Familial infantile thrombotic thrombocytopenic purpura.

PURPOSE: To further define familial infantile thrombotic thrombocytopenic purpura and clarify its pathophysiology, we describe a family with two infants presenting with this rare syndrome. RESULTS: Complete, but temporary remission followed the transfusion of whole blood in the first sibling and fresh frozen plasma (FFP) in the second. Periodic FFP transfusions have kept the surviving proband in a prolonged clinical remission. The presence of unusually large von Willebrand factor multimers was demonstrated in the proband and the processing activity of these large multimers was found to be normal. CONCLUSION: The occurrence of this rare disorder, in siblings who are products of a consanguinous union, suggests an as yet uncharacterized genetic defect.

Pretreatment with 1,25(OH)2D3 protects from Cytoxan-induced alopecia without protecting the leukemic cells from Cytoxan.

The authors previously demonstrated that pretreatment with 1,25(OH)2D3 protects from Cytoxan-induced alopecia in the rat model. The current study was designed to investigate whether 1,25(OH)2D3 protects the transplantable rat chloroleukemia (C51) cells from the cytotoxic effects of Cytoxan. In vitro, 4-hydroperoxycyclophosphamide had a dose-dependent cytotoxic effect on C51 cells. In separate experiments, preincubation with 1,25(OH)2D3 did not protect the C51 cells from the cytotoxic effects of 4-hydroperoxycyclophosphamide. In vivo, 4 groups of 10 5-day-old rats were treated as follows: Groups 1 and 2 received 0.2 micrograms of 1,25(OH)2D3 topically in ethanol daily starting on day 5 through day 10. Groups 3 and 4 received ethanol topically similarly. On day 7, all rats received 1 x 10(5) C51 cells intraperitoneally. On day 11, groups 1 and 3 received 35 mg/kg Cytoxan intraperitoneally. All rats in groups 2 and 4 were dead of leukemia by day 34. In groups 1 and 3, only 1 of 10 and 2 of 10 rats died of leukemia, respectively. Alopecia developed in all rats in group 3. In contrast, all rats in group 1 were protected from Cytoxan-induced alopecia. These results indicate that, in vivo, pretreatment with 1,25(OH)2D3 does not protect the rat chloroleukemia cells from the cytotoxic effect of Cytoxan, while protecting from Cytoxan-induced alopecia.

Protection from chemotherapy-induced alopecia by 1,25-dihydroxyvitamin D3.

We have previously reported that several biological agents, when given simultaneously with cytosine arabinoside or cytoxan, will protect from cytosine arabinoside-induced but not from cytoxan-induced alopecia. In the present study we used the secosteroid 1,25-dihydroxyvitamin D3 in a different timing schedule to protect from chemotherapy-induced alopecia. In three separate experiments, 0.2 microgram of topical, 1,25-dihydroxyvitamin D3 protected rats from alopecia induced by etoposide, cytoxan, and an Adriamycin-cytoxan combination. In another experiment, 0.1 microgram protected rats from etoposide-induced alopecia at the site of application. 1,25-Dihydroxyvitamin D3 may offer a new and exciting approach to the prevention of chemotherapy-induced alopecia.

Protection from chemotherapy-induced neutropenia by ImuVert.

Neutropenia is a major cause of chemotherapy-induced morbidity. This study examines the effect of ImuVert on the course of neutropenia from Cytoxan and Adriamycin. In adult rats treated with either Cytoxan or Adriamycin and ImuVert, the absolute neutrophil count remained above 7,000 and 2,000, respectively, for the duration of ImuVert therapy. In contrast, in the control Cytoxan group the absolute neutrophil count reached 300/mm3 on day 5 and remained below 1,000 from day 4 through day 7. In the Adriamycin control group, the nadir absolute neutrophil count reached 600/mm3 on day 7 and remained below 100 from day 6 through day 8. C3H/EJ mice (endotoxin-hyporesponsive) treated with Cytoxan developed an absolute neutrophil count below 1,000 from day 4 through day 7. In contrast, in the group concomitantly treated with ImuVert, the nadir absolute neutrophil count remained below 1,000 only on days 4 and 5. The authors conclude that stimulation of endogenous cytokine production by ImuVert may provide a potentially useful approach to bone marrow rescue from chemotherapy.

Interleukin 1 protects hair follicles from cytarabine (ARA-C)-induced toxicity in vivo and in vitro.

ImuVert, a biologic response modifier, and interleukin 1 (IL 1) have been shown to protect the young rat from alopecia induced by cytarabine (ARA-C). In the present study the inhibition by ARA-C of DNA synthesis in hair follicles (HFs) and the protective effect of ImuVert and IL 1 were investigated in vivo and in vitro. Both ImuVert and IL 1 were equally effective in protecting rats from ARA-C-induced alopecia. DNA synthesis in HFs isolated from ARA-C-treated animals was 10-20% of untreated controls. Follicles isolated from animals given either ImuVert or IL 1 before ARA-C exhibited normal DNA synthesis. In vitro, the incubation of normal rat HF with ARA-C resulted in 80% inhibition of [3H]-thymidine uptake. Preincubation of the follicles for 1 hr with IL 1 before the addition of ARA-C completely blocked the inhibition. Preincubation with imuVert, however, was less effective in blocking the inhibition from ARA-C.

Protection from 1-beta-D-arabinofuranosylcytosine-induced alopecia by epidermal growth factor and fibroblast growth factor in the rat model.

The present study was designed to examine the effect of epidermal growth factor (EGF) and fibroblast growth factor (FGF) on 1-beta-D-arabinofuranosylcytosine (ARA-C)- and cyclophosphamide-induced alopecia in the young rat model. Seven-day-old rats were given ARA-C with or without human or murine EGF daily for 7 days. Alopecia was scored on the 12th day of the experiment. Both human and murine EGF protected rats from ARA-C-induced alopecia. The topical application of murine EFG in dimethyl sulfoxide offered significant protection limited to the treated area. In other experiments the administration of acidic FGF (aFGF) with ARA-C resulted in protection from alopecia limited to the site of FGF injection. Neither EGF nor FGF had any influence on alopecia from cyclophosphamide. It is concluded that both EGF and FGF are effective in protecting against ARA-C-induced alopecia in the rat model.

Treatment with ImuVert/N-acetylcysteine protects rats from cyclophosphamide/cytarabine-induced alopecia.

Chemotherapy-induced alopecia is a distressing problem to the cancer patient for which currently there is no effective preventive measure. Recently ImuVert, a biologic response modifier, has been shown to protect from cytarabine-induced alopecia in the young rat model, but not from alopecia induced by cyclophosphamide. In the present study, the rat model was used to examine the effect of N-acetylcysteine on the course of alopecia from cyclophosphamide and of ImuVert plus N-acetylcysteine on alopecia induced by cytarabine-cyclophosphamide combination. The following observations were made: (1) Cyclophosphamide-induced alopecia could be effectively prevented by N-acetylcysteine, administered parenterally or applied topically in liposomes. (2) Alopecia caused by the combination of cyclophosphamide and cytarabine could be prevented by the parenteral or topical administration of ImuVert plus N-acetylcysteine. The potential applicability of these observations to the clinical settings remains to be determined.

Interleukin 1 protects from cytosine arabinoside-induced alopecia in the rat model.

Protection from cytosine arabinoside-induced alopecia by ImuVert has recently been reported in a rat model. ImuVert, a biologic response modifier, is capable of activating mononuclear cells causing release of various cytokines. In the present study, using the young rat model, recombinant human IL 1 beta produced excellent protection from cytosine arabinoside-induced alopecia. Mouse recombinant tumor necrosis factor gave definite but modest protection whereas human tumor necrosis factor gave none. It is concluded that the protection from alopecia by ImuVert is mediated by cytokines, primarily IL 1.

Synergism of ara-C-imuvert combination in aborting the development of transplanted chloroleukemia in the rat.

The development of chloroleukemia in the rat after transplantation of C51 chloroma cells can be aborted by treatment with an exogenous source of differentiating activity (DA) or by stimulation of endogenous DA production by the administration of Imuvert, a biologic response modifier. Success or failure of treatment was determined by the ratio of DA to leukemic cell load. Accordingly, in the present study we examined the hypothesis that treatment with cytosine arabinoside (ARA-C) to reduce the leukemic cell load coupled with Imuvert would be more effective than either given alone. Treatment with ARA-C-Imuvert aborted the development of chloroleukemia in 79% of the animals in contrast to 0%, 9% and 45% in animals treated with buffer, ARA-C, or Imuvert respectively. These results suggest that stimulation of endogenous DA in combination with cytoreductive agents offers a potentially curative treatment for myelogenous leukemia.

Interaction of chloramphenicol and metabolites with colony stimulating factors: possible role in chloramphenicol-induced bone marrow injury.

We have recently demonstrated that two chloramphenicol (CAP) metabolites known to be produced by intestinal bacteria, dehydro-CAP (DH-CAP) and nitrophenylaminopropane (NPAP), are much more cytotoxic to bone marrow in vitro than CAP itself. Since colony stimulating factors (CSFs) play an essential role in hematopoietic cell growth, toxicity from CAP metabolites could also involve interaction with CSF or CSF-producing cells. In the present study, we found that increasing concentrations of rhGM-CSF or rhG-CSF completely reversed the inhibitory effect of CAP (2 x 10(-4) M) on human CFU-GM growth and on the growth of KG-1 cells. GM-CSF also reversed the inhibitory effect of CAP on HL-60 cells. Inhibition by DH-CAP (50% at 5 x 10(-7) M), nitroso-CAP (NO-CAP) (60% at 5 x 10(-6) M) and NPAP (35% at 10(-5) M) was not affected by either CSF. In addition to their inhibitory effect on cell growth, DH-CAP (5 x 10(-6) M) and NO-CAP (5 x 10(-6) M) inhibited CSF production by buffy coat cells 50-70% without affecting cell viability. Neither CAP nor NPAP inhibited CSF production. It is suggested that the dual toxic-inhibitory effect of some intestinal metabolites of CAP such as DH-CAP on hematopoietic cell growth on the one hand, and on CSF production on the other, renders them very potent as potential mediators of CAP induced aplastic anemia.

Protection from chemotherapy-induced alopecia in a rat model.

Alopecia (hair loss) is among the most distressing side effects of cancer chemotherapy. Little progress has been made, however, in its prevention or treatment, partly because of the lack of suitable experimental model. In recent work on the treatment of myelogenous leukemia in the rat, the following observations were made: (i) treatment of 8-day-old rats with cytosine arabinoside consistently produced alopecia, and (ii) ImuVert, a biologic response modifier derived from the bacterium Serratia marcescens, uniformly produced complete protection against the alopecia. In subsequent experiments, both cyclophosphamide and doxorubicin also produced alopecia in this model, and the doxorubicin-induced alopecia was prevented by treatment with ImuVert. The potential relevance of these observations to chemotherapy-induced alopecia in the clinical setting should be examined.

Treatment with Imuvert aborts development of chloroleukemia in newborn rats.

We have previously demonstrated that the successful transfer of rat chloroleukemia (Mia C51) cells to newborn rats is related to the host's inability to generate adequate levels of differentiation factor (DF). Thus, when the appropriate amount of DF was injected into rats bearing MIA C51 cells, the development of chloroleukemia was aborted. In the present study, we provide evidence that stimulation of endogenous differentiation activity (DA) production by the administration of a biologic response modifier (Imuvert) will like-wise abort the development of chloroleukemia. Imuvert at 50 micrograms/ml had no direct effect on growth, viability, or differentiation of MIA C51 cells. However, when monocytes from young rats or adult rats were stimulated with Imuvert in vivo or in vitro, there was significant increase in DA production. Treatment of young rats with Imuvert aborted the development of chloroleukemia from transplanted MIA C51 cells. It is concluded that stimulation of endogenous DA production may provide a potentially useful approach in the treatment of leukemia.

Application of thermospray liquid chromatography-mass spectrometry for the analysis of chloramphenicol and three related compounds.

The thermospray (TS) liquid chromatographic-mass spectrometric analysis of the antibiotic chloramphenicol and three related compounds is presented. The three additional compounds are dehydrochloramphenicol, aminodehydrochloramphenicol, and nitrophenylaminopropanediol. Baseline separation of the four compounds is achieved. The TS mass spectrum of each of the four compounds includes a prominent [MH]+ ion plus some fragment ion peaks.

Congenital neutropenia: a case study.

Infantile genetic agranulocytosis (IGA) has a high morbidity and mortality rate due to severe neutropenia. The pathogenetic mechanisms of this syndrome have not been elucidated. However, a recent clinical trial with recombinant human granulocyte-colony-stimulating factor (rhG-CSF) has shown a dramatic increase in the absolute neutrophil count in patients with IGA. This suggests that these patients have either a lack of granulocyte-colony-stimulating factor (G-CSF) or have a defect in the G-CSF receptors. A clinical trial of recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF) in an infant with IGA is reported in this article. A marked eosinophilic response was observed without any increase in the absolute neutrophil count (ANC). In an effort to elucidate the pathogenetic mechanism underlying IGA, we examined (a) the in vitro response of patient's CFU-GM to rhGM-CSF and to rhG-CSF and (b) the ability of patient's monocytes to produce G-CSF. Our results tend to support the thesis that the defect in IGA is at the G-CSF receptor level. We also found a lack of correlation between in vivo and in vitro response to rhGM-CSF.

Chloramphenicol toxicity: 25 years of research.

Significant progress has been made in recent years in the understanding of the pathogenesis of the two types of hematologic toxicity from chloramphenicol. The common, dose-dependent, reversible bone marrow suppression from chloramphenicol is a consequence of mitochondrial injury. The greater erythroid susceptibility to chloramphenicol appears to be a function of the endogenous mitochondrial amino acid pools. The pathogenesis of aplastic anemia from chloramphenicol treatment remains uncertain. A large body of indirect evidence favors a complex mechanism involving metabolic transformation of the p-NO2 group of chloramphenicol by the predisposed subject, leading to the production of a toxic intermediate causing stem cell damage. A concept is presented wherein metabolites of chloramphenicol generated by intestinal bacteria undergo further metabolic transformation in the bone marrow with in situ production of toxic intermediate. This concept of the marrow being both the metabolic site for the offending agent as well as the target for its toxic metabolites will likely apply to other potential myelotoxins.

Drug-induced red cell dyscrasias.

Major advances have been made in recent years in our understanding of the pathogenetic mechanisms of drug-induced blood dyscrasias, particularly those involving the red cell. Among the latter, hemolytic anemia is the most common. Drug-induced red cell destruction may occur on an immune basis or through disruption by the drug of red cell metabolism. The immunological basis of drug-induced hemolysis is reviewed with emphasis on the clinical and laboratory manifestations, differential diagnosis and the major mechanisms involved. Drug-induced oxidative hemolysis both in normal individuals and in those with certain enzymopathies, notably glucose-6-phosphate dehydrogenase deficiency, is summarized. Drugs may also produce red cell dyscrasias by acting on the immature erythroid compartment. Some of these inhibit erythroid growth by as yet poorly understood mechanisms. Others exert more specific metabolic effects in erythroid precursors. These include drugs which interfere with DNA synthesis causing megaloblastic erythropoiesis and those which disrupt mitochondrial function and the synthesis of heme manifested by sideroblastic erythropoiesis. A brief consideration of heme biosynthesis and the action of drugs which are associated with sideroblastic anemia, including the antituberculous agents, lead, alcohol and chloramphenicol is presented. Finally, where pertinent, an updated listing of drugs involved in red cell dyscrasias is included.

Treatment with monocyte-derived partially purified GM-CSF but not G-CSF aborts the development of transplanted chloroleukemia in rats.

We have previously demonstrated that cultured rat chloroleukemia cells, MIA C51, will terminally differentiate to macrophages when treated with rat lung-conditioned medium in vitro and in vivo. In the present study we fractionated rat monocyte-conditioned medium by ultrafiltration according to molecular size. The fraction with molecular weight (mol wt) 30 to 50 Kd containing partially purified granulocyte-macrophage colony-stimulating factor (GM-CSF) activity caused the differentiation of C51 cells to macrophages in vitro and in diffusion chambers in vivo. Treatment of young rats with this fraction aborted the development of chloroleukemia from transplanted C51 cells. In contrast, the fraction with mol wt 10 to 30 Kd containing virtually all the G-CSF activity exhibited no differentiation activity either in vitro or in vivo. It is concluded that in this rat myelogenous leukemia model partially purified GM-CSF but not G-CSF contains the effector molecule(s) causing terminal differentiation of C51 cells and tumor cell rejection.

Aerobic nitroreduction of dehydrochloramphenicol by bone marrow.

It has been previously demonstrated that dehydrochloramphenicol (DH-CAP), a bacterial metabolite of chloramphenicol, induces DNA single strand breaks in intact cells and is profoundly more cytotoxic than chloramphenicol (CAP). In view of previous observations relating genotoxicity of nitrocompounds to their nitroreduction by the target tissue, we studied the nitroreduction of DH-CAP by human and rabbit bone marrow. Nitroreduction by tissue homogenates was determined by the Bratton Marshall colorimetric assay and by high-performance liquid chromatography (HPLC). Nitroreduction of DH-CAP by bone marrow cell homogenates was observed under aerobic conditions and the reduction was both cell concentration- and time-dependent. The formation of the amino product aminodehydrochloramphenicol was confirmed by HPLC. Reduction by other tissues including human liver, Raji cells, and HL-60 tumors was also observed. These results suggest that genotoxicity of DH-CAP may be related to its nitroreduction by the target tissue with in situ production of toxic intermediates. Together with previous studies, these observations lend support to the thesis that the p-NO2 group may be the structural feature underlying aplastic anemia from CAP.