Expression of hepcidin, the hepatic hormone controlling iron homeostasis, is regulated by bone morphogenetic protein (BMP) signaling. We sought to identify which BMP type II receptor expressed in ...hepatocytes, ActR2a or BMPR2, is responsible for regulating hepcidin gene expression. We studied Bmpr2 heterozygous mice (Bmpr2+/−), mice with hepatocyte-specific deficiency of BMPR2, mice with global deficiency of ActR2a, and mice in which hepatocytes lacked both BMPR2 and ActR2a. Hepatic hepcidin messenger RNA (mRNA) levels, serum hepcidin and iron levels, and tissue iron levels did not differ in wild-type mice, Bmpr2+/− mice, and mice in which either BMPR2 or ActR2a was deficient. Deficiency of both BMP type II receptors markedly reduced hepatic hepcidin gene expression and serum hepcidin levels leading to severe iron overload. Iron injection increased hepatic hepcidin mRNA levels in mice deficient in either BMPR2 or ActR2a, but not in mice deficient in both BMP type II receptors. In addition, in mouse and human primary hepatocytes, deficiency of both BMPR2 and ActR2a profoundly decreased basal and BMP6-induced hepcidin gene expression. These results suggest that BMP type II receptors, BMPR2 and ActR2a, have redundant roles in the regulation of hepatic hepcidin gene expression and iron metabolism.
•Presence of either ActR2a or BMPR2 in hepatocytes is sufficient to maintain hepatic hepcidin gene expression and iron metabolism.•Deficiency of both BMP type II receptors in hepatocytes induces iron overload.
The bone morphogenetic protein (BMP) type II receptor (BMPR2) has a long cytoplasmic tail domain whose function is incompletely elucidated. Mutations in the tail domain of BMPR2 are found in familial ...cases of pulmonary arterial hypertension. To investigate the role of the tail domain of BMPR2 in BMP signaling, we generated a mouse carrying a Bmpr2 allele encoding a non-sense mediated decay-resistant mutant receptor lacking the tail domain of Bmpr2. We found that homozygous mutant mice died during gastrulation, whereas heterozygous mice grew normally without developing pulmonary arterial hypertension. Using pulmonary artery smooth muscle cells (PaSMC) from heterozygous mice, we determined that the mutant receptor was expressed and retained its ability to transduce BMP signaling. Heterozygous PaSMCs exhibited a BMP7‑specific gain of function, which was transduced via the mutant receptor. Using siRNA knockdown and cells from conditional knockout mice to selectively deplete BMP receptors, we observed that the tail domain of Bmpr2 inhibits Alk2‑mediated BMP7 signaling. These findings suggest that the tail domain of Bmpr2 is essential for normal embryogenesis and inhibits Alk2‑mediated BMP7 signaling in PaSMCs.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Bone morphogenetic protein (BMP) signaling contributes to the development of cardiac hypertrophy. However, the identity of the BMP type I receptor involved in cardiac hypertrophy and the underlying ...molecular mechanisms are poorly understood. By using quantitative PCR and immunoblotting, we demonstrated that BMP signaling increased during phenylephrine-induced hypertrophy in cultured neonatal rat cardiomyocytes (NRCs), as evidenced by increased phosphorylation of Smads 1 and 5 and induction of Id1 gene expression. Inhibition of BMP signaling with LDN193189 or noggin, and silencing of Smad 1 or 4 using small interfering RNA diminished the ability of phenylephrine to induce hypertrophy in NRCs. Conversely, activation of BMP signaling with BMP2 or BMP4 induced hypertrophy in NRCs. Luciferase reporter assay further showed that BMP2 or BMP4 treatment of NRCs repressed atrogin-1 gene expression concomitant with an increase in calcineurin protein levels and enhanced activity of nuclear factor of activated T cells, providing a mechanism by which BMP signaling contributes to cardiac hypertrophy. In a model of cardiac hypertrophy, C57BL/6 mice treated with angiotensin II (A2) had increased BMP signaling in the left ventricle. Treatment with LDN193189 attenuated A2-induced cardiac hypertrophy and collagen deposition in left ventricles. Cardiomyocyte-specific deletion of BMP type I receptor ALK2 (activin-like kinase 2), but not ALK1 or ALK3, inhibited BMP signaling and mitigated A2-induced cardiac hypertrophy and left ventricular fibrosis in mice. The results suggest that BMP signaling upregulates the calcineurin/nuclear factor of activated T cell pathway via BMP type I receptor ALK2, contributing to cardiac hypertrophy and fibrosis.
Resumen: Los trabajos sobre el jesuita chileno Manuel Lacunza han tratado aspectos biográficos, teológicos y editoriales, los cuales se han centrado en los temas escatológicos de La Venida del Mesías ...en Gloria y Majestad y en las ideas religiosas de su autor. Además, se suma a esto que los investigadores han abordado las diferentes ediciones de la obra y la recepción que tuvo esta tanto en Europa como en América, dejando un vacío en lo referente a las ideas científicas presentes en ella, las cuales están contenidas en el tercer tomo. Por lo tanto, en este artículo abordaremos las referencias astronómicas incluidas en el tratado teológico de Lacunza, la cual fue la ciencia que el jesuita chileno sintió afición durante su estadía en Chile como en Italia, desde la perspectiva de la historia de la ciencia. Para este cometido se analizará la ciencia y su vinculación con la religión en el pensamiento del teólogo jesuita proponiendo que no hubo una contradicción o conflicto en ambas vías.
Blood transfusion is a lifesaving treatment for hemorrhagic shock. During storage, red blood cells (RBC) undergo progressive deleterious functional, biochemical and structural alterations, which are ...collectively termed the “storage lesion”. The association between transfusion of blood stored for more than 14 days and adverse clinical outcomes (increased infection, multi-organ failure and mortality) is controversial. Studying mice with hemorrhagic shock, we recently found that resuscitation with blood stored for prolonged periods (SRBC) was associated with worse outcomes than was resuscitation with fresh blood (FRBC). The mechanisms responsible for the adverse effects associated with transfusion of SRBC are incompletely characterized. However, it is known that transfusion of SRBC increases plasma levels of hemoglobin (Hb), which can scavenge vascular nitric oxide (NO). Intravenous infusion of a solution containing cell-free Hb induces systemic hypertension in wild-type (WT) mice, but not in mice that are congenitally deficient in NO synthase 3 (NOS3-/-). In the present study, we sought to determine if NOS3-/- mice are protected from the adverse effects associated with resuscitation of hemorrhagic shock with SRBC.
Leukoreduced, packed RBC from WT C57BL6 mice were stored with 14% CPDA-1 anticoagulant at 4°C for either ≤24 h (FRBC) or 2 weeks (SRBC). Mice, of each genotype that were not subjected to hemorrhagic shock or resuscitation, served as control groups. Anesthetized WT mice and NOS3-/- mice (on a C57BL6 background) were bled to a mean arterial pressure (MAP) of 40 mmHg over 10 min. After 90 min of hemorrhagic shock, mice were resuscitated with FRBC or SRBC. Survival rates for up to 7 days were determined. In addition, blood and tissue samples were collected at 4 h after resuscitation to measure plasma markers of liver and kidney injury, plasma Hb and interleukin 6 (IL-6) levels, tissue IL-6 mRNA levels, and pulmonary myeloperoxidase activity and mRNA levels. All data are expressed as mean±SD.
Baseline MAP under anesthesia was higher in NOS3-/- mice than in WT mice (111±4 vs 83±5 mmHg; P<0.01). After hemorrhagic shock and resuscitation with either FRBC or SRBC, MAP returned to the respective baseline values in each genotype. Survival rates at one week did not differ between WT or NOS3-/- mice resuscitated with either FRBC or SRBC. In both genotypes, after hemorrhagic shock, resuscitation with FRBC elevated plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities similarly as compared to a control group (P<0.01). Plasma levels of ALT and AST were greater after resuscitation with SRBC than after resuscitation with FRBC in both genotypes at 4 h (P<0.01). Resuscitation with SRBC, but not FRBC, increased plasma blood urea nitrogen and creatinine levels similarly in WT and NOS3-/- mice. Plasma Hb levels were greater in mice resuscitated with SRBC than in those resuscitated with FRBC in both genotypes at 4 h after resuscitation (WT: 202±79 vs 5±2 µM, respectively; NOS3-/-: 240±51 vs 14±3 µM, respectively; P<0.001 for both). At 4 h after resuscitation, plasma IL-6 levels were greater in mice treated with SRBC than in mice treated with FRBC in both genotypes (WT: 0.8±0.1 vs 0.5±0.1 ng/ml, respectively; NOS3-/-: 0.8±0.1 vs 0.4±0.1 ng/ml; P<0.01 for both). In both WT and NOS3-/- mice, IL-6 mRNA levels were greater in the liver, kidney, and spleen after resuscitation with SRBC than after resuscitation with FRBC (P<0.01, all values differ SRBC vs FRBC for both genotypes). Pulmonary myeloperoxidase activity and mRNA levels were greater in both genotypes after resuscitation with SRBC than mice resuscitated with FRBC (P<0.01, all values differ SRBC vs FRBC for both genotypes).
Survival rate after hemorrhagic shock did not differ in WT and NOS3-/- mice resuscitated with either FRBC or SRBC. Resuscitation with SRBC induced greater tissue injury and a more marked inflammatory response than did resuscitation with FRBC, but there was no difference between the genotypes. Our data suggest that mice with NOS3 deficiency are not protected from the adverse effects associated with resuscitation of hemorrhagic shock with SRBC. These findings suggest that the adverse effects of transfusing blood stored for prolonged periods in mice with hemorrhagic shock are not exclusively attributable to scavenging of NOS3-generated NO by increased plasma Hb levels.
No relevant conflicts of interest to declare.
Abstract 992
Hepcidin regulates iron metabolism by reducing duodenal iron absorption and iron release from macrophages and hepatocytes. In inflammatory states, including infection, neoplasia, and ...heart failure, cytokines induce hepcidin synthesis leading to the development of anemia of inflammation. The regulation of hepcidin gene expression by bone morphogenetic proteins (BMPs), members of the TGFβ family of growth factors, has been extensively investigated. In contrast, less is known about the regulation of hepcidin gene expression by other stimuli, including TGFβ itself. Although TGFβ expression is increased in inflammatory states, the role of TGFβ in the induction of hepcidin gene expression is controversial. To further elucidate the role TGFβ in iron metabolism, we investigated the regulation of hepcidin gene expression in the hepatoma cell line, HepG2.
HepG2 cells were incubated with TGFβ (0.1, 0.5, 1, 2.5, and 5 ng/ml) for varying durations. RNA was extracted for measurement of levels of mRNAs encoding hepcidin, PAI-1 (a TGFβ-target gene), and Id-1 (a BMP-target gene). Cellular proteins were extracted to measure levels of phosphorylated TGFβ-responsive SMADs (using antibodies directed against phosphorylated SMAD2 or SMAD3) and levels of phosphorylated BMP-responsive SMADs (using antibodies directed to phosphorylated SMADs 1 and 5, SMAD1/5). The mechanisms by which TGFβ regulates hepcidin were investigated by pretreating cells with cycloheximide, an inhibitor of protein synthesis (50 μg/mL); Noggin (250 ng/mL) or LDN-193189 (100 nM), inhibitors of BMP signaling; or SB-431542 (5 μM), an inhibitor of the TGFβ type 1 receptor, Alk5. In additional experiments, HepG2 cells were transfected with an siRNA directed against Alk5, 72 hours before exposure to TGFβ.
In HepG2 cells, TGFβ induced hepcidin gene expression in a time- and dose-dependent manner: hepcidin mRNA levels were maximal at 2 hours after stimulation with TGFβ (1 ng/ml) and declined thereafter. Incubation of HepG2 cells increased PAI-1 and Id-1 mRNA levels, although increased PAI-1 mRNA levels persisted for at least 8 hours whereas Id-1 mRNA levels peaked at 2 hours. Cycloheximide did not block the ability of TGFβ to induce expression of genes encoding hepcidin, PAI-1, or Id-1. TGFβ induced phosphorylation of SMADs 2 and 3, as well as SMAD1/5. Pretreatment of HepG2 cells with LDN-193189 (at concentrations that inhibit all four BMP type I receptors, as well as Alk1 which is a target of both BMPs and TGFβ) did not block the ability of TGFβ to induce hepcidin or Id-1 gene expression or phosphorylation of SMADs 2, 3, or 1/5. Pretreatment with Noggin gave similar results. Inhibition of Alk5 with SB-421542 blocked the ability of TGFβ to induce expression of genes encoding hepcidin, PAI-1, and Id-1, as well as phosphorylation of SMADs 2, 3, or 1/5. TGFβ-stimulated hepcidin gene expression was inhibited by siRNA-mediated knockdown of Alk5.
In HepG2 cells, TGFβ induces hepcidin gene expression via a mechanism which requires Alk5. Although, in addition to phosphorylation of SMADs 2 and 3, TGFβ induces phosphorylation of BMP-responsive SMADs, the failure of cycloheximide to inhibit the induction of hepcidin gene expression by TGFβ suggests that synthesis of BMPs is not required. Moreover, the inability of LDN-193189 to inhibit TGFβ-stimulated hepcidin gene expression suggests against a role for activation of Alk1 by TGFβ. Taken together our findings suggest that TGFβ stimulates hepcidin gene expression via a mechanism that requires Alk5 and may be mediated by signaling either via SMADs 2 and 3 or SMAD1/5. Targeting the regulation of hepcidin gene expression by TGFβ may offer a novel therapeutic approach to the anemia of inflammation.
No relevant conflicts of interest to declare.
Se sostiene tradicionalmente que la física es la ciencia que estudia los estados de la materia y que procura explicar de manera racional los fenómenos naturales mediante el uso de complejas ...ecuaciones matemáticas. No obstante, consideramos que esta ciencia puede tener otros usos más allá del mero y objetivo estudio de la realidad natural. Este fue el caso de la expedición científica española, al mando del navegante y científico italiano Alejandro Malaspina, la cual recorrió los dominios de ultramar de la Corona española entre los años 1789 a 1794 y efectuó trabajos científicos en diversas disciplinas, siendo la física una de ellas. En este sentido, la física en la comisión hispana tuvo carices de tipo ideológico, práctico y teórico-experimental. El primero fue dado por el vínculo de esta ciencia con aspectos políticos, civilizatorios y de prosperidad nacional, los que estaban en directa concordación con el ethos de la Ilustración europea. El segundo, en tanto, estuvo en directa relación con los trabajos propiamente científicos; vale decir que se analiza la física en concordancia con el uso de instrumentos científicos, la medición y cuantificación de fenómenos de la naturaleza y el empleo de una metodología de acuerdo con la ciencia del siglo xviii. Por último, el aspectoteórico-experimental estaba condicionado por las explicaciones de tipo físico y teórico que se dieron a ciertos elementos de la naturaleza y por la elaboración de experimentos y experiencias sobre los estados de la materia, mientras transcurrió el viaje por el Océano Atlántico y el Océano Pacífico.
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