Protein diaphanous homolog 1 (Diaph1) promotes myofibroblastic activation of hepatic stellate cells by regulating Rab5a activity and TGFß receptor endocytosis.

TGFß induces the differentiation of hepatic stellate cells (HSCs) into tumor-promoting myofibroblasts but underlying mechanisms remain incompletely understood. Because endocytosis of TGFß receptor II (TßRII), in response to TGFß stimulation, is a prerequisite for TGF signaling, we investigated the role of protein diaphanous homolog 1 (known as Diaph1 or mDia1) for the myofibroblastic activation of HSCs. Using shRNA to knockdown Diaph1 or SMIFH2 to target Diaph1 activity of HSCs, we found that the inactivation of Diaph1 blocked internalization and intracellular trafficking of TßRII and reduced SMAD3 phosphorylation induced by TGFß1. Mechanistic studies revealed that the N-terminal portion of Diaph1 interacted with both TßRII and Rab5a directly and that Rab5a activity of HSCs was increased by Diaph1 overexpression and decreased by Diaph1 knockdown. Additionally, expression of Rab5aQ79L (active Rab5a mutant) increased whereas the expression of Rab5aS34N (inactive mutant) reduced the endosomal localization of TßRII in HSCs compared to the expression of wild-type Rab5a. Functionally, TGFß stimulation promoted HSCs to express tumor-promoting factors, and α-smooth muscle actin, fibronection, and CTGF, markers of myofibroblastic activation of HSCs. Targeting Diaph1 or Rab5a suppressed HSC activation and limited tumor growth in a tumor implantation mouse model. Thus, Dipah1 and Rab5a represent targets for inhibiting HSC activation and the hepatic tumor microenvironment.

Single-Atom Pd Nanozyme for Ferroptosis-Boosted Mild-Temperature Photothermal Therapy.

Photothermal therapy (PTT) is an extremely promising tumor therapeutic modality. However, excessive heat inevitably injures normal tissues near tumors, and the damage to cancer cells caused by mild hyperthermia is easily repaired by stress-induced heat shock proteins (HSPs). Thus, maximizing the PTT efficiency and minimizing the damage to healthy tissues simultaneously by adopting appropriate therapeutic temperatures is imperative. Herein, an innovative strategy is reported: ferroptosis-boosted mild PTT based on a single-atom nanozyme (SAzyme). The Pd SAzyme with atom-economical utilization of catalytic centers exhibits peroxidase (POD) and glutathione oxidase (GSHOx) mimicking activities, and photothermal conversion performance, which can result in ferroptosis featuring the up-regulation of lipid peroxides (LPO) and reactive oxygen species (ROS). The accumulation of LPO and ROS provides a powerful approach for cleaving HSPs, which enables Pd SAzyme-mediated mild-temperature PTT.

p300 Acetyltransferase Is a Cytoplasm-to-Nucleus Shuttle for SMAD2/3 and TAZ Nuclear Transport in Transforming Growth Factor ß-Stimulated Hepatic Stellate Cells.

Nuclear translocation of mothers against decapentaplegic homolog 2/3 (SMAD2/3), core transcription factors of transforming growth factor ß (TGF-ß) signaling, is critical for hepatic stellate cell (HSC) differentiation into metastasis-promoting myofibroblasts. SMAD2/3 have multiple coactivators, including WW domain-containing transcription regulator protein 1 (WWTR1 or TAZ) and p300 acetyltransferase. In the nucleus, TAZ binds to SMAD2/3 to prevent SMAD2/3 nuclear export. However, how TAZ and SMAD2/3 enter the nucleus remains poorly understood because neither contains a nuclear localization signal (NLS), an amino acid sequence tagging proteins for nuclear transport. p300 is an NLS-containing large scaffold protein, so we hypothesized that SMAD2/3 and TAZ may undergo nuclear import through complexing with p300. Coimmunoprecipitation, immunofluorescence, and nuclear fractionation assays revealed that TGF-ß1 promoted binding of SMAD2/3 and TAZ to p300 and that p300 inactivation disrupted TGF-ß1-mediated SMAD2/3 and TAZ nuclear accumulation. Deleting the p300 NLS blocked TGF-ß1-induced SMAD2/3 and TAZ nuclear transport. Consistently, p300 inactivation suppressed TGF-ß1-mediated HSC activation and transcription of genes encoding tumor-promoting factors, such as connective tissue growth factor, Tenascin C, Periostin, platelet-derived growth factor C, and fibroblast growth factor 2, as revealed by microarray analysis. Chromatin immunoprecipitation-real-time quantitative PCR showed that canonical p300-mediated acetylation of histones also facilitated transcription in response to TGF-ß1 stimulation. Interestingly, although both TGF-ß1-mediated and stiffness-mediated HSC activation require p300, comparison of gene expression data sets revealed that transcriptional targets of TGF-ß1 were distinct from those of stiffness-p300 mechanosignaling. Lastly, in tumor/HSC coinjection and intrasplenic tumor injection models, targeting p300 of activated-HSC/myofibroblasts by C646, short hairpin RNA, or cre-mediated gene disruption reduced tumor and liver metastatic growth in mice. Conclusion: p300 facilitates TGF-ß1-stimulated HSC activation by both noncanonical (cytoplasm-to-nucleus shuttle for SMAD2/3 and TAZ) and canonical (histone acetylation) mechanisms. p300 is an attractive target for inhibiting HSC activation and the prometastatic liver microenvironment.

Hyperthermic intraperitoneal chemotherapy plus high-frequency diathermic therapy followed by intravenous chemotherapy versus intravenous chemotherapy alone for postoperative adjuvant treatment of gastrointestinal cancer: a comparative research study.

PURPOSE: To evaluate the therapeutic efficacy and toxicity of hyperthermic intraperitoneal chemotherapy (HIPEC) plus high-frequency diathermic therapy (HFDT) followed by intravenous chemotherapy vs intravenous chemotherapy alone for adjuvant treatment of postoperative gastrointestinal neoplasms. METHODS: Fifty-two gastrointestinal carcinoma patients who were radically operated were enrolled and divided into the treatment group and the control group. In the treatment group, 25 patients were treated with combination of HIPEC+HFDT and subsequent intravenous chemotherapy, while in the control group 27 patients received intravenous chemotherapy alone. Post-therapeutic complications and adverse reactions, time to progression (TTP) and overall survival (OS) were compared between these two groups. RESULTS: Difference in toxic reactions between the two groups was not statistically significant (p>0.05). Postoperative progression- free survival (PFS) rate at 12 and 40 months after radical surgery was 72.0 and 54.0% respectively in the treatment group, and 65.8 and 11.5% respectively in the control group (p=0.108). TTP was statistically significantly longer in the treatment group than in the control group (median TTP 40.1 vs 18.5 months, p=0.027). Postoperative OS at 12 and 20 months after radical surgery was 88.0 and 78.0% respectively in the treatment group and 92.6 and 72.7% in the control group, without significant difference. CONCLUSION: After radical surgery, combination of HIPEC+HFDT and subsequent intravenous chemotherapy brings about superior PFS compared with intravenous adjuvant chemotherapy alone, while having no more complications and adverse reactions.

Effect of arctiin on glomerular filtration barrier damage in STZ-induced diabetic nephropathy rats.

Diabetic nephropathy (DN) is the major life-threatening complication of diabetes. Abnormal permeability of glomerular basement membrane plays an important role in DN pathogenesis. This study was performed to assess the effect of arctiin, the lignan constituent from Arctium lappa L., on metabolic profile and aggravation of renal lesions in a rat model of streptozotocin (STZ)-induced DN. STZ-induced diabetic rats were treated with arctiin at the dosage of 60 or 40 mg/kg/day via intraperitoneal injection for 8 weeks. Blood glucose and 24-h urinary albumin content were measured, and kidney histopathological changes were monitored. RT-PCR and immunohistochemistry were used to detect the mRNA and protein levels of nephrin, podocin and heparanase (HPSE) in the kidney cortex of rats, respectively. Treatment with arctiin significantly decreased the levels of 24-h urinary albumin, prevented the sclerosis of glomeruli and effectively restored the glomerular filtration barrier damage by up-regulating the expression of nephrin and podocin and down-regulating HPSE level. Our studies suggest that arctiin might be beneficial for DN. The effects of arctiin on attenuating albuminuria and glomerulosclerosis are possibly mediated by regulating the expression of nephrin and podocin and HPSE in STZ-induced diabetic rats.

Core-Shell Au@Pd Bimetallic Nanozyme Mediated Mild Photothermal Therapy through Reactive Oxygen Species-Regulating Tumor Thermoresistance.

Mild photothermal therapy (mPTT), which circumvents the limitations of conventional photothermal therapy, is emerging and exhibits remarkable potential in clinical applications. Nevertheless, mPTT is not able to efficiently eradicate tumors because its therapeutic efficacy is dramatically diminished by stress-induced heat shock proteins (HSP). Herein, a core-shell structured Au@Pd (AP) bimetallic nanozyme was fabricated for reactive oxygen species (ROS) augmentation-induced mPTT. The nanocatalytic AP nanozymes with photothermal conversion performance harbor multienzymatic (catalase, oxidase, and peroxidase) activities to induce ROS storm formation. The generated ROS could suppress the heat-defense response of tumor cells by cleaving HSP. Overall, our work highlights a ROS-regulating strategy to counteract hyperthermia-associated resistance in mPTT.

Related Factor Analysis and Nursing Strategies of Diarrhea in Critically Ill Patients with Enteral Nutrition.

Objective: To explore the related factors of diarrhea in critically ill patients with enteral nutrition (EN) in the intensive care unit (ICU). Methods: This single-center retrospective study analyzed the occurrence of intolerant diarrhea in ICU patients receiving EN treatment in our hospital. By collecting clinical data, univariate and multivariate logistic regression analysis was used to screen the risk factors for diarrhea. Results: Among 120 patients included in the study, 68 (48.33%) had diarrhea. Age (OR = 2.599, P = 0.027), use of antibiotics (OR = 3.496, P = 0.007), ICU hospitalization time (OR = 1.311, P = 0.001), and mechanical ventilation time (OR = 1.273, P = 0.035) were all independent risk factors for diarrhea in EN. Conclusion: Older age, frequent use of antibiotics, long ICU stay, and mechanical ventilation time can lead to diarrhea in ICU patients receiving EN treatment. It is necessary to effectively analyze the above independent factors and implement targeted interventions to improve the incidence of diarrhea in patients.

Surface-Functionalized NdVO4:Gd3+ Nanoplates as Active Agents for Near-Infrared-Light-Triggered and Multimodal-Imaging-Guided Photothermal Therapy.

Development of nanotheranostic agents with near-infrared (NIR) absorption offers an effective tool for fighting malignant diseases. Lanthanide ion neodymium (Nd3+)-based nanomaterials, due to the maximum absorption at around 800 nm and unique optical properties, have caught great attention as potential agents for simultaneous cancer diagnosis and therapy. Herein, we employed an active nanoplatform based on gadolinium-ion-doped NdVO4 nanoplates (NdVO4:Gd3+ NPs) for multiple-imaging-assisted photothermal therapy. These NPs exhibited enhanced NIR absorption and excellent biocompatibility after being grafted with polydopamine (pDA) and bovine serum albumin (BSA) layers on their surface. Upon expose to an 808 nm laser, these resulting NPs were able to trigger hyperthermia rapidly and cause photo-destruction of cancer cells. In a xenograft tumor model, tumor growth was also significantly inhibited by these photothermal agents under NIR laser irradiation. Owing to the multicomponent nanostructures, we demonstrated these nanoagents as being novel contrast agents for in vivo magnetic resonance (MR) imaging, X-ray computed tomography (CT), photoacoustic (PA) imaging, and second biological window fluorescent imaging of tumor models. Thus, we believe that this new kind of nanotherapeutic will benefit the development of emerging nanosystems for biological imaging and cancer therapy.

Nitric oxide-mediated regulation of mitochondrial protective autophagy for enhanced chemodynamic therapy based on mesoporous Mo-doped Cu9S5 nanozymes.

The depletion of reactive oxygen species (ROS) by glutathione (GSH) and oxidative stress induced protective autophagy severely impaired the therapeutic effect of chemodynamic therapy (CDT). Therefore, how to construct a CDT treatment nanosystem with high yield and full utilization of ROS in tumor site is the main issue of CDT. Herein, a multifunctional cascade bioreactor based on mesoporous Mo-doped Cu9S5 (m-MCS) nanozymes loaded with L-Arginine (LA), abbreviated as m-MCS@LA, is constructed for realizing enhanced CDT promoted by ultrasound (US) triggered gas therapy. The m-MCS based on the catalytic performance of multivalent metal ions, which were served as nanozymes, exhibit enhanced Fenton-like and glutathione (GSH) peroxidase-like activities in comparison to Cu9S5 nanoparticles without Mo-doping. Once placed in tumor microenvironment (TME), the existence of redox couples (Cu+/Cu2+ and Mo4+/Mo6+) in m-MCS enabled it to react with hydrogen peroxide (H2O2) to generate ·OH for achieving CDT effect via Fenton-like reaction. Meanwhile, m-MCS could consume overexpressed GSH in tumor microenvironment (TME) to alleviate antioxidant capability for enhancing CDT effect. Moreover, m-MCS with mesoporous structure could be employed as the carrier to load natural nitric oxide (NO) donor LA. US as the excitation source with high tissue penetration can trigger m-MCS@LA to produce NO. As the gas transmitter with physiological functions, NO could play dual roles to kill cancer cells through gas therapy directly, and enhance CDT effect by inhibiting protective autophagy simultaneously. As a result, this US-triggered and NO-mediated synergetic cancer chemodynamic/gas therapy based on m-MCS@LA NPs can effectively eliminate primary tumor and achieved tumor-specific treatment, which provide a possible strategy for developing more effective CDT in future practical applications. STATEMENT OF SIGNIFICANCE: The depletion of reactive oxygen species (ROS) by glutathione (GSH) and oxidative stress induced protective autophagy severely impaired the therapeutic effect of chemodynamic therapy (CDT). Herein, a multifunctional cascade bioreactor based on mesoporous Mo-doped Cu9S5 (m-MCS) nanozymes loaded with L-Arginine (m-MCS@LA) is constructed for realizing enhanced CDT promoted by ultrasound (US) triggered gas therapy. The m-MCS with double redox couples presents the enhanced enzyme-like activities to perform cascade reactions for reducing GSH and generating ROS. LA loaded by m-MCS can produce NO triggered by US to inhibit the mitochondria protective autophagy for reactivating mitochondria involved apoptosis pathway. The US-triggered and NO-mediated CDT based on m-MCS@LA can effectively eliminate primary tumor through the high yield and full utilization of ROS.

In vitro and in vivo assessment of the protective effect of sufentanil in acute lung injury.

OBJECTIVES: To investigate the mechanisms underlying the protective effect of sufentanil against acute lung injury (ALI). MATERIAL AND METHODS: Rats were administered lipopolysaccharide (LPS) by endotracheal instillation to establish a model of ALI. LPS was used to stimulate BEAS-2B cells. The targets and promoter activities of IκB were assessed using a luciferase reporter assay. Apoptosis of BEAS-2B cells was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling. RESULTS: Sufentanil treatment markedly reduced pathological changes in lung tissue, pulmonary edema and secretion of inflammatory factors associated with ALI in vivo and in vitro. In addition, sufentanil suppressed apoptosis induced by LPS and activated NF-κB both in vivo and in vitro. Furthermore, upregulation of high mobility group box protein 1 (HMGB1) protein levels and downregulation of miR-129-5p levels were observed in vivo and in vitro following sufentanil treatment. miR-129-5p targeted the 3' untranslated region and its inhibition decreased promoter activities of IκB-α. miR-129-5p inhibition significantly weakened the protective effect of sufentanil on LPS-treated BEAS-2B cells. CONCLUSION: Sufentanil regulated the miR-129-5p/HMGB1 axis to enhance IκB-α expression, suggesting that sufentanil represents a candidate drug for ALI protection and providing avenues for clinical treatment.

Focal Adhesion Kinase Promotes Hepatic Stellate Cell Activation by Regulating Plasma Membrane Localization of TGFß Receptor 2.

Transforming growth factor ß (TGFß) induces hepatic stellate cell (HSC) differentiation into tumor-promoting myofibroblast, although underlying mechanism remains incompletely understood. Focal adhesion kinase (FAK) is activated in response to TGFß stimulation, so it transmits TGFß stimulus to extracellular signal-regulated kinase and P38 mitogen-activated protein kinase signaling. However, it is unknown whether FAK can, in return, modulate TGFß receptors. In this study, we tested whether FAK phosphorylated TGFß receptor 2 (TGFßR2) and regulated TGFßR2 intracellular trafficking in HSCs. The FAKY397F mutant and PF-573,228 were used to inhibit the kinase activity of FAK, the TGFßR2 protein level was quantitated by immunoblotting, and HSC differentiation into myofibroblast was assessed by expression of HSC activation markers, alpha-smooth muscle actin, fibronectin, or connective tissue growth factor. We found that targeting FAK kinase activity suppressed the TGFßR2 protein level, TGFß1-induced mothers against decapentaplegic homolog phosphorylation, and myofibroblastic activation of HSCs. At the molecular and cellular level, active FAK (phosphorylated FAK at tyrosine 397) bound to TGFßR2 and kept TGFßR2 at the peripheral plasma membrane of HSCs, and it induced TGFßR2 phosphorylation at tyrosine 336. In contrast, targeting FAK or mutating Y336 to F on TGFßR2 led to lysosomal sorting and degradation of TGFßR2. Using RNA sequencing, we identified that the transcripts of 764 TGFß target genes were influenced by FAK inhibition, and that through FAK, TGFß1 stimulated HSCs to produce a panel of tumor-promoting factors, including extracellular matrix remodeling proteins, growth factors and cytokines, and immune checkpoint molecule PD-L1. Functionally, targeting FAK inhibited tumor-promoting effects of HSCs in vitro and in a tumor implantation mouse model. Conclusion: FAK targets TGFßR2 to the plasma membrane and protects TGFßR2 from lysosome-mediated degradation, thereby promoting TGFß-mediated HSC activation. FAK is a target for suppressing HSC activation and the hepatic tumor microenvironment.

[Study on the extraction of total flavonoids from Mentha spicata by ultrasonic method].

The optimal conditions of total flavonoids extraction from Mentha spicata were identified by orthogonal test. The results showed that the ethanol concentration and the ultrasonic time had significant effect on the extraction of total flavonoids and the optimum conditions of extraction were A3B3C1D1, which were as follows: the ethanol concentration was 50%, the ultrasonic time was 40 min, the solid-liquid ratio was 1 g: 50 mL and the temperature of extraction was 60 degrees C, On the basis of the extracting method above, the content of total flavonoids in Mentha spicata was 0.0951 mg/mg. The linear equation was A = 11.51143C - 0.00029, r = 0.9999. This method is simple, quick and accurate with good prosperity.

[Determination of trace lead in traditional Chinese herbal medicine Astragalus by microwave digestion-CTAB enhancing-continual flow ingection hydride generation-ICP-AES].

A new method using microwave digestion technique was developed for the determination of lead in Astragalus by CTAB enhancing-continual flow hydride generation-inductively coupled plasma atomic emission spectrometry (HG-ICP-AES). The experimental conditions of microwave digestion and hydride generation were optimized. This method shows a linear range of 0.23-800 microg x L(-1) and the correlation coefficient is 0.999 9. It is satisfactory to apply the microwave digestion procedure to the determination of Pb under the optimized conditions. The detection limit of the method is 0.23 microg x L(-1) and the RSD is 1.02%. The recovery obtained is 98.8%-100.1%. The results show that this method is rapid and simple with low environmental contamination and complete digestion of samples.