From "wet" matrices to "dry" blood spot sampling strategy: a versatile LC-MS/MS assay for simultaneous monitoring caffeine and its three primary metabolites in preterm infants.

OBJECTIVES: To update traditional "wet" matrices to dried blood spot (DBS) sampling, based on the liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) technique, and develop a method for simultaneous analyzing caffeine and its three primary metabolites (theobromine, paraxanthine, and theophylline), supporting routine therapeutic drug monitoring (TDM) for preterm infants. METHODS: DBS samples were prepared by a two-step quantitative sampling method, i.e., volumetric sampling of a quantitative 10 µL volume of peripheral blood and an 8 mm diameter whole punch extraction by a methanol/water (80/20, v/v) mixture containing 125 mM formic acid. Four paired stable isotope labeled internal standards and a collision energy defect strategy were applied for the method optimization. The method was fully validated following international guidelines and industrial recommendations on DBS analysis. Cross validation with previously developed plasma method was also proceeded. The validated method was then implemented on the TDM for preterm infants. RESULTS: The two-step quantitative sampling strategy and a high recovery extraction method were developed and optimized. The method validation results were all within the acceptable criteria. Satisfactory parallelism, concordance, and correlation were observed between DBS and plasma concentrations of the four analytes. The method was applied to provide routine TDM services to 20 preterm infants. CONCLUSIONS: A versatile LC-MS/MS platform for simultaneous monitoring caffeine and its three primary metabolites was developed, fully validated, and successfully applied into the routine clinical TDM practices. Sampling method switching from "wet" matrices to "dry" DBS will facilitate and support the precision dosing of caffeine for preterm infants.

Icariin alleviates diabetic renal interstitial fibrosis aggravation by inhibiting miR-320a-3p targeting BMP6.

Diabetic nephropathy is a common complication of diabetes, accumulating evidence underscores the pivotal role of tubulointerstitial fibrosis in the progression of diabetic nephropathy. However, the underlying mechanisms remain incompletely understood. Although the mechanisms in diabetic nephropathy fibrosis have been the focus of many studies, only limited information is currently available concerning microRNA regulation in tubulointerstitial fibrosis. In this study, we aimed to investigate the roles of miR-320a-3p and bone morphogenetic protein-6 (BMP6) in tubulointerstitial fibrosis. After inducing fibrosis with high glucose in HK-2 cells, we found that miR-320a-3p is significantly up-regulated, whereas BMP6 is markedly down-regulated. These changes suggest close link between miR-320a-3p and BMP6 in tubulointerstitial fibrosis. To elucidate this phenomenon, miR-320a-3p mimic, inhibitor and siBMP6 were employed. We observed in miR-320a-3p mimic group the fibrosis marker include alpha smooth muscle actin and type I collagen was significantly up-regulated, whereas BMP6 exhibited the opposite trend. Additionally, we found icariin could alleviate tubulointerstitial fibrosis by downregulation the miR-320a-3p expression. In conclusion, miR-320a-3p promotes tubulointerstitial fibrosis during the development of DN by suppressing BMP signal pathway activity via inhibiting BMP6 expression. Suggesting that miR-320a-3p represents a potential therapeutic target for tubulointerstitial fibrosis induced by diabetic nephropathy.

Clinical observations and mechanistic insights of traditional Chinese medicine in the management of diabetic retinopathy.

CONTEXT: Diabetic retinopathy (DR) is one of the leading causes of vision impairment and blindness among diabetic patients globally. Despite advancements in conventional treatments, the quest for more holistic approaches and fewer side effects persists. Traditional Chinese medicine (TCM) has been used for centuries in managing various diseases, including diabetes and its complications. OBJECTIVE: This review evaluated the efficacy and underlying mechanisms of TCM in the management of DR, providing information on its potential integration with conventional treatment methods. METHODS: A comprehensive literature review was conducted using PubMed, Web of Science, and the China National Knowledge Infrastructure (CNKI) with the search terms 'traditional Chinese medicine', 'diabetic retinopathy', 'clinical efficacies' and their combinations. Studies published before 2023 without language restriction were included, focusing on clinical trials and observational studies that assessed the effectiveness of TCM in DR treatment. RESULTS: The review synthesized evidence of empirical traditional Chinese formulas, traditional Chinese patent medicines, and isolated phytochemicals on DR treatment. The key mechanisms identified included the reduction of oxidative stress, inflammation, and neovascularization, as well as the improvement in neurovascular functionality and integrity of the retinal blood barrier. CONCLUSIONS: TCM shows promising potential to manage DR. More large-scale, randomized controlled trials are recommended to validate these findings and facilitate the integration of TCM into mainstream DR treatment protocols.

Advances of traditional Chinese medicine preclinical mechanisms and clinical studies on diabetic peripheral neuropathy.

CONTEXT: Diabetic peripheral neuropathy (DPN) results in an enormous burden and reduces the quality of life for patients. Considering there is no specific drug for the management of DPN, traditional Chinese medicine (TCM) has increasingly drawn attention of clinicians and researchers around the world due to its characteristics of multiple targets, active components, and exemplary safety. OBJECTIVE: To summarize the current status of TCM in the treatment of DPN and provide directions for novel drug development, the clinical effects and potential mechanisms of TCM used in treating DPN were comprehensively reviewed. METHODS: Existing evidence on TCM interventions for DPN was screened from databases such as PubMed, the Cochrane Neuromuscular Disease Group Specialized Register (CENTRAL), and the Chinese National Knowledge Infrastructure Database (CNKI). The focus was on summarizing and analyzing representative preclinical and clinical TCM studies published before 2023. RESULTS: This review identified the ameliorative effects of about 22 single herbal extracts, more than 30 herbal compound prescriptions, and four Chinese patent medicines on DPN in preclinical and clinical research. The latest advances in the mechanism highlight that TCM exerts its beneficial effects on DPN by inhibiting inflammation, oxidative stress and apoptosis, endoplasmic reticulum stress and improving mitochondrial function. CONCLUSIONS: TCM has shown the power latent capacity in treating DPN. It is proposed that more large-scale and multi-center randomized controlled clinical trials and fundamental experiments should be conducted to further verify these findings.

Evidence of traditional Chinese medicine for treating type 2 diabetes mellitus: from molecular mechanisms to clinical efficacy.

CONTEXT: The global prevalence of type 2 diabetes mellitus (T2DM) has increased significantly in recent decades. Despite numerous studies and systematic reviews, there is a gap in comprehensive and up-to-date evaluations in this rapidly evolving field. OBJECTIVE: This review provides a comprehensive and current overview of the efficacy of Traditional Chinese Medicine (TCM) in treating T2DM. METHODS: A systematic review was conducted using PubMed, Web of Science, Wanfang Data, CNKI, and Medline databases, with a search timeframe extending up to November 2023. The search strategy involved a combination of subject terms and free words in English, including 'Diabetes,' 'Traditional Chinese Medicine,' 'TCM,' 'Hypoglycemic Effect,' 'Clinical Trial,' and 'Randomized Controlled Trial.' The studies were rigorously screened by two investigators, with a third investigator reviewing and approving the final selection based on inclusion and exclusion criteria. RESULTS: A total of 108 relevant papers were systematically reviewed. The findings suggest that TCMs not only demonstrate clinical efficacy comparable to existing Western medications in managing hypoglycemia but also offer fewer adverse effects and a multitarget therapeutic approach. Five main biological mechanisms through which TCM treats diabetes were identified: improving glucose transport and utilization, improving glycogen metabolism, promoting GLP-1 release, protecting pancreatic islets from damage, and improving intestinal flora. CONCLUSIONS: TCM has demonstrated significant protective effects against diabetes and presents a viable option for the prevention and treatment of T2DM. These findings support the further exploration and integration of TCM into broader diabetes management strategies.

Shenqi Fuzheng injection restores the sensitivity to gefitinib in non-small cell lung cancer by inhibiting the IL-22/STAT3/AKT pathway.

CONTEXT: Non-small cell lung cancer (NSCLC) is the most common type of lung cancer. Gefitinib is a first-line treatment for NSCLC. However, its effectiveness is hindered by the development of drug resistance. At present, Shenqi Fuzheng injection (SFI) is widely accepted as an adjuvant therapy in NSCLC. OBJECTIVE: This study investigates the molecular mechanism of SFI when combined with gefitinib in regulating cell progression among EGFR-TKI-resistant NSCLC. MATERIALS AND METHODS: We established gefitinib-resistant PC9-GR cells by exposing gefitinib escalation from 10 nM with the indicated concentrations of SFI in PC9 cells (1, 4, and 8 mg/mL). Quantitative real-time polymerase chain reaction was performed to assess gene expression. PC9/GR and H1975 cells were treated with 50 ng/mL of interleukin (IL)-22 alone or in combination with 10 mg/mL of SFI. STAT3, p-STAT3, AKT, and p-AKT expression were evaluated using Western blot. The effects on cell proliferation, clonogenicity, and apoptosis in NSCLC cells were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), colony formation and flow cytometry assays. RESULTS: SFI treatment alleviated the development of gefitinib resistance in NSCLC. PC9/GR and H1975 cells treated with SFI significantly exhibited a reduction in IL-22 protein and mRNA overexpression levels. SFI effectively counteracted the activation of the STAT3/AKT signaling pathway induced by adding exogenous IL-22 to PC9/GR and H1975 cells. Moreover, IL-22 combined with gefitinib markedly increased cell viability while reducing apoptosis. In contrast, combining SFI with gefitinib and the concurrent treatment of SFI with gefitinib and IL-22 demonstrated the opposite effect. DISCUSSION AND CONCLUSION: SFI can be a valuable therapeutic option to address gefitinib resistance in NSCLC by suppressing the IL-22/STAT3/AKT pathway.

Cholesterol promotes EGFR-TKIs resistance in NSCLC by inducing EGFR/Src/Erk/SP1 signaling-mediated ERRα re-expression.

BACKGROUND: The use of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) brings remarkable benefits for the survival of patients with advanced NSCLC harboring EGFR mutations. Unfortunately, acquired resistance seems to be inevitable and limits the application of EGFR-TKIs in clinical practice. This study reported a common molecular mechanism sustaining resistance and potential treatment options to overcome EGFR-TKIs resistance. METHODS: EGFR-TKIs resistant NSCLC cells were established and confirmed by MTT assay. Cholesterol content was detected and the promotional function of cholesterol on NSCLC growth was determined in vivo. Then, we identified ERRα expression as the downstream factor of cholesterol-mediated drug resistance. To dissect the regulatory mechanism, we conducted experiments, including immunofluorescence, co-immunoprecipitation, luciferase reporter assay and chromatin immunoprecipitation assay. RESULTS: Long-term exposure to EGFR-TKIs generate drug resistance with the characteristic of cholesterol accumulation in lipid rafts, which promotes EGFR and Src to interact and lead EGFR/Src/Erk signaling reactivation-mediated SP1 nuclear translocation and ERRα re-expression. Further investigation identifies ERRα as a target gene of SP1. Functionally, re-expression of ERRα sustains cell proliferation by regulating ROS detoxification process. Lovastatin, a drug used to decrease cholesterol level, and XCT790, an inverse agonist of ERRα, overcome gefitinib and osimertinib resistance both in vitro and in vivo. CONCLUSIONS: Our study indicates that cholesterol/EGFR/Src/Erk/SP1 axis-induced ERRα re-expression promotes survival of gefitinib and osimertinib-resistant cancer cells. Besides, we demonstrate the potential of lowing cholesterol and downregulation of ERRα as effective adjuvant treatment of NSCLC.

Population pharmacokinetic modeling of caffeine in preterm infants with apnea of prematurity: New findings from concomitant erythromycin and AHR genetic polymorphisms.

Current standard-dose caffeine therapy results in significant intersubject variability. The aims of this study were to develop and evaluate population pharmacokinetic (PPK) models of caffeine in preterm infants through comprehensive screening of covariates and then to propose model-informed precision dosing of caffeine for this population. A total of 129 caffeine concentrations from 96 premature neonates were incorporated into this study. Comprehensive medical record and genotype data of these neonates were collected for analysis. PPK modeling was performed by a nonlinear mixed effects modeling program (NONMEM). Final models based on the current weight (CW) or body surface area (BSA) were evaluated via multiple graphic and statistical methods. The model-informed dosing regimen was performed through Monte Carlo simulations. In addition to CW or BSA, postnatal age, coadministration with erythromycin (ERY), and aryl hydrocarbon receptor coding gene (AHR) variant (rs2158041) were incorporated into the final PPK models. Multiple evaluation results showed satisfactory prediction performance and stability of the CW- and BSA-based models. Monte Carlo simulations demonstrated that trough concentrations of caffeine in preterm infants would be affected by concomitant ERY therapy and rs2158041 under varying dose regimens. For the first time, ERY and rs2158041 were found to be associated with the clearance of caffeine in premature infants. Similar predictive performance and stability were obtained for both CW- and BSA-based PPK models. These findings provide novel insights into caffeine precision therapy for preterm infants.

Development and Validation of Machine Learning Models to Predict Epidermal Growth Factor Receptor Mutation in Non-Small Cell Lung Cancer: A Multi-Center Retrospective Radiomics Study.

OBJECTIVE: To develop and validate a generalized prediction model that can classify epidermal growth factor receptor (EGFR) mutation status in non-small cell lung cancer patients. METHODS: A total of 346 patients (296 in the training cohort and 50 in the validation cohort) from four centers were included in this retrospective study. First, 1085 features were extracted using IBEX from the computed tomography images. The features were screened using the intraclass correlation coefficient, hypothesis tests and least absolute shrinkage and selection operator. Logistic regression (LR), decision tree (DT), random forest (RF), and support vector machine (SVM) were used to build a radiomics model for classification. The models were evaluated using the following metrics: area under the curve (AUC), calibration curve (CAL), decision curve analysis (DCA), concordance index (C-index), and Brier score. RESULTS: Sixteen features were selected, and models were built using LR, DT, RF, and SVM. In the training cohort, the AUCs was .723, .842, .995, and .883; In the validation cohort, the AUCs were .658, 0567, .88, and .765. RF model with the best AUC, its CAL, C-index (training cohort=.998; validation cohort=.883), and Brier score (training cohort=.007; validation cohort=0.137) showed a satisfactory predictive accuracy; DCA indicated that the RF model has better clinical application value. CONCLUSION: Machine learning models based on computed tomography images can be used to evaluate EGFR status in patients with non-small cell lung cancer, and the RF model outperformed LR, DT, and SVM.

Relationship Between Neutrophil-To-Lymphocyte Ratio and Brain Metastasis in Non-Small Cell Lung Cancer Patients.

OBJECTIVE: To investigate the relationship between the neutrophil-to-lymphocyte ratio (NLR) of patients with non-small cell lung cancer (NSCLC) and their risk of developing brain metastases after adjusting for confounding factors. METHODS: A retrospective observational study of the general data of patients with NSCLC diagnosed from January 2016 to December 2020. Multivariate logistic regression was used to calculate the dominance ratio (OR) with 95% confidence interval (CI) for NLR and NSCLC brain metastases with subgroup analysis. Generalized summation models and smoothed curve fitting were used to identify whether there was a nonlinear relationship between them. RESULTS: In all 3 models, NLR levels were positively correlated with NSCLC brain metastasis (model 1: OR: 1.12, 95% CI: 1.01-1.23, P = .025; model 2: OR: 1.16, 95% CI: 1.04-1.29, P = .007; model 3: OR: 1.20, 95% CI: 1.05-1.37, P = .006). Stratified analysis showed that this positive correlation was present in patients with adenocarcinoma (LUAD) and female patients (LUAD: OR: 1.30, 95% CI: 1.10-1.54, P = .002; female: OR: 1.52, 95% CI: 1.05-2.20, P = .026), while there was no significant correlation in patients with squamous carcinoma (LUSC) and male patients (LUSC: OR:0.76,95% CI:0.38- 1.53, P = .443; male: OR:1.13, 95% CI:0.95-1.33, P = .159). CONCLUSION: This study showed that elevated levels of NLR were independently associated with an increased risk of developing brain metastases in patients with NSCLC, and that this correlation varied by TYPE and SEX, with a significant correlation in female patients and patients with LUAD.

MiR-21-3p triggers cardiac fibroblasts pyroptosis in diabetic cardiac fibrosis via inhibiting androgen receptor.

AIMS/HYPOTHESIS: MicroRNA-21 has been implicated in diabetic complication, including diabetic cardiomyopathy. However, there is limited information regarding the biological role of the miR-21 passenger strand (miR-21-3p) in diabetic cardiac fibrosis. The aim of this study was to investigate the role of miR-21-3p and its target androgen receptor in STZ-induced diabetic cardiac fibrosis. METHODS: The pathological changes and collagen depositions was analyzed by HE, Sirius Red staining and Masson's Trichrome Staining. MiR-21-3p, AR, NLRP3, caspase1 and collagen I expression were analyzed by western blotting, immunohistochemistry, immunofluorescence, qRT-PCR, miR one step qRT-PCR, respectively. A luciferase reporter assay was used to verify the interaction between miR-21 and the 3' untranslated region (3'UTR) of AR. RESULTS: Our results indicated that miR-21-3p level was up-regulated, while AR was decreased in STZ-induced diabetic cardiac fibrosis tissues and cardiac fibroblast. High glucose triggers cardiac fibroblasts pyroptosis and collagen deposition. Gain-of-function and loss-of-function assays demonstrated that miR-21-3p mediated the crucial role in diabetic cardiac fibrosis. Our results show that miR-21-3p bound to the 3'UTR of AR post-transcriptionally repressed its expression. We also found AR, which regulates cardiac fibroblasts pyroptosis and collagen deposition through caspase1 signaling. CONCLUSIONS: /interpretation: Taken together, our study showed that miR-21-3p aggravates STZ-induced diabetic cardiac fibrosis through the caspase1 pathways by suppressing AR expression.

PPARα agonist fenofibrate relieves acquired resistance to gefitinib in non-small cell lung cancer by promoting apoptosis via PPARα/AMPK/AKT/FoxO1 pathway.

Recent studies show that intracellular accumulation of cholesterol leads to acquired resistance to gefitinib in non-small cell lung cancer (NSCLC) cells. In this study we investigated how to regulate the cholesterol levels in gefitinib-resistant NSCLC cells. We showed that intracellular cholesterol levels in gefitinib-resistant cell lines (PC-9/GR, H1975, H1650, and A549) were significantly higher than that in gefitinib-sensitive cell line (PC-9). Treatment with gefitinib (5 µM) significantly increased intracellular cholesterol levels in PC-9/GR, H1975, and H1650 cells. Gefitinib treatment downregulated the expression of PPARα, LXRα, and ABCA1, leading to dysregulation of cholesterol efflux pathway. We found that a lipid-lowering drug fenofibrate (20, 40 µM) dose-dependently increased the expression of PPARα, LXRα, and ABCA1, decreased the intracellular cholesterol levels, and enhanced the antiproliferative effects of gefitinib in PC-9/GR, H1975, and H1650 cells. We revealed that fenofibrate increased the gefitinib-induced apoptosis via regulating the key proteins involved in the intrinsic apoptosis pathway. In PC-9/GR, H1975 and H1650 cells, fenofibrate dose-dependently increased the expression of AMPK, FoxO1, and decreased the expression of AKT, which were remarkably weakened by knockdown of PPARα. In PC-9/GR cell xenograft mice, combined administration of gefitinib (25 mg · kg-1 · d-1) and fenofibrate (100 mg · kg-1 · d-1) caused remarkable inhibition on tumor growth as compared to treatment with either drug alone. All the results suggest that fenofibrate relieves acquired resistance to gefitinib in NSCLC by promoting apoptosis via regulating PPARα/AMPK/AKT/FoxO1 pathway. We propose that combination of gefitinib and fenofibrate is a potential strategy for overcoming the gefitinib resistance in NSCLC.

Cerebrospinal fluid metabolic profiling reveals divergent modulation of pentose phosphate pathway by midazolam, propofol and dexmedetomidine in patients with subarachnoid hemorrhage: a cohort study.

BACKGROUND: Agitation is common in subarachnoid hemorrhage (SAH), and sedation with midazolam, propofol and dexmedetomidine is essential in agitation management. Previous research shows the tendency of dexmedetomidine and propofol in improving long-term outcome of SAH patients, whereas midazolam might be detrimental. Brain metabolism derangement after SAH might be interfered by sedatives. However, how sedatives work and whether the drugs interfere with patient outcome by altering cerebral metabolism is unclear, and the comprehensive view of how sedatives regulate brain metabolism remains to be elucidated. METHODS: For cerebrospinal fluid (CSF) and extracellular space of the brain exchange instantly, we performed a cohort study, applying CSF of SAH patients utilizing different sedatives or no sedation to metabolomics. Baseline CSF metabolome was corrected by selecting patients of the same SAH and agitation severity. CSF components were analyzed to identify the most affected metabolic pathways and sensitive biomarkers of each sedative. Markers might represent the outcome of the patients were also investigated. RESULTS: Pentose phosphate pathway was the most significantly interfered (upregulated) pathway in midazolam (p = 0.0000107, impact = 0.35348) and propofol (p = 0.00000000000746, impact = 0.41604) groups. On the contrary, dexmedetomidine decreased levels of sedoheptulose 7-phosphate (p = 0.002) and NADP (p = 0.024), and NADP is the key metabolite and regulator in pentose phosphate pathway. Midazolam additionally augmented purine synthesis (p = 0.00175, impact = 0.13481) and propofol enhanced pyrimidine synthesis (p = 0.000203, impact = 0.20046), whereas dexmedetomidine weakened pyrimidine synthesis (p = 0.000000000594, impact = 0.24922). Reduced guanosine diphosphate (AUC of ROC 0.857, 95%CI 0.617-1, p = 0.00506) was the significant CSF biomarker for midazolam, and uridine diphosphate glucose (AUC of ROC 0.877, 95%CI 0.631-1, p = 0.00980) for propofol, and succinyl-CoA (AUC of ROC 0.923, 95%CI 0.785-1, p = 0.000810) plus adenosine triphosphate (AUC of ROC 0.908, 95%CI 0.6921, p = 0.00315) for dexmedetomidine. Down-regulated CSF succinyl-CoA was also associated with favorable outcome (AUC of ROC 0.708, 95% CI: 0.524-0.865, p = 0.029333). CONCLUSION: Pentose phosphate pathway was a crucial target for sedatives which alter brain metabolism. Midazolam and propofol enhanced the pentose phosphate pathway and nucleotide synthesis in poor-grade SAH patients, as presented in the CSF. The situation of dexmedetomidine was the opposite. The divergent modulation of cerebral metabolism might further explain sedative pharmacology and how sedatives affect the outcome of SAH patients.

Therapeutic drug monitoring of caffeine and its primary metabolites in plasma using LC-ESI-MS/MS for apnea of prematurity treatment: Evaluation of ultrapure water as a surrogate matrix.

The growing evidence has endorsed the view that therapeutic drug monitoring of caffeine for apnea of prematurity is helpful for dose tailoring when the therapeutic response is lacking or toxicity is suspected. However, plasma without caffeine is difficult to obtain. Therefore, a method was developed and validated to measure caffeine and its three primary metabolites (paraxanthine, theobromine and theophylline) using LC-ESI-MS/MS in human plasma and several surrogate matrices. The chromatographic separation of analytes was finally achieved on a Waters Symmetry C18 (4.6 × 75 mm, 3.5 µm) column. Several strategies were successfully applied to overcome the matrix effects: (a) appropriate dilution for sample cleanup; (b) a starting lower proportion of organic phase; and (c) multiple individual stable-labeled isotopic internal standards. The parallelism between the authentic matrix and surrogate matrices was convincing. The recovery of the analytes in both human plasma and rat plasma was acceptable over the linear range (0.500-50.0 µg/ml for caffeine and 0.0100-1.00 µg/ml for three metabolites). The method was successfully applied in 118 samples from 74 preterm infants with apnea of prematurity. The rat plasma or ultrapure water as a surrogate matrix is worthy of recommendation for routine therapeutic drug monitoring of caffeine.

DNMT1 deregulation of SOCS3 axis drives cardiac fibroblast activation in diabetic cardiac fibrosis.

Cardiac fibrosis is one of the main pathological manifestations of diabetic cardiomyopathy (DCM). Cardiac fibroblast activation is a key effector of cells resulting in diabetic cardiac fibrosis. However, the underlying mechanism of cardiac fibroblast activation and diabetic cardiac fibrosis remains unclear. Accumulating evidence suggests that DNA methylation alterations play a central role in cardiac fibroblast activation. In this study, we demonstrated that DNA methyltransferase 1 (DNMT1)-mediated suppression of cytokine signaling 3 (SOCS3) promoter hypermethylation leads to downregulation of SOCS3 expression in diabetic cardiac fibrosis. High glucose-induced expression of DNMT1 was increased in cardiac fibroblasts, while the expression of SOCS3 was decreased. Downregulation of SOCS3 facilitated activation of STAT3 to promote cardiac fibroblast activation and collagen deposition. Genetic or pharmacological inactivation of DNMT1 reversed the activated phenotype of cardiac fibroblasts. Clinically, we observed a significant inverse correlation between DNMT1 and SOCS3 expression levels, and loss of SOCS3 expression or increased expression of DNMT1. Taken together, these findings identify DNMT1 silencing of SOCS3 axis as a driver of cardiac fibroblast activation in diabetic cardiac fibrosis. These results provide a scientific and new explanation of the underlying mechanism of diabetic cardiac fibrosis.

Icariin ameliorates endothelial dysfunction in type 1 diabetic rats by suppressing ER stress via the PPARα/Sirt1/AMPKα pathway.

Icariin (ICA), as a flavonoid glycoside, is associated with the improvement of vascular complications in diabetes. However, its protective mechanisms remain to be well-established. Here, we tested the hypothesis that ICA attenuates vascular endothelial dysfunction by inhibiting endoplasmic reticulum (ER) stress in type 1 diabetes. In streptozotocin-induced diabetic rats, ICA positively affected acetylcholine-induced vasodilation and phenylephrine-induced vasoconstriction in aortas. ICA treatment significantly attenuated ER stress in diabetic rats and high-glucose induced human umbilical vein endothelial cells. Incubation with ICA in vitro attenuated vascular reactivity in diabetic rats, which was blocked by the ER stress inducer, and peroxisome proliferator-activated receptor α (PPARα), sirtuin1 (Sirt1), or AMP-activated protein kinase-α (AMPKα) inhibitors. Western blot showed that ICA activated the PPARα/Sirt1/AMPKα pathway, which contributed to reducing ER stress and activating endothelial nitric oxide synthase in vivo and vitro. Our results implicate that ICA normalizes ER stress to attenuate endothelial dysfunction by the regulation of the PPARα/Sirt1/AMPKα pathway.

DNA methyltransferase-1 inactivation of androgen receptor axis triggers homocysteine induced cardiac fibroblast autophagy in diabetic cardiac fibrosis.

Diabetic cardiac fibrosis is one of the main pathological manifestations of diabetic cardiomyopathy (DCM). Cardiac fibroblast autophagy plays critical roles in diabetic cardiac fibrosis, however, the underlying mechanism of cardiac fibroblast autophagy and diabetic cardiac fibrosis still largely unknown. The aim of the study was to investigate the mechanism of DNMT1 mediated DNA methylation alterations control cardiac fibroblast autophagy in diabetic cardiac fibrosis. We employed streptozotocin (STZ)-induced rats DCM, DCM patient and Hcy induced cardiac fibroblast autophagy. Heart tissue sections were stained with H&E, Sirius Red and Masson's trichrome stain. The expression of DNMT1, AR, Collagen genes mRNA was detected by qRT-PCR. MSP and BSP detected the methylation status of the AR promoter. The expression of DNMT1, AR, Collagen and autophagy-related proteins were detected by Western blotting, Immunofluorescence, Immunohistochemistry. Gain and loss function of AR and DNMT1 in cardiac fibroblast was analyzed. DNMT1 inhibition or knockdown elevated the expression of AR in cardiac fibroblast. Furthermore, we found that AR negatively regulation of Hcy induced cardiac fibroblast autophagy. We demonstrated that DNMT1 enhances cardiac fibroblast autophagy in diabetic cardiac fibrosis through inhibiting AR axis. In conclusion, our results provide new insight into the DNMT1 inactivation of AR axis triggers cardiac fibroblast autophagy in diabetic cardiac fibrosis.

Icariin Attenuates Diabetic Cardiomyopathy and Downregulates Extracellular Matrix Proteins in Heart Tissue of Type 2 Diabetic Rats.

OBJECTIVE: Diabetic cardiomyopathy (DCM) is a serious complication of type 2 diabetes mellitus (T2DM), resulting in unfavorable prognosis. Icariin (ICA) is a major flavonoid isolated from the traditional oriental herbal medicine Epimedium that has been recently proved to show potential therapeutic efficacy on T2DM. The aim of this study was to investigate the underlying mechanism of how ICA improved DCM in rat models. METHODS: To corroborate myocardial improvement by ICA, we managed to establish the T2DM rat model by streptozotocin (STZ) administration and high-glucose-high-fat diet. RESULTS: The rats with T2DM showed severe insulin resistance, left ventricular dysfunction, aberrant lipids deposition, cardiac inflammation, and fibrosis compared with the control group. All these pathological symptoms were ameliorated by the treatment of ICA. The levels of extracellular matrix proteins of heart tissue significantly declined in ICA-treated rats. CONCLUSION: ICA may exert as a protector in T2DM-induced DCM by reducing extracellular matrix proteins in the heart tissue, implicating its potential role for the treatment of human DCM.

High cholesterol in lipid rafts reduces the sensitivity to EGFR-TKI therapy in non-small cell lung cancer.

Overcoming EGFR-TKI resistant which has the initial enthusiasm over substantial clinical responses is a formidable challenge on nowadays. In this study, we showed that cholesterol level in lipid rafts in gefitinib resistant non-small cell lung cancer (NSCLC) cell lines was remarkably higher than gefitinib sensitive cell line, and depletion of cholesterol increased gefitinib sensitivity. Furthermore, cholesterol-depleted enhanced gefitinib inhibit phosphorylation of EGFR, Akt-1, MEK1/2, and ERK1/2 and these were reversed in cholesterol add-back experiments. Gefitinib resistant cell lines showed high affinity of gefitinib and EGFR when cholesterol was depleted. Therefore, targeting cholesterol combined with EGFR-TKI is potentially a novel therapeutic strategy for gefitinib resistant treatment.

Therapeutic Drug Monitoring of Voriconazole in Children from a Tertiary Care Center in China.

Voriconazole is a broad-spectrum triazole antifungal and the first-line treatment for invasive aspergillosis (IA). The aim of this research was to study the dose adjustments of voriconazole as well as the affecting factors influencing voriconazole trough concentrations in Asian children to optimize its daily administration. Clinical data were analyzed of inpatients 2 to 14 years old who were subjected to voriconazole trough concentration monitoring from 1 June 2015 to 1 December 2017. A total of 138 voriconazole trough concentrations from 42 pediatric patients were included. Voriconazole trough concentrations at steady state ranged from 0.02 to 9.35 mg/liter, with high inter- and intraindividual variability. Only 50.0% of children achieved the target range (1.0 to 5.5 mg/liter) at initial dosing, while 35.7% of children were subtherapeutic, and 14.3% of children were supratherapeutic at initial dosing. There was no correlation between initial trough concentrations and initial dosing. A total of 28.6% of children (12/42) received an adjusted dose according to trough concentrations. Children <6, 6 to 12, and >12 years old required a median oral maintenance dose to achieve the target range of 11.1, 7.2, and 5.3 mg/kg twice daily, respectively (P = 0.043). The average doses required to achieved the target range were 7.7 mg/kg and 5.6 mg/kg, respectively, and were lower than the recommended dosage (P = 0.033 and 0.003, respectively). Affecting factors such as administration routes and coadministration with proton pump inhibitors (PPIs) explained 55.3% of the variability in voriconazole exposure. Therapeutic drug monitoring (TDM) of voriconazole could help to individualize antifungal therapy for children and provide guidelines for TDM and dosing optimization in Asian children.