New Approach for Targeted Treatment of Mild COVID-19 by Honeysuckle through Network Pharmacology Analysis.

OBJECTIVE: To investigate the potential pharmacological value of extracts from honeysuckle on patients with mild coronavirus disease 2019 (COVID-19) infection. METHODS: The active components and targets of honeysuckle were screened by Traditional Chinese Medicine Database and Analysis Platform (TCMSP). SwissADME and pkCSM databases predict pharmacokinetics of ingredients. The Gene Expression Omnibus (GEO) database collected transcriptome data for mild COVID-19. Data quality control, differentially expressed gene (DEG) identification, enrichment analysis, and correlation analysis were implemented by R toolkit. CIBERSORT evaluated the infiltration of 22 immune cells. RESULTS: The seven active ingredients of honeysuckle had good oral absorption and medicinal properties. Both the active ingredient targets of honeysuckle and differentially expressed genes of mild COVID-19 were significantly enriched in immune signaling pathways. There were five overlapping immunosignature genes, among which RELA and MAP3K7 expressions were statistically significant (P < 0.05). Finally, immune cell infiltration and correlation analysis showed that RELA, MAP3K7, and natural killer (NK) cell are with highly positive correlation and highly negatively correlated with hematopoietic stem cells. CONCLUSION: Our analysis suggested that honeysuckle extract had a safe and effective protective effect against mild COVID-19 by regulating a complex molecular network. The main mechanism was related to the proportion of infiltration between NK cells and hematopoietic stem cells.

Ascorbic acid supports ex vivo generation of plasmacytoid dendritic cells from circulating hematopoietic stem cells.

Plasmacytoid dendritic cells (pDCs) constitute a rare type of immune cell with multifaceted functions, but their potential use as a cell-based immunotherapy is challenged by the scarce cell numbers that can be extracted from blood. Here, we systematically investigate culture parameters for generating pDCs from hematopoietic stem and progenitor cells (HSPCs). Using optimized conditions combined with implementation of HSPC pre-expansion, we generate an average of 465 million HSPC-derived pDCs (HSPC-pDCs) starting from 100,000 cord blood-derived HSPCs. Furthermore, we demonstrate that such protocol allows HSPC-pDC generation from whole-blood HSPCs, and these cells display a pDC phenotype and function. Using GMP-compliant medium, we observe a remarkable loss of TLR7/9 responses, which is rescued by ascorbic acid supplementation. Ascorbic acid induces transcriptional signatures associated with pDC-specific innate immune pathways, suggesting an undescribed role of ascorbic acid for pDC functionality. This constitutes the first protocol for generating pDCs from whole blood and lays the foundation for investigating HSPC-pDCs for cell-based immunotherapy.

High-throughput drug screening identifies the ATR-CHK1 pathway as a therapeutic vulnerability of CALR mutated hematopoietic cells.

Mutations of calreticulin (CALR) are the second most prevalent driver mutations in essential thrombocythemia and primary myelofibrosis. To identify potential targeted therapies for CALR mutated myeloproliferative neoplasms, we searched for small molecules that selectively inhibit the growth of CALR mutated cells using high-throughput drug screening. We investigated 89 172 compounds using isogenic cell lines carrying CALR mutations and identified synthetic lethality with compounds targeting the ATR-CHK1 pathway. The selective inhibitory effect of these compounds was validated in a co-culture assay of CALR mutated and wild-type cells. Of the tested compounds, CHK1 inhibitors potently depleted CALR mutated cells, allowing wild-type cell dominance in the co-culture over time. Neither CALR deficient cells nor JAK2V617F mutated cells showed hypersensitivity to ATR-CHK1 inhibition, thus suggesting specificity for the oncogenic activation by the mutant CALR. CHK1 inhibitors induced replication stress in CALR mutated cells revealed by elevated pan-nuclear staining for γH2AX and hyperphosphorylation of RPA2. This was accompanied by S-phase cell cycle arrest due to incomplete DNA replication. Transcriptomic and phosphoproteomic analyses revealed a replication stress signature caused by oncogenic CALR, suggesting an intrinsic vulnerability to CHK1 perturbation. This study reveals the ATR-CHK1 pathway as a potential therapeutic target in CALR mutated hematopoietic cells.

Inferring the dynamics of mutated hematopoietic stem and progenitor cells induced by IFNα in myeloproliferative neoplasms.

Classical BCR-ABL-negative myeloproliferative neoplasms (MPNs) are clonal disorders of hematopoietic stem cells (HSCs) caused mainly by recurrent mutations in genes encoding JAK2 (JAK2), calreticulin (CALR), or the thrombopoietin receptor (MPL). Interferon α (IFNα) has demonstrated some efficacy in inducing molecular remission in MPNs. To determine factors that influence molecular response rate, we evaluated the long-term molecular efficacy of IFNα in patients with MPN by monitoring the fate of cells carrying driver mutations in a prospective observational and longitudinal study of 48 patients over more than 5 years. We measured the clonal architecture of early and late hematopoietic progenitors (84 845 measurements) and the global variant allele frequency in mature cells (409 measurements) several times per year. Using mathematical modeling and hierarchical Bayesian inference, we further inferred the dynamics of IFNα-targeted mutated HSCs. Our data support the hypothesis that IFNα targets JAK2V617F HSCs by inducing their exit from quiescence and differentiation into progenitors. Our observations indicate that treatment efficacy is higher in homozygous than heterozygous JAK2V617F HSCs and increases with high IFNα dose in heterozygous JAK2V617F HSCs. We also found that the molecular responses of CALRm HSCs to IFNα were heterogeneous, varying between type 1 and type 2 CALRm, and a high dose of IFNα correlates with worse outcomes. Our work indicates that the long-term molecular efficacy of IFNα implies an HSC exhaustion mechanism and depends on both the driver mutation type and IFNα dose.

Increased insulin and GLUT2 gene expression and elevated glucokinase activity in ß-like cells of islets of langerhans differentiated from human haematopoietic stem cells on treatment with Costus igneus leaf extract.

In the quest to understand lost ß-cells regeneration in the diabetic condition, we have demonstrated successful differentiation of human haematopoietic stem cells (HSCs) to functional ß-like cells. Costus igneus (Ci) leaf extract is known to exhibit anti-diabetic properties by lowering the blood glucose level as demonstrated in mice models. To establish the anti-diabetic properties of Ci leaf extract on human subjects, we studied the effect of Ci on these differentiated ß-like cells. Ci leaf extract showed its anti-diabetic property through elevated glucokinase activity which catalyzes the rate-limiting step of glucose catabolism in ß-like cells and acts as a sensor for insulin production while decreasing the glucose-6-phosphatase activity. Upon increasing the concentrations of Ci leaf extract (25, 65, 105, 145, 185 µg/ml) and glucose concentrations (5.5, 11.1, and 25 mM) Ci leaf extract treated ß-like cells showed enhanced glucokinase and decreased glucose-6-phosphatase activities and an exponential rise in gene expressions of INS and GLUT2 was observed. The present study shows enhanced INS and GLUT2 gene expression and elevated glucokinase activity in ß-like cells differentiated from HSCs upon treatment with Ci leaf extract explain the anti-diabetic property of Ci leaf extract. This extract can be effectively used in the management of diabetes.

Comprehensive transcriptome analysis of erythroid differentiation potential of olive leaf in haematopoietic stem cells.

Anaemia is one of the leading causes of disability in young adults and is associated with increased morbidity and mortality in elderly. With a global target to reduce the disease burden of anaemia, recent researches focus on novel compounds with the ability to induce erythropoiesis and regulate iron homeostasis. We aimed to explore the biological events and potential polypharmacological effects of water-extracted olive leaf (WOL) on human bone marrow-derived haematopoietic stem cells (hHSCs) using a comprehensive gene expression analysis. HPLC analysis identifies six bioactive polyphenols in the WOL. Treatment with WOL for 12 days regulated gene expressions related to erythroid differentiation, oxygen homeostasis, iron homeostasis, haem metabolism and Hb biosynthesis in hHSCs. Functional clustering analysis reveals several major functions of WOL such as ribosomal biogenesis and mitochondrial translation machinery, glycolytic process, ATP biosynthesis and immune response. Additionally, the colonies of both primitive and mature erythroid progenitors, CFU-E and BFU-E, were significantly increased in WOL-treated hHSCs. The expressions of erythroid markers, CD47, glycophorin A (GYPA), and transferrin receptor (TFRC) and adult Hb subunits-HBA and HBB were also confirmed in immunofluorescent staining and flow cytometer analysis in WOL-treated hHSCs. It is well known that induction of lineage-specific differentiation, as well as the maturation of early haematopoietic precursors into fully mature erythrocytes, involves multiple simultaneous biological events and complex signalling networks. In this regard, our genome-wide transcriptome profiling with microarray study on WOL-treated hHSCs provides general insights into the multitarget prophylactic and/or therapeutic potential of WOL in anaemia and other haematological disorders.

Nicotinamide riboside attenuates age-associated metabolic and functional changes in hematopoietic stem cells.

With age, hematopoietic stem cells (HSC) undergo changes in function, including reduced regenerative potential and loss of quiescence, which is accompanied by a significant expansion of the stem cell pool that can lead to haematological disorders. Elevated metabolic activity has been implicated in driving the HSC ageing phenotype. Here we show that nicotinamide riboside (NR), a form of vitamin B3, restores youthful metabolic capacity by modifying mitochondrial function in multiple ways including reduced expression of nuclear encoded metabolic pathway genes, damping of mitochondrial stress and a decrease in mitochondrial mass and network-size. Metabolic restoration is dependent on continuous NR supplementation and accompanied by a shift of the aged transcriptome towards the young HSC state, more youthful bone marrow cellular composition and an improved regenerative capacity in a transplant setting. Consequently, NR administration could support healthy ageing by re-establishing a more youthful hematopoietic system.

Radix Astragali polysaccharide RAP directly protects hematopoietic stem cells from chemotherapy-induced myelosuppression by increasing FOS expression.

Radix Astragali polysaccharide RAP has been reported to play a crucial role in hematopoiesis without a clear mechanism. In this study, RAP's effects to enhance the recovery of cyclophosphamide (Cy)-suppressed bone marrow and blood cells is confirmed in vivo first. Confocal micrographs demonstrated the interesting direct binding of FITC-RAP to hematopoietic stem cells (HSC) in bone marrow. RAP protects both mice and human HSC in terms of cell morphology, proliferation, and apoptosis. RNA-sequencing and shRNA approaches revealed FOS to be a key regulator in RAP's protection. These evidences provide an unreported mechanism that RAP directly protects hematopoietic stem cells from chemotherapy-induced myelosuppression by increasing FOS expression.

Polysaccharide-rich extract from Polygonatum sibiricum protects hematopoiesis in bone marrow suppressed by triple negative breast cancer.

Polysaccharide is one of main components in Polygonatum sibiricum (PS), which is an herbal medicine widely used in East Asia. Polysaccharides from Polygonatum sibiricum has been shown to exhibit multiple biological activities, such as anti-diabetes, anti-inflammation, antioxidant, immunity modulation, and anticancer. Since hematopoietic system is one of determinant factors in cancer control, we here explored the effect of polysaccharide-rich extract from Polygonatum sibiricum (PREPS) on hematopoiesis in the mice bearing triple negative breast cancer (TNBC). We found that the 4T1 TNBC tumor significantly increased myeloid cells in peripheral blood, bone marrow and spleen, while decreasing bone marrow hematopoietic stem and progenitor cells (HSPCs), indicative of an inhibition of medullary hematopoiesis. When 4T1 TNBC tumor-bearing mice were treated with PREPS, the percentage of myeloid cells within tumor-infiltrating immune cells was reduced. In addition, PREPS also inhibited hematopoietic cell expansion in the spleen, which was induced by TNBC tumors. Importantly, PREPS markedly increased HSPCs and common lymphoid progenitors in the bone marrow that had been suppressed by TNBC tumors. These findings suggest that PREPS protect hematopoiesis inhibited by TNBC tumors in the bone marrow. Although PREPS alone did not achieve statistical significance in the suppression of TNBC tumor growth, it may have a long-lasting anti-tumor effect to assist TNBC therapies by sustaining hematopoiesis and lymphoid regeneration in bone marrow.

Yin and Yang: The dual effects of interferons on hematopoiesis.

Interferons are an ancient and well-conserved group of inflammatory cytokines most famous for their role in viral immunity. A decade ago, we discovered that interferons also play an important role in the biology of hematopoietic stem cells (HSCs), which are responsible for lifelong blood production. Though we have learned a great deal about the role of interferons on HSC quiescence, differentiation, and self-renewal, there remains some controversy regarding how interferons impact these stem cells, with differing conclusions depending on experimental models and clinical context. Here, we review the contradictory roles of Type 1 and 2 interferons in hematopoiesis. Specifically, we highlight the roles of interferons in embryonic and adult hematopoiesis, along with short-term and long-term adaptive and maladaptive responses to inflammation. We discuss experimental challenges in the study of these powerful yet short-lived cytokines and strategies to address those challenges. We further review the contribution by interferons to disease states including bone marrow failure and aplastic anemia as well as their therapeutic use to treat myeloproliferative neoplasms and viral infections, including SARS-CoV2. Understanding the opposing effects of interferons on hematopoiesis will elucidate immune responses and bone marrow failure syndromes, and future therapeutic approaches for patients undergoing HSC transplantation or fighting infectious diseases and cancer.

Effect of Tussilago farfara L. Polysaccharides on the Content of Hematopoietic Stem Cells (CD117+34+) in the Bone Marrow of Mice Treated with Cyclophosphamide.

Myelotoxicity is a serious side effect of anticancer drugs. The search for drugs that can reduce the hematological complications of chemotherapy through modulation of hematopoietic stem cells is an urgent task of oncopharmacology. In the present study we showed that administration of Tussilago farfara L. polysaccharides to C57BL/6 mice treated with cyclophosphamide can increase the number of hematopoietic stem cells (CD117+34+) in the bone marrow.

Guilu Erxian Glue () Inhibits Chemotherapy-Induced Bone Marrow Hematopoietic Stem Cell Senescence in Mice May via p16INK4a-Rb Signaling Pathway.

OBJECTIVE: To evaluate the effect of Guilu Erxian Glue (, GEG) on cyclophosphamide (CTX)-induced bone marrow hematopoietic stem cells (HSCs) senescence in mice and explore the underlying mechanism. METHODS: The H22 liver cancer ascites lump model was established in male Kunming mice by injecting intraperitoneally (i.p.) with 5 × 106/mL H22 cells per mouse. Fifty tumor-bearing mice were divided into the control, model, pifithrin-α, GEG, and GEG+pifithrin-α groups using a random number table, 10 mice in each group. CTX (100 mg/kg i.p.) was administrated to mice from day 1 to day 3 (d1-d3) continuously except for the control group. The mice in the pifithrin-α, GEG and GEG+pifithrin-α groups were treated with pifithrin-α (2.2 mg/(kg·d) i.p.) for 6 consecutive days (d4-d9), GEG (9.5 g/(kg·d) i.p.) for 9 consecutive days (d1-d9), and GEG plus pifithrin-α, respectively. HSCs were collected after 9-d drug treatment. The anti-aging effect of GEG was studied by cell viability, cell cycle, and ß -galactosidase (ß -gal) assays. The mRNA and protein expressions of cyclin-dependent kinase 2 (CDK2), CDK4, inhibitor of cyclin-dependent kinase 4a encoding the tumor suppressor protein p16 (p16INK4a), p21Cip1/Waf1, p53, and phosphorylated retinoblastoma (pRb) were evaluated by quantitative real-time reverse transcription-polymerase chain reaction and semi-quantitative Western blot, respectively. RESULTS: Compared with the model group, GEG increased cell viability as well as proliferation (P<0.05 or P<0.01) and reduced ß -gal expression. Furthermore, GEG significantly decreased the expressions of p16INK4a, p53 and p21Cip1/Waf1 proteins, and increased the expressions of CDK2, CDK4 and pRb proteins compared with the model group (P<0.05 or P<0.01). CONCLUSION: GEG can alleviate CTX-induced HSCs senescence in mice, and the p16INK4a-Rb signaling pathway might be the underlying mechanism.

Furostanol Saponins from Trillium tschonoskii Promote the Expansion of Human Cord Blood Hematopoietic Stem and Progenitor Cells.

Trillium tschonoskii is a medicinal plant known to biosynthesize steroidal saponins. A phytochemical investigation of the rhizomes of T. tschonoskii led to the isolation of nine new furostanol saponins (1-9) and 11 known analogues (10-20). Five of these new compounds were shown to have hydroxy groups at the C-5 and C-6 positions, while two possess a rare aglycone containing carbonyl groups at the C-16 and C-22 positions as well as a Δ17(20) double bond, and the others have conjugated double bonds in the E-ring or have different sugar chains at the C-3 position. All the isolates were tested for their effect on the expansion of human cord blood (CB) CD34+ hematopoietic stem and progenitor cells. It was found that CB CD34+ cells treated with compounds 6, 7, 9, 10, 14, 15, and 19 showed increased numbers of rigorously phenotype-defined hematopoietic stem cells. Notably, compounds 9, 10, 13, and 14 demonstrated an enhanced ability to increase the percentages and numbers of CB CD34+CD38- cells and multipotential progenitors. The present study is the first to report that furostanol saponins from T. tschonoskii rhizomes can promote hematopoietic stem/progenitor cell (HSPC) expansion.

Development of Small Molecule MEIS Inhibitors that modulate HSC activity.

Meis1, which belongs to TALE-type class of homeobox gene family, appeared as one of the key regulators of hematopoietic stem cell (HSC) self-renewal and a potential therapeutical target. However, small molecule inhibitors of MEIS1 remained unknown. This led us to develop inhibitors of MEIS1 that could modulate HSC activity. To this end, we have established a library of relevant homeobox family inhibitors and developed a high-throughput in silico screening strategy against homeodomain of MEIS proteins using the AutoDock Vina and PaDEL-ADV platform. We have screened over a million druggable small molecules in silico and selected putative MEIS inhibitors (MEISi) with no predicted cytotoxicity or cardiotoxicity. This was followed by in vitro validation of putative MEIS inhibitors using MEIS dependent luciferase reporter assays and analysis in the ex vivo HSC assays. We have shown that small molecules named MEISi-1 and MEISi-2 significantly inhibit MEIS-luciferase reporters in vitro and induce murine (LSKCD34l°w cells) and human (CD34+, CD133+, and ALDHhi cells) HSC self-renewal ex vivo. In addition, inhibition of MEIS proteins results in downregulation of Meis1 and MEIS1 target gene expression including Hif-1α, Hif-2α and HSC quiescence modulators. MEIS inhibitors are effective in vivo as evident by induced HSC content in the murine bone marrow and downregulation of expression of MEIS target genes. These studies warrant identification of first-in-class MEIS inhibitors as potential pharmaceuticals to be utilized in modulation of HSC activity and bone marrow transplantation studies.

Expansion and preservation of the functional activity of adult hematopoietic stem cells cultured ex vivo with a histone deacetylase inhibitor.

Attempts to expand ex vivo the numbers of human hematopoietic stem cells (HSCs) without compromising their marrow repopulating capacity and their ability to establish multilineage hematopoiesis has been the subject of intense investigation. Although most such efforts have focused on cord blood HSCs, few have been applied to adult HSCs, a more clinically relevant HSC source for gene modification. To date, the strategies that have been used to expand adult HSCs have resulted in modest effects or HSCs with lineage bias and a limited ability to generate T cells in vivo. We previously reported that culturing umbilical cord blood CD34+ cells in serum-free media supplemented with valproic acid (VPA), a histone deacetylase inhibitor, and a combination of cytokines led to the expansion of the numbers of fully functional HSCs. In the present study, we used this same approach to expand the numbers of adult human CD34+ cells isolated from mobilized peripheral blood and bone marrow. This approach resulted in a significant increase in the numbers of phenotypically defined HSCs (CD34+CD45RA-CD90+D49f+). Cells incubated with VPA also exhibited increased aldehyde dehydrogenase activity and decreased mitochondrial membrane potential, each functional markers of HSCs. Grafts harvested from VPA-treated cultures were able to engraft in immune-deficient mice and, importantly, to generate cellular progeny belonging to each hematopoietic lineage in similar proportion to that observed with unmanipulated CD34+ cells. These data support the utility of VPA-mediated ex vivo HSC expansion for gene modification of adult HSCs.

Extracellular Adenosine Triphosphate (eATP) and Its Metabolite, Extracellular Adenosine (eAdo), as Opposing "Yin-Yang" Regulators of Nlrp3 Inflammasome in the Trafficking of Hematopoietic Stem/Progenitor Cells.

Nlrp3 inflammasome plays a pleiotropic role in hematopoietic cells. On the one hand, physiological activation of this intracellular protein complex is crucial to maintaining normal hematopoiesis and the trafficking of hematopoietic stem progenitor cells (HSPCs). On the other hand, its hyperactivation may lead to cell death by pyroptosis, and prolonged activity is associated with sterile inflammation of the BM and, as a consequence, with the HSPCs aging and origination of myelodysplasia and leukemia. Thus, we need to understand better this protein complex's actions to define the boundaries of its safety window and study the transition from being beneficial to being detrimental. As demonstrated, the Nlrp3 inflammasome is expressed and active both in HSPCs and in the non-hematopoietic cells that are constituents of the bone marrow (BM) microenvironment. Importantly, the Nlrp3 inflammasome responds to mediators of purinergic signaling, and while extracellular adenosine triphosphate (eATP) activates this protein complex, its metabolite extracellular adenosine (eAdo) has the opposite effect. In this review, we will discuss and focus on the physiological consequences of the balance between eATP and eAdo in regulating the trafficking of HSPCs in an Nlrp3 inflammasome-dependent manner, as seen during pharmacological mobilization from BM into peripheral blood (PB) and in the reverse mechanism of homing from PB to BM and engraftment. We propose that both mediators of purinergic signaling and the Nlrp3 inflammasome itself may become important therapeutic targets in optimizing the trafficking of HSPCs in clinical settings.

A novel hiPSC-derived system for hematoendothelial and myeloid blood toxicity screens identifies compounds promoting and inhibiting endothelial-to-hematopoietic transition.

The exposure to toxic environmental and pharmaceutical substances can pose a long-term risk to human's health. In this study, we sought to investigate the potential of our recently developed method for induction of myeloid hematoendothelial and blood cells by overexpression of two transcription factors, GATA2 and ETV2, in human induced pluripotent stem cells (hiPSCs) for toxicity screening. For the primary screen in a high-throughput format, we selected twenty-two chemicals with various degrees of cytotoxicity available from the NIEHS National Toxicology Program (Tox21). The compounds were applied during the endothelial-to-hematopoietic transition and to differentiated myeloid progenitors growing in suspension. The system was capable of identifying compounds with both inhibitory and favorable effects on hematopoietic network, changes in expression of hematopoietic markers, and mitochondrial and cytotoxicity. The findings were confirmed and further investigated by secondary screens, colony forming cell assay, and gene expression profiling. The hematoendothelial toxicity of 5-fluorouracil, berberine chloride, and benzo(a)pyrene is characterized by the inhibition of cell division and a shift of hematopoietic programming to non-hemogenic endothelial and mesenchymal fates. This study demonstrates the feasibility of transcription factor (TF)-based differentiation systems to monitor endothelial and hematotoxicity and serves as an informative platform for screening myelosuppressive or stimulatory drugs and mechanistic studies of their action.

Fetal liver Mll-AF4+ hematopoietic stem and progenitor cells respond directly to poly(I:C), but not to a single maternal immune activation.

T(4;11) MLL-AF4 acute leukemia is one of the most aggressive malignancies in infant and pediatric populations. Epidemiological and functional studies have highlighted the influence of an overstimulation of the immune system on leukemia development. This study aimed at assessing if the cell-of-origin of t(4;11) MLL-AF4 acute leukemia is sensitive to a viral or bacterial mimic and if maternal immune activation can lead to a full-blown leukemia. To answer this, we used the Mll-AF4 pre-leukemia mouse model that initiates the expression of Mll-AF4 in the first definitive hematopoietic cells formed during embryonic development. We observed an increase in proliferation upon hematopoietic differentiation of fetal liver Mll-AF4+ Lineage-Sca1+ckit+ (LSK) cells exposed to the immune stimulants, poly(I:C) or LPS/lipopolysaccharide. This was accompanied by increased expression of a subset of MLL-AF4 signature genes and members of the Toll-like receptor signaling pathways in fetal liver Mll-AF4+ LSK exposed to poly(I:C), suggesting that the cell-of-origin responds to inflammatory stimuli. Maternal immune activation using a single dose of poly(I:C) did not lead to the development of leukemia in Mll-AF4+ and control offspring. Instead, aging MLL-AF4+ mice showed an increased proportion of T-lymphoid cells in the spleen, lost their B-lymphoid bias, and had decreased frequencies of hematopoietic stem and multipotent progenitor cells. Overall, this study suggests that the fetal liver Mll-AF4+ LSK cells are sensitive to direct exposure to inflammatory stimuli, especially poly(I:C); however, maternal immune activation induced by a single exposure to poly(I:C) is not sufficient to initiate MLL-AF4 leukemogenesis.

Cytotoxic and Proapoptotic Activity of Sanguinarine, Berberine, and Extracts of Chelidonium majus L. and Berberis thunbergii DC. toward Hematopoietic Cancer Cell Lines.

Isoquinoline alkaloids belong to the toxic secondary metabolites occurring in plants of many families. The high biological activity makes these compounds promising agents for use in medicine, particularly as anticancer drugs. The aim of our study was to evaluate the cytotoxicity and proapoptotic activity of sanguinarine, berberine, and extracts of Chelidonium majus L. and Berberis thunbergii DC. IC10, IC50, and IC90 doses were established toward hematopoietic cancer cell lines using trypan blue staining. Alterations in the expression of 18 apoptosis-related genes in cells exposed to IC10, IC50, and IC90 were evaluated using real-time PCR. Sanguinarine and Chelidonium majus L. extract exhibit significant cytotoxicity against all studied cell lines. Lower cytotoxic activity was demonstrated for berberine. Berberis thunbergii DC. extract had no influence on cell viability. Berberine, sanguinarine, and Chelidonium majus L. extract altered the expression of apoptosis-related genes in all tested cell lines, indicating the induction of apoptosis. The presented study confirmed the substantial cytotoxicity and proapoptotic activity of sanguinarine, berberine, and Chelidonium majus L. extract toward the studied hematopoietic cell lines, which indicates the utility of these substances in anticancer therapy.

Ipomeolides A and B, Resin Glycosides from Ipomoea pes-caprae and Combination Therapy of Ipomeolide A with Doxorubicin.

Two new resin glycosides, ipomeolides A (1) and B (2), both with an unusual nonlinear heteropentasaccharide core, along with five known compounds were isolated from the n-hexane/CHCl3 (1:1) extract of the aerial parts of Ipomoea pes-caprae. Ipomeolides A (1) and B (2) are macrolactone analogues of the rare (11 R)-jalapinolic acid, and macrolactonization occurred at C-2 of the second saccharide moiety. Compounds 1 and 2 show structural variation even in the pentasaccharide core. The structures of 1 and 2 were established by a combination of spectroscopic techniques as well as chemical modifications such as acetyl and acetonide derivatives as well as hydrolysis products. The new glycosidic acid was named ipomeic acid (1c). Compounds 1, 1b, and 2b were evaluated for cytotoxicity against human tumor cell lines. Compounds 1b and 2b were not effective on epithelial cells, but affected survival of K-562, which is of hematopoietic origin. A sublethal concentration of compound 1 (4 µM) when used in combination with 1 µM doxorubicin, an anticancer agent, significantly enhanced cytotoxicity to tumor cells. Such combined synergistic potency against leukemia cells and the absence of effects on epithelial cells may be beneficial for chemotherapy with minimal side effects to treat CML (chronic myeloid leukemia) malignancies.