Benzene-induced Aplastic Anemia Research Articles

Effects of Cyclosporine and Azacitidine on Some Hematologic and ‎Biochemical Parameters of Benzene-Induced Aplastic Anemia in Rats

Aplastic anemia, marked by deficiencies in hematopoietic stem cells, leads to peripheral blood pancytopenia and hypocellular bone ‎marrow. This study aimed to evaluate the therapeutic efficacy of cyclosporine and azacitidine, administered either alone or in combination, in rats with benzene-induced aplastic ‎anemia, focusing on restoring normal blood cell levels and preventing disease complications. Thirty adult female Wistar rats ‎(Rattus ‎norvegicus)‎ were randomly divided into five groups: negative control (C-, untreated), positive control (C+, induced aplastic anemia with ‎distilled water), cyclosporine-treated (CsA, 5.86 mg/kg), azacitidine-treated (Aza, 5.75 mg/kg), and combination-treated (CsA+Aza, 3.68 ‎mg/kg each). Benzene (1940 mg/kg) was administered orally for fifteen days to induce aplastic anemia. Post a 30-day treatment period, ‎evaluations included differential WBC and reticulocyte counts, serum IL-2 levels, and alkaline phosphatase (ALP) activity. Results ‎showed significant improvements in WBC% and reticulocyte% in all treated groups compared to the C+ group, with the combination-‎treated group showing the highest enhancement. IL-2 levels in the combination group were significantly reduced compared to other ‎treatment groups, aligning closely with the negative control. The ALP activity was significantly higher in both the cyclosporine and ‎azacitidine-treated groups compared to the positive control, with the combination group showing a marked increase over the azacitidine ‎group but no significant difference from the cyclosporine group and negative control.‎ In conclusion, the study demonstrates the potential therapeutic benefits of cyclosporine and azacitidine in treating benzene-induced ‎aplastic anemia in rats. The combination therapy, in particular, showed improved efficacy in all tested parameters, suggesting a potential ‎strategy for dose reduction and toxicity mitigation.

Let-7e-5p, a promising novel biomarker for benzene toxicity, is involved in benzene-induced hematopoietic toxicity through targeting caspase-3 and p21

Benzene is a common industrial chemical and environmental pollutant. However, the mechanism of hematotoxicity caused by exposure to low doses of benzene is unknown. Let-7e-5p pathway regulatory networks were constructed by bioinformatics analysis using a benzene-induced aplastic anemia (BIAA) mouse model. The MTT assay, EdU staining, flow cytometric analysis, dual luciferase reporter gene assay, and RIP assay were utilized to evaluate the effects of benzoquinone (1,4-BQ) on let-7e-5p pathway. This study consisted of 159 workers with a history of low-level benzene exposure and 159 workers with no history of benzene exposure. After the confounding factors were identified, the associations between let-7e-5p expression and hematotoxicity were assessed by multiple linear regression. Furthermore, we used four machine learning algorithms (decision trees, neural network, Bayesian network, and support vector machines) to construct a predictive model for detecting benzene-causing hematotoxicity in workers. In this study, compared with respective controls, let-7e-5p expression was decreased in BIAA mice and benzene-exposed workers. After 1,4-BQ exposure, let-7e-5p overexpression negatively regulated caspase-3 and p21 expression, protected cells from apoptosis, and facilitated cell proliferation. RIP assays, and dual luciferase reporter gene assays confirmed that let-7e-5p could target p21 and caspase-3 and regulate the cell cycle and apoptosis. The support vector machines classifier achieved the best prediction of benzene-induced hematotoxicity (prediction accuracy = 88.27, AUC = 0.83) by statistically characterizing the internal dose of benzene exposure and the oxidative stress index, as well as the expression levels of let-7e-5p pathway-related genes in benzene-exposed workers. Let-7e-5p may be a potential therapeutic target of benzene-induced hematotoxicity, provide a basis for evaluating the health hazards of long-term and low-dose benzene exposure in workers, and supply a reference for revising occupational health standards.

LincRNA-p21 promotes p21-mediated cell cycle arrest in benzene-induced hematotoxicity by sponging miRNA-17-5p

Benzene is widely employed in manufacturing and causes hematotoxic effects and leukemia in humans. A long intergenic noncoding RNA (lincRNA)-microRNA (miRNA)-mRNA coexpression and competing endogenous RNA (ceRNA) regulatory network was constructed by bioinformatics analysis based on a benzene-induced aplastic anemia (BIAA) mouse model. In this population-based study, we observed a trend consistent with that in the BIAA mice: lincRNA-p21 and p21 were upregulated, while miRNA-17-5p expression was downregulated in benzene-exposed workers. Moreover, multiple linear regressions indicated that lincRNA-p21 was negatively associated with white blood cell (WBC) counts. Predictive thresholds of hematotoxicity were identified by ROC curve analysis with S-phenylmercapturic acid (SPMA) and lincRNA-p21 showing a better predictive ability than the other parameters and the combination of SPMA and lincRNA-p21 exhibiting the highest predictive value for hematotoxicity. LincRNA-p21 was predominantly present in the cytoplasm of bone marrow cells (BMCs) and K562 cells as assessed by fluorescence in situ hybridization (FISH). Upon exploring the underlying mechanism by which lincRNA-p21 mediates benzene-induced hematotoxicity, we observed that the negative regulation of 1,4-benzoquinone (1,4-BQ) on cell cycle arrest and inhibition of K562 cell proliferation was partially relieved by lincRNA-p21 knockdown, which can inhibit the expression of P21 and thereby suppress the toxic effects of 1,4-BQ. Finally, dual-luciferase reporter gene and RIP assay showed that, by acting as a sponge, lincRNA-p21 reduced the activity of miRNA-17-5p and consequently increased the expression of p21. In conclusion, our research suggested that benzene induces hematotoxicity via the lincRNA-p21/miRNA-17-5p/p21 signaling which might contribute to the underlying mechanism of lincRNA-p21 in benzene-induced hematotoxicity. Therefore, lincRNA-p21 can serve as a potential biomarker for the early detection of hematopoiesis inhibition in individuals with long-term exposure to low-dose benzene.

MiR-146b-5p regulates bone marrow mesenchymal stem cell differentiation by SIAH2/PPARγ in aplastic anemia children and benzene-induced aplastic anemia mouse model

ABSTRACT This study aimed to reveal the mechanism of miR-146b-5p in the differentiation of bone marrow mesenchymal stem cells (BMSCs) derived from children with aplastic anemia (AA). Here, we found that miR-146b-5p was highly expressed in BMSCs from children with AA, and the BMSCs surface markers expressions in BMSCs derived from children with AA and the healthy controls exerted no significant differences. Besides, the overexpression of miR-146b-5p in normal human-derived BMSCs promoted the adipogenic differentiation of BMSCs. Furthermore, miR-146b-5p negatively regulated SIAH2 luciferase activity, and the interference with miR-146b-5p reduced the stability of PPARγ protein and inhibited SIAH2-mediated ubiquitination of PPARγ protein. Besides, the interference with miR-146b-5p was beneficial for ameliorating AA in a mouse model of AA. Overall, our results found that miR-146b-5p was highly expressed in BMSCs from children with AA, and our further studies indicated that miR-146b-5p improved AA via promoting SIAH2-mediated ubiquitination of PPARγ protein.

Epimedium Polysaccharide Ameliorates Benzene-Induced Aplastic Anemia in Mice.

Benzene (BZ) is an important occupational and environmental pollutant. Exposure to BZ may cause aplastic anemia which is characterized as bone marrow hematopoietic failure. In order to reduce the harmful effects of this pollutant, it is necessary to identify additional preventative measures. In this study, we investigated the protective effects of epimedium polysaccharide (EPS), a natural compound with antioxidant and immune-enhancing potency, on aplastic anemia induced by benzene exposure in mice. Male CD-1 mice were randomly divided into five groups including control, BZ (880 mg/kg), LE (EPS low-dose, 20 mg/kg + BZ), ME (EPS middle-dose, 100 mg/kg + BZ), and HE (EPS high-dose, 200 mg/kg + BZ) groups. Animals were exposed to BZ by subcutaneous injection in the presence or absence of EPS via oral administration. All mice were treated 3 times a week for 8 consecutive weeks to develop a mouse model of benzene-induced aplastic anemia (BIAA). Results showed that BZ induced a significant decrease in both white and red blood cells, platelet counts, and hemoglobin level compared with that in the control group (p < 0.01). Treatment of EPS led to a protective effect against these changes particularly in the highest-dose group (HE, p < 0.01). EPS also recovered the decreased number of nucleated cells in peripheral blood cell smears and femur biopsies by BZ exposure. The increased level of reactive oxygen species (ROS) in bone marrow mononuclear cells (BMMNCs) in mice from the BZ group was significantly lower (p < 0.01) in the mice from the highest concentration of EPS (HE) group when compared with that from the control group. In addition, BZ exposure led to a significant increase in the apoptosis rate in BMMNCs which was prevented by EPS in a dose-dependent manner (p < 0.01). The antiapoptosis effect of EPS was through reversing apoptotic proteins such as BAX, Caspase-9 and Caspase-3, and Bcl-2. Finally, EPS treatment partially restored the levels of T cells and the different subtypes except CD80+ and CD86+ compared with the BZ group (HE, p < 0.05). These results suggest that EPS has protective effects against BIAA via antioxidative stress, immune modulation, and antiapoptosis mechanisms.

Effects of Angelica Polysaccharide on telomere length in mice with benzene-induced aplastic anemia

Effects of Angelica Polysaccharide on telomere length in mice with benzene-induced aplastic anemia

Benzene's toxicity: a consolidated short review of human and animal studies by HA Khan

Khan's review is a brief summary of the complex field of study revolving around bone marrow toxicity and leukemogenesis observed in people chronically exposed to benzene. These comments are intended to demonstrate the use of the Kahn review as a launching pad for an in-depth analysis of the several related areas that must be fully explored to understand benzene-related diseases. The accumulated evidence demonstrates that benzene-induced bone marrow damage results from the production of hematotoxins that are metabolic products of benzene metabolism. The metabolism of benzene is described with respect to the formation benzene metabolites with emphasis on phenol and hydroquinone, which are the major metabolites, the significance of the formation of glutathione conjugates, the activity of NAD(P)H:quinone oxidoreductase (NQO1), and the ring opening products. Results are shown suggesting that oxidative stress induced by benzene metabolites is likely to be a significant factor in damaging DNA in bone marrow cells. Although a variety of effects on bone marrow can be demonstrated it is not yet clear which metabolites are most important in either benzene-induced aplastic anemia or leukemia. Benzene metabolism alone is insufficient to fully describe benzene toxicity. The impact of benzene metabolites on bone marrow cells must be fully explored to determine how benzene exposure can result in decreased viability or genetic toxicity to cells in the bone marrow.

A Novel Ex Vivo Immunotherapy for Some Hematopoietic and Blood Deficient Disorders.

We have developed a novel cell-based immunotherapy for treatment of some hematopoietic and blood deficient diseases such as aplastic anemia, chemotherapy-induced severe myelosuppression, idiopathic thrombocytopenia purpura and autoimmunity-induced cytopenia. Autologous and/or allogeneic peripheral blood mononuclear cells were cultured in vitro with a combination of cytokines and a calcium mobilizing agent for 2 days before given to patients via intravenous infusion. The immunotherapy has been shown to have potent activities in stimulating multi-lineage hematopoiesis and blood production including platelet production, which remains a major clinical problem to be solved. The immunotherapy is more effective for treatment of chronic and severe bone marrow failure and inefficient blood production than currently available growth factors of G-CSF, GM-CSF, Erythropoietin and IL-11. The mechanism of the immunotherapy is yet completely clear to us, however, some evidence suggests that in vitro activated immune cells produce and secrete multiple cytokines, working in concert, these cytokines released by the infused cells in organs important for hematopoiesis and blood production such as bone marrow, liver and spleen have remarkable effects on target cells, resulting in improved hematopoiesis, blood cell differentiation and maturation. In the preliminary clinical studies, more than 100 patients with aplastic anemia, severe chemotherapy-induced myelosuppression, systemic lupus erythematosus-associated cytopenia and idiopathic thrombocytopenia refractory to conventional therapies have been treated with the immunotherapy and the results have been encouraging. In severe idiopathic and benzene-induced aplastic anemia, 90% patients have complete or partial remission after the immunotherapy and one and half year survival is 90%. We have used 2–5x108in vitro activated allogeneic immune cells per infusion per day for 5 consecutive days, followed by small numbers of autologous infusions (1 to 10 million from approximately 50 ml of peripheral blood, depending on the severity of the disease, once a week for 4 weeks). This cycle of therapy is repeated till patient's absolute neutrophil count is more than 0.5x109/L. The duration of the immunotherapy required for patients with AA to significantly improve ranges from two months to two years depending on the severity of the disease. Idiopathic thrombocytopenia is as difficult as aplastic anemia to treat for the immunotherapy and also requires relatively long time (several months to a year) for patients to respond to the therapy. Approximately 50% adult patients treated with the immunotherapy have complete or partial remission. In severe myelosuppression induced by chemotherapy in leukemia patients, the immunotherapy is highly effective and capable of reducing infection, bleeding and blood transfusion. The recovery of severe myelosuppression (from a few days to a month depending on the severity) after the immunotherapy is much quicker than that of aplastic anemia and idiopathic thrombocytopenia purpura. In conclusion, animal and preliminary human clinical studies suggest that the immunotherapy is highly effective for some bone marrow failure and blood deficient disorders, which are usually difficult to treat with the conventional therapies. The immunotherapy described here merits further investigation.

Expression of T cell receptor Vbeta subfamily gene in patients with benzene-induced aplastic anemia

目的 比较苯中毒所致再生障碍性贫血(再障)患者与原发再障患者外周血T细胞受体(TCR)Vβ亚家族基因不同期间的表达.方法采用RT-PCR扩增1例苯中毒所致再障患者与1例原发再障患者发病初期和骨髓恢复后的不同期间外周血单个核细胞的TCRVβ24个亚家族的互补决定区3(CDR3),产物进一步经基因扫描分析T细胞的克隆性,并与10名正常人比较.结果健康人表达全部Vβ亚家族,PCR产物呈多峰图像(多克隆).苯中毒所致再障患者发病初期出现Vβ表达受抑制的情况不明显,但PCR产物出现多个单峰图像(寡克隆)或双峰图像(双克隆)的异常克隆性改变占41%,提示苯对机体的刺激作用而引起机体的相应的细胞免疫反应.随着病情恢复重新出现多克隆.原发再障患者发病初期出现9个亚家族表达受抑制,但其克隆性的改变绝大多数表现为多克隆图像,提示免疫功能不同程度受损,在骨髓恢复后可见受抑制的亚家族基因重新得到恢复,出现多克隆的多表达图像.结论苯中毒再障和原发再障患者外周血TCR Vβ亚家族基因表达有所差异,为进一步研究两者的免疫诊治提供了参考。

Effects of Sheng-Mai injection on the PRPP synthetase activity in BFU-Es and CFU-Es from bone marrows of mice with benzene-induced aplastic anemia

160 Kunming mice were divided at random into 3 groups. Group 1: normal control (40 mice). Group 2: aplastic anemia (AA) control (60 mice); benzene inhalation was carried out for 2.5 months and sterilized normal saline was injected ip. for another 6 weeks. Group 3: treated AA (60 mice); benzene was administered by inhalation in a similar manner, Sheng-Mai Injection (SMI) was administered ip. for 6 weeks after the AA models were established. SMI is a famous Chinese traditional prescription of Panax ginseng C.A. Meyer (0.1g/ml), Ophiopogon japonicus (Thunb.) Ker-Gawl (0.312g/ml) and Fructus Schisandrae (0.158g/ml). Activities of phosphoribosylpyrophosphate (PRPP) synthetase in BFU-Es and CFU-Es were estimated by ion pair reversed phase HPLC (IPrHPLC). Accompanying the sharp drop in counts of erythroid progenitor cells, the PRPP synthetase activity in CFU-Es of AA mice was reduced significantly (P<0.01), whereas there were no remarkable changes of this enzyme activity in their BFU-Es compared with the control group. Both the counts of erythroid progenitor cells and PRPP synthetase activity in CFU-Es returned nearly to normal levels following treatment with SMI of mice in Group 3 (P<0.01). Our results suggest that the attenuation of PRPP synthetase activity in peripheral erythrocytes of AA patients may originate from the weakening of activity of this enzyme in CFU-Es from their bone marrow. The impairment of PRPP formation would explain ATP depletion and disorders of energy metabolism in AA erythrocytes. SMI can distinctly increase the reduced quantity of erythroid progenitor cells and promote rapid restoration of PRPP synthetase activity in CFU-Es of AA mice.

Inhibition of the conversion of pre-interleukins-1α and 1β to mature cytokines by p-benzoquinone, a metabolite of benzene

Chronic exposure of humans to benzene causes severe bone marrow cell depression leading to aplastic anemia. Marrow stromal macrophage dysfunction and deficient interleukin-1 production has been reported for patients with severe aplastic anemia. The stromal macrophage, a target of benzene toxicity, is involved in hematopoietic regulation through the synthesis of several cytokines including interleukin-1, which is required for production by stromal fibroblasts of a number of cytokines required for the survival of hematopoietic progenitor cells. We have previously demonstrated that hydroquinone, a major toxic metabolite of benzene in marrow, prevents the proteolytic conversion of 31 kDa pre-interleukin-1α to the 17 kDa cytokine by calpain in purified murine stromal macrophages. Furthermore, stromal macrophages from benzene-treated mice produce the 31 kDa pre-interleukin-1α when stimulated in culture with endotoxin, but cannot convert the precursor to interleukin-1α. In this report, we show that 1,4-benzoquinone, the oxidation product of hydroquinone in the cell, causes a concentration-dependent inhibition of highly purified human platelet calpain with an IC 50 of 3 μM. Hydroquinone also inhibits the processing of pre-interleukin-1β by interleukin-1β convertase. The addition of 2 μM hydroquinone to B1 cells that undergo autocrine stimulation by interleukin-1β resulted in the cessation of autocrine cell growth and interleukin-1β secretion into the culture medium, as determined by Western immunoblots of the culture supernatants. Purified converting enzyme treated with 3 μM benzoquinone was incapable of converting 31 kDa recombinant pre-interleukin-1β to the 17 kDa mature cytokine as analyzed by polyacrylamide gel electrophoresis and Western immunoblotting. These findings support our observations in a mouse model that benzene-induced bone marrow cell depression results from a lack of interleukin-1α subsequent to an inhibition by benzoquinone of calpain, the protease required for converting pre-interleukin-1α to active cytokine. The results may provide a basis for studying benzene-induced aplastic anemia in a mouse model.

The toxicology of benzene.

Benzene is metabolized, primarily in the liver, to a series of phenolic and ring-opened products and their conjugates. The mechanism of benzene-induced aplastic anemia appears to involve the concerted action of several metabolites acting together on early stem and progenitor cells, as well as on early blast cells, such as pronormoblasts and normoblasts to inhibit maturation and amplification. Benzene metabolites also inhibit the function of microenvironmental stromal cells necessary to support the growth of differentiating and maturing marrow cells. The mechanism of benzene-induced leukemogenesis is less well understood. Benzene and its metabolites do not function well as mutagens but are highly clastogenic, producing chromosome aberrations, sister chromatid exchange, and micronuclei. Benzene has been shown to be a multi-organ carcinogen in animals. Epidemiological studies demonstrate that benzene is a human leukemogen. There is need to better define the lower end of the dose-response curve for benzene as a human leukemogen. The application of emerging methods in biologically based risk assessment employing pharmacokinetic and mechanistic data may help to clarify the uncertainties in low-dose risk assessment.

The Toxicology of Benzene

Benzene is metabolized, primarily in the liver, to a series of phenolic and ring-opened products and their conjugates. The mechanism of benzene-induced aplastic anemia appears to involve the concerted action of several metabolites acting together on early stem and progenitor cells, as well as on early blast cells, such as pronormoblasts and normoblasts to inhibit maturation and amplification. Benzene metabolites also inhibit the function of microenvironmental stromal cells necessary to support the growth of differentiating and maturing marrow cells. The mechanism of benzene-induced leukemogenesis is less well understood. Benzene and its metabolites do not function well as mutagens but are highly clastogenic, producing chromosome aberrations, sister chromatid exchange, and micronuclei. Benzene has been shown to be a multi-organ carcinogen in animals. Epidemiological studies demonstrate that benzene is a human leukemogen. There is need to better define the lower end of the dose-response curve for benzene as a human leukemogen. The application of emerging methods in biologically based risk assessment employing pharmacokinetic and mechanistic data may help to clarify the uncertainties in low-dose risk assessment.

Mechanism of Benzene-Induced Aplastic Anemia

Mechanism of Benzene-Induced Aplastic Anemia