Anaemia of cancer: an overview of mechanisms involved in its pathogenesis.

Anaemia is a common complication in cancer patients. The decrease in haemoglobin is associated with an impaired quality of life, poorer response to therapy and worse prognosis. Numerous factors are involved in the physiopathology of cancer-related anaemia. Some factors such as bleeding, bone marrow infiltration, the effects of chemoradiotherapy and associated nutritional deficiencies are related to the disease itself. In addition, the interaction of the immune system with iron metabolism and erythropoiesis has been shown to be an important factor in the development of anaemia in cancer patients and can be seen in the action of several cytokines on different iron-homeostasis and erythrocyte-cell-production pathways. Some inhibitory cytokines, such as tumour necrosis factor-alpha and interleukin-1, act on the suppression of erythroid precursor cells and erythropoietic production and response; others, such as interleukins 1 and 6 and hepcidin, impair iron metabolism, causing iron to be diverted from erythropoiesis and retained within the reticuloendothelial system. The main mechanisms involved in the development of cancer-related anaemia are discussed in this review.

Anaemia in patients with cancer: role of inflammatory activity on iron metabolism and severity of anaemia.

Hepcidin has been proposed as an important factor in the pathogenesis of the anaemia of chronic disease (ACD). The aim of this study was to assess the relationship between anaemia and inflammatory activity in patients with solid tumours. Patients were classified as having iron deficiency anaemia (IDA) (hypoferremia and hypoferretinemia), ACD (hypoferremia, normal or increased serum ferritin) and anaemia related to cancer (ARC) (no abnormalities in iron status). Serum pro-hepcidin, IL-6, C-reactive protein (CRP) and iron status parameters were measured using commercial kits. CRP and IL-6 levels were significantly higher in patients with ACD when compared to IDA, ARC and non anaemic patients (P < 0.005). Serum pro-hepcidin levels were not different among all studied groups (P = 0.138). A negative correlation was observed between haemoglobin and serum ferritin, CRP and IL-6 levels only in group of ACD. Serum pro-hepcidin concentrations were not correlated with degree of anaemia or iron metabolism parameters. According to our results the inflammatory activity represented by high levels of IL-6 and CRP are involved in the pathogenesis of ACD, probably due to the action of inflammation on iron metabolism, but not in ARC. It was not possible to demonstrate a significant effect of pro-hepcidin on the anaemia in cancer patients.

Evaluation of reticulated platelets in patients with sickle cell diseases.

BACKGROUND AND PURPOSE: Reticulated platelet (RP) count provides an estimate of thrombopoiesis. The objective was to evaluate RP in patients in different stages of sickle cell disease (SCD) and to determine the relationship between interleukin-6 (IL-6), interleukin-3 (IL-3) and thrombopoietin (TPO) and RP count and degree of activation. METHODS: Eighty-nine adult patients with SCD were studied: 38 were in the steady state, 27 in hemolytic crisis (HC) and 24 in vaso-occlusive crisis (VOC). RPs and activated platelets were analyzed by flow cytometry. Soluble P-selectin, IL-6, IL-3 and thrombopoietin (TPO) levels were measured by ELISA tests. RESULTS: The patients in VOC had a higher absolute number of RPs and CD62P+ platelets than did the control group or patients in the steady state. A significant correlation was observed between the absolute number of CD62P+ platelets and RPs in patients in the steady state, HC and VOC. In the steady-state group of patients, the level of soluble P-selectin was found to be dependent on the RP values. IL-3 and TPO serum levels were higher in patients in the steady state, HC and VOC than in the control group. IL-6 serum levels were higher in HC and VOC patients than in the control group and higher in patients in the steady state than in the VOC group. CONCLUSION: Our results suggest that PRs contribute to the vaso-occlusive process in sickle cell disease. Increased interleukin serum levels probably indicate that inflammatory process is involved in the vascular-occlusive phenomenon. However, it appears that these inflammatory mediators do not have an effect on thrombopoiesis in sickle-cell-disease patients.

Hematopoietic progenitor cells (HPC) and immature reticulocytes evaluations in mobilization process: new parameters measured by conventional blood cell counter.

Monitoring the timing of leukapheresis in peripheral blood stem cells (PBSC) mobilization is an important clinical decision that requires an accurate analytical tool. The present study assessed hematopoietic progenitor cells (HPC) and immature reticulocyte fraction (IRF) counts provided by a routine automated blood counter as potential parameters for predicting the appropriate time for harvesting. The HPC and IRF values were compared with white blood cell (WBC) and CD34+ cell counts obtained by flow cytometry in 30 adult patients with hematological malignancies undergoing PBSC mobilization. It was observed that there was a significant correlation between HPC counts and CD34(+) cells in peripheral blood counts (r=0.61, P=0.0003) and between the number of HPC and CD34+cells collected by leukapheresis (r=0.5733, P=0.0009). Comparing HPC, IRF, WBC, and CD34+ cells parameters as a sign of hematological recovery showed that the raise in immature reticulocytes counts preceded the increase of WBC (P=0.0002), HPC (P=0.0001), and CD34(+) (P=0.0001) cells in peripheral blood counts. According to our results, HPC and IRF parameters may be integrated into clinical protocols to evaluate the timing of leukapheresis. IRF, as previously demonstrated in bone marrow transplantation, is the earliest sign of hematopoietic recovery in mobilization process.

High levels of neopterin and interleukin-3 in sickle cell disease patients.

Sickle cell disease (SCD) is recognized as a chronic inflammatory condition. Cytokines are released in response to stress or pathological situations, and influence hematopoiesis. The aim of this study was to evaluate interleukin-3 (IL-3), interferon-gamma (IFN-gamma), and neopterin (NP) levels in steady-state patients with sickle cell anemia (SS) (n = 35) and SC hemoglobinopathy (n = 15) in order to verify the possible action of those cytokines and NP on iron metabolism and hematopoiesis. Serum IL-3 concentration was higher in SS and SC groups than in controls, whereas IFN-gamma levels did not differ among groups. SS patients presenting hemoglobin fetal (HbF) >or=8.5% had significantly higher IL-3 levels than those with HbF <8.5% (P = 0.0338). No correlation was observed among inflammatory and iron metabolism parameters. Significant correlations were observed between IL-3 and Hb levels (r = 0.4633, P = 0.0457), and IL-3 and HbF levels (r = 0.6011, P = 0.0065). A negative correlation was observed between IL-3 and reticulocyte counting (r = -0.4632, P = 0.0457) only in the SS group. NP levels were significantly higher in the SS and SC groups than in controls, but did not differ between SS and SC. No correlation was observed between NP and iron metabolism parameters. These data suggest that IL-3 stimulates hematopoiesis, and that SS patients, even in steady state, have macrophage/monocyte activation (represented by high levels of NP) that probably contributes to their chronic inflammatory condition.

Haptoglobin phenotype appears to affect the pathogenesis of American trypanosomiasis.

In Latin America, 16 million-18 million people are thought to be infected with Trypanosoma cruzi, the parasite that causes American trypanosomiasis. The pathophysiology of this disease, particularly that of its chronic phase, has yet to be fully elucidated. The major function of haptoglobin, an acute-phase plasma protein found in three different phenotypes (Hp1-1, Hp2-1 and Hp2-2), is to bind to free haemoglobin and so prevent the accumulation of reactive hydroxyl radicals and renal damage. The haptoglobin phenotype present can influence the severity and progression of many diseases, including infectious ones. The aim of the present study was to see if any haptoglobin phenotype could be associated with any of the various clinical forms of American trypanosomiasis, and so explore the possibility that haptoglobin and iron metabolism have a role in the pathophysiology of this disease. The Brazilian subjects investigated were either suffering from the 'indeterminate' (N=16), chronic cardiac (N=34), chronic digestive (N=13) or chronic 'combined' (i.e. cardiac plus digestive; N=29) forms of the disease or were apparently healthy blood donors from the same region as the patients (N=197). Haptoglobin phenotypes were determined by polyacrylamide-gel electrophoresis. Among the iron-related parameters investigated in the patients, only total iron-binding capacity and the serum concentration of haptoglobin differed significantly with haptoglobin phenotype. Compared with its frequency in the healthy controls, the Hp2-2 phenotype was much more frequent in the patients with any form of American trypanosomiasis, in the patients with the indeterminate form of the disease, and in the patients with the chronic combined form (P

Reticulocyte evaluation in alpha(+)-thalassemia.

Although it is almost certain that alpha(+)-thalassemia protects against malaria, the mechanisms for that are still unknown. It has been suggested that an increased number of young circulating red blood cells in alpha(+)-thalassemic children, as a result of some degree of ineffective erythropoiesis, could be related to the high frequencies of the alpha(+)-thalassemic allele in malaria endemic areas. Reticulocyte evaluation in this condition, however, has been poorly performed so far. Our objective was to determine the reticulocyte number and maturation degree, in addition to the soluble transferrin receptor and serum erythropoietin levels, in alpha(+)-thalassemia heterozygotes, comparing them with normal alpha-genotype controls. One hundred twenty-one alpha(+)-thalassemia carriers (-alpha(3.7)/alphaalpha) and 249 controls (alphaalpha/alphaalpha), all of them with normal serum ferritin levels, were subclassified according to age (1-5, 6-10, 11-15, 16-20, and over 20 years old). Reticulocyte analyzes were carried out by flow cytometry and sTfR and s-Epo levels determined by immunonephelometry and chemiluminescence, respectively. The comparisons did not show any significant difference between thalassemics and controls regarding the reticulocyte parameters [percentages and absolute values, P = 0.2643 and 0.5421; high, medium, and low maturation degree, P = 0.2579, 0.2196, and 0.4192; RET maturity index (RMI), P = 0.2471, respectively], as well as the s-Epo levels (P = 0.5711). The sTfR concentrations were higher in the thalassemic group (P = 0.0001), but statistical significance was due only to the 1-5 and over 20 subgroups (P = 0.0082 and 0.0436, respectively). The results found here are compatible with a compensated erythropoiesis and do not confirm the hypothesis mentioned above.

Platelet larger cell ratio (P-LCR) in patients with dyslipidemia.

We have described preliminary results of platelet larger cell ratio (P-LCR) analysis, provided by an automated hematology analyzer in patients with hypercholesterolemia and/or hypertriglyceremia. P-LCR values were significantly higher in patients (n = 41) than in control normal group (n = 20) (P < 0.0001). Larger platelets are more reactive and contribute to vasooclusive events in patients with dyslipidemia. P-LCR may be used as an indicator of risk factor for thromboembolic ischemic events.

Effect of hydroxyurea on G gamma chain fetal hemoglobin synthesis by sickle-cell disease patients

Hydroxyurea is used for sickle-cell disease patients in order to increase fetal hemoglobin synthesis and consequently decrease the severity of pain episodes. Fetal hemoglobin, which is formed by gamma-globin chains A and G, is present in a constant composition throughout fetal development: about 75 percent of Ggamma and 25 percent of Agamma. In contrast, adult red cells contain about 40 percent of Ggamma and 60 percent of Agamma. In the present study, we analyzed the effect of hydroxyurea induction on the gamma chain composition of fetal hemoglobin in 31 sickle-cell disease patients treated with hydroxyurea. The control group was composed of 30 sickle-cell disease patients not treated with hydroxyurea in clinical steady state. The patients were older than 13 years and were not matched for age. All patients were seen at Hemocentro/UNICAMP and Boldrini Infantile Center, Campinas, SP, Brazil. The levels of total hemoglobin were significantly higher in patients treated with hydroxyurea (mean ± SD, 9.6 ± 2.16 g/dl) than in untreated patients (8.07 ± 0.91 g/dl). Fetal hemoglobin levels were also higher in treated patients (14.16 ± 8.31 percent) than in untreated patients (8.8 ± 4.09 percent), as was the Ggamma/Agamma ratio (1.45 ± 0.78 vs 0.98 ± 0.4, P < 0.005). The increase in the Ggamma/Agamma ratio in patients treated with hydroxyurea suggests the prevalence of a pattern of fetal hemoglobin synthesis, whereas patients not treated with hydroxyurea maintain the adult pattern of fetal hemoglobin synthesis. Because no correlation was observed between the Ggamma/Agamma ratio and total hemoglobin or fetal hemoglobin levels, the increase in Ggamma chain synthesis may not imply a higher production of hemoglobin

Effect of hydroxyurea on G gamma chain fetal hemoglobin synthesis by sickle-cell disease patients.

Hydroxyurea is used for sickle-cell disease patients in order to increase fetal hemoglobin synthesis and consequently decrease the severity of pain episodes. Fetal hemoglobin, which is formed by gamma-globin chains A and G, is present in a constant composition throughout fetal development: about 75% of Ggamma and 25% of Agamma. In contrast, adult red cells contain about 40% of Ggamma and 60% of Agamma. In the present study, we analyzed the effect of hydroxyurea induction on the gamma chain composition of fetal hemoglobin in 31 sickle-cell disease patients treated with hydroxyurea. The control group was composed of 30 sickle-cell disease patients not treated with hydroxyurea in clinical steady state. The patients were older than 13 years and were not matched for age. All patients were seen at Hemocentro/UNICAMP and Boldrini Infantile Center, Campinas, SP, Brazil. The levels of total hemoglobin were significantly higher in patients treated with hydroxyurea (mean +/- SD, 9.6+/-2.16 g/dl) than in untreated patients (8.07+/-0.91 g/dl). Fetal hemoglobin levels were also higher in treated patients (14.16+/-8.31%) than in untreated patients (8.8+/-4.09%), as was the Ggamma/Agamma ratio (1.45+/-0.78 vs 0.98+/-0.4, P < 0.005). The increase in the Ggamma/Agamma ratio in patients treated with hydroxyurea suggests the prevalence of a pattern of fetal hemoglobin synthesis, whereas patients not treated with hydroxyurea maintain the adult pattern of fetal hemoglobin synthesis. Because no correlation was observed between the Ggamma/Agamma ratio and total hemoglobin or fetal hemoglobin levels, the increase in Ggamma chain synthesis may not imply a higher production of hemoglobin.

Reticulocyte parameters and hemoglobin F production in sickle cell disease patients undergoing hydroxyurea therapy.

Hemoglobin F (HbF) is an effective inhibitor of HbS polymerization. Hydroxyurea (HU) is used to increase HbF synthesis and improve the clinical course of sickle cell disease (SCD) patients. We studied a series of laboratory parameters concerning HbF production and reticulocyte response, and compared data between two groups: 1) 13 SCD patients treated with HU, and 2) 33 untreated SCD patients. Higher values of Hb concentration, mean cell volume (MCV), mean cell hemoglobin (MCH), mean reticulocyte volume (MRV), HbF concentration, percentage of F-cells, and amount of HbF/F-cells were observed in the treated group of patients. There was no correlation between Hb and HbF elevations. The reticulocyte count, immature reticulocyte count, mean fluorescence index (MFI), and neutrophil count were significantly lower in treated patients. Taken together, these findings suggest that a decreased hemolytic process occurred in patients undergoing HU treatment. There was a significant correlation between MCV and HbF, between MRV and HbF, and between MRV and F-cell in patients taking HU. These data indicate that macroreticulocytes correspond to F-reticulocytes, and that an increase in MRV in SCD patients using HU may be an indirect signal of F-cell production. The concentration of HbF/F-cells was higher in patients treated with HU, but this increase apparently was independent of F-cell production. Reticulocyte (RTC) parameters, as assessed by hematological analyzers, may be useful for following erythropoietic changes in patients receiving HU, and can indirectly indicate HbF and F-cell production induced by HU therapy.

Immature reticulocytes as an early predictor of engraftment in autologous and allogeneic bone marrow transplantation.

The purpose of this study was to evaluate reticulocyte parameters by means of flow cytometric reticulocyte counting in a group of patients who had undergone autologous and allogeneic bone marrow transplantation (BMT). The pattern of reticulocyte response and the predictive value of absolute neutrophil count (ANC), platelet count, number of CD34+ cell infused and graft source for reticulocyte response were studied. We compared absolute reticulocyte count (RetAbs), mean fluorescence index (MFI) and mean reticulocyte volume/mean corpuscular volume (MRV/MCV) ratio with conventional criteria (ANC and platelet count) in 22 allogeneic and 20 autologous BMT recipients. An abrupt increase in MRV/MCV ratio or a rise in MFI value were the earliest signs of erythropoietic recovery following allogeneic transplantation (63.6 and 22.8% of cases, respectively). In 13.6% of the cases, both parameters were observed simultaneously. All but three autologous transplant recipients showed changes in reticulocyte parameters earlier than ANC recovery. Granulocyte recovery and peripheral blood progenitor cells (PBPC) graft were predictive variables for RetAbs response in allogeneic transplant recipients. In the autologous group, predictive variables for RetAbs response were a high number of CD34+ infused cells and platelet recovery. An increase in the immature reticulocyte population is the earliest sign of haematopoietic recovery following BMT.

[Clinical aspects and complete blood counts in children exposed to HIV-1: comparison between infected patients and seroreverters] / Aspectos clínicos e o hemograma em crianças expostas ao HIV-1: comparação entre pacientes infectados e soro-reversores.

OBJECTIVE: To analyze the evolution of clinical and hematological aspects of children exposed to the vertical transmission of HIV-1, comparing infected patients with uninfected ones or seroreverters. METHODS: Prospective, descriptive, longitudinal study. We analyzed 79 children born from HIV-1 infected mothers, under clinical follow up from March, 1996 until November, 1997, at the Immunodeficiency division of the Hospital de Clínicas da Unicamp (State University Hospital of Campinas). RESULTS: Failure to thrive was observed in both groups, but was greater among seroreverters. Among the infected children, 23 mothers did not use AZT during pregnancy, 16 of them (61.5%) had been breastfed, four were classified into clinical category N, seven into A and fifteen into B. Clinical manifestations in patients younger than one year were seen in 18 infected children (69.2%). Anemia was observed in 73.1% of the infected group and in 41.5% of the seroreverters (P<0.008). The comparison between the groups showed that the most common hematologic alterations in the infected children was microcytosis and hypochromia (P<0.05), lymphopenia between 15 and 18 months (P<0.05), monocytosis between 9 and 12 months (P<0.05) and a tendency towards high ferritin levels, with no statistical significance. CONCLUSIONS: Microcytic and hypochromic anemia were observed in both groups: iron deficiency in the uninfected children, and chronic disease anemia in the infected ones. The infected children presented with monocytosis and lymphopenia at an earlier stage.

Soluble transferrin receptor in sickle cell diseases: correlation with spleen function.

OBJECTIVE: To correlate spleen function with soluble transferrin receptor (sTfR) levels and red cell ferritin (RCF) values in patients with sickle cell diseases. DESIGN: Prospective study. LOCATION: University Hospital, School of Medical Sciences, State University of Campinas; a tertiary hospital. PARTICIPANTS: 60 patients with sickle cell diseases, in a steady state, who had not received blood transfusions for 3 months; 28 normal individuals with no clinical or laboratory signs of anemia. MEASUREMENTS: Determination of serum iron, transferrin iron-binding capacity, serum ferritin, RCF and sTfR. Evaluation of spleen function: erythrocytes with pits were quantified. RESULTS: Patients with sickle cell anemia had sTfR levels significantly higher than in normal individuals or those with HbSC (p=0.0001) and there was an inverse correlation between sTfR and fetal Hb (p=0.0016). RCF values were significantly higher in sickle cell anemia patients than in normal individuals or those with HbSC (p=0.0001), and there was a correlation between RCF and pitted erythrocytes (p=0.0512). CONCLUSION: The association between sTfR and fetal Hb confirms the contribution of fetal Hb to improving the hemolytic state by minimizing the consequent reactive erythrocyte expansion. High sTfR levels are not related to the degree of spleen function deficiency seen in sickle cell disease patients. The deficiency in the exocytosis process of the spleen occurring in sickle cell anemia patients may contribute to their accumulation of RCF.

Comparison of red cell distribution width and a red cell discriminant function incorporating volume dispersion for distinguishing iron deficiency from beta thalassemia trait in patients with microcytosis.

The red cell distribution width (RDW), and another red cell discriminant function incorporating RDW (MCV2 x RDW/Hgb x 100) were determined in a group of 30 patients with iron deficiency anemia, 30 patients with beta thalassemia trait, and 30 normal subjects. Both RDW and (MCV2 x RDW/Hgb x 100) mean values were significantly higher in iron deficiency anemia than in beta thalassemia trait (p < 0.001). Taking RDW equal or above 21.0 percent among microcytic anemia patients, we identified correctly 90.0 percent of patients with iron deficiency anemia. The sensitivity and specificity of the test were 90.0 percent (IC 95 percent: 0.75-0.98) and 77.0 percent (IC 95 percent: 0.60-0.88), respectively. RDW values below 21.0 percent identified correctly 77.0 percent of beta thalassemia trait with a sensitivity and a specificity of 77.0 percent (IC 95 percent: 0.60-0.88) and 90.0 percent (IC 95 percent: 0.75-0.96), respectively. Taking values of (MCV2 x RDW/Hgb x 100) above and below 80.0 percent as indicative of iron deficiency and beta thalassemia trait, respectively, we identified correctly 97.0 percent of those patients in each group. Both sensitivity and specificity were 97.0 percent (IC 95 percent: 0.84-0.99). These results indicated that the red cell discriminant function incorporating volume dispersion (MCV2 x RDW/Hgb x 100) is a highly sensitive and specific method in the initial screening of patients with microcytic anemia and is better than RDW in differentiating iron deficiency anemia from beta thalassemia trait.

Hb Lepore Baltimore (delta 50Ser beta 86Ala) identified by DNA analysis in a Brazilian family.

Genomic DNA from two Brazilian hemoglobin (Hb) Lepore heterozygotes of Italian ancestry have been studied in order to identify the Hb Lepore type and to sequence the breakpoint region. The two genes were sequenced after PCR amplification and had the delta globin sequence up to exon 2 codon 68 while the first specific base for the beta globin gene was at codon 86 of the second exon; between the two ends, they had 51 base pairs in common with the delta and beta globin genes. These data indicate that the mutation was of the Hb Lepore Baltimore type. The Lepore chromosome haplotype was different from that previously described in members of a Spanish family with Hb Lepore Baltimore. These data suggest that independent mutations have given rise to Hb Lepore Baltimore in different regions of the world.

Diferenciaçäo entre talassemia beta-heterozigótica e anemia ferropriva / Differentiation between heterozygotic beta-thalassemia and iron deficiency anemia

Várias funcöes discriminantes baseadas nos parâmetros hematimétricos fornecidos pelos contadores eletrônicos de células têm sido desenvolvidas e utilizadas nas diferenciaçäo de várias formas de anemia. A eficiência de alguns desses critérios comumente empregados na distinçäo entre a talassemia beta-heterozigótica e a anemia ferropriva foi testada em 192 indivíduos talassêmicos e em 72 adultos com anemia ferropriva. Foram avaliados: 1) funçäo discriminante de England e Fraser: VCM - (5xHb) - H - 8,4;2) HCM/H; 3) relaçäo de Mentzer: VCM/H; 4) (VCM)² x HCM; e 5) número de hemácias. Os resultados obtidos demonstraram falha na distinçäo entre estas duas entidades, em percentagens variáveis de 9 a 90//, sendo que a equaçäo de England e Fraser demonstrou os resultados mais satisfatórios, enquanto a relaçäo de Mentzer apresentou maiores índices de erro no diagnóstico da anemia ferropriva. Assim, embora algumas dessas fármulas possam ser utilizadas em programa de triagem, em casos individuais a diferenciaçäo entre talassemia beta-heterozigótica e anemia ferropriva deve ser sempre realizada com dosagens de HbA2, ferro sérico, capacidade máxima de transporte de ferro e ferritina

[Differentiation between heterozygotic beta-thalassemia and iron deficiency anemia]. / Diferenciação entre talassemia beta-heterozigótica e anemia ferropriva.

Several laboratory tests have been proposed for the differentiation of beta thalassemia from iron deficiency, including decision functions based on red blood cells indices generated by electronic cell counters. The accuracy of these screening methods was assessed in 192 patients with microcytosis known to be secondary to beta thalassemia minor and 72 patients with iron deficiency. The functions evaluated were: 1) discriminant function of England an Fraser: MCV--(5xHb)--RBC--8.4; 2) ratio of MCH/RBC; 3) ratio of MCV/RBC; 4) ratio of (MCV)2 x MCH and 5) the erythrocyte count. The discriminant function of England and Fraser showed the highs, percentage of correct distinction between iron deficiency and beta thalassemia minor, although diagnosis errors occurred in 10%. Mentzer ratio MCV/RBC detected all cases of beta thalassemia but was a poor index for iron deficiency detection. The tested discriminant function, with exception of the Mentzer ratio, although not sufficiently accurate for definitive diagnosis, appears to be a useful technique in the initial screening of patients with microcytosis.