Erythropoiesis Inhibiting Factor(s) in Intact and Haemolyzed Red Blood Cells

A previous report have shown that both intact red blood cells (RBC) and an equal number of haemolyzed cells were able to inhibit erythropoiesis on ESF-induced erythropoiesis, suggesting a cell compound to be responsible for the inhibitory effect. Haeme and haeme compounds have been found to stimulate or inhibit both erythropoiesis and haeme synthesis. The present work presents data on the inhibitory effect of haemolyzed RBC and compounds from intact RBC on erythropoiesis. The inhibiting factor was found to be of a small molecular size and of the same range as a urinary erythropoiesis inhibiting factor (EIF). The inhibitor did not contain haeme. Both Fe²⁺ and Fe³⁺ were tested, showing no reduction of the ⁵⁹Fe incorporation into RBC in the test animals. The inhibition could not be due to dilution of the ⁵⁹Fe with unlabelled iron from the haemolyzed cells. On the contrary, Fe³⁺-ions rather stimulated erythropoiesis, probably due to increased amounts of available iron. Haemolysates were prepared from RBC with different amounts of immature cells. With increasing amounts of reticulocytes, a reduction of the inhibitory effect occurred. Also foetal cells showed less inhibition than an equal amount of adult cells. After high speed centrifugation, the inhibitory effect of haemolysates was found in the supernatant, while ghost cells exerted no inhibition. No species differences were found using both exhypoxic polycythaemic mice and rats. An inhibiting factor was liberated into the incubation medium when RBC were incubated for 20 h. No haemolysis occurred during the incubation period. Mature, adult RBC therefore contain a substance which is different from haeme, with a negative feedback on erythropoiesis.     

Erythropoiesis Inhibiting Factor (EIF) The Inhibitory Effect of Oestrogens on Erythropoiesis and the Content of Oestrogens in the Urinary EIF

An erythropoiesis inhibiting factor (EIF) with a low molecular weight has been demonstrated in urine from normal healthy individuals, indicating a dual, humoral regulatory mechanism of the erythropoietic homeostasis. Since oestrogens are known to inhibit erythropoiesis and are excreted in the urine, the question was raised whether the EIF preparations could contain eostrogens in such amounts that this could explain the inhibitory effect. Fractionated eostrogen determinations in EIF preparations revealed only micro-quantities of the different oestrogen-metabolites. An inhibitory effect on erythropoiesis was obtained with commercial oestrogens only in doses 10⁴-10⁵ times higher than the oestrogen amounts found in the EIF preparations. It is therefore most unlikely that oestrogens are causing the inhibitory effect of EIF on erythropoiesis.     

Erythropoiesis in the Anaemia of Chronic Disease

The amount and effectiveness of erythropoiesis was measured using ⁵⁹Fe in 10 patients with the anaemia of chronic disease and in 10 iron deficient patients with a comparable degree of anaemia. In both conditions the anaemia was the result of the failure of the marrow to compensate for a modest degree of peripheral haemolysis but ineffective erythropoiesis was significantly greater in iron deficiency than in chronic disease. The results suggest that although the peripheral blood picture is similar in both conditions the anaemia of chronic disease cannot be attributed simply to iron deficient erythropoiesis.     

Erythropoiesis-inhibiting factor(s) (EIF): methodologic studies.

Erythropoiesis-inhibiting factors (EIF) have been demonstrated in plasma from hypertransfused animals and from polycythemic individuals during periods of hyperoxia, but there is a decided discrepancy in the data published. In the present paper methodologic variations of a bioassay for demonstrating the erythropoiesis-inhibiting factor are discussed. In these studies no inhibitor of erythropoiesis could be demonstrated in plasma from hypoxia-induced polycythemic mice (HPM) on posthypoxic day 5. Injections of RBC or an equal amount of hemolyzed RBC were capable of suppressing the stimulatory effects of ESF, indicating that a red cell constituent may be responsible for the inhibitory effect observed. Transfusion-induced polycythemic mice (TPM) were therefore considered to be less suitable for demonstrating erythropoiesis inhibitors. Our results from testing several doses of a urinary EIF in normal mice, TPM and HPM, indicated that the HPM provided the most sensitive assay system. A similar effect was obtained with hypoxia-induced polycythemic rats. The most marked effect was seen in HPM when the EIF was injected shortly before administering the ESF, while the effect was less pronounced when the EIF was injected 24 hr before or after the ESF.     

Erythropoiesis in primary (idiopathic) osteomyelofibrosis: Quantification,PCNA-reactivity,and prognostic impact

In 64 patients with primary (idiopathic) osteomyelofibrosis (OMF), a morphometric analysis has been performed on bone marrow trephine biopsies following sequential doubleimmunostaining with monoclonal antibodies against proliferating cell nuclear antigen (PCNA) and erythroid precursor cells (glycophorin C). The purpose of this study was to quantify erythropoiesis and its PCNA-staining capacity and, further, to determine the impact of these parameters for the development of anemia and for prognosis. In comparison with a control group (15 patients), a significant reduction in the number of erythronormoblasts could be demonstrated, associated with an increase in PCNA-labelling. Moreover, significant correlations between the amount of nucleated erythroid marrow cells and degree of anemia (hemoglobin level, hematocrit, erythrocyte count) and survival could be calculated. Adverse relationships were assessed between number of erythroid cells, thrombocyte count, and spleen size, and also argyrophilic (reticulin/collagen) fiber density. These interactions were thought to reflect the biological behaviour of the disease process, i.e., the progression or extent of myeloid metaplasia. Our findings support ferrokinetic studies suggesting erythroid hypoplasia as one of the major causes of anemia in OMF. The remarkable high PCNA-labelling index of the macrocytic-megaloblastoid appearing erythropoiesis is probably caused by an overexpression of this marker protein. A comparative evaluation of Ki-67 antigen immunostaining in splenic tissue (myeloid metaplasia) and of the PCNA-labelling in pernicious anemia lend support to the assumption of an undue prolongation of the S-phase generated by secondary folate (hematinic) deficiency. © 1994 Wiley-Liss, Inc.     

Studies on the Erythropoiesis Inhibiting Factor in the Plasma of Animals with Transfusion Polycythemia

Further investigations of the action of polycythemic plasma filtrate were madeusing Fe⁵⁹ and with detailed examination of the blood and bone marrow. Thesestudies confirmed the appearance of an active thermostabile plasma factor(erythropoiesis inhibitor) which depressed erythropoiesis in normal rabbits orrats. The plasma obtained from bilaterally nephrectomized sheep subjectedto transfusion polycythemia also contained the erythropoiesis inhibitor. Blockade of the reticuloendothelial system using trypan blue, in sheep, inducedproduction of the active substance similar or identical with the erythropoiesisinhibitor produced after transfusion polycythemia.

Submitted on November 2, 1960 Accepted on October 10, 1961     

Fetal Erythropoiesis after Allogeneic Bone Marrow Transplantation Estimated by the Peripheral Blood Erythrocytes Containing Hemoglobin F (F-cells)

During bone marrow engraftment following BMT there is a re-establishment of fetal erythropoiesis, expressed by the increase of F-cells. This seems to depend on several factors such as underlying disease, conditioning before therapy and other mechanisms concerning both the donor and the recipient bone marrow. The aim of this work was to study the factors influencing F-cell production during bone marrow engraftment following transplantation. We studied 28 patients who underwent allogeneic bone marrow transplantation, for various hematological malignancies (CML, AML, ALL, CMML and SAA). F-cells were estimated on peripheral blood smears by indirect immunofluorescence. Overall, there was an F-cell increase after BMT in comparison with values before BMT; this increase was significant on days 15–50 (p <.01). F-cell on days 18, 25, 32 and 40 following transplantation were significantly higher (p <.01) in patients who have had increased F-cell numbers post-chemotherapy before BMT, compared with the patients who did not show any increase of the F-cell number post chemotherapy. During the first month following transplantation (day 7 to day 40) patients who were transplanted from high F-cell donors failed to show any significant differences in their F-cell numbers in comparison to those transplanted from low F-cell donors. However, the F-cell increase became significantly higher in the former group between days 50 and 120. This observation implies that the stressed erythropoiesis of the initial phase does not allow revealing the varying F-cell production of the capacities donor bone marrow, while later, when the graft has settled, the high F-cell donors reveal this property of the host.     

Erythropoiesis inhibiting factor(s) in intact and haemolyzed red blood cells.

A previous report have shown that both intact red blood cells (RBC) and an equal number of haemolyzed cells were able to inhibit erythropoiesis on ESF-induced erythropoiesis, suggesting a cell compound to be responsible for the inhibitiory effect. Haeme and haeme compounds have been found to stimulate or inhibit both erythropoiesis and haeme synthesis. The present work presents data on the inhibitory effect of haemolyzed RBC and compounds from intact RBC on erythropoiesis. The inhibiting factor was found to be of a small molecular size and of the same range as a urinary erythorpoiesis inhibiting factor (EIF). The inhibitor did not contain haeme. Both Fe+2 and Fe+3 were tested, showing no reduction of the 59-Fe incorporation into RBC in the test animals. The inhibition could not be due to dilution of the 59-Fe with unlabelled iron from the haemolyzed cells. On the contrary, Fe+3-ions rather stimulated erythropoiesis, probably due to increased amounts of available iron. Haemolysates were prepared from RBC with different amounts of immature cells. With increasing amounts of reticulocytes, a reduction of the inhibitory effect occurred. Also foetal cells showed less inhibition than an equal amount of adult cells. After high speed centrifugation, the inhibitory effect of haemolysates was found in the supernatant, while ghost cells exerted no inhibition. No species differences were found using both exhypoxic polycythaemic mice and rats. An inhibiting factor was liberated into the incubation medium when RBC were incubated for 20 h. No haemolysis occurred during the incubation period. Mature, adult RBC therefore contain a substance which is different from haeme, with a negative feedback on erythropoiesis.     

Humoral Factor(s) in Experimental Hypersplenism

A type of experimental hypersplenism characterized by splenomegaly andthrombocytopenia has been produced in the rat by the repeated intraperitoneal injection of methylcellulose. The urine of these animals was collectedand given through a gastric tube to another group of normal rats for a periodof 4 weeks. The results were a marked and rapidly developing thrombocytopenia, a delayed but definite anemia with mild reticulocytosis and leukocytosis. When the administration of urine was discontinued, the anemia regressed,but the thrombocytopenia persisted unmodified for 2 weeks. When the urineof hypersplenic rats was again given to this group for 6 additional weeks, itfailed to induce anemia or to change the persistent thrombocytopenia. Intragastric administration of urine from normal rats, from rats made anemic bytotal body radiation and from the hypersplenic group after splenectomy toother groups of normal rats, failed to produce the same changes and onlyinduced moderate leukocytosis.

On the basis of these results, it is postulated that in experimental methylcellulose-hypersplenism in the rat there is a humoral factor(s) responsiblefor the thrombocytopenia, that this humoral factor(s) is eliminated in theurine and that when such urine is given to normal rats, it is responsible forthe thrombocytopenia and partially for the anemia that are observed. Suchfactor(s) are in some important way related to the presence of the spleen.

Submitted on October 16, 1959 Accepted on March 8, 1960     

Erythropoiesis in Sickle Cell Anaemia during Acute Infection and Crisis

Erythropoiesis was evaluated in 37 patients with sickle cell anaemia, 26 of them children under 12 years of age. Mean haemoglobin, haematocrit, reticulocyte, and erythropoietin levels were similar for 11 who were asymptomatic, 11 with infections, and 12 in vaso-occlusive crisis. Mean haemoglobin, haematocrit, and reticulocyte values were significantly lower and the mean erythropoietin level significantly higher for three patients in aplastic crisis. Reticulocyte counts reflected erythropoietic activity during the asymptomatic state but were variable during infection and crisis. No erythropoietic inhibitory activity was found in any of the four clinical states. It has been suggested that erythropoietin production decreases during infection. Patients in this study responded appropriately to stress, showing no decrease in erythropoietic activity during acute infection or crisis.     

Acquired,Transient Factor X (Stuart Factor) Deficiency in a Patient with Mycoplasma Pneumonial Infection

A case of severe haemorrhagic diathesis due to acquired deficiency of factor X (both immunologically and in procoagulant activity) is presented. The clinical and serological features of this case indicated mycoplasma pneumonial infection. Factor X in the peripheral blood did not appear to be influenced by administration of vitamin K, prothrombin-complex concentrate, fresh plasma or fresh whole blood. Circulating inhibitors of blood coagulation were absent and systemic amyloidosis could not be demonstrated. After 20 d, factor X spontaneously returned to normal. In view of the absence of other known causes of factor X deficiency, a possible relationship with mycoplasma pneumonial infection is suggested.     

脑梗死患者血浆EDRF、vWF、TNF含量变化探讨

目的观察脑梗死患者内皮细胞舒张因子(EDRF),血管性假血友病因子(vWF)及肿瘤坏死因子(TNF)血浆中的含量。方法测定42例急性期脑梗死患者,38例恢复期脑梗死患者及40例正常人血浆E-DRFv、WF、TNF含量,并分析其相互关系。结果(1)急性期脑梗死患者vWF、TNF较对照组明显升高(P<0.01),EDRF明显降低(P<0.01)。(2)大梗死组与小梗死组相比较,vWF、TNF水平升高和EDRF水平下降均有显著差异(P<0.01)。(3)急性期脑梗死患者vWF升高与TNF呈正相关(r=0.72、P<0.01),vWF升高与E-DRF呈负相关(r=-0.74,P<0.01)。(4)恢复期脑梗死患者vWF、TNF及EDRF与对照组比较,无显著差异(P>0.05)。结论急性脑梗死患者存在内皮细胞损伤和功能障碍。     

非诺贝特抑制脂肪细胞组织因子表达

为探讨动脉粥样硬化兔脂肪细胞组织因子表达及非诺贝特对其的影响 ,将 15只兔随机分为正常组、动脉粥样硬化组和非诺贝特组 ,非诺贝特组在高胆固醇饮食第 9周起加用非诺贝特 (每天 30mg/kg)干预 4周 ,采用逆转录聚合酶链反应测定脂肪细胞组织因子的表达。结果发现 ,高胆固醇饮食可显著升高血清总胆固醇 (P <0 .0 5 ) ,甘油三酯无明显升高 ;加用非诺贝特治疗 4周 ,总胆固醇和甘油三酯均无明显改变。动脉粥样硬化组脂肪细胞组织因子表达明显高于正常组 (1.0 81± 0 .0 11比 0 .939± 0 .0 18,P <0 .0 1) ,非诺贝特治疗 4周后组织因子表达较动脉粥样硬化组显著降低 (0 .893± 0 .0 2 2比 1.0 81± 0 .0 11,P <0 .0 1)。结果提示 ,动脉粥样硬化组兔脂肪细胞表达组织因子明显增加 ,非诺贝特能抑制其表达 ,提示非诺贝特可能具有抗血栓作用     

Recombinant human thrombopoietin (TPO) stimulates erythropoiesis by inhibiting erythroid progenitor cell apoptosis

Thrombopoietin (TPO) has been reported to stimulate erythropoiesis, but the stimulatory mechanism has not been defined. To address this issue, we performed serum-free cell-culture experiments with recombinant human TPO and purified human progenitor cells. We found that TPO alone was able to stimulate the megakaryocyte colony formation in serum-free cultures, but erythroid colonies were never observed. Only in the presence of EPO (erythropoietin) +IL-3 was TPO able to stimulate a small increase (∼25%) in erythroid colony formation. Accordingly, we hypothesized that TPO might have an effect on erythroid progenitor cell viability, rather than a direct stimulatory effect. To test this idea, CD34⁺ cells were cultured for 7 d in serum-free methylcellulose in the presence or absence of TPO, after which time KL+ EPO was added to the cultures. Cells which were pre-cultured for 7 d in the presence of TPO gave rise to approximately 6 times as many burst forming unit-erythroid (BFU-E) colonies as cells which were pre-cultured in the absence of TPO. Further, when primitive CD34⁺, Kit⁺ MNC were cultured for 3–7 d under serum-free conditions in the presence or absence of TPO, significantly fewer cells cultured in the presence of TPO displayed apoptotic changes when compared to cells cultured in the absence of TPO. Taken together, these results suggest that TPO has little direct stimulatory effect on erythroid progenitor cells, but might indirectly enhance erythropoiesis by preventing very early erythroid progenitor cells from undergoing apoptotic cell death.     

结核性脑膜炎脑脊液中粒细胞集落刺激因子的测定

本文采用ＥＬＩＳＡ法检测２３例结核性脑膜炎及２１例非结核性脑膜炎脑脊液中的Ｇ－ＣＳＦ。结脑组（０．６５＋０．３２ＯＤ）明显高于非结脑组（０．１５±０．１１ＯＤ）（Ｐ＜０．０１）。诊断结脑的敏感性为９１．３％，特异性为９０．５％。Ｇ－ＣＳＦ在抗感染的非特异性细胞免疫中起重要作用，细菌及其产物是刺激和调节Ｇ－ＣＳＦ产生的主要物质。