PDHA1 Alleviates Myocardial Ischemia-Reperfusion Injury by Improving Myocardial Insulin Resistance During Cardiopulmonary Bypass Surgery in Rats.

OBJECTIVE: Cardiopulmonary bypass (CPB) is a requisite technique for thoracotomy in advanced cardiovascular surgery. However, the consequent myocardial ischemia-reperfusion injury (MIRI) is the primary culprit behind cardiac dysfunction and fatal consequences post-operation. Prior research has posited that myocardial insulin resistance (IR) plays a vital role in exacerbating the progression of MIRI. Nonetheless, the exact mechanisms underlying this phenomenon remain obscure. METHODS: We constructed pyruvate dehydrogenase E1 α subunit (PDHA1) interference and overexpression rats and used ascending aorta occlusion in an in vivo model of CPB-MIRI. We devised an in vivo model of CPB-MIRI by constructing rat models with both pyruvate dehydrogenase E1α subunit (PDHA1) interference and overexpression through ascending aorta occlusion. We analyzed myocardial glucose metabolism and the degree of myocardial injury using functional monitoring, biochemical assays, and histological analysis. RESULTS: We discovered a clear downregulation of glucose transporter 4 (GLUT4) protein content expression in the CPB I/R model. In particular, cardiac-specific PDHA1 interference resulted in exacerbated cardiac dysfunction, significantly increased myocardial infarction area, more pronounced myocardial edema, and markedly increased cardiomyocyte apoptosis. Notably, the opposite effect was observed with PDHA1 overexpression, leading to a mitigated cardiac dysfunction and decreased incidence of myocardial infarction post-global ischemia. Mechanistically, PDHA1 plays a crucial role in regulating the protein content expression of GLUT4 on cardiomyocytes, thereby controlling the uptake and utilization of myocardial glucose, influencing the development of myocardial insulin resistance, and ultimately modulating MIRI. CONCLUSION: Overall, our study sheds new light on the pivotal role of PDHA1 in glucose metabolism and the development of myocardial insulin resistance. Our findings hold promising therapeutic potential for addressing the deleterious effects of MIRI in patients.

Camrelizumab combined with apatinib in patients with first-line platinum-resistant or PD-1 inhibitor resistant recurrent/metastatic nasopharyngeal carcinoma: a single-arm, phase 2 trial.

Immunotherapy combined with antiangiogenic targeted therapy has improved the treatment of certain solid tumors, but effective regimens remain elusive for refractory recurrent/metastatic nasopharyngeal carcinoma (RM-NPC). We conducted a phase 2 trial to evaluate the safety and activity of camrelizumab plus apatinib in platinum-resistant (cohort 1, NCT04547088) and PD-1 inhibitor resistant NPC (cohort 2, NCT04548271). Here we report on the primary outcome of objective response rate (ORR) and secondary endpoints of safety, duration of response, disease control rate, progression-free survival, and overall survival. The primary endpoint of ORR was met for cohort 1 (65%, 95% CI, 49.6-80.4, n = 40) and cohort 2 (34.3%; 95% CI, 17.0-51.8, n = 32). Grade ≥ 3 treatment-related adverse events (TRAE) were reported in 47 (65.3%) of 72 patients. Results of our predefined exploratory investigation of predictive biomarkers show: B cell markers are the most differentially expressed genes in the tumors of responders versus non-responders in cohort 1 and that tertiary lymphoid structure is associated with higher ORR; Angiogenesis gene expression signatures are strongly associated with ORR in cohort 2. Camrelizumab plus apatinib combination effectiveness is associated with high expression of PD-L1, VEGF Receptor 2 and B-cell-related genes signatures. Camrelizumab plus apatinib shows promising efficacy with a measurable safety profile in RM-NPC patients.

A new prognostic model for predicting outcomes of patients with recurrent or metastatic nasopharyngeal carcinoma receiving subsequent line (≥2 lines) anti-programmed death-1 monotherapy.

OBJECTIVES: About 17.7-34.0 % of patients with recurrent or metastatic nasopharyngeal carcinoma (RM-NPC) responded well to anti-PD-1 monotherapy. We sought to establish a nomogram to estimate the progression-free survival (PFS) of RM-NPC patients receiving subsequent-line anti-PD-1 monotherapy. MATERIALS AND METHODS: This cohort study investigated consecutive RM-NPC patients undergoing anti-PD-1 monotherapy. A nomogram was developed in the training cohort (n = 161), using a Cox multivariate model with backward stepwise inclusion, and was validated in the validation cohort (n = 69). Its predictive accuracy was assessed using a concordance index (C-index) and calibration curve. The primary endpoint was PFS. Secondary endpoints included the objective response rate (ORR), disease control rate (DCR), and overall survival (OS). RESULTS: Liver metastasis, albumin, lactate dehydrogenase, monocyte-to-lymphocyte ratio, and plasma Epstein-Barr virus DNA were used to develop a nomogram that could separate patients into favourable- and unfavourable-prognosis groups. The C-index in the training and validation cohort were 0.70 and 0.68, respectively, which was confirmed by calibration curves. Median PFS (mPFS) was lower for the unfavourable-prognosis than for the favourable-prognosis group (1.80 vs 4.93; hazard ratio 2.49 [95 % confidence interval: 1.78-3.49]; p < 0.001), across all subgroups. OS exhibited the same pattern. The ORR and DCR were markedly lower in the unfavourable-prognosis than in the favourable-prognosis group. All results were confirmed in the validation cohort. CONCLUSION: Our model is a reliable prognostic indicator of PFS in RM-NPC patients undergoing anti-PD-1 monotherapy, allowing robust estimation of the immunotherapy benefit an individual might derive.

Exploring biomarkers for prognosis and neoadjuvant chemosensitivity in rectal cancer: Multi-omics and ctDNA sequencing collaboration.

Introduction: This study aimed to identified the key genes and sequencing metrics for predicting prognosis and efficacy of neoadjuvant chemotherapy (nCT) in rectal cancer (RC) based on genomic DNA sequencing in samples with different origin and multi-omics association database. Methods: We collected 16 RC patients and obtained DNA sequencing data from cancer tissues and plasma cell-free DNA before and after nCT. Various gene variations were analyzed, including single nucleotide variants (SNV), copy number variation (CNV), tumor mutation burden (TMB), copy number instability (CNI) and mutant-allele tumor heterogeneity (MATH). We also identified genes by which CNV level can differentiate the response to nCT. The Cancer Genome Atlas database and the Clinical Proteomic Tumor Analysis Consortium database were used to further evaluate the specific role of therapeutic relevant genes and screen out the key genes in multi-omics levels. After the intersection of the screened genes from differential expression analysis, survival analysis and principal components analysis dimensionality reduction cluster analysis, the key genes were finally identified. Results: The genes CNV level of principal component genes in baseline blood and cancer tissues could significantly distinguish the two groups of patients. The CNV of HSP90AA1, EGFR, SRC, MTOR, etc. were relatively gained in the better group compared with the poor group in baseline blood. The CNI and TMB was significantly different between the two groups. The increased expression of HSP90AA1, EGFR, and SRC was associated with increased sensitivity to multiple chemotherapeutic drugs. The nCT predictive score obtained by therapeutic relevant genes could be a potential prognostic indicator, and the combination with TMB could further refine prognostic prediction for patients. After a series of analysis in multi-omics association database, EGFR and HSP90AA1 with significant differences in multiple aspects were identified as the key predictive genes related to prognosis and the sensitivity of nCT. Discussion: This work revealed that effective combined application and analysis in multi-omics data are critical to search for predictive biomarkers. The key genes EGFR and HSP90AA1 could serve as an effective biomarker to predict prognose and neoadjuvant chemosensitivity.

Identification and Validation in a Novel Quantification System of Ferroptosis Patterns for the Prediction of Prognosis and Immunotherapy Response in Left- and Right-Sided Colon Cancer.

Background: This study aimed to establish a novel quantification system of ferroptosis patterns and comprehensively analyze the relationship between ferroptosis score (FS) and the immune cell infiltration (ICI) characterization, tumor mutation burden (TMB), prognosis, and therapeutic sensitivity in left-sided and right-sided colon cancers (LCCs and RCCs, respectively). Methods: We comprehensively evaluated the ferroptosis patterns in 444 LCCs and RCCs based on 59 ferroptosis-related genes (FRGs). The FS was constructed to quantify ferroptosis patterns by using principal component analysis algorithms. Next, the prognostic value and therapeutic sensitivities were evaluated using multiple methods. Finally, we performed weighted gene co-expression network analysis (WGCNA) to identify the key FRGs. The IMvigor210 cohort, TCGA-COAD proteomics cohort, and Immunophenoscores were used to verify the predictive abilities of FS and the key FRGs. Results: Two ferroptosis clusters were determined. Ferroptosis cluster B demonstrated a high degree of congenital ICI and stromal-related signal enrichment with a poor prognosis. The prognosis, response of targeted inhibitors, and immunotherapy were significantly different between high and low FS groups (HSG and LSG, respectively). HSG was characterized by high TMB and microsatellite instability-high subtype with poor prognosis. Meanwhile, LSG was more likely to benefit from immunotherapy. ALOX5 was identified as a key FRG based on FS. Patients with high protein levels of ALOX5 had poorer prognoses. Conclusion: This work revealed that the evaluation of ferroptosis subtypes will contribute to gaining insight into the heterogeneity in LCCs and RCCs. The quantification for ferroptosis patterns played a non-negligible role in predicting ICI characterization, prognosis, and individualized immunotherapy strategies.

Identification and quantification of immune infiltration landscape on therapy and prognosis in left- and right-sided colon cancer.

BACKGROUND: The left-sided and right-sided colon cancer (LCCs and RCCs, respectively) have unique molecular features and clinical heterogeneity. This study aimed to identify the characteristics of immune cell infiltration (ICI) subtypes for evaluating prognosis and therapeutic benefits. METHODS: The independent gene datasets, corresponding somatic mutation and clinical information were collected from The Cancer Genome Atlas and Gene Expression Omnibus. The ICI contents were evaluated by "ESTIMATE" and "CIBERSORT." We performed two computational algorithms to identify the ICI landscape related to prognosis and found the unique infiltration characteristics. Next, principal component analysis was conducted to construct ICI score based on three ICI patterns. We analyzed the correlation between ICI score and tumor mutation burden (TMB), and stratified patients into prognostic-related high- and low- ICI score groups (HSG and LSG, respectively). The role of ICI scores in the prediction of therapeutic benefits was investigated by "pRRophetic" and verified by Immunophenoscores (IPS) (TCIA database) and an independent immunotherapy cohort (IMvigor210). The key genes were preliminary screened by weighted gene co-expression network analysis based on ICI scores. And they were further identified at various levels, including single cell, protein and immunotherapy response. The predictive ability of ICI score for prognosis was also verified in IMvigor210 cohort. RESULTS: The ICI features with a better prognosis were marked by high plasma cells, dendritic cells and mast cells, low memory CD4+ T cells, M0 macrophages, M1 macrophages, as well as M2 macrophages. A high ICI score was characterized by an increased TMB and genomic instability related signaling pathways. The prognosis, sensitivities of targeted inhibitors and immunotherapy, IPS and expression of immune checkpoints were significantly different in HSG and LSG. The genes identified by ICI scores and various levels included CA2 and TSPAN1. CONCLUSION: The identification of ICI subtypes and ICI scores will help gain insights into the heterogeneity in LCC and RCC, and identify patients probably benefiting from treatments. ICI scores and the key genes could serve as an effective biomarker to predict prognosis and the sensitivity of immunotherapy.

Cerebral infarction after cardiac surgery.

Cerebral infarction, a common central nervous system complication after adult cardiac surgery, is one of the main factors leading to the poor prognosis of cardiac surgery patients besides cardiac insufficiency. However, there is currently no effective treatment for cerebral infarction. Therefore, early prevention and diagnosis of postoperative cerebral infarction are particularly important. There are many factors and mechanisms during and after cardiac surgery that play an important role in the occurrence of postoperative cerebral infarction, such as intraoperative embolism, systemic inflammatory response syndrome, atrial fibrillation, temperature regulation, blood pressure control, use of postoperative blood products, and so forth. The mechanism by which most risk factors act on the human body, leading to postoperative cerebral infarction, is not well understood, and further research is needed. Therefore, this paper aims to summarize and explain the relevant risk factors, mechanisms, clinical signs, imaging characteristics, and early diagnosis methods of cerebral infarction complications after cardiac surgery, and provides useful data for the establishment of related diagnosis and treatment standards.

Prognostic Immunity and Therapeutic Sensitivity Analyses Based on Differential Genomic Instability-Associated LncRNAs in Left- and Right-Sided Colon Adenocarcinoma.

Background: The purpose of our study was to develop a prognostic risk model based on differential genomic instability-associated (DGIA) long non-coding RNAs (lncRNAs) of left-sided and right-sided colon cancers (LCCs and RCCs); therefore, the prognostic key lncRNAs could be identified. Methods: We adopted two independent gene datasets, corresponding somatic mutation and clinical information from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Identification of differential DGIA lncRNAs from LCCs and RCCs was conducted with the appliance of "Limma" analysis. Then, we screened out key lncRNAs based on univariate and multivariate Cox proportional hazard regression analysis. Meanwhile, DGIA lncRNAs related prognostic model (DRPM) was established. We employed the DRPM in the model group and internal verification group from TCGA for the purpose of risk grouping and accuracy verification of DRPM. We also verified the accuracy of key lncRNAs with GEO data. Finally, the differences of immune infiltration, functional pathways, and therapeutic sensitivities were analyzed within different risk groups. Results: A total of 123 DGIA lncRNAs were screened out by differential expression analysis. We obtained six DGIA lncRNAs by the construction of DRPM, including AC004009.1, AP003555.2, BOLA3-AS1, NKILA, LINC00543, and UCA1. After the risk grouping by these DGIA lncRNAs, we found the prognosis of the high-risk group (HRG) was significantly worse than that in the low-risk group (LRG) (all p < 0.05). In all TCGA samples and model group, the expression of CD8+ T cells in HRG was lower than that in LRG (all p < 0.05). The functional analysis indicated that there was significant upregulation with regard to pathways related to both genetic instability and immunity in LRG, including cytosolic DNA sensing pathway, response to double-strand RNA, RIG-Ⅰ like receptor signaling pathway, and Toll-like receptor signaling pathway. Finally, we analyzed the difference and significance of key DGIA lncRNAs and risk groups in multiple therapeutic sensitivities. Conclusion: Through the analysis of the DGIA lncRNAs between LCCs and RCCs, we identified six key DGIA lncRNAs. They can not only predict the prognostic risk of patients but also serve as biomarkers for evaluating the differences of genetic instability, immune infiltration, and therapeutic sensitivity.

Deubiquitinating Enzymes Orchestrate the Cancer Stem Cell-Immunosuppressive Niche Dialogue: New Perspectives and Therapeutic Potential.

Cancer stem cells (CSCs) are sparks for igniting tumor recurrence and the instigators of low response to immunotherapy and drug resistance. As one of the important components of tumor microenvironment, the tumor associated immune microenvironment (TAIM) is driving force for the heterogeneity, plasticity and evolution of CSCs. CSCs create the inhibitory TAIM (ITAIM) mainly through four stemness-related signals (SRSs), including Notch-nuclear factor-κB axis, Hedgehog, Wnt and signal transducer and activator of transcription. Ubiquitination and deubiquitination in proteins related to the specific stemness of the CSCs have a profound impact on the regulation of ITAIM. In regulating the balance between ubiquitination and deubiquitination, it is crucial for deubiquitinating enzymes (DUBs) to cleave ubiquitin chains from substrates. Ubiquitin-specific peptidases (USPs) comprise the largest family of DUBs. Growing evidence suggests that they play novel functions in contribution of ITAIM, including regulating tumor immunogenicity, activating stem cell factors, upregulating the SRSs, stabilizing anti-inflammatory receptors, and regulating anti-inflammatory cytokines. These overactive or abnormal signaling may dampen antitumor immune responses. The inhibition of USPs could play a regulatory role in SRSs and reversing ITAIM, and also have great potential in improving immune killing ability against tumor cells, including CSCs. In this review, we focus on the USPs involved in CSCs signaling pathways and regulating ITAIM, which are promising therapeutic targets in antitumor therapy.

Prognostic Immune-Related Analysis Based on Differentially Expressed Genes in Left- and Right-Sided Colon Adenocarcinoma.

BACKGROUND: Colon adenocarcinoma (COAD) can be divided into left-sided and right-sided COAD (LCCs and RCCs, respectively). They have unique characteristics in various biological aspects, particularly immune invasion and prognosis. The purpose of our study was to develop a prognostic risk scoring model (PRSM) based on differentially expressed immune-related genes (IRGs) between LCCs and RCCs, therefore the prognostic key IRGs could be identified. METHODS: The gene sets and clinical information of COAD patients were derived from TCGA and GEO databases. The comparison of differentially expressed genes (DEGs) of LCCs and RCCs were conducted with appliance of "Limma" analysis. The establishment about co-expression modules of DEGs related with immune score was conducted by weighted gene co-expression network analysis (WGCNA). Furthermore, we screened the module genes and completed construction of gene pairs. The analysis of the prognosis and the establishment of PRSM were performed with univariate- and lasso-Cox regression. We employed the PRSM in the model group and verification group for the purpose of risk group assignment and PRSM accuracy verification. Finally, the identification of the prognostic key IRGs was guaranteed by the adoption of functional enrichment, "DisNor" and protein-protein interaction (PPI). RESULTS: A total of 215 genes were screened out by differential expression analysis and WGCNA. A PRSM with 16 immune-related gene pairs (IRGPs) was established upon the genes pairing. Furthermore, we confirmed that the risk score was an independent factor for survival by univariate- and multivariate-Cox regression. The prognosis of high-risk group in model group (P < 0.001) and validation group (P = 0.014) was significantly worse than that in low-risk group. Treg cells (P < 0.001) and macrophage M0 (P = 0.015) were highly expressed in the high-risk group. The functional analysis indicated that there was significant up-regulation with regard of lymphocyte and cytokine related terms in low-risk group. Finally, we identified five prognostic key IRGs associated with better prognosis through PPI and prognostic analysis, including IL2RB, TRIM22, CIITA, CXCL13, and CXCR6. CONCLUSION: Through the analysis and screening of the DEGs between LCCs and RCCs, we constructed a PRSM which could predicate prognosis of LCCs and RCCs, and five prognostic key IRGs were identified as well. Therefore, the basis for identifying the benefits of immunotherapy and immunomodulatory was built.

Ultrasound Elastic Parameters Predict Central Lymph Node Metastasis of Papillary Thyroid Carcinoma.

BACKGROUND: This study aimed to explore the new factors that can predict central lymph node metastasis (CLNM) of papillary thyroid carcinoma (PTC) independently from ultrasound characteristics, elastic parameters, and endocrine indicators. METHODS: A total of 391 patients with PTC undergoing thyroidectomy and prophylactic central lymph node dissection from January 2017 to June 2019 were collected to determine the independent predictors of CLNM by single-factor and multivariate logistic regression analysis. RESULTS: Multivariate logistic regression analysis showed 9 independent predictors of CLNM, age, male, tumors in the middle or lower poles (without tumors in the isthmus), tumors in the isthmus, multiple tumors, and maximum tumor diameter measured by ultrasound, microcalcification, visible surrounding blood flow signal, and the maximum value of elastic modulus (Emax).We used the aforementioned factors to establish a scoring prediction model: predictive score Y(P) = 1/[1 + exp (1.444 + 0.084 ∗ age - 0.834 ∗ men - 0.73 ∗ multifocality - 2.718 ∗ tumors in the isthmus - 0.954 ∗ tumors in the middle or lower poles - 0.086 ∗ tumor maximum diameter - 1.070 ∗ microcalcification - 0.892 ∗ visible surrounding blood flow signal - 0.021 ∗ Emax)]. The area under the curve of the receiver operating characteristic was 0.827. It was found that 0.524 was the highest index of Youden, and the best cutoff value for predicting CLNM. When Y(P)≥0.524, the risk of CLNM in patients with PTC is predicted to be high. Predictive accuracy was 78.5% and 72.4% in the internal validation group and 78.6% in the external validation group. CONCLUSIONS: These data indicate that the scoring prediction model could provide a scientific and quantitative way to predict CLNM in patients with PTC.

Clinical Significance And Integrative Analysis Of Kinesin Family Member 18B In Lung Adenocarcinoma.

BACKGROUND: Kinesin family member 18B (KIF18B) is a member of the kinesin-8 superfamily, and functions as an oncogene in human cancers. However, its expression profile and role in lung adenocarcinoma (LUAD) remain unclear. MATERIALS AND METHODS: We examined the expression profile of KIF18B using quantitative real-time reverse transcription polymerase chain reaction and immunohistochemistry in fresh clinical samples. Using data downloaded from the Cancer Genome Atlas database and Gene Expression Omnibus, we explored the clinical significance of KIF18B, potential mechanisms of its dysregulation and its underlying biological function in LUAD. RESULTS: KIF18B was significantly over-expressed in LUAD tissues relative to normal tissues. High KIF18B expression was associated with smoking history, positive nodal invasion, advanced clinical stage, death status and poorer prognosis. Cox regression analyses revealed that KIF18B overexpression was an independent prognostic biomarker for poor overall survival (OS) and recurrence-free survival in LUAD. In addition, KIF18B mutation was observed in 2.2% of LUAD cases. DNA copy number variation was correlated with upregulated expression of KIF18B in LUAD tissues and cell lines. The methylation level of some KIF18B DNA CpG sites was negatively associated with its mRNA expression. KIF18B was predictively targeted by miR-125a-5p, which was downregulated in LUAD tissues, inversely correlated with KIF18B mRNA expression and significantly associated with poor OS. Furthermore, gene set enrichment analysis revealed that genes positively co-expressed with KIF18B were mainly enriched in cell cycle signaling pathways. CONCLUSION: Our results indicate that KIF18B is a promising prognostic biomarker for LUAD. DNA amplification, hypomethylation as well as miR-125a-5p downregulation may be involved in the mechanism of KIF18B dysregulation in LUAD. KIF18B might function as a novel oncogene through cell cycle regulation pathways in LUAD.

Role of Phosphorylated AMP-Activated Protein Kinase (AMPK) in Myocardial Insulin Resistance After Myocardial Ischemia-Reperfusion During Cardiopulmonary Bypass Surgery in Dogs.

BACKGROUND The aim of this study was to determine the role of AMP-activated protein kinase (AMPK) in myocardial insulin resistance after myocardial ischemia-reperfusion during cardiopulmonary bypass surgery in dogs. MATERIAL AND METHODS Twenty-four mongrel dogs were randomly assigned to 4 groups. The control group did not undergo aortic cross-clamping; the model group underwent 60 mins of aortic cross-clamping with 150 ml cardioplegic solution. The treatment group, the inhibition group respectively with 0.11mg/kg AICAR (AMPK agonist) in 150 ml cardioplegic solution and 0.11mg/kg Compound C (AMPK inhibitor) in 150 ml cardioplegic solution. The blood flow was determined and left ventricular myocardial tissue were taken at pre-bypass, 15, 60, and 90 min after aorta declamping, respectively. Expression of AMPK mRNA, p-AMPK and GLUT-4 proteins was determined by RT-PCR, IHC and WB. RESULTS Compared with the control group, receiving 60 min ischemia at 15 min after reperfusion, Myocardial Glucose Extraction Ratio were significantly decreased in the other 3 groups, it was significantly decreased from 20.0% to 1.2% at 60 min of reperfusion, and recovered to 6.1% after 90 min reperfusion in model group, while recovered to 4.1%, 12.0% after 90 min reperfusion respectively exposed to Compound C and AICAR. The expressions of p-AMPK, GLUT-4 protein and AMPK mRNA in myocardium were decreased in different experiment groups, but these changes occurred to a lesser extent in the treatment group. CONCLUSIONS The inability of GLUT-4 expression induced by the decreases in p-AMPK protein expression that may be one of the reasons for myocardial insulin resistance.

Effect of Si-RNA-silenced HIF-1α gene on myocardial ischemia-reperfusion-induced insulin resistance.

OBJECTIVE: To explore the effect (expression and implication) of hypoxia-inducible factor-1α (HIF-1α) silence induced by siRNA on the myocardial ischemia-reperfusion-induced insulin resistance in adult rats. METHODS: One-step enzymolysis method was used to isolate adult rat cardiomyocytes; adult rat cardiomyocytes were cultured; HIF-1α gene-specific Si-RNA was constructed and transfected into rat cardiomyocytes using liposome method. Myocardial IRI model was prepared. HIF-1α and glucose transporter 4 (GLUT-4) mRNA expression was detected by RT-PCR; distribution of GLUT-4 protein expression in adult rat cardiomyocytes was detected by immunofluorescence; Western blot was used for the detection of HIF-1α protein expression; isotope tracer assay was used to detect the changes in cell glucose (Glu) uptake rate. RESULTS: This method can stably get 85% to 90% active calcium tolerant adult rat cardiac myocytes, and the cultured cells were proved to be cardiomyocytes. After experiencing ischemia-reperfusion injury, HIF-1α mRNA expression levels in adult rat hypoxia cardiomyocytes had different degrees of increase compared with the control group (compared with the control group, P < 0.05). Compared with the model group, HIF-1α mRNA expression levels after ischemia and reperfusion in HIF-1αsi-RNA group and empty-vector group were lower than that in the control group and the model group; the expression reached the peak after 60 min of reperfusion, which did not change significantly in the control group. Expression of HIF-1α protein in myocardial cells was quite low in the control group; in the model group and intervention group, only after hypoxia-ischemia for 60 min, expression bands could be detected; especially in the model group, the expression had been increased until 60 min after reperfusion and began to decline from the time point of 180 min after reperfusion, but was still higher than that in the control group; in the intervention and empty-vector groups, it also increased rapidly at 60 min, but the expression was significantly lower than that in the model group; at 180 min after reperfusion, its protein expression peaked; while at 8 h after reperfusion, all the expression was extremely low. Compared with the control group, Glut4 mRNA expression in model group, transfected group and empty-vector group was reduced at the time points of T1-T4 (P < 0.05); the decline was the most significant at the time points of T1 and T2, followed by slightly increase at T3 and gradual recovery at T4; Compared with model group, Glut4 mRNA expression in transfection group was significantly reduced (P < 0.05); the decline was the most obvious at T1-T2, and then there was an increasing trend and it was recovered at T5 point. After experiencing ischemia, GLUT-4 protein expression changing trend was as follows: it was significantly reduced on the cell membrane, which was the most obvious from T1 to T3 and began to improve at T3, but still had not reached the level in the control group; it had been reached the levels of the control group at T5. After HIF-1αsi-RNA transfection and ischemia, GLUT-4 protein expression was increased in plasma and reduced on cell membrane; the decline was slightly improved at T3 and recovered to control distribution level at T5. After cardiac ischemia-reperfusion, glucose uptake rate decreased to varying degrees in myocardial cells and reached the lowest value after 60 min of ischemia, then gradually increased. After 8 h of reperfusion, the level in model group returned to the control level; compared with the model group, glucose concentration increased more serious in transfection group and empty-vector group after reperfusion. CONCLUSION: HIF-1α played a central regulatory role in this mechanism; HIF-1α may be one of the molecular mechanisms triggering myocardial IR.

Expression of metallothionein and Nrf2 pathway genes in lung cancer and cancer-surrounding tissues.

BACKGROUND: Nuclear factor (erythroid-derived 2)-like (Nrf)2 and metallothionein have been implicated in carcinogenesis. This study investigated the expression of Nrf2 and of Nrf2-targeted genes (NQO1 and GCLC) and the genes for the metallothionein (MT) isoforms (MT-1A and MT-2A) in human lung cancer and cancer-surrounding tissues. METHODS: Surgically removed lung cancer samples (n = 80) and cancer-surrounding tissues (n = 38) were collected from Zunyi Medical College Hospital, China. Total RNA was extracted, purified, and used for real-time reverse transcription-PCR analysis of interested genes. RESULTS: Expression of the Nrf2-targed genes NQO1 and GCLC tended to be higher (30 to 60%) in lung cancers, but was not significantly different from that in peri-cancer tissues. By contrast, expression of the genes for M)-1A, MT-2A, and the metal transcription factor MTF-1 were three-fold to four-fold lower in lung cancers. CONCLUSION: In surgical samples of lung cancer, MT expression was generally downregulated, whereas Nrf2 expression tended to be upregulated. These changes could play an integral role in lung carcinogenesis.

Rare syringyl acylated flavonol glycosides from the aerial parts of Leonurus japonicus Houtt.

Five new syringyl acylated flavonol glycosides, named leonurusoides A (1), B (2), C (3), D (4), and E (5), together with one known one 6 were obtained from the aerial parts of Leonurus japonicus. Their structures were elucidated by chemical and spectroscopic methods (UV, IR, HRESI-TOF-MS, 1D and 2D NMR). Compounds 1-6 showed triglyceride (TG) accumulation inhibitory effects in free fatty acid-induced HepG2 cells.

[Influence of siRNA-mediated PPARgamma gene knockdown on insulin resistance induced by myocardial ischemia-reperfusion in rats].

OBJECTIVE: To observe the influenece of siRNA-mediated PPARgamma gene knockdown on insulin resistance induced by myocardial ischemia-reperfusion in adult rats. METHOD: The targeting PPARgamma siRNA was synthesized. The myocardial cells of adult rats were isolated and cultured. They were divided into four groups: IRI group, siRNA-PPARgamma group, empty group and blank control group. Two groups of rat cardiac cells were transfected with PPARgamma-targeting siRNA (siRNA-PPARgamma group), or empty small interfering RNA (NC group), respectively. Real-time quantitive PCR was performed to detect the mRNA levels of PPARgamma and GLUT-4. PPARgamma protein expression level was determined with Western blot test. The uptake rate of glucose was determined by the isotope tracer method. RESULT: The PPARgamma mRNA and protein expression of IRI group were significantly higher than those in blank control group (P < 0.05). The PPARgamma mRNA and protein expression of siRNA-PPARgamma group were significantly less than those in blank control and IRI group (P < 0.01). There was no significant difference in the PPARgamma mRNA and protein expression between the blank group and IRI group. The mRNA expression of GLUT-4 in blank control was no significant difference at each time point. The mRNA expression of GLUT-4 in IRI group was significantly less at 0 min, but increased gradually over the following time point. Finally, The mRNA expression of GLUT-4 in IRI group restored the same level as blank control. There was no significant difference in the GLUT-4 mRNA expression between the empty group and IRI group. The GLUT-4 mRNA expression in siRNA-PPARgamma group was significantly less than that in IRI group or NC group (P < 0.05), and recovered more slowly than IRI group. After given insulin, The uptake rate of glucose in siRNA-PPARgamma group was significantly less at each time point compared with those in IRI group (P < 0.05), declined by 49.78%, 38.94%, 18.61%, 11.54% at 0 min, 15 min, 1 h, 2 h, respectively. At 6 h time point, the uptake rate of glucose in siRNA-PPARgamma group reached the same level as IRI group. There was no significant difference was observed in the uptake rate of glucose between the empty group and IRI group. CONCLUSION: The siRNA-mediated PPARgamma gene knockdown may enhance the myocardial insulin resistance. The molecular mechanisms that trigger myocardial cell insulin resistance might because the silence of PPARgamma expression decreasing the expression of GLUT-4 and decline its transportation from cytoplasm to membrane.

Asymmetric dimethylarginine reduced erythrocyte deformability in streptozotocin-induced diabetic rats.

To investigate the effect of asymmetric dimethylarginine on erythrocyte deformability in streptozotocin-induced diabetic rats, a single intraperitoneal injection of streptozotocin (STZ, 65 mg/kg) in male Sprague-Dawley rats was carried out to induce diabetes and normal erythrocytes were incubated with asymmetric dimethylarginine or aortic rings from diabetic rats in the presence of L-arginine or vitamin E. We found that erythrocyte deformability was significantly decreased in diabetic rats. The levels of asymmetric dimethylarginine in plasma and erythrocytes of diabetic rats were elevated significantly from 2-week diabetic duration to 8-week diabetic duration. Nitric oxide in erythrocytes was decreased at 8-week diabetic duration while plasma nitric oxide remained unchanged all along. The content of malondialdehyde in erythrocytes of diabetic rats was increased. After incubation of erythrocytes with asymmetric dimethylarginine (10(-6) M) for 30 min, erythrocyte deformability and nitric oxide level in erythrocytes were decreased markedly. Reactive oxygen species and malondialdehyde production in erythrocytes were promoted by asymmetric dimethylarginine. Both L-arginine and vitamin E reversed the effects of asymmetric dimethylarginine. After incubation of erythrocytes with aortic rings from diabetic rats, erythrocyte deformability was decreased, which was attenuated by L-arginine. These results indicated that reduction of erythrocyte deformability in diabetic rats was associated with promoted oxidant stress as well as impaired nitric oxide synthesis by elevation of asymmetric dimethylarginine.