Objective:Previous studies suggest a complex interaction between 3,3',5-triiodo-L-thyronine (T3) and inflammation, but this relationship remains unclear. This study investigates the association between free triiodothyronine (FT3) levels and inflammatory markers in the US population using National Health and Nutrition Examination Survey (NHANES) data. Methods:This study analyzed NHANES data from three cycles between 2007 and 2012, using Spearman correlation tests and multivariate linear regression. Subgroup analyses and interaction tests were conducted based on age, gender, race, body mass index (BMI), thyroid-stimulating hormone, and free thyroxine (T4) to determine the correlations between FT3 and seven systemic inflammatory markers: C-reactive protein (CRP), neutrophil-to-lymphocyte ratio (NLR), platelet-to-white blood cell ratio (PWR), platelet-to-lymphocyte ratio (PLR), monocyte-to-lymphocyte ratio (MLR), systemic immune-inflammation index (SII), and systemic inflammation response index (SIRI). Results:A total of 2306 participants were included in this study. Univariate correlation analysis showed that CRP, NLR, MLR, and PLR were significantly negatively correlated with FT3 levels (all p  < 0.05). After adjusting for confounders, FT3 levels were significantly negatively associated with CRP, NLR, and PLR (all p  < 0.05). Subgroup analysis showed that age significantly modified the associations between FT3 and systemic inflammatory markers, such as CRP, NLR, MLR, SII, and SIRI (p-interaction < 0.05). The inverse associations were more consistent among individuals aged ≥ 65 years. Conclusion:In the general population, FT3 levels exhibit inverse associations with systemic inflammatory markers.

Background: While elevated thyroid-stimulating hormone (TSH) is a known risk factor for differentiated thyroid cancer (DTC), it remains unclear whether autoimmune thyroiditis (AT) modifies this association. Clarifying this interaction is critical for personalized risk assessment and TSH suppression therapy. Methods: This study performed a retrospective analysis including 2425 participants who underwent thyroidectomy for thyroid nodules. Participants were categorized based on histological AT diagnosis and thyroid peroxidase antibody (TPOAb) and thyroglobulin antibody (TgAb) levels. Multivariable logistic regression models were used to assess the association between thyroid parameters and DTC risk, stratified by AT and autoantibody status. Results: The prevalence of histological diagnosed AT, TgAb-positivity, and TPOAb-positivity among DTC patients was 31.58%, 13.68%, and 18.76%, respectively. An increase in one standard deviation in TSH, thyrotrophic thyroxine resistance index (TT4RI), and TSH index (TSHI) was associated with an elevated risk of DTC in euthyroid individuals without AT or positive thyroid autoantibodies. A positive and nonlinear association between TSH and DTC risk in euthyroid patients without AT was identified, with inflection points at TSH levels of 1.32 mIU/L. In subgroups characterized by concurrent AT, TgAb-positivity, or TPOAb-positivity, these thyroid parameters showed no statistically significant correlation with DTC risk. Conclusions: The association between TSH and DTC risk varies according to autoimmune thyroiditis status. These findings highlight the importance of considering thyroid autoimmunity in thyroid cancer risk assessment and warrant prospective evaluation to determine its potential implications for TSH suppressive therapy.

The role of thyroid-stimulating hormone receptor (TSHR) in macrophages on adipose tissue metabolic disorders remains unclear. We generated macrophage-specific TSHR knockout mice (LysM-Cre Tshrfl/fl) using the Cre/LoxP system and induced subclinical hypothyroidism (SCH) via methimazole treatment. Metabolic/inflammatory phenotypes were assessed via glucose/insulin tolerance tests, histology, and molecular analyses. Mitochondrial function and proinflammatory polarization were examined in primary peritoneal macrophages and RAW264.7 cells under TSH stimulation. The results showed macrophage TSHR deletion attenuated SCH-induced insulin resistance, inflammatory infiltration, and adipocyte hypertrophy. TSH triggered proinflammatory polarization via TSHR-dependent mitochondrial permeability transition pore (mPTP) overactivation, oxidative stress, and impaired electron transport chain function. Cyclophilin D (CypD), a key mPTP regulator, mediated TSH-induced mitochondrial dysfunction. Pharmacological CypD inhibition with cyclosporine A(CsA) reversed TSH-driven inflammation and metabolic deficits in vitro and in vivo. We conclude that TSH promotes adipose tissue dysfunction via macrophage TSHR signaling by enhancing CypD acetylation to disrupt mitochondrial homeostasis and drive pro-inflammatory polarization, unveiling the TSHR-CypD axis as a therapeutic target for SCH-related metabolic disorders.

Papillary thyroid cancer (PTC) progression is characterized by lymph node metastasis and thyroid capsular invasion. This study aimed to identify high-risk PTC populations for these events based on peripheral blood test parameters and to determine the associated factors. This retrospective study analyzed data from 4557 PTC patients. Principal component analysis (PCA) and cluster analysis were performed on 45 peripheral blood test results. High- and low-risk clusters were defined based on metastasis and invasion prevalence. Univariate and multivariate analyses identified parameters significantly differentiating the clusters, examining their association with tumor progression. Preoperative blood tests stratified patients into two distinct clusters. Cluster 0 demonstrated significantly higher rates of metastasis and invasion than Cluster 1, defining it as the high-risk group. PCA identified four principal components significantly differentiating the clusters. Analysis of these components revealed key peripheral blood parameters. Multivariable logistic regression identified six parameters associated with increased risk of Cluster 0: alanine aminotransferase, free triiodothyronine, thrombin time, hemoglobin, hematocrit, and leukocyte count. Conversely, aspartate aminotransferase and neutrophil count were associated with decreased risk. These findings suggest that peripheral blood parameters may provide insights into the progression of thyroid tumors and highlight potential avenues for exploring the underlying mechanisms of PTC. However, given the retrospective nature of this study and the potential for selection bias, further prospective studies are necessary to validate these results and confirm their predictive value in clinical practice.

Background m6A modification is important in cancer progression. In the research, we explored the functions and mechanisms of WTAP (the key component of m6A) in HCC. Methods Expression of WTAP and ITGB4 was examined by qRT-PCR, western blot and IHC assays. Tumor cell proliferation, apoptosis, metastasis and stemness were investigated via colony formation, flow cytometry, transwell and sphere formation assays. The interaction between WTAP and ITGB4 was estimated by RIP, meRIP and dual-luciferase reporter experiments. Results ITGB4 was upregulated in HCC. Knockdown of ITGB4 repressed HCC cell growth, motility and stemness, accelerated HCC cell apoptosis in vitro, and blocked tumorigenesis in vivo. Through SRAMP website and a series experiments, WTAP catalysed m6A modification on ITGB4 mRNA, recognized by the reader YTHDF1, leading to increased mRNA stability. WTAP overexpression aggravated the malignant behaviors and stemness of HCC cells, with ITGB4 deficiency abrogated the effects. In addition, we demonstrated that WTAP could activate FAK/PI3K/AKT signaling pathway via regulating ITGB4 expression. Conclusion WTAP mediated the m6A methylation modification of ITGB4 to promote HCC progression and tumor stemness, providing a new idea for HCC therapy.

Lymph node metastasis (LNM) and thyroid capsular invasion (CI) are the main pathological features leading to poor prognosis of differentiated thyroid cancer (DTC), and there is a lack of effective diagnostic methods before surgery. Therefore, this study was designed to analyze a large number of preoperative clinical features of DTC and identify factors closely related to those two pathological features. 4557 patients with DTC, postoperative pathological results showed LNM in 2146 cases and CI in 2783 cases were retrospectively included. The preoperative blood, urine, serum laboratory test and ultrasound of thyroid were performed for data collection. A total of 74 clinical features were analyzed by the methods of principal component analysis (PCA), and key principal components were extracted for regression analysis of LNM and CI as well as subgroup analysis. 11 key clinical features were used for principal component analysis, and 6 principal components PC0-PC5 were finally obtained. PC0 is mainly composed of prothrombin time and international normalized ratio, and the score represents better coagulation function and has a protective effect on LNM. PC1 is mainly composed of thyroid peroxidase antibody and thyroid texture, and the score represents the severity of thyroiditis and has a protective effect on LNM and CI. Thyroiditis and coagulation function were identified by principal component analysis as protective and risk factors for adverse pathology of DTC, meaning they were closely related to tumor metastasis and invasion.

Objective Hashimoto's thyroiditis (HT) is a common disease characterized by autoimmune injury of the thyroid. Its pathogenesis entails complex interactions among hereditary predisposition, immune disorders and environmental factors. In recent years, viral infection has attracted much attention as a potential environmental trigger, but the role genes associated with HT remain unclear. Methods In this study, COVID-19-related genes were combined with transcriptome data (GSE29315, GSE138198) of HT patients in the GEO database. Key genes were selected using machine learning (LASSO, SVM, and RF), GO/KEGG enrichment, and GSEA. Its function was confirmed by single-cell sequencing and ssGSEA immunoinfiltration analysis. Results A total of 16 co-expressed genes of HT and viral infection were identified. KEGG and GO enrichment results showed that these genes were significantly enriched in inflammatory signaling, viral defense and immune cell activation pathways. After screening by machine learning algorithm, four key genes (IFITM3, IFI44L, CCL3, OAS1) were finally identified as the common diagnostic markers of HT and viral infection, and the ROC curve also showed good diagnostic performance. In addition, single cell sequencing further confirmed its high expression in thyroid tissue and immune infiltrating cells. Conclusion A virus-triggered autoimmune cascade involving IFITM3, IFI44L, CCL3, and OAS1 may precipitate HT. These four genes constitute robust, multi-omics biomarkers for early diagnosis and targeted therapy of HT following viral infection.

OBJECTIVE:The study aimed (1) to investigate the association between aggressive clinicopathological characteristics and the American Thyroid Association (ATA) recurrence risk classification in differentiated thyroid cancer (DTC) patients, and (2) to investigate the prognostic value of preoperative thyroid parameters. METHODS:A total of 3833 patients histologically confirmed DTC were recruited. Preoperative clinical and postoperative pathologic data were retrospectively collected. Participants were stratified into low recurrence risk and intermediate-to-high recurrence risk groups based on the ATA risk stratification system. RESULTS:The study cohort had a mean age of 48.87 ± 8.08 years, and 1,465 (76.82%) were female. Male (OR = 1.37, p = .024), aged 52 years and older (OR = 2.01, p < .001), larger tumor size (OR = 3.71, p = 0.011), nerve invasion (OR = 6.69, p = .004), margin involvement (OR = 5.46, p < .001), multifocality (OR = 3.71, p < .001), and bilaterality (OR = 3.95, p < .001) were identified as risk factors for a higher ATA recurrence risk classification, in addition to established factors such as lymph node metastasis and angioinvasion, after adjusting for potential confounding variables. Higher preoperative levels of free triiodothyronine (FT3), FT3 to free thyroxine (FT3/FT4), and lower thyroid feedback quantile-based index (TFQI) levels were associated with a higher ATA recurrence risk classification. The comprehensive predictive model incorporating these variables demonstrated excellent discrimination (AUC = 0.836). Furthermore, higher FT3/FT4 levels and lower TFQI levels were associated with higher risk of lymph node metastases and angioinvasion. CONCLUSIONS:Factors such as male sex, older age, multifocality, bilaterality, margin involvement, nerve invasion, larger tumor size, and preoperative thyroid parameters serve as complementary predictors for higher ATA recurrence risk in DTC, in addition to conventional risk factors. These insights contribute to a more nuanced understanding and optimization of current risk stratification methodologies.

Background:Recent studies suggest connection between the thyroid stimulating hormone (TSH) and insulin resistance (IR). Adipose tissue is one of insulin's target tissues. However, currently the regulatory mechanism of TSH on the adipose tissue is not fully investigated yet. Methods:We constructed a subclinical hypothyroidism (SCH) mouse model induced by methimazole with elevated TSH levels and then observed its metabolic profile, adipose tissue IR, and the adipose tissue macrophages (ATMs) phenotype. In vitro, we treated RAW264.7 cells and bone marrow-derived macrophages (BMDM) to assess the effect of TSH on macrophage polarization and explore the specific underlying mechanisms. Results:SCH mice exhibited a poorer metabolic profile and an advanced adipose tissue IR. Meanwhile, the number of M1 ATMs was increased in SCH mice adipose tissue. In vitro, TSH induced endoplasmic reticulum stress in macrophages, which activated the GRP78-ATF6-CHOP signaling pathway, and further promoted M1 macrophage polarization. 4-phenylbutyric acid (4-PBA), an endoplasmic reticulum stress inhibitor, corrected the polarization imbalance of ATMs in SCH mice adipose tissue and improved adipose tissue dysfunction and IR. Conclusion:TSH activated endoplasmic reticulum stress in macrophages, which induced the polarization of ATMs toward a pro-inflammatory M1 phenotype and promotes adipose tissue IR. Our findings highlight the possible relationship of TSH with immunity and metabolism.

Background: Traditional Chinese medicine (TCM) offers the advantage of effectively relieving rheumatoid arthritis (RA) with minimal side effects. The Juanbi recipe is a commonly utilized TCM treatment for RA, yet its pharmacological mechanism remains unclear. Network pharmacology serves as an effective tool for identifying pharmaceutical ingredients and potential therapeutic targets of TCM, thereby uncovering its mechanisms. This study aimed to identify the core target genes and explore the mechanisms underlying the treatment of RA with the Juanbi recipe. Methods: This study adopted the method of network pharmacology to filter key gene targets of Juanbi recipe in RA treatment. Single-cell ribonucleic acid (RNA) sequencing data was used to screen the key genes to form the core genes of Juanbi recipe in RA treatment. The molecular docking technique was used to verify the core target genes and explore the mechanisms of Juanbi recipe in RA treatment. The RA model of mice was induced by the collagen-induced arthritis and the effect of Juanbi recipe was evaluated by intragastric administrating of extraction of Juanbi recipe. Enzyme linked immunosorbent assay was used to analysis serum inflammatory factors. Hematoxylin and eosin staining was used to evaluate inflammation and immunohistochemical (IHC) staining was used to evaluate core target genes and pathways in synovium of ankle. Results: This study screened out 281 active molecules in Juanbi recipe, found 105 key target genes of Juanbi recipe in RA treatment, and drew an "ingredient - molecule - gene" diagram. Juanbi recipe reduced the levels of serum interleukin (IL)-1 and IL-6, the inflammatory infiltration in synovium, demonstration that Juanbi recipe reduced both systemic and synovial inflammatory response. Single cell RNA sequencing data were used to select six core target genes and six core active molecules of Juanbi recipe in RA treatment. The pathways of Juanbi recipe in RA treatment involved in activator protein-1 (AP-1) and nuclear factor kappa B (NF-kappa B) pathway. Results of western blot and IHC staining showed that Juanbi recipe decreased the expressions of c-jun and p65, which demonstrated that Juanbi recipe inhibited the expression of AP-1 and Conclusions: The core active molecules of Juanbi recipe could inhibit key factors of AP-1 and NF-kappa B pathway to inhibit the inflammation, which played a protective role in RA.

In recent years, the incidence of thyroid cancer has been increasing, posing a significant public health problem. This study aims to assess the burden of thyroid cancer in China and globally between 1990 and 2021. It seeks to elucidate trends in incidence, prevalence, mortality, and disability-adjusted life years (DALYs), while also identifying variations across different age groups and genders. Data were extracted from the Global Burden of Disease (GBD) database from 1990 to 2021, focusing on thyroid cancer indicators in China and globally. We calculated age-standardised incidence (ASIR), age-standardised prevalence (ASPR), age-standardised mortality (ASMR), and age-standardised DALY rate (ASDR) for each age group, and analyzed the annual percentage change (APC) in trends over the study period using Joinpoint regression models. In China, the incidence of thyroid cancer increased by 295.70% between 1990 and 2021, with a significant annual increase of 2.242%. The global incidence increased by 177.62% and the APC was 1.139%. While ASMR declined slightly both in China and globally, ASPR showed a substantial increase. It is worth noting that the burden of thyroid cancer is higher in women, and the increased risk of developing the disease has been more pronounced in men in recent years. The findings highlight the need for targeted prevention strategies, improved diagnostics to avoid overdiagnosis, and equitable allocation of public health resources to address the growing thyroid cancer challenge.

BackgroundNon-invasive prognostic predictors for rare pancreatic neuroendocrine tumors (PNETs) are lacking. We aimed to approach the prognostic value of preoperative systemic inflammatory markers in patients with PNETs.MethodsThe clinical data of 174 patients with PNETs undergoing surgical treatment were retrospectively analyzed to explore the correlation of neutrophil to lymphocyte ratio (NLR), platelet to lymphocyte ratio (PLR), lymphocyte to monocyte ratio (LMR), and platelet to white blood cell ratio (PWR) with clinicopathological parameters and the progression of tumor after the operation. The optimal cutoff values for predictors and the area under the curve (AUC) of the receiver operating characteristic (ROC) were estimated. Univariate and multivariate Cox proportional hazards models were used to assess the relation between NLR, LMR, PLR, and progression-free survival (PFS), examined by the Kaplan–Meier and log-rank tests.ResultsThe scores of the NLR (P = 0.039) and PLR (P = 0.011) in the progression group were significantly higher than those in the progression-free group, and the LMR was significantly lower than those in the progression-free group (P = 0.001). The best cutoff values of NLR, LMR, and PLR before operation were 2.28, 4.36, and 120.91. The proportions of tumor progression in the high NLR group (P = 0.007) and high PLR group (P = 0.013) obviously increased, and the proportion of tumor development in the low LMR group was higher than that in the high LMR group (P < 0.001). The K-M survival curve showed that the progression-free survival rate was lower in the high NLR group (P = 0.004), the low LMR group (P < 0.001), and the high PLR group (P = 0.018). The results of the multivariate Cox proportional hazards model suggested that preoperative LMR (HR = 3.128, 95% CI: 1.107~8.836, P = 0.031) was an independent predictor of PFS.ConclusionThe markers of systemic inflammation, especially LMR, can predict the postoperative progression of PNETs.

In recent years, there has been an increasing incidence of metabolic syndrome, type 2 diabetes, and cardiovascular events related to insulin resistance. As one of the target organs for insulin, adipose tissue is essential for maintaining in vivo immune homeostasis and metabolic regulation. Currently, the specific adipose tissue mechanisms involved in insulin resistance remain incompletely understood. There is increasing evidence that the process of insulin resistance is mostly accompanied by a dramatic increase in the number and phenotypic changes of adipose tissue macrophages (ATMs). In this review, we discuss the origins and functions of ATMs, some regulatory factors of ATM phenotypes, and the mechanisms through which ATMs mediate insulin resistance. We explore how ATM phenotypes contribute to insulin resistance in adipose tissue. We expect that modulation of ATM phenotypes will provide a novel strategy for the treatment of diseases associated with insulin resistance.

Background: The metalloprotease ADAMTS-7 (a disintegrin and metalloproteinase with thrombospondin type 1 motif 7) is a novel locus associated with human coronary atherosclerosis. ADAMTS-7 deletion protects against atherosclerosis and vascular restenosis in rodents. Methods: We designed 3 potential vaccines consisting of distinct B cell epitopic peptides derived from ADAMTS-7 and conjugated with the carrier protein KLH (keyhole limpet hemocyanin) as well as aluminum hydroxide as an adjuvant. Arterial ligation or wire injury was used to induce neointima in mice, whereas ApoE-/- and LDLR-/- (LDLR [low-density lipoprotein receptor]) mice fed a high-fat diet were applied to assess atherosclerosis. In addition, coronary stent implantation was performed on vaccine-immunized Bama miniature pigs, followed by optical coherence tomography to evaluate coronary intimal hyperplasia. Results: A vaccine, ATS7vac, was screened out from 3 candidates to effectively inhibit intimal thickening in murine carotid artery ligation models after vaccination. As well, immunization with ATS7vac alleviated neointima formation in murine wire injury models and mitigated atherosclerotic lesions in both hyperlipidemic ApoE-/- and LDLR-/- mice without lowering lipid levels. Preclinically, ATS7vac markedly impeded intimal hyperplasia in swine stented coronary arteries, but without significant immune-related organ injuries. Mechanistically, ATS7vac vaccination produced specific antibodies against ADAMTS-7, which markedly repressed ADAMTS-7–mediated COMP (cartilage oligomeric matrix protein) and TSP-1 (thrombospondin-1) degradation and subsequently inhibited vascular smooth muscle cell migration but promoted re-endothelialization. Conclusions: ATS7vac is a novel atherosclerosis vaccine that also alleviates in-stent restenosis. The application of ATS7vac would be a complementary therapeutic avenue to the current lipid-lowering strategy for atherosclerotic disease.

钠-葡萄糖协同转运蛋白2（SGLT2）抑制剂已成为治疗心力衰竭（心衰）的新兴药物,但其作用机制尚不清楚。健康人心肌细胞仅表达SGLT1,SGLT2定位在肾脏近曲小管和心外膜脂肪组织（EAT）,在疾病状态下高表达。舒张性心衰患者发生EAT堆积,EAT高表达SGLT2和分泌脂肪细胞因子,介导心肌纤维化和心肌肥厚;心肌细胞高表达SGLT1介导细胞内Na + 超载。基于SGLT2/1在心脏和肾脏的分布,我们推测SGLT2抑制剂治疗心衰的潜在作用机制主要涉及心脏血流动力学和心脏代谢重构,EAT-SGLT2可能是心脏代谢重构防治的重要靶点。

Objective The aims of the present study were to investigate gender differences in the clinicopathological features, distant metastasis and prognosis of pancreatic neuroendocrine neoplasms (pNENs) in a Chinese population, and to identify any important gaps in the classification and management of pNENs relative to gender. Methods Retrospective collection of the clinicopathological data of 193 patients with pathologically confirmed pNENs were analyzed and follow up was extended to observe the prognosis of the disease. Differences between genders in basic characteristics, clinical symptoms, comorbidities, and tumor parameters were analyzed. Results There was no significant difference in females and males, however, moderately higher for females (52.8% vs. 47.2%), with the largest subgroup being 40~60 years of age (54.9%). Age at onset (P=0.002) and age at diagnosis (P=0.005) were both younger in females compared to males. Males lived more in urban areas and females lived more in rural areas (P=0.047). The proportion of smokers and alcohol drinkers was significantly higher in males than in females (P < 0.001). Non-functional pNENs were more frequent in males and functional pNENs in females (P=0.032). In women, functional status of the tumor was significantly associated with metastatic outcome (P=0.007) and functional tumors proved to be a protective factor compared to non-functional tumors (OR=0.090,95% CI: 0.011~ 0.752). There were no gender differences in tumor size, location, grade, stage or prognosis. Conclusions Gender differences in some clinicopathological features, and distant metastasis in patients with pNENs were identified, which suggested certain management details that justified emphasis based on gender.

ObjectiveThe preoperative nutritional status of cancer patients is closely related to prognosis. The prognostic nutritional index (PNI) has been shown to predict the prognosis of a variety of tumors, but its study in pancreatic neuroendocrine neoplasms (pNENs) is lacking. The aim of the present study is to investigate the predictive value of the preoperative PNI for postoperative progression in patients with pNENs.MethodsThe medical records of 181 patients with pNENs, who underwent surgery, were retrospectively analyzed. A time-dependent receiver operating characteristic (ROC) curve was plotted to determine the optimal cut-off value of the preoperative PNI. Correlations between the preoperative PNI and clinicopathological parameters were analyzed using multiple linear regression. A Kaplan-Meier curve was applied to assess the progression-free survival (PFS) rate, which was tested using a log rank. Univariate and multivariate Cox proportional risk regression models were used to analyze the predictive value of the preoperative PNI on prognosis.ResultsThe optimal cut-off value of the preoperative PNI was 48.275. The patients were divided into a high PNI group (PNI > 48.275, n = 92) and a low PNI group (PNI ≤ 48.275, n = 89). The proportion of patients with tumor progression after surgery was significantly higher in the low PNI group compared with that in the high PNI group (P = 0.004). The Kaplan-Meier curve showed that the PFS rate after surgery was significantly lower in the low PNI group compared with that in the high PNI group (P = 0.026). The preoperative PNI was an independent predictor of PFS (HR: 2.727, 95% CI: 1.174∼6.333, P = 0.020).ConclusionThe preoperative PNI has a predictive value for postoperative progression in patients with pNENs.

Although curcumin (CUR) has many advantages, its hydrophobicity and instability limit its application. In this study, the anti-lipotoxic injury activity of CUR-loaded nanoparticles (CUR-NPs) and the corresponding mechanism were examined in palmitate (PA)-treated cardiomyocytes. An amphiphilic copolymer was selected as the vehicle material, and CUR-NPs with suitable sizes were prepared under optimized conditions. Cellular uptake was examined by confocal laser scanning microscopy, and cell proliferation inhibition rate was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetra bromide (MTT) assay. The terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay was used to detect cell apoptosis. The protein expression was detected by western blot. Exposure to PA reduces the proliferation of cardiomyocytes, but this effect was strongly reversed by CUR-NPs. In addition, our data showed that CUR-NPs strongly inhibited cell apoptosis in PA-treated cardiomyocytes. Furthermore, CUR-NPs remarkably increased the expression of LC3-II, as well as inhibited the expression of p-PERK, p-eIF2α, and ATF4 in PA-treated cardiomyocytes. Salubrinal (an eIF2α inhibitor) blocked the protective effect of CUR-NPs against PA-induced cardiomyocyte injury. Our results suggested that CUR-NPs can activated the autophagy pathway and protect myocardial cells from apoptosis, and these effects may be mediated by the eIF2α-related endoplasmic reticulum stress signaling pathway.

Myocardial infarction is a major cause of premature death in adults. Compromised cardiac function after myocardial infarction leads to chronic heart failure with systemic health complications and a high mortality rate1. Effective therapeutic strategies are needed to improve the recovery of cardiac function after myocardial infarction. More specifically, there is a major unmet need for a new class of drugs that can improve cardiomyocyte contractility, because inotropic therapies that are currently available have been associated with high morbidity and mortality in patients with systolic heart failure2,3 or have shown a very modest reduction of risk of heart failure4. Microtubule detyrosination is emerging as an important mechanism for the regulation of cardiomyocyte contractility5. Here we show that deficiency of microtubule-affinity regulating kinase 4 (MARK4) substantially limits the reduction in the left ventricular ejection fraction after acute myocardial infarction in mice, without affecting infarct size or cardiac remodelling. Mechanistically, we provide evidence that MARK4 regulates cardiomyocyte contractility by promoting phosphorylation of microtubule-associated protein 4 (MAP4), which facilitates the access of vasohibin 2 (VASH2)-a tubulin carboxypeptidase-to microtubules for the detyrosination of α-tubulin. Our results show how the detyrosination of microtubules in cardiomyocytes is finely tuned by MARK4 to regulate cardiac inotropy, and identify MARK4 as a promising therapeutic target for improving cardiac function after myocardial infarction.

ETHNOPHARMACOLOGICAL RELEVANCE:Gnaphalium affine D. Don is an important Traditional Chinese herbal Medicine (TCM) used to treat hyperuricemia, asthma, rheumatic arthritis, antitussive, expectorant and cardiovascular in folk medicine because of anti-inflammatory and anti-oxidant activity. The aim of this study was to investigate the potential beneficial effect of G. affine extract (GAE) on hydrogen peroxide (H2O2)-induced apoptosis and explore the possible underlying mechanism in cardiomyocyte.MATERIALS AND METHODS:The ingredients of GAE were isolated and tentatively identified by HPLC-ESI-Q-Qribatrip-MS/MS. The cardioprotective and anti-oxidant effects of GAE were evaluated in the experimental model with H2O2 induced apoptosis in H9c2 cells. H9c2 cells were pretreated for 3 h with or without GAE or with GAE plus PX866 (PI3K inhibitor), then exposed to H2O2 for 6 h, H9c2 cells viability were detected by CCK8 kit, the content of intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) and intracellular superoxide dismutase (SOD) activity were measured by the commercial biochemical kits, western blotting, immunohistochemical (IHC), immunofluorescence (IF) and reverse transcription-polymerase chain reaction (RT-PCR) assays were performed to evaluate the proteins and mRNA expression, propidium iodide (PI) staining was adopted to indicate H9c2 cells apoptosis.RESULTS:Firstly, seventeen polyphenols and flavonoids compounds with the characteristics of anti-inflammatory and anti-oxidant in GAE were tentatively identified by HPLC-ESI-Q-Qribatrip-MS/MS. In the experimental model, GAE not only significantly improved cells viability, but also showed anti-oxidant effects through improving SOD activity, up-regulating nuclear factor E2-related factor 2 (Nrf2), and decreasing intracellular concentration of ROS and MDA and the proteins expression of p47phox, p67phox and gp91phox. On the other hand, GAE revealed anti-apoptotic effect through up-regulating the expression of B-cell lymphoma-2 (Bcl-2), down-regulating Bcl2-associated X (BAX) and cleaved-caspase 3. Furthermore, GAE significantly facilitated phosphorylation of AKT and glycogen synthase kinase-3 beta (GSK-3β) but not AMPK, while the effects were blocked by PX866 (PI3K inhibitor).CONCLUSIONS:Our data suggested that GAE showed strong anti-oxidant effect to ameliorate oxidative stress and attenuate apoptosis induced by H2O2 in H9c2 cells by targeting PI3K/AKT/GSK-3β signaling pathway.