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Interior tomography by rotary computed tomography (RCT) is an effective method to improve the detection efficiency and achieve high-resolution imaging for the region of interest (ROI) within a large-scale object. However, because only the X-rays through the ROI can be received by detector, the projection data is inevitably truncated, resulting in truncation artifacts in the reconstructed image. When the ROI is totally within the object, the solution of the problem is not unique, which is named interior problem. Fortunately, projection completion (PC) is an effective technique to solve the interior problem. In this study, we proposed a multi source translation CT based PC method (mSTCT-PC) to cope with the interior problem. Firstly, mSTCT-PC employs multi-source translation to sparsely obtain the global projection which covered the whole object. Secondly, the sparse global projection is utilized to fill up the truncated projection of ROI. The global projection and truncated projection are obtained under the same geometric parameters. Therefore, it omits the registration of projection. To verify the feasibility of this method, simulation and practical experiments were implemented. Compared with the results of ROI reconstructed by filtered back-projection (FBP), simultaneous iterative reconstruction technique-total variation (SIRT-TV) and the multi-resolution based method (mR-PC), the proposed mSTCT-PC is good at mitigating truncation artifacts, preserving details and improving the accuracy of ROI images.
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Although osimertinib, an EGFR tyrosine kinase inhibitor, has become the standard therapy for treating non-small cell lung cancer (NSCLC) patients with EGFR-activating mutation, upregulation of MCL-1 induces acquired resistance to osimertinib. Bufalin, a natural digoxin-like ingredient isolated from a traditional Chinese medicine Chan Su, has been shown to downregulate MCL-1 in NSCLC cells. However, whether bufalin reverses this acquired resistance to osimertinib in NSCLC cells remains unclear. In this study, bufalin reduced cell viability and promoted apoptosis in osimertinib-resistant cells. Moreover, co-treatment with bufalin and osimertinib restored the sensitivity of osimertinib-resistant cells to osimertinib-induced growth regression and apoptosis in vitro and in vivo. Mechanistically, MEK/ERK-dependent MCL-1 phosphorylation and Ku70-mediated MCL-1 overexpression confer osimertinib resistance in EGFR-mutant NSCLC cells. In osimertinib-resistant cells, bufalin modulates Ku70-mediated MCL-1 degradation, but not MEK/ERK/MCL-1 signaling. In conclusion, our study suggests that bufalin eliminates resistance to osimertinib by inhibiting Ku70-mediated MCL-1 overexpression, indicating that a combination of osimertinib and bufalin could be an effective additional treatment to overcome acquired resistance to osimertinib in NSCLC cells.
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Acrilamidas/uso terapéutico , Compuestos de Anilina/uso terapéutico , Antineoplásicos/uso terapéutico , Bufanólidos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Genes erbB-1/genética , Neoplasias Pulmonares/tratamiento farmacológico , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Animales , Bufanólidos/uso terapéutico , Carcinoma de Pulmón de Células no Pequeñas/genética , Resistencia a Antineoplásicos/efectos de los fármacos , Técnica del Anticuerpo Fluorescente , Etiquetado Corte-Fin in Situ , Neoplasias Pulmonares/genética , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Trasplante de NeoplasiasRESUMEN
BACKGROUND: Biallelic variants in OGDHL, encoding part of the α-ketoglutarate dehydrogenase complex, have been associated with highly heterogeneous neurological and neurodevelopmental disorders. However, the validity of this association remains to be confirmed. A second OGDHL patient cohort was recruited to carefully assess the gene-disease relationship. METHODS: Using an unbiased genotype-first approach, we screened large, multiethnic aggregated sequencing datasets worldwide for biallelic OGDHL variants. We used CRISPR/Cas9 to generate zebrafish knockouts of ogdhl, ogdh paralogs, and dhtkd1 to investigate functional relationships and impact during development. Functional complementation with patient variant transcripts was conducted to systematically assess protein functionality as a readout for pathogenicity. RESULTS: A cohort of 14 individuals from 12 unrelated families exhibited highly variable clinical phenotypes, with the majority of them presenting at least one additional variant, potentially accounting for a blended phenotype and complicating phenotypic understanding. We also uncovered extreme clinical heterogeneity and high allele frequencies, occasionally incompatible with a fully penetrant recessive disorder. Human cDNA of previously described and new variants were tested in an ogdhl zebrafish knockout model, adding functional evidence for variant reclassification. We disclosed evidence of hypomorphic alleles as well as a loss-of-function variant without deleterious effects in zebrafish variant testing also showing discordant familial segregation, challenging the relationship of OGDHL as a conventional Mendelian gene. Going further, we uncovered evidence for a complex compensatory relationship among OGDH, OGDHL, and DHTKD1 isoenzymes that are associated with neurodevelopmental disorders and exhibit complex transcriptional compensation patterns with partial functional redundancy. CONCLUSIONS: Based on the results of genetic, clinical, and functional studies, we formed three hypotheses in which to frame observations: biallelic OGDHL variants lead to a highly variable monogenic disorder, variants in OGDHL are following a complex pattern of inheritance, or they may not be causative at all. Our study further highlights the continuing challenges of assessing the validity of reported disease-gene associations and effects of variants identified in these genes. This is particularly more complicated in making genetic diagnoses based on identification of variants in genes presenting a highly heterogenous phenotype such as "OGDHL-related disorders".
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Proteínas , Pez Cebra , Animales , Humanos , Frecuencia de los Genes , Complejo Cetoglutarato Deshidrogenasa/genética , Complejo Cetoglutarato Deshidrogenasa/metabolismo , Fenotipo , Proteínas/genética , Pez Cebra/genéticaRESUMEN
FINCA syndrome [MIM: 618278] is an autosomal recessive multisystem disorder characterized by fibrosis, neurodegeneration and cerebral angiomatosis. To date, 13 patients from nine families with biallelic NHLRC2 variants have been published. In all of them, the recurrent missense variant p.(Asp148Tyr) was detected on at least one allele. Common manifestations included lung or muscle fibrosis, respiratory distress, developmental delay, neuromuscular symptoms and seizures often followed by early death due to rapid disease progression.Here, we present 15 individuals from 12 families with an overlapping phenotype associated with nine novel NHLRC2 variants identified by exome analysis. All patients described here presented with moderate to severe global developmental delay and variable disease progression. Seizures, truncal hypotonia and movement disorders were frequently observed. Notably, we also present the first eight cases in which the recurrent p.(Asp148Tyr) variant was not detected in either homozygous or compound heterozygous state.We cloned and expressed all novel and most previously published non-truncating variants in HEK293-cells. From the results of these functional studies, we propose a potential genotype-phenotype correlation, with a greater reduction in protein expression being associated with a more severe phenotype.Taken together, our findings broaden the known phenotypic and molecular spectrum and emphasize that NHLRC2-related disease should be considered in patients presenting with intellectual disability, movement disorders, neuroregression and epilepsy with or without pulmonary involvement.
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Discapacidad Intelectual , Trastornos del Movimiento , Humanos , Progresión de la Enfermedad , Fibrosis , Células HEK293 , Discapacidad Intelectual/genética , Fenotipo , Convulsiones/genética , SíndromeRESUMEN
The LHR has an essential role in sexual development and reproductive function, and its transcription is subjected to several modes of regulation. In this study, we investigated PC4 coactivator function in the control of LHR transcription. Knockdown of PC4 by siRNA inhibited the LHR basal promoter activity and trichostatin A (TSA)-induced gene transcriptional activation and expression in MCF-7 cells. While overexpression of PC4 alone had no effect on the LHR gene, it significantly enhanced Sp1- but not Sp3-mediated LHR transcriptional activity. PC4 directly interacts with Sp1 at the LHR promoter, and this interaction is negatively regulated by PC4 phosphorylation. The coactivator domain (22-91 aa) of PC4 and DNA binding domain of Sp1 are essential for PC4/Sp1 interaction. ChIP assay revealed significant occupancy of PC4 at the LHR promoter that increased upon TSA treatment. Disruption of PC4 expression significantly reduced TSA-induced recruitment of TFIIB and RNAP II, at the promoter. PC4 functions are beyond TSA-induced phosphatase release, PI3K-mediated Sp1 phosphorylation, and HDAC1/2/mSin3A co-repressor release indicating its role as linker coactivator of Sp1 and the transcriptional machinery. These findings demonstrated a critical aspect of LHR modulation whereby PC4 acts as a coactivator of Sp1 to contribute to the human of LHR transcription.
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Proteínas de Unión al ADN/metabolismo , Receptores de HL/genética , Factor de Transcripción Sp1/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética/fisiología , Neoplasias de la Mama , Línea Celular Tumoral , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Femenino , Humanos , Ácidos Hidroxámicos/farmacología , Regiones Promotoras Genéticas/fisiología , Estructura Terciaria de Proteína , Inhibidores de la Síntesis de la Proteína/farmacología , ARN Polimerasa II/metabolismo , ARN Interferente Pequeño , Factor de Transcripción Sp1/química , Factor de Transcripción Sp1/genética , Factor de Transcripción TFIIB/metabolismo , Factores de Transcripción/química , Factores de Transcripción/genética , Transcripción Genética/efectos de los fármacosRESUMEN
BACKGROUND: In previous studies on the application of cyclin-dependent kinase 4/6 (CDK4/6) inhibitors combined with endocrine therapy in advanced breast cancer, the outcomes of overall survival (OS) were inconsistent. This systematic review and meta-analysis aimed to further evaluate the clinical efficacy and safety of CDK4/6 inhibitors combined with endocrine therapy on patients with hormone receptor (HR)-positive and human epidermal growth factor receptor 2 (HER2)-negative advanced breast cancer. METHODS: Randomized controlled trials (RCTs) comparing CDK4/6 inhibitors plus endocrine therapy and endocrine therapy alone in patients with HR-positive and HER2-negative advanced breast cancer were searched in the databases of PubMed, Embase, Cochrane Library, China National Knowledge Infrastructure (CNKI), WANFANG and China Science and Technology Journal Database (VIP) up to November 2022. Hazard ratios (HRs) and confidence intervals (CI) of OS, progression-free survival (PFS), the time from randomization to the first recorded disease progression while the patient was receiving next-line therapy or death from any cause (PFS2), time to first subsequent chemotherapy after discontinuation (TTC), and objective response rate (ORR), clinical benefit rate (CBR), safety indicators were extracted. Stata 14.0 software was used for meta analysis and the Cochrane risk-of-bias tool 2.0 was used to evaluate the bias risk. RESULTS: A total of 9 RCTs with 4,920 participants were included. The addition of CDK4/6 inhibitors to endocrine therapy significantly prolonged OS (HR 0.76; 95% CI: 0.69-0.84; P<0.001), regardless of the application in first-line and second-line treatment, compared with endocrine therapy alone. Similar benefit was observed in PFS (HR 0.56; 95% CI: 0.52-0.60; P<0.001). Moreover, the CDK4/6 inhibitors group improved results of ORR [relative risk (RR) 1.43; 95% CI: 1.27-1.62; P<0.001], CBR (RR 1.24; 95% CI: 1.08-1.41; P<0.01 and RR 1.11; 95% CI: 1.06-1.18; P<0.001), PFS2 (HR 0.68; 95% CI: 0.60-0.76; P<0.001) and TTC (HR 0.65; 95% CI: 0.58-0.72; P<0.001). One of the included RCTs had performance bias. Publication bias was not significant. CONCLUSIONS: CDK4/6 inhibitors combined with endocrine therapy effectively prolong OS, PFS, PFS2, and TTC, and also improve ORR and CBR in patients with HR-positive, HER2-negative advanced breast cancer, and the safety was within the controllable range.
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Neoplasias de la Mama , Quinasa 4 Dependiente de la Ciclina , Quinasa 6 Dependiente de la Ciclina , Inhibidores de Proteínas Quinasas , Femenino , Humanos , Neoplasias de la Mama/tratamiento farmacológico , Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 6 Dependiente de la Ciclina/antagonistas & inhibidores , Supervivencia sin Progresión , Resultado del Tratamiento , Inhibidores de Proteínas Quinasas/uso terapéuticoRESUMEN
Non-alcoholic fatty liver disease (NAFLD) has been considered to be one of the most common chronic liver diseases. However, no validated pharmacological therapies have been officially proved in clinic due to its complex pathogenesis. The purpose of this study was to examine the protective effects of Corilagin (referred to Cori) against NAFLD in mice under a high fat diet (HFD) condition. Mice were fed either a normal control diet (NCD) or HFD with or without Cori (5 or 10 mg/kg body weight) for 15 weeks. In our results, Cori treatment significantly attenuated HFD-induced hepatic steatosis, high NAFLD activity score (NAD) and liver injury. Consistently, Cori treatment remarkably alleviated HFD-induced hepatic lipid accumulation (e.g., triglycerides (TG) and total cholesterol (TC) contents in liver), and improved plasma lipid concentrations (e.g., plasma TG, TC, low-density lipoprotein cholesterol (LDL-c), high-density lipoprotein cholesterol (HDL-c)). Moreover, Cori treatment ameliorated NAFLD associated metabolic disorders such as glucose intolerance and insulin resistance in HFD-fed mice. Additionally, Cori treatment dramatically changed HFD-induced liver gene expression profiles, and identified overlapped differentially expressed genes (DEGs) between NCD vs. HFD group and HFD vs. HCR (high fat diet plus treatment with Cori) group. With these DEGs, we observed a marked enrichment of Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, which were closely associated with the metabolic balance in liver. Particularly, we found several potential hub proteins against NAFLD development with analyses of protein-protein interaction (PPI) network and qPCR assays. Collectively, our results revealed the important protective effects of Cori against the progress of NAFLD, which was probably mediated through improving dysregulated lipid metabolism and insulin resistance in HFD-fed mice. Additionally, Cori-dependent overlapped DEGs might serve as a featured NAFLD-associated gene expression signature for the diagnosis, treatment, as well as drug discovery and development of NAFLD in the near future.
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OBJECTIVE: To investigate the effects of Astragalus polysaccharide (APS) on proliferation of basal-like breast cancer cell line MDA-MB-468 cells and Akt phosphorylation in MDA-MB-468 cells. METHODS: APS at different concentrations was used to culture MDA-MB-468 cells for different time periods, and then proliferation of MDA-MB-468 cells was assayed using methyl thiazolyl tetrazolium (MTT) assay to determine the time- and dose-dependent effects of APS. For observing the effects of APS on phosphor-Akt (p-Akt), in-cell Western blot method was used after 1, 2, 4 and 7 d of culture in APS to detect protein expressions of p-Akt (Thr308) and p-Akt (Ser473). Protein levels of the key targets in p53/murine double minute 2 (MDM2) signaling pathway, such as p53, MDM2 and phosphatase and tensin homolog deleted on chromosome ten (PTEN) were also detected. After PTEN gene was silenced by small interfering RNA (siRNA) in MDA-MB-468 cells, expressions of p-Akt (Thr308 and Ser473) were assayed by the in-cell Western blot method after 2 d of APS treatment. RESULTS: APS at 1 and 0.5 mg/mL concentrations effectively inhibited the proliferation of MDA-MB-468 cells and was used in subsequent tests. Compared with the control group, APS decreased the protein expression of p-Akt (Thr308) in MDA-MB-468 cells after 1-, 2-, 4- and 7-day culture, and also decreased the protein expression of p-Akt (Ser473) and up-regulated the protein expression of MDM2 in MDA-MB-468 cells after 1- and 2-day culture. Expressions of p53 and PTEN were up-regulated after 7 d of APS culture. After silencing PTEN gene by siRNA, APS could not mediate Akt phosphorylation. CONCLUSION: APS can inhibit proliferation of basal-like breast cancer cell line MDA-MB-468, and down-regulate the expression of Akt phosphorylation. The antiproliferation mechanisms may be related to its effects of up-regulating the expressions of p53 and PTEN by regulating p53/MDM2 positive and negative feedback loops.
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Astragalus propinquus/química , Neoplasias de la Mama/patología , Proliferación Celular/efectos de los fármacos , Polisacáridos/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Antineoplásicos Fitogénicos/farmacología , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Regulación hacia Abajo , Femenino , Humanos , Fosfohidrolasa PTEN/metabolismo , Fosforilación/efectos de los fármacos , Polisacáridos/aislamiento & purificación , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
This investigation attempted to discern whether formononetin restrained progression of triple-negative breast cancer (TNBC) by blocking lncRNA AFAP1-AS1-miR-195/miR-545 axis. We prepared TNBC cell lines (i.e. MDA-MB-231 and BT-549) and normal human mammary epithelial cell line (i.e. MCF-10A) in advance, and the TNBC cell lines were, respectively, transfected by pcDNA3.1-lncRNA AFAP1-AS1, si-lncRNA AFAP1-AS1, pcDNA6.2/GW/EmGFP-miR-545 or pcDNA6.2/GW/EmGFP-miR-195. Resistance of TNBC cells in response to 5-Fu, adriamycin, paclitaxel and cisplatin was evaluated through MTT assay, while potentials of TNBC cells in proliferation, migration and invasion were assessed via CCK8 assay and Transwell assay. Consequently, silencing of lncRNA AFAP1-AS1 impaired chemo-resistance, proliferation, migration and invasion of TNBC cells (P<0.05), and over-expression of miR-195 and miR-545, which were sponged and down-regulated by lncRNA AFAP1-AS1 (P<0.05), significantly reversed the promoting effect of pcDNA3.1-lncRNA AFAP1-AS1 on proliferation, migration, invasion and chemo-resistance of TNBC cells (P<0.05). Furthermore, CDK4 and Raf-1, essential biomarkers of TNBC progression, were, respectively, subjected to target and down-regulation of miR-545 and miR-195 (P<0.05), and they were promoted by pcDNA3.1-lncRNA AFAP1-AS1 at protein and mRNA levels (P<0.05). Additionally, formononetin significantly decreased expressions of lncRNA AFAP1-AS1, CDK4 and Raf-1, while raised miR-195 and miR-545 expressions in TNBC cells (P<0.05), and exposure to it dramatically contained malignant behaviors of TNBC cells (P<0.05). In conclusion, formononetin alleviated TNBC malignancy by suppressing lncRNA AFAP1-AS1-miR-195/miR-545 axis, suggesting that molecular targets combined with traditional Chinese medicine could yield significant clinical benefits in TNBC.
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Resistencia a Antineoplásicos/efectos de los fármacos , Isoflavonas/farmacología , MicroARNs/metabolismo , ARN Largo no Codificante/genética , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Animales , Antineoplásicos/farmacología , Línea Celular Tumoral , Progresión de la Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Ratones Endogámicos BALB C , MicroARNs/efectos de los fármacos , MicroARNs/genética , ARN Largo no Codificante/efectos de los fármacos , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patologíaRESUMEN
BACKGROUND: Polyacrylamide hydrogel (PAAG) was used as an injectable implant for augmentation mammoplasty for over 30 years, but its use was ceased due to various related complications. The only way to treat these complications is PAAG removal, but this causes breast ptosis, nipple retraction, breast asymmetry, and skin laxity. OBJECTIVES: This article reports a new technique for breast reshaping after PAAG removal without prosthesis implantation. METHOD: From January 2015 to June 2018, twenty-three patients underwent periareolar mammoplasty with the tissue folding technique (PMTFT) for breast reshaping after PAAG removal. Postoperative breast shape and the degree of satisfaction of the patients were evaluated during follow-up. RESULTS: All patients recovered well without severe complications. All patients were satisfied with their postoperative breast shape and their symptoms were relieved after surgery. CONCLUSIONS: PMTFT provides satisfactory postoperative breast shape results. Economical, practical, and technical advantages were found over traditional prosthesis-mediated breast reconstruction. PMTFT can be an ideal surgical choice in appropriate cases.
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BACKGROUND: Chronic refractory plasma cell mastitis (CRPCM) is an aseptic inflammation of the breast with a chronic course of the disease, extended treatment cycle (months to years), with a high recurrence rate. The objective of this study was to evaluate the efficacy of continuous postoperative negative pressure irrigation assisted mammaplasty (CPNPIAM) in treating CRPCM. METHODS: Between 2016 and 2018, 36 CRPCM patients receiving CPNPIAM were enrolled in this study. CPNPIAM mainly involved complete lesion removal, immediate breast mammaplasty, and continuous postoperative negative pressure irrigation. The age of the patients, local symptoms, history of treatment, the duration of the disease before surgery, hospitalization period, related risk factors, the success rate, the recurrence rate and patients' overall satisfaction ratings were analyzed in the article. RESULTS: Patients were aged between 22 and 53 years (mean 34.64 years). All patients had a history of conservative treatment or simple drainage. Local symptoms included inflammatory mass (n=36, 100%), abscess (n=33, 92%), nipple discharge (n=7, 19%), inflammatory plaque (n=34, 94%), and sinus tract formation (n=19, 53%). The lesion sizes ranged from 3 to 10 cm (mean 5.13 cm) in diameter. The mean hospitalization period was 8.42 days. The success rate was 100% (36/36) and the recurrence rate was 0% (0/36) at a 3-month follow-up. The patients' overall satisfaction ratings were "very good" (n=22, 61%), "good" (n=12, 33%), and "moderate" (n=2, 6%) with no poor or unsatisfactory ratings. CONCLUSIONS: CPNPIAM is an effective way of treating CRPCM, and showed a high success rate, a low recurrence rate, and high patient satisfaction.
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We have demonstrated that silencing of luteinizing hormone receptor (LHR) gene transcription is mediated via a proximal Sp1 site at its promoter. Trichostatin A (TSA) induced histone acetylation and gene activation in JAR cells that prevailed in the absence of changes in Sp1/Sp3 expression, their binding activity, disassociation of the histone deacetylase/mSin3A complex from the Sp1 site, or demethylation of the promoter. This indicated a different mechanism involved in TSA-induced derepression. The present studies have revealed that phosphatidylinositol 3-kinase/protein kinase Czeta (PI3K/PKCzeta)-mediated Sp1 phosphorylation accounts for Sp1 site-dependent LHR gene activation. TSA caused marked phosphorylation of Sp1 at serine 641 in JAR and MCF-7 cells. Blockade of PI3K or PKCzeta activity by specific inhibitors, kinase-deficient mutants, or small interfering RNA abolished the effect of TSA on the LHR gene and Sp1 phosphorylation. PKCzeta was shown to associate with Sp1, and this association was enhanced by TSA. Sp1 phosphorylation at serine 641 was required for the release of the pRb homologue p107 from the LHR gene promoter, while p107 acted as a repressor of the LHR gene. Inhibition of PKCzeta activity blocked the dissociation of p107 from the LHR gene promoter and markedly reduced Sp1 phosphorylation and transcription. These results have demonstrated that phosphorylation of Sp1 by PI3K/PKCzeta is critical for TSA-activated LHR gene expression. These studies have revealed a novel mechanism of TSA action through derecruitment of a repressor from the LHR gene promoter in a PI3K/PKCzeta-induced Sp1 phosphorylation-dependent manner.
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Inhibidores de Histona Desacetilasas , Fosfatidilinositol 3-Quinasas/metabolismo , Proteína Quinasa C/metabolismo , Receptores de HL/genética , Proteína p107 Similar a la del Retinoblastoma/metabolismo , Factor de Transcripción Sp1/metabolismo , Transcripción Genética , Acetilación/efectos de los fármacos , Animales , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HeLa , Histonas/metabolismo , Humanos , Ácidos Hidroxámicos/farmacología , Células Jurkat , Modelos Genéticos , Fosforilación/efectos de los fármacos , Fosfoserina/metabolismo , Ratas , Proteínas Represoras/metabolismo , Activación Transcripcional , Células Tumorales CultivadasRESUMEN
LH receptor (LHR) gene transcription is subject to repression/derepression through various modes and multiple effectors. Epigenetic silencing and activation of the LHR is achieved through coordinated regulation at both histone and DNA levels. The LHR gene is subject to repression by deacetylation and methylation at its promoter region, where a HDAC/mSin3A repressor complex is anchored at Sp1 sites. The present studies revealed that protein kinase C (PKC) alpha/ERK signaling is important for the activation of LHR promoter activity, and the increase of endogenous transcripts induced by phorbol-12-myristate-13-acetate (PMA) in HeLa cells. Whereas these effects were attributable to PKCalpha activity, the ERK pathway was the downstream effector in LHR activation. PMA caused a significant enhancement of Sp1 phosphorylation at serine residue (s), which was blocked by PKCalpha or ERK inhibition. The interaction of activated phosphorylated ERK with Sp1 and ERK's association with the LHR promoter points to Sp1 as a direct target of ERK. After Sp1 phosphorylation, the HDAC1/mSin3A repressor complex dissociated from Sp1 sites, histone 3 was acetylated, and transcription factor II B and RNA polymerase II were recruited. In addition, overexpression of a constitutively active PKCalpha (PKCalpha CA) strongly activated LHR transcription in MCF-7 cells (devoid of PKCalpha), induced Sp1 phosphorylation at serine residue (s) and caused derecruitment of HDAC1/mSin3A complex from the promoter. These effects were negated by cotransfection of a dominant-negative PKCalpha. In conclusion, these studies have revealed a novel regulatory signaling mechanism of transcriptional control in which the LHR is derepressed through PKCalpha/ERK-mediated Sp1 phosphorylation, causing the release of HDAC1/mSin3A complex from the promoter.
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Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Regulación de la Expresión Génica , Histona Desacetilasas/metabolismo , Proteína Quinasa C-alfa/fisiología , Receptores de HL/genética , Proteínas Represoras/metabolismo , Sitios de Unión , Células HeLa , Histona Desacetilasa 1 , Humanos , Complejos Multiproteicos/metabolismo , Fosforilación , Regiones Promotoras Genéticas/efectos de los fármacos , Unión Proteica , Proteína Quinasa C-alfa/metabolismo , Complejo Correpresor Histona Desacetilasa y Sin3 , Factor de Transcripción Sp1/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Transcripción Genética/efectos de los fármacos , Células Tumorales CultivadasRESUMEN
Corilagin (Cori) possesses multiple biological activities. To determine whether Cori can exert protective effects against nonalcoholic fatty liver disease (NAFLD) and its potential mechanisms. C57BL/6 mice were fed with high-fat diet (HFD) alone or in combination with Cori (20 mg/kg, i.p.) and AML12 cells were exposed to 200 µM PA/OA with or without Cori (10 µM or 20 µM). Phenotypes and key indicators relevant to NAFLD were examined both in vivo and in vitro. In this study, Cori significantly ameliorated hepatic steatosis, confirmed by improved serum lipid profiles, and hepatic TC, TG contents, and the gene expression related to lipid metabolism in livers of HFD mice. Moreover, Cori attenuated HFD-mediated autophagy (including mitophagy) blockage by restoring autophagic flux, evidenced by increased number of autophagic double vesicles containing mitochondria, elevated LC3II protein levels, decreased p62 protein levels, as well as enhanced colocalization of autophagy-related protein (LC3, Parkin) and mitochondria. In accordance with this, Cori also reduced the accumulation of ROS and MDA levels, and enhanced the activities of antioxidative enzymes including SOD, GSH-Px, and CAT. In addition, Cori treatment improved mitochondrial dysfunction, evidenced by increased mitochondrial membrane potential (ΔΨm). In parallel with this, Cori decreased mitochondrial DNA oxidative damage, while increased mitochondrial biogenesis related transcription factors expression, mitochondrial DNA content and oxygen consumption rate (OCR). In conclusion, these results demonstrate that Cori is a potential candidate for the treatment of NAFLD via diminishing oxidative stress, restoring autophagic flux, as well as improving mitochondrial functions.
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Growth hormone is one of the most important hormones, which is involved in many reproductive processes of giant panda Ailuropoda melanoleuca. In this study, the mature peptide of A. melanoleuca growth hormone (AmGH) was successfully expressed and secreted in Pichia pastoris under the control of AOX1 promoter. The expression condition for AmGH in P. pastoris, such as the expression time, pH value and methanol concentration in the BMMY were optimized and the AmGH expression level is about 100 mg/L using GS115 recombinant under optimized condition (96 h of 1.5% methanol induction). The secreted nascent AmGH were purified using ammonium sulfate fractionation. The mature AmGH protein exhibited a molecular mass of approximately 22 kDa on SDS-PAGE. This study would provide a new opportunity for large-scale expression and purification of AmGH, which might facilitate studies on the biological activity of AmGH.
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Hormona del Crecimiento/aislamiento & purificación , Animales , Secuencia de Bases , Western Blotting , Cartilla de ADN , Electroforesis en Gel de Poliacrilamida , Hormona del Crecimiento/química , Hormona del Crecimiento/genética , Pichia/genética , Reacción en Cadena de la Polimerasa , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , UrsidaeRESUMEN
OBJECTIVE: To reveal the characteristics of gene expression in adrenal gland of H22 tumor mice with typical syndromes and in different liver cancer stages. METHODS: By the quantitative four diagnosis and syndrome differentiation methods and GeneChip Mouse Exon 1.0 ST Array, we observed adrenal gland gene expression in H22 tumor mice with pathogenic factor-toxin predominance syndrome and qi deficiency syndrome in the earlier stage, yang-qi deficiency syndrome in the intermediate stage, and qi-yin-yang deficiency syndrome in the advanced stage. Genes highly expressed and remarkably different were analyzed in this study. RESULTS: A total of seventy-three up-regulated coincident genes and twenty-six down-regulated coincident genes in different stages were investigated in the study. Up-regulated coincident genes included Hp, C3, Anxa1, Procr, C2, Il4ra, Cd14, Ptprc, Cd52, C4b, Eno3, Xdh, Gpx3, and so on. Down-regulated coincident genes included nervous system function-related genes such as Plp1, Mbp, Aldh1a1, Cck, Atn1, genes associated with electrolyte metabolism such as Aldh1a1 and Slc22a17, genes related to signal transduction such as Cxcr4, Spag5 and Stmn3, etc, and genes related to transcriptional control and protein biosynthesis such as Hspa1a, Dnajb1, Thra, Hhex and so on. CONCLUSION: With the development of the tumorigenesis, the symptoms and signs and differentially expressed genes in adrenal gland of H22 tumor mice can be measured. Up-regulated and down-regulated coincident genes may be the features of H22 tumor mice different from those of normal mice.
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Glándulas Suprarrenales/metabolismo , Diagnóstico Diferencial , Perfilación de la Expresión Génica , Neoplasias Hepáticas Experimentales/genética , Medicina Tradicional China , Animales , Regulación Neoplásica de la Expresión Génica , Masculino , Ratones , Análisis de Secuencia por Matrices de Oligonucleótidos , Distribución Aleatoria , SíndromeRESUMEN
Curcumin is a wellknown phenolic substance and has many pharmacological effects associated with metabolism. However, the exact molecular mechanisms underlying this process have yet to be determined. The Notch pathway is a signal transduction pathway involved in energy metabolism. The present study aimed to investigate the effects of curcumin administration on glucoselipid metabolism in rats subjected to a high fat diet, and investigate changes in Notch1 signaling. SpragueDawley rats (n=40) were randomly divided into four groups (10 rats/group): Control diet group, high fat diet group, high fat diet plus curcumin low dose group and high fat diet plus curcumin high dose group. Following 8 weeks of treatment with curcumin (100 mg/kg in the low dose group and 200 mg/kg in the high dose group), serum metabolic markers and hepatic gene expression patterns were investigated. No differences in body weight following 8 weeks of curcumin administration (P>0.05) were observed; however, curcumin treatment did reduce visceral fat levels (periepididymal and perirenal), and decreased cholesterol, triglyceride and lowdensity lipoprotein levels in serum compared with the high fat diet rats that did not receive curcumin (P<0.05, P<0.01). An oral glucose tolerance test and an intraperitoneal insulin tolerance test revealed that insulin resistance was reduced (P<0.05 or P<0.01) and tissue section analysis revealed that hepatosteatosis was attenuated following treatment with curcumin. Furthermore, the protein expression of Notch1 and its downstream target Hes1 were suppressed. These effects were also in parallel with an upregulation of fatty acid oxidationassociated gene expression, including peroxisome proliferatoractivated receptor (PPAR)α, carnitine palmitoyltransferase 1 and PPARγ (P<0.05). In addition, curcumin administration led to a downregulation in the expression of lipogenic genes, including sterol regulatory elementbinding protein, fatty acid synthase and acetylCoA carboxylase (P<0.05). The expression of inflammationassociated genes, including nuclear factorκB, tumor necrosis factorα and prostaglandinendoperoxide synthase 2 were also suppressed. The results of the present study suggest that the hepatic Notch1 pathway can be suppressed via curcumin treatment, which may ameliorate fatty liver and insulin resistance in rats subjected to a high fat diet.
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Curcumina/farmacología , Receptor Notch1/metabolismo , Transducción de Señal/efectos de los fármacos , Animales , Glucemia , Peso Corporal/efectos de los fármacos , Modelos Animales de Enfermedad , Hígado Graso/metabolismo , Hígado Graso/patología , Resistencia a la Insulina , Grasa Intraabdominal/efectos de los fármacos , Metabolismo de los Lípidos/efectos de los fármacos , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Masculino , Modelos Biológicos , Tamaño de los Órganos/efectos de los fármacos , RatasRESUMEN
The luteinizing hormone receptor (LHR) is essential for sexual development and reproduction in mammals. We have established that Sp1 has a central role in derepression of LHR gene transcription induced by Trichostatin A (TSA) in MCF7 cells. Moreover, the co-activator PC4 which associates directly with Sp1 at the LHR promoter is essential for TSA-mediated LHR transcription. This study explores interactions of PC4 with histone proteins, which presumably triggers chromatin modifications during LHR transcriptional activation. TSA treatment of MCF7 cells expressing PC4-Flag protein induces acetylation of histone 3 (H3) and immunoprecipitation (IP) studies revealed its interaction with PC4-Flag protein. MS/MS analysis of the protein complex obtained after IP from TSA treated samples detected H3.3 acetylated at K9, K14, K18, K23 and K27 as a PC4 interacting protein. The association of PC4 with H3.3 was corroborated by IP and re-ChIP using H3.3 antibody. Similarly, IP and re-ChIP showed association of PC4 with H3 acetylated protein. Knockdown of PC4 in MCF7 cells reduced H3.3 enrichment, H3 acetylation at the Lys sites and LHR promoter activity in TSA treated cells despite an increase in H3 and H3.3 protein induced by TSA, linking PC4 to H3 acetylation and LHR transcription. Depletion of H3.3 A/B in MCF7 cells impair chromatin accessibility and enrichment of Pol II and TFIIB at the LHR promoter and its activation, resulting in marked reduction of LHR gene expression. Together, these findings point to the critical role of PC4 and its association with acetylated H3.3 in TSA-induced LHR gene transcription.
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Proteínas de Unión al ADN/metabolismo , Histonas/metabolismo , Receptores de HL/genética , Activación Transcripcional , Acetilación , Histonas/química , Humanos , Ácidos Hidroxámicos/farmacología , Células MCF-7 , ARN Mensajero/metabolismo , Receptores de HL/metabolismo , Espectrometría de Masas en TándemRESUMEN
Objective. The randomized controlled trial was to evaluate the efficacy of topical Chinese herbal Zhangpi Ointment for hydroxyurea-induced leg ulcers in patients with myeloproliferative neoplasms. Patients and Methods. This single-center, prospective, randomized, open-label, controlled clinical trial conducted at Shanghai Ninth People's Hospital enrolled 54 patients with hydroxyurea-induced leg ulcers. Patients were randomly assigned to the control group (n = 27) treated with chlorhexidine dressing or the intervention group (n = 27) treated with the Zhangpi Ointment. Finally, 26 patients in the control group and 23 patients in the intervention group completed 8 weeks of observation. Results. The rate of complete healing was 100% for the intervention group, which was significantly higher than that of the control group (96.15%) (P<0.05). Furthermore, the intervention group achieved a significantly higher rate of wound healing (95.56%) than the control group (69.02%) at week 4 (P<0.01). The intervention group took 34 ± 5 days to achieve complete healing while the control group took 41 ± 7 days (P < 0.01). Moreover, grade 3/4 side effects were observed in neither group. Conclusion. The Zhangpi Ointment is effective in promoting the healing of hydroxyurea-induced leg ulcers in patients with myeloproliferative neoplasms, providing a therapeutic option for a condition that is recalcitrant to conventional therapy.
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BACKGROUND: The application of breast-conserving surgery (BCS) on patients with locally advanced breast cancer (LABC) with good response to neoadjuvant chemotherapy (NACT) still remains controversial. The objective in this study is to analyze the safety of BCS in the management of LABC in patients with good response to NACT. METHODS: We searched the electronic databases of Medline (Pubmed) and Cochrane Library for reports on local recurrence (LR), regional recurrence (RR), distant recurrence (DR), 5-year disease-free survival (DFS) or 5-year overall survival (OS) in patients with LABC receiving BCS or mastectomy (MT) and with good response to NACT. Based on the research results, we conducted a meta-analysis using Review Manager 5.3. RESULTS: Our study showed that 16 studies with a combined total of 3531 patients, of whom 1465 patients underwent BCS, whereas 2066 patients underwent MT. There was no significant heterogeneity among these studies (Q statistic: P = .88; I = 0%). Patients with good response to NACT showed no significant difference in LR and RR [odd ratio (OR)â=â0.83; 95% confidence interval (CI): 0.60-1.15; Pâ=â.26; ORâ=â0.56; 95% CI: 0.33-0.93; Pâ=â.03], while we figured out a lower DR (ORâ=â0.51; 95% CI: 0.42-0.63; Pâ<â.01), a higher DFS (ORâ=â2.35; 95% CI: 1.84 to 3.01, Pâ<â.01) and a higher OS (ORâ=â2.12; 95% CI: 1.51 to 2.98, Pâ<â.01) in BCS compared with MT. CONCLUSION: This meta-analysis concluded that BCS was a safe surgery for patients with LABC and had good response to NACT.