertel, Columbus, Ohio
23. Hypothalamic-pituitary regulation in familial goiter
Lymphocyte-mediated cytotoxic effect upon tumor cells was studied in children with neoplastic diseases. The assay tumor cells were cultured from operative specimens of neuroblastomas, Wilms" tumors, and rhabdomyosarcomas by an in vitro technique which produced monolayers. The target cells were then labeled with radioactive chromium (51Cr). Lymphocytes isolated from the peripheral blood of children with previously diagnosed malignancies were tested for cytotoxic activity against these tagged tumor cells. The lympbocytes were first purified by the Ficoll-Hypaque density gradient separation method. Cytotoxicity was quantitated by measuring the amount of 51Cr released from the lysis of tagged tumor cells after incubation with different concentrations of lymphocytes. The isotope assays were consistently repeatable and without subjective error. The activity of the lymphocytes from 20 children was compared to the anti-tumor lymphocyte activity of their own parents and that of unrelated normal controls. The patients' lymphocytes showed greater cytotoxic effects upon the tumor cells than did those of their parents or normal controls. Lymphocytes from tumor-bearing children showed the greatest cytotoxic effects upon the tumor cells and that effect was proportionate to the concentration of lymphocytes present. To assess whether serum factors influenced lymphocyte antitumor activity, sera from patients and controls were introduced into the test system. When serum from tumor-bearing patients was added for preincubation of the tumor cells, the cytotoxic activity of lymphocytes decreased. It appears that in the malignan-
Amelia V. Agustin,* Anita Perrieelli,* and Thomas P. Foley, Jr., Pittsburgh, Pa. Neuroendocrine regulation of thyrotropin (TSH) and prolactin (PR) release was studied in two patients with familial goiter secondary to a defect of thyroxin synthesis, with the onset of goiter in early childhood. Patient A. S. was hypothyroid after two months off thyroid therapy; Patient K. A. is euthyroid on no medications (Table I). TSH and PR levels were measured by radioimmunoassay. Serum was obtained q 30' for 180 minutes after L-dopa (500 mg. p.o.) and q 15' x 4, q 30' x 4 to 180 min. after IV infusion of TRH (7/zg per kilogram). Nocturnal samples were collected q 30' from 1130 to 0730, then q 15' to 0830. The results are summarized as shown in Table ii. During nocturnal sampling of Patient K. A. between 0530 and 0830 the mean TSH decreased by 33 per cent of the mean TSH between 1130 and 0500. SUMMARY: 1. Dopaminergic stimulation will decrease the elevated TSH levels in patients with familial goiter; the decrease is greater in the euthyroid patient. 2. TSH responses to TRH are greater in the hypothyroid patient. 3. Normal PR levels are suppressed after L-dopa and stimulated after TRH. COMMENT: Dr. Pena asked why an increase in T 4 w a s noted. Dr. Agustin suggested that this might be related to iodine deficiency.
23. Table I
Pt.
CA I T41I PBI I
A.S. K.A.
7.6 15.1
TSH
RAIt31uptake
pKCIO 4
probabledefect
I
1.7 4.8
5.5 9.7
31.3 38.5
6 hr: 6 hr:
74% 78%
24 hr: 24 hr:
67% + dischg. Organification 65% -- dischg. Defective thyroglobulin
23. Table II
Patient A. S.
Test L-dopa TRH L-dopa TRH
TSH (p,U/ml.) TSH (/zU/ml.) PR (ng.lmL) PR (ng./ml.)
Decrease: Increase: Decrease: Increase:
38.5to 40.9 to 4.6 to 5.0to
11.8 297 (2.5 35.0
Patient K. A. atl80' at 45' at 90' at 30'
31.3to 26.8 to 7.4 to .7.9to
<2.5 166 (2.5 47.5
atlS0min. at 45 rain. at 90 rain. at 30min.
a peak response from baseline of 1.41 + 0.41 N moles per milligram creatine (M _+ range). Urinary cyclic AMP response to PTH was equally reduced regardless of age, birth weight, or presence of hypocalcemia. Phosphaturic response to PTH was difficult to demonstrate in premature infants and on the first day of life. A definite serum calcemic response occurred with a rise of 1.22 + 0.70 rag. per 100 ml. (M-• range) 12 hours postinfusion. Three premature infants demonstrated a 20-fold greater response to glucagon (30/~g per kilogram S.C. than to the PTH (5 U per kilogram per hour) infusion, with peak delta responses of 24.4 • 3.9 vs. 1.30 • 0.40 N moles per milligram creatine (M • range) respectively. Our studies suggest that in prematures the cyclic AMP response of the proximal renal tubule to PTH is substantially less developed than is that of the liver to glucagon. Cyclic AMP excretion does not reflect the hypoparathyroid state of premature infants. The reduced renal response to PTH may contribute to hypocalcemia but other factors must be involved. COMMENT: The relationship between the renal cyclic AMP response to PTH and the phosphaturic response was not studied. Urinary cyclic AMP excretion of hypocalcemic newborn infants did not differ from that of normocalcemic infants, suggesting that the hypocalcemia was not caused by hypoparathyroidism or that cyclic AMP excretion in premature infants is not regulated by parathormone.
19. Thymus transplant in lymphopenic immune deficiency (Nezelof's syndrome) D. G. Tubergen, Ann Arbor, Mich. A 5-year-old girl presented with a history of autoimmune hemolytic anemia, thrombocytopenic purpura, juvenile rheumatoid arthritis, and chronic respiratory infection. A brother died at age 18 months with anemia, thrombocytopenia and progressive pulmonary disease. He was lymphopenic and at autopsy was found to have a hypoplastic thymus with absent Hassall's corpuscles. Laboratory studies on our patient demonstrated normal immunoglobulin levels with antibody activity against bacterial and viral antigens. Lymphopenia, 400 to 1,500/ cmm, was persistent and there was no reaction to delayed hypersensitivity skin tests. In vitro response to phytohemagglutinin (PHA) was very low. Challenge with dinitrochlorobenzene two weeks after sensitization was associated with erythema and induration. No circulating lymphotoxins could be demonstrated. Transfer fa~ztor was administered once with no change in clinical condition or in skin test reactivity over a 3 month period. Transplantation of thymic tissue from a 17-week-old fetus was associated with a rise in in vitro lymphocyte response to PHA to normal levels in two days. There was no increase in circulating lymphocyte count. The PHA response was transient with decreased responsiveness evident by two weeks after transplant; by eight weeks the PHA response was only twice background. The transplant had no apparent effect on her clinical course. There was no evidence of graft vs. host disease. The rapid change in PHA responsiveness following transplantation is most compatible with a humoral effect of the thymus gland, The transient nature of the response suggests graft failure which may have had an immunologic basis. COMMENT: Dr. Lauren Pachman commented on the importance of this patient in confirming that transfer factor did not
Abstracts
915
work in thymic dysplasia and in documenting a return of PHA responsiveness following thymic transplant.
20. Correction of cyclic neutropenia L. M. Paehman, Chicago, IlL, D. W. Golde,* San Francisco, Calif., R. Barron,* and A. D. Schwartz, i2hicago, Ill. This severely neutropenic child had, since birth, cyclic episodes of aphthous ulcers, otitis media, cervical adenitis, and pneumonia. Serial total absolute granulocyte (PMN) counts documented a cyclic neutropenia--range 0 to 1,500 PMNs--occurring at 3 week intervals. Assays of PMN, thymic cell, complement, and antibody function were normal. An examination of bone marrow at the cycle's nadir demonstrated a maturation arrest at the myelocyte stage, in vitro bone marrow culture with liquid and semisolid media documented a maturation arrest at the myelocyte stage o f PMNs, with a marked increase in proliferative capacity. The patient received epinephrine, typhoid vaccine, and infusions of normal plasma without increase in PMN number. Normal donors were given typhoid vaccine and plasmaphoresed 60 minutes later. Their response to the typhoid vaccine was observed: local inflammation, rise in PMNs, and increase in antibody titer. On two separate occasions, post-typhoid plasma (10 c.c. per kilo) was given intravenously to the patient 10 days before his next anticipated PMN peak. On both occasions there was an increase in absolute PMNs within 24 hours, lasting 8 to 10 days. Sequential bone marrow studies showed progressive depletion in myelocytes with a shift to metamyelocytes and mature PMNs, accompanied by clinical improvement. The patient appears to have a defect in PMN maturation which is transiently corrected following infusion of plasma obtained from donors responsive to typhoid vaccine. COMMENT: Although the patient is able to produce the PMNstimulating factor, the conditions which regulate the production or release of the "factor" have not been defined. The physiologic characteristics and chemical identification of the "factor" are presently under investigation. It is not known which portion of the plasma contains the "factor"; it is not present in significant amounts in fresh frozen plasma.
21. VitaminE deficiency in Type H congenital dyserythropoietic anemia S. O.'Regan,* D. K. Melhorn,* A. J. Newman,* and R. C. Graham,* Cleveland, Ohio (Introduced by S. Gross, Cleveland, Ohio) An 8-year-old girl with Type II congenital dyserythropoietic anemia, characterized by morphologic abnormalities of erythroid precursors, marked nuclear and cytoplasmic membrane abnormalities shown by electron microscopy, immunologic alterations, hyperbilirubinemia, and chronic anemia, was found to be vitamin E deficient. Nutritional history and vitamin E absorption studies indicated that neither dietary lack nor intestinal malabsorption was the cause of the deficiency. Striking changes in the patient's hematologic status following "administration of vitamin E included rise in hemoglobin, decrease in bilirubin and reticulocyte count, and a marked increase in red cell survival. Erythrocyte phospholipids, decreased
916
Abstracts
The Journal of Pediatrics June 1974
while the patient was vitamin E deficient, returned toward normal levels during therapy. The most notable increase was seen in the phosphatidyl ethanolamine fraction. However, hematologic improvement was not complete, immunologic abnormalities persisted, and morphologic aberrations in erythrocyte precursors actually increased during vitamin E therapy. It is therefore concluded that vitamin E deficiency played a role in the production of the child's hematologic disorder, and may have been a result of the increased utilization of the vitamin in stabilization of defective cellular membranes.
cies of children there is demonstrable anti-tumor cell activity by lymphocytes which is countered by a patient serum factor capable of blocking it. COMMENT: It was asked if blocking antibodies were present; one serum was found to possess blocking antibody. In response to the question concerning the usefulness of this test to monitor therapy, it was aswered that follow-up was not yet of sufficient duration; several patients in the group had died. The question was raised whether 5tCr release from the target cells was related to toxicity; Dr. Ertel answered that with increased cell destruction there was increased 51Cr release. Dr. Howard asked if this reaction was specific; did lymphocytes from normal people kill tumor cells as well? Dr. Ertel replied that normal cells were capable of cytotoxicity but at a lower level.
22. L ymphocyte-mediated cytotoxicity in children's tumors I. J. Ertel, Columbus, Ohio
23. Hypothalamic-pituitary regulation in familial goiter
Lymphocyte-mediated cytotoxic effect upon tumor cells was studied in children with neoplastic diseases. The assay tumor cells were cultured from operative specimens of neuroblastomas, Wilms" tumors, and rhabdomyosarcomas by an in vitro technique which produced monolayers. The target cells were then labeled with radioactive chromium (51Cr). Lymphocytes isolated from the peripheral blood of children with previously diagnosed malignancies were tested for cytotoxic activity against these tagged tumor cells. The lympbocytes were first purified by the Ficoll-Hypaque density gradient separation method. Cytotoxicity was quantitated by measuring the amount of 51Cr released from the lysis of tagged tumor cells after incubation with different concentrations of lymphocytes. The isotope assays were consistently repeatable and without subjective error. The activity of the lymphocytes from 20 children was compared to the anti-tumor lymphocyte activity of their own parents and that of unrelated normal controls. The patients' lymphocytes showed greater cytotoxic effects upon the tumor cells than did those of their parents or normal controls. Lymphocytes from tumor-bearing children showed the greatest cytotoxic effects upon the tumor cells and that effect was proportionate to the concentration of lymphocytes present. To assess whether serum factors influenced lymphocyte antitumor activity, sera from patients and controls were introduced into the test system. When serum from tumor-bearing patients was added for preincubation of the tumor cells, the cytotoxic activity of lymphocytes decreased. It appears that in the malignan-
Amelia V. Agustin,* Anita Perrieelli,* and Thomas P. Foley, Jr., Pittsburgh, Pa. Neuroendocrine regulation of thyrotropin (TSH) and prolactin (PR) release was studied in two patients with familial goiter secondary to a defect of thyroxin synthesis, with the onset of goiter in early childhood. Patient A. S. was hypothyroid after two months off thyroid therapy; Patient K. A. is euthyroid on no medications (Table I). TSH and PR levels were measured by radioimmunoassay. Serum was obtained q 30' for 180 minutes after L-dopa (500 mg. p.o.) and q 15' x 4, q 30' x 4 to 180 min. after IV infusion of TRH (7/zg per kilogram). Nocturnal samples were collected q 30' from 1130 to 0730, then q 15' to 0830. The results are summarized as shown in Table ii. During nocturnal sampling of Patient K. A. between 0530 and 0830 the mean TSH decreased by 33 per cent of the mean TSH between 1130 and 0500. SUMMARY: 1. Dopaminergic stimulation will decrease the elevated TSH levels in patients with familial goiter; the decrease is greater in the euthyroid patient. 2. TSH responses to TRH are greater in the hypothyroid patient. 3. Normal PR levels are suppressed after L-dopa and stimulated after TRH. COMMENT: Dr. Pena asked why an increase in T 4 w a s noted. Dr. Agustin suggested that this might be related to iodine deficiency.
23. Table I
Pt.
CA I T41I PBI I
A.S. K.A.
7.6 15.1
TSH
RAIt31uptake
pKCIO 4
probabledefect
I
1.7 4.8
5.5 9.7
31.3 38.5
6 hr: 6 hr:
74% 78%
24 hr: 24 hr:
67% + dischg. Organification 65% -- dischg. Defective thyroglobulin
23. Table II
Patient A. S.
Test L-dopa TRH L-dopa TRH
TSH (p,U/ml.) TSH (/zU/ml.) PR (ng.lmL) PR (ng./ml.)
Decrease: Increase: Decrease: Increase:
38.5to 40.9 to 4.6 to 5.0to
11.8 297 (2.5 35.0
Patient K. A. atl80' at 45' at 90' at 30'
31.3to 26.8 to 7.4 to .7.9to
<2.5 166 (2.5 47.5
atlS0min. at 45 rain. at 90 rain. at 30min.