Induction of erythroid differentiation in K562 cells by inhibitors of inosine monophosphate dehydrogenase

The effects of three inhibitors of inosine monophosphate (IMP) dehydrogenase on a human erythroleukemic cell line, K562, were studied. Following incubation with these inhibitors, K562 cells underwent differentiation and accumulated hemoglobins. The induction of hemoglobin accumulation was dose dependent; maximum induction was observed at 100, 25, and 3 microM, respectively, for ribavirin, tiazofurin, and mycophenolic acid. The induction was associated with reduction of intracellular GTP content and was blocked by adding guanosine within 24 h after adding inducer. The effective dose for half-maximum induction by ribavirin was 3 times less than that for 50% inhibition of K562 proliferation; however, for tiazofurin and mycophenolic acid, it closely approximated the concentrations which suppressed cellular proliferation. Ribavirin was sequestered preferentially inside the K562 cells, and the induction by ribavirin had a greater than 30-fold increase in hemoglobin. Studies with isoelectric focusing, globin chain analyses, and immunochemical assays indicated that both A gamma and G gamma were detected and that the hemoglobin produced in the ribavirin-treated cells consisted of approximately 60% fetal hemoglobin and its acetylated equivalents. The adult-type alpha globin was found, while no beta globin chains were demonstrated. Thus, accumulation of fetal hemoglobin and production of alpha globin chain in ribavirin-treated cells are different from the pattern of hemoglobins induced by hemin.     

血速升颗粒联合铁剂治疗胃癌切除术后贫血的疗效及安全性比较

目的评价血速升颗粒联合蔗糖铁治疗胃癌切除术后合并缺铁性贫血患者的疗效及安全性。方法选取胃癌手术后合并缺铁性贫血的患者98例随机分为联合组和单药组,两组患者术后均给予蔗糖铁治疗,联合组加用血速升颗粒,比较治疗前后贫血指标的变化及药物不良反应情况。结果治疗后,联合组血红蛋白含量、红细胞压积、血清铁蛋白水平等较单药组增高,且差异有统计学意义。联合组治疗贫血总有效率及显效率高于单药组,且差异均有统计学意义。两组治疗后,中医症状记分均较治疗前减小。用药过程中,两组不良反应比较,差异无统计学意义。结论血速升颗粒联合蔗糖铁治疗可改善胃癌手术后合并缺铁性贫血患者的血红蛋白含量、红细胞压积、红细胞平均血红蛋白量、血清铁蛋白水平,并可改善贫血症状,较单用蔗糖铁效果更佳,且联合用药不增加不良反应发生率。     

Pharmacodynamic model for chemotherapy-induced anemia in rats

Anticancer agents often cause bone marrow toxicity resulting in progressive anemia which may influence the therapeutic effects of erythropoietic-stimulating agents. The objective of this study was to develop a pharmacodynamic (PD) model to describe chemotherapy-induced anemia in rats. Anemia was induced in male Wistar rats with a single intravenous (i.v.) injection of 60 mg/kg carboplatin. Hematological responses including reticulocytes, red blood cells (RBC), hemoglobin, and endogenous rat erythropoietin (EPO) were measured for up to 4 weeks. A catenary, lifespan-based, indirect response model served as a basic PD model to represent erythroid cellular populations in the bone marrow and blood involved in erythropoiesis. The model assumed that actively proliferating progenitor cells in the bone marrow are sensitive to anti-cancer agents and subject to an irreversible removal process. The removal rate of the target cells is proportional to drug activity concentrations and the cell numbers. An additional RBC loss from the circulation resulting from thrombocytopenia was described by a first-order process. The turnover process of rat EPO and EPO-mediated feedback inhibition mechanism regulated by hemoglobin changes were incorporated. Reticulocyte counts decreased rapidly and reached a nadir by day 3 after administration of carboplatin and returned to the baseline by day 13. This was followed by a gradual increase and the rebound peak occurred at about day 15. The hemoglobin nadir was approximately 9 g/dl observed at about 11–13 days compared to its normal value of 13 g/dl and hemoglobin returned to the baseline by day 30. The increase in endogenous rat EPO mirrored inversely hemoglobin changes and the maximum increase was observed soon after the hemoglobin nadir. The carboplatin-treated rats exhibited progressive anemia. The proposed model adequately described the time course of hematological changes after carboplatin in rats and can be a useful tool to explore potential strategies for the management of anemia caused by chemotherapy.     

The diagnostic plot

Iron balance is regulated by the rate of erythropoiesis and the size of the iron stores. Anemia that accompanies infection, inflammation, and cancer (anemia of chronic disease) features normal or increased iron stores, although patients may have functional iron deficiency, namely, an imbalance between iron requirements of the erythroid marrow and the actual supply. The proportion of hypochromic red cells and the hemoglobin content of reticulocytes are direct indicators of functional iron deficiency. Biochemical markers, especially the soluble transferrin receptor/log ferritin ratio (ferritin index), are useful indicators of the iron supply to erythropoiesis. The relationship between functional iron deficiency (reticulocyte hemoglobin content) and iron supply to erythropoiesis (ferritin index) can be described in a diagnostic plot. In normoproliferative and hypoproliferative erythropoiesis, the plot allows the differentiation of classic iron deficiency from anemia of chronic disease and the combined state of functional iron deficiency with anemia of chronic disease. The therapeutic implications of the plot are to differentiate patients into those who should be administered iron supplements, epoetin, or a combination of epoetin and iron. In patients receiving epoetin therapy, the plot is an important tool for monitoring erythropoietic activity, functional iron deficiency, and adequate iron stores for new red cell production. Enhanced erythropoiesis is reflected quantitatively by the ferritin index vector. A transgression of the 1.5 (3.2) cut-off value for the ferritin index indicates that extra doses of iron need to be administered to increase the body's iron stores. A lack of increase or a reticulocyte hemoglobin content below 28 picograms indicates functional iron deficiency. The diagnostic plot is a model for differentiating iron-deficient states and predicting those patients who will respond to epoetin therapy.     

血红蛋白水平影响食管癌放疗疗效的前瞻性研究

目的探讨促红细胞生成素干预对食管癌放疗疗效的影响情况。方法2000年7月~2001年02月,54例经病理证实的中晚期食管癌接受放射治疗的病例,治疗前血红蛋白浓度均小于100g/L,随机分成两组,一组为对照组,一组为治疗组。两组均采用6MV或者18MVX射线放射治疗,食管癌原发灶剂量60~68Gy,区域淋巴结剂量50~60Gy。对照组放疗过程中予以支持治疗,而干预组除支持治疗外,每周注射促红细胞生成素,3000I U/次,2/周,同时口服铁剂。放射治疗前、后均测定血红蛋白量。两组病例合并行影响生存时间因素的COX回归分析;对比两组局部控制率和生存率的差别。结果病变长度、血红蛋白变化值为生存时间的影响因素;干预组的局部控制时间与生存时间优于对照组。结论促红素的使用有益于食管癌放疗患者血红蛋白水平的提升,从而改善其局部控制率和生存率。     

Regulation of i- and I-antigen expression in the K562 cell line

Expression of i- and I-antigen in K562 cultured under different conditions of culture was investigated. Under standard conditions of culture, i-antigen expression was very high (100% of i-labeled cells) in contrast to I-antigen expression of which was very low (2 to 5% of I-labeled cells). The addition of hemin to K562 cells did not modify the mean antigenic density or the proportion of i- and I-labeled cells. In contrast, sodium butyrate elicited an important increase in the proportion of cells exhibiting I-antigen associated to a decrease of i-antigenic density. The effect of butyrate was reversible and dependent upon de novo protein and messenger RNA synthesis since it was abolished in the presence of cycloheximide or actinomycin D. The stimulation of i-antigen conversion to I-antigen elicited by butyrate cannot be directly related to an induction of differentiation since evidence in this sense is lacking; in fact, butyrate did not increase the hemoglobin content of K562 cells. The passage from exponential to stationary phase of growth (cell density inhibition) was associated with an increase in I-antigen expression and a slight decrease in i-antigen density on the surface of K562 cells.     

Epoetin alfa 40000 U once weekly and intravenous iron supply in solid tumor patients: early increase of hemoglobin level during chemotherapy

The objective of this observational study was the early evaluation of the impact, a week after the first administration of epoetin alfa 40000 U once weekly and i.v. dose of 62.5 mg sodium ferric gluconate for seven days in improving hemoglobin levels in cancer patients affected by mild/moderate or severe anemia during chemotherapy. Twenty patients affected by solid tumors who received epoetin alfa 40000 U once weekly and daily i.v. sodium ferric gluconate for one week were evaluated: 90% of the patients showed hemoglobin increase, with a median level of hemoglobin increase of 0.73 g/L from baseline, and 50% of them showing a hemoglobin increase > 1 gr/L. The treatment was well tolerated and no adverse event was observed. The early increase of hemoglobin level from baseline is interesting and suggestive for the possibility of achieving an adequate hemoglobin level with a short-term treatment. It is still necessary to further explore the real need of iron supplementation to maintain adequate erythropoiesis prior and during epoetin therapy.     

Induction of differentiation in erythroleukemic K562 cells by γ-irradiation

Various agents have been shown to induce differentiation in neoplastic cells. The present study aimed at investigating comparable phenomena induced by high doses of γ-irradiation in the presence of physiological factors. The erythroleukemic K562 cells were γ-irradiated or treated with cytosine-arabinoside (Ara-C), and examined for cell size, protein content, acetylcholinesterase (AChE)-activity and hemoglobin synthesis in relation to mitotic activity. At doses above 10 Gy, differentiation was induced, as recognized by elevated AChE-activity, accompanied by an increase in cell size and protein content and cessation of cell proliferation. Moreover, irradiation, as well as Ara-C, induced hemoglobin synthesis when cultures were supplemented with hemin prior to treatment. It is suggested that the basic mechanisms of differentiation induction are similar for ionizing radiation and certain chemical agents and are related to continued growth of essential cytoplasmic constituents during inhibition of mitotic activity.     

Effect of carboxyhemoglobin on tumor oxygen unloading capacity in patients with squamous cell carcinoma of the head and neck.

Hemoglobin and blood gas parameters, with special attention to the influence of carboxyhemoglobin, were studied in 115 head and neck cancer patients undergoing radiotherapy. In 712 weekly blood samples, the values of total hemoglobin, carboxyhemoglobin (CO-Hb), and p50 were measured and the total oxygen content in the arterial and tumor venous blood was estimated. The difference between these values express the tumor oxygen unloading capacity (t-OUC). CO-Hb ranged from 0-12% and showed a significant inverse relationship with t-OUC. This was caused by a reduced amount of effective hemoglobin combined with a left shift of the oxyhemoglobin dissociation curve (reduced p50). Overall, the tumor oxygen utilization decreased from 70% to 52% as a function of an increase in CO-Hb from 0 to 12%.     

Anemia in malignant tumor diseases. V. The relation of tumor-induced hypotransferrinemia and the degree of anemia

Transferrin is a growth factor in malignancy. In this function, transferrin is taken up into the proliferating malignant cell. The tumor-induced loss of circulating transferrin results in a hypotransferrinemia which correlates with tumor mass and proliferation rate. The cellular uptake of iron into the erythropoietic precursors depends on the presence of iron-saturated transferrin. Thus, iron utilization for the hemoglobin synthesis correlates with the transferrin concentration in blood. In 256 patients with malignancies of different histological types and different tumor extension a strong correlation was found between the degree of tumor-induced hypotransferrinemia and anemia. This correlation between transferrin concentration and hemoglobin concentration could be demonstrated in the different histological tumor entities. Tumor progression was accompanied by a progressive fall in transferrin concentration and hemoglobin concentration. By contrast, tumor remission achieved by an effective antineoplastic therapy resulted in an improvement of hypotransferrinemia and anemia. These variations in the two parameters were found to be strongly correlated. We conclude from our data that tumor-induced loss of transferrin is one of the most important factors responsible for the development of anemia in malignancy.     

Cellular kinetics of transforming growth factor-beta induced hemoglobin accumulation in the HEL erythroleukemia cell line

Transforming growth factor-beta 1 (TGF beta 1) can induce hemoglobin accumulation in a clone of the human HEL erythroleukemia cell line. This clone has previously been designated as HEL-T. The effect of TGF beta 1 was reversible and it had to be continuously present for the maximal number of cells to become positive for hemoglobin. The TGF beta 1 effect was blocked by phorbol ester and partially blocked by the calmodulin antagonist W-7, but not by dexamethasone. Simultaneous exposure to gamma-interferon, IL-1, IL-6, IL-3 and GM-CSF had no significant effect on TGF beta induced hemoglobin accumulation. However, when TGF beta was combined with TNF alpha, it was observed that there was approximately a 10-15% reduction in benzidine-positive cells. Cell-cycle analysis revealed no significant long-term alterations in any of the compartments. Analysis of the TGF beta 1 effect on 10 different HEL-T-derived clones revealed that the number of benzidine-positive cells ranged from 12 to 70% after 5 days of continuous exposure. Cell proliferation was similarly differentially affected. Another HEL cell line, designated as W-HEL, did not accumulate hemoglobin in the presence of TGF beta 1, but did have an increase in alpha-globin RNA expression.     

Differential expression of transglutaminase in human erythroleukemia cells in response to retinoic acid

Two human erythroleukemia cell lines, HEL and K562, express transglutaminase activity. The enzyme was identified as a tissue transglutaminase following chromatographic purification. All-trans-retinoic acid (10 microM) stimulated differentiation in HEL cells as judged by a 4-fold increase in hemoglobin content and a reduction in cell proliferation. The transglutaminase activity increased 9-fold. This increase in transglutaminase was the result of a pretranslational regulation of the gene as revealed by Northern blot analysis of mRNA. These changes were not a result of cell apoptosis, since parallel DNA degradation catalyzed by a Ca2(+)-dependent endonuclease could not be demonstrated. The K562 cells, in contrast, showed no transglutaminase induction following exposure to retinoic acid and displayed no changes in maturation markers or cell growth.     

Changes in plasma iron levels following Carboplatin administration

Carboplatin-induced changes in plasma iron levels and the related erythropoiesis impairment were investigated in 32 neoplastic patients for a total of 64 courses of chemotherapy. Iron showed a significant increase over pretreatment levels starting from day 1 after carboplatin administration (p< 0.001). Return to pre-treatment levels was achieved on day 14. Hemoglobin decreased significantly on day 7 (p< 0.05) and further on days 14 and 21 (p< 0.001). In patients undergoing 3 consecutive cycles of chemotherapy, basal hemoglobin before the 2nd cycle was significantly lower than before the 1st (p< 0.05), whereas before the 3rd cycle the levels were similar to those before the 2nd. Hemoglobin time-course did not differ among the three cycles. No relationship was observed between maximum iron levels and hemoglobin at minimum levels, nor between pre-treatment hemoglobin levels and severity of chemotherapy-induced subacute anemia. These results suggest that neither pre-treatment hemoglobin nor the entity of iron increase are predictive of the need of blood transfusion. Moreover, the absence of correlation between iron increase and hemoglobin decrease suggests that the toxic block on erythroid maturation is not the only mechanism with which platinum compounds interfere with iron metabolism. It is possible that the bivalent platinum ion may displace competitively iron from its binding sites, either on proteins or on cells.     

The antileukemic alkaloid fagaronine and the human K 562 leukemic cells: effects on growth and induction of erythroid differentiation

In view of new antitumor compounds which could exert their therapeutic effect through a combination of cell growth inhibition and cell maturation, we describe here the effects of a novel antileukemic alkaloid, fagaronine, on the growth and the induction of hemoglobin synthesis in the K 562 cell line. We found that fagaronine, after 3 days, reduces in a concentration dependent relationship the cell growth rate without lethality and this effect on the cell growth is irreversible. Reducing the cell growth rate by 50% (IC50 = 3 X 10(-6)M) is sufficient to induce an optimal amount of hemoglobin synthesis (75% benzidine-positive cells, 13-15 pg hemoglobin/cell) after 4 days of culture. Considering the variation of the total intracellular protein content during the response, it appears that fagaronine stimulated mainly hemoglobin synthesis, and to a lesser extent non-hemoglobin proteins. These results suggest that the novel antileukemic alkaloid, fagaronine, can be considered as a potent inducer of differentiated-associated properties in the human K 562 leukemic cells.     

Near-infrared characterization of breast tumors in vivo using spectrally-constrained reconstruction

Multi-wavelength Near-Infrared (NIR) Tomography was utilized in this study to non-invasively quantify physiological parameters of breast tumors using direct spectral reconstruction. Frequency domain NIR measurements were incorporated with a new spectrally constrained direct chromophore and scattering image reconstruction algorithm, which was validated in simulations and experimental phantoms. Images of total hemoglobin, oxygen saturation, water, and scatter parameters were obtained with higher accuracy than previously reported. Using this spectral approach, in vivo NIR images are presented and interpreted through a series of case studies (n=6 subjects) having differing abnormalities. The corresponding mammograms and ultrasound images are also evaluated. Three of six cases were malignant (infiltrating ductal carcinomas) and showed higher hemoglobin (34-86% increase), a reduction in oxygen saturation, an increase in water content as well as scatter changes relative to surrounding normal tissue. Three of six cases were benign, two of which were diagnosed with fibrocystic disease and showed a dominant contrast in water, consistent with fluid filled cysts. Scatter amplitude was the main source of contrast in the volunteer with the benign condition fibrosis, which typically contains denser collagen tissue. The changes monitored correspond to physiological changes associated with angiogenesis, hypoxia and cell proliferation anticipated in cancers. These changes represent potential diagnostic indicators, which can be assessed to characterize breast tumors.     

Induction of erythroid differentiation in murine erythroleukemia cells by N-substituted polymethylene diamides.

Various N-substituted polymethylene diamides were synthesized and tested for their potency to induce erythroid differentiation in murine erythroleukemia cells. N,N,N',N'-tetramethyl-1.6-hexane-dicarboxamide (IIc) was the most potent inducer among 15 compounds tested. The effectiveness of this compound was similar to that of hexamethylene bisacetamide (HMBA). HMBA has a different amide linkage order from that of IIc. HMBA and IIc at a concentration of 5 mM had similar effects on the cell growth rate and induced a similar frequency of benzidine-positive cells. However, hemoglobin production was 1.5 times more effective with IIc. Polymethylene diester, diamide, dihydrazide and dianilide had no effect on the induction of hemoglobin synthesis. The N-alkylated amide group appears to be required for induction of differentiation in murine erythroleukemia cells.     

Antioxidant properties of Rajgira (Amaranthus paniculatus) leaves and potential synergy in chemoprevention

In recent years there has been a substantial increase in the use of functional foods for disease control. Fruits and vegetables produce phytochemicals such as flavonoids and antioxidants which can lower oxidative stress and reduce the risk of chronic ailments like cancer. The aim of the present study was to investigate the antioxidant capacity and the possible protective effects of Amaranthus paniculatus leaves on the antioxidant defense system in Ehrlich's ascites carcinoma (EAC) -treated mice. Oral administration of the leaf extract at different doses caused a significant decrease in tumor volume, viable cell count and tumor weight and elevated the life span of EAC bearing mice. It also showed an improved antioxidant potential as evidenced by a significant increase in the cellular antioxidant defense system such as catalase, superoxide dismutase and reduced glutathione and also significantly reduced the levels of TBARS. The levels of RBC, hemoglobin and lymphocyte count were altered in EAC bearing mice and were reverted back to near normal levels after the treatment with the leaf extracts. Their adequate content of total phenolics and flavonoids, DPPH scavenging activity which further suggests that the extracts exert a significant protection against oxidative stress conditions.     

Assessment of hematologic effects and fatigue in cancer patients with chemotherapy-induced anemia given darbepoetin alfa every two weeks

The objective of this ongoing trial is to study the ability of darbepoetin alfa to reverse chemotherapy-induced anemia in cancer patients, and to relate improvement in hemoglobin with changes in fatigue and functional capacity. Eligible subjects had a nonmyeloid malignancy, were receiving multicycle chemotherapy, and were anemic, as defined by a screening hemoglobin < or = 11 g/dL. Darbepoetin alfa was administered at a starting dosage of 3 microg/kg every 2 weeks for up to eight doses (16 weeks) in an open-label, noncomparative setting. A total of 194 oncology practices contributed 1,173 subjects to this interim analysis. The mean increase in hemoglobin was 1.7 g/dL (95% CI: 1.6, 1.8) to last value on study (intent-to-treat analysis) and 2.1 g/dL (95% CI: 1.9, 2.2) for those patients receiving the full 16 weeks of therapy.The Kaplan-Meier estimate of the proportion of subjects with a hematopoietic response (increase in hemoglobin > or = 2 g/dL and/or hemoglobin value > or = 12 g/dL) was 84% (95% CI:81,86).Subjects in the lower baseline hemoglobin category (< 10 g/dL) tended to have a greater hemoglobin response during treatment. The Functional Assessment of Cancer Therapy-Fatigue (FACT-Fatigue) subscale score increased by a mean of 6.8 points (26%) during the study, and improvements in fatigue paralleled the increases observed in hemoglobin. Study treatment-related toxicity was minimal, with the most common event being injection-site pain, seen in 2% of subjects. Experience to date with an every-2-week regimen of darbepoetin alfa indicated efficacy comparable to historical experience with weekly and 3-times-weekly regimens of epoetin alfa in treating chemotherapy-induced anemia in cancer subjects.     

早期鼻咽癌66例放射治疗结果和预后因素分析

目的　分析早期鼻咽癌治疗结果和预后影响因素。方法　采用 Ｋａｐｌａｎ Ｍｅｉｅｒ 及 Ｃｏｘ 回归方法回顾性分析接受放射治疗的早期鼻咽癌６６ 例。结果　全组５ 年无瘤生存率、局部控制率、远地转移率分别为７１ ．６ ％ ,８７ ．８ ％ 和１７ ．４ ％ 。鼻咽肿物放射治疗 Ｄ Ｔ ＜ ４０ Ｇｙ 消退者在生存率和远地转移的控制上均差于 Ｄ Ｔ ＞ ４０ Ｇｙ 消退者,二者差异有显著性（ Ｐ＜ ０ ．０５） ;在局部控制率上,二者差异无显著性;外周血血红蛋白低于１１０ ｇ／ Ｌ 者无瘤生存率、局部控制率均差于血红蛋白＞ １１０ ｇ／ Ｌ 者,且差异有显著性（ Ｐ＜ ０ ．０５） ;而二者远地转移率差异无显著性（ Ｐ＞ ０ ．０５） 。结论　鼻咽肿物对射线越敏感越容易发生远地转移,生存率越差;血红蛋白水平越低,局部控制率和生存率越差。     

Anemia in cervical cancer patients

Iron deficiency and tumor bleeding are common causes of anemia in cervical cancer. Anemia has a negative prognostic influence, and its correction is thought to improve prognosis; therefore, most patients are treated with either transfusion and/or erythropoietin. At present little is known about the value of iron stores replenishment to increase hemoglobin levels in this setting. Untreated cervical cancer patients with a hemoglobin <12 g/dL were randomized to intramuscular iron or to transfusion. Iron dose was calculated according to [weight (kg)x(15--actual Hb)x2.4]+500 and administered by injections of 200 mg daily. In both arms, patients who did not achieve at least 10 g/dL hemoglobin before or during chemoradiation were transfused. Patients received standard pelvic radiation plus six weekly doses of cisplatin. Hematic counts were performed before starting chemoradiation and weekly thereafter. Fifteen patients were studied; six were assigned to iron and nine to transfusion. Mean basal hemoglobin levels were 9.9 and 9.5 g/dL respectively. Total iron, saturation index, binding capacity, and ferritin were within normal limits, although there was a high variability among the patients. The mean total dose of iron administered was 1229 mg. Two weeks after randomization, hemoglobin increased to 10.9 and 10.2 g/dL respectively. At wk 1 of treatment and thereafter, levels were higher in the iron arm, in whom the values were close or higher than 12 g/dL (p=0.03). The median number of units transfused were 0 in the iron group and 2 in the transfusion (p=0.02) arm. Parenteral iron seems to be effective to increase hemoglobin in cervical cancer patients.