ObjectiveTo examine the influences of Shenfu injection on the endogenous metabolic byproducts in the myocardium of the rat model exhibiting chronic heart failure， thus deciphering the therapeutic mechanism of the Qi-reinforcing and Yang-warming method. MethodsSD rats were randomly allocated into a control group and a modeling group. Chronic heart failure with heart-Yang deficiency syndrome in rats was modeled by multi-point subcutaneous injection of isoproterenol， and the rats were fed for 14 days after modeling. The successfully modeled rats were randomized into model， Shenfu injection （6.0 mL·kg^-1）， and trimetazidine （10 mg·kg^-1） groups and treated with corresponding agents for 15 days. The control group and the model group were injected with equal doses of normal saline， and the samples were collected after the intervention was completed. Cardiac color ultrasound was performed. Hematoxylin-eosin （HE） staining was used to observe histopathological morphology， and the serum level of N-terminal pro-brain natriuretic peptide （NT-proBNP） was assessed by enzyme-linked immunosorbent assay （ELISA）. The mitochondrial morphological and structural changes of cardiomyocytes were observed by transmission electron microscopy， and the metabolic profiling was carried out by ultra high performance liquid chromatography-quantitative exactive-mass spectrometry （UHPLC-QE-MS）. Differential metabolites were screened and identified by orthogonal partial least squares-discriminant analysis （OPLS-DA） and other methods， and then the MetaboAnalyst database was used for further screening. The relevant biological pathways were obtained through pathway enrichment analysis. The receiver operating characteristic （ROC） curve was established to evaluate the diagnostic value of each potential biomarker for myocardial injury and the evaluation value for drug efficacy. ResultsThe results of color ultrasound showed that Shenfu Injection improved the cardiac function indexes of model rats （P<0.05）. The results of HE staining showed that Shenfu injection effectively alleviated the pathological phenomena such as myocardial tissue structure disorder and inflammatory cell infiltration in model rats. The results of ELISA showed that Shenfu injection effectively regulated the serum NT-proBNP level in the model rats. Transmission electron microscopy （TEM） showed that Shenfu injection effectively restored the mitochondrial morphological structure. The results of metabolomics showed that the metabolic phenotypes of myocardial samples presented markedly differences between groups. Nine differential metabolites could be significantly reversed in the Shenfu injection group， involving three metabolic pathways： pyruvate metabolism， histidine metabolism， and citric acid cycle （TCA cycle）. The results of ROC analysis showed that the area under the curve （AUC） values of all metabolites were between 0.75 and 1.0， indicating that the differential metabolites had high diagnostic accuracy for myocardial injury， and the changes in their expression levels could be used as potential markers for efficacy evaluation. ConclusionShenfu injection significantly alleviated the damage of cardiac function， myocardium， and mitochondrial structure in the rat model of chronic heart failure with heart-Yang deficiency syndrome by ameliorating energy metabolism remodeling. Reinforcing Qi and warming Yang is a key method for treating chronic heart failure with heart-Yang deficiency syndrome.

Energy metabolism reconstruction is the new target of the treatment of heart failure. By combing the researches of Chinese medicinals for energy metabolism reconstruction of heart failure， it was found that Chinese medicinal compound formula and single Chinese medicinal have a certain role in regulating energy metabolism， mainly through three aspects， including the optimization of substrate utilization， improvement of mitochondrial structure， function， and homeostasis， and improvement of mitochondrial energy transport， so as to make the energy metabolism of the cardiomyocyte adjusted in the direction of beneficial to the organism， increasing the supply of energy， and improving the cardiac function.

ObjectiveThis study aims to explore the mechanism and targets of Shenfu Injection in regulating glycolysis to intervene in myocardial fibrosis in chronic heart failure based on the hypoxia-inducible factor-1α （HIF-1α）/ 6-phosphofructo-2-kinase/fructose-2，6-bisphosphatase 3 （PFKFB3） signaling pathway. MethodsA rat model of chronic heart failure was established by subcutaneous injection of isoproterenol （ISO）. After successful modeling， the rats were randomly divided into the Sham group， Model group， Shenfu injection （SFI， 6 mL·kg^-1） group， and inhibitor （3PO， 35 mg·kg^-1） group， according to a random number table， and they were treated for 15 days. Cardiac function was evaluated by echocardiography， and serum N-terminal pro-brain natriuretic peptide （NT-proBNP） levels were detected by enzyme-linked immunosorbent assay （ELISA）. Fasting body weight and heart weight were measured， and the heart index （HI） was calculated. Pathological changes in myocardial tissue were observed by hematoxylin-eosin （HE） and Masson staining， and the fibrosis rate was calculated. Biochemical assays were used to determine serum levels of glucose （GLU）， lactic acid （LA）， and pyruvic acid （PA）. Western blot was used to analyze the expression of proteins related to the HIF-1α/PFKFB3 signaling pathway （HIF-1α and PFKFB3）， glycolysis-related proteins （HK1， HK2， PKM2， and LDHA）， and fibrosis-related proteins ［transforming growth factor （TGF）-β₁， α-smooth muscle actin （α-SMA）， and Collagen type Ⅰ α1 （ColⅠA1）］. Real-time PCR was used to detect the mRNA expression of HIF-1α and PFKFB3 in myocardial tissue. ResultsCompared with the Sham group， the Model group showed significantly decreased left ventricular ejection fraction （LVEF）， left ventricular shortening fraction （LVFS）， interventricular septal thickness （IVSd）， and interventricular septal strain （IVSs） （P<0.05）， while left ventricular internal dimension at end-diastole （LVDd） and end-systole （LVIDs） were increased （P<0.05）. Serum NT-proBNP levels were significantly increased （P<0.01）， and body weight was decreased. Heart weight was increased， and the HIT index was increased （P<0.05）. Myocardial tissue exhibited inflammatory cell infiltration and collagen fiber deposition， and the fibrosis rate was significantly increased （P<0.05）. Serum GLU was decreased （P<0.05）， while LA and PA levels were increased （P<0.05）. Protein expressions of HIF-1α， PFKFB3， HK1， HK2， PKM2， LDHA， TGF-β₁， α-SMA， and ColⅠA1， as well as the mRNA expression of HIF-1α and PFKFB3 were increased （P<0.05）. Compared with the Model group， both the SFI group and 3PO groups showed significant improvements in LVEF， LVFS， IVSd， and IVSs （P<0.05） and decreases in LVDd， LVIDs， and NT-proBNP levels （P<0.05）. Body weight was significantly increased. Heart weight was significantly decreased， and the HIT index was significantly decreased （P<0.05）. Inflammatory cell infiltration， collagen fiber deposition， and the fibrosis rate were significantly decreased （P<0.05）. Serum GLU levels were significantly increased （P<0.05）， while LA and PA levels were decreased （P<0.05）. Expressions of glycolysis-related proteins， fibrosis-related proteins， and HIF-1α/PFKFB3 pathway-related proteins and mRNAs were significantly suppressed （P<0.05）. ConclusionSFI improves cardiac function in chronic heart failure by downregulating the expression of HIF-1α/PFKFB3 signaling pathway-related proteins， regulating glycolysis， and inhibiting myocardial fibrosis.

BACKGROUND: The incidence of leptomeningeal metastasis (LM) in patients with advanced non-small cell lung cancer (NSCLC) is increasing gradually. However, it poses therapeutic challenges due to limited effective interventions. Intrathecal Pemetrexed (IP) holds broad application prospects in the therapeutic domain of LM. This study aims to evaluate the efficacy, safety, and optimal combination strategies of IP in NSCLC-LM patients with epidermal growth factor receptor (EGFR) mutation-positive status, with the aim of providing real-world data support for exploring more precise personalized treatment strategies for these patients. METHODS: 104 EGFR-mutated NSCLC-LM patients who received IP treatment at Affiliated Nanjing Drum Tower Hospital of Nanjing University Medical School from January 2018 to June 2024 were analyzed retrospectively. Clinical parameters, treatment regimens, and survival outcomes were collected. The overall survival (OS), progression-free survival (PFS), clinical response rate and adverse events (AEs) were evaluated. RESULTS: The cohort demonstrated a median PFS of 9.6 months and OS of 13.0 months with 6-month and 1-year OS rates of 80.8% and 56.5%, respectively. Clinical response was observed in 77.9% of patients. The common AEs were myelosuppression (58.7%) and elevation of hepatic aminotransferases (25.0%). Nine (8.7%) patients experienced grade 4 myelosuppression and recovered to normal after receiving symptomatic treatment. Subgroup analyses revealed prolonged OS in patients with Karnofsky performance status (KPS) ≥60 versus <60 (14.4 vs 9.0 months, P=0.0022) and those receiving Bevacizumab therapy versus not (19.2 vs 10.5 months, P=0.0011). CONCLUSIONS IP exhibits promising efficacy and manageable toxicity in EGFR-mutated NSCLC-LM patients. When combined with Bevacizumab, it exerts synergistic antitumor effects with the potential to further improve clinical outcomes.
Humans ; Pemetrexed/therapeutic use* ; Carcinoma, Non-Small-Cell Lung/pathology* ; Male ; Female ; Middle Aged ; Lung Neoplasms/pathology* ; ErbB Receptors/genetics* ; Aged ; Mutation ; Adult ; Retrospective Studies ; Injections, Spinal ; Meningeal Neoplasms/genetics* ; Treatment Outcome ; Aged, 80 and over

Loss of protein homeostasis is a hallmark of cellular senescence, and ribosome pausing plays a crucial role in the collapse of proteostasis. However, our understanding of ribosome pausing in senescent cells remains limited. In this study, we utilized ribosome profiling and G-quadruplex RNA immunoprecipitation sequencing techniques to explore the impact of RNA G-quadruplex (rG4) on the translation efficiency in senescent cells. Our results revealed a reduction in the translation efficiency of rG4-rich genes in senescent cells and demonstrated that rG4 structures within coding sequence can impede translation both in vivo and in vitro. Moreover, we observed a significant increase in the abundance of rG4 structures in senescent cells, and the stabilization of the rG4 structures further exacerbated cellular senescence. Mechanistically, the RNA helicase DHX9 functions as a key regulator of rG4 abundance, and its reduced expression in senescent cells contributing to increased ribosome pausing. Additionally, we also observed an increased abundance of rG4, an imbalance in protein homeostasis, and reduced DHX9 expression in aged mice. In summary, our findings reveal a novel biological role for rG4 and DHX9 in the regulation of translation and proteostasis, which may have implications for delaying cellular senescence and the aging process.
G-Quadruplexes ; Cellular Senescence ; Ribosomes/genetics* ; Humans ; Animals ; Mice ; DEAD-box RNA Helicases/genetics* ; Protein Biosynthesis ; RNA/chemistry* ; Neoplasm Proteins

Objective : To explore the effect of Wilms′ tumor 1-associated protein(WTAP) on tissue collagen deposition in pulmonary fibrosis caused by bleomycin. Methods : 60 mice were randomly divided into four groups: control group(Control group), Bleomycin-induced pulmonary fibrosis group(BLM group), pulmonary fibrosis lentivirus empty vector control group(BLM+LV-NC group), pulmonary fibrosis WTAP lentivirus group virus group(BLM+LV-WTAP group). Experimental pulmonary fibrosis mouse model was established by subcutaneous injection of bleomycin(35 mg/kg) into the abdomen, twice a week for a total of 8 times. After modeling, Western Blot was used to detect the protein expression of fibrosis-related markers α-smooth muscle actin(α-SMA), type I collagen(Collagen Ⅰ), fibronectin(Fibronectin), and WTAP protein. Masson staining and Sirius Red staining were used to detect collagen deposition. RT-qPCR was used to detect WTAP mRNA expression, WTAP lentivirus infection effect, and Collagen Ⅰ mRNA expression. Results: Compared with the Control group, the expression of pulmonary fibrosis markers α-SMA(P<0.001), Collagen Ⅰ(P<0.001), and Fibronectin(P<0.01) protein in the BLM group all increased. Masson staining(P<0.001) and Sirius Red staining(P<0.001) confirmed that significant collagen deposition occurred in the lung tissue of the BLM group. In addition, the expression of WTAP protein in the lung tissue of the BLM group increased(P<0.01). Compared with the Control group, the expression of WTAP mRNA in the BLM group increased(P<0.001). Compared with the BLM+LV-NC group, the expression of WTAP mRNA in the tissues of the BLM+LV-WTAP group decreased(P<0.001), proving that virus infection is effective. After infection with WTAP lentivirus, collagen fiber deposition decreased(P<0.001), Collagen Ⅰ mRNA(P<0.001) level decreased, and protein(P<0.001) expression decreased in the BLM+LV-WTAP group. Conclusion Knocking down of WTAP can reduce collagen deposition in bleomycin-induced pulmonary fibrosis tissue in mice and improve experimental pulmonary fibrosis.

Objective: To investigate the effect of sirtuin 2(SIRT2) on the proliferation and migration of cardiac fibroblasts(CFs)in C57BL/6 mice under angiotensin II(Ang Ⅱ) stimulation. Methods : The hearts were taken from 1 to 2 days C57BL/6 milk mice. After cutting and digesting, CFs were extracted by different adhesion centrifugation. After CFs attachment, the cells were cultured under control medium and Ang Ⅱ(100 nmol/L) medium and treated using OE-SIRT2 plasmid to overexpression the SIRT2 gene. RT-qPCR was used to detect mRNA expression of SIRT2 proliferating cell nuclear antigen(PCNA), periostin(POSTN)and type Ⅰ collagen procollagen A1(Col1A1), Western blot assay was used to measure the protein expression levels of SIRT2, PCNA, POSTN and Col1A1, CCK-8 assay and EdU assay were used to evaluate CFs proliferation rate, Transwell experiment was used to assess CFs migration activity. Results: Compared with control group, Ang Ⅱ stimulation led to down-regulation of SIRT2 expression in CFs, increased collagen expression, and promoted CFs proliferation and migration. The expression of SIRT2 was up regulated in CFs treated with OE-SIRT2 plasmid under Ang Ⅱ stimulation, Col1A1, POSTN and PCNA expression was down regulated, and CFs proliferation and migration ability decreased. Conclusion Overexpression of SIRT2 can inhibit the proliferation and migration of CFs under Ang Ⅱ stimulation, indicating that SIRT2 may be a key regulatory point in the onset and progression of cardiac fibrosis.

Objective To assess the clinical and cranial CT/MR features of leptomeningeal metastasis(LM)from lung adenocar-cinoma with non-enhanced T2-fluid attenuated inversion recovery(T2-FLAIR)hyperintensity on the brainstem surface.Methods Eleven cases with LM from lung adenocarcinoma with non-enhanced T2-FLAIR hyperintensity on the brainstem surface confirmed by cere-brospinal fluid cytology were analyzed retrospectively.The clinical features included the pathological classification of the primary tumor,genetic testing results,and treatment methods.The imaging features included lesion location,MRI(T1 and T2)signals,diffusion weighted imaging(DWI)signal,enhancement patterns,CT density characteristics.Results The features of non-enhanced T2-FLAIR hyperintensity on the brainstem surface of 11 cases among 98 patients(11.2％)with LM from lung adenocarcinoma were reviewed.All patients were lung adenocarcinomas with epidermal growth factor receptor(EGFR)gene mutation and were diagnosed with LM after receiving EGFR-tyrosine kinase inhibitors(EGFR-TKIs)targeted therapy.There were 5 cases located in the pons,2 cases in the midbrain and pons,and 4 cases involving the midbrain,pons and medulla oblongata with the symmetrical band-like high signal on non-enhanced T2-FLAIR imaging.On T1WI,1 case showed slightly high signal,10 cases showed equal signal.On DWI,4 cases showed high signal,7 cases showed equal signal,and on enhanced T1WI,1 case showed mild enhancement,and 10 cases showed no enhancement.Additionally,on cranial CT scans,1 case showed calcification on the brainstem.Conclusion Non-enhanced T2-FLAIR hyperintensity on the brainstem surface may be more common among LM patient from lung adenocarcinoma with EGFR gene muta-tions and TKIs treatment,with the most frequent site being the pons.The features may include non-enhanced T2-FLAIR high signal,non-enhancement on enhanced T1 WI,restricted diffusion on DWI,and calcifications on CT imaging.

Objective To evaluate both the causal relationship between plasma metabolites and the risk of knee osteoarthritis(KOA)and the potential mediating or masking effect of immune cells using Mendelian randomization(MR)systems.Methods The GWAS data on 1400 plasma metabolites,731 immune cell traits and KOA was retrieved from the genome-wide association study(GWAS)database.Two-way MR analysis was used to evaluate the causal relationship between plasma metabolism and KOA.Two-step mediation MR analysis was conducted to evaluate immune cell traits that might have mediating or masking effects.Results After sensitivity analysis and screening,65 plasma metabolites and 35 immune cell traits were found to have causal relationships with KOA(P＜0.05).Mediation analysis found that CD45RA+CD28-CD8br％CD8br had a mediating effect in the causal relationship between three metabolites(2-hydroxyhi-ppurate,X-07765,X-23739)and the risk of KOA.2-hydroxyhippurate(salicylic acid)exerted a masking effect,and the effect ratio was 0.0412.Conclusion A variety of plasma metabolites and immune cell traits are causally related to KOA,which should not be regarded as a simple degenerative joint disease.The protective effect of salicylic acid against KOA may be weakened by its role in inducing the differentiation of Treg cells,which is worthy of more studies.

Objective:To retrospectively analyze the treatment status of tyrosine kinase inhibitors (TKI) in newly diagnosed patients with chronic myeloid leukemia (CML) in China.Methods:Data of chronic phase (CP) and accelerated phase (AP) CML patients diagnosed from January 2006 to December 2022 from 77 centers, ≥18 years old, and receiving initial imatinib, nilotinib, dasatinib or flumatinib-therapy within 6 months after diagnosis in China with complete data were retrospectively interrogated. The choice of initial TKI, current TKI medications, treatment switch and reasons, treatment responses and outcomes as well as the variables associated with them were analyzed.Results:6 893 patients in CP ( n=6 453, 93.6%) or AP ( n=440, 6.4%) receiving initial imatinib ( n=4 906, 71.2%), nilotinib ( n=1 157, 16.8%), dasatinib ( n=298, 4.3%) or flumatinib ( n=532, 7.2%) -therapy. With the median follow-up of 43 ( IQR 22-75) months, 1 581 (22.9%) patients switched TKI due to resistance ( n=1 055, 15.3%), intolerance ( n=248, 3.6%), pursuit of better efficacy ( n=168, 2.4%), economic or other reasons ( n=110, 1.6%). The frequency of switching TKI in AP patients was significantly-higher than that in CP patients (44.1% vs 21.5%, P<0.001), and more AP patients switched TKI due to resistance than CP patients (75.3% vs 66.1%, P=0.011). Multi-variable analyses showed that male, lower HGB concentration and ELTS intermediate/high-risk cohort were associated with lower cytogenetic and molecular responses rate and poor outcomes in CP patients; higher WBC count and initial the second-generation TKI treatment, the higher response rates; Ph + ACA at diagnosis, poor PFS. However, Sokal intermediate/high-risk cohort was only significantly-associated with lower CCyR and MMR rates and the poor PFS. Lower HGB concentration and larger spleen size were significantly-associated with the lower cytogenetic and molecular response rates in AP patients; initial the second-generation TKI treatment, the higher treatment response rates; lower PLT count, higher blasts and Ph + ACA, poorer TFS; Ph + ACA, poorer OS. Conclusion:At present, the vast majority of newly-diagnosed CML-CP or AP patients could benefit from TKI treatment in the long term with the good treatment responses and survival outcomes.