Enhanced hemocompatibility and rapid magnetic anastomosis of electrospun small-diameter artificial vascular grafts.

Background: Small-diameter (<6 mm) artificial vascular grafts (AVGs) are urgently required in vessel reconstructive surgery but constrained by suboptimal hemocompatibility and the complexity of anastomotic procedures. This study introduces coaxial electrospinning and magnetic anastomosis techniques to improve graft performance. Methods: Bilayer poly(lactide-co-caprolactone) (PLCL) grafts were fabricated by coaxial electrospinning to encapsulate heparin in the inner layer for anticoagulation. Magnetic rings were embedded at both ends of the nanofiber conduit to construct a magnetic anastomosis small-diameter AVG. Material properties were characterized by micromorphology, fourier transform infrared (FTIR) spectra, mechanical tests, in vitro heparin release and hemocompatibility. In vivo performance was evaluated in a rabbit model of inferior vena cava replacement. Results: Coaxial electrospinning produced PLCL/heparin grafts with sustained heparin release, lower platelet adhesion, prolonged clotting times, higher Young's modulus and tensile strength versus PLCL grafts. Magnetic anastomosis was significantly faster than suturing (3.65 ± 0.83 vs. 20.32 ± 3.45 min, p < 0.001) and with higher success rate (100% vs. 80%). Furthermore, magnetic AVG had higher short-term patency (2 days: 100% vs. 60%; 7 days: 40% vs. 0%) but similar long-term occlusion as sutured grafts. Conclusion: Coaxial electrospinning improved hemocompatibility and magnetic anastomosis enhanced implantability of small-diameter AVG. Short-term patency was excellent, but further optimization of anticoagulation is needed for long-term patency. This combinatorial approach holds promise for vascular graft engineering.

Fabrication of Decellularized Spleen Matrix Derived from Rats.

Liver transplantation is the primary treatment for end-stage liver disease. However, the shortage and inadequate quality of donor organs necessitate the development of alternative therapies. Bioartificial livers (BALs) utilizing decellularized liver matrix (DLM) have emerged as promising solutions. However, sourcing suitable DLMs remains challenging. The use of a decellularized spleen matrix (DSM) has been explored as a foundation for BALs, offering a readily available alternative. In this study, rat spleens were harvested and decellularized using a combination of freeze-thaw cycles and perfusion with decellularization reagents. The protocol preserved the microstructures and components of the extracellular matrix (ECM) within the DSM. The complete decellularization process took approximately 11 h, resulting in an intact ECM within the DSM. Histological analysis confirmed the removal of cellular components while retaining the ECM's structure and composition. The presented protocol provides a comprehensive method for obtaining DSM, offering potential applications in liver tissue engineering and cell therapy. These findings contribute to the development of alternative approaches for the treatment of end-stage liver disease.

Sodium alginate/sodium lignosulfonate hydrogel based on inert Ca2+ activation for water conservation and growth promotion.

Soil degradation has become a major global problem owing to the rapid development of agriculture. The problems of soil drought and decreased soil fertility caused by soil degradation severely affect the development of the agricultural and forestry industries. In this study, we designed sodium alginate (SA)/sodium lignosulfonate (SLS) hydrogel based on the activation and crosslinking of inert Ca2+. CaCO3 and SA were mixed, and then, inert Ca2+ was activated to prepare a gel with a stable structure and a uniform interior and exterior. The crosslinking activated by inert Ca2+ enhanced the stability of the hydrogel, and the optimal swelling rate of the hydrogel reached 28.91 g/g, thereby effectively improving the water-holding capacity of the soil (77.6-108.83 g/kg). SLS was degraded into humic acid (HA) and gradually released, demonstrating a positive growth-promoting effect in plant growth experiments. The SA/SLS hydrogel can be used for soil water retention and mitigation to significantly decrease the water loss rate of soil. This study will assist in addressing soil drought and fertility loss.

RGO-Induced Flower-like Ni-MOF In Situ Self-Assembled Electrodes for High-Performance Hybrid Supercapacitors.

Currently, the primary bottlenecks that hinder the widespread application of supercapacitors are low energy density and narrow potential windows. Herein, the hybrid supercapacitor with high energy density and wide potential window is constructed via an in situ self-assembly method employing RGO-induced flower-like MOF(Ni). Benefiting from the synergistic effect between RGO and MOF(Ni), the interfacial interactions are effectively improved, and the contact area with the electrolyte is enhanced, which increases the ion transfer kinetics and overall electrochemical performance. The MOF(Ni)@RGO electrode exhibits a specific capacitance of 1267.73 F g-1 at a current density of 1 A g-1. Crucially, the assembled MOF(Ni)@RGO//BC with a broad potential window and good stability employing a MOF(Ni)@RGO anode and biomass carbon cathode, combined with a 2 M PVA-KOH gel-electrolyte, achieves a maximum energy density of 70.16 Wh kg-1 at a power density of 2200.09 W kg-1, outperforming most reported supercapacitors. This hybrid supercapacitor exhibits excellent stability and high energy density, providing a novel strategy for further large-scale applications.

Clinical value of ultrasound in adult Wilms' tumor patient with uremia: A case report and literature review.

RATIONALE: Wilms' tumor (WT) is the most common pediatric kidney malignancy and is rarely found in adults. Nonspecific clinical symptoms and imaging features often lead to delayed diagnosis or misdiagnosis of adult WT, resulting in poor clinical outcomes. Ultrasound (US), as an efficient and noninvasive examination method, has been widely used in clinical diagnosis and treatment. Therefore, various US evidence is meaningful to improve understanding of adult WT characteristics in ultrasound. PATIENT CONCERNS: A 45-year-old female patient with uremia (regular hemodialysis for 13 years) with painless gross hematuria was diagnosed with a right kidney tumor penetrating to the lung. Preoperatively, B-mode ultrasonography showed an ill-defined hyperechoic mass in the right kidney, which revealed an unclear border, uneven internal echoes, and calcification. Besides, the internal blood flow signal of the tumor was detected. Contrast-enhanced ultrasound (CEUS) showed an uneven hyper-enhancement in the tumor ("fast in and slow out"). Contrast-enhanced computed tomography of the kidney indicated a similar result as the CEUS. Moreover, the chest CT identified multiple pulmonary metastatic nodules. DIAGNOSES: An ultrasound-guided percutaneous core needle biopsy of the tumor proceeded to make a definite diagnosis of adult WT (epithelial type). INTERVENTIONS: The patient was treated with tislelizumab. OUTCOMES: No progress was found to date. LESSONS: We report the first case in which CEUS was performed in an adult WT patient with uremia and multiple pulmonary metastases. The features obtained by the US can help in the diagnosis of adult WT and direct further diagnostic procedures.

Cinnamaldehyde Inhibits Postharvest Gray Mold on Pepper Fruits via Inhibiting Fungal Growth and Triggering Fruit Defense.

Gray mold infected with Botrytis cinerea frequently appears on fruits and vegetables throughout the supply chain after harvest, leading to economic losses. Biological control of postharvest disease with phytochemicals is a promising approach. CA (cinnamaldehyde) is a natural phytochemical with medicinal and antimicrobial activity. This study evaluated the effect of CA in controlling B. cinerea on fresh pepper fruit. CA inhibited B. cinerea growth in vitro significantly in a dose- (0.1-0.8 mM) and time-dependent (6-48 h) manner, with an EC50 (median effective concentration) of 0.5 mM. CA induced the collapse and breakdown of the mycelia. CA induced lipid peroxidation resulting from ROS (reactive oxygen species) accumulation in mycelia, further leading to cell leakage, evidenced by increased conductivity in mycelia. CA induced mycelial glycerol accumulation, resulting in osmotic stress possibly. CA inhibited sporulation and spore germination resulting from ROS accumulation and cell death observed in spores. Spraying CA at 0.5 mM induced a defense response in fresh pepper fruits, such as the accumulation of defense metabolites (flavonoid and total phenols) and an increase in the activity of defense enzymes (PAL, phenylalanine ammonia lyase; PPO, polyphenol oxidase; POD, peroxidase). As CA is a type of environmentally friendly compound, this study provides significant data on the activity of CA in the biocontrol of postharvest gray mold in peppers.

Fusion of the molecular adjuvant C3d to cleavage-independent native-like HIV-1 Env trimers improves the elicited antibody response.

An effective HIV vaccine likely requires the elicitation of neutralizing antibodies (NAbs) against multiple HIV-1 clades. The recently developed cleavage-independent native flexibly linked (NFL) envelope (Env) trimers exhibit well-ordered conformation and elicit autologous tier 2 NAbs in multiple animal models. Here, we investigated whether the fusion of molecular adjuvant C3d to the Env trimers can improve B- cell germinal center (GC) formation and antibody responses. To generate Env-C3d trimers, we performed a glycine-serine- based (G4S) flexible peptide linker screening and identified a linker range that allowed native folding. A 30-60- amino- acid- long linker facilitates Env-to-C3d association and achieves the secretion of well-ordered trimers and the structural integrity and functional integrity of Env and C3d. The fusion of C3d did not dramatically affect the antigenicity of the Env trimers and enhanced the ability of the Env trimers to engage and activate B cells in vitro. In mice, the fusion of C3d enhanced germinal center formation, the magnitude of Env-specific binding antibodies, and the avidity of the antibodies in the presence of an adjuvant. The Sigma Adjuvant System (SAS) did not affect the trimer integrity in vitro but contributed to altered immunogenicity in vivo, resulting in increased tier 1 neutralization, likely by increased exposure of variable region 3 (V3). Taken together, the results indicate that the fusion of the molecular adjuvant, C3d, to the Env trimers improves antibody responses and could be useful for Env-based vaccines against HIV.

Design and Functional Characterization of HIV-1 Envelope Protein-Coupled T Helper Liposomes.

Functionalization of experimental HIV-1 virus-like particle vaccines with heterologous T helper epitopes (T helper VLPs) can modulate the humoral immune response via intrastructural help (ISH). Current advances in the conjugation of native-like HIV-1 envelope trimers (Env) onto liposomes and encapsulation of peptide epitopes into these nanoparticles renders this GMP-scalable liposomal platform a feasible alternative to VLP-based vaccines. In this study, we designed and analyzed customizable Env-conjugated T helper liposomes. First, we passively encapsulated T helper peptides into a well-characterized liposome formulation displaying a dense array of Env trimers on the surface. We confirmed the closed pre-fusion state of the coupled Env trimers by immunogold staining with conformation-specific antibodies. These peptide-loaded Env-liposome conjugates efficiently activated Env-specific B cells, which further induced proliferation of CD4+ T cells by presentation of liposome-derived peptides on MHC-II molecules. The peptide encapsulation process was then quantitatively improved by an electrostatically driven approach using an overall anionic lipid formulation. We demonstrated that peptides delivered by liposomes were presented by DCs in secondary lymphoid organs after intramuscular immunization of mice. UFO (uncleaved prefusion optimized) Env trimers were covalently coupled to peptide-loaded anionic liposomes by His-tag/NTA(Ni) interactions and EDC/Sulfo-NHS crosslinking. EM imaging revealed a moderately dense array of well-folded Env trimers on the liposomal surface. The conformation was verified by liposomal surface FACS. Furthermore, anionic Env-coupled T helper liposomes effectively induced Env-specific B cell activation and proliferation in a comparable range to T helper VLPs. Taken together, we demonstrated that T helper VLPs can be substituted with customizable and GMP-scalable liposomal nanoparticles as a perspective for future preclinical and clinical HIV vaccine applications. The functional nanoparticle characterization assays shown in this study can be applied to other systems of synthetic nanoparticles delivering antigens derived from various pathogens.

Tumor-infiltrating lymphocyte signature in epithelial and stromal compartments of an esophageal squamous cell carcinoma acidic microenvironment mediated by MCT4.

Tumor-infiltrating lymphocytes (TILs), including but not limited to neutrophils, M2 macrophages, cytotoxic CD8 T cells and dendritic cells, will play a role in the acidic tumor microenvironment mediated by monocarboxylate transporter 4 (MCT4) in esophageal squamous cell carcinoma (ESCC). However, the roles they play and their significance in ESCC remain less clear. To understand the clinicopathological and prognostic significance of neutrophils, M2 macrophages, CD8 T cells and dendritic cells in the tumor acidic microenvironment mediated by MCT4, we investigated the distribution of these TILs in the epithelial and stromal compartments of ESCC by means of multiplexed immunohistochemistry on a tissue microarray containing 87 paired dots of ESCC and its adjacent normal tissue (ANT) and an additional 6 cases of unpaired ESCC dots. The density of cells stained with MCT4 in the epithelium was significantly associated with overall survival. Dendritic cells stained with S100 in epithelial compartmentalization were found to markedly correlate with clinical stage and tumor invasion depth. No other significant association could be identified in terms of prognostic and clinicopathological significance. The potential correlation between the number of cells stained with MCT4 versus the number of TILs was also explored, showing that only in epithelial cells were there significant and positive correlations identified between the number of cells stained with MCT4 versus the number of neutrophils stained with CD15, M2 macrophages stained with CD163 and CD8 T cells stained by CD8a. However, no significant correlation was found along the stromal line. Together, the data we described here, although somewhat discouraging, showed that in epithelial cells from which ESCC originated, acidicity mediated by MCT4 may be responsible for lactate release and may have an effect on the infiltration of TILs we assessed.

Proteomic profiling of plasma exosomes reveals CD82 involvement in the development of esophageal squamous cell carcinoma.

The Xinjiang Uygur autonomous region has a high incidence of esophageal cancer. For the early diagnosis of patients with esophageal squamous cell carcinoma (ESCC), exosomes were isolated and quantified by liquid chromatography tandem mass spectrometry ((LC-MS/MS) with data independent acquisition (DIA) from the peripheral blood of patients with benign esophageal disease (BED), esophageal intraepithelial neoplasia (EIN) and ESCC. A total of 1117 proteins were identified in the above 9 samples. The proteomic results showed that the quantity of CD82 in exosomes of EIN was significantly higher than that in patients with BED and ESCC. Meanwhile, our ELISA test verified our proteomic results. In addition, the immunohistochemical results showed high CD82 expression in adjacent normal tissues and low expression in ESCC tissues. CD82 expression in ESCC tissues was negatively correlated with tumor stage and the expression of PKM2, and the high expression of CD82 combined with low expression of PKM2 in ESCC tissues suggested a good prognosis. To further clarify the tumor suppressive mechanism of CD82, the TIMER and TISDB databases were analyzed, and CD82 expression in tumor tissues was found to be related to the infiltration of immune cells. CD82 in exosomes is involved in the development of ESCC. SIGNIFICANCE: Xinjiang is a high incidence area of ESCC. When diagnosed in the middle and late stages of the disease, the prognosis of patients is poor. Exosomes provide the possibility of relatively noninvasive and early detection of esophageal carcinogenesis. To the best of our knowledge, this was the first study using the DIA technique to analyze the exosomal proteins of patients with different stages of ESCC. The proteins identified in the exosomes in these three groups could provide insights for understanding how exosomes promote the occurrence of ESCC, the antitumour mechanism of humans and the early diagnosis of ESCC.

Regional anticoagulation magnetic artificial blood vessels constructed by heparin-PLCL core-shell nanofibers for rapid deployment of veno-venous bypass.

Veno-venous bypass (VVB) is necessary for maintaining hemodynamic and internal environment stabilities in complex liver surgeries. However, the current VVB strategies require systematic anticoagulation and are time-consuming, leading to unexpected complications. This study aims to overcome these limitations by using a novel magnetic artificial blood vessel constructed with heparin-PLCL core-shell nanofibers. Coaxial electrospinning was used to fabricate core-shell nanofibers with heparin encapsulated into the core layer. The microstructure, physical and chemical properties, hemocompatibility, and heparin release behavior were characterized. The regional anticoagulation magnetic artificial vessel was constructed with these nanofibers and used to perform VVB in a rat liver transplantation model for in vivo evaluation. The core-shell nanofibers appeared smooth and uniform without apparent defects. Fluorescence and TEM images indicated that heparin was successfully encapsulated into the core layer. In addition, the in vitro heparin release test presented a two-phase release profile, burst release at day 1 and sustained release from days 2 to 14, which resulted in better hemocompatibility. The VVB could be rapidly deployed in 3.65 ± 0.83 min by the magnetic artificial vessel without systemic anticoagulation. Moreover, the novel device could reduce portal pressure and abdominal organ congestion, protect intestinal function, and increase the survival rate of liver transplantation with a long anhepatic phase from 0 to 65%. In summary, VVB can be rapidly deployed using regional anticoagulation magnetic artificial blood vessels without systemic anticoagulation, which is promising for improving patient outcomes after complex liver surgery.

The Defensive Role of Endogenous H2S in Brassica rapa against Mercury-Selenium Combined Stress.

Plants are always exposed to the environment, polluted by multiple trace elements. Hydrogen sulfide (H2S), an endogenous gaseous transmitter in plant cells, can help plant combat single elements with excess concentration. Until now, little has been known about the regulatory role of H2S in response to combined stress of multiple elements. Here we found that combined exposure of mercury (Hg) and selenium (Se) triggered endogenous H2S signal in the roots of Brasscia rapa. However, neither Hg nor Se alone worked on it. In roots upon Hg + Se exposure, the defensive role of endogenous H2S was associated to the decrease in reactive oxygen species (ROS) level, followed by alleviating cell death and recovering root growth. Such findings extend our knowledge of plant H2S in response to multiple stress conditions.

Near-infrared fluorescence imaging with indocyanine green for assessment of donor livers in a rat model of ischemia-reperfusion.

BACKGROUND: Although marginal donor livers expand the donor pool, an ideal method for quantitatively evaluating the quality of donor livers has not been developed. This study aimed to explore the feasibility of indocyanine green (ICG) fluorescence imaging for estimating liver function in an ischemia-reperfusion model. METHODS: Forty-eight rats were randomly and evenly divided into 8 groups: the control group and the experimental groups (I-VII). The portal vein blocking period was 0 min, 10 min, 20 min, 30 min, 40 min, 50 min and 60 min. After blood flow was reestablished and the hemodynamics stabilized, ICG was injected through the dorsal penile vein as a bolus, and the fluorescence signal was recorded for 30 min in real time. The fluorescence intensity (FI) curve of the liver was fitted with an asymptotic regression model. Fresh liver tissues and serum were obtained from the middle lobe of the liver on postoperative day (POD) 1 and POD 7 for histopathological evaluation and liver function tests. RESULTS: The growth rate of the FI curve, parameter b3, decreased from groups I to VII. According to the two sudden changes in b3 (20 min, 50 min), the experimental groups could be classified into 3 groups (A, B and C). Hepatocytes in groups I-II showed slight edema, group III began to show obvious hepatocyte edema and vacuolar degeneration, and in groups VI-VII, severe hepatocyte degeneration, necrosis and large inflammatory cell infiltration were observed. Suzuki's scores in the 3 groups were also significantly different (P < 0.01). At the same time, the serum liver function in the experimental groups showed a significant increase on POD 1 and a decrease on POD 7. The alanine aminotransferase (ALT), aspartate aminotransferase (AST), and total bilirubin (TB) levels of groups A, B, and C were significantly different on POD 1 (P < 0.05), and the ALT and direct bilirubin (DB) levels were significantly different on POD 7 (P < 0.05); the lactic dehydrogenase (LDH) level of the group C was significantly higher than that of the groups A and B on POD 1 and POD 7. Meanwhile, the 7-day survival rate of the rats in group C was poor compared to that of the rats in groups A and B (58.3% vs. 100% vs. 100%). CONCLUSION: ICG fluorescence imaging is effective for estimating the degree of liver damage and grading in an ischemia-reperfusion model. It probably has the potential for use in assessing the quality of the donor liver in liver transplantation.

[Corrigendum] Plasma­derived exosomal pyruvate kinase isoenzyme type M2 accelerates the proliferation and motility of oesophageal squamous cell carcinoma cells.

Subsequently to the publication of the above article, the authors have realized that Fig. 5B on p. 8 was compiled erroneously, in the sense that the two immunohistochemical images selected for Fig 5B did not correspond to each other, meaning they were not derived from the same field under the microscope. This error was inadvertently made during the preparation of the manuscript. A corrected version of Fig. 5, showing the correct data for the expression of PKM2 in NAT in Fig. 5B, is shown on the next page. This inadvertent error did not affect the conclusions reported in this paper, and all the authors agree with this Corrigendum. The authors sincerely thank the Editor of Oncology Reports for presenting them with the opportunity to publish this Corrigendum, and apologize to the readership of the journal for any inconvenience caused. [the original article was published in Oncology Reports 46: Article no. 216, 2021; DOI: 10.3892/or.2021.8167].

Plasma­derived exosomal pyruvate kinase isoenzyme type M2 accelerates the proliferation and motility of oesophageal squamous cell carcinoma cells.

Exosomal pyruvate kinase isoenzyme type M2 (PKM2) has been found to play a key role in the progression of human hepatocarcinoma. However, exosomal PKM2 (especially plasma­derived exosomal PKM2), in patients with oesophageal squamous cell carcinoma (ESCC) has not been well defined. In the present study, plasma­derived exosomes were isolated from healthy controls and patients with ESCC, and identified by transmission electronic microscopy, western blotting, nano­flow cytometry, nanoparticle tracking and phagocytosis analysis; exosomal PKM2 was detected by western blotting and ELISA. In addition, changes in cellular proliferation and motility in recipient cells (Eca109) were assessed using Cell Counting Kit­8, colony formation, wound­healing and Transwell assays. The PKM2 content was higher in exosomes from patients with ESCC than in those from healthy donors. Furthermore, exosomes from patients with ESCC enhanced the proliferation and motility of ESCC cells in vitro. Notably, PKM2 was found to be transferred by exosomes, and was able to act by activating STAT3. To verify the association between PKM2 and STAT3, immunohistochemistry was employed to analyse the protein levels of PKM2 and pSTAT3Tyr705. These data revealed that PKM2 and pSTAT3Tyr705 were upregulated and associated with overall survival in patients with ESCC. Therefore, the present study highlights that exosomes from patients with ESCC enhance the migration and invasiveness of ESCC cells by transferring PKM2.

Shikonin Inhibits Tumor Growth of ESCC by suppressing PKM2 mediated Aerobic Glycolysis and STAT3 Phosphorylation.

Background: Shikonin, a small molecule inhibitor of pyruvate kinase 2 (PKM2), has been demonstrated to play the antitumor effect in various cancers. However, the specific effects and related regulatory mechanism of Shikonin in esophageal squamous cell carcinoma (ESCC) have not been clearly declared. Materials and methods: Cell viability was valued through 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Glucose consumption, lactate production, glycolytic intermediates and pyruvate kinase enzymatic activity were measured using corresponding assay kits. Patient-derived xenograft (PDX) models were constructed to observe the anti-ESCC effect of Shikonin in vivo. PKM2, p-PKM2, signal transducer and activator of transcription 3 (STAT3), p-STAT3, glucose transporter 1 (GLUT1) and hexokinase 2 (HK2) in ESCC tissues were assessed by western blot. The expression of PKM2, p-PKM2, p-STAT3, GLUT1 and HK2 was assessed by immunohistochemistry (IHC) in ESCC tissue based on PDXs. Results: Shikonin effectively inhibited cell proliferation in dose-dependent and time-dependent manner compared with the control group. The detection of glycolysis showed that Shikonin suppressed the glucose consumption, lactate production, glycolytic intermediates and pyruvate kinase enzymatic activity. Furthermore, Shikonin not only inhibited the growth of ESCC, but also decreased the expression of p-PKM2 and p-STAT3 in vivo. Finally, Shikonin suppressed the expression of GLUT1 and HK2 proteins which are related to glycolysis. Conclusion: Shikonin has a significant antitumor effect in the ESCC by suppressing PKM2 mediated aerobic glycolysis and regulating PKM2/STAT3 signal pathway.

CALM1 promotes progression and dampens chemosensitivity to EGFR inhibitor in esophageal squamous cell carcinoma.

BACKGROUND: Calmodulin1 (CALM1) has been identified as one of the overexpression genes in a variety of cancers and EGFR inhibitor have been widely used in clinical treatment but it is unknown whether CALM1 and epidermal growth factor receptor (EGFR) have a synergistic effect in esophageal squamous cell carcinoma (ESCC). The aim of the present study was to explore the synergistic effects of knock-out CALM1 combined with EGFR inhibitor (Afatinib) and to elucidate the role of CALM1 in sensitizing the resistance to Afatinib in ESCC. METHOD: Immunohistochemistry (IHC) and qRT-PCR were used to examine the expression of CALM1 and EGFR in ESCC tissues. Kaplan-Meier survival analysis was used to analyze the clinical and prognostic significance of CALM1 and EGFR expression in ESCC. Furthermore, to evaluate the biological function of CALM1 in ESCC, the latest gene editing technique CRISPR/Cas9(Clustered regularly interspaced short palindromic repeats)was applied to knockout CALM1 in ESCC cell lines KYSE150, Eca109 and TE-1. MTT, flow cytometry, Transwell migration, scratch wound-healing and colony formation assays were performed to assay the combined effect of knock-out CALM1 and EGFR inhibitor on ESCC cell proliferation and migration. In addition, nude mice xenograft model was used to observe the synergistic inhibition of knock-out CALM1 and Afatinib. RESULTS: Both CALM1 and EGFR were found to be significantly over-expressed in ESCC compared with paired normal control. Over-expressed CALM1 and EGFR were significantly associated with clinical stage, T classification and poor overall prognosis, respectively. In vitro, the combined effect of knock-out CALM1 mediated by the lentivirus and EGFR inhibitor was shown to be capable of inhibiting the proliferation, inducing cell cycle arrest at G1/S stage and increasing apoptosis of KYSE-150 and Eca109 cells; invasion and migration were also suppressed. In vivo, the results of tumor weight and total fluorescence were markedly reduced compared with the sgCtrl-infected group and sgCAML1 group. CONCLUSION: Our data demonstrated that knock-out of CALM1 could sensitize ESCC cells to EGFR inhibitor, and it may exert oncogenic role via promotion of EMT. Taken together, CALM1 may be a tempting target to overcome Afatinib resistance.

Prognostic significance of pyruvate kinase M2 expression in esophageal squamous cell carcinoma and its meta-analysis.

BACKGROUND: Pyruvate kinase 2 (PKM2) is a key enzyme in the glycolysis pathway and has been reported to be associated with the development of esophageal squamous cell carcinoma (ESCC). However, the prognostic value of PKM2 in ESCC remains undetermined. METHODS: This study aimed to investigate the clinicopathological significance of PKM2 expression in ESCC. A comprehensive and systematic literature search was conducted using the PubMed, Embase, Medline, and Cochrane library databases. The quality of studies and potential for bias were appraised, and meta-analysis was performed to assess the prognostic impact of PKM2 on overall survival (OS). RESULTS: A total of 5 studies with 781 participants were eligible and enrolled. Patients with high PKM2 expression were associated with poor prognosis in ESCC [hazard ratio (HR) =1.72, 95% confidence interval (CI): 1.41-2.09; P<0.01]. Furthermore, upregulated PKM2 was significantly associated with lymph node metastasis [odds ratio (OR) =2.38, 95% CI: 1.68-3.35; P<0.01], clinical stage (OR =3.29, 95% CI: 2.27-4.77; P<0.01), and tumor (T) classification (OR =2.92, 95% CI: 2.05-4.16, P<0.01). DISCUSSION: High PKM2 expression denotes worse OS in ESCC patients, and correlates with the lymph node metastasis, clinical stage, and T classification. However, further studies are warranted to assess how PKM2 can be implemented as a reliable staging element in clinical practice and whether it could provide a new target for therapeutic intervention.

CXCL6 fuels the growth and metastases of esophageal squamous cell carcinoma cells both in vitro and in vivo through upregulation of PD-L1 via activation of STAT3 pathway.

CXCL6, contraction of C-X-C motif chemokine ligand 6, whose biological roles have been rarely described in esophageal squamous cell carcinoma (ESCC). To understand the clinicopathological and biological roles played by CXCL6 in the growth and metastasis of ESCC, immunohistochemistry was used to detect the expression of CXCL6 in ESCC tissues, totaling 105 cases; and the correlation was statistically analyzed between CXCL6 expression and clinicopathological parameters. The role mediated in migration and invasion was evaluated using wound-healing and Transwell assays. MTT and flow cytometry were used to assay the proliferative variation. In vivo, tail vein injection model was established in nude mice xenografted with human ESCC cell lines whose CXCL6 were artificially manipulated. It was found that relative to normal control, CXCL6 was profoundly higher in ESCC; upregulated CXCL6 only significantly correlated with differentiation degree. In vitro, CXCL6 was found to promote the proliferation, migration, and invasion of ESCC cells; which was fully corroborated by nude mice experiment that CXCL6 can promote the growth and metastases of ESCC cells in vivo. Mechanistically, CXCL6 was discovered to be capable of promoting epithelial-mesenchymal transition and upregulating PD-L1 expression through activation of the STAT3 pathway. Collectively, all the data we showed here demonstrate that CXCL6 can enhance the growth and metastases of ESCC cells both in vivo and in vitro.

Downregulation of GSK-3ß Expression via Ultrasound-Targeted Microbubble Destruction Enhances Atherosclerotic Plaque Stability in New Zealand Rabbits.

Accumulating evidence suggests that atherosclerosis (AS) is the underlying cause of vascular diseases, including heart disease and stroke. Ultrasound-targeted microbubble destruction (UTMD) technology provides a tolerable, efficient and effective system for drug delivery and gene transfection, which has broad application prospects in the treatment of AS. In addition, glycogen synthase kinase (GSK)-3ß has been implicated as a potentially valuable therapeutic agent for AS treatment; however, the specific molecular mechanisms remain unknown. Therefore, this study was conducted to explore the effect of downregulation of GSK-3ß expression via UTMD on atherosclerotic plaque stability. We established a THP-1 macrophage-derived foam cell model in vitro and an atherosclerotic plaque model in the right common carotid artery of New Zealand rabbits. We determined levels of the relevant vulnerable plaque stability elements. The results indicate that GSK-3ß was upregulated in the foam cells and in atherosclerotic rabbits. Downregulation of GSK-3ß expression by UTMD suppressed vulnerable plaque factors and inflammation in vitro and in vivo, changed the cytoskeleton of the foam cells in vitro, increased Young's modulus and decreased the peak intensity of atherosclerotic plaque in vivo. Moreover, GSK-3ß inhibition by UTMD did not influence the viability of the foam cells. Collectively, our results indicate that GSK-3ß could be a potential target for anti-atherogenic interventions and, in particular, can improve the stability of AS plaques in combination with UTMD.