Adel A. Yunis

Isolation of colony stimulating factor from human milk

Human milk contains colony stimulating factor (CSF), a polypeptide growth factor, which stimulates in in vitro bone marrow culture proliferation and differentiation of colony forming granulocytic macrophage progenitor cells (CFU-GM) to form colonies. This activity was not found in either bovine milk or colostrum when assayed in human or mouse bone marrow cells. The human milk CSF activity is destroyed by treatment with proteases. However, neither 6M urea, 4M guanidine hydrochloride, 5 mM dithiothreitol, nor exposure to pH 2 will inactivate the milk derived CSF. Gel filtration and isoelectric focusing indicate that human milk CSF differs biochemically from the other CSFs isolated from various sources and has a molecular weight between 250,000 and 240,000 and an isoelectric point between 4.4 and 4.9.

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 1β 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.—Jimenez, J. J.; Wong, G. H. W.; Yunis, A. A. Interleukin 1 protects from cytosine arabinoside‐induced alopecia in the rat model. FASEB J. 5: 2456–2458; 1991.

Treatment with imuvert/JV-acetylcysteine protects rats from cyclophosphamide/ cytarabine-induced alopecia: Original article

Chemotherapy-induced alopecia is a distressing problem to the cancer patient for which currently there is no effective preventive measure. Recently hnuVert, 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, administeredparenterally 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 setting remains to be determined.

Differential effects of chloramphenicol and its nitrosoanalogue on protein synthesis and oxidative phosphorylation in rat liver mitochondria.

Abstract Protein synthesis and oxidative phosphorylation were chosen as measures to study the differences between the effects of chloramphenicol (CAP) and its derivative nitroso-chloramphenicol (NO-CAP) on rat liver mitochondria. [ 14 C]Leucine incorporation into mitochondrial protein was inhibited 83 per cent by 30 μM CAP and was equally inhibited by a similar concentration of thiamphenicol; 30μM NO-CAP, however, inhibited [ 14 C]leucine incorporation only 34 per cent and 30μM nitrosobenzene had no effect. A millimolar concentration of CAP was required to inhibit oxidative phosphorylation, whereas 75 μM NO-CAP was inhibitory. Unlike CAP, NO-CAP at 100 μM slightly inhibited state 4 respiration with glutamate as substrate, but slightly activated it with succinate. Respiratory state 3 with glutamate was completely inhibited by 75 μM NO-CAP, whereas the same concentration of CAP was only 10 per cent inhibitory. With succinate, 250 μM NO-CAP was required to inhibit state 3, whereas 600 μM CAP had no effect. The uncoupled state triggered by 2,4-dinitrophenol in the presence of either glutamate or succinate was inhibited totally by NO-CAP, but not by CAP. The inhibition by NO-CAP was mitochondrial protein dependent, for more NO-CAP was required for inhibition with a larger amount of protein. NO-CAP effects could be prevented or released by cysteine, but not by washing. Oxidative phosphorylation was also inhibited by another nitroso compound, nitrosobenzene, which, however, did not affect mitochondrial protein synthesis. The results indicate that, unlike CAP, NO-CAP is a potent inhibitor of the energy conserving mechanism.

The isolation and characterization of a colony stimulating factor from human lung

Serum-free conditioned medium from human lung obtained at autopsy provides a rich source of colony stimulating factor which stimulates granulocytic and macrophagic colony growth in both mouse and human bone marrow. The appearance of the factor is enhanced by endotoxin and inhibited by either puromycin or actinomycin D. Human lung colony stimulating factor is stable at the pH range of 6.5-10 and temperature of 56 degrees C for 30 min. It is resistant to trypsin and neuraminidase but is sensitive to subtilisin, chymotrypsin and periodate. It shows heterogeneity on Sephadex gel filtration with two activity peaks having molecular weight of 200 000 and 40 000, respectively. Upon gel electrophoresis, human lung colony stimulating factor migrates in the alpha-globulin post-albumin region. Using the combination procedures of hydroxyapatite chromatography and preparative polyacrylamide gel electrophoresis a 600-fold purification was achieved with a final specific activity of 6-10(5) units per mg protein. The purified colony stimulating factor is very labile; however, the activity can be stabilized by the addition of gelatin or bovine serum albumin at the concentration of 0.1% and 0.2 mg/ml, respectively.

The effect of chloramphenicol treatment on ferrochelatase activity in dogs

Abstract Chloramphenicol fed to dogs at a dose of 100 mg/kg/day resulted in a decrease in bone marrow ferrochelatase activity to 5–35% of control levels. Chloramphenicol in concentrations of 25 and 50 μg/ml had no effect on ferrochelatase activity in vitro .

Enzymes of glycogen metabolism in white blood cells III. Heterogeneity of glycogen phosphorylase from rat chloroma

Summary 1. Glycogen phosphorylase ( α -1,4-glucan:orthophosphate glucosyltransferase (EC 2.4.1.1)) of rat chloroma has been purified and partially characterized. 2. Column chromatography on o -(diethylaminoethyl)-cellulose yields 2 phosphorylase peaks both of which are active without adenosine 5′-phosphate. The identity of these 2 peaks has been ascertained by rechromatography on O -(diethylaminoethyl)-cellulose and electrophoresis on acrylamide gel. 3. There appears to be some distinct differences in the properties of the enzyme from the 2 peaks. 4. The finding of 2 isoenzymes of phosphorylase in rat chloroma represents the second example of heterogeneity of this enzyme, the first example being that of phosphorylase from rabbit heart.

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 uncharactcrized genetic defect.

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.

Characteristics of fibrinolysin secreted by cultured rat breast carcinoma cells.

Summary Fibrinolysin secreted by cultured R2426 rat breast carcinoma cells has been purified and partially characterized. Three methods of assay were used: the hydrolysis of [ 125 I]fibrinogen and N-p -tosyl- l -arginine methyl ester (TAME), and clearing of fibrin-agar plates (heated and unheated). Two types of fibrinolytic activity were identified: a plasminogen activator and a direct fibrinolysin. The fibrinolysin had direct hydrolytic action on plasminogen-free fibrinogen, fibrin, and TAME. The products of fibrinogen hydrolysis by the tumor cell fibrinolysin differed from those of human plasmin. It was heat stable (60°C, 12 h), and was inhibited by e-aminocaproic acid and soybean trypsin inhibitor. Ethylenediaminetetraacetate (EDTA) inhibited the action of the tumor cell enzyme on TAME, but not on human fibrinogen. The direct fibrinolytic activity could be separated from plasminogen activator activity by gel filtration on Sephadex G-200, and the molecular weight was estimated to be approx. 110 000 D.