RESUMEN
When cells are stressed, DNA from energy-producing mitochondria can leak out and drive inflammatory immune responses if not cleared. Cells employ a quality control system called autophagy to specifically degrade damaged components. We discovered that mitochondrial transcription factor A (TFAM)-a protein that binds mitochondrial DNA (mtDNA)-helps to eliminate leaked mtDNA by interacting with the autophagy protein LC3 through an autolysosomal pathway (we term this nucleoid-phagy). TFAM contains a molecular zip code called the LC3 interacting region (LIR) motif that enables this binding. Although mutating TFAM's LIR motif did not affect its normal mitochondrial functions, more mtDNA accumulated in the cell cytoplasm, activating inflammatory signalling pathways. Thus, TFAM mediates autophagic removal of leaked mtDNA to restrict inflammation. Identifying this mechanism advances understanding of how cells exploit autophagy machinery to selectively target and degrade inflammatory mtDNA. These findings could inform research on diseases involving mitochondrial damage and inflammation.
Asunto(s)
Autofagia , ADN Mitocondrial , Proteínas de Unión al ADN , Inflamación , Mitocondrias , Proteínas Mitocondriales , Factores de Transcripción , ADN Mitocondrial/genética , ADN Mitocondrial/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ADN/genética , Inflamación/metabolismo , Inflamación/patología , Inflamación/genética , Animales , Humanos , Proteínas Mitocondriales/metabolismo , Proteínas Mitocondriales/genética , Mitocondrias/metabolismo , Mitocondrias/genética , Ratones , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Unión Proteica , Citoplasma/metabolismo , Lisosomas/metabolismo , Transducción de Señal , Células HEK293 , Ratones Endogámicos C57BL , Proteínas del Grupo de Alta MovilidadRESUMEN
ATP9A, a lipid flippase of the class II P4-ATPases, is involved in cellular vesicle trafficking. Its homozygous variants are linked to neurodevelopmental disorders in humans. However, its physiological function, the underlying mechanism as well as its pathophysiological relevance in humans and animals are still largely unknown. Here, we report two independent families in which the nonsense mutations c.433C>T/c.658C>T/c.983G>A (p. Arg145*/p. Arg220*/p. Trp328*) in ATP9A (NM_006045.3) cause autosomal recessive hypotonia, intellectual disability (ID) and attention deficit hyperactivity disorder (ADHD). Atp9a null mice show decreased muscle strength, memory deficits and hyperkinetic movement disorder, recapitulating the symptoms observed in patients. Abnormal neurite morphology and impaired synaptic transmission are found in the primary motor cortex and hippocampus of the Atp9a null mice. ATP9A is also required for maintaining neuronal neurite morphology and the viability of neural cells in vitro. It mainly localizes to endosomes and plays a pivotal role in endosomal recycling pathway by modulating small GTPase RAB5 and RAB11 activation. However, ATP9A pathogenic mutants have aberrant subcellular localization and cause abnormal endosomal recycling. These findings provide strong evidence that ATP9A deficiency leads to neurodevelopmental disorders and synaptic dysfunctions in both humans and mice, and establishes novel regulatory roles for ATP9A in RAB5 and RAB11 activity-dependent endosomal recycling pathway and neurological diseases.
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Trastorno por Déficit de Atención con Hiperactividad , Animales , Humanos , Ratones , Trastorno por Déficit de Atención con Hiperactividad/metabolismo , Endosomas/metabolismo , Transporte de Proteínas , Proteínas de Unión al GTP rab5/genética , Proteínas de Unión al GTP rab5/metabolismoRESUMEN
Objective: To investigate whether postoperative anion gap (AG) is associated with short- and long-term mortality in patients following cardiac surgery. Materials and methods: We conducted a retrospective cohort study of adults who underwent cardiac surgery from the Medical Information Mart for Intensive Care - III database. The generalized additive model (GAM), logistic regression, and Cox regression were performed to assess the correlations between AG levels and in-hospital, 90-day, and 4-year mortality. Linear regression was used to evaluate the associations between AG and length of stay (LOS). Results: Totally, 6,410 subjects were enrolled in this study and classified into tertiles based on the initial AG levels. The GAM indicated a positive association between initial AG and in-hospital mortality after adjusting for potential confounders. Multivariate logistic analysis revealed that the risk of in-hospital mortality was higher among patients in tertile 2 (OR 2.05, 95% CI 1.11-3.76, P = 0.021) and tertile 3 (OR 4.51, 95% CI 2.57-7.91, P < 0.001) compared with those in tertile 1. For 90-day and 4-year mortality, multivariate Cox regression found similar associations between AG tertiles and mortality. The LOS in ICU and hospital also increased as AG tertiles increased. The E-value indicated robustness to unmeasured confounders. Conclusion: This study found a positive association between postoperative AG levels and short- and long-term mortality among patients after cardiac surgery. This relationship warrants further research.
RESUMEN
METHODS: A rat hyperalgesia model was induced using an intraplantar injection of Freund's complete adjuvant (FCA) or an intrathecal injection of IL-6. Mechanical allodynia was evaluated using von Frey filament tests after intrathecal injections of T-5224 (c-Fos/AP-1 inhibitor), minocycline (Mino, a specific microglia inhibitor), L-2-aminoadipic acid (LAA, an astroglial toxin), PKCε inhibitor peptide, APTSTAT3-9R (STAT3 inhibitor), or anti-IL-6 antibody. The c-Fos, GFAP, Iba-1, PKCε, STAT3, pSTAT3Tyr705 and pSTAT3Ser727, and IL-6 expression at the spinal cord level was assessed by Western blot analysis. The interactive effects of PKCε and STAT3 were determined using immunofluorescence staining and immunoprecipitation in vivo and in vitro. Interleukin-6 promoter activity was examined using luciferase assays. RESULTS: T-5224, Mino, and LAA attenuated FCA- or IL-6-mediated inflammatory pain, with a decrease in c-Fos, GFAP, Iba-1, PKCε, and IL-6 expression. PKCε inhibitor peptide and APTSTAT3-9R reversed FCA-induced nociceptive behavior, while decreasing pSTAT3Ser727, IL-6, c-Fos, GFAP, and Iba-1 expression and PKCε and STAT3 coexpression. Interleukin-6 promoter activity increased in the presence of PKCε and STAT3. The interaction with PKCε increased on phosphorylating STAT3 at Ser727 but not at Tyr705. CONCLUSION: STAT3 phosphorylation at Ser 727 and the interaction with PKCε contribute to hyperalgesia via the IL-6-mediated signaling pathway, thus regulating neuron-glia crosstalk during inflammatory pain.
Asunto(s)
Hiperalgesia , Interleucina-6 , Animales , Hiperalgesia/metabolismo , Interleucina-6/metabolismo , Neuroglía/metabolismo , Neuronas/metabolismo , Fosforilación , Proteína Quinasa C-epsilon/metabolismo , Proteína Quinasa C-epsilon/farmacología , Ratas , Médula Espinal/metabolismoRESUMEN
OBJECTIVE: To investigate impact of ulinastatin (UTI) on sigma-1 receptor (σ1R) and binding immunoglobulin protein (BiP) after cerebral ischemia/reperfusion injury. METHODS: The middle cerebral artery occlusion (MCAO) model was used to induce cerebral ischemia/reperfusion injury. Eighty male Sprague Dawley rats were randomly divided into 6 groups: control, MCAO, MCAO+50,000 U/kg UTI, MCAO+100,000 U/kg UTI, MCAO+200,000 U/kg UTI, MCAO+300,000 U/kg UTI. At 24 and 48 hours after MCAO, infarct volume, neurological dysfunction, and grip strength test were measured, and level of σ1R and BiP proteins was further detected using Western blot. Molecular docking assays were carried out to verify interaction between σ1R, BiP, and UTI. The serum concentration of BiP and the binding assay between σ1R, BiP, and UTI were determined using enzyme-linked immunosorbent assay. RESULTS: UTI increased the modified neurological severity score and upregulated σ1R and BiP expression in the cerebral cortex after MCAO. The grip strength of forelimbs increased significantly in the MCAO+200,000 U/kg UTI and MCAO+300,000 U/kg UTI groups compared with the MCAO group, while BiP serum levels remained unchanged. The molecular docking assay indicated putative binding between σ1R, BiP, and UTI. The binding assay also revealed that both σ1R and BiP could be combined with UTI. CONCLUSIONS: UTI displays a neuroprotective effect via upregulation of σ1R and BiP during ischemia/reperfusion injury, suggesting that UTI modulates σ1R and BiP and their interaction may provide a novel insight into potential therapeutic mechanisms for stroke.
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Isquemia Encefálica , Proteínas de Choque Térmico , Fármacos Neuroprotectores , Receptores sigma , Daño por Reperfusión , Animales , Isquemia Encefálica/tratamiento farmacológico , Glicoproteínas , Proteínas de Choque Térmico/metabolismo , Inmunoglobulinas/metabolismo , Inmunoglobulinas/uso terapéutico , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Masculino , Simulación del Acoplamiento Molecular , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Ratas , Ratas Sprague-Dawley , Receptores sigma/metabolismo , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Daño por Reperfusión/prevención & control , Receptor Sigma-1RESUMEN
Hyperglycemia-induced oxidative stress plays important roles in the development of non-alcoholic fatty liver disease (NAFLD), which is a common complication in diabetic patients. The Nrf2-Keap1 pathway is important for cell antioxidant protection, while its role in exogenous antioxidant mediated protection against NAFLD is unclear. We thus, postulated that antioxidant treatment with allopurinol (ALP) may attenuate diabetic liver injury and explored the underlying mechanisms. Control (C) and streptozotocin (STZ)-induced diabetes rats (D) were untreated or treated with ALP for 4 weeks starting at 1 week after diabetes induction. Serum levels of alanine aminotransferase (ALT) and aspartate transaminase (AST), production of lipid peroxidation product malondialdehyde (MDA), and serum superoxide dismutase (SOD) were detected. Liver protein expressions of cleaved-caspase 3, IL-1ß, nuclear factor-erythroid-2-related factor-2 (Nrf2), heme oxygenase-1 (HO-1), P62, Kelch-like ECH-associated protein 1 (Keap1), and LC3 were analyzed. In vitro, cultured rat normal hepatocytes BRL-3A were grouped to normal glucose (5.5 mM, NG) or high glucose (25 mM, HG) and treated with or without allopurinol (100 µM) for 48 h. Rats in the D group demonstrated liver injury evidenced as increased serum levels of ALT and AST. Diabetes increased apoptotic cell death, enhanced liver protein expressions of cleaved-caspase 3 and IL-1ß with concomitantly increased production of MDA while serum SOD content was significantly reduced (all P < 0.05 vs C). In the meantime, protein levels of Nrf2, HO-1, and P62 were reduced while Keap1 and LC3 were increased in the untreated D group as compared to control (P < 0.05 vs C). And all the above alterations were significantly attenuated by ALP. Similar to our findings obtained from in vivo study, we got the same results in in vitro experiments. It is concluded that ALP activates the Nrf2/p62 pathway to ameliorate oxidative stress and liver injury in diabetic rats.
Asunto(s)
Alopurinol/uso terapéutico , Diabetes Mellitus Tipo 1/complicaciones , Inhibidores Enzimáticos/uso terapéutico , Lesión Pulmonar/prevención & control , Factor 2 Relacionado con NF-E2/efectos de los fármacos , Animales , Antioxidantes/metabolismo , Apoptosis/efectos de los fármacos , Línea Celular , Diabetes Mellitus Experimental/tratamiento farmacológico , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Pruebas de Función Hepática , Lesión Pulmonar/etiología , Masculino , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Estrés Oxidativo/efectos de los fármacos , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND: The glucocorticoid receptor (GR) plays a role in inflammatory pain modulation. However, the specific role played by interleukin 6 receptor (IL-6R) in these processes remains elusive. The present study aimed to investigate the extent of inflammation induced by IL-6R and GR. METHODS: Male Wistar rats were treated with Freund's complete adjuvant to induce right hind paw inflammation. The levels of IL-6Rα and GR were evaluated in the spinal cord and dorsal root ganglion using Western blot and immunofluorescence assays. Subsequently, we examined the nociceptive behavioral changes following the binding of IL-6R with a GR agonist and/or antagonist, as well as the concentration levels of IL-6 and soluble IL-6R (sIL-6R) in the serum and cerebrospinal fluid. Moreover, the spinal levels of IL-6, IL-6Rα, gp130, JAK2, pJAK2, STAT3, pSTAT3, c-fos, GFAP, and Iba-1 were assessed following anti-IL-6R antibody, sgp130, and dexamethasone intrathecal administration. RESULTS: Right hind paw inflammation resulted in significant upregulation of IL-6Rα expression in spinal nociceptive neurons, astrocytes, and microglia cells, as well as increased of IL-6Rα and GR colocalization. Notably, anti-IL-6R or dexamethasone attenuated the nociceptive behavior in a dose-dependent manner. Isobologram analysis indicated the sub-additive effects with a concomitant decrease in the spinal levels of IL-6, pJAK2, pSTAT3, c-fos, GFAP, and Iba-1 and increase in the sIL-6R level. CONCLUSION: The enhanced mechanical sensitivity accompanying the increase of IL-6Rα and GR was attenuated by anti-IL-6R and dexamethasone application, and the sub-additive effects were regulated by the decreased activation of neurons and glial cells and modulated by IL-6/JAK2/STAT3 signaling pathway, which might be attributed to IL-6 induced trans-signaling.
Asunto(s)
Inflamación/metabolismo , Dolor/metabolismo , Receptores de Glucocorticoides/metabolismo , Receptores de Interleucina-6/metabolismo , Analgésicos/farmacología , Animales , Inflamación/inmunología , Microglía/metabolismo , Neuroglía/metabolismo , Neuronas/metabolismo , Dolor/tratamiento farmacológico , Ratas Wistar , Receptores de Interleucina-6/inmunología , Médula Espinal/metabolismoRESUMEN
Allopurinol (ALP) attenuates oxidative stress and diabetic cardiomyopathy (DCM), but the mechanism is unclear. Activation of nuclear factor erythroid 2-related factor 2 (Nrf2) following the disassociation with its repressor Keap1 under oxidative stress can maintain inner redox homeostasis and attenuate DCM with concomitant attenuation of autophagy. We postulated that ALP treatment may activate Nrf2 to mitigate autophagy over-activation and consequently attenuate DCM. Streptozotocin-induced type 1 diabetic rats were untreated or treated with ALP (100 mg/kg/d) for 4 weeks and terminated after heart function measurements by echocardiography and pressure-volume conductance system. Cardiomyocyte H9C2 cells infected with Nrf2 siRNA or not were incubated with high glucose (HG, 25 mmol/L) concomitantly with ALP treatment. Cell viability, lactate dehydrogenase, 15-F2t-Isoprostane and superoxide dismutase (SOD) were measured with colorimetric enzyme-linked immunosorbent assays. ROS, apoptosis, was assessed by dihydroethidium staining and TUNEL, respectively. The Western blot and qRT-PCR were used to assess protein and mRNA variations. Diabetic rats showed significant reductions in heart rate (HR), left ventricular eject fraction (LVEF), stroke work (SW) and cardiac output (CO), left ventricular end-systolic volume (LVVs) as compared to non-diabetic control and ALP improved or normalized HR, LVEF, SW, CO and LVVs in diabetic rats (all P < .05). Hearts of diabetic rats displayed excessive oxidative stress manifested as increased levels of 15-F2t-Isoprostane and superoxide anion production, increased apoptotic cell death and cardiomyocytes autophagy that were concomitant with reduced expressions of Nrf2, heme oxygenase-1 (HO-1) and Keap1. ALP reverted all the above-mentioned diabetes-induced biochemical changes except that it did not affect the levels of Keap1. In vitro, ALP increased Nrf2 and reduced the hyperglycaemia-induced increases of H9C2 cardiomyocyte hypertrophy, oxidative stress, apoptosis and autophagy, and enhanced cellular viability. Nrf2 gene silence cancelled these protective effects of ALP in H9C2 cells. Activation of Nrf2 subsequent to the suppression of Keap1 and the mitigation of autophagy over-activation may represent major mechanisms whereby ALP attenuates DCM.
Asunto(s)
Alopurinol/farmacología , Cardiomiopatías Diabéticas/patología , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo/efectos de los fármacos , Proteína Sequestosoma-1/metabolismo , Animales , Apoptosis/efectos de los fármacos , Línea Celular , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/diagnóstico por imagen , Diabetes Mellitus Experimental/patología , Diabetes Mellitus Experimental/fisiopatología , Cardiomiopatías Diabéticas/complicaciones , Cardiomiopatías Diabéticas/diagnóstico por imagen , Cardiomiopatías Diabéticas/fisiopatología , Diástole/efectos de los fármacos , Glucosa/toxicidad , Ventrículos Cardíacos/diagnóstico por imagen , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/patología , Ventrículos Cardíacos/fisiopatología , Hemodinámica/efectos de los fármacos , Hiperglucemia/complicaciones , Hiperglucemia/patología , Hiperglucemia/fisiopatología , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Masculino , Modelos Biológicos , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Sístole/efectos de los fármacosRESUMEN
OBJECTIVE: To analyze the effect of dexmedetomidine on the inflammatory factors level and cognitive function after femoral head replacement in elderly patients. METHODS: From January 2016 to December 2017, 60 elderly patients(more than 60 years old, and Grade I to II of ASA) treated with femoral head replacement were divided into three groups, and 20 in each group. All patients received midazolam, fentanyl, etomidate, cisatracurium anesthesia induction and sevoflurane inhalation anesthesia maintenance. The patients in group B and group C were first given 1.0 µg·kg⻹ of dexmedetomidine 10 minutes during the operation. The maintenance volume was 0.3 µg·kg⻹·h⻹ of dexmedetomidine(in group B) and 0.6 µg·kg⻹·h⻹ of dexmedetomidine(in group C) by pumping. The same amount of saline was given to the patients in group A in the same way. The time of extubation, wakefulness and recovery, the simple intelligent mental state score (MMSE), the incidence of postoperative cognitive dysfunction (POCD) and the levels of interleukin-6 (IL-6), interleukin-10 (IL-10) and S100ß protein expression in the 3 groups were compared. RESULTS: There were significant differences in the time of spontaneous breathing recovery, eye opening tome and the time of extubation, as well as the dosage of propofol among the three groups(P<0.05). On the 1st, 3rd and 7th day after operation, there was a significant difference in MMSE score of group B and group C compared with that of group A(P<0.05), and MMSE score in group C was significantly higher than that of group B(P<0.05). The incidence of POCD was 0.0% (0/20) and the incidence of adverse reactions was 30%(6/20) in group C, but those were 25% (5/20) and 0.0% (0/20) in group A and 5% (1/20) and 10% (2/20) respectively in group B. The difference was statistically significant (P<0.05). Before induction of anesthesia, there was no significant difference in the levels of IL-6, IL-10 and S100ß protein among the three groups(P>0.05); but one hour after the operation, the levels of IL-6 IL-10 and S100ß protein in group B and group C was statistically different from those in group A(P<0.05). The IL-6 and S100ß protein in group C were significantly lower than those in group B (P<0.05), and IL-10 was significantly higher than that in group B (P<0.05). CONCLUSIONS: For elderly patients operated for femoral head replacement, dexmedetomidine can reduce the level of inflammatory factors level and propofol consumption, and the incidence of postoperative POCD is low, indicating a dose dependence of dexmedetomidine. But it is necessary to choose the right dose according to the patient's condition.
Asunto(s)
Cognición , Anciano , Delirio , Dexmedetomidina , Humanos , Interleucina-6 , Persona de Mediana Edad , SevofluranoRESUMEN
Analgesics and antineoplastic drugs are often used concurrently for cancer patients. Our previous study reported that gap junctions composed of connexin32 (Cx32) was implicated in the effect of analgesics on cisplatin cytotoxicity. However, the effect of analgesic on the most widely expressed connexin (Cx), connexin43 (Cx43), and whether such effect mediates the influence on chemotherapeutic efficiency remain unknown. By manipulation of Cx43 expression or gap junction function, we found that there were gap junction-dependent and independent effect of Cx43 on temozolomide (TMZ) sensitivity in U87 glioblastoma cells. Studies on survival and apoptosis showed widely used analgesic tramadol significantly reduced TMZ-induced cytotoxicity in control and negative control cells but not shCx43-transfected cells. Proliferation assay demonstrated tramadol suppressed TMZ-induced cytotoxicity only on high density (with gap junction formation) but not on low density (without gap junction formation). Tramadol inhibited dye-coupling through gap junctions between U87 cells. Tramadol treatment for 72 h did not alter Cx43 expression, but decreased Cx43 phosphorylation accompanied with reduced p-ERK and p-JNK. Our results indicated that long-term treatment with tramadol reduced TMZ cytotoxicity in U87 cells by suppressing Cx43-composed gap junctions, suggesting identification and usage of antinociceptive drugs which do not downregulate connexin activity should have beneficial therapeutic consequences.
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Neoplasias Encefálicas/tratamiento farmacológico , Comunicación Celular/efectos de los fármacos , Conexina 43/metabolismo , Dacarbazina/análogos & derivados , Uniones Comunicantes/efectos de los fármacos , Glioblastoma/tratamiento farmacológico , Tramadol/farmacología , Analgésicos Opioides/farmacología , Antineoplásicos Alquilantes/farmacología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Comunicación Celular/genética , Línea Celular Tumoral , Conexina 43/genética , Dacarbazina/administración & dosificación , Dacarbazina/farmacología , Interacciones Farmacológicas , Uniones Comunicantes/genética , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patología , Humanos , Temozolomida , Tramadol/administración & dosificación , TransfecciónRESUMEN
Acute kidney injury (AKI) represents a group of complicated syndromes with a high mortality rate. The administration of adipose-derived mesenchymal stem cells (ADMSCs) has been tested as a possible treatment method for AKI. The long-term evaluation of AKI induced by ischemia/reperfusion (IR) and the probable renal protection of ADMSCs are limited. In this study we have established a rat AKI model induced by IR and investigated the possible protective effects of ADMSCs. Adult Sprague-Dawley (SD) rats were divided into three groups (n = 6/each group). The MOCK group was as the normal control. Rats in the IR-AKI and IR-AKI+ADMSCs groups were subjected to IR injury by clamping both renal pedicles for 40 minutes. Rats in the MOCK and IR-AKI groups were injected with PBS via the tail vein as negative treatment controls. Rats in the IR-AKI+ADMSCs group received ADMSCs therapy (2 × 106 cells were injected into the rats via the tail vein). We found that ADMSC transplantation restored the pathologic morphology induced by IR-AKI to normal compared with the MOCK group, suggesting the reparative function of ADMSCs in kidney tissues. Compared with IR-induced AKI alone, ADMSC treatment significantly decreased the number of apoptotic cells, the level of total urinary protein and serum creatinine, the expression of pro-inflammatory cytokines (IL-6, TNF-α, IL-1ß, IFN-γ, TNF-α, IFN-γ, and TGF-ß), and the inflammation-associated proteins (HGF and SDF1), but increased the expression of the anti-inflammatory cytokine, IL-10, and the anti-apoptotic regulator, Bcl-2. Our data have indicated that ADMSC transplantation may protect against IR-induced AKI by anti-apoptotic and anti-inflammatory effects.
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Lesión Renal Aguda/terapia , Tejido Adiposo/citología , Riñón/irrigación sanguínea , Trasplante de Células Madre Mesenquimatosas/métodos , Daño por Reperfusión/complicaciones , Lesión Renal Aguda/etiología , Lesión Renal Aguda/inmunología , Lesión Renal Aguda/patología , Animales , Apoptosis , Modelos Animales de Enfermedad , Inflamación , Riñón/inmunología , Riñón/patología , Pruebas de Función Renal , Masculino , Ratas Sprague-DawleyRESUMEN
Isoflurane postconditioning (IsoPostC) attenuates myocardial ischaemia/reperfusion injury (IRI). Signal transducer and activator of transcription-3 (STAT3) is critical in ischaemic postconditioning cardioprotection, which can be regulated by the Brahma-related gene (Brg1) and nuclear factor-erythroid 2-related factor 2 (Nrf2), although they are both reduced in diabetic hearts. We hypothesized that reduced Brg1/Nrf2 and STAT3 activation may jeopardize IsoPostC-mediated cardioprotection in diabetic hearts. In the present study, Langendorff-perfused, non-diabetic (control) and 8-week-old streptozotocin-induced Type 1 diabetic rat hearts were subjected to 30 min of global ischaemia and 120 min of reperfusion without or with IsoPostC, which was achieved by administering emulsified isoflurane (2.0%, v/v) in Krebs-Henseleit (KH) solution immediately at the onset of reperfusion for 10 min and switching to KH solution perfusion alone thereafter. Cultured H9C2 cells were exposed to normal glucose (NG, 5.5 mM) or high glucose (HG, 30 mM) and subjected to hypoxia/reoxygenation (HR) in the presence or absence of IsoPostC. Diabetic rats displayed larger post-ischaemic myocardial infarction and more severe haemodynamic dysfunction, associated with increased myocardial oxidative stress and reduced cardiac Brg1, Nrf2 and STAT3 phosphorylation/activation (p-STAT3), compared with controls. These changes were reversed/prevented by IsoPostC in control but not in diabetic rats. In H9C2 cells exposed to NG but not HG, IsoPostC significantly attenuated HR-induced cellular injury and superoxide anion production with increased Brg1, Nrf2 and p-STAT3. These beneficial effects of IsoPostC were abolished by Brg1, Nrf2 or STAT3 gene knockdown. Brg1 or Nrf2 gene knockdown abolished IsoPostC-induced STAT3 activation. N-acetylcysteine restored Brg1, Nrf2 and p-STAT3, and IsoPostC-induced protection in H9C2 cells exposed to HG and HR. In conclusion, IsoPostC confers cardioprotection through Brg1/Nrf2/STAT3 signalling, and impairment of this pathway may be responsible for the loss of IsoPostC cardioprotection in diabetes.
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ADN Helicasas/metabolismo , Diabetes Mellitus Experimental/tratamiento farmacológico , Isoflurano/farmacología , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Factor 2 Relacionado con NF-E2/metabolismo , Proteínas Nucleares/metabolismo , Factor de Transcripción STAT3/metabolismo , Transducción de Señal/efectos de los fármacos , Factores de Transcripción/metabolismo , Animales , Diabetes Mellitus Experimental/complicaciones , Modelos Animales de Enfermedad , Poscondicionamiento Isquémico/métodos , Isoflurano/administración & dosificación , Masculino , Infarto del Miocardio/metabolismo , Daño por Reperfusión Miocárdica/metabolismo , Ratas Sprague-Dawley , EstreptozocinaRESUMEN
Cisplatin, an important chemotherapeutic agent against testicular germ cell cancer, induces testicular toxicity on Leydig and Sertoli cells, leading to serious side-effects such as azoospermia and infertility. In a previous study, it was found that simvastatin enhanced the sensitivity of Leydig tumor cells to chemotherapeutic toxicity through the enhancement of gap junction functions. In the present study, the effect of simvastatin on the sensitivity of normal Sertoli cells to cisplatin and the role of gap junctions in such effects was investigated. The results showed that, simvastatin attenuated cisplatin toxicity only when cells exhibited high-density culture where gap junctional formation was possible. When gap junction function was decreased by the gap junction inhibitor or by siRNA targeting connexin 43, the protective effect of simvastatin to cisplatin toxicity was substantially attenuated. Simvastatin also enhanced gap junction functions between Sertoli cells. This effect was mediated by the reduction of PKC-mediated connexin phosphorylation, thereby increasing connexin 43 membrane localization. Thus, simvastatin-induced enhancement of gap junctionmediated intercellular communication attenuated cisplatin toxicity on Sertoli cells. This result indicated that enhancement of gap junction function by simvastatin may have bilateral beneficial effects on cisplatinbased chemotherapy, enhancing cisplatin killing on cancer while ameliorating the reproduction toxicity.
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Antineoplásicos/toxicidad , Cisplatino/toxicidad , Uniones Comunicantes/efectos de los fármacos , Células de Sertoli/efectos de los fármacos , Simvastatina/farmacología , Animales , Comunicación Celular/efectos de los fármacos , Línea Celular , Conexina 43/metabolismo , Masculino , Ratones , Fosforilación/efectos de los fármacos , Proteína Quinasa C/metabolismo , Células de Sertoli/fisiologíaRESUMEN
OBJECTIVE: To examine the effect of acute incisional pain on the expression of connexin 43 in rat spinal cord dorsal horn. METHODS: Eighty rats were assigned into control group without any treatment and incisional pain group with incision surgery. For paw incisions, a 1-cm longitudinal incision was made through the skin and fascia of the plantar aspect of the right hind paw. After surgery, the 50% paw withdrawal threshold (PWT) was assessed in response to a tactile stimulus with calibrated von Frey monofilaments at 1, 2, 4 and 24 h, respectively. The spinal cord dorsal horn of rats was isolated at 1, 2, and 4 h after the surgery to assess the expression of connexin 43 using Western blotting and immunofluorescence assay. RESULTS: The 50% PWT of the rats was significantly decreased after the incision surgery, and this decrement was the most obvious at 2 and 4 h. Western blotting and immunofluorescence assay showed that the expression of connexin 43 in the spinal cord dorsal horn was significantly increased in rats receiving the surgery especially at 2 and 4 h after the surgery. CONCLUSION: Incision surgery induces an significant increase in connexin 43 expression in rat spinal cord dorsal horn, suggestting an potential role of connexin43 in postoperative incisional pain.
Asunto(s)
Conexina 43/metabolismo , Dolor Postoperatorio/metabolismo , Asta Dorsal de la Médula Espinal/metabolismo , Animales , Ratas , Ratas Sprague-DawleyRESUMEN
UNLABELLED: To investigate the effects of Ulinastatin (UTI) in cerebral ischemia-reperfusion (IR) injury in rats and whether this effect might be related to Aquaporin 4 (AQP4), one hundred and eighty adult male Sprague Dawley (SD) rats, weighing 230-280 g, were randomly divided into 3 groups: sham (S) group, IR group and UTI (U) group. Every group was further divided into 3 sub-groups: 6 h group, 24 h group and 48 h group. The transient focal IR injury was induced by inserting a silicone-coater monofilament nylon suture (0.28 mm) from the right external carotid artery to the origin of the left middle cerebral artery. The suture was removed after 2 h to allow reperfusion. UTI treatment group was injected with UTI 100000 u/kg at the beginning of the reperfusion period, while S group and IR group were injected with the same volume of saline. Samples were taken according to the reperfusion time (6 h, 12 h and 24 h). Infract volume was measured by triphenyl tetrazolium chloride staining, and brain water content was determined by wet-dry weight method and neurological scores were checked with a five-point scale. Expression levels of AQP4 were checked with immunohistochemistry and Western blot. RESULTS: Compared with S group, the infarct volume, water content, neurological scores and AQP4 levels in the rat brain tissues were significantly increased in IR group. Administration of UTI significantly decreased the infarct volume, water content of the brain tissue and neurological scores. Moreover, the expression levels of AQP4 were also down-regulated by UTI treatment. CONCLUSION: UTI improves cerebral IR injury in rats potentially via decreasing the expression levels of AQP4.
RESUMEN
OBJECTIVE: Bumetanide has been reported to attenuate ischemia-evoked cerebral edema. However, whether bumetanide can protect cerebral ischemia-reperfusion injury (IRI) in vivo is unclear. In the present study, we aim to determine whether intravenously injection bumetanide can attenuate cerebral IRI and if its protection effect might be related to the modification of cerebral NKCC1 and KCC2 protein expression. METHODS: Focal cerebral ischemia was induced by occluding the right middle cerebral artery (MCAO) for 2-h, followed by 3-h, 24-h or 48-h of reperfusion respectively. Brain edema, neurological deficits, and infarction volume were determined by (wet weights-dry weights)/dry weights×100, 5-point neurological function score evaluation system, and TTC staining, respectively. The expression levels of NKCC1 and KCC2 were determined by immunohistochemical staining. RESULTS: Reperfusion increased brain edema, neurological deficits, and infarction volume. Bumetanide decreased brain edema, attenuated the neurological defects and reduced post-ischemic cerebral infarction. Cerebral ischemia-reperfusion injury increased NKCC1 expression level and decreased KCC2 expression level. Interestingly, bumetanide down-regulated the NKCC1 protein expression level without changing the KCC2 protein expression level in rat brain cortex. CONCLUSION: These results suggest that bumetanide protects focal cerebral ischemia-reperfusion injury in rat, which might through the inhibition of NKCC1.
Asunto(s)
Isquemia Encefálica/prevención & control , Bumetanida/uso terapéutico , Daño por Reperfusión/prevención & control , Inhibidores del Simportador de Cloruro Sódico y Cloruro Potásico/uso terapéutico , Animales , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Bumetanida/administración & dosificación , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Modelos Animales de Enfermedad , Inyecciones Intravenosas , Masculino , Ratas , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Inhibidores del Simportador de Cloruro Sódico y Cloruro Potásico/administración & dosificación , Miembro 2 de la Familia de Transportadores de Soluto 12/metabolismo , Simportadores/metabolismo , Factores de Tiempo , Cotransportadores de K ClRESUMEN
Ischemic stroke is the most common type of stroke and brings about a big disease burden because of high mortality and disability in China. Tetrandrine, a bisbenzylisoquinoline alkaloid isolated from the Chinese herb Radix Stephania tetrandra, has been demonstrated to possess anti-inflammatory and free radical scavenging effects and even regulate astrocyte activation, but the possible role of tetrandrine in ameliorating cerebral ischemia/reperfusion injury of ischemic stroke remains unknown. The aim of this study was to determine the effects of tetrandrine on neurological injury and differential proteomic changes induced by transient reversible middle cerebral artery occlusion (MCAO) in mice. Male Balb/c mice were divided into sham (n = 30), MCAO + saline as control (n = 30), and MCAO + Tet as tetrandrine-treated (n = 30) groups. Mice in the control and tetrandrine-treated groups underwent 120 min of MCAO following reperfusion. Immediately and 2 h after MCAO, the mice received either normal saline (sham operated and control groups) or tetrandrine (tetrandrine-treated group) intraperitoneally. Neurological defects, brain water content, and infarct volume at 24 h after stoke were used to evaluate neurological injury extent. Treatment with tetrandrine not only mitigated cerebral neurological deficits (P < 0.05) and infarct size (P < 0.01), but also decreased brian edema in the ischemic brain (P < 0.05). Then, fluorescence two-dimensional difference in gel electrophoresis was used to detect our systematic differential profiling of proteomic changes responding to tetrandrine administration. We validated that the expression of GRP78, DJ-1 and HYOU1 was associated with neuroprotective effect of tetrandrine in MCAO model by Western blotting. These findings indicate a potential neuroprotective role of tetrandrine for ischemic stroke and yield insights into cellular and molecular mechanisms of tetrandrine taking place in ischemic stroke.
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Bencilisoquinolinas/farmacología , Isquemia Encefálica/metabolismo , Modelos Animales de Enfermedad , Electroforesis en Gel Bidimensional/métodos , Infarto de la Arteria Cerebral Media/metabolismo , Proteómica , Daño por Reperfusión/metabolismo , Animales , Chaperón BiP del Retículo Endoplásmico , Masculino , Espectrometría de Masas , Ratones , Ratones Endogámicos BALB CRESUMEN
BACKGROUND: The transcription factor nuclear factor κB (NF-κB) regulates the expression of numerous proinflammatory factors that may exacerbate the response to acute injury. We investigated the effect of an inhibitory NF-κB decoy oligodeoxynucleotide on proinflammatory factor expression and pulmonary function after acute lung contusion in rabbits. METHODS: Thirty-minutes after acute lung contusion, the NF-κB decoy or a scrambled control oligodeoxynucleotide was injected via the jugular vein. Blood samples were collected for blood gas analysis and plasma tumor necrosis factor α, interleukin 1ß (IL-1ß), IL-13, and IL-10 were measured by enzyme-linked immunosorbent assay at 1, 2, 3, and 4 hours after contusion. In addition, NF-κB protein expression in lung tissue was detected by Western blot analysis. RESULTS: The blood PO2 decreased immediately after lung contusion, whereas PAO2 increased significantly, indicative of disrupted respiratory function. Respiratory function improved after sense NF-κB decoy injection but not after injection of the inactive scrambled form. Injection of NF-κB decoy resulted in significant inhibition of NF-κB protein expression in lung tissue and a reduction in the serum concentrations of proinflammatory cytokines tumor necrosis factor α and IL-1ß compared with those of control rabbits injected with the scrambled decoy. In contrast, serum levels of the anti-inflammatory cytokines IL-10 and IL-13 increased after decoy injection compared with those of control animals and rabbits injected with the scrambled decoy. CONCLUSION: The sense NF-κB decoy protected respiratory function and reduced serum proinflammatory factor secretion after acute lung contusion. Inhibition of NF-κB may allow for preservation of pulmonary function for patients with acute lung injury.
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Lesión Pulmonar/prevención & control , Oligodesoxirribonucleótidos/administración & dosificación , Transfección/métodos , Animales , Western Blotting , Citocinas/sangre , Modelos Animales de Enfermedad , Inyecciones Intravenosas , Pulmón/metabolismo , Pulmón/ultraestructura , Lesión Pulmonar/genética , Lesión Pulmonar/fisiopatología , Microscopía Electrónica de Transmisión , Oligodesoxirribonucleótidos/biosíntesis , Oligodesoxirribonucleótidos/genética , Conejos , Factores de Transcripción/genéticaRESUMEN
OBJECTIVE: Recently, evidence has accumulated to show that cation-chloride cotransporters (CCCs) participate in the modulation of pain transmission at the spinal cord level. To investigate whether CCC inhibitors might affect surgical pain, we examined the effect of furosemide in a rat incisional pain model. DESIGN: We examined pain thresholds using von Frey filaments in intact and incision model Sprague-Dawley rats before and after intrathecal furosemide administration (100 µg/20 µL). Twenty-four rats were divided into four groups (N=6), groups A and B were intact rats, which received furosemide and its solvent, respectively. Groups C and D were incision rats, which received furosemide and its solvent, respectively. Non-parametric tests were used to calculate pain thresholds, and P<0.05 was considered significant. RESULTS: Furosemide decreased the pain threshold (vs solvent) in intact rats for 2 hours after administration and caused excitatory behavior. However, furosemide increased the pain threshold in incision model rats ([1] at the incision point: at 20 minutes, 2 hours, 3 hours, 4 hours, and on the second to fifth days after incision; [2] at the remote point: at 20 minutes, 3-5 hours, and from the 3rd to the 7th day after incision) and improved wound recovery. CONCLUSIONS: Intrathecal administration of the CCC inhibitor furosemide had antinociceptive effects in rats with incisional pain. Furosemide may be a novel treatment for postoperative pain.