Nitric Oxide Dependent Metabolism Regulates the Proliferation and Differentiation of Stress Erythroid Progenitors

Abstract Infection and tissue damage induce inflammation, which increases myelopoiesis at the expense of steady state erythropoiesis. Stress erythropoiesis is induced to compensate for the loss of erythroid output until the inflammation is resolved and bone marrow erythropoiesis can resume. Steady state erythropoiesis constantly produces erythrocytes, while stress erythropoiesis generates a bolus of new erythrocytes through the rapid expansion of immature progenitor cells which is followed by the synchronous differentiation of progenitors. We hypothesized that the proliferation of early progenitor cells and their transition to differentiation is regulated by changes in metabolism. Metabolomics and isotope tracing analysis was performed to assess the intracellular metabolic profiles in proliferating progenitors isolated from in vitro stress erythropoiesis cultures. We observed an active engagement of glucose metabolism in glycolysis and anabolic biosynthesis, while the levels of TCA intermediates suggested that TCA cycle and mitochondrial respiration were blocked. Concomitantly, inducible nitric oxide synthase (iNOS) was induced in progenitor cells to increase the production of nitric oxide (NO), which was demonstrated to be crucial for proliferating progenitor metabolism. Inhibition or genetic mutation of iNOS decreased NO levels resulting in the suppression of progenitor proliferation in vitro and in vivo. As evaluated by RNA-seq, inhibition of iNOS suppressed cell proliferation-related pathways including cell cycle and nucleotide metabolism, while upregulating erythroid differentiation genes. These data suggest that iNOS-derived NO production establishes a metabolism that promotes the proliferation of progenitor cells while inhibiting their differentiation. Notably, proliferating progenitor cells displayed low levels of the metabolite itaconate and decreased expression of Immunoresponsive gene 1 (Irg1), the enzyme that catalyzes itaconate synthesis from cis-aconitate. Further analysis showed that the addition of 4-Octyl itaconate (OI), a cell-permeable itaconate derivative, inhibited iNOS-derived NO production by activating nuclear factor erythroid 2-related factor 2 (Nrf2), which in turn impaired progenitor expansion. These results indicate that itaconate production is inhibited to enable the accumulation of NO and the NO dependent metabolism required for progenitor cell proliferation during the initial expansion stage of stress erythropoiesis. In contrast, the transition to differentiation is marked by elevated itaconate synthesis, Nrf2 activation, and attenuated iNOS expression. We hypothesized that the inhibition of NO production alters metabolism and in concert with new cell signaling removes the NO-dependent inhibition of erythroid program, which allows the differentiation of progenitor cells. We tested this mechanism by examining the effects of iNOS inhibitors and mutants in iNOS, Irg1 and Nrf2 on progenitor cells isolated from differentiation cultures. iNOS deficiency led to the activation of erythroid transcriptional program, and increased numbers of mature progenitors as well as stress BFU-Es. In contrast, Irg1 and Nrf2 mutants showed impaired transition to erythroid differentiation, while they had elevated iNOS expression and NO production. Further analysis showed that treatment with either OI or iNOS inhibitor inhibited NO production in Irg1 and Nrf2 deficient progenitors, and consequently rescued the defects in erythroid differentiation. These data support a model in which inflammation inhibits steady state erythropoiesis, while at the same time promoting stress erythropoiesis to maintain homeostasis. Our work reveals a dynamic and tight coordination between pro-inflammatory signals and progenitor cell metabolism in regulating the proliferation and differentiation of stress erythroid progenitors, and highlights the therapeutic potential of targeting metabolic and inflammatory signaling pathways in inflammatory anemia. Disclosures Paulson: Forma Therapeutics: Consultancy.

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Erythropoietin can promote erythroid progenitor survival by repressing apoptosis through Bcl-XL and Bcl-2

Blood ◽

10.1182/blood.v88.5.1576.1576 ◽

1996 ◽

Vol 88 (5) ◽

pp. 1576-1582 ◽

Cited By ~ 243

Author(s):

M Silva ◽

D Grillot ◽

A Benito ◽

C Richard ◽

G Nunez ◽

...

Keyword(s):

Progenitor Cells ◽

Apoptotic Cell ◽

Ectopic Expression ◽

Erythroid Differentiation ◽

Erythroid Progenitors ◽

Erythroid Progenitor ◽

Survival Factor ◽

Erythroid Progenitor Cells ◽

Survival Signal ◽

Proliferation And Differentiation

Abstract Erythropoietin (Epo), the hormone that is the principal regulator of red blood cell production, interacts with high-affinity receptors on the surface of erythroid progenitor cells and maintains their survival. Epo has been shown to promote cell viability by repressing apoptosis; however, the molecular mechanism involved is unclear. In the present studies we have examined whether Epo acts as a survival factor through the regulation of the bcl-2 family of apoptosis-regulatory genes. We addressed this issue in HCD-57, a murine erythroid progenitor cell line that requires Epo for proliferation and survival. When HCD-57 cells were cultured in the absence of Epo, Bcl-2 and Bcl-XL but not Bax were downregulated, and the cells underwent apoptotic cell death. HCD-57 cells infected with a retroviral vector encoding human Bcl-XL or Bcl-2 rapidly stopped proliferating but remained viable in the absence of Epo. Furthermore, endogenous levels of bcl-2 and bcl-XL were downregulated after Epo withdrawal in HCD-57 cells that remained viable through ectopic expression of human Bcl-XL, further indicating that Epo specifically maintains the expression of bcl-2 and bcl-XL. We also show that HCD-57 rescued from apoptosis by ectopic expression of Bcl-XL can undergo erythroid differentiation in the absence of Epo, demonstrating that a survival signal but not Epo itself is necessary for erythroid differentiation of HCD-57 progenitor cells. Thus, we propose a model whereby Epo functions as a survival factor by repressing apoptosis through Bcl-XL and Bcl-2 during proliferation and differentiation of erythroid progenitors.

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Erythropoietin can promote erythroid progenitor survival by repressing apoptosis through Bcl-XL and Bcl-2

Blood ◽

10.1182/blood.v88.5.1576.bloodjournal8851576 ◽

1996 ◽

Vol 88 (5) ◽

pp. 1576-1582 ◽

Cited By ~ 10

Author(s):

M Silva ◽

D Grillot ◽

A Benito ◽

C Richard ◽

G Nunez ◽

...

Keyword(s):

Progenitor Cells ◽

Apoptotic Cell ◽

Ectopic Expression ◽

Erythroid Differentiation ◽

Erythroid Progenitors ◽

Erythroid Progenitor ◽

Survival Factor ◽

Erythroid Progenitor Cells ◽

Survival Signal ◽

Proliferation And Differentiation

Erythropoietin (Epo), the hormone that is the principal regulator of red blood cell production, interacts with high-affinity receptors on the surface of erythroid progenitor cells and maintains their survival. Epo has been shown to promote cell viability by repressing apoptosis; however, the molecular mechanism involved is unclear. In the present studies we have examined whether Epo acts as a survival factor through the regulation of the bcl-2 family of apoptosis-regulatory genes. We addressed this issue in HCD-57, a murine erythroid progenitor cell line that requires Epo for proliferation and survival. When HCD-57 cells were cultured in the absence of Epo, Bcl-2 and Bcl-XL but not Bax were downregulated, and the cells underwent apoptotic cell death. HCD-57 cells infected with a retroviral vector encoding human Bcl-XL or Bcl-2 rapidly stopped proliferating but remained viable in the absence of Epo. Furthermore, endogenous levels of bcl-2 and bcl-XL were downregulated after Epo withdrawal in HCD-57 cells that remained viable through ectopic expression of human Bcl-XL, further indicating that Epo specifically maintains the expression of bcl-2 and bcl-XL. We also show that HCD-57 rescued from apoptosis by ectopic expression of Bcl-XL can undergo erythroid differentiation in the absence of Epo, demonstrating that a survival signal but not Epo itself is necessary for erythroid differentiation of HCD-57 progenitor cells. Thus, we propose a model whereby Epo functions as a survival factor by repressing apoptosis through Bcl-XL and Bcl-2 during proliferation and differentiation of erythroid progenitors.

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Inverse PAR1 Activity of Hematopoietic Stem Cells and BM Stromal Cells Mediates G-CSF-Induced Mobilization By Regulation of Nitric Oxide Generation

Blood ◽

10.1182/blood.v128.22.3370.3370 ◽

2016 ◽

Vol 128 (22) ◽

pp. 3370-3370 ◽

Cited By ~ 1

Author(s):

Neta Nevo ◽

Shiri Gur-Cohen ◽

Orit Kollet ◽

Francesca Avemaria ◽

Seymen Avci ◽

...

Keyword(s):

Nitric Oxide ◽

Stem Cells ◽

Stem Cell ◽

Steady State ◽

Progenitor Cells ◽

Small Animal ◽

Predictive Marker ◽

No Production ◽

Hematopoietic Stem ◽

Neutrophil Recovery

Abstract Hematopoietic stem and progenitor cell (HSPC) egress from the bone marrow (BM) to the circulation is tightly regulated and is accelerated during stress conditions, a process utilized for BM harvest. Recently, we demonstrated that mouse long term repopulating hematopoietic stem cell (LT-HSC) BM retention and their rapid release to the blood circulation are governed by a switch in nitric oxide (NO) generation via distinct coagulation-related protease activated receptor 1 (PAR1) cascades (Gur-Cohen S. et al., NM, 2016). Herein we report that surface PAR1 expression can be exploited and serve as a positive predictive marker for the efficiency of human CD34+ HSPC mobilization among healthy donors in clinical G-CSF-induced mobilization for matched allogeneic transplantations. We found that PAR1 expression on circulating leukocytes before G-CSF administration was positively correlated with higher yields of mobilized leukocytes after stimulation (P<0.001). In addition, the percentage of PAR1-expressing CD34+ HSPCs in the blood and their absolute numbers before initiation of mobilization positively correlated with peripheral blood leukocyte counts (P<0.001) and yield of mobilized CD34+ HSPCs (P=0.002). Importantly, tracing the consequences of the follow-up BM transplantation, we found accelerated neutrophil recovery in patients transplanted with mobilized cells expressing higher surface PAR1 levels at baseline (P=0.074), shortening the time for neutrophil recovery (neutrophil count>0.5 x 109/L) from an average of 14.6 days to an average of 11 days post-transplant. In addition, a trend of accelerated platelet production was documented to be related with higher PAR1 expression by circulating leukocytes prior to G-CSF stimulation. Consequently, poor mobilizer donors were characterized by extremely low surface PAR1 expression on circulating CD34+ cells prior to G-CSF stimulations. Herein we present a case report of a thrombophilic donor carrying the MTHFR mutation, expressing exceedingly low PAR1 levels at baseline, with the outcome of inadequate numbers of mobilized CD34+ HSPC in the blood following G-CSF treatments. To further gain insight into the role played by PAR1 signaling in the regulation of G-CSF-induced HSPC mobilization, we used mice as a functional preclinical small animal model. We found that antagonizing PAR1 signaling attenuated both steady state release and G-CSF-induced HSPC mobilization. Furthermore, co-administration of G-CSF with PAR1 antagonist attenuated secretion of BM stromal CXCL12 and abrogated upregulation of surface CXCR4 and PAR1 expression by BM HSPCs, all leading to significantly reduced HSPC migration, differentiation and mobilization. In support, PAR1-/- mice failed to efficiently mobilize HSPCs in response to G-CSF compared to wild type counterparts. Enforced HSPC recruitment by G-CSF treatments dramatically accelerated PAR1-dependent NO production by eNOS, known to promote TACE-mediated EPCR shedding and rapid LT-HSC mobilization. Concomitantly, circulating steady state and G-CSF-mobilized stem cells lack surface EPCR expression. Intriguingly, while EPCR expression by primitive BM stem cells was transiently reduced after G-CSF treatments, antagonizing PAR1 signaling along with G-CSF stimuli blocked NO generation and synchronically expanded BM EPCR+ LT-HSC and their supportive stromal progenitor cells (MSPCs), as confirmed by increased repopulation in transplanted mice. Finally, we report an inverse PAR1 expression and regulation by BM HSPC and stromal MSPCs in mediating G-CSF-induced mobilization. G-CSF induced elevation of PAR1 expression on BM HSPCs, providing the driving force for their enhanced NO mediated migration, proliferation, differentiation and recruitment to the circulation. Contrary, the levels of PAR1 expression were reduced on MSPCs in response to G-CSF treatment, and following NO generation by eNOS activity and CXCL12 secretion, resulted in reduced mesenchymal differentiation leading to accumulated numbers of immature mesenchymal (CFU-F) and osteoblast (CFU-OB) progenitor cells. Taken together, our study identifies and highlights inverse PAR1 signaling and NO generation as essential regulator of G-CSF induced HSPC mobilization and MSPC development opening new avenues to advance therapeutics for enhancing clinical G-CSF induced stem cell mobilization and transplantation protocols. Disclosures Ruf: Iconic Therapeutics: Consultancy.

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AMD3100 Signals Via the Nervous System, Inducing Release to the Circulation of Bone Marrow SDF-1, Which Is Crucial for Progenitor Cell Mobilization.

Blood ◽

10.1182/blood.v108.11.1315.1315 ◽

2006 ◽

Vol 108 (11) ◽

pp. 1315-1315 ◽

Cited By ~ 1

Author(s):

Ayelet Dar ◽

Alexander Kalinkovich ◽

Neta Netzer ◽

Raanan Margalit ◽

Amir Schajnovitz ◽

...

Keyword(s):

Bone Marrow ◽

Nervous System ◽

Steady State ◽

Progenitor Cells ◽

Peripheral Blood ◽

Neutralizing Antibodies ◽

Progenitor Cell ◽

Hematopoietic Progenitor Cells ◽

Hematopoietic Progenitor

Abstract AMD3100, a bicyclam antagonist of the chemokine receptor CXCR4 in vitro, has been shown to induce rapid mobilization of human and murine maturing leukocytes and immature hematopoietic stem and progenitor cells in vivo. In addition, AMD3100 combined with G-CSF, synergistically augments mobilization of human progenitor cells (Broxmeyer & Srour et al, JEM, 2005). However, the mechanism of AMD3100-induced mobilization is currently poorly understood. We report that AMD3100-induced mobilization in mice was accompanied with rapid increase in functional SDF-1 concentrations in the circulation and their parallel decrease in the bone marrow within 1 hour. Biotinylated SDF-1 (bSDF-1) directly injected into the femur was detected in the peripheral blood, adjacent bones and spleen as early as 10 minutes post administration. Interestingly, AMD3100 induced significant elevations in bone marrow-derived bSDF-1 concentrations in the peripheral blood. Similarly, G-CSF induced mobilization was initiated (24 hours post a single injection of G-CSF), by SDF-1 release to the circulation. Administration of neutralizing antibodies against CXCR4 to either untreated or AMD3100 treated mice markedly reduced SDF-1 levels in the peripheral blood, coinciding with increased retention levels of this ligand in the bone marrow. In vitro, AMD3100 directly induced SDF-1 release from the human osteoblast cell line MG-63 in a bell shaped dose response. Inhibition of CXCR4-dependent release of SDF-1 during homeostasis or upon treatment with AMD3100, correlated with selective reduction in recruitment of hematopoietic progenitor cells but not mature leukocytes to the circulation. Importantly, injection of neutralizing antibodies against SDF-1 (but not matched control antibodies) resulted in decreased steady state egress and AMD3100-induced mobilization of hematopoietic progenitor cells. Rapid recruitment (within 1 hour) of hematopoietic progenitor cells and maturing leukocytes out of the bone marrow as well as SDF-1 release were dependent on signals from the nervous system. Administration of the b2 adrenergic agonist (clenbuterol) inhibited endogenous SDF-1 and exogenous bSDF-1 release to the circulation and reduced progenitor cell egress, both during steady state and AMD3100-induced mobilization, while administration of the b2 adrenergic antagonist (propranolol) resulted in opposite effects. Based on our results we propose a model in which egress and mobilization of immature progenitor cells differs from that of maturing leukocytes and is more dependent on SDF-1/CXCR4 interactions. In addition to hematopoietic progenitor cells, also bone marrow stromal cells induce homeostatic secretion of SDF-1, which is increased during mobilization and stress induced recruitment. Secretion of this ligand is also CXCR4-dependent, revealing orchestrated mutual and reciprocal SDF1/CXCR4 interactions and a cross-talk with the nervous system, which regulates progenitor cell egress and recruitment.

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Integrated FGF and BMP signaling controls the progression of progenitor cell differentiation and the emergence of pattern in the embryonic anterior pituitary

Development ◽

10.1242/dev.125.6.1005 ◽

1998 ◽

Vol 125 (6) ◽

pp. 1005-1015 ◽

Cited By ~ 16

Author(s):

J. Ericson ◽

S. Norlin ◽

T.M. Jessell ◽

T. Edlund

Keyword(s):

Progenitor Cells ◽

Anterior Pituitary ◽

Progenitor Cell ◽

Cell Types ◽

Bmp Signaling ◽

Cell Identity ◽

Coordinate Control ◽

Signaling Function ◽

Proliferation And Differentiation

The mechanisms by which inductive signals control the identity, proliferation and timing of differentiation of progenitor cells in establishing spatial pattern in developing vertebrate tissues remain poorly understood. We have addressed this issue in the embryonic anterior pituitary, an organ in which distinct hormone cell types are generated in a precise temporal and spatial order from an apparently homogenous ectodermal primordium. We provide evidence that in this tissue the coordinate control of progenitor cell identity, proliferation and differentiation is imposed by spatial and temporal restrictions in FGF- and BMP-mediated signals. These signals derive from adjacent neural and mesenchymal signaling centers: the infundibulum and ventral juxtapituitary mesenchyme. The infundibulum appears to have a dual signaling function, serving initially as a source of BMP4 and subsequently of FGF8. The ventral juxtapituitary mesenchyme appears to serve as a later source of BMP2 and BMP7. In vitro, FGFs promote the proliferation of progenitor cells, prevent their exit from the cell cycle and contribute to the specification of progenitor cell identity. BMPs, in contrast, have no apparent effect on cell proliferation but instead appear to act with FGFs to control the initial selection of thyrotroph and corticotroph progenitor identity.

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Heterozygous Mutations in DDX41 Cause Erythroid Progenitor Cell Defects

Blood ◽

10.1182/blood-2021-153544 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 148-148

Author(s):

Timothy M Chlon ◽

Emily Stepanchick ◽

Analise Sulentic ◽

Kathleen Hueneman ◽

Daniel Starczynowski

Keyword(s):

Bone Marrow ◽

Myelodysplastic Syndrome ◽

Progenitor Cells ◽

Liquid Culture ◽

Erythroid Differentiation ◽

Erythroid Progenitors ◽

Erythroid Progenitor ◽

Colony Assays

Abstract Germline mutations in the RNA Helicase gene DDX41 cause inherited susceptibility to Myelodysplastic Syndrome (MDS) and Acute Myeloid Leukemia (AML). These mutations are always heterozygous and are typically frameshifts, causing loss of protein expression. We recently reported that at least one functional copy of DDX41 is essential for hematopoiesis, and that DDX41 is required for ribosome biogenesis. While biallelic DDX41 mutations cause dramatic defects in hematopoiesis, the role of heterozygous mutations in Myelodysplastic Syndrome pathogenesis is not yet understood. Recent clinical studies have pointed out that some patients bearing germline DDX41 mutations have idiopathic cytopenias of unknown significance (ICUS) prior to MDS onset, suggesting that underlying hematopoietic defects precede and potentially contribute to the onset of MDS/AML (Choi et al., Haemotologica 2021). It has also been noted that the majority of DDX41-mutant MDS patients have refractory anemia, indicating that the erythroid lineage is particularly effected in these patients (Sebert et al., Blood 2019). Since ribosome defects are a common cause of inherited anemias and also contribute to MDS pathogenesis, we characterized the effect of heterozygous DDX41 mutations on erythropoiesis in murine and human models. Mice that have been transplanted with Ddx41 +/- bone marrow develop anemia at 12-15 months post-transplant, indicating that detection of erythroid defects in vivo is aging-dependent. We characterized the effect of heterozygosity of Ddx41 on erythroid progenitor function in vitro and found that Ddx41 +/- bone marrow from young mice yields fewer BFU-E in colony assays but comparable numbers of myeloid colonies. Liquid culture erythroid differentiation of Ddx41 +/- bone marrow produces fewer CD71+ Ter119+ progenitors than controls. To characterize the effect of heterozygous DDX41 mutations on human erythropoiesis, we generated induced pluripotent stem cells bearing heterozygous frameshift mutations in DDX41 using CRISPR. We found that these DDX41 +/- iPSC lines produced CD43+/CD34+ hematopoietic progenitor cells (HPC) with equal efficiency as unmodified control iPSC. However, once these HPC were induced to differentiate down the erythroid lineage in liquid culture, they made fewer CD71+ GLYA+ erythroid progenitors and fewer hemoglobinized cells. The DDX41 +/- HPC also produced fewer BFU-E in colony assays. Mechanistically, we found that the in vitro-derived erythroid progenitors from both mice and human iPSC had decreased protein translation, suggesting that ribosome defects underlie the observed erythroid differentiation defects. In diseases such as Diamond Blackfan Anemia and Dyskeratosis Congenita, ribosome defects lead to p53 activation which reduces cell cycle progression in erythroid progenitors. To test the role of p53 in the erythroid defects caused by Ddx41 heterozygosity, we crossed Ddx41 +/- mice with p53-knockout mice and found that loss of p53 fully rescued the BFU-E colony formation of Ddx41 +/- bone marrow HPC. We confirmed this finding using CRISPR-mediated knockout of p53 in Ddx41 +/- BM HPC. Collectively, these results suggest that a mild ribosome defect in DDX41 +/- HPC causes a deficit in erythropoiesis that results in anemia with aging. It is likely that this anemia causes stress in the bone marrow and a selective environment in which malignant hematopoietic stem and progenitor cells arise, leading to MDS and AML. Disclosures Starczynowski: kurome Inc: Consultancy.

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Bone Marrow Niches for Skeletal Progenitor Cells and their Inhabitants in Health and Disease

Current Stem Cell Research & Therapy ◽

10.2174/1574888x14666190123161447 ◽

2019 ◽

Vol 14 (4) ◽

pp. 305-319 ◽

Cited By ~ 4

Author(s):

Marietta Herrmann ◽

Franz Jakob

Keyword(s):

Bone Marrow ◽

Progenitor Cells ◽

Progenitor Cell ◽

Adult Stem Cells ◽

Current Knowledge ◽

Cell Populations ◽

Surface Markers ◽

Bone Marrow Niches

The bone marrow hosts skeletal progenitor cells which have most widely been referred to as Mesenchymal Stem or Stromal Cells (MSCs), a heterogeneous population of adult stem cells possessing the potential for self-renewal and multilineage differentiation. A consensus agreement on minimal criteria has been suggested to define MSCs in vitro, including adhesion to plastic, expression of typical surface markers and the ability to differentiate towards the adipogenic, osteogenic and chondrogenic lineages but they are critically discussed since the differentiation capability of cells could not always be confirmed by stringent assays in vivo. However, these in vitro characteristics have led to the notion that progenitor cell populations, similar to MSCs in bone marrow, reside in various tissues. MSCs are in the focus of numerous (pre)clinical studies on tissue regeneration and repair.Recent advances in terms of genetic animal models enabled a couple of studies targeting skeletal progenitor cells in vivo. Accordingly, different skeletal progenitor cell populations could be identified by the expression of surface markers including nestin and leptin receptor. While there are still issues with the identity of, and the overlap between different cell populations, these studies suggested that specific microenvironments, referred to as niches, host and maintain skeletal progenitor cells in the bone marrow. Dynamic mutual interactions through biological and physical cues between niche constituting cells and niche inhabitants control dormancy, symmetric and asymmetric cell division and lineage commitment. Niche constituting cells, inhabitant cells and their extracellular matrix are subject to influences of aging and disease e.g. via cellular modulators. Protective niches can be hijacked and abused by metastasizing tumor cells, and may even be adapted via mutual education. Here, we summarize the current knowledge on bone marrow skeletal progenitor cell niches in physiology and pathophysiology. We discuss the plasticity and dynamics of bone marrow niches as well as future perspectives of targeting niches for therapeutic strategies.

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Effects of chitosan on nitric oxide production and inducible nitric oxide synthase activity and mRNA expression in weaned piglets

Czech Journal of Animal Science ◽

10.17221/8405-cjas ◽

2018 ◽

Vol 60 (No. 8) ◽

pp. 359-366

Author(s):

J. Li ◽

B. Shi ◽

S. Yan ◽

L. Jin ◽

Y. Guo ◽

...

Keyword(s):

Nitric Oxide ◽

Mrna Expression ◽

Inducible Nitric Oxide Synthase ◽

No Production ◽

Weaned Piglets ◽

Inos Activity ◽

Oxide Synthase ◽

Inducible Nitric Oxide

The effects of chitosan on nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) activity and gene expression in vivo or vitro were investigated in weaned piglets. In vivo, 180 weaned piglets were assigned to five dietary treatments with six replicates. The piglets were fed on a basal diet supplemented with 0 (control), 100, 500, 1000, and 2000 mg chitosan/kg feed, respectively. In vitro, the peripheral blood mononuclear cells (PBMCs) from a weaned piglet were cultured respectively with 0 (control), 40, 80, 160, and 320 µg chitosan/ml medium. Results showed that serum NO concentrations on days 14 and 28 and iNOS activity on day 28 were quadratically improved with increasing chitosan dose (P < 0.05). The iNOS mRNA expressions were linearly or quadratically enhanced in the duodenum on day 28, and were improved quadratically in the jejunum on days 14 and 28 and in the ileum on day 28 (P < 0.01). In vitro, the NO concentrations, iNOS activity, and mRNA expression in unstimulated PBMCs were quadratically enhanced by chitosan, but the improvement of NO concentrations and iNOS activity by chitosan were markedly inhibited by N-(3-[aminomethyl] benzyl) acetamidine (1400w) (P < 0.05). Moreover, the increase of NO concentrations, iNOS activity, and mRNA expression in PBMCs induced by lipopolysaccharide (LPS) were suppressed significantly by chitosan (P < 0.05). The results indicated that the NO concentrations, iNOS activity, and mRNA expression in piglets were increased by feeding chitosan in a dose-dependent manner. In addition, chitosan improved the NO production in unstimulated PBMCs but inhibited its production in LPS-induced cells, which exerted bidirectional regulatory effects on the NO production via modulated iNOS activity and mRNA expression.

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New Cyclic Diarylheptanoids from Platycarya strobilacea

Molecules ◽

10.3390/molecules25246034 ◽

2020 ◽

Vol 25 (24) ◽

pp. 6034

Author(s):

Wen-bing Ding ◽

Rui-yuan Zhao ◽

Guan-hua Li ◽

Bing-lei Liu ◽

Hua-liang He ◽

...

Keyword(s):

Nitric Oxide ◽

Pc12 Cells ◽

Stem Bark ◽

No Production ◽

Electronic Circular Dichroism ◽

Pheochromocytoma Pc12 ◽

Pheochromocytoma Pc12 Cells ◽

Induced Apoptosis ◽

Circular Dichroïsm

Five new cyclic diarylheptanoids (platycary A–E, compounds 1–5) and three previously identified analogues (i.e., phttyearynol (compound 6), myricatomentogenin (compound 7), and juglanin D (compound 8)) were isolated from the stem bark of Platycarya strobilacea. The structures of these compounds were determined using NMR, HRESIMS, and electronic circular dichroism (ECD) data. The cytotoxicity of compounds 1–5 and their ability to inhibit nitric oxide (NO) production, as well as protect against the corticosterone-induced apoptosis of Pheochromocytoma (PC12) cells, were evaluated in vitro using the appropriate bioassays. Compounds 1 and 2 significantly inhibited the corticosterone-induced apoptosis of PC12 cells at a concentration of 20 μΜ.

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