Integrated single-cell and bulk RNA-Seq analysis enhances prognostic accuracy of PD-1/PD-L1 immunotherapy response in lung adenocarcinoma through necroptotic anoikis gene signatures.

In addition to presenting significant diagnostic and treatment challenges, lung adenocarcinoma (LUAD) is the most common form of lung cancer. Using scRNA-Seq and bulk RNA-Seq data, we identify three genes referred to as HMR, FAM83A, and KRT6A these genes are related to necroptotic anoikis-related gene expression. Initial validation, conducted on the GSE50081 dataset, demonstrated the model's ability to categorize LUAD patients into high-risk and low-risk groups with significant survival differences. This model was further applied to predict responses to PD-1/PD-L1 blockade therapies, utilizing the IMvigor210 and GSE78220 cohorts, and showed strong correlation with patient outcomes, highlighting its potential in personalized immunotherapy. Further, LUAD cell lines were analyzed using quantitative PCR (qPCR) and Western blot analysis to confirm their expression levels, further corroborating the model's relevance in LUAD pathophysiology. The mutation landscape of these genes was also explored, revealing their broad implication in various cancer types through a pan-cancer analysis. The study also delved into molecular subclustering, revealing distinct expression profiles and associations with different survival outcomes, emphasizing the model's utility in precision oncology. Moreover, the diversity of immune cell infiltration, analyzed in relation to the necroptotic anoikis signature, suggested significant implications for immune evasion mechanisms in LUAD. While the findings present a promising stride towards personalized LUAD treatment, especially in immunotherapy, limitations such as the retrospective nature of the datasets and the need for larger sample sizes are acknowledged. Prospective clinical trials and further experimental research are essential to validate these findings and enhance the clinical applicability of our prognostic model.

Differential clinical outcomes after 3 versus 5 years in a comparison of preoperative chemotherapy with and without radiotherapy in locally advanced rectal cancer: A national cohort propensity score-matched study.

Background: Preoperative chemotherapy alone might be a good alternative to preoperative chemoradiotherapy for patients with locally advanced rectal cancer, yet long-term real-world data from the same cohort are lacking. Methods: Patients diagnosed with stage II-III rectal adenocarcinoma from 2011 to 2015 were randomly sampled from the SEER-Plus database to evaluate the superiority of preoperative chemoradiotherapy versus preoperative chemotherapy alone. Findings: A total of 1314 eligible patients were enrolled, with a median follow-up of 74.0 months. At 3-year follow-up, neither overall survival (OS) nor cancer-specific survival (CSS) was significantly different between the two treatment groups. At 5-year follow-up, CSS was similar across groups (HR 0.768, 95% CI 0.532-1.108; P = 0.156), but the 5-year OS was significantly better in the preoperative chemoradiotherapy group than in the preoperative chemotherapy group (HR 0.682, 95% CI 0.538-0.866; P = 0.002). Besides, the landmark analysis indicated a direct contrast in the CSS within 3 years (HR 1.101, 95% CI 0.598-2.029; P = 0.756) versus that at 3-5 years (HR 0.597, 95% CI 0.377-0.948; P = 0.027). The landmark analysis also showed directly contrasting OS outcomes within 3 years (HR 0.761, 95% CI 0.533-1.086; P = 0.130) versus those at 3-5 years (HR 0.621, 95% CI 0.451-0.857; P = 0.003). Interpretation: In patients with locally advanced rectal cancer under real-world treatment practices, the addition of preoperative radiotherapy to chemotherapy improves survival outcomes at 3-5 years' follow-up but not at 3-year follow-up.

Rab26 alleviates sepsis-induced immunosuppression as a master regulator of macrophage ferroptosis and polarization shift.

Macrophage dysfunction is a significant contributor to more than 70 % of sepsis-related deaths, specifically secondary bacterial infections, during the immunosuppression stage of sepsis. Nevertheless, the role of Rab26 in this context remains unclear. In this study, we observed a substantial decrease in Rab26 expression in macrophages during the immunosuppressive phase of sepsis, which was also found to be suppressed by high extracellular levels of HMGB1. During the progression of sepsis, Rab26 deficiency promotes a polarization shift from the M1 to the M2-like phenotype in macrophages, rendering them susceptible to ferroptosis. Subsequent experimentation has revealed that Rab26 deficiency facilitates the degradation of GPX4, thereby aggravating macrophage ferroptosis through the upregulation of levels of lipid ROS, MDA, and ferrous iron induced by RSL3, a ferroptosis inducer. Additionally, Rab26-deficient mice in the immunosuppressed phase of sepsis exhibit heightened susceptibility to secondary infections, leading to exacerbated lung tissue damage and increased mortality rate. Overall, these findings indicate that Rab26 plays a crucial role in sepsis-induced macrophage immunosuppression by regulating macrophage ferroptosis and polarization. Hence, it represents a potential novel target for sepsis therapy.

Silk Fibroin-Modified Liposome/Gene Editing System Knocks out the PLK1 Gene to Suppress the Growth of Lung Cancer Cells.

Polo-like protein kinase 1 (PLK1) plays a key role in lung cancer cell mitosis. The knockout of PLK1 gene by the CRISPR-Cas9 system can effectively inhibit the proliferation of tumor cells, but there is no suitable vector for in vivo delivery. In this study, CRISPR-Cas9 gene knockout plasmids encoding sgRNA, Cas9 and green fluorescent protein were constructed. Then, the plasmids were packaged with liposome (Lip) and cholesterol-modified Antheraea pernyi silk fibroin (CASF) to obtain the CASF/Lip/pDNA ternary complex. The CASF/Lip/pDNA complex was transfected into lung cancer cells A549 to investigate the transfection efficiency, the PLK1 gene knockout effect and the inhibitory effect on lung cancer cells. The results showed that the transfection efficiency of the CASF/Lip/pDNA complex was significantly higher than that of the Lip/pDNA binary complex, and the expression of PLK1 in cells transfected with CASF/Lip/pDNA complexes was significantly lower than that in cells transfected with Lip/pDNA complexes. The CASF/Lip/pDNA complex significantly increased the apoptosis rate and decreased the proliferation activity of lung cancer cells compared with Lip/pDNA complexes. The cytotoxicity of the complexes was evaluated by coculture with the human bronchial epithelial cells BEAS2B. The results showed that CASF/Lip/pDNA complexes exhibited lower cytotoxicity than Lip/pDNA complexes. The fibroin-modified liposome/PLK1 gene knockout system not only effectively inhibited the growth of lung cancer cells but also showed no obvious toxicity to normal cells, showing potential for clinical application in lung cancer therapy.

Zigui-Yichong-Fang protects against cyclophosphamide-induced premature ovarian insufficiency via the SIRT1/Foxo3a pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Zigui-Yichong-Fang (ZGYCF) is a traditional Chinese medicine prescription for the treatment of infertility and premature ovarian insufficiency (POI). It is clinically used to regulate hormone levels, improve ovarian reserve and increase pregnancy rate. However, the exact mechanism of action is not yet clear. AIMS OF THE STUDY: This study aimed to explore the potential impact and mechanism of ZGYCF on POI, and provide a scientific basis for its clinical application. MATERIALS AND METHODS: UHPLC‒MS/MS was used to identify the main compounds of ZGYCF. Female 8-week-old C57BL/6N mice were randomized into four group containing the vehicle control (Veh) group, the cyclophosphamide (CTX) model group, the low-dose ZGYCF (CTX-ZG-L) group and the high-dose ZGYCF (CTX-ZG-H) group. A mouse POI model was induced with a single intraperitoneal injection of CTX, and the therapeutic effects of different doses of ZGYCF on POI were evaluated according to the ovarian weight coefficient, serum AMH, serum E2, ovarian histomorphology and follicle counts. After the dose screening experiment, the CTX-ZG-L group was renamed the CTX-ZG group and subjected to follow-up experiments. RNA-seq was used to explore the mechanism of POI and the therapeutic mechanism of ZGYCF on POI in Veh group, CTX group and CTX-ZG group. The mechanism of action of ZGYCF on POI were determined by measuring serum hormone level, histomorphology, follicle counts, protein expression and acetylation modification in groups of Veh, CTX, CTX-ZG and CTX-ZG-Nam (SIRT1 inhibitor). RESULTS: A total of 37 compounds in ZGYCF were identified. ZGYCF attenuated the morphological changes in ovarian tissue in POI model mice, increased serum AMH and E2 levels, reduced the damage to primordial follicles and other follicles at all stages, and protected ovarian reserve. RNA-seq results suggested that the genes expression of the PI3K signaling and apoptosis signaling pathways was increased in POI mice, while ZGYCF upregulated SIRT1 gene and the expression of estradiol, apoptosis inhibition and other signaling pathway genes. Immunohistochemical staining, TUNEL staining, Western blot analysis and immunoprecipitation results showed that in CTX group, SIRT1 expression and Foxo3a nuclei localization were decreased, while Ac-Foxo3a, p-AKT, p-Foxo3a and apoptotic markers were upregulated. After administration of ZGYCF, these conditions were reversed, however, after treatment with the SIRT1 inhibitor, the results were opposite to those of ZGYCF. CONCLUSIONS: Acetylated Foxo3a plays an important role in the occurrence of POI. ZGYCF improves the ovarian reserve of CTX-induced POI mice by activating SIRT1-mediated deacetylation of Foxo3a, and played a role in the treatment of POI. SIRT1 may be a novel target for ZGYCF to ameliorate POI.

A Native Drug-Free Macromolecular Therapeutic to Trigger Mutual Reinforcing of Endoplasmic Reticulum Stress and Mitochondrial Dysfunction for Cancer Treatment.

Drug-free macromolecular therapeutics are promising alternatives to traditional drugs. Nanomedicines with multiple organelles targeting can potentially increase the efficacy. Herein, a drug-free macromolecular therapeutic was designed to formulate endoplasmic reticulum (ER) and mitochondria dual-targeting nanoparticles (EMT-NPs), which can synergistically elicit ER stress and mitochondrial dysfunction. In vitro experiments indicated that EMT-NPs could effectively enter ER and mitochondria at an approximate ratio of 2 to 3. Subsequently, EMT-NPs could upregulate ER stress-related protein expression (IRE1α, CHOP), boosting calcium ion (Ca2+) efflux and activating the caspase-12 signaling cascade in cancer cells. In addition, EMT-NPs induced direct oxidative stress in mitochondria; some mitochondrial-related apoptotic events such as decreased mitochondrial membrane potential (MMP), upregulation of Bax, cytochrome c release, and caspase-3 activation were also observed for tumor cells upon incubation with EMT-NPs. Furthermore, the leaked Ca2+ from ER could induce mitochondrial Ca2+ overloading to further augment cancer cell apoptosis. In brief, mitochondrial and ER signaling networks collaborated well to promote cancer cell death. Extended photoacoustic and fluorescence imaging served well for the treatment of in vivo patient-derived xenografts cancer model. This drug-free macromolecular strategy with multiple subcellular targeting provides a potential paradigm for cancer theranostics in precision nanomedicine.

Nocardia pulmonis sp. nov., an actinomycete isolated from a patient with pulmonary infection.

A novel bacterial strain, CDC141T, was isolated from sputum samples of a patient with pulmonary infection in Hainan Province, PR China. We performed a polyphasic study to assess the taxonomic position of the new species. Based on the results of 16S rRNA gene sequence analyses, strain CDC141T belonged to the genus Nocardia with the highest sequence similarity to Nocardia nova NBRC 15556T (98.84 %) and Nocardia macrotermitis RB20T (98.54 %). The dapb1 gene sequence-based phylogenetic and phylogenomic trees further showed that the novel strain was clustered in a distinct clade adjacent to Nocardia pseudobrasiliensis DSM 44290T. The DNA G+C content of strain CDC141T was 68.57 mol%. The genomic diversity analysis revealed low average nucleotide identity and in silico DNA‒DNA hybridization values (<84.7 and <28.9 %, respectively) with its closest relative. Growth occurred at 20-40 °C, pH 6.0-9.0 and with NaCl concentrations of 0.5-2.5 % (w/v). The main fatty acids of strain CDC141T were C16 : 0, C18 : 0 10-methyl, TBSA, C16 : 1 ω6c/C16 : 1 ω7c, C18 : 1 ω9c, C18 : 0, C17 : 1 iso I/anteiso B and C17 : 0. The polar lipid profile was dominated by diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannoside, unidentified glycolipids, unidentified phospholipids and unidentified lipids. MK8 (H4ω-cycl) and MK8 (H4) were the major respiratory quinones. These characteristics were consistent with the typical chemotaxonomic properties of members of the genus Nocardia. Based on the results of phenotypic and genetic analyses, strain CDC141T was identified as representing a new species of the genus Nocardia, with the proposed name Nocardia pulmonis sp. nov. (CDC141T=JCM 34955T=GDMCC 4.207T).

Plasmid containing VEGF-165 and ANG-1 dual genes packaged with fibroin-modified PEI to promote the regeneration of vascular network and dermal tissue.

Reducing the cytotoxicity of cationic polymers is the major issue to their use as a gene delivery carrier. In this study, plasmids containing encoding vascular endothelial cell growth factor 165 and angiopoietin-1 were packaged with the conjugates of cationic fibroin (CSF) and polyethylenimine (PEI), instead of packaging pDNA with PEI alone, to prepare nanocomplexes (CSF+PEI)/pDNA. The complexes were loaded into a silk fibroin scaffold to enhance its function to induce microvascular network generation and dermal tissue regeneration. The results of transfecting EA.hy926 cells with the complexes in vitro showed that (CSF+PEI)/pDNA had a stronger transfection ability than PEI/pDNA. Importantly, compared with PEI as the gene carrier alone, the cell viability was significantly increased and the cytotoxicity was effectively reduced after the conjugate of CSF and PEI was used as the gene carrier. The results of angiogenesis in chick embryo chorioallantoic membranes showed that compared with scaffolds loaded with PEI/pDNA, the neovascularization ratio in scaffolds loaded with (CSF+PEI)/pDNA was significantly increased. In vivo experimental results of scaffolds implantation for full-thickness skin defects in SD rats showed that, compared with loading PEI/pDNA complex, loading (CSF+PEI)/pDNA complex in the scaffold more effectively promoted the formation of vascular network in the scaffold and accelerated the regeneration of dermal tissue. The gene delivery system established in this study has application potential not only in the regeneration of vascular-containing tissues, but also in tumor gene therapy.

Accurate, Sensitive, and Rapid Detection of Pseudomonas aeruginosa Based on CRISPR/Cas12b with One Fluid-Handling Step.

Pseudomonas aeruginosa is a major bacterial pathogen causing nosocomial infections and accounts for morbidity and mortality among patients with cystic fibrosis. An accurate, sensitive, and rapid method to detect P. aeruginosa is critical for the early control of infection and patient management. In this study, we established a P. aeruginosa clustered regularly interspaced short palindromic repeats testing in one pot (CRISPR-top) assay which detected P. aeruginosa with one fluid-handling step in one tube. The reaction was performed isothermally within 1 h; thus, specific instruments were not required. The optimal reaction conditions of this assay were determined to be a temperature of 55°C; working concentrations of 1 µM for the forward inner primer and backward inner primer, 0.5 µM for the loop forward primer and loop backward primer, and 0.25 µM for the forward outer primer and backward outer primer; as well as a 2 µM concentration single-stranded DNA reporter molecules. In terms of specificity, our assay showed 100% inclusivity and exclusivity among 48 strains, including 15 P. aeruginosa clinical isolates and 33 non-P. aeruginosa strains. The limit of detection of our method was 10 copies per reaction mixture. Forty-six human sputum specimens from patients with respiratory symptoms were tested. Using the results of quantitative real-time PCR as the gold standard, our method showed 85.3% (29/34) sensitivity, 100% (12/12) specificity, a positive predictive value of 100% (29/29), and a negative predictive value of 70.6% (12/17). In summary, the P. aeruginosa CRISPR-top assay developed in the present study is a high-efficiency alternative tool for the accurate and rapid detection of P. aeruginosa, especially in resource-limited settings. IMPORTANCE This study reports a P. aeruginosa CRISPR-top assay which can precisely identify P. aeruginosa using nucleic acids from pure cultures or clinical samples in one pot with one fluid-handling step. The P. aeruginosa CRISPR-top reaction is suitable for on-site testing, and its diagnostic performance can be compared with that of qPCR.

Chemical immune conization of precancerous cervical lesions awakens immune cells and restores normal HPV negative and abnormal proliferation.

Background: Cervical cancer is one of the most common and deadly cancers in women, which is closely linked to the persistent infection of high-risk human papillomavirus (HPV). Current treatment of cervical cancer involves radical hysterectomy, radiotherapy, and chemotherapy or a combination. Objective: We investigated if hapten-enhanced intratumoral chemotherapy (HEIC) was effective in boosting immunity for effective treatment of precancerous cervical lesions and HPV infection. Study design: We used single-cell RNA sequencing (scRNA-Seq) to obtain transcriptome profiles of 40,239 cells from biopsies of precancerous cervical lesions from the cervix directly from one patient before the start of HEIC and approximately 1 week after HEIC. The blood samples were taken at the same time as biopsies. We compared the expression characteristics of malignant epithelial cells and immune cells, including epithelial cells, endothelial cells (ECs), fibroblasts, mural cells, T cells, B cells, T and NK neutrophils, mast cells, microparticles (MPs), and platelets, as well as the dynamic changes in cell percentage and cell subtype heterogeneity. Results: Intratumoral injection of chemotherapy drug plus hapten induces an acute immune response in precancerous cervical lesions with HPV and further awakens immune cells to prevent the abnormal proliferation of the precancerous cells. Conclusion: HEIC provides a potential treatment method for cervical cancer and HPV infection tailored to each patient's condition.

Research on application of tumor treating fields in glioblastoma: A bibliometric and visual analysis.

Background: Glioblastoma, one of the common tumors of the central nervous system (CNS), is prone to recurrence even after standard treatment protocols. As an innovative physiotherapy method emerging in recent years, the tumor treating fields (TTFields) technique has been approved for the treatment of glioblastoma due to its non-invasive and portable features. The purpose of this study is to visualize and analyze the scientific results and research trends in TTFields therapy for glioblastoma. Methods: Publications related to TTFields therapy for glioblastoma were searched in the Web of Science Core Collection (WoSCC) database in September 2022. A bibliometric and visual analysis of publications in this field was performed mainly using CiteSpace and R software for country/region, author, journal, reference and keyword. Results: A total of 618 publications in this field were retrieved, and 248 were finally obtained according to the search criteria, including 159 articles (64.11%) and 89 reviews (37.89%). The cumulative number of publications increased year by year, with an average growth rate (AGR) of 28.50%. The test results of Pearson correlation coefficient showed a high positive correlation between publications and citations (r=0.937, p<0.001). The USA had the largest number of publications (123, 49.60%), followed by Germany (32, 12.90%) and China (30, 12.10%). As for the country/region collaborations, the USA cooperated most closely with other countries/regions, followed by Germany and China. The degree of collaboration (DC) between countries/regions was 25.81%. The institutions with the largest number of publications were Tel Aviv Univ (10), Harvard Med Sch (10) and Novocure Ltd (10). Moreover, Wong E (18) possessed the greatest number of publications, followed by Weinberg U (11) and Kirson E (10). The DC between authors was 97.58%. STUPP R (236) was the most cited author followed by KIRSON ED (164) and GILADI M (104). JOURNAL OF NEURO-ONCOLOGY (22) was the journal with the largest number of published publications (75), followed by FRONTIERS IN ONCOLOGY (15) and CANCERS (13). The top 10 keywords that occurred frequently included glioblastoma (156), tumor treating field (152), temozolomide (134), randomized phase III (48), brain (46), survivor (46), cancer (44), trial (42), alternating electric field (42) and radiotherapy (36). Furthermore, cluster analysis was performed on the basis of keyword co-occurrence, and finally 15 clusters were formed to determine the current research status and future development trend of TTFields therapy for glioblastoma. Conclusion: TTFields has been increasingly known as the fourth novel physical anti-tumor therapy in addition to surgery, radiotherapy and anti-tumor drugs. Cooperation and communication between countries/regions need to be enhanced in future research. Several studies have demonstrated the therapeutic potential of TTFields in glioma, and its application alone or in combination with other treatments has become a current research hotspot.

Multifunctional ß-Cyclodextrin-Poly(ethylene glycol)-Cholesterol Nanomicelle for Anticancer Drug Delivery.

Nanoparticle drug delivery systems have drawn considerable attention worldwide due to their unique characteristics and advantages in anticancer drug delivery. Herein, the curcumin (Cur) loaded nanomicelles with two-stage drug release behavior were developed. ß-Cyclodextrin (ß-CD) and cholesterol were conjugated onto both ends of the poly(ethylene glycol) (PEG) chain to obtain an amphiphilic ß-CD-PEG-Chol. The Cur was loaded into the cavities of ß-CD and nanomicelle when the ß-CD-PEG-Chol self-assembled to the Cur@ß-CD-PEG-Chol nanomicelles (Cur@CPC NMs). These Cur@CPC NMs are spherical particles with a particle size of 120.9 nm. The Cur drug loading capacity of Cur@CPC NMs are 61.6 ± 6.9 mg/g. The release behavior of Cur from Cur@CPC NMs conformed to a two-stage mode of "burst-release followed by sustained-release". The prepared Cur@CPC NMs possess high storage stability and excellent hemocompatibility. Moreover, these Cur@CPC NMs exhibit satisfactory antioxidant activity and anticancer activity, resulting in significant reduction in intracellular H2O2-induced ROS and a nearly 50% lethality rate of HepG-2 cells. Meanwhile, the Cur@CPC NMs show good anti-inflammatory activity, by which the secretion of inflammatory factors of IL-6 and TNF-α are inhibited. Overall, the developed Cur@CPC NMs show application prospects in anticancer drug delivery systems.

Sensitization effect of kaempferol from persimmon leaves on HepG2 hepatoma cells with ABT-199 resistance and its molecular mechanisms.

ABT-199 (venetoclax) is the first-in-class selective B-cell lymphoma 2 (BCL2) inhibitor, which is known to be ineffective towards liver cancer cells. Here, we investigated the efficacy and the underlying molecular processes of the sensitization effect of kaempferol isolated from persimmon leaves (KPL) on the ABT-199-resistant HepG2 cells. The effects of various doses of KPL coupled with ABT-199 on the proliferation of HepG2 cells and on the H22 liver tumor-bearing mouse model were examined, as well as the underlying mechanisms. Our findings showed that ABT-199 alone, in contrast to KPL, had no significant impact on hepatoma cell growth, both in vitro and in vivo. Interestingly, the combination therapy showed significantly higher anti-hepatoma efficacy. Mechanistic studies revealed that combining KPL and ABT-199 may promote both early and late apoptosis, as well as decrease the mitochondrial membrane potential in HepG2 cells. Western blot analysis showed that combination of KPL and ABT-199 significantly reduced the expression of the anti-apoptotic proteins Bcl-2, Bcl-xL, and Mcl-1, raised the expression of Bax and cleaved caspase 3, and enhanced cytochrome C release and Bax translocation. Therefore, KPL combined with ABT-199 has a potential application prospect in the treatment of hepatocellular carcinoma.

The role of interaction between autophagy and apoptosis in tumorigenesis (Review).

Autophagy is a highly conserved process that maintains cellular homeostasis during evolution. Autophagy can occur in the form of macroautophagy, microautophagy or molecular chaperone autophagy, among which macroautophagy is the most common. Apoptosis exists in all kinds of cell organisms, and is a kind of programmed cell death which is regulated by pro­apoptotic factors and anti­apoptotic signals. The main biological feature of apoptosis is the activation of caspase. Apoptosis is induced by a variety of cell signals, such as endoplasmic reticulum stress, induction of toxic substances, stimulation of pathogenic microorganisms and DNA damage. Inextricable links are found between autophagy and apoptosis. Studies have found that numerous of the autophagy molecules and autophagy signaling pathways involved in the process of autophagy are related to apoptosis. In addition to regulating autophagy, the autophagy signaling pathway also regulates apoptosis. The interaction between the two can achieve a dynamic balance to certain extent, which maintains the basic physiological functions of cells and reduces the damage to the body under stress. Disease occurs when the balance between autophagy and apoptosis is disrupted. Tumors form due to the ability of cells to avoid apoptosis. Autophagy is closely related to apoptosis, there must be a close connection between the three. In the present review, the mechanism between autophagy and apoptosis and the impact of their interaction on tumorigenesis shall be discussed.

Evaluation of Tumor Resection Effect of Color Doppler Ultrasound Positioning Guided Breast-Conserving Surgery Using Nano-Contrast Agent.

This study aimed to explore the clinical value of Fe3O4-based magnetic lipid nano-contrast agent in breast-conserving surgery for breast cancer using color doppler ultrasound positioning and to analyze the tumor resection effect of breast-conserving surgery. On account of Fe3O4 magnetic nanoparticles prepared by the chemical co-precipitation method, magnetic lipid ultrasonic microbubbles (MLU-MBs) were prepared by mechanical oscillation method after surface modification using polyethylene terephthalate (PET). Characterization and analysis of the prepared MLU-MBs were performed using scanning electron microscopy (SEM), energy spectrometer, transmission electron microscopy (TEM), and Fourier infrared spectroscopy (FIRS). A high-frequency alternating magnetic field was used to detect the heating of MLU-MBs and the color ultrasound machine was applied to observe the enhancement effect of the MLU-MBs on rabbit liver images. 92 patients undergoing breast-conserving surgery for breast cancer were taken as the research objects and were divided into a nano group (MLU-MB as contrast agent) and a control group (conventional contrast agent) according to the differences of intraoperative contrast agents, with 46 cases in each group. Before the surgery, both groups of patients were positioned and marked the tumor boundary under ultrasound. The differences in tumor volume (TV), amount of tissue removed, resection rate, and positive rate (PR) of resection margins were compared between the two groups. The results showed that the Fe3O4 magnetic nanoparticles were particles with an average particle size of about 15nm, and their iron and oxygen percentages were consistent with the content of Fe3O4. The MLU-MBs were spherical particles of about 1120nm, containing phosphorus (P), oxygen(O), and Ferrum (Fe). Under 30A and 220kHz of output current and frequency, the temperature rise of the MLU-MBs suspension with different concentrations was 10 ~ 60°C, and the temperature was constant after heating for 45 minutes. Compared with the rabbit liver parenchyma, the image was greatly enhanced. TV, the amount of tissue removed, the resection rate, and the PR of resection margins in the nano group were obviously lower than those in the control group (P<0.05). It showed that MLU-MBs with good image enhancement effect on account of Fe3O4 magnetic nanoparticles were successfully prepared and it could effectively reduce the PR of normal tissues and the positive margin of two-fold resection during breast-conserving surgery for breast cancer and showed good accuracy and stability.

Framework Nucleic Acids Enabled Pulmonary Artery Endothelial Cell Growth Inhibition by Targeting microRNA-152.

Pulmonary artery vascular endothelial dysfunction plays a pivotal role in the occurrence and progression of pulmonary vascular remodeling (PVR). To address this, aberrantly expressed non-coding microRNAs (miRNAs) are excellent therapeutic targets in human pulmonary artery endothelial cells (HPAECs). Here, we discovered and validated the overexpression of miRNA-152 in HPAECs under hypoxia and its role in endothelial cell dysfunction. We constructed a framework nucleic acid nanostructure that harbors six protruding single-stranded DNA segments that can fully hybridize with miRNA-152 (DNT-152). DNT-152 was efficiently taken up by HPAECs with increasing time and concentration; it markedly induced apoptosis, and inhibited HPAEC growth under hypoxic conditions. Mechanistically, DNT-152 silenced miRNA-152 expression and upregulated its target gene Meox2, which subsequently inhibited the AKT/mTOR signaling pathway. These results indicate that miRNA-152 in HPAECs may be an excellent therapeutic target against PVR, and that framework nucleic acids with carefully designed sequences are promising nanomedicines for noncancerous cells and diseases.

Isoflavaspidic Acid PB Extracted from Dryopteris fragrans (L.) Schott Inhibits Trichophyton rubrum Growth via Membrane Permeability Alternation and Ergosterol Biosynthesis Disruption.

Isoflavaspidic acid PB (PB), a phloroglucinol derivative extracted from aerial parts of Dryopteris fragrans (L.) Schott, had antifungal activity against several dermatophytes. This study was aimed at exploring the antifungal mechanism of PB against Trichophyton rubrum (T. rubrum). The effectiveness of PB in inhibiting T. rubrum growth was detected by time-kill kinetics study and fungal biomass determination. Studies on the mechanism of action were investigated using scanning electron microscopy (SEM), transmission electron microscopy (TEM), sorbitol and ergosterol assay, nucleotide leakage measurement, and UPLC-based test and enzyme-linked immunosorbent assay. Fungicidal activity of PB was concentration- and time-dependent at 2 × MIC (MIC: 20 µg/mL) after 36 h. The total biomass of T. rubrum was reduced by 64.17%, 77.65%, and 84.71% in the presence of PB at 0.5 × MIC, 1 × MIC, and 2 × MIC, respectively. SEM analysis showed that PB changed mycelial morphology, such as shrinking, twisting, collapsing, and even flattening. TEM images of treated cells exhibited abnormal distributions of polysaccharide particles, plasmolysis, and cytoplasmic content degradation accompanied by plasmalemma disruption. There were no changes in the MIC of PB in the presence of sorbitol. However, the MIC values of PB were increased by 4-fold with exogenous ergosterol. At 4 h and 8 h, PB increased nucleotide leakage. Besides, ergosterol content in T. rubrum membrane treated with PB at 0.5 × MIC, 1 × MIC, and 2 × MIC was decreased by 9.58%, 15.31%, and 76.24%, respectively. There was a dose-dependent decrease in the squalene epoxidase (SE) activity. And the reduction in the sterol 14α-demethylase P450 (CYP51) activity was achieved after PB treatments at 1 × MIC and 2 × MIC. These results suggest that PB displays nonspecific action on the cell wall. The membrane damaging effects of PB were attributed to binding with ergosterol to increase membrane permeability and interfering ergosterol biosynthesis involved with the reduction of SE and CYP51 activities. Further study is needed to develop PB as a natural antifungal candidate for clinical use.

MUC21 controls melanoma progression via regulating SLITRK5 and hedgehog signaling pathway.

Mucins are heavily glycosylated proteins secreted by various cell types, to protect the epithelial surface of the gastrointestinal tract from damage. Currently, increasing studies provided evidence to suggest that mucins play an essential role in regulating tumor progression. However, the role of mucins and the underpinning mechanism of how mucins drive melanoma progression remains elusive. In this study, we first demonstrated that mucin 21 (MUC21) expression was significantly upregulated in metastatic melanoma tissues, and a higher MUC21 expression resulted in poor overall survival in melanoma patients by The Cancer Genome Atlas database analysis. In vitro, MUC21 overexpression markedly promoted proliferative properties and aggressive behavior of melanoma cell A375 and A875, as assessed by Cell Counting Kit-8 and transwell assay. In mechanism, we proved that MUC21 suppressed expression of SLITRK5, an integral membrane protein, leading to activation of prosurvival hedgehog pathway and sustained melanoma development. More importantly, we found that combination of hedgehog pathway inhibitor cyclopamine and chemotherapy revealed an improved anticancer effect in MUC21 overexpression xenograft model. Altogether, our study described a novel role of MUC21 in regulating tumor progression, which offers a promising target for melanoma diagnosis and therapy.

Estradiol Aggravate Nocardia farcinica Infections in Mice.

Males are generally more susceptible to Nocardia infection than females, with a male-to-female ratio of 2 and higher clinical disease. 17ß-Estradiol has been implicated in affecting the sex-based gap by inhibiting the growth of N. brasiliensis in experiments, but the underlying mechanisms have not yet been fully clarified. In the present study, however, we report increased severity in N. farcinica IFM 10152-infected female mice compared with male mice with increased mortality, elevated lung bacterial loads and an exaggerated pulmonary inflammatory response, which was mimicked in ovariectomized female mice supplemented with E2. Similarly, the overwhelming increase in bacterial loads was also evident in E2-treated host cells, which were associated with downregulating the phosphorylation level of the MAPK pathway by binding the estrogen receptor. We conclude that although there are more clinical cases of Nocardia infection in males, estrogen promotes the survival of the bacteria, which leads to aggravated inflammation in females. Our data emphasize the need to include and separately analyze both sexes in future studies of Nocardia to understand the sex differences in immune responses and disease pathogenesis.

MiR-3682 promotes the progression of hepatocellular carcinoma (HCC) via inactivating AMPK signaling by targeting ADRA1A.

INTRODUCTION AND OBJECTIVES: This study aimed to investigate miR-3682 as a biomarker in hepatocellular carcinoma (HCC). MATERIALS AND METHODS: MiRNA and RNA profiles of 375 HCC tissues and 50 normal liver samples were downloaded from The Cancer Genome Atlas (TCGA) database. Multivariate Cox regression and Kaplan-Meier analyses were applied to examine the prognostic value of factors. Target genes of miR-3682 were analyzed by TargetScan and dual-luciferase reporter assay. Online Database for Annotation, Visualization, and Integrated Discovery (DAVID) to perform KEGG pathway enrichment. Cell counting kit-8, colony formation and migration and invasion assays were performed to analyze biological behaviors of HCC cells. RESULTS: MiR-3682 was identified to be highly expressed in HCC tissues and cell lines. And miR-3682 was negatively and independently associated with the outcome of HCC patients. Inhibition of miR-3682 suppressed HCC cell viability and mobility. ADRA1A, predicted and confirmed as the novel target of miR-3682, was an independent and positive prognostic predictor for HCC. In addition, the knockdown of ADRA1A partially offset the inhibitory effect of miR-3682 inhibitor on the growth and mobility of HCC cells. DAVID enrichment and western blot of key signaling-related proteins analyses revealed that miR-3682 inactivated 5'-AMP-activated protein kinase (AMPK) signaling by negatively regulating ADRA1A. Mechanically, it was partially through suppressing AMPK signaling via targeting ADRA1A that miR-3682 supported the HCC cell malignant phenotype. CONCLUSIONS: This study implicates that miR-3682 plays an oncogenetic role in HCC and can be considered a novel therapeutic target and prognostic indicator of HCC.