RESUMEN
BACKGROUND: Adverse environmental exposure during the prenatal period can lead to diseases in the offspring, including hypertension. Whether or not the hypertensive phenotype can be transgenerationally transmitted is not known. METHODS: Pregnant Sprague Dawley rats were intraperitoneally injected with lipopolysaccharide (LPS) on gestation days 6, 8, 10, and 12 to generate the prenatal LPS exposure model. Blood pressure was monitored by both telemetry and tail-cuff method. RNA sequencing was performed to analyze transcriptome alteration in the kidney of the third generation. Tempol and spironolactone were used to test the potential preventative and therapeutic effect of targeting reactive oxygen species and mineralocorticoid receptor signaling, respectively. Molecular biological experiments were performed to illustrate the mechanism of epigenetic and transcription regulation. RESULTS: Prenatal LPS exposure can impair the ability to excrete a salt load and induce hypertension from the first to the third generations, with the fourth and fifth generations, inducing salt-sensitive hypertension. Compared with control pups, the transcriptome in the kidney of the hypertensive third-generation prenatal LPS-exposed offspring have upregulation of the Ras-related C3 botulinum toxin substrate 1 (Rac1) gene and activation of mineralocorticoid receptor signaling. Furthermore, we found that LPS exposure during pregnancy triggered oxidative stress that upregulated KDM3B (histone lysine demethylase 3B) in the oocytes of first-generation female rats, leading to an inheritable low level of H3K9me2 (histone H3 lysine 9 dimethylation), resulting in the transgenerational upregulation of Rac1. Based on these findings, we treated the LPS-exposed pregnant rats with the reactive oxygen species scavenger, tempol, which successfully prevented hypertension in the first-generation offspring and the transgenerational inheritance of hypertension. CONCLUSIONS: These findings show that adverse prenatal exposure induces transgenerational hypertension through an epigenetic-regulated mechanism and identify potentially preventive and therapeutic strategies for hypertension.
Asunto(s)
Hipertensión , Efectos Tardíos de la Exposición Prenatal , Animales , Óxidos N-Cíclicos , Femenino , Histona Demetilasas , Histonas , Hipertensión/inducido químicamente , Hipertensión/genética , Histona Demetilasas con Dominio de Jumonji , Lipopolisacáridos/toxicidad , Lisina , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Efectos Tardíos de la Exposición Prenatal/etiología , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno , Receptores de Mineralocorticoides/genética , Marcadores de Spin , Espironolactona , Proteína de Unión al GTP rac1/genéticaRESUMEN
The adverse environment in early life can modulate adult phenotype, including blood pressure. Our previous study shows, in a rat streptozotocin (STZ)-induced maternal diabetes model, fetal exposure to maternal diabetes is characterized by established hypertension in the offspring. However, the exact mechanisms are not known. Our present study found, as compared with male control mother offspring (CMO), male diabetic mother offspring (DMO) had higher blood pressure with arterial dysfunction, i.e., decreased acetylcholine (Ach)-induced vasodilation. But there is no difference in blood pressure between female CMO and DMO. The decreased Ach-induced vasodilation was related to decreased nitric oxide (NO) production in the endothelium, not NO sensitivity in vascular smooth muscle because sodium nitroprusside (SNP)-mediated vasodilation was preserved; there was decreased NO production and lower eNOS phosphorylation in male DMO. The reactive oxygen species (ROS) level was increased in male DMO than CMO; normalized ROS levels with tempol increased NO production, normalized Ach-mediated vasodilation, and lowered blood pressure in male DMO rats. It indicates that diabetic programming hypertension is related to arterial dysfunction; normalizing ROS might be a potential strategy for the prevention of hypertension in the offspring.
Asunto(s)
Diabetes Mellitus Experimental/complicaciones , Diabetes Gestacional , Endotelio Vascular/fisiopatología , Hipertensión/etiología , Arteria Mesentérica Superior/fisiopatología , Efectos Tardíos de la Exposición Prenatal , Factores de Edad , Animales , Presión Arterial , Glucemia/metabolismo , GMP Cíclico/metabolismo , Diabetes Mellitus Experimental/sangre , Diabetes Mellitus Experimental/fisiopatología , Diabetes Gestacional/sangre , Diabetes Gestacional/fisiopatología , Endotelio Vascular/metabolismo , Femenino , Hipertensión/metabolismo , Hipertensión/fisiopatología , Masculino , Arteria Mesentérica Superior/metabolismo , Óxido Nítrico/metabolismo , Estrés Oxidativo , Embarazo , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismo , Factores Sexuales , VasodilataciónRESUMEN
Maternal exposure to fine particulate matter (PM2.5) causes hypertension in offspring. However, paternal contribution of PM2.5 exposure to hypertension in offspring remains unknown. In the present study, male Sprague-Dawley rats were treated with PM2.5 suspension (10 mg/ml) for 12 weeks and/or fed with tap water containing an antioxidant tempol (1 mM/L) for 16 weeks. The blood pressure, 24 h-urine volume and sodium excretion were determined in male offspring. The offspring were also administrated with losartan (20 mg/kg/d) for 4 weeks. The expressions of angiotensin II type 1 receptor (AT1R) and G-protein-coupled receptor kinase type 4 (GRK4) were determined by qRT-PCR and immunoblotting. We found that long-term PM2.5 exposure to paternal rats caused hypertension and impaired urine volume and sodium excretion in male offspring. Both the mRNA and protein expression of GRK4 and its downstream target AT1R were increased in offspring of PM2.5-exposed paternal rats, which was reflected in its function because treatment with losartan, an AT1R antagonist, decreased the blood pressure and increased urine volume and sodium excretion. In addition, the oxidative stress level was increased in PM2.5-treated paternal rats. Administration with tempol in paternal rats restored the increased blood pressure and decreased urine volume and sodium excretion in the offspring of PM2.5-exposed paternal rats. Treatment with tempol in paternal rats also reversed the increased expressions of AT1R and GRK4 in the kidney of their offspring. We suggest that paternal PM2.5 exposure causes hypertension in offspring. The mechanism may be involved that paternal PM2.5 exposure-associated oxidative stress induces the elevated renal GRK4 level, leading to the enhanced AT1R expression and its-mediated sodium retention, consequently causes hypertension in male offspring.
Asunto(s)
Hipertensión/etiología , Material Particulado/efectos adversos , Exposición Paterna/efectos adversos , Efectos Tardíos de la Exposición Prenatal , Receptor de Angiotensina Tipo 1/metabolismo , Animales , Presión Sanguínea , Femenino , Quinasa 4 del Receptor Acoplado a Proteína-G/metabolismo , Hipertensión/metabolismo , Masculino , Estrés Oxidativo , Embarazo , Ratas Sprague-Dawley , Sodio/orinaRESUMEN
The antiviral cGMP-AMP (cGAMP)-stimulator of interferon genes (STING) pathway is well characterized in mammalian cells. However, whether this pathway also plays a role in insect antiviral immunity is unknown. In this study, we found that cGAMP is produced in silkworm (Bombyx mori) cells infected with nucleopolyhedrovirus (NPV). In searches for STING-related sequences, we identified BmSTING, a potential cGAMP sensor in B. mori We observed that BmSTING overexpression effectively inhibits NPV replication in silkworm larvae, whereas dsRNA-mediated BmSTING knockdown resulted in higher viral load. Cleavage and nuclear translocation of BmRelish, a NF-κB-related transcription factor, was also observed when BmSTING was overexpressed and was enhanced by cGAMP stimulation or viral infection of B. mori larvae. Moreover, we identified a caspase-8-like protein (BmCasp8L) as a BmSTING-interacting molecule and as a suppressor of BmSTING-mediated BmRelish activation. Interestingly, cGAMP stimulation decreased BmCasp8L binding to BmSTING and increased BmRelish activity. Of note, an interaction between death-related ced-3/Nedd2-like caspase (BmDredd) and BmSTING promoted BmRelish cleavage for efficient antiviral signaling and protection of insect cells from viral infection. Our findings have uncovered BmSTING as a critical mediator of antiviral immunity in the model insect B. mori and have identified several BmSTING-interacting proteins that control antiviral defenses.
Asunto(s)
Bombyx/inmunología , Bombyx/virología , Caspasas Iniciadoras/inmunología , Proteínas de Insectos/inmunología , FN-kappa B/inmunología , Nucleopoliedrovirus/inmunología , Nucleótidos Cíclicos/inmunología , Animales , Inmunidad Innata , Transducción de SeñalRESUMEN
AIMS: Exposure to maternal diabetes is associated with increased prevalence of hypertension in the offspring. The mechanisms underlying the prenatal programming of hypertension remain unclear. Because endoplasmic reticulum (ER) stress plays a key role in vascular endothelial dysfunction in hypertension, we investigated whether aberrant ER stress causes endothelial dysfunction and high blood pressure in the offspring of dams with diabetes. METHODS AND RESULTS: Pregnant Sprague-Dawley rats were intraperitoneally injected with streptozotocin (35 mg/kg) or citrate buffer at Day 0 of gestation. Compared with control mother offspring (CMO), the diabetic mother offspring (DMO) had higher blood pressure and impaired endothelium-dependent relaxation in mesenteric arteries, accompanied by decreased AMPK phosphorylation and PPARδ expression, increased ER stress markers, and reactive oxygen species (ROS) levels. The inhibition of ER stress reversed these aberrant changes in DMO. Ex vivo treatment of mesenteric arteries with an AMPK agonist (A769662) or a PPARδ agonist (GW1516) improved the impaired EDR in DMO and reversed the tunicamycin-induced ER stress, ROS production, and EDR impairment in mesenteric arteries from CMO. The effects of A769662 were abolished by co-treatment with GSK0660 (PPARδ antagonist), whereas the effects of GW1516 were unaffected by Compound C (AMPK inhibitor). CONCLUSION: These results suggest an abnormal foetal programming of vascular endothelial function in offspring of rats with maternal diabetes that is associated with increased ER stress, which can be ascribed to down-regulation of AMPK/PPARδ signalling cascade.
Asunto(s)
Diabetes Mellitus , Hipertensión , PPAR delta , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Diabetes Mellitus/metabolismo , Regulación hacia Abajo , Estrés del Retículo Endoplásmico , Endotelio Vascular/metabolismo , Femenino , PPAR delta/genética , PPAR delta/metabolismo , PPAR delta/farmacología , Embarazo , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismoRESUMEN
AIMS: The proliferation of VSMCs is the pathologic basis for intimal hyperplasia after angioplasty in diabetic patients. Translocator protein (TSPO), located in the outer mitochondrial membrane, has been found to regulate redox intermediate components in cell dysfunction. We hypothesized that TSPO may regulate VSMC proliferation and migration, and be involved in the intimal hyperplasia after angioplasty in diabetes. MATERIALS AND METHODS: Cell proliferation was measured by cell counting and MTT assays. Cell migration was measured by Transwell® and scratch-wound assays. TSPO expression in arteries of rats and high glucose-treated A10 cells were detected by immunoblotting and immunofluorescence staining. Neointimal formation of carotid artery was induced by balloon injury in type 2 diabetic rat. KEY FINDINGS: TSPO expression was increased in the arterial samples from diabetic rats and A10 cells treated with high glucose. Down-regulation of TSPO expression by siRNA decreased the high-glucose-induced VSMC proliferation and migration in A10 cells. This phenomenon could be simulated by using TSPO ligands, PK 11195 and Ro5-4864. cGMP/PKG signals were involved in the TSPO ligand action, since in the presence of cGMP or PKG inhibitor ODQ or KT5823 respectively, the effect of PK 11195 on VSMC proliferation was blocked. Furthermore, PK 11195 significantly inhibited neointimal formation by the inhibition of VSMC proliferation. SIGNIFICANCE: This study suggests that TSPO inhibition suppresses the proliferation and migration of VSMCs induced by hyperglycemia, consequently, preventing atherosclerosis and restenosis after angioplasty in diabetic conditions. TSPO may be a potential therapeutic target to reduce arterial remodeling induced by angioplasty in diabetes.
Asunto(s)
Proteínas Portadoras/metabolismo , Hiperplasia/metabolismo , Receptores de GABA-A/metabolismo , Animales , Benzodiazepinonas/farmacología , Arterias Carótidas/patología , Proteínas Portadoras/fisiología , Movimiento Celular , Proliferación Celular , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Modelos Animales de Enfermedad , Hiperplasia/prevención & control , Isoquinolinas/farmacología , Ligandos , Masculino , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/patología , Neointima/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores de GABA-A/fisiologíaRESUMEN
The relevance of protein tyrosine phosphatase (PTP) to host-pathogen interaction is highlighted in mammalian studies, whereas less is known in insects. Here we presented the categorization of the PTP complement of silkworm and characterized their homologous relationship with human and fruit fly PTPs. Among the 36 PTP genes, ptp-h, which was proposed to be the origin of baculovirus ptp belongs to atypical VH1-like dual-specific PTP subset and encodes a catalytic active protein. The maximum expression level of Bmptp-h was at 5th instar and in fat body. Bombyx mori nucleopolyhedrovirus (BmNPV) infection potently induced its expression in silkworm larvae and in BmE cells. Knock-down of Bmptp-h by RNA interference significantly inhibited viral replication, and over-expression enhanced viral replication as determined by viral DNA abundance and BmNPV-GFP positive cells. These results suggest that BmPTP-h might be one of the host factors that is beneficial to baculovirus infection by promoting viral replication.
Asunto(s)
Baculoviridae/fisiología , Bombyx/inmunología , Infecciones por Virus ADN/metabolismo , Proteínas de Insectos/metabolismo , Proteínas Tirosina Fosfatasas/metabolismo , Animales , Células Cultivadas , Drosophila , Perfilación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Especificidad del Huésped , Interacciones Huésped-Patógeno , Humanos , Proteínas de Insectos/genética , Proteínas Tirosina Fosfatasas/genética , Homología de Secuencia de Aminoácido , Replicación Viral/genéticaRESUMEN
Intestinal immune response is a front line of host defense. The host factors that participate in intestinal immunity response remain largely unknown. We recently reported that Translationally Controlled Tumor Protein (BmTCTP) was obtained by constructing a phage display cDNA library of the silkworm midgut and carrying out high throughput screening of pathogen binding molecules. To further address the function of BmTCTP in silkworm intestinal immunity, transgenic RNAi silkworms were constructed by microinjection piggBac plasmid to Dazao embryos. The antimicrobial capacity of transgenic silkworm decreased since the expression of gut antimicrobial peptide from transgenic silkworm was not sufficiently induced during oral microbial challenge. Moreover, dynamic ERK phosphorylation from transgenic silkworm midgut was disrupted. Taken together, the innate immunity of intestinal was suppressed through disruption of dynamic ERK phosphorylation after oral microbial infection as a result of RNAi-mediated knockdown of midgut TCTP in transgenic silkworm.
Asunto(s)
Animales Modificados Genéticamente/genética , Animales Modificados Genéticamente/inmunología , Bombyx/genética , Bombyx/inmunología , Proteínas de Insectos/genética , Intestinos/inmunología , Interferencia de ARN/inmunología , Animales , Silenciador del Gen/inmunología , Inmunidad Innata , Proteínas de Insectos/inmunología , Larva/genética , Larva/inmunología , Sistema de Señalización de MAP Quinasas/genética , Sistema de Señalización de MAP Quinasas/inmunología , Plásmidos/genéticaRESUMEN
Paralytic peptide (PP) activates innate immunity of silkworm Bombyx mori, inducing production of anti-microbial peptides (AMPs) and phagocytosis-related proteins; however the signal pathways of PP-dependent immune responses are not clear. In present study, we characterized BmE cells as a PP-responsive cell line by examining the expression of AMP genes and activation of p38 mitogen-activated protein kinase (p38 MAPK) under PP stimulation, and we also found PP directly binds to BmE cell membrane. Then we found that PP-dependent expression of AMP genes is suppressed by tyrosine kinase inhibitor (genistein) both in BmE cells and in fat body of silkworm larvae. Moreover, the specific tyrosine kinase epidermal growth factor receptor (EGFR) inhibitor (AG1478) attenuates PP-induced expression of AMP genes in BmE cells and fat body of silkworm and RNA interference (RNAi) to BmEGFR also suppresses PP-induced expression of AMP genes. Furthermore, the PP-induced p38 MAPK phosphorylation is inhibited by AG1478. Our results suggest that BmE cells can be used as a cell model to investigate the signal pathway of PP-dependent humoral immune response and receptor tyrosine kinase EGFR/p38 MAPK pathway is involved in the production of AMPs induced by PP.
Asunto(s)
Bombyx/inmunología , Receptores ErbB/inmunología , Inmunidad Humoral/efectos de los fármacos , Proteínas de Insectos/inmunología , Neuropéptidos/farmacología , Animales , Larva/inmunologíaRESUMEN
Insect gut immunity is the first line of defense against oral infection. Although a few immune-related molecules in insect intestine has been identified by genomics or proteomics approach with comparison to well-studied tissues, such as hemolymph or fat body, our knowledge about the molecular mechanism underlying the gut immunity which would involve a variety of unidentified molecules is still limited. To uncover additional molecules that might take part in pathogen recognition, signal transduction or immune regulation in insect intestine, a T7 phage display cDNA library of the silkworm midgut is constructed. By use of different ligands for biopanning, Translationally Controlled Tumor Protein (TCTP) has been selected. BmTCTP is produced in intestinal epithelial cells and released into the gut lumen. The protein level of BmTCTP increases at the early time points during oral microbial infection and declines afterwards. In vitro binding assay confirms its activity as a multi-ligand binding molecule and it can further function as an opsonin that promotes the phagocytosis of microorganisms. Moreover, it can induce the production of anti-microbial peptide via a signaling pathway in which ERK is required and a dynamic tyrosine phosphorylation of certain cytoplasmic membrane protein. Taken together, our results characterize BmTCTP as a dual-functional protein involved in both the cellular and the humoral immune response of the silkworm, Bombyx mori.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Bombyx/inmunología , Bombyx/metabolismo , Inmunidad Celular , Inmunidad Humoral , Proteínas de Insectos/metabolismo , Animales , Péptidos Catiónicos Antimicrobianos/biosíntesis , Fenómenos Fisiológicos Bacterianos , Bombyx/microbiología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Hemocitos/inmunología , Hemocitos/microbiología , Mucosa Intestinal/metabolismo , Ligandos , Fagocitosis/inmunología , Transducción de Señal/inmunología , Proteína Tumoral Controlada Traslacionalmente 1RESUMEN
Malpighian tubules (MTs) are highly specific organs of arthropods (Insecta, Myriapoda and Arachnida) for excretion and osmoregulation. In order to highlight the important genes and pathways involved in multi-functions of MTs, we performed a systematic proteomic analysis of silkworm MTs in the present work. Totally, 1,367 proteins were identified by one-dimensional gel electrophoresis coupled with liquid chromatography-tandem mass spectrometry, and as well as by Trans Proteomic Pipeline (TPP) and Absolute protein expression (APEX) analyses. Forty-one proteins were further identified by two-dimensional gel electrophoresis. Some proteins were revealed to be significantly associated with various metabolic processes, organic solute transport, detoxification and innate immunity. Our results might lay a good foundation for future functional studies of MTs in silkworm and other lepidoptera.