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Osteoarthritis (OA) is a degenerative joint disease primarily affecting the elderly. It is characterized by the progressive decline of joint cartilage and alterations in the underlying bone. Several probiotic strains have exhibited immunomodulatory and anti-inflammatory properties. Here, we examined the functions of live and dead Clostridium butyricum GKB7 (GKB7-L and GKB7-D) in a preclinical anterior cruciate ligament transection (ACLT)-enhanced OA procedure. Oral administration of GKB7-L and GKB7-D ameliorated ACLT-induced bone pain as assessed by weight-bearing behavioral testing but did not affect body weight. Micro-computed tomography (CT) results showed that GKB7-L and GKB7-D diminished ACLT-induced bone destruction and loss. GKB7-L and GKB7-D-enriched therapies also reduced ACLT-induced production of the pro-inflammatory cytokines interleukin (IL)-1ß and tumor necrosis factor (TNF)-α, as well as the chondrolytic factor matrix metalloproteinase (MMP)-3, leading to inhibition of aggrecan and collagen type II degradation and thereby blocking cartilage breakdown. We therefore suggest that oral supplementation with GKB7-L or GKB7-D can be beneficial in the prevention and treatment of OA.
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We used gastric cancer cell line AGS and clinical samples to investigate the roles of mitochondrial DNA (mtDNA) alterations and mitochondrial respiratory dysfunction in gastric adenocarcinoma (GAC). A total of 131 clinical samples, including 17 normal gastric mucosa (N-GM) from overweight patients who had received sleeve gastrectomy and 57 paired non-cancerous gastric mucosae (NC-GM) and GAC from GAC patients who had undergone partial/subtotal/total gastrectomy, were recruited to examine the copy number and D310 sequences of mtDNA. The gastric cancer cell line AGS was used with knockdown (KD) mitochondrial transcription factor A (TFAM) to achieve mitochondrial dysfunction through a decrease of mtDNA copy number. Parental (PT), null-target (NT), and TFAM-KD-(A/B/C) represented the parental, control, and TFAM knocked-down AGS cells, respectively. These cells were used to compare the parameters reflecting mitochondrial biogenesis, glycolysis, and cell migration activity. The median mtDNA copy numbers of 17 N-GM, 57 NC-GM, and 57 GAC were 0.058, 0.055, and 0.045, respectively. The trend of decrease was significant (p = 0.030). In addition, GAC had a lower mean mtDNA copy number of 0.055 as compared with the paired NC-GM of 0.078 (p < 0.001). The mean mtDNA copy number ratio (mtDNA copy number of GAC/mtDNA copy number of paired NC-GM) was 0.891. A total of 35 (61.4%) GAC samples had an mtDNA copy number ratio ≤0.804 (p = 0.017) and 27 (47.4%) harbored a D310 mutation (p = 0.047), and these patients had shorter survival time and poorer prognosis. After effective knockdown of TFAM, TFAM-KD-B/C cells expressed higher levels of hexokinase II (HK-II) and v-akt murine thymoma viral oncogene homolog 1 gene (AKT)-encoded AKT, but lower levels of phosphorylated pyruvate dehydrogenase (p-PDH) than did the NT/PT AGS cells. Except for a higher level of p-PDH, the expression levels of these proteins remained unchanged in TFAM-KD-A, which had a mild knockdown of TFAM. Compared to those of NT, TFAM-KD-C had not only a lower mtDNA copy number (p = 0.050), but also lower oxygen consumption rates (OCR), including basal respiration (OCRBR), ATP-coupled respiration (OCRATP), reserve capacity (OCRRC), and proton leak (OCRPL)(all with p = 0.050). In contrast, TFAM-KD-C expressed a higher extracellular acidification rate (ECAR)/OCRBR ratio (p = 0.050) and a faster wound healing migration at 6, 12, and 18 h, respectively (all with p = 0.050). Beyond a threshold, the decrease in mtDNA copy number, the mtDNA D310 mutation, and mitochondrial dysfunction were involved in the carcinogenesis and progression of GACs. Activation of PDH might be considered as compensation for the mitochondrial dysfunction in response to glucose metabolic reprogramming or to adjust mitochondrial plasticity in GAC.
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Adenocarcinoma/cirugía , ADN Mitocondrial/genética , Proteínas de Unión al ADN/genética , Mitocondrias/metabolismo , Proteínas Mitocondriales/genética , Obesidad/cirugía , Neoplasias Gástricas/cirugía , Factores de Transcripción/genética , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Línea Celular Tumoral , Movimiento Celular , Variaciones en el Número de Copia de ADN , Femenino , Gastrectomía , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Glucólisis , Humanos , Masculino , Persona de Mediana Edad , Mitocondrias/genética , Obesidad/genética , Obesidad/metabolismo , Biogénesis de Organelos , Pronóstico , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Análisis de SupervivenciaRESUMEN
Fast radio bursts are bright, unresolved, non-repeating, broadband, millisecond flashes, found primarily at high Galactic latitudes, with dispersion measures much larger than expected for a Galactic source. The inferred all-sky burst rate is comparable to the core-collapse supernova rate out to redshift 0.5. If the observed dispersion measures are assumed to be dominated by the intergalactic medium, the sources are at cosmological distances with redshifts of 0.2 to 1 (refs 10 and 11). These parameters are consistent with a wide range of source models. One fast burst revealed circular polarization of the radio emission, but no linear polarization was detected, and hence no Faraday rotation measure could be determined. Here we report the examination of archival data revealing Faraday rotation in the fast radio burst FRB 110523. Its radio flux and dispersion measure are consistent with values from previously reported bursts and, accounting for a Galactic contribution to the dispersion and using a model of intergalactic electron density, we place the source at a maximum redshift of 0.5. The burst has a much higher rotation measure than expected for this line of sight through the Milky Way and the intergalactic medium, indicating magnetization in the vicinity of the source itself or within a host galaxy. The pulse was scattered by two distinct plasma screens during propagation, which requires either a dense nebula associated with the source or a location within the central region of its host galaxy. The detection in this instance of magnetization and scattering that are both local to the source favours models involving young stellar populations such as magnetars over models involving the mergers of older neutron stars, which are more likely to be located in low-density regions of the host galaxy.
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BACKGROUND: Hyperglycemia evoked oxidative stress contributing to diabetes (DM)-induced voiding dysfunction. We explored whether antioxidant sulforaphane,a NF-E2-related nuclear factor erythroid-2 (Nrf-2) activator, may ameliorate DM-induced bladder dysfunction. METHODS: DM was induced by streptozotocin and sulforaphanewas administered before DM induction.Bladder reactive oxygen species (ROS) were determined by an ultrasensitive chemiluminescence analyzer. Mitochondrial function index mitochondrial Bax and cytosolic cytochrome c, antioxidant defense Nrf-2/HO-1, endoplasmic reticulum stress marker ATF-6/CHOP, and caspase 3/PARP were evaluated by Western blot. RESULTS: DM increased Keap1 and reduced Nrf-2 expression, associated with increase of bladder ROS, mitochondrial Bax translocation, cytosolic cytochrome c release, ATF-6/CHOP, caspase-3/PARP in bladders which resulted in voiding dysfunction by increased intercontraction intervals and micturition duration. However, sulforaphanesignificantly increased nuclear Nrf-2/HO-1axis expression, decreased bladder ROS amount, mitochondrial Bax translocation, cytochrome c release, ATF-6/CHOP and caspase 3/PARP/apoptosis, thereby improved the voiding function by the shortened intercontraction intervals and micturition duration. CONCLUSION: We suggest that sulforaphanevia activating Nrf-2/HO-1 signaling preserved mitochondrial function and suppressed DM-induced ROS, endoplasmic reticulum stress, apoptosis and voiding dysfunction.
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Diabetes Mellitus Experimental , Isotiocianatos/uso terapéutico , Mitocondrias/efectos de los fármacos , Micción/efectos de los fármacos , Animales , Apoptosis , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Estrés del Retículo Endoplásmico , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Mitocondrias/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo , Ratas , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , SulfóxidosRESUMEN
Repetitive hypoxic preconditioning (HP) enforces protective effects to subsequently severe hypoxic/ischemic stress. We hypothesized that HP may provide protection against ischemia/reperfusion (I/R) injury in rat livers via hypoxia-induced factor-1 alpha (HIF-1α)/reactive oxygen species (ROS)-dependent defensive mechanisms. Female Wistar rats were exposed to hypoxia (15 h/day) in a hypobaric hypoxic chamber (5500 m) for HP induction, whereas the others were kept in sea level. These rats were subjected to 45 min of hepatic ischemia by portal vein occlusion followed by 6 h of reperfusion. We evaluated HIF-1α in nuclear extracts, MnSOD, CuZnSOD, catalase, Bad/Bcl-xL/caspase 3/poly-(ADP-ribose)-polymerase (PARP), mitochondrial Bcl-xL, and cytosolic cytochrome C expression with Western blot and nitroblue tetrazolium/3-nitrotyrosine stain. Kupffer cell infiltration and terminal deoxynucleotidyl transferase-mediated nick-end labeling method apoptosis were determined by immunocytochemistry. The ROS value from liver surface and bile was detected by an ultrasensitive chemiluminescence-amplification method. Hepatic function was assessed with plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels. HP increased nuclear translocation of HIF-1α and enhanced Bcl-xL, MnSOD, CuZnSOD, and catalase protein expression in a time-dependent manner. The response of HP enhanced hepatic HIF-1α, and Bcl-xL expression was abrogated by a HIF-1α inhibitor YC-1. Hepatic I/R increased ROS levels, myeloperoxidase activity, Kupffer cell infiltration, ALT and AST levels associated with the enhancement of cytosolic Bad translocation to mitochondria, release of cytochrome C to cytosol, and activation of caspase 3/PARP-mediated apoptosis. HP significantly ameliorated hepatic I/R-enhanced oxidative stress, apoptosis, and mitochondrial and hepatic dysfunction. In summary, HP enhances HIF-1α/ROS-dependent cascades to upregulate mitochondrial Bcl-xL protein expression and to confer protection against I/R injury in the livers.
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Hepatopatías , Daño por Reperfusión , Animales , Apoptosis , Femenino , Hipoxia , Subunidad alfa del Factor 1 Inducible por Hipoxia , Mitocondrias , Ratas , Ratas Wistar , Regulación hacia ArribaRESUMEN
5-Methoxytryptophan (5-MTP) is a tryptophan metabolite with recently discovered anti-inflammatory and tumor-suppressing activities. Its synthesis is catalyzed by a hydroxyindole O-methyltransferase (HIOMT)-like enzyme. However, the exact identity of this HIOMT in human cells remains unclear. Human HIOMT exists in several alternatively spliced isoforms, and we hypothesized that 5-MTP-producing HIOMT is a distinct isoform. Here, we show that human fibroblasts and cancer cells express the HIOMT298 isoform as contrasted with the expression of the HIOMT345 isoform in pineal cells. Sequencing analysis of the cloned isoforms revealed that HIOMT298 is identical to the sequence of a previously reported truncated HIOMT isoform. Of note, HIOMT298 expression was reduced in cancer cells and tissues. Stable transfection of A549 cancer cells with HIOMT298 restored HIOMT expression to normal levels, accompanied by 5-MTP production. Furthermore, HIOMT298 transfection caused a tryptophan-metabolic switch from serotonin to 5-MTP production. To determine the in vivo relevance of this alteration, we compared growth and lung metastasis of HIOMT298-transfected A549 cells with those of vector- or untransfected A549 cells as controls in a murine xenograft model. Of note, the HIOMT298-transfected A549 cells exhibited slower growth and lower metastasis than the controls. Our findings provide insight into the crucial role of HIOMT298 in 5-MTP production in cells and in inhibiting cancer progression and highlight the potential therapeutic value of 5-MTP for managing cancer.
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Acetilserotonina O-Metiltransferasa/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Serotonina/metabolismo , Triptófano/análogos & derivados , Triptófano/metabolismo , Animales , Apoptosis , Proliferación Celular , Humanos , Masculino , Ratones , Ratones SCID , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Observations of 21-cm radio emission by neutral hydrogen at redshifts z approximately 0.5 to approximately 2.5 are expected to provide a sensitive probe of cosmic dark energy. This is particularly true around the onset of acceleration at z approximately 1, where traditional optical cosmology becomes very difficult because of the infrared opacity of the atmosphere. Hitherto, 21-cm emission has been detected only to z = 0.24. More distant galaxies generally are too faint for individual detections but it is possible to measure the aggregate emission from many unresolved galaxies in the 'cosmic web'. Here we report a three-dimensional 21-cm intensity field at z = 0.53 to 1.12. We then co-add neutral-hydrogen (H i) emission from the volumes surrounding about 10,000 galaxies (from the DEEP2 optical galaxy redshift survey). We detect the aggregate 21-cm glow at a significance of approximately 4sigma.
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BACKGROUND: 14-3-3σ is implicated in promoting tumor development of various malignancies. However, the clinical relevance of 14-3-3σ in hepatocellular carcinoma (HCC) tumor progression and modulation and pathway elucidation remain unclear. METHODS: We investigated 14-3-3σ expression in 109 HCC tissues by immunohistochemistry. Overexpression and knockdown experiments were performed by transfection with cDNA or siRNA. Protein expression and cell migration were determined by Western blot and Boyden chamber assay. RESULTS: In this study, we found that 14-3-3σ is abundantly expressed in HCC tumors. Stable or transient overexpression of 14-3-3σ induces the expression of heat shock factor-1α (HSF-1α) and heat shock protein 70 (HSP70) in HCC cells. Moreover, expression of 14-3-3σ significantly correlates with HSF-1α/HSP70 in HCC tumors and both 14-3-3σ and HSP70 overexpression are associated with micro-vascular thrombi in HCC patients, suggesting that 14-3-3σ/HSP70 expression is potentially involved in cell migration/invasion. Results of an in vitro migration assay indicate that 14-3-3σ promotes cell migration and that 14-3-3σ-induced cell migration is impaired by siRNA knockdown of HSP70. Finally, 14-3-3σ-induced HSF-1α/HSP70 expression is abolished by the knockdown of ß-catenin or activation of GSK-3ß. CONCLUSIONS: Our findings indicate that 14-3-3σ participates in promoting HCC cell migration and tumor development via ß-catenin/HSF-1α/HSP70 pathway regulation. Thus, 14-3-3σ alone or combined with HSP70 are potential prognostic biomarkers for HCC.
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Proteínas 14-3-3/metabolismo , Biomarcadores de Tumor/metabolismo , Carcinoma Hepatocelular/genética , Exorribonucleasas/metabolismo , Proteínas HSP70 de Choque Térmico/biosíntesis , Neoplasias Hepáticas/genética , Anciano , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Movimiento Celular/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Proteínas HSP70 de Choque Térmico/metabolismo , Factores de Transcripción del Choque Térmico , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Pronóstico , Transducción de Señal , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
Cyclooxygenase-2(COX-2) overexpression promotes inflammation and tumorigenesis. COX-2 expression in response to diverse stimuli is tightly controlled to avoid persistent overexpression. 5-methoxyindole metabolites of L-tryptophan represent a new class of compounds that control COX-2 expression at the transcriptional level. Two of the metabolites, the newly discovered 5-methoxytryptophan (5-MTP, also known as cytoguardin) and N-acetyl 5-methoxytryptamine (melatonin) are the focus of this review. 5-MTP is produced by mesenchymal cells such as fibroblasts via 5-hydroxytryptophan (5-HTP). It inhibits COX-2 transcriptional activation induced by diverse proinflammatory and mitogenic factors. Cancer cells are deficient in cytoguardin production which contributes to COX-2 overexpression. Fibroblast-generated 5-MTP is capable of restoring the control of COX-2 overexpression in cancer cells. 5-MTP blocks cancer cell migration and invasion in vitro and inhibits tumor growth and cancer metastasis in a xenograft model. Melatonin possesses similar COX-2 suppressing and anti-cancer properties albeit at supra-pharmacological concentrations. By contrast, 5-hydroxyindole metabolites of L-tryptophan such as 5-hydroxytryptamine (serotonin), 5-hydroxytryptophol and other serotonin catabolites do not control COX-2 expression. 5-hydroxytryptophan inhibits COX-2 expression through conversion to 5-MTP. The physiological relevance of 5-MTP as an endogenous regulator of inflammation and cancer metastasis remains to be investigated. On the other hand, 5-methoxyindole metabolites of tryptophan are valuable lead compounds for development of new anti-inflammatory drugs and cancer chemoprevention.
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Ciclooxigenasa 2/biosíntesis , Indoles/metabolismo , Neoplasias/genética , Triptófano/metabolismo , Carcinogénesis/genética , Ciclooxigenasa 2/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Inflamación/genética , Melatonina/genética , Melatonina/metabolismo , Metástasis de la Neoplasia , Serotonina/metabolismoRESUMEN
Benzophenones (BPs) are widely used as photoinitiators (PIs) or printing inks in food packaging, which may migrate into foods. However, the toxicity information of some BP analogues, such as 4,4'-bis(diethylamino)-benzophenone (DEAB), 4-phenylbenzophenone (4-PBP), 4 (hydroxymethyl)benzophenone (4-HMBP), those are used as PIs is lacking. Developmental toxicity is a health concern associated with PIs exposure. Recently, alternative non-in vivo methods have been proposed to evaluate the concerned chemicals or better understand the modes of action of certain toxicological endpoints. In this study, using in silico methods, we predicted that BP, DEAB, 4-PBP and 4-HMBP might exhibit developmental toxicity. However, we found that only DEAB is strong embryotoxic and disturbs the early differentiation of mouse embryonic stem cells into three germ layers and cardiomyocytes. DEAB treatment also prevented cardiomyocyte differentiation in human induced pluripotent stem cells (hiPSCs) on day 10. However, BP, 4-PBP and 4-HMBP had no similar effects on cardiomyocyte differentiation on day 10. Transcriptomic analysis revealed that treatment with DEAB significantly decreased the mRNA levels of differentiation-related transcription factors SOX17 and FOXA1, in hiPSCs on day 4. Furthermore, DEAB treatment caused tail malformations and yolk sac edema in zebrafish embryos. To conclude, DEAB may be embryotoxic because it disturbs the early differentiation of stem cells. Further studies are warranted to better understand the health effects of DEAB exposure.
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AIMS: The results of our earlier studies suggested that 14-3-3ε is involved in cancer cell survival and growth. However, it is not clear whether 14-3-3ε plays a role in tumour metastasis and patient outcome. The aim of this study was to determine whether 14-3-3ε is a marker for predicting hepatocellular carcinoma (HCC) metastasis and survival. METHODS AND RESULTS: One hundred and fourteen patients with tissue-diagnosed primary HCC were followed for an average of 58.6 months. 14-3-3ε in liver tissues was analysed by immunohistochemistry, and quantified by a Quick score system. Correlation of 14-3-3ε with patient survival and metastasis was analysed with a Wilcoxon signed rank test, Kaplan-Meier survival curves, and Cox proportional hazard regression. Seventy-one of 114 patients (62.3%) had a significant increase of 14-3-3ε expression in HCC tissues, whereas normal tissues expressed weak or undetectable 14-3-3ε. Elevated 14-3-3ε expression was significantly associated with shortened overall survival and progression-free survival. Furthermore, 14-3-3ε overexpression increased the risk of metastasis 4.6-fold. CONCLUSIONS: Overexpression of 14-3-3ε in primary HCC tissues predicts a high risk of extrahepatic metastasis and worse survival, and is a potential therapeutic target.
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Proteínas 14-3-3/metabolismo , Carcinoma Hepatocelular/mortalidad , Carcinoma Hepatocelular/secundario , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/patología , Biomarcadores de Tumor/análisis , Carcinoma Hepatocelular/metabolismo , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Neoplasias Hepáticas/metabolismo , Estadificación de Neoplasias , PronósticoRESUMEN
5-methoxytryptophan (5-MTP) is a recently discovered tryptophan (Trp) metabolite with anti-inflammatory and tumor-suppressing actions. Its synthesis is catalyzed by hydroxyindole O-methyltransferase (HIOMT). HIOMT levels were reported to be decreased in some patients with colorectal, pancreatic and breast cancer. It is unclear whether tissue HIOMT levels is altered in hepatocellular carcinoma (HCC). It is also unclear whether serum 5-MTP concentration is influenced by HCC. In this study, 150 HCC and adjacent normal liver tissues and serum samples were obtained from the HCC biobank established by a prospective multicenter study. Serum samples from 47 healthy subjects were included as a reference. HIOMT mRNA was measured by real time PCR. Serum 5-MTP and selected Trp metabolites were analyzed by quantitative LC-MS. HCC tissue HIOMT mRNA levels adjusted for adjacent normal tissue HIOMT mRNA levels was associated with overall and relapse-free (RF) survival. Combined serum 5-MTP or tissue HIOMT mRNA and serum kynurenine (Kyn) analysis predicted prolonged overall and RF survival following liver resection. A high serum 5-MTP or tissue HIOMT mRNA and low serum Kyn is associated with long-term survival. In conclusion, tumor tissue HIOMT mRNA and serum 5-MTP are potential biomarkers of HCC, especially when analyzed in combination with serum Kyn.
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Chronic kidney disease (CKD) is a commonly occurring complex renal syndrome that causes overall mortality in many diseases. The clinical manifestations of CKD include renal tubulointerstitial fibrosis and loss of renal function. Metallothionein-I/II (MT-I/II) is potentially expressed in the liver and kidney, and possesses antioxidant and metal detoxification properties. However, whether MT-I/II expression is associated with the prognosis of nephropathy remains unknown. In this study, we investigated the MT-I/II level in human CKD, using immunohistochemistry. MT-I/II is located on the proximal tubules and is notably reduced in patients with CKD. MT-I/II expression was significantly correlated with the functional and histological grades of CKD. In an aristolochic acid (AAI)-induced nephropathy mouse model, MT-I/II was abundantly increased after AAI injection for 7 days, but decreased subsequently compared to that induced in the acute phase when injected with AAI for 28 days. Furthermore, we found that ammonium pyrrolidinedithiocarbamate (PDTC) restored AAI-induced MT-I/II reduction in HK2 cells. The injection of PDTC ameliorated AAI-induced renal tubulointerstitial fibrosis and reduced the concentrations of blood urea nitrogen and creatinine in mouse sera. Taken together, our results indicate that MT-I/II reduction is associated with advanced CKD, and the retention of renal MT-I/II is a potential therapeutic strategy for CKD.
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Túbulos Renales Proximales/metabolismo , Túbulos Renales Proximales/fisiopatología , Metalotioneína/efectos adversos , Metalotioneína/metabolismo , Insuficiencia Renal Crónica/inducido químicamente , Insuficiencia Renal Crónica/metabolismo , Insuficiencia Renal Crónica/fisiopatología , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
OBJECTIVES: The impact of viral eradication on hepatic angiogenesis is unknown. This study aimed to analyze the correlations of liver angiogenesis with liver fibrosis progression or regression in chronic hepatitis C (CHC) after viral eradication. METHODS: From 2003 to 2020, a cohort of 130 eligible participants underwent paired percutaneous liver biopsies (median = 48 months apart; range = 46-62) at the treatment baseline and after sustained virological response to CHC treatment at the tertiary referral center. The collagen proportionate area (CPA) of liver tissue sections was determined using picrosirius red staining through digital image analysis. CD34 and α-smooth muscle actin (α-SMA) phenotypically quantitated liver angiogenesis and myofibroblasts, respectively, through immunohistochemistry staining, to correlate the total, portal, and extraportal liver angiogenesis with fibrogenesis. RESULTS: Paired histology manifested significant regressions in fibrosis stages, and necroinflammatory grades (both P < 0.001). The median of changes in CPAs (follow-up minus baseline) was -6.12% (interquartile range = -12.35 to -2.05%). The median of CPA changes per year was -1.38%/year (interquartile range = -2.98 to -0.51%/year). The significance of declines in total CD34 [coefficient (95% confidence interval), 5.577 (3.286-7.868); P < 0.001] outweighed α-SMA declines, when explaining (R2 = 0.522; adjusted R2 = 0.502) the CPA declines through multiple regression analysis adjusting for other histological variables. CONCLUSION: Through viral eradication in CHC, the downregulated liver angiogenesis significantly explains the CPA regression.
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Hepatitis C Crónica , Trasplante de Hígado , Regulación hacia Abajo , Hepatitis C Crónica/diagnóstico , Hepatitis C Crónica/tratamiento farmacológico , Hepatitis C Crónica/patología , Humanos , Hígado/patología , Cirrosis Hepática/diagnóstico , Cirrosis Hepática/patologíaRESUMEN
BACKGROUND: Triple progressive thermopreconditioning (3PTP) may induce high Hsp-70 expression to maintain cardiac function. We suggest that 3PTP may reduce myocardial ischemia/reperfusion (I/R) injury during organ transplantation through Bag3/Hsp-70 mediated defense mechanisms. METHODS: Male Wistar rats were divided into sham control group and 72 h after 3PTP in a 42°C water bath (3PTP) group. Rats underwent 60 min of ischemia by occlusion of the left anterior descending coronary artery followed by 240 min reperfusion. Hemodynamic parameters, including the electrocardiogram, microcirculation, heart rate, left ventricular end-diastolic pressure, maximal rate of rise (+dp/dt), and fall (-dp/dt) in the left ventricular pressure for index of contraction and relaxation were determined. Myocardial infarct size was evaluated by the Evans blue-2,3,5-triphenyltetrazolium chloride method. 3PTP-induced protective mechanisms were determined by Western blot and immunohistochemistry. RESULTS: Cardiac I/R depressed cardiac microcirculation, induced S-T segment elevation, and R-R and P-R interval elongation increased infarct size associated with erythrocyte extravasation, leukocytes and macrophage/monocyte infiltration, granulocyte colony-stimulating factor, poly(ADP-ribose) polymerase 1 stain, and transferase-mediated dUTP-biotin nick end labeling positive cells. However, 3PTP evoked significant cardioprotection against I/R injury, characterized by the increased +dp/dt value and the decreased elevated left ventricular end-diastolic pressure, erythrocyte extravasation, leukocyte and macrophage/monocyte infiltration, granulocyte colony-stimulating factor expression, poly(ADP-ribose) polymerase 1 expression, transferase-mediated dUTP-biotin nick end labeling positive cells, and fragmentation and infarct area. In addition, 3PTP increased Hsp-70 and Bag3 expression and decreased Bax/Bcl-2 ratio, but did not affect the Beclin-1 and LC3-II/LC3-I ratio in the heart with I/R injury. CONCLUSIONS: 3PTP therapies may through Bag3 upregulation alleviate I/R injury-induced left ventricular structural deterioration and dysfunction.
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Ventrículos Cardíacos/patología , Precondicionamiento Isquémico Miocárdico , Contracción Miocárdica/fisiología , Daño por Reperfusión Miocárdica/fisiopatología , Disfunción Ventricular Izquierda/prevención & control , Proteínas Adaptadoras Transductoras de Señales/fisiología , Animales , Apoptosis , Proteínas Reguladoras de la Apoptosis/fisiología , Factor Estimulante de Colonias de Granulocitos/farmacología , Masculino , Microcirculación , Daño por Reperfusión Miocárdica/patología , Proteínas Proto-Oncogénicas c-bcl-2/análisis , Ratas , Ratas WistarRESUMEN
Embryonic stem (ES) cells are an important source of stem cells in tissue engineering and regenerative medicine because of their high self-renewal capacities and differentiation potentials. However, the detailed molecular mechanisms controlling the differentiation and renewal programs in ES cells remained unclear. One of the difficulties in understanding these mechanisms substantially results from the low efficacies of gene manipulation by delivering exogenous gene expression or knockdown of endogenous gene expression with small interfering RNA (siRNA) in ES cells. Here we describe an optimized protocol for efficiently transfecting mouse ES cells by Effectene, a liposome-based method. The high transfection efficiency in mouse ES cells is demonstrated in this chapter by (1) achieving a percentage of enhanced green fluorescence protein (EGFP) expression in >98% embryoid bodies after introducing plasmids encoding the protein; (2) decreased SOX-2 and Oct-3/4 expression and subsequent morphological evidences of cell differentiation after introducing siRNA expression for suppressing SOX-2 and Oct-3/4, which are known to be essential for maintenance of stem cell properties in mouse ES cells; and (3) overexpression or attenuated expression of 14-3-3σ to regulate cell proliferation of mouse ES cells.
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Diferenciación Celular , Células Madre Embrionarias de Ratones/citología , Células Madre Embrionarias de Ratones/metabolismo , Transfección , Animales , Diferenciación Celular/genética , Línea Celular , Proliferación Celular , Expresión Génica , Genes Reporteros , Ratones , Plásmidos/genética , ARN Interferente Pequeño/genética , Transfección/métodosRESUMEN
5-methoxytryptophan (5-MTP) is a newly discovered tryptophan metabolite which controls stress-induced inflammatory signals. To determine whether 5-MTP protects against stress-induced mesenchymal stem cell (MSC) senescence, we incubated bone marrow-derived MSC (BM-MSC) in high-glucose medium or regular medium for 2 weeks followed by addiction of 5-MTP (10 µM) or vehicle for 48 h. 5-MTP reduced p16 and p21 expression, senescence-associated ß-Gal (SA-ß-Gal) and IL-6 secretion and increased BrdU incorporation. 5-MTP exerted a similar effect on BM-MSC senescence induced by a sublethal concentration of H2O2. 5-MTP enhanced FoxO3a expression and increased superoxide dismutase and catalase activities in HG BM-MSCs. Silencing of FoxO3a with siRNA abrogated 5-MTP-mediated reduction of SA-ß-Gal and IL-6 secretion but not p21 or p16. Since mechanistic target of rapamycin (mTOR) is involved in cellular senescence, we determined whether 5-MTP influences mTOR expression. Our data reveal that mTOR protein level was depressed in HG-MSC which was rescued by 5-MTP. Rapamycin abrogated 5-MTP-mediated suppression of p16, p21, SA-ß-Gal and IL-6 and rise of BrdU incorporation. Our findings suggest that 5-MTP protects MSCs against stress-induced senescence via FoxO3a and mTOR upregulation and has potential to improve cell expansion for cell therapy.
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Proteína Forkhead Box O3/metabolismo , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Sustancias Protectoras/farmacología , Estrés Fisiológico , Serina-Treonina Quinasas TOR/metabolismo , Triptófano/análogos & derivados , Adipogénesis , Diferenciación Celular , Proliferación Celular/efectos de los fármacos , Senescencia Celular/efectos de los fármacos , Glucosa/metabolismo , Glucosa/farmacología , Humanos , Peróxido de Hidrógeno/metabolismo , Células Madre Mesenquimatosas/citología , Osteogénesis , Transducción de Señal/efectos de los fármacos , Estrés Fisiológico/efectos de los fármacos , Triptófano/farmacologíaRESUMEN
Cisplatin-induced nephrotoxicity leaded to apoptosis of tubular epithelial cells (ECs) and tubulointerstitial fibrosis through ROS stress and inflammatory cytokines. Tubulointerstitial fibrosis caused by cisplatin might be via activation of resident fibroblasts and epithelial-mesenchymal transition (EMT) of tubular ECs. Inflammatory niche was crucial for progression of fibroblast activation or EMT. It had been reported that M1/M2 macrophage polarization regulated pro-inflammation or pro-resolving phase in damage repairing. However, the role of macrophage polarization on cisplatin-induced EMT of tubular ECs had not been well elucidated. In this study, we used co-cultured cell model and condition medium to examine the interaction between tubular ECs, fibroblasts and M1/M2 macrophages. Our data showed that cisplatin alone induced incomplete EMT of tubular ECs, whereas fibroblasts co-cultured with cisplatin-treated ECs could lead to fibroblast activation by detection of α-SMA and collagen-1. Moreover, decrease of iNOS and increase of argenase-1 and CD206 expression indicated that macrophages co-cultured with cisplatin-treated ECs would turn to M2 phenotype. Finally, we found that condition medium of M2 macrophages could promote complete EMT of cisplatin-treated ECs. Taken together, cisplatin created an inflammatory niche via tubular ECs to activate fibroblasts and stimulated M2 macrophage polarization. M2 macrophages could turn back to promote EMT of cisplatin-treated ECs. These results revealed the cooperative roles of tubular ECs, fibroblast and M2 macrophages to facilitate the progression of renal fibroblasis.
RESUMEN
14-3-3σ overexpression results in enhanced hepatocellular carcinoma (HCC) cell migration and HCC tumor vascular-invasion is significantly associated with 14-3-3σ expression. However, increased expression of 14-3-3σ paradoxically suppresses in vitro cell invasion of HCC. We hypothesize that surrounding tumor-associated stromal cells play a crucial role in 14-3-3σ-regulated HCC cell invasion. In this study, H68 fibroblasts, THP-1 and phorbol-12-myristate-13-acetate (PMA)-treated THP-1 (PMA-THP-1) cells were incubated with conditioned media of control (control-CM) and 14-3-3σ-overepxressing cells (14-3-3σ-CM), followed by co-culture with HCC cells. Invasiveness of HCC cells was examined by a Boyden chamber assay. HCC cells co-cultured with 14-3-3σ-CM treated cells significantly enhanced their invasive ability compared with control-CM treated cells. Moreover, incubation with 14-3-3σ-CM induced differential expression profiles of matrix metalloproteinases (MMPs) in fibroblasts (MMP-1, MMP-2, MMP-9, MMP-12 and MMP-14), THP-1 (MMP-1 and MMP-12) and PMA-THP-1 cells (MMP-2, MMP-12 and MMP-14). In contrast, silencing of 14-3-3σ by siRNA significantly abolished 14-3-3σ-CM induced MMPs. In addition, treatment with recombinant 14-3-3σ (r14-3-3σ) protein exhibits a similar expression profile of MMPs induced by 14-3-3σ-CM in fibroblasts, THP-1 and PMA-THP-1 cells. Finally, knockdown of aminopeptidase N (APN) significantly abrogated r14-3-3σ induced expression of MMPs in HS68 fibroblasts. These results suggest that HCC-secreted 14-3-3σ promotes expression of MMPs in cancerous surrounding cells via an APN dependent mechanism. 14-3-3σ has a paracrine effect in educating stromal cells in tumor-associated microenvironment.