G. Izak

Suppressive Effect of Ferritin on in Vitro Lymphocyte Function

Summary. This study describes the effect of ferritin on lymphocyte function in vitro. Peripheral blood lymphocytes isolated from normal donors were incubated with purified human splenic ferritin, and the mitogenic effect of phytohaemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM) and mixed lymphocyte reaction (MLR) were assessed by the uptake of 3H‐thymidine (3H‐TdR). Ferritin (0·25–5·0 μg/ml culture) caused a marked suppression of PHA and Con A blastogenesis but had no suppressive effect on PWM‐induced transformation. Maximal suppression was obtained at a ferritin concentration of 1 μg/ml and this was not enhanced by increasing ferritin concentrations. Ferritin also reduced the Con A capping phenomenon in normal lymphocytes from 22% to 6%, suppressed the MLR reaction but had no effect on the ability of normal lymphocytes to form E, EA and EAC rosettes or on in vitro lymphocyte cytoxicity against the K‐562 cell line. Visual proof of the suppressive effect of ferritin on mitogen induced blastogenesis was provided by scanning electron microscopy, and direct evidence for the ability of lymphocytes to bind ferritin was obtained from studies with radioiodine labelled ferritin. The above findings indicate that ferritin suppresses certain parameters of T‐lymphocyte function in vitro. The relation of the present findings to recognized abnormalities of T‐cell function encountered in certain neoplastic disorders associated with high serum ferritin levels is at present unknown.

Prevalence and causes of anemia in elderly hospitalized patients.

The prevalence and causes of anemia have been studied in 104 patients over 60 years of age admitted to a general medical ward in Jerusalem. In males and females, mean hemoglobin levels were about 1 g less than in the corresponding groups of healthy younger controls. A primary nutritional anemia could not be implicated in any of the 15 patients with hemoglobins below 11 g/dl. The most important causes of anemia were chronic renal failure, metastatic carcinoma, gastrointestinal bleeding, and infection. Conversely, in diseases with no adverse effect on erythropoiesis such as chronic ischemic heart disease, hypertension and diabetes, hemoglobin levels were equal to those of the younger controls. These findings indicate that although diminished serum iron and RBC folate levels may occasionally be found in elderly subjects, nutritional deficiency is seldom responsible for anemia in this age group in Israel- and anemia when present is often the manifestation of a chronic underlying disease.

Folic and folinic acid in maternal and foetal blood.

SEVERAL investigators have shown that cord blood contains a higher concentration of vitamin B,, than thc maternal blood (Chow, Barrows and Ling, 1951; Karlin, 1956; Sadovsky, Bercovici, Rachmilewitz, Grossowicz and Aronovitch, 1959). This finding presumably indicates the increased demand of the developing foetus for the vitamin, leading occasionally to depletion of the maternal stores and ultimately to anaemia. It is reasonable to expect that a depletion in maternal folic acid may develop by the same mechanism, especially as macrocytic anaemia in pregnancy is known to respond to treatment with folic acid much more readily than to vitamin BIZ. Baker, Ziffer, Pasher and Sobotka (1958) compared folic acid levels in foetal and in matcrnal sera, using a thermophlic bacillus as a test organism, and found hgher values in the foetal sera. Folic acid consists of a variety of metabolites of both high (folinic and tetrahydrofolic acid) and low (pteroylglutamic acid) co-factor activity. It is a plausible assumption that in the foetus with its high metabolic and haematopoietic activity, the active forms of folic acid mainly would be found; a study was thereforc undertaken to determine the various folic acid metabolites in foetal and maternal blood. Since the red cells contain much more folic acid than the serum, folic and folinic acid were determined in whole blood. The results of this study are presented in this paper.

The effect of acute haemorrhage and acute haemolysis on the intestinal iron absorption in the rat.

As the total body pool of iron is extremely small, a very sensitive and accurate mechanism is required to preserve the balance between the amount of iron entering the body and that being excreted. Although much effort has been concentrated on clarifying the nature of this regulating mechanism, the conclusions arrived at by different investigators vary widely. It has been stated that the body iron pool is maintained by controlled absorption, while excretion plays only a minor role (Moore, 1955; Pirzio–Biroli, Bothwell and Finch, 1958; Pirzio–Biroli and Finch, 1960). It has been proposed by some investigators that the iron content, or the degree of saturation of iron–binding protein in the intestinal epithelium are responsible for the amount of iron absorbed from the gut (Hahn, Bale, Ross, Balfour and Whipple, 1943; Granick, 1946; Stewart, Yuile, Claiborne, Snowman and Whipple, 1950). Others have denied the importance of these mechanisms (Brown, Dubach and Moore, 1958) and have advocated, instead, the importance of erythropoietic activity in the regulation of iron absorption (Chapelle, Gabrio, Stevens and Finch, 1955; Bothwell, Pirzio–Biroli and Finch, 1958; Reynafarje, Lozano and Valdivieso, 1959). Still other workers have demonstrated that iron is lost from the body in desquamated intestinal epithelium in a selective manner so that needed iron is conserved and excess eliminated (Conrad and Crosby, 1963; Crosby, 1963).

The Absorption of Milk-Bound Pteroylglutamic Acid from Small Intestine Segments

Summary Binding tritiated pteroylglutam-ic acid to cow or to goat milk protein was associated with a substantial reduction in its absorption from rat jejunum, while the ileal absorption increased markedly. There was no difference in the amount of folate absorbed whether it was bound to cow milk or to goat milk. Despite the fourfold higher folate binding capacity of goat milk than of cow milk, there was no difference in the amount of folate absorbed after their administration. Folate depletion or folate overload had no effect on the amount of the milk-bound folate absorbed, while such treatments decreased the absorption of free folate.

Folate Activity in Tissues of Rats on Folate-Free Diets.∗

Summary Rats kept on a folate-free diet were found to have decreased folate activity in kidneys, livers and blood in comparison with control animals kept on a mixed standard diet. In the kidneys the drop in folate activity amounted to 70% after 10 days, and to about 85% after 20 days. No appreciable differences were encountered, regardless of whether or not the kidneys were autolyzed prior to the bioassay, or whether the bio-assay was performed with L. casei or P. cerevisiae. A 40% drop in folate activity was found in livers of animals kept on a folate-free diet for 10 days; the drop amounted to 80% after 20 days. These results were obtained when autolyzed tissues were employed in the bioassay. Livers assayed without prior autolysis showed very low activity with P. cerevisiae because of the high contents of methyl-tetrahydrofolate of the liver. After autolysis similar activities were obtained with both assays. Decreased blood folate activity was found in rats only after they had been kept for 20 days on the deficient diet. Thus, decreased folate activity in the blood indicates a severe depletion of the tissue stores. Rats kept on a synthetic diet supplemented with PGA (20 or 4 mg/kg diet) showed a higher liver and blood folate activity than control rats fed on mixed standard diet. The livers of both groups showed almost double folate activity when assayed on autolyzed samples than when done without such treatment. Autolysis presumably made available to L. casei forms of folates (e.g., polyglutamates) to which it could not respond prior to the treatment. The valuable technical assistance of Mrs. M. Jab-lonska is gratefully acknowledged.

Folate Activity in Rat Tissue Before and After Pteroylglutamic Acid Load.

Summary Folate activity was determined in the whole blood and tissues of 70 normal rats, before and after loading with PGA using 3 different bio-assays simultaneously. L. casei was used to measure “total folate activity,” S. faecalis to estimate monoglutamate activity as well as that of the reduced forms of PGA, and P. cerevisiae to determine only the reduced folate activity. L. casei activity before loading with PGA was about 25 times higher in the peripheral blood than when measured with S. faecalis and P. cerevisiae. On the other hand, the ratio of activity with L. casei, S. faecalis and P. cerevisiae in liver, kidney, spleen and intestines was only 1.4:1:1. It was concluded that the reduced forms constitute over 50% of the folate derivatives in rat tissues. Intraperitoneal injection of 2 μ/g PGA was followed by a temporary elevation of the activity with the L. casei and S. faecalis assays which disappeared within 6–24 hr after the injection. No significant increase was noted in the P. cerevisiae assay following PGA load.

Morphological-Biochemical Correlations in Rabbit Red Cell Precursors Synchronized by Actinomycin Administration

Summary. By making use of the selective toxicity of Actinomycin D, homogenous erythroid populations of various stages of maturation had been obtained in the rabbit. Comparison of the morphological characteristics of red cell precursors with in vitro synthetic activity as measured by the rate of incorporation of 32P, 59Fe and [2‐14C]glycine into DNA, RNA, haem and protein respectively, revealed the following:

The Effect of Acute Haemorrhage and Haemolysis on Folate Metabolism in the Rat

It has been reported by several investigators that increased blood production accompanying haemolytic disease is usually associated with folate deficiency (Chanarin, Dacie and Mollin, 1959). In many such instances the bone marrow shows megaloblastic transformation which reverts to normal following treatment with folic acid (Chanarin et al., 1959). It therefore seemed of interest to study the effect of experimentally induced bone marrow proliferation on the requirement and distribution of various folate metabolites in the blood, serum, bone marrow and liver tissue.

Utilization of Yeast Polyglutamate Folates in Man

Summary Ingestion by healthy humans of small amounts of polyglutamate folates from yeast, equivalent to 300 μg of monoglutamate folate and containing 30 μg of “free folate,” resulted in an appreciable elevation of the serum folate corresponding to 300 μg of synthetic pteroylmonoglutamate (PGA). Ingestion of higher amounts of polyglutamate folate did not result in higher serum folate elevations than did 300 μg. It is concluded that small amounts of polyglutamate folate from yeast are fully utilized, presumably by deconjugation in the gut prior to absorption. The relative ineffectiveness of larger doses of polyglutamate folates from yeast may be due to limiting conjugase activity in the gut, unfavorable conditions for its activity (such as unsuitable pH) or to an inhibitor of the enzyme present in impure preparations. The able technical assistance of Mrs. M. Jablonska, H. Barromi, and M. Ben-Basat is gratefully acknowledged. This investigation received financial support from the World Health Organization, Geneva, and from the Ministry of Health, Government of Israel, Jerusalem.