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Early metastasis of pancreatic cancer (PC) leads to high mortality, and the underlying mechanism of metastasis remains unclear. Tumor necrosis factor superfamily member 9 (TNFSF9) is associated with poor prognosis in PC. Here, we investigated the effect of TNFSF9 on PC proliferation and apoptosis, and focused on the effect of TNFSF9 on PC metastasis and its potential mechanism. We found that TNFSF9 promotes PC metastasis in vivo and in vitro, and may be partially dependent on the Wnt/Snail signaling pathway. In addition, TNFSF9 also regulates the release of cytokines IL-10 and transforming growth factor-β (TGF-β) in pancreatic cancer cells through Wnt signaling to induce the M2 polarization of macrophages and promote the migration of PC cells. Overall, our study found that TNFSF9 may directly promote PC metastasis or indirectly promote PC metastasis through macrophage M2 polarization. Our study provides a new costimulatory target for the treatment of PC.

As an important transcription factor, zinc-finger and BTB domain-containing 7B (ZBTB7C) plays an important role in a variety of tumors. However, its relationship with human immunity is unclear. This article aims to study its differential expression and survival across cancers and explore the relationships between its differential expression and the tumor microenvironment and immune cell infiltration. In this study, we used R software to process The Cancer Genome Atlas (TCGA) data and explored the expression pattern and prognostic value of ZBTB7C across cancers. Next, we comprehensively explained the important role of ZBTB7C in several tumor types in terms of tumor mutational burden (TMB), microsatellite instability (MSI) and immune cell infiltration. In general, the expression level of ZBTB7C in tumor tissues was lower than that in normal tissues. Highly expressed ZBTB7C was beneficial to the survival of patients with colon adenocarcinoma (COAD), lymphoid neoplasm diffuses large B cell lymphoma (DLBC), esophageal carcinoma (ESCA) and mesothelioma (MESO). Multivariate analysis showed that the expression of ZBTB7C was an independent prognostic factor in COAD and MESO. In COAD, the expression of ZBTB7C was positively correlated with both TMB and MSI. In colorectal cancer (CRC), there was a significant positive correlation between ZBTB7C expression and immune cell infiltration, especially the infiltration of mast cells and B cells. In conclusion, ZBTB7C can be used as a potential therapeutic target across cancers and is related to immune cell infiltration.

Poly-L-lysine (PLL) is a promising candidate for the treatment of pulmonary infection with lower occurrence of drug-resistance due to its unique antibacterial mechanisms. Dry powder inhalations (DPIs) are considered as the first choice for formulating PLL to treat pulmonary infection on account of direct delivery and satisfactory stability. However, hygroscopicity of PLL limited its therapeutic effect on pulmonary infection when PLL developed into DPIs. The hygroscopicity caused two obstacles including the low drug deposition in the lower respiratory tract and undesirable aerosolization performance deterioration. In this study, PLL was co-spray-dried with L-leucine (LL) to achieve moisture-resistance and desirable aerosolization performance. The ratio of PLL and LL was optimized to obtain particles with different morphology, hygroscopicity and aerodynamic properties. The obtained PLL DPIs were suitable for inhalation with a corrugated surface formed by hydrophobic LL. The anti-hygroscopicity, aerosolization performance and rheological properties of P

Objective Parkinson’s disease (PD) is the second most prevalent neurodegenerative disease after Alzheimer's disease (AD) worldwide. The prevalence of small intestinal bacterial overgrowth (SIBO) in PD patients is high. We conducted this comprehensive systematic review and meta-analysis to determine the association between SIBO and PD. Methods A comprehensive literature search of the PubMed, Cochrane Library and EMBASE databases was performed to identify studies correlating SIBO with PD. Studies were screened, and relevant data were extracted and analysed. We calculated the pooled prevalence of SIBO in all individuals with PD and compared the prevalence of SIBO between the two groups to calculate an odds ratio (OR) and 95% confidence interval (CI). Egger’s test was performed to assess publication bias. Results Eleven studies with 973 participants met the inclusion criteria. The pooled prevalence of SIBO in patients with PD was 46% (95% CI 36–56). A random-effects model was applied given the heterogeneity (I2 = 83%) detected among the studies. Egger’s test indicated no publication bias (p = 0.0657). Subgroup analyses showed that the prevalence of SIBO was greater in studies including patients diagnosed using the lactulose hydrogen breath test (LBT) (51%, 95% CI 37–65) than in those including patients diagnosed using the glucose hydrogen breath test (GBT) (35%, 95% CI 20–50), and the prevalence of SIBO in PD was highest (55%, 95% CI 38–72) in patients diagnosed by the LBT and GBT. The prevalence of SIBO was 52% (95% CI 40–64) among patients from Western countries and 33% (95% CI 22–43) among patients from Eastern countries. The pooled OR of SIBO in PD patients compared with healthy controls was 5.22 (95% CI 3.33–8.19, p Conclusion In conclusion, our meta-analysis found a strong association between SIBO and PD with approximately half of PD patients testing positive for SIBO. These relationships significantly differed based on diagnostic test and geographic area.

Background Increasing evidence has shown that cytolytic activity (CYT) is a new immunotherapy biomarker that characterises the antitumour immune activity of cytotoxic T cells and macrophages. In this study, we established a prognostic model associated with CYT. Methods A prognostic model based on CYT-related genes was developed. Furthermore, aberrant expression of genes of the model in colon cancer (CC) was identified by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry (IHC) assays. Next, the correlation between the model and T-cell infiltration in the CC microenvironment was analysed. The Tumour Immune Dysfunction and Exclusion (TIDE) algorithm and subclass mapping were used to predict clinical responses to immune checkpoint inhibitors. Results In total, 280 of the 1418 genes were differentially expressed based on CYT. A prognostic model (including HOXC8 and MS4A2) was developed based on CYT-related genes. The model was validated using the testing set, the whole set and a Gene Expression Omnibus (GEO) cohort (GSE41258). Gene set enrichment analysis (GSEA) and other analyses showed that the levels of immune infiltration and antitumour immune activation in low-risk-score tumours were greater than those in high-risk-score tumours. CC patients with a low-risk-score showed more promise in the response to anti-immune checkpoint therapy. Conclusions Overall, our model may precisely predict the overall survival of CC and reflect the strength of antitumour immune activity in the CC microenvironment. Furthermore, the model may be a predictive factor for the response to immunotherapy.

Additional file 2: Table S2: Primer sequences for RT-PCR.

Additional file 3: Table S3: Clinicopathological parameters of CC patients in the CC cohort for IHC.

Additional file 6: Figure S1: Flow chart of the study. Five public colon cancer-related datasets were chosen for this study. First, we obtained CYT-related DEGs. Next, a two-gene prognostic model associated with CYT was established and validated. Finally, we verified the correlation between the model and T-cell infiltration. Figure S2: a-b LASSO Cox analysis identified the two genes most correlated with overall survival (OS). c-d Effect of HOXC8 and MS4A2 expression on the OS of CC patients in the whole TCGA cohort. Figure S3: Kaplan-Meier survival, risk score and time-dependent receiver operating characteristic (ROC) curves of the model for the testing cohort a-c and whole cohort d-f in CC. a, d Overall survival (OS) was significantly higher in the low-risk-score group than in the high-risk-score group. b, e The relationship between the risk score (upper panel) and expression of two prognostic immune genes (lower panel) is shown. c, f Time-dependent ROC curve analysis. Relationship between the expression of the risk score and g the TNM stage, h T stage, i M stage and j N stage in the IHC cohort.

Additional file 1: Table S1: Clinical features of CC patients in training set and testing set.

Additional file 4: Table S4: The univariate Cox HR regression result of selected genes.

Additional file 5: Table S5: The specific statistical parameters of HOXC8 and MS4A2 in each group.

Downregulation of Carnitine palmitoyltransferase-2 (CPT2) has been shown to be highly associated with the progression of several cancers, but little known about its expression, biological functions and mechanisms in colorectal cancer (CRC).Bioinformatics analysis of The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) data sets was used to explore the expression of CPT2, the relationship between CPT2 expression and clinicopathologic features, as well as the overall survival of CRC. Cox's proportional hazards regression model was used to analyze independent prognostic factors of CRC. In vitro, CRC tissues were analyzed by RT-qPCR, IHC, IF and western blotting to verify CPT2 expression. Colony formation, CCK-8, cell cycle, apoptosis, transwell and wound healing assays were performed to examine the functions of CPT2 in CRC. In vivo, nude mouse xenograft experiment was used to further examine the effect of CPT2 on tumorigenesis. Furthermore, gene set enrichment analysis (GSEA) was conducted to explore the downstream pathway of CPT2. The regulation of p53 pathway by CPT2 was verified by RT-qPCR and Western blotting.CPT2 expression was frequently downregulated in CRC and correlated with poor prognosis. Low CPT2 expression was significantly associated with age, lymph node metastasis, distant metastasis and TMN stage. Univariate and multivariate analysis indicated that low CPT2 expression was an independent prognostic factor for poorer overall survival. Functionally, overexpression of CPT2 in CRC cells induced growth suppression, cell cycle arrest at the G1 phase, enhanced apoptosis and reduced cell migration and invasion. Conversely, knockdown of CPT2 contributed to cell proliferation, migration and invasion, increased the proportion of S phase cells, decreased the proportion of G1 phase cells and inhibited apoptosis. Mechanistically, we found that CPT2 overexpression can increase p53 expression by activating p-p53, leading to p21, Bax, cleaved caspase-9, cleaved caspase-3 and cleaved PARP activation and Bcl2, MDM2 deactivation, thereby inhibiting tumor proliferation and promoting apoptosis. CPT2 knockdown yielded opposite results.These findings suggest that CPT2 may be a novel prognostic marker of CRC and downregulation of CPT2 can promote proliferation and inhibit apoptosis through p53 pathway in CRC. Strategies targeting CPT2 may be developed as therapies for CRC.

The effect of tumor necrosis factor superfamily member 9 (TNFSF9) on the metastasis of pancreatic cancer (PC) and the underlying mechanism remain unclear. We studied the expression of TNFSF9 in pancreatic cancer and its relationship with immune cells. We further explored the effect of TNFSF9 on pancreatic cancer metastasis by inducing macrophage polarization, and evaluated the expression of Src/FAK/p-Akt/IL-1β signals in macrophages after knocking down TNFSF9. The data shows that TNFSF9 expression is elevated in pancreatic cancer and is related to the poor prognosis of patients with pancreatic cancer. In addition, TNFSF9 may induce the M2 polarization of macrophages through Src/FAK/p-Akt/IL-1β signals, thereby promoting the migration of pancreatic cancer cells. In conclusion, our data reveals that TNFSF9 may become a predictive biomarker of pancreatic cancer and provides a new intervention target for the immunotherapy of pancreatic cancer.

Currently, colorectal cancer (CRC) predictions are based on an early diagnosis and the tumor-node-metastasis (TNM) stage, but the outcomes of patients with the same cancer type are difficult to predict. Novel molecular tests for the early diagnosis and stratification of CRC patients must be devised. After our initial bioinformatics screen, we examined zinc finger and BTB domain-containing 7C (ZBTB7C). To date, few studies have investigated ZBTB7C in CRC, necessitating further analyses of its expression and regulatory mechanism in CRC. ZBTB7C mRNA and protein expression was detected in CRC and corresponding non-CRC tissues. We evaluated the relationship between clinical prognosis and ZBTB7C protein levels using Cox regression analysis and Kaplan-Meier curves. A receiver operating characteristic (ROC) curve was generated to verify the diagnostic performance of ZBTB7C levels in CRC. Several bioinformatics techniques were applied to analyze the potential molecular mechanism of ZBTB7C. Low mRNA and protein levels of ZBTB7C were detected in tumor tissues from CRC patients. The survival curve predicted a poor prognosis for CRC patients exhibiting low ZBTB7C expression (P=0.001). According to the univariate Cox regression analysis, older age, a high TNM stage and low ZBTB7C expression were responsible for poor outcomes in CRC patients. The multivariate analysis further revealed ZBTB7C as an independent prognostic factor for CRC (P=0.015). The area under the curve of ZBTB7C expression for CRC diagnosis was 0.970 (95% confidence interval, 0.9447-0.9946; P < 0.0001). According to in silico analyses, genes coexpressed with ZBTB7C are associated mainly with the Ras and Wnt signaling pathways. Overall, ZBTB7C is downregulated in CRC and represents an early diagnostic marker and independent prognostic factor for CRC. ZBTB7C may be functionally mediated by different pathways or targeting miRNAs.

Stroke is a major global health problem, with the prevalence and economic burden predicted to increase due to aging populations in western society. Following stroke, numerous biochemical alterations occur and damage can spread to nearby tissue. This zone of “at risk” tissue is termed the peri-infarct zone (PIZ). As the PIZ contains tissue not initially damaged by the stroke, it is considered by many as salvageable tissue. For this reason, much research effort has been undertaken to improve the identification of the PIZ and to elucidate the biochemical mechanisms that drive tissue damage in the PIZ in the hope of identify new therapeutic targets. Despite this effort, few therapies have evolved, attributed in part, to an incomplete understanding of the biochemical mechanisms driving tissue damage in the PIZ. Magnetic resonance imaging (MRI) has long been the gold standard to study alterations in gross brain structure, and is frequently used to study the PIZ following stroke. Unfortunately, MRI does not have sufficient spatial resolution to study individual cells within the brain, and reveals little information on the biochemical mechanisms driving tissue damage. MRI results may be complemented with histology or immuno-histochemistry to provide information at the cellular or sub-cellular level, but are limited to studying biochemical markers that can be successfully “tagged” with a stain or antigen. However, many important biochemical markers cannot be studied with traditional MRI or histology/histochemical methods. Therefore, we have developed and applied a multi-modal imaging platform to reveal elemental and molecular alterations that could not previously be imaged by other traditional methods. Our imaging platform incorporates a suite of spectroscopic imaging techniques; Fourier transform infrared imaging, Raman spectroscopic imaging, Coherent anti-stoke Raman spectroscopic imaging and X-ray fluorescence imaging. This approach does not preclude the use of traditional imaging techniques, and rather it should be use to complement traditional methods such as MRI or histology and immunohistochemistry, to gain a greater insight into disease mechanisms. We demonstrate the potential of this approach by characterizing biochemical alterations within the PIZ 24 h after the induction of photothrombotic stroke in mice. Substantial molecular and elemental alterations were identified in the PIZ 24 h after stroke that are consistent with tissue swelling and edema, but not oxidative stress. This reveals important mechanistic information, that could not previously be obtained, which should be considered in future studies aimed at developing therapeutic intervention from this model.

Clathrin-mediated endocytosis (CME) is an important entry pathway for viruses. Here, we applied click chemistry to covalently immobilize reovirus on surfaces to study CME during early host-pathogen interactions. To uncouple chemical and physical properties of viruses and determine their impact on CME initiation, we used the same strategy to covalently immobilize nanoparticles of different sizes. Using fluorescence live microscopy and electron microscopy, we confirmed that clathrin recruitment depends on particle size and discovered that the maturation into clathrin-coated vesicles (CCVs) is independent from cargo internalization. Surprisingly, we found that the final size of CCVs appears to be imprinted on the clathrin coat at early stages of cargo-cell interactions. Our approach has allowed us to unravel novel aspects of early interactions between viruses and the clathrin machinery that influence late stages of CME and CCVs formation. This method can be easily and broadly applied to the field of nanotechnology, endocytosis, and virology.

The cellular environment impacts a myriad of cellular functions by providing signals that can modulate cell phenotype and function. Physical cues such as topography, roughness, gradients, and elasticity are of particular importance. Thus, synthetic substrates can be potentially useful tools for exploring the influence of the aforementioned physical properties on cellular function. Many micro- and nanofabrication processes have been employed to control substrate characteristics in both 2D and 3D environments. This review highlights strategies for modulating the physical properties of surfaces, the influence of these changes on cell responses, and the promise and limitations of these surfaces in in-vitro settings. While both hard and soft materials are discussed, emphasis is placed on soft substrates. Moreover, methods for creating synthetic substrates for cell studies, substrate properties, and impact of substrate properties on cell behavior are the main focus of this review.

The chemical basis for the alteration of the refractive properties of an intraocular lens with a femtosecond laser was investigated. Three different microscope setups have been used for the study: Laser Induced Fluorescence (LIF) microscopy, Raman microscopy and coherent anti-Stokes Raman Scattering (CARS) microscopy. Photo-induced hydrolysis of polymeric material in aqueous media produces two hydrophilic functional groups: acid group and alcohol group. The spectral signatures identify two of the hydrophilic polar molecules as N-phenyl-4-(phenylazo)-benzenamine (C18H15N3) and phenazine-1-carboxylic acid (C13H8N2O2). The change in hydrophilicity results in a negative refractive index change in the laser-treated areas.

A gel electrolyte membrane is obtained through the absorption of a carbamate-modified liquid disiloxane-containing lithium bis(trifluoromethane)sulfonimide (LiTFSI) by using macroporous poly(vinylidene fluoride-hexafluoropropylene) (PVDF-HFP) membranes. The porous membranes are prepared by means of a phase inversion technique. The resulting gel electrolyte membrane is studied by using differential scanning calorimetry, Fourier-transform infrared (FTIR) spectroscopy, and microscope mapping through coherent anti-Stokes Raman scattering (CARS) confocal microscopy and impedance spectroscopy. The ionic conductivity of the gel electrolyte is 10(-4) S cm(-1) at 20 °C. FTIR spectroscopy reveals interactions between LiTFSI and the carbonyl moiety of the disiloxane. No interactions between LiTFSI and PVDF-HFP or between disiloxane and PVDF-HFP are detected by FTIR spectroscopy. Furthermore, the distribution of the α and β/γ phases of PVDF-HFP and the homogeneous distribution of disiloxane/LiTFSI in the gel electrolyte membranes are examined by FTIR mapping. CARS confocal microscopy is used to image the three-dimensional interconnectivity, which reveals a reticulated structure of macrovoids in the porous PVDF-HFP framework. Owing to properties such as electrochemical and thermal stability of the disiloxane-based liquid electrolyte and the mechanical stability of the porous PVDF-HFP membrane, the gel electrolyte membranes presented herein are promising candidates for applications as electrolytes/separators in lithium-ion batteries.

2010 WILEY-VCH Verlag Gm Mechanical interactions between cells and their environment play an important role in regulating many cellular functions, such as migration, differentiation, and proliferation. They are also involved in complex processes like tissue formation during development of multicellular organisms. During the last two decades, several methods have been developed to visualize and measure traction forces produced by single cells. Initial experiments were performed by growing fibroblasts on a thin silicon rubber substrate that was deformed by forces exerted by migrating cells. Subsequently, elastic polyacrylamide substrates embedded with fluorescent latex beads were used to get an insight into the force vectors exerted by the cells. These methods were further improved by producing regular arrays of traceable markers in the elastic substrates allowing a quantification of traction forces exerted by adhering cells. A different approach has used arrays of elastic posts positioned on a flat 2D substrate, coated with extracellular-matrix (ECM)molecules. Because each post effectively acts like a harmonic spring, local traction forces at multiple cell adhesion sites could then be measured by simply monitoring post deflections. A modification of this assay was recently applied to measure contraction forces of single cardiomyocytes. Although these different approaches have provided valuable insight into our understanding of cellular forces, they have the disadvantage that the cells are forced into a 2D growth pattern, which significantly differs from the in vivo situation. Therefore, the development of in vitro model systems that capture more of the complexity present in 3D tissue scaffolds are highly desirable. Technologies for fabricating macroscopic tissue architectures in the millimeter and micrometer range have become available only during recent years. In contrast, methods to reliably produce flexible 3D cellular environments in the micrometer to nanometer range are still missing. Ideally, these scaffolds should have a controllable distribution of cell-substrate contact sites and an adjustable stiffness. In this Communication, we show that such biocompatible scaffolds can be fabricated by means of direct laser writing (DLW) and subsequent surface functionalization. We furthermore demonstrate that these scaffolds can be rhythmically deformed by single beating cardiomyocytes. In addition, we quantitatively evaluate the involved contraction forces by calibrating these structures using atomic force microscopy (AFM). In particular, with the method presented here, beamlike structures with defined geometries and physiologically relevant stiffness values can be produced; thus, these scaffolds structurally and mechanically mimic large macromolecules surrounding cells in 3D tissues, such as collagen fibrils. In essence, DLW (see the Experimental section) can be thought of as a pencil of light in three dimensions. Any connected structure envisioned on a computer can easily be implemented by scanning a photoresist relative to the focus of a femtosecond-laser beam (Fig. 1a). After development of a negative resist, sufficiently exposed regions correspond to the remaining polymer (Fig. 1b). Lithography of 3D polymeric templates by DLW with lateral feature sizes down to the 100 nm range has become routine in the field of nanophotonics. Along these lines, fully 3D (Fig. 1c) scaffolds can be fabricated. Sample footprints of mm up to cm dimensions can be realized by combining 300mm 300mm areas that are accessible within the piezo scanning range. The height of the structures is limited only by the working distance of the microscope lens (80mm in our case). Typical writing times for the structures shown in this Communication are less than 10min. Here, we employ the commercially available resist system Ormocomp (obtained from Micro Resist Technology GmbH). Ormocomp is a member of Ormocer, a class of inorganic (Si O Si)–organic hybrid polymer systems, previously developed at the Fraunhofer Institute for Silica Research (Wurzburg, Germany). Its detailed chemical composition is proprietary. With Ormocomp as a photoresist, complex fully 3D structures (Fig. 1a–c) can be easily manufactured and, after functionalization with fibronectin, directly used for cell culture (Fig. 1d). To demonstrate that single cells can lead to noticeable deformation of the 3D Ormocomp scaffolds, primary cardiomyocytes are isolated from chicken embryos (see the

Graded distributions of proteins are pivotal for many signaling processes during development, such as morphogenesis, cell migration, and axon guidance. Here, we describe a technique to fabricate substrate-bound stepwise protein gradients by means of a microfluidic network etched into a silicon wafer with an array of parallel 14-micrometer–wide channels, which can be filled with a series of arbitrarily chosen protein solutions. In a subsequent microcontact printing step, the protein pattern is transferred onto a surface and is used as a substrate for cell culture. Cellular responses to a defined microscopic pattern of a protein, such as guided axonal outgrowth and directed migration, cell polarization, changes in morphology, and signaling, can be thus studied in a controlled in vitro environment.