Hemophagocytic lymphohistiocytosis is a life-threatening systemic hyperinflammatory disorder with primary and secondary forms. Primary hemophagocytic lymphohistiocytosis is associated with inherited ...defects in various genes that affect the immunological cytolytic pathway. Secondary hemophagocytic lymphohistiocytosis is not inherited, but complicates various medical conditions including infections, autoinflammatory/autoimmune diseases, and malignancies. When senescence-accelerated mice (SAMP1/TA-1) with latent deterioration of immunological function and senescence-resistant control mice (SAMR1) were treated repeatedly with lipopolysaccharide, SAMP1/TA-1 mice displayed the clinicopathological features of hemophagocytic lymphohistiocytosis such as hepatosplenomegaly, pancytopenia, hypofibrinogenemia, hyperferritinemia, and hemophagocytosis. SAMR1 mice showed no features of hemophagocytic lymphohistiocytosis. Lipopolysaccharide induced upregulation of proinflammatory cytokines such as interleukin-1β, interleukin-6, tumor necrosis factor-α, and interferon-γ, and interferon-γ-inducible chemokines such as c-x-c motif chemokine ligands 9 and 10 in the liver and spleen in both SAMP1/TA-1 and SAMR1 mice. However, upregulation of proinflammatory cytokines and interferon-γ-inducible chemokines in the liver persisted for longer in SAMP1/TA-1 mice than in SAMR1 mice. In addition, the magnitude of upregulation of interferon-γ in the liver and spleen after lipopolysaccharide treatment was greater in SAMP1/TA-1 mice than in SAMR1 mice. Furthermore, lipopolysaccharide treatment led to a prolonged increase in the proportion of peritoneal M1 macrophages and simultaneously to a decrease in the proportion of M2 macrophages in SAMP1/TA-1 mice compared with SAMR1 mice. Lipopolysaccharide appeared to induce a hyperinflammatory reaction and prolonged inflammation in SAMP1/TA-1 mice, resulting in features of secondary hemophagocytic lymphohistiocytosis. Thus, SAMP1/TA-1 mice represent a useful mouse model to investigate the pathogenesis of bacterial infection-associated secondary hemophagocytic lymphohistiocytosis.
Lipopolysaccharide (LPS) treatment induced hemophagocytic lymphohistiocytosis in senescence-accelerated mice (SAMP1/TA-1), but not in senescence-resistant control mice (SAMR1). SAMP1/TA-1 treated ...with LPS exhibited functional impairment of the hematopoietic microenvironment, which disrupted the dynamics of hematopoiesis. Macrophages are a major component of the bone marrow (BM) hematopoietic microenvironment, which regulates hematopoiesis. Qualitative and quantitative changes in activated macrophages in LPS-treated SAMP1/TA-1 are thought to contribute to the functional deterioration of the hematopoietic microenvironment. Thus, we examined the polarization of pro-inflammatory (M1) and anti-inflammatory (M2) macrophages, and the dynamics of macrophage production in the BM of SAMP1/TA-1 and SAMR1 after LPS treatment. After LPS treatment, the proportions of M1 and M2 macrophages and the numbers of macrophage progenitor (CFU-M) cells increased in both SAMP1/TA-1 and SAMR1. However, compared to the SAMR1, the increase in the M1 macrophage proportion was prolonged, and the increase in the M2 macrophage proportion was delayed. The increase in the number of CFU-M cells was prolonged in SAMP1/TA-1 after LPS treatment. In addition, the levels of transcripts encoding an M1 macrophage-inducing cytokine (interferon-γ) and macrophage colony-stimulating factor were markedly increased, and the increases in the levels of transcripts encoding M2 macrophage-inducing cytokines (interleukin (IL)-4, IL-10, and IL-13) were delayed in SAMP1/TA-1 when compared to SAMR1. Our results suggest that LPS treatment led to the severely imbalanced polarization of activated M1/M2 macrophages accompanied by a prolonged increase in macrophage production in the BM of SAMP1/TA-1, which led to the impairment of the hematopoietic microenvironment, and disrupted the dynamics of hematopoiesis.
Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening systemic hyper-inflammatory disorder. The mortality of HLH is higher in the elderly than in young adults. Senescence-accelerated mice ...(SAMP1/TA-1) exhibit characteristic accelerated aging after 30 weeks of age, and HLH-like features, including hematopoietic organ damage, are seen after lipopolysaccharide (LPS) treatment. Thus, SAMP1/TA-1 is a useful model of hematological pathophysiology in the elderly with HLH. In this study, dosing of SAMP1/TA-1 mice with LPS revealed that the suppression of myelopoiesis and B-lymphopoiesis was more severe in aged mice than in young mice. The bone marrow (BM) expression of genes encoding positive regulators of myelopoiesis (G-CSF, GM-CSF, and IL-6) and of those encoding negative regulators of B cell lymphopoiesis (TNF-α) increased in both groups, while the expression of genes encoding positive-regulators of B cell lymphopoiesis (IL-7, SDF-1, and SCF) decreased. The expression of the GM-CSF-encoding transcript was lower in aged mice than in young animals. The production of GM-CSF by cultured stromal cells after LPS treatment was also lower in aged mice than in young mice. The accumulation of the TNF-α-encoding transcript and the depletion of the IL-7-encoding transcript were prolonged in aged mice compared to young animals. LPS dosing led to a prolonged increase in the proportion of BM M1 macrophages in aged mice compared to young animals. The expression of the gene encoding p16
and the proportion of β-galactosidase- and phosphorylated ribosomal protein S6-positive cells were increased in cultured stromal cells from aged mice compared to those from young animals, while the proportion of Ki67-positive cells was decreased in stromal cells from aged mice. Thus, age-related deterioration of stromal cells probably causes the suppression of hematopoiesis in aged mice. This age-related latent organ dysfunction may be exacerbated in elderly people with HLH, resulting in poor prognosis.
Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening systemic hyperinflammatory disorder. We found recently that repeated lipopolysaccharide (LPS) treatment induces HLH-like features in ...senescence-accelerated mice (SAMP1/TA-1) but not in senescence-resistant control mice (SAMR1). In this study, we analyzed the dynamics of hematopoiesis in this mouse model of HLH. When treated repeatedly with LPS, the numbers of myeloid progenitor cells (CFU-GM) and B-lymphoid progenitor cells (CFU-preB) in the bone marrow (BM) rapidly decreased after each treatment in both strains. The number of CFU-GM in SAMP1/TA-1 and SAMR1, and of CFU-preB in SAMR1, returned to pretreatment levels by 7 days after each treatment. However, the recovery in the number of CFU-preB in SAMP1/TA-1 was limited. In both strains, the BM expression of genes encoding positive regulators of myelopoiesis (granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), and interleukin (IL)-6), and negative regulators of B lymphopoiesis (tumor necrosis factor (TNF)-α) was increased. The expression of genes encoding positive regulators of B lymphopoiesis (stromal-cell derived factor (SDF)-1, IL-7, and stem cell factor (SCF)) was persistently decreased in SAMP1/TA-1 but not in SAMR1. Expression of the gene encoding p16
INK4a
and the proportion of β-galactosidase-positive cells were increased in cultured stromal cells obtained from LPS-treated SAMP1/TA-1 but not in those from LPS-treated SAMR1. LPS treatment induced qualitative changes in stromal cells, which comprise the microenvironment supporting appropriate hematopoiesis, in SAMP1/TA-1; these stromal cell changes are inferred to disrupt the dynamics of hematopoiesis. Thus, hematopoietic tissue is one of the organs that suffer life-threatening damage in HLH.
Previously, we established a three-dimensional (3D) bone marrow culture system that maintains normal hematopoiesis, including prolongation of hematopoietic stem cell proliferation and ...differentiation. To analyze the role of bone marrow stromal cells that compose the microenvironment, the growth of a leukemic cell line (K562) in the 3D condition and with arginine deprivation stress was compared with two-dimensional stromal cell monolayers (2D) and suspension cultures without stromal cells (stroma (−)). Arginine is essential for the proliferation and differentiation of erythrocytes. The proliferation and differentiation of K562 cells cultured in the 3D system were stabilized compared with cells in 2D or stroma (−). Furthermore, the number of K562 cells in the G0/G1 phase in 3D was increased significantly compared with cells grown in 2D or stroma (−). Interestingly, the mRNA expression of various hematopoietic growth factors of stromal cells in 3D was not different from 2D, even though supportive activity on K562 cell growth was observed in the arginine deprivation condition. Thus, the hematopoietic microenvironment involves multi-dimensional and complex systems including biochemical and physiochemical factors that regulate quiescence, proliferation, activation, and differentiation of normal hematopoietic cells and cloned leukemic cells. Our 3D culture system may be a valuable new tool for investigating leukemic cell-stromal cell interactions in vitro.
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•The autonomous hyperproliferation of K562 cells was suppressed in the 3D niche.•K562 cell differentiation was induced in the 3D niche despite the lack of arginine.•Increased K562 cells in the G0/G1 phase were observed in the 3D stromal niche.•Stroma in the 3D niche affects leukemic cell growth by controlling the cell cycle.
Chronic lipopolysaccharide (LPS) exposure to mice reduces the lymphoid compartment and skews the hematopoietic cell compartment toward myeloid-cells, which is considered to be a direct effect of LPS ...on hematopoietic stem cells. However, the effect of chronic LPS exposure on stromal-cells, which compose the hematopoietic microenvironment, has not been elucidated. Here, we investigated early- and late-phase effects of repeated LPS exposure on stromal-cells. During the early phase, when mice were treated with 5 or 25 µg LPS three times at weekly intervals, the numbers of myeloid-progenitor (colony forming unit-granulocyte macrophage (CFU-GM)) cells and B lymphoid-progenitor (CFU-preB) cells in the bone-marrow (BM) rapidly decreased after each treatment. The number of CFU-GM cells recovered from the initial decrease and then increased to levels higher than pretreatment levels, whereas the number of CFU-preB cells remained lower than pretreatment levels. In the BM, expression of genes for positive-regulators of myelopoiesis including granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), and interleukin (IL)-6 and negative-regulators of B lymphopoiesis including tumor necrosis factor (TNF)-α was up-regulated, whereas expression of positive-regulators of B lymphopoiesis including stromal cell-derived factor (SDF)-1, IL-7, and stem cell factor (SCF) was down-regulated. During the late phase, the number of CFU-preB cells remained lower than pretreatment levels 70 d after the first treatments with 5 and 25 µg LPS, whereas the number of CFU-GM cells returned to pretreatment levels. IL-7 gene expression in the BM remained down-regulated, whereas gene-expression levels of SDF-1 and SCF were restored. Thus, chronic LPS exposure may impair stromal-cell function, resulting in prolonged suppression of B lymphopoiesis, which may appear to be senescence similar to the hematological phenotype.
Tetrahydrobiopterin (BH4) is an essential cofactor of nitric oxide synthase (NOS) and aromatic amino acid hydroxylases. BH4 and 7,8-dihydrobiopterin (BH2) are metabolically interchangeable at the ...expense of NADPH. Exogenously administered BH4 can be metabolized by the body, similar to vitamins. At present, synthetic BH4 is used as an orphan drug for patients with inherited diseases requiring BH4 supplementation. BH4 supplementation has also drawn attention as a means of treating certain cardiovascular symptoms, however, its application in human patients remains limited. Here, we tracked biopterin (BP) distribution in blood, bile, urine, liver, kidney and brain after BH4 administration (5 mg/kg rat, i.v.) with or without prior treatment with probenecid, a potent inhibitor of uptake transporters particularly including organic anion transporter families such as OTA1 and OAT3. The rapid excretion of BP in urine was driven by elevated blood concentrations and its elimination reached about 90% within 120 min. In the very early period, BP was taken up by the liver and kidney and gradually released back to the blood. BH4 administration caused a considerable decrease in the BH4% in blood BP as an inevitable compensatory process. Probenecid treatment slowed down the decrease in blood BP and simultaneously inhibited its initial rapid excretion in the kidney. At the same time, the BH4% was further lowered, suggesting that the probenecid-sensitive BP uptake played a crucial role in BH2 scavenging in vivo. This suggested that the overproduced BH2 was taken up by organs by means of the probenecid-sensitive process, and was then scavenged by counter-conversion to BH4 via the BH4 salvage pathway. Taken together, BH4 administration was effective at raising BP levels in organs over the course of hours but with extremely low efficiency. Since a high BH2 relative to BH4 causes NOS dysfunction, the lowering of the BH4% must be avoided in practice, otherwise the desired effect of the supplementation in ameliorating NOS dysfunction would be spoiled.
Aims: The crew members of an endurance motor sports rally team developed acute high-altitude illness (AHAI)during the first year of a route that included driving at high altitude for three days. ...Therefore, a health care manager (HCM) was assigned to the team to help prevent AHAI during the second and third years. We evaluated theeffect of the HCM on the prevalence of AHAI.Methods: The numbers of individuals who presented at the official medical tent of the rally were recorded. Reports from another team without an HCM served as the control.Results: The rates of visits were essentially the same between the two teams (5 20.8% of 24 vs. 3 20.0% of15) respectively during 2016. These values were reduced in the team with the HCM during 2017 and 2018, butdid not change in the control team (1 4% of 25 and 0 0% of 25, respectively vs. 3 18.8% of 16; P = 0.0128Fisher exact test).Conclusions: An HCM with knowledge of AHAI was effective in preventing the development of AHAI amongendurance rally crews.
Tetrahydrobiopterin (BH
4
) is a common coenzyme of phenylalanine-, tyrosine-, and tryptophan hydroxylases, alkylglycerol monooxygenase, and NO synthases (NOS). Synthetic BH
4
is used medicinally for ...BH
4
-responsive phenylketonuria and inherited BH
4
deficiency. BH
4
supplementation has also drawn attention as a therapy for various NOS-related cardio-vascular diseases, but its use has met with limited success in decreasing BH
2
, the oxidized form of BH
4
. An increase in the BH
2
/BH
4
ratio leads to NOS dysfunction. Previous studies revealed that BH
4
supplementation caused a rapid urinary loss of BH
4
accompanied by an increase in the blood BH
2
/BH
4
ratio and an involvement of probenecid-sensitive but unknown transporters was strongly suggested in these processes. Here we show that OAT1 and OAT3 enabled cells to take up BP (BH
4
and/or BH
2
) in a probenecid-sensitive manner using rat kidney slices and transporter-expressing cell systems, LLC-PK1 cells and
Xenopus
oocytes. Both OAT1 and OAT3 preferred BH
2
and sepiapterin as their substrate roughly 5- to 10-fold more than BH
4
. Administration of probenecid acutely reduced the urinary exclusion of endogenous BP accompanied by a rise in blood BP in vivo. These results indicated that OAT1 and OAT3 played crucial roles: (1) in determining baseline levels of blood BP by excluding endogenous BP through the urine, (2) in the rapid distribution to organs of exogenous BH
4
and the exclusion to urine of a BH
4
excess, particularly when BH
4
was administered, and (3) in scavenging blood BH
2
by cellular uptake as the gateway to the salvage pathway of BH
4
, which reduces BH
2
back to BH
4
.
Hematopoiesis in the bone marrow (BM) and spleen is controlled by stromal cells. Inflammation promotes myelopoiesis and simultaneously suppresses B lymphopoiesis. However, the role of the reciprocal ...regulation of myelopoiesis and B lymphopoiesis by stromal cells during inflammation is not fully understood. We investigated inflammation-induced alteration of hematopoietic regulation in lipopolysaccharide (LPS)-treated mice. C57BL/6 female mice were intravenously injected with a single, 5-µg dose of LPS, which induced a rapid decrease in the number of granulocyte-macrophage progenitors (colony-forming unit granulocyte-macrophage; CFU-GM) and B cell progenitors (CFU-preB) in BM. The CFU-GM count rapidly recovered, whereas the recovery of CFU-preB was delayed. LPS induced a marked increase in the number of CFU-GM but not in the number of CFU-preB in spleen. After LPS treatment, gene expression levels of positive regulators of myelopoiesis such as granulocyte colony-stimulating factor (G-CSF), interleukin (IL)-6, and granulocyte-macrophage colony-stimulating factor (GM-CSF) in BM and spleen were markedly upregulated whereas levels of positive regulators for B lymphopoiesis such as stromal cell-derived factor (SDF)-1, stem cell factor (SCF), and IL-7 remained unchanged. Meanwhile, the negative regulator of B lymphopoiesis tumor necrosis factor (TNF)-α was markedly up-regulated. The number of CFU-GM in S-phase in BM increased after LPS treatment, whereas the number of CFU-preB in S-phase decreased. These results suggest that LPS-activated stromal cells induce positive-dominant regulation of myelopoiesis and negative-dominant regulation of B lymphopoiesis, which facilitates emergency myelopoiesis during inflammation by suppressing B lymphopoiesis, thereby contributing to the host defense against infection.
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