Myelosuppressed Mice Research Articles

Blood-supplementing effect of low molecular weight peptides of E-Jiao on chemotherapy-induced myelosuppression: evaluation of pharmacological activity and identification of bioactive peptides released in vivo.

Background: Equus asinus L. [Equidae; Asini Corri Colla] (donkey-hide gelatin, E-Jiao) is a traditional Chinese medicine renowned for its exceptional blood-supplementing effect. However, the specific components that contribute to its efficacy remain elusive. This study aimed to demonstrate that peptides are responsible for E-Jiao's blood-supplementing effect and to explore the specific peptides contributing to its efficacy. Methods: The low molecular weight peptides of E-Jiao (LMEJ) were obtained using an in vitro digestion method. LMEJ and peptides in the rat bloodstream were characterized by peptidomics analysis. The blood-supplementing effect of LMEJ was assessed using blood-deficient zebrafish and mouse models. The effect of the peptides detected in rat blood was evaluated using the same zebrafish model, and network pharmacology analysis was performed to investigate the underlying mechanisms. Results: A total of 660 unique peptides were identified within LMEJ. Both E-Jiao and LMEJ significantly alleviated myelosuppression in mice but only LMEJ attenuated myelosuppression in zebrafish. After the administration of E-Jiao to rats, 67E-Jiao-derived peptides were detected in the bloodstream, 41 of which were identical to those identified in LMEJ. Out of these 41 peptides, five were synthesized. Subsequent verification of their effects revealed that two of them were able to alleviate myelosuppression in zebrafish. Network pharmacology study suggested that E-Jiao may exert a blood-supplementing effect by regulating signaling pathways such as JAK-STAT, IL-17 and others. These results indicated that peptides are at least partially responsible for E-Jiao's efficacy. Conclusion: This study provides a crucial foundation for further exploration of the bioactive components of E-Jiao.

Investigating the protective effects of Astragalus polysaccharides on cyclophosphamide-induced bone marrow suppression in mice and bone mesenchymal stem cells

BackgroundThis study determines the role and mechanism of APS in cyclophosphamide-induced myelosuppression in mice and bone mesenchymal stem cells (BMSCs) cell model. MethodsCy-induced myelosuppression mice and BMSCs cell model were established. Fifty C57BL/6 mice (weighing 20 ± 2 g) were randomly divided into five groups. Femur and tibia samples, bone marrow samples, and blood samples were collected 3 days after the last injection of Cy. Histopathology changes and cell apoptosis were detected. Cell viability, apoptosis, cycle distribution, reactive oxygen species activity, osteogenesis ability, and protein levels were detected. γ-H2AX and senescence-associated β-galactosidase activity expression was detected by immunofluorescence. Cy-induced senescence and Wnt/β-catenin related protein levels were detected using western blotting. ResultsThe results showed that APS effectively induced Cy-induced histological injury and cell apoptosis rate. After treated with APS, ROS and ALP levels were significantly increased. In BMSCs, cell viability, apoptosis, and cell cycle distribution were also influenced by APS treatment. Compared with the control group, cell viability was significantly increased, the cell apoptosis rate was decreased while the number of cells remained in the G0-G1 phase was increased. Meanwhile, ROS levels were significantly increased in APS group. Cell senescence and Wnt/β-catenin related protein (γ-H2AX, SA-β-gal, p21, p16, p-β-catenin/ β-catenin, c-Myc, and AXIN2) levels were also altered both in vivo and in vitro. Interestingly, the effects of APS were reversed by BML-284. ConclusionOur results indicate that APS protected Cy-induced myelosuppression through the Wnt/β-catenin pathway and APS is a potential therapeutic drug for Cy-induced myelosuppression.

Effect of ginsenoside Rg1 on hematopoietic stem cells in treating aplastic anemia in mice via MAPK pathway.

Aplastic anemia (AA) presents a significant clinical challenge as a life-threatening condition due to failure to produce essential blood cells, with the current therapeutic options being notably limited. To assess the therapeutic potential of ginsenoside Rg1 on AA, specifically its protective effects, while elucidating the mechanism at play. We employed a model of myelosuppression induced by cyclophosphamide (CTX) in C57 mice, followed by administration of ginsenoside Rg1 over 13 d. The investigation included examining the bone marrow, thymus and spleen for pathological changes via hematoxylin-eosin staining. Moreover, orbital blood of mice was collected for blood routine examinations. Flow cytometry was employed to identify the impact of ginsenoside Rg1 on cell apoptosis and cycle in the bone marrow of AA mice. Additionally, the study further evaluated cytokine levels with enzyme-linked immunosorbent assay and analyzed the expression of key proteins in the MAPK signaling pathway via western blot. Administration of CTX led to significant damage to the bone marrow's structural integrity and a reduction in hematopoietic cells, establishing a model of AA. Ginsenoside Rg1 successfully reversed hematopoietic dysfunction in AA mice. In comparison to the AA group, ginsenoside Rg1 provided relief by reducing the induction of cell apoptosis and inflammation factors caused by CTX. Furthermore, it helped alleviate the blockade in the cell cycle. Treatment with ginsenoside Rg1 significantly alleviated myelosuppression in mice by inhibiting the MAPK signaling pathway. This study suggested that ginsenoside Rg1 addresses AA by alleviating myelosuppression, primarily through modulating the MAPK signaling pathway, which paves the way for a novel therapeutic strategy in treating AA, highlighting the potential of ginsenoside Rg1 as a beneficial intervention.

Sequential administration of delta-tocotrienol ameliorates radiation-induced myelosuppression in mice and non-human primates through inducing G-CSF production

To date only four recombinant growth factors, including Filgrastim (rhG-CSF), have been approved by FDA as radiomitigators to ameliorate hematopoietic acute radiation syndrome (H-ARS). These approved agents are not stable under room-temperature, needing to be stored at 2–8 °C, and would not be feasible in a mass casualty scenario where rapid and cost-effective intervention is crucial. Delta-tocotrienol (δ-T3H), the most potent G–CSF–inducing agent among vitamin E isoforms, exhibited efficiency and selectivity on G-CSF production in comparison with TLR and STING agonists in mice. Five-dose δ-T3H was utilized as the optimal therapeutic regimen due to long-term G-CSF production and the best peripheral blood (PB) recovery of irradiated mice. Comparable with rhG-CSF, sequential administration of δ-T3H post-irradiation improved hematologic recovery and accelerated the regeneration of hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) in the bone marrow (BM) and spleen of 6.5Gy irradiated mice; and consistently enhanced repopulation of BM-HSCs. In 4.0Gy irradiated nonhuman primates, δ-T3H exhibited comparable efficacy as rhG-CSF to promote PB recovery and colony-formation of BM-HPCs. Altogether, we demonstrated that sequential administration of delta-tocotrienol ameliorates radiation-induced myelosuppression in mice and non-human primates through inducing G-CSF production, indicated δ-T3H as a promising radiomitigator for the management of H-ARS, particularly in a mass casualty scenario.

Mechanism of herb pair containing Astragali Radix and Spatholobi Caulis in the treatment of myelosuppression based on network pharmacology and experimental investigation

Ethnopharmacological relevanceThe Astragali Radix and Spatholobi Caulis herb pair (ARSC) is one of the most commonly used herbal combinations for bone marrow suppression. According to traditional Chinese medicine, Astragali Radix strengthens the spleen and replenishes qi, while Spatholobi Caulis is a hematinic agent that promotes blood circulation and enrichment. The compatibility of the two helps the body to tonify the spleen and kidneys and compensate for visceral deficiencies. However, the multi-target mechanism of ARSC in bone marrow suppression has remained largely unknown. Aim of the studyThe aim of this study is to explore the key targets and signaling pathways of the traditional Chinese herbal pair ARSC for the treatment of bone marrow suppression. Materials and methodsThe active components of ARSC and targets for myelosuppression were screened using network databases. Cytoscape 3.8.0 was used to construct compound-target, compound-disease-target and protein-protein interaction (PPI) networks. Go-function and pathway enrichment analyses were performed to explore the potential mechanism. In vivo animal experiments were conducted to verify the molecular mechanisms. ResultsThe 36 active compounds were identified from the ARSC, and a total of 108 genes involved in myelosuppression were screened. VEGFA, IL6, TNF, JUN, STAT3, PTGS2, CASP3 and MMP9 genes were identified as potential drug targets in the PPI network analyzed by CytoHubba. Enrichment analysis indicated that ARSC may treat myelosuppression through various biological processes, such as apoptosis, TNF-α signaling pathway via NF-κB, PI3K/AKT/mTOR signaling pathway, IL6/JAK/STAT3 signaling pathway, P53 signaling pathway and G2/M checkpoint signaling pathway. The results of the experiment showed that the aqueous extract of ARSC significantly alleviated myelosuppression, reduced the apoptosis rate of bone marrow cells, upregulated the mRNA expression levels of TNF-α, IL-6 and VEGF, and promoted NF-κB phosphorylation in myelosuppressed mice. ConclusionsThis study identified the active components and relevant mechanisms of ARSC in the treatment of myelosuppression. Our findings predicted that ARSC could treat bone marrow suppression through multiple components, multiple targets and multiple pathways. Pharmacological experiments showed that ARSC alleviated fluorouracil-induced myelosuppression by reducing the apoptosis rate of bone marrow cells and regulating the TNF-α/NF-κB signaling pathway.

Effect of Carica papaya Leaf Juice on Bone Marrow Aspirate of Busulfan-induced Chronic Bone Marrow Aplasia in Mice

Introduction: Aplastic anemia is a disorder characterized by peripheral pancytopenia and hypoplastic bone marrow. Papaya, botanical name Carica Papaya, is a tropical fruit with a wide range of benefits. In past couple of years, researches have been carried out on beneficial outcomes of papaya leaf juice on blood and bone marrow, particularly platelet count. Aim: Taking into consideration the beneficial effects of papaya leaves on bone marrow, current research was done to probe the effects of C.papaya leaf juice on bone marrow differential cell count and cellularity in myelosuppressed mice where busulfan was used to induce myelosuppression. Study Design: Experimental. Place and Duration of Study: our study was performed in Post Graduate Medical Institute, Lahore in one year time. Materials & Methods : It was an animal experimental study of 23 days in which 36 adult healthy swiss albino mice weighing 30-35 g of either sex were segregated into three groups with 12 mice in each group. Busulfan at a dose of 14mg/kg was given orally to the busulfan group to induce myelosuppression and to the C.papaya +busulfan group on 8 occassions (0, 3,8,11,15,18,21&23) for a duration of 23 days. To observe the myeloprotective effect of Carica papaya leaf juice 10ml/kg of juice was given to the C.papaya + busulfan group daily for 23 days. After the final dose of busulfan and Carica papaya leaf juice on day 23, the samples for bone marrow aspirate were collected on same day. Results: The current study has revealed that C.papaya leaf juice not only prevented a decrease in all parameters of bone marrow aspirate but also maintained their normal values close to normal. The megakaryopoiesis was however found to be significantly higher than the normal. Keeping in view these beneficial effects carica papaya leaf juice can be used in chemotherapy induced thrombocytopenia and other related clinical conditions. Conclusion: it can be concluded from this study that C.papaya can be used in all such conditions that lead to fall in chemotherapy induced thrombocytopenia because of its property to increase megakaryopoiesis. Keywords: C.papaya, Busulfan, Megakaryopoiesis, Bone marrow failure.

The Antiapoptotic Effect of Danggui Buxue Tang and Its Main Components on Hematopoietic Cells in Mice with Bone Marrow Suppression

To explore the hematopoiesis protection effect of Danggui Buxue Tang (DBT) and its main components Angelica polysaccharide (APS) and Astragalus polysaccharide (ASPS) on myelosuppression mice, and the mechanism of anti-apoptosis of Meg-01 cells. Mice were radiated with 4 Gy of 137Csγ ray to establish the model of radiation-induced myelosuppression. DBT, APS or ASPS (10 mg/kg) were injected into irradiated mice. Peripheral blood cell counts were performed on mice before radiation (day 0) and day 7, 14 and 21 after radiation. On the 21st day, poor plasma platelets were collected from mice to detect TPO concentration and then the mice were sacrificed. The femoral bone marrow cells were cultured for colony cell forming units (CFU). Meg-01 cells were cultured without FBS for 24 h to induce apoptosis, and then treated with DBT/APS/ASPS for 72 h. Flow cytometry (FCM) was used to detect early apoptosis (Annexin V), mitochondrial membrane potential (JC-1) and the expression of Caspase-3 to analyze the effect of DBT/APS/ASPS on cell apoptosis. DBT can stimulate the recovery of white blood cells (WBC), red blood cells (RBC) and platelets (PLT) of myelosuppression mice, especially for WBC and PLT (P<0.01, P<0.05). Compared with the control group, the number of BFU-E, CFU-MK and CFU-GM increased after adding DBT (BFU-E & CFU-GM: P<0.05； CFU-MK: P<0.01). The effect of DBT on blood TPO concentration in mice was not obvious (P=0.89). RBC, WBC and PLT were increased in APS group compared with control group (P<0.05). WBC increased after the treatment of ASPS (P<0.05). APS stimulated the formation of CFU-F, CFU-MK and CFU-GM (P<0.05). Only CFU-GM increased in ASPS group(P<0.05). Besides, DBT decreased the apoptosis of Meg-01 cells (P<0.05). The early apoptosis rate and total death rate in APS (100 μg/ml) group were lower than that of control group (P<0.01, P<0.05). The early apoptosis rate of ASPS (100 μg/ml) group was lower than that of control group (P<0.05). JC-1 and Caspase-3 showed that APS (100 μg/ml) significantly reduced apoptosis rate (P<0.01, P<0.05). DBT has protective effect on hematopoietic system, especially WBC and PLT, and has anti-apoptotic effect on Meg-01. It was found that the above effects of DBT were mainly caused by APS, and its anti-apoptosis mechanism was carried out mainly through JC-1 and Caspase-3 pathways.

Carbohydrates and ginsenosides in shenmai injection jointly improve hematopoietic function during chemotherapy-induced myelosuppression in mice

BackgroundShenmai injection (SMI), a traditional Chinese medicine (TCM) injection prepared from Red ginseng and Ophiopogon japonicus, is widely used in clinics to treat chemotherapy-induced myelosuppression. Similar to other TCM injections, SMI contains a high amount of carbohydrates (fructose, sucrose, and maltose) in addition to the bioactive substances, specifically ginsenosides (Rg1, Re, and Rb1). To date, the role of these carbohydrates in the hematopoietic function of SMI remains unclear.PurposeWe aimed to investigate the hematopoietic effects and potential mechanisms of SMI and its components, focusing on the carbohydrates present in SMI.Experimental design/methodsFirst, we evaluated the hematopoietic effect of SMI on 5-fluorouracil (5-FU)-induced myelotoxicity in a tumor-bearing mouse model. Then we prepared mixtures of ginsenosides and carbohydrates according to their proportions in SMI and evaluated their hematopoietic function in mice with 5-FU-induced myelosuppression. Finally, hematopoiesis-related molecular networks were built based on RNA sequencing (RNA-seq) of the bone marrow stromal cells (BMSCs), and the potential mechanisms of carbohydrates and ginsenosides were evaluated.ResultsSMI attenuated 5-FU-induced myelotoxicity in tumor-bearing mice. Both ginsenosides and carbohydrates increased the bone marrow nucleated cell (BMNC) count and improved the bone marrow morphology in myelosuppressive mice; they promoted the proliferation of BMSCs derived from those myelosuppressive mice. Bioinformatics analyses revealed ECM-receptor interaction, Hippo signaling, and Wnt signaling are common pathways regulated by both ginsenosides and carbohydrates; Gstt1, Gstp2, Gsta4 and Oplah in Glutathione metabolism pathway and Cd19, Cd79a, and Cd79b in B cell receptor pathway are uniquely regulated genes related to carbohydrates but not ginsenosides.ConclusionsCarbohydrates may collaborate with ginsenosides and contribute to the hematopoietic function of SMI. Carbohydrates could be considered as a bioactive component in this TCM injection.Graphical

Effect of Ligustri Lucidi Fructus on myelosuppression in mice induced by cytoxan.

In this study, we aimed to demonstrate the therapeutic effect of Ligustri Lucidi Fructus on chemotherapy-induced myelosuppression and elucidate its mechanism. A pharmacological study was conducted to investigate the mechanism of the inhibiting effects of Ligustri Lucidi Fructus on cyclophosphamide-induced bone marrow suppression in mice. HPLC was used to measure the chemical components. We demonstrated that medium and high doses of Ligustri Lucidi Fructus increased the amount of white blood cells and bone marrow nucleated cells (p < 0.05) in the cyclophosphamian-induced mouse model, and at the same time reduced granulocyte-macrophage-colony stimulating factor and thrombopoietin in the serum of myelosuppression mice (p < 0.01). Medium and high doses of Ligustri Lucidi Fructus can also adjust the thymus index and spleen index(p < 0.05). Ligustri Lucidi Fructus regulates the balance of bcl-2/bax, inhibits the expression of Caspase-3 and meanwhile stimulates the expression of mitogen-activated protein (MEK) and phospho extracellular regulated protein kinases (p-ERK) on the MAPK pathway. Five chemical constituents of Ligustri Lucidi Fructus, which may be related to myelosuppression, were analyzed. The content of specnuezhenide was 0.281%, that of ligustroflavone was 0.004%, that of salidroside was 0.094%, that of hydroxytyrosol was 0.060% and that of tyrosol was 0.069%. The effect of Ligustri Lucidi Fructus on myelosuppression after chemotherapy may be related to its multicomponent and multitarget nature. Ligustri Lucidi Fructus may be a promising potential drug for treatment after chemotherapy.

Animal Model Establishment and Its Mechanism of Cytarabine-Iduced Myelosuppression

To establish an optimized model of bone marrow suppression induced by cytarabine (Ara-C) in C57BL/6 mice and preliminarily explore the mechanism of myelosuppression based on the cycle and apoptosis of BMNC. C57BL/6 mice were intraperitoneally injected with Ara-C 50, 100 and 200 mg/kg for 7 days, respectively. The survival rate and body weight of C57BL/6 mice were monitored. The number of peripheral blood cells and bone marrow nucleated cells (BMNC) was detected, and the morphology of bone marrow, thymus and spleen were measured on the 7th, 14th and 21st day of the experiment. The cycle and apoptosis of BMNC were also detected by flow cytometry. Ara-C 200 mg/kg caused 46.7% mortality in mice, and other doses had no significant effect on mortality. All doses of Ara-C induced bone marrow suppression in mice, as shown by a decrease in the number of peripheral blood cells (WBC, Neu, RBC, PLT) and BMNC (P<0.05), decrease in bone marrow hyperplasia, accompanied by immunosuppression and compensatory hematopoiesis of the spleen, and the above manifestations and duration were dose-dependent. Among them, the myelosuppression caused by Ara-C 50 mg/kg recovered quickly, and caused by Ara-C 200 mg/kg was too severe. The result of flow cytometry showed that Ara-C could cause S and G2/m arrest and increased apoptosis in BMNC. Ara-C can induce myelosuppression in mice with a dose-dependent severity and duration, and the model of myelosuppression with Ara-C 100 mg/kg is more optimized. The mechanism is related to the inhibition of BMNC proliferation and the promotion of apoptosis.

Quercetin supplementation attenuates cisplatin induced myelosuppression in mice through regulation of hematopoietic growth factors and hematopoietic inhibitory factors

The present study investigated the effects of quercetin on cisplatin (CDDP)-induced common side effect, myelosuppression, and the possible mechanisms in Balb/c mice. The mice were randomly treated with CDDP alone or in combination with quercetin for 14 days. Quercetin was given by intraperitoneal injection (10 mg/kg, 3 times a week; IQ) or by a diet containing 0.1% or 1% quercetin (LQ and HQ, respectively). We found that quercetin supplementation especially HQ and IQ, significantly restored the decrease in number of bone marrow cells, total white blood cells, red blood cells and platelets, and the body weight in mice exposed to CDDP (P≤.05). Similar trends were observed in the number of neutrophils, lymphocytes and monocytes in the plasma. HQ and IQ also increased the levels of hematopoietic growth factors (HGFs), especially in granulocyte-macrophage-colony stimulating factor and IL-9 (P<.05), but decreased the levels of hematopoietic inhibitory factors (HIFs) and oxidative stress in the plasma and the bone marrow in CDDP-exposed mice. Furthermore, both quercetin and quercetin-3-O-glucuronide (Q3G) significantly increase cell viability and inhibited apoptosis at 48 or 72 h (P≤.05), accompanied by increasing HGF levels and decreasing HIF levels in the cultured medium in 32D cells exposed to CDDP. IL-9 siRNA transfection suppressed the effects of quercetin and Q3G on cell viability (P≤.05) in32D cells. In conclusion, our results indicate that quercetin attenuates CDDP-induced myelosuppression through the mechanisms associated with regulation of HGFs and HIFs.

Psoralen suppresses the phosphorylation of amyloid precursor protein (APP) to inhibit myelosuppression

This study aims to explore the effect of Psoralen on myelosuppression, and investigating the mechanism involved in. The mesenchymal stem cells (MSCs) were treated with CTX to construct cell model of myelosuppression, and then with APP knockdown or overexpression transfection. Cell proliferation, cell apoptosis, bone growth factors, and hematopoietic growth factors were identified. The animal model of myelosuppression syndrome was established by intraperitoneal injection of cyclophosphamide (CTX) into C57BL/6 mice, and then with APP knockdown transfection. The effect of Psoralen on myelosuppression mice with APP knockdown was explored, including observin the number of hematopoietic stem cells and bone marrow MSCs, detecting the degree of osteoporosis and the number of osteoclasts. The expression of phosphorylation-amyloid precursor protein (p-APP), bone growth factors, and hematopoietic growth factors were also examined. We found that CTX treatment inhibited cell proliferation, induced cell apoptosis, promoted p-APP/APP, and inhibited the expression of aph-1 homolog A (APH-1α), presenilin enhancer-2 (PEN-2), the receptor of advanced glycation endproducts (RAGE). Psoralen pretreatment effectively promoted cell proliferation, suppressed cell apoptosis, inhibited p-APP/APP and stimulated the expression of APH-1α, PEN-2, RAGE compared with CTX treatment. After APP knockdown, cell proliferation was inhibited, and cell apoptosis was increased. The release of bone growth factors and hematopoietic growth factors was decreased. Psoralen pretreatment could reverse the effect of APP knockdown on MSCs and myelosuppression mice. In conclusion, Psoralen treatment inhibited cell apoptosis and regulated bone growth factors and hematopoietic growth factors in myelosuppression syndrome by suppressing the phosphorylation of APP.

Hirsutine, a novel megakaryopoiesis inducer, promotes thrombopoiesis via MEK/ERK/FOG1/TAL1 signaling

BackgroundThrombocytopenia (TP) remains a challenge in clinical hematology. TP may have serious consequences, such as recurrent skin and mucosal bleeding and increased risk of intracranial and internal organ hemorrhage. However, effective and safe therapeutic drugs for the long-term management of TP are still lacking. PurposeThis study aimed to identify more effective active compounds for TP therapy. MethodsLiquid chromatography-mass spectrometry-nuclear magnetic resonance analysis was used to confirm the medicinal species and chemical structure of Hirsutine (HS). The proliferation of HS was examined by Cell Counting Kit (CCK-8) assay on cells lines. The effect of HS on megakaryocyte differentiation was analyzed by evaluating the expression of CD41, CD42b, and DNA ploidy via flow cytometry (FCM). The morphology of megakaryocytes and intermediate cells was observed using an optical microscope. K562 cells were then stained with Giemsa and benzidine. qRT-PCR was used to examine the mRNA expression of GATA-1, GATA-2, FOG-1, TAL-1, RUNX-1, NF-E2, and KLF-1 in K562 cells. Protein levels of the transcription factors were analyzed by western blotting. An MEK inhibitor was used to verify the relationship between the MEK/ERK signaling pathway and CD41/CD42b (FCM), FOG-1, and TAL-1. The Kunming thrombocytopenia mouse model was established by X-ray irradiation (4 Gy) and used to test HS activity and related hematopoietic organ index in vivo. Finally, computer simulations of molecular docking were used to predict the binding energies between HS-MEK and HS-ERK. ResultsWe preliminarily identified HS by screening a plant-sourced compound library for natural compounds with megakaryocytic differentiation and maturation (MKD/MKM)-promoting activity. We found that HS not only enhanced MKD/MKM of K562 and Meg01 cells, but also suppressed the decline of peripheral platelet levels in X-ray-induced myelosuppressive mice. In addition, HS promoted MKD via activation of MEK-ERK-FOG1/TAL1 signaling, which may be the key molecular mechanism of HS action in TP treatment. Molecular docking simulations further verified that HS targets the signaling protein MEK with high-affinity. ConclusionIn this study, we report for the first time that hirsutine boosts MKD/MKM through the MEK/ERK/FOG1/TAL1 signaling pathway and thus represents a promising treatment option for TP.

Prevention Effect of Protopanaxadiol-Type Saponins Saponins and Protopanaxatriol-Type Saponins on Myelosuppression Mice Induced by Cyclophosphamide.

Ginsenosides from ginseng are used as a therapeutic agent for various diseases. They enhance the immunomodulatory effect in cyclophosphamide (CP)-treated tumor disease. The structural characteristics of steroidal saponins are mainly divided into protopanaxadiol-type saponin (PDS) and protopanaxatriol-type saponin (PTS). At present, few researchers have studied which kind of saponin plays a more important role, thus, we compared the prevention effect of PDS and PTS on myelosuppression mice induced by CP. The components and contents of saponin and monosaccharide were analyzed by using ultra high performance liquid chromatography-charged aerosol detector (UPLC-CAD) and reversed phase-high performance liquid chromatography (RP-HPLC), respectively. Thirty-two mice were randomly divided into four groups, including control, model (CP), CP+PDS, and CP+PTS. The mice were orally administered with PDS or PTS for 28 days and then injected with CP saline solution on 25, 26, 27, and 28 days at a dose of 50 mg × kg−1. After the end of modeling, the whole blood of mice from the ophthalmic venous plexus was collected to detect routine blood tests, inflammatory cytokines, and hematopoiesis-related cytokines. Cell cycle and the apoptosis of bone marrow in the right femur were detected. The spleen and thymus were used to calculate the organ index and histological examination, and splenocytes were used to detect the percentage of CD4+ and CD25+ T cells. In the saponins analysis, PDS mainly included the Rb1, Rc, Rb2, and Rd of protopanaxadiol-type ginsenosides (accounted for 91.64%), and PTS mainly included the Re, Rg1, and Rf of protopanaxatriol-type ginsenosides (accounted for 75.46%). The animal results showed that both PDS and PTS improved the most indicators of myelosuppression mice induced by CP, including increased weight, blood cell numbers, hematopoiesis-related cytokines, and inflammatory cytokines; promoted the cell cycle of bone marrow and inhibited the apoptosis of bone marrow; elevated the spleen and thymus indexes and CD4+ count of splenocytes. The prevention effect of PDS was better than PTS in some indicators, such as red blood cells, hemoglobin, interleukin (IL)-1β, IL-4, IL-10, tumor necrosis factor-α, CD4+, and thymus index. These results suggest both PDS and PTS can prevent myelosuppression of mice induced by CP. Meanwhile, PDS and its metabolite showed higher bioavailability and bioactivity compared with PTS.

Structural Characterization of Polysaccharides Isolated from Panax notoginseng Medicinal Residue and Its Protective Effect on Myelosuppression Induced by Cyclophosphamide.

This study aims to establish the isolation and purification method of polysaccharides from medicinal residue of Panax notoginseng (PPN). The structure and protective effect of PPN on myelosuppression mice were investigated. One neutral polysaccharide (NPPN) and five acidic polysaccharides (APPN I, APPN II-A, APPN II-B, APPN III-A, and APPN III-B) were obtained. The results confirmed that NPPN, APPN I and APPN II-A are glycan with 1, 4 main chains. APPN III-A is a glycan. APPN II-B and APPN III-B are homogalacturonan pectin with 1, 4 main chains. This study demonstrated that NPPN played a bone marrow protective role in myelosuppression mice induced by cyclophosphamide. NPPN could relieve cell cycle arrest, reduce the apoptosis rate of marrow cells, and improve granulocyte-macrophage colony-stimulating (GM-CSF), thermoplastic polyolefin (TPO) and erythropoietin (EPO) serum level, which contributes to promoting the proliferation of hematopoietic cells.

Effect of Carica papaya Leaf Juice on Blood Cell Count of Busulfan-Induced Chronic Bone Marrow Aplasia in Mice

Introduction: Aplastic anemia is a syndrome of bone marrow failure comprising of peripheral pancytopenia and marrow hypoplasia. Recently, research has shown papaya leaf juice to increase blood cells counts.&#x0D; Aims &amp; Objectives: This research was planned to investigate the effect of papaya leaf juice on blood cell counts in busulfan induced myelosuppressed mice.&#x0D; Place and duration of study: Post Graduate Medical Institute, Lahore from September 2015 to December 2015. Material &amp;Methods: This animal experimental study comprised of 36 adult healthy Swiss albino mice weighing 30-35 g, divided randomly into three equal groups. Busulfan was given orally (14mg/kg) to induce myelosuppression to busulfan group and Carica papaya+ busulfan group on 8 occasions over a period of 23 days. Carica papaya leaf juice (10ml/kg) was given to the Carica papaya+ busulfan group daily for 23 days to observe its role in preventing myelosuppression. Normal group was given distilled water in an equivalent amount. One day after the final dose of busulfan and Carica papaya, the animals were sampled for blood cell counts. Data was analyzed using SPSS version 20. A p-value &lt; 0.05 and below was considered as significant.&#x0D; Results: The Carica papaya leaf juice not only prevented significant fall in hemoglobin (10.86 ± 1.455 g/dl), RBC (9.54 ± 0.954× 106/mm3) and platelet count (958.92 ± 176.22 x103/mm3) vs (9.70 ± 0.904,11.24 ± 0.952 g/dl), RBC count (6.5025 ± 1.43,10.43 ± 0.234× 106/mm3) and platelet count (627.83 ± 151.63, 3881.08 ± 83.13 x103/mm3) in busulfan group but also kept them close to normal range. Total leukocyte count in C.papaya treated group improved significantly to 8116.67± 1540.85/mm3 vs (4516.67±789.51, 6666.67±798.10/mm3) busulfan treated and normal groups respectively.&#x0D; Conclusion: The Carica papaya leaf juice possesses significant myeloprotective activity. It may be a good candidate for prevention of chemotherapy induced pancytopenia

The Involvement of Macrophage Colony Stimulating Factor on Protein Hydrolysate Injection Mediated Hematopoietic Function Improvement.

Protein hydrolysate injection (PH) is a sterile solution of hydrolyzed protein and sorbitol that contains 17 amino acids and has a molecular mass of 185.0–622.0 g/mol. This study investigated the effect of PH on hematopoietic function in K562 cells and mice with cyclophosphamide (CTX)-induced hematopoietic dysfunction. In these myelosuppressed mice, PH increased the number of hematopoietic cells in the bone marrow (BM) and regulated the concentration of several factors related to hematopoietic function. PH restored peripheral blood cell concentrations and increased the numbers of hematopoietic stem cells and progenitor cells (HSPCs), B lymphocytes, macrophages, and granulocytes in the BM of CTX-treated mice. Moreover, PH regulated the concentrations of macrophage colony stimulating factor (M-CSF), interleukin (IL)-2, and other hematopoiesis-related cytokines in the serum, spleen, femoral condyle, and sternum. In K562 cells, the PH-induced upregulation of hematopoiesis-related proteins was inhibited by transfection with M-CSF siRNA. Therefore, PH might benefit the BM hematopoietic system via the regulation of M-CSF expression, suggesting a potential role for PH in the treatment of hematopoietic dysfunction caused by cancer therapy.

TPGS-Modified Long-Circulating Liposomes Loading Ziyuglycoside I for Enhanced Therapy of Myelosuppression.

BackgroundZiyuglycoside I (ZgI), an active ingredient isolated from traditional Chinese medicine Sanguisorba officinalis L, has been demonstrated to increase the leucocytes and protect hematopoietic stem cells. However, the poor solubility and a short half-life of ZgI limit its bioavailability and efficacy. The D-α-tocopherol polyethylene glycol 1000 succinate (TPGS) has been widely used to increase the solubility, improve the encapsulation rate, and extend the half-life of drugs.MethodsHere, we formulated the TPGS-modified long-circulating liposomes loading ZgI with a sustained drug release and enhanced therapy for myelosuppression. ZgI-TPGS-liposomes were manufactured using a thin-film hydration technique, followed by characterizations of physicochemical properties, including the particle size, zeta potential, TEM, SEM, FTIR, XRD, stability, drug loading (DL), encapsulation efficiency (EE). The in vitro and in vivo delivery efficiency were further evaluated by cellular uptake, in vitro drug release and in vivo pharmacokinetics. Finally, therapeutic effect on myelosuppression was investigated.ResultsThe ZgI-TPGS-liposomes had an particle size of 97.89 ± 1.42 nm and ZP of −28.65 ± 0.16 mV. It exhibited DL of 9.06 ± 0.76% and EE of 92.34 ± 3.83%, along with excellent storage stability, cellular uptake and sustained drug release to free ZgI and liposomes without TPGS. Additionally, the TPGS modified liposomes significantly enhanced the therapeutic effect of ZgI on CTX induced myelosuppression, which can be confirmed in the apoptosis inhibition and cell viability promotion of CTX injured HSPC-1 cells. Also, the mice in vivo pharmacodynamics demonstrated that TPGS liposomes promoted ZgI increasing the numbers of leucocytes and neutrophils in myelosuppression mice induced by CTX.ConclusionOur research suggest that TPGS-modified long-circulating liposomes loading ziyuglycoside I has potential application in myelosuppression therapy.

Effect of astragalus polysaccharide on Hematopoiesis and Thrombopoiesis In Vivo and In Vitro

Objective:Astragali Radix is a main component of DangGui BuXue Tang (DBT), a popular Chinese Medicine formula used for promotion of “blood production”. Astragalus polysaccharide (APS) is the main active compound of Astragali Radix. Our previous studies have demonstrated the hematopoietic effect of DBT (Yang et al, J Ethnopharmacol 2009). The study is to investigate the effects of APS on hematopoiesis and thrombopoiesis in vivo and in vitro. Further, we determined the mechanism in this study. Methods: Irradiation-induced myelosuppression mice were divided into three groups of 6 subjects each. Three groups were treated with APS (10 mg/kg/day), thrombopoietin (TPO, 1μg/kg/day) and vehicle respectively. Peripheral blood cell counts from each group were then compared at different time-points. Colony-forming unit (CFU) assays were used to determine the mechanism of APS on the proliferation and differentiation of hematopoietic cells. Finally, using the HL-60 cell as model, we analyzed the cellular apoptosis progression with and without APS by Annexin V, Caspase 3 and JC-1. Results: APS enhanced platelets and red blood cells (RBC) of the myelosuppression mice recovery in vivo, except white blood cells (WBC). The platelet count in APS-treated mice were higher than that in control mice on day 7, day 14 and day 21(p< 0.05). Meanwhile, the RBC count in APS-treated mice was higher than that in control mice on day 21(p=0.003). Morphological examination showed that APS treatment significantly protected the hematopoietic cells from the irradiation-induced apoptosis . Moreover, APS promoted the formation of all lineages of CFU in vitro . Last, we test the expression of Annexin V, Caspase-3 and JC-1 via flow cytometry and found that APS significantly reduced cell apoptosis in HL-60 cells. Conclusions: APS can promote hematopoiesis and thrombopoiesis of irradiation mouse model in vivo and in vitro . APS promotes the proliferation of peripheral blood cells by promoting the proliferation of hematopoietic stem/progenitors and inhibiting apoptosis of hematopoietic cells. DisclosuresNo relevant conflicts of interest to declare.

BET-Inhibition By JQ1 Promotes Proliferation and Self-Renewal Capacity of Hematopoietic Stem Cells

Background/significance: Bromodomain and extra terminal domain (BET) proteins have been shown to be critical for maintaining the function of leukemia stem cells. Although BET-inhibitors are in clinical development and show promising activity in different hematologic malignancies, a systematic analysis of the consequences of pharmacological BET-inhibition on healthy hematopoietic (stem) cells is outstanding.Methods: C57BL/6J (WT) or B6.SJL-Ptprca Pepcb/BoyJ mice, which carry the congenic pan-leukocyte marker CD45.1 were treated for 3 weeks with the BET-Inhibitor JQ1 or a placebo. For competitive repopulation transplantations, bone marrow from treated CD45.1 mice was transplanted together with WT competitor cells (CD45.2) into lethally irradiated WT mice (9 Gy) in different ratios. Engraftment of all major hematopoietic lineages was monitored in peripheral blood (PB) and BM using differential blood counting and flow cytometry. For the determination of HSC frequency in vivo, extreme limiting dilution analysis (ELDA) was carried out by transplanting 2x103, 2x104 or 2x105 JQ1- or placebo-treated bone marrow cells from WT mice together with 2x105 bone marrow cells from CD45.2 mice. Engraftment was assumed when CD45.1+ myeloid cells were present in the peripheral blood in frequencies > 1 %.Colony formation assays were used to measure the abundance of colony forming progenitor cells (M3434) as well as megakaryocytic progenitors (MegaCult-C). For analysis of recovery after myelosuppression, JQ1- or placebo-treated WT mice received sublethal irradiation (5 Gy) and hematopoietic repopulation was monitored. For analysis of HSC proliferation, 1 mg BrdU was injected 24 h, 12 h, and 6 h before sacrifice.Results: As a first step, we analyzed the mRNA expression patterns of Brd2-4 in all major hematopoietic subpopulations, because BET-family members represent the main molecular targets of JQ1. Expression of Brd4 mRNA is most abundant in megakaryocytes (MK), followed by B cells, T cells and hematopoietic stem cells (HSC) and expression levels of Brd2+3 show similar distribution.JQ1-treatment decreases the numbers of pre-, immature and mature B cells while numbers of early pro-B cells remain constant (pre-B cells: 125 ± 7 vs. 52 ± 5 and immature B cells: 121 ± 9 vs. 26 ± 5 x104 cells/tibia; n=4; *p<0.05). Furthermore, it increases apoptosis in all T cell subsets, all together leading to reduced cellularity in thymus, BM and spleen. Intriguingly, JQ1 induces a 3-fold increase in the number of hematopoietic stem and progenitor cells (25 ± 2 vs. 70 ± 5 x103 LSK/femur; n=5; *p<0.05) and mobilizes HSC into the peripheral blood (4 ± 1 vs. 28 ± 2 x106 CFU/ml blood; n=5; *p<0.05). Consistently, extreme limiting dilution analysis (ELDA) showed a 3-fold increase in stem cell frequencies in the bone marrow upon JQ1-treatment (1/19480 vs. 1/6284; n=8-12; *p<0.05).We found that the elevated HSC numbers arise from increased cycling of LT-HSC, as these cells incorporate more BrdU upon JQ1-treatment (29 ± 2 vs. 41 ± 3 % BrdU+ LT-HSC in bone marrow; n=5; *p<0.05). Moreover and in contrast to placebo, JQ1-treated bone marrow cells preserve their colony-forming properties during several rounds of replating in vitro, indicating sustained stemness (18 ± 4 vs. 77 ± 10 CFU/1x104 bone marrow cells after third replating; n=3-8; *p<0.05). Due to increased numbers of HSC, JQ1-treated bone marrow engrafts better after competitive stem cell transplantation and repopulates the hematopoietic system significantly faster after sublethal myeloablation. Six weeks after irradiation, total leukocyte counts of JQ1-treated mice fully recover to levels before irradiation (94 ± 7 % recovery; n=5; *p<0.05), whereas recovery is slower in the control group (47 ± 5 % recovery; n=5; *p<0.05).Conclusions: JQ1 induces proliferation and mobilization of HSC, thereby increasing their number in bone marrow and peripheral blood. Therefore, BET-inhibition might benefit patients after myelosuppression or stem cell transplantation. DisclosuresLoges:BerGenBio ASA: Research Funding.