Neuroprotective effects of anti-TRAIL-ICG nanoagent and its multimodal imaging evaluation in cerebral ischemia-reperfusion injury.

Cerebral ischemia-reperfusion injury (CIRI) is a major challenge to neuronal survival in acute ischemic stroke (AIS). However, effective neuroprotective agents remain to be developed for the treatment of CIRI. In this work, we have developed an Anti-TRAIL protein-modified and indocyanine green (ICG)-responsive nanoagent (Anti-TRAIL-ICG) to target ischemic areas and then reduce CIRI and rescue the ischemic penumbra. In vitro and in vivo experiments have demonstrated that the carrier-free nanoagent can enhance drug transport across the blood-brain barrier (BBB) in stroke mice, exhibiting high targeting ability and good biocompatibility. Anti-TRAIL-ICG nanoagent played a better neuroprotective role by reducing apoptosis and ferroptosis, and significantly improved ischemia-reperfusion injury. Moreover, the multimodal imaging platform enables the dynamic in vivo examination of multiple morphofunctional information, so that the dynamic molecular events of nanoagent can be detected continuously and in real time for early treatment in transient middle cerebral artery occlusion (tMCAO) models. Furthermore, it has been found that Anti-TRAIL-ICG has great potential in the functional reconstruction of neurovascular networks through optical coherence tomography angiography (OCTA). Taken together, our work effectively alleviates CIRI after stoke by blocking multiple cell death pathways, which offers an innovative strategy for harnessing the apoptosis and ferroptosis against CIRI.

Experimental Investigation for Proton Exchange Membrane Fuel Cells in Marine Salt Spray Environment.

In the maritime setting, Proton Exchange Membrane Fuel Cells (PEMFCs) are subjected to salt spray, posing a risk of contaminating internal components and leading to irreversible degradation in the performance of the PEMFCs. Thus, it is crucial to assess the impact of sodium chloride contamination on PEMFC operation. To address challenges related to prolonged cycle times, high costs, and intricate sample preparation in sodium chloride contamination experiments for PEMFCs, this Article replicates the marine atmospheric conditions using a standard salt spray experimental chamber. The liquid nitrogen fracture method is employed for cost-effective and efficient preparation of experimental samples. The meteorological environment with varying salt content in the salt spray is achieved through precise control of sodium chloride concentration. The Article systematically presents the salt spray experimental method for the membrane electrode assembly (MEA) of PEMFCs. A dedicated salt spray experimental rig was constructed to validate this method for the MEA of PEMFCs. The results indicate that the salt spray experimental method for the MEA of PEMFCs can effectively explore internal component contamination and is well-suited for analyzing the physicochemical effects of NaCl on MEA components, along with their microscopic characterization under salt spray conditions.

ScRNA-seq revealed the tumor microenvironment heterogeneity related to the occurrence and metastasis in upper urinary tract urothelial carcinoma.

Metastasis is the greatest clinical challenge for UTUCs, which may have distinct molecular and cellular characteristics from earlier cancers. Herein, we provide single-cell transcriptome profiles of UTUC para cancer normal tissue, primary tumor lesions, and lymphatic metastases to explore possible mechanisms associated with UTUC occurrence and metastasis. From 28,315 cells obtained from normal and tumor tissues of 3 high-grade UTUC patients, we revealed the origin of UTUC tumor cells and the homology between metastatic and primary tumor cells. Unlike the immunomicroenvironment suppression of other tumors, we found no immunosuppression in the tumor microenvironment of UTUC. Moreover, it is imperative to note that stromal cells are pivotal in the advancement of UTUC. This comprehensive single-cell exploration enhances our comprehension of the molecular and cellular dynamics of metastatic UTUCs and discloses promising diagnostic and therapeutic targets in cancer-microenvironment interactions.

Long-term Intravital Investigation of an Orthotopic Glioma Mouse Model via Optical Coherence Tomography Angiography.

BACKGROUND/AIM: Probing brain tumor microvasculature holds significant importance in both basic cancer research and medical practice for tracking tumor development and assessing treatment outcomes. However, few imaging methods commonly used in clinics can noninvasively monitor the brain microvascular network at high precision and without exogenous contrast agents in vivo. The present study aimed to investigate the characteristics of microvasculature during brain tumor development in an orthotopic glioma mouse model. MATERIALS AND METHODS: An orthotopic glioma mouse model was established by surgical orthotopic implantation of U87-MG-luc cells into the mouse brain. Then, optical coherence tomography angiography (OCTA) was utilized to characterize the microvasculature progression within 14 days. RESULTS: The orthotopic glioma mouse model evaluated by bioluminescence imaging and MRI was successfully generated. As the tumor grew, the microvessels within the tumor area slowly decreased, progressing from the center to the periphery for 14 days. CONCLUSION: This study highlights the potential of OCTA as a useful tool to noninvasively visualize the brain microvascular network at high precision and without any exogenous contrast agents in vivo.

Irreversible Electroporation-Induced Inflammation Facilitates Neutrophil-Mediated Drug Delivery to Enhance Pancreatic Cancer Therapy.

Pancreatic cancer is a deadly disease with a five-year overall survival rate of around 11%. Chemotherapy is a cornerstone in the treatment of this malignancy, but the intratumoral delivery of chemotherapy drugs is impaired by the highly fibrotic tumor-associated stroma. Irreversible electroporation (IRE) is an ablative technique for treating locally advanced pancreatic cancer. During a typical IRE procedure, high-intensity electric pulses are released to kill tumor cells through the irreversible disruption of the cytoplasm membranes. IRE also induces rapid tumor infiltration by neutrophils and offers an opportunity for neutrophil-mediated drug delivery. We herein showed that the IRE-induced neutrophil trafficking was facilitated by the upregulation of neutrophil chemotaxis and migration as well as the release of several chemoattractants. Doxorubicin-loaded bovine serum albumin nanoparticles were prepared and loaded into neutrophils at a ratio of 9.9 ± 1.2 to 11.7 ± 2.0 pg of doxorubicin per cell. The resultant formulation (NP@NEs) efficiently accumulated in the IRE-treated KPC-A377 murine pancreatic tumors with an uptake value of 10.7 ± 1.5 (percent of injected dose per gram of tissue, abbreviated as %ID/g) at 48 h after intravenous injection. In both Panc02 and KPC-A377 murine pancreatic tumor models, the combination of IRE + NP@NEs inhibited tumor growth more effectively than either monotherapy. The tumors treated with the combination also exhibited the lowest frequency of Ki67+ proliferating cells and the highest abundance of terminal deoxynucleotidyl transferase dUTP nick end labeling+ (TUNEL+) apoptotic cells among the experiment groups. Minimal treatment-associated toxicity was observed. Our findings suggest that neutrophil-mediated delivery of chemotherapy drugs is a useful tool to enhance the response of pancreatic cancer to IRE.

Internal Light Sources-Mediated Photodynamic Therapy Nanoplatforms: Hope for the Resolution of the Traditional Penetration Problem.

Photodynamic therapy (PDT) is an alternative cancer treatment technique with a noninvasive nature, high selectivity, and minimal adverse effects. The indispensable light source used in PDT is a critical factor in determining the energy conversion of photosensitizers (PSs). Traditional light sources are primarily concentrated in the visible light region, severely limiting their penetration depth and making them prone to scattering and absorption when applied to biological tissues. For that reason, its efficacy in treating deep-seated lesions is often inadequate. Self-exciting PDT, also known as auto-PDT (APDT), is an attractive option for circumventing the limited penetration depth of traditional PDT and has acquired significant attention. APDT employs depth-independent internal light sources to excite PSs through resonance or radiative energy transfer. APDT has considerable potential for treating deep-tissue malignancies. To facilitate many researchers' comprehension of the latest research progress in this field and inspire the emergence of more novel research results. This review introduces internal light generation mechanisms and characteristics and provides an overview of current research progress based on the recently reported APDT nanoplatforms. The current challenges and possible solutions of APDT nanoplatforms are also presented and provide insights for future research in the final section of this article.

Altered NCR3 Splice Variants May Result in Deficient NK Cell Function in Renal Cell Carcinoma Patients.

BACKGROUND/AIM: The natural killer (NK) cell function of patients with malignant tumours may be suppressed by deficiency, and the poor prognosis of renal cell carcinoma (RCC) patients may be due to escape from NK cell cytotoxicity, especially with respect to natural cytotoxicity receptors (NCRs) on the NK cell surface. However, the specific mechanism remains unclear. Therefore, in this study, we sought to explore the role of NCR, especially NCR3 splice variants, in the process of NK cell deficiency in RCC patients. MATERIALS AND METHODS: We used flow cytometry to analyse the phenotype of NK cells from the peripheral blood and kidney tumour tissue of RCC patients. The NKp30-mediated NK cell killing function was measured by antibody-dependent cell-mediated cytotoxicity (ADCC) in NK and RCC cell coincubation. We extracted RNA from the peripheral blood mononuclear cells (PBMCs) of RCC patients and renal carcinoma tissue and carried out real-time quantitative PCR to detect the mRNA levels of NKp30a, NKp30b and NKp30c. mRNA expression levels of cytokines (IL-6, IL-8, IL-10, IL-18 and TGF-ß) based on RNA extracted from renal carcinoma tissue and adjacent normal kidney tissues were also measured by real-time quantitative PCR. RESULTS: Regarding the phenotype of NK cells in RCC patients, the proportion of NK cells in tumour tissue was significantly reduced, with changes in the NK cell proportion being most obvious in NKp30+ NK cells. Furthermore, the results of the ADCC function assay showed limited NKp30+ NK cell-mediated cytotoxicity in RCC patients. Through real-time quantitative PCR, we found lower expression of NKp30a and NKp30b, the immunostimulatory splice variants of NCR3 encoding NKp30, in RCC patients. Moreover, expression of activating cytokines (IL-6 and IL-8) in renal cancer tissue was decreased, though inhibitory cytokine (TGF-ß) expression remained unchanged, which may result in an immunosuppressive cytokine microenvironment. CONCLUSION: Decreased expression of immunostimulatory NCR3 splice variants and the inhibitory cytokine microenvironment in RCC patients may contribute to deficient NK cell cytotoxicity and renal carcinoma cell immune escape from NK cell killing, which may provide a theoretical basis for finding new immunotherapeutic targets for RCC.

NOTCH3 promotes docetaxel resistance of prostate cancer cells through regulating TUBB3 and MAPK signaling pathway.

Docetaxel is the preferred chemotherapeutic agent in patients with castrate-resistant prostate cancer (CRPC). However, patients eventually develop docetaxel resistance and in the absence of effective treatment options. Consequently, it is essential to investigate the mechanisms generating docetaxel resistance and develop novel alternative therapeutic targets. RNA sequencing was undertaken on docetaxel-sensitive and docetaxel-resistant prostate cancer (PCa) cells. Subsequently, chemoresistance, cancer stemness, and lipid metabolism were investigated. To obtain insight into the precise activities and action mechanisms of NOTCH3 in docetaxel-resistant PCa, immunoprecipitation, mass spectrometry, ChIP, luciferase reporter assay, cell metabolism, and animal experiments were performed. Through RNA sequencing analysis, we found that NOTCH3 expression was markedly higher in docetaxel-resistant cells relative to parental cells, and that this trend was continued in docetaxel-resistant PCa tissues. Experiments in vitro and in vivo revealed that NOTCH3 enhanced stemness, lipid metabolism, and docetaxel resistance in PCa. Mechanistically, NOTCH3 is bound to TUBB3 and activates the MAPK signaling pathway. Moreover, NOTCH3 was directly regulated by MEF2A in docetaxel-resistant cells. Notably, targeting NOTCH3 and the MEF2A/TUBB3 signaling axis was related to docetaxel chemoresistance in PCa. Overall, these results demonstrated that NOTCH3 fostered stemness, lipid metabolism, and docetaxel resistance in PCa via the TUBB3 and MAPK signaling pathways. Therefore, NOTCH3 may be employed as a prognostic biomarker in PCa patients. NOTCH3 could be a therapeutic target for PCa patients, particularly those who have developed docetaxel resistance.

Effects of Cathode GDL Gradient Porosity Distribution along the Flow Channel Direction on Gas-Liquid Transport and Performance of PEMFC.

The gas diffusion layer (GDL) is an important component of proton exchange membrane fuel cells (PEMFCs), and its porosity distribution has considerable effects on the transport properties and durability of PEMFCs. A 3-D two-phase flow computation fluid dynamics model was developed in this study, to numerically investigate the effects of three different porosity distributions in a cathode GDL: gradient-increasing (Case 1), gradient-decreasing (Case 3), and uniform constant (Case 2), on the gas-liquid transport and performance of PEMFCs; the novelty lies in the porosity gradient being along the channel direction, and the physical properties of the GDL related to porosity were modified accordingly. The results showed that at a high current density (2400 mA·cm-2), the GDL of Case 1 had a gas velocity of up to 0.5 cm·s-1 along the channel direction. The liquid water in the membrane electrode assembly could be easily removed because of the larger gas velocity and capillary pressure, resulting in a higher oxygen concentration in the GDL and the catalyst layer. Therefore, the cell performance increased. The voltage in Case 1 increased by 8% and 71% compared to Cases 2 and 3, respectively. In addition, this could ameliorate the distribution uniformity of the dissolved water and the current density in the membrane along the channel direction, which was beneficial for the durability of the PEMFC. The distribution of the GDL porosity at lower current densities had a less significant effect on the cell performance. The findings of this study may provide significant guidance for the design and optimization of the GDL in PEMFCs.

A novel scheme for the utilization of Cu slag flotation tailings in preparing internal electrolysis materials to degrade printing and dyeing wastewater.

About 60 million tons of Fe-rich Cu slag (IRCS) are generated annually worldwide during Cu slag flotation and cause irreversible water and soil pollution. Current research provides an environmentally friendly technology, the preparation of internal electrolysis materials (IEMs) through the carbothermal reduction of IRCS, for the degradation of printing and dyeing wastewater. XRD and SEM-EDS indicated that carbothermal reduction could promote the conversion of fayalite to zero-valent iron (ZVI), and ZVI could effectively form IEM with residual carbon. The degradation capacity of IEM for methylene blue (MB) was remarkably improved compared with raw IRCS after roasting for 60 min at 1100 °C with 35% anthracite dosage. MB degradation efficiency improved by increasing the IEM dosage and reaction temperature and decreasing the MB concentration and solution pH. FTIR, XRD, SEM-EDS, and XPS all detected the formation of Fe oxide or Fe hydroxide. UV-vis and TOC demonstrated that the characteristic groups of MB were destroyed and resulted in the mineralization of MB. MB degradation could be attributed to the Fe2+, [H], and ·OH produced by the galvanic reaction induced by IEM. Overall, this study offers theoretical guidance in the treatment of printing and dyeing wastewater and the reuse of IRCS.

Study protocol for a single-centre non-inferior randomised controlled trial on a novel three-dimensional matrix positioning-based cognitive fusion-targeted biopsy and software-based fusion-targeted biopsy for the detection rate of clinically significant prostate cancer in men without a prior biopsy.

INTRODUCTION: The classical pathway for diagnosing prostate cancer is systematic 12-core biopsy under the guidance of transrectal ultrasound, which tends to underdiagnose the clinically significant tumour and overdiagnose the insignificant disease. Another pathway named targeted biopsy is using multiparametric MRI to localise the tumour precisely and then obtain the samples from the suspicious lesions. Targeted biopsy, which is mainly divided into cognitive fusion method and software-based fusion method, is getting prevalent for its good performance in detecting significant cancer. However, the preferred targeted biopsy technique in detecting clinically significant prostate cancer between cognitive fusion and software-based fusion is still beyond consensus. METHODS AND ANALYSIS: This trial is a prospective, single-centre, randomised controlled and non-inferiority study in which all men suspicious to have clinically significant prostate cancer are included. This study aims to determine whether a novel three-dimensional matrix positioning cognitive fusion-targeted biopsy is non-inferior to software-based fusion-targeted biopsy in the detection rate of clinically significant cancer in men without a prior biopsy. The main inclusion criteria are men with elevated serum prostate-specific antigen above 4-20 ng/mL or with an abnormal digital rectal examination and have never had a biopsy before. A sample size of 602 participants allowing for a 10% loss will be recruited. All patients will undergo a multiparametric MRI examination, and those who fail to be found with a suspicious lesion, with the anticipation of half of the total number, will be dropped. The remaining participants will be randomly allocated to cognitive fusion-targeted biopsy (n=137) and software-based fusion-targeted biopsy (n=137). The primary outcome is the detection rate of clinically significant prostate cancer for cognitive fusion-targeted biopsy and software-based fusion-targeted biopsy in men without a prior biopsy. The clinically significant prostate cancer will be defined as the International Society of Urological Pathology grade group 2 or higher. ETHICS AND DISSEMINATION: Ethical approval was obtained from the ethics committee of Shanghai East Hospital, Tongji University School of Medicine, Shanghai, China. The results of the study will be disseminated and published in international peer-reviewed journals. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov Registry (NCT04271527).

CHD1 and SPOP synergistically protect prostate epithelial cells from DNA damage.

BACKGROUND: Recent genomic profiling has identified a subtype of prostate cancer (PCa) characterized by two key genetic alterations: missense mutation of speckle-type POZ protein (SPOP) and homozygous deletion of chromodomain helicase DNA-binding protein 1 (CHD1). Mutually exclusive with E26 transformation-specific (ETS) rearrangements, this subtype displays high genomic instability. Previous studies indicate that deficient SPOP or CHD1 alone leads to feeble prostate abnormalities and each protein is involved in DNA damage response (DDR). It remains to be determined whether CHD1 and SPOP cooperate to suppress prostate tumorigenesis and DDR. METHODS: Prostate-specific single or double knockout of Spop and Chd1 was generated with the Cre/loxP system in mice. Wild-type or mutant SPOP (F102C, F133V) overexpression and CHD1 knockdown with short hairpin RNA were created in human benign prostatic hyperplasia cell line BPH1. The levels of DNA damage and homologous recombination repair were measured by immunofluorescence staining of γH2AX and RAD51, respectively. RESULTS: Spop/Chd1 double-knockout mice displayed prostatic intraepithelial neoplasia at both young (3 months) and old (12 months) ages and failed to generate prostate adenocarcinoma. Compared with wild-type or single-knockout mice, the double-knockout prostate harbored moderately higher proliferating cells and dramatically augmented the level of γH2AX staining, although androgen receptor-positive cells and apoptotic cells remained at a similar level. In BPH1 cell line, SPOP mutant overexpression and CHD1 silencing synergistically sensitized the cells to DNA damage by camptothecin, an inducer of double-strand breaks. CONCLUSIONS: Our results indicate that SPOP and CHD1 can synergistically promote repair of naturally occurring or chemically induced DNA damages in prostate epithelial cells. Regarding the progression of the SPOP/CHD1 subtype of PCa, other functionally complementary drivers warrant further identification. The clinical implication is that this subtype of PCa may be particularly sensitive to poly(ADP-ribose) polymerase inhibitors or DNA-damaging agents.

Portfolio Targeting Strategy To Realize the Assembly and Membrane Fusion-Mediated Delivery of Gold Nanoparticles to Mitochondria for Enhanced NIR Photothermal Therapies.

Targeting mitochondria has always been a challenging goal for therapeutic nanoparticle agents due to their heterotypic features and size, which usually lead to a lysosome/endosome endocytosis pathway. To overcome this limitation, in this work, a portfolio targeting strategy combining a small targeting molecule with a biomembrane was developed. Modification of small targeting molecule H2N-TPP on gold nanoparticles (GNPs) could not only facilitate the mitochondrial targeting but could also induce gold nanoparticle assembly. Therefore, the GNPs were endowed with good absorption and photothermal conversion abilities in the near-infrared (NIR) region. Meanwhile, a biomimetic strategy was adopted by wrapping the gold nanoparticle assembly (GNA) with cancer cell membranes (CCMs), which helped the GNA enter the prostatic cancer cell via a homotypic membrane-fusion process to avoid being trapped in endosomes/lysosomes. Thereafter, the GNA remaining in the cytoplasm could reach mitochondria more efficiently via guidance from H2N-TPP molecules. This "biomembrane-small molecule" combination targeting process was evidenced by fluorescence microscopy, and the highly efficient photothermal ablation of prostatic tumors in vivo was demonstrated. This portfolio targeting strategy could be extended to various nanodrugs/agents to realize an accurate subcellular targeting efficiency for cancer treatments or cell detections.

Polymorphonuclear MDSCs are enriched in the stroma and expanded in metastases of prostate cancer.

Myeloid-derived suppressor cells with polymorphonuclear morphology (PMN-MDSCs) contribute to the progression and immune evasion of prostate cancer. However, the spatial distribution of tumor-infiltrating PMN-MDSCs in primary and metastatic prostate cancer, especially in the context of comparison between the epithelial and stromal compartments of the tumor, has not been characterized. Here, we describe a multicolor immunofluorescence staining study of 90 primary tumors, 37 lymph node metastases (all with matched primary tumors) and 35 bone metastases using archived samples. CD11b+ CD15+ cells were identified as PMN-MDSCs and pan-cytokeratin+ cells were identified as prostate epithelial cells. We found that, in both primary tumor and metastases, PMN-MDSCs infiltrate much more readily in the stromal area compared with the epithelial area of the tumor regions. In comparison to the stromal area of primary tumors, the stromal area of either lymph node metastases or bone metastases was infiltrated with more PMN-MDSCs. In primary tumors, stromal PMN-MDSCs were associated with vascularization, segmented neutrophils, patient age and close juxtaposition to neoplastic epithelial cells. These results reveal the stroma rather than the epithelia of prostate cancer as the major hotbed for PMN-MDSCs and support the role of PMN-MDSCs in the metastatic progression of prostate cancer.

Intercellular Crosstalk of Mesenchymal Stem Cells with Prostate Cancer Cells via Microvesicles Loaded with Magnetic Nanocubes for Targeted Magnetic Hyperthermia.

The targeted delivery of nanomedicines into solid tumors remains challenging in cancer treatment. Stem cells with tumortropic migration ability are promising as biocarriers to transport nanomedicines. The transportation of nanomedicines into cancer cells is the key step for tumor targeted delivery via stem cells. In this study, we designed a magnetic nanocube (scMNP) loaded in mesenchymal stem cells for magnetic hyperthermia of prostate cancer, and the delivery and transportation pathways into the cancer cells were fully investigated. The MSCs acted as the carrier of the loaded scMNPs along with the upregulation of CXCR4 for the migration to cancer cells. The therapeutic effect was mainly due to scMNPs via magnetic hyperthermia. Stem cell-derived microvesicles containing scMNPs played an essential role in the crosstalk between stem cells and cancer cells for targeted delivery. Both in vitro and in vivo studies demonstrated that the system showed satisfactory therapeutic efficiency under magnetic hyperthermia therapy. Our investigation presents a comprehensive study of magnetic nanoparticles in combination with MSCs and their extracellular microvesicles and is promising as an effective strategy for magnetic hyperthermia therapy of prostate cancer.

Copper-64 Labeled PEGylated Exosomes for In Vivo Positron Emission Tomography and Enhanced Tumor Retention.

Exosomes have attracted tremendous attention due to their important role in physiology, pathology, and oncology, as well as promising potential in biomedical applications. Although great efforts have been dedicated to investigating their biological properties and applications as natural cancer drug-delivery systems, the systemic biodistribution of exosomes remains underexplored. In addition, exosome-based drug delivery is inevitably hindered by the robust liver clearance, leading to suboptimal tumor retention and therapeutic efficiency. In this study, we report one of the first examples using in vivo positron emission tomography (PET) for noninvasive monitoring of copper-64 (64Cu)-radiolabeled polyethylene glycol (PEG)-modified exosomes, achieving excellent imaging quality and quantitative measurement of blood residence and tumor retention. PEGylation not only endowed exosomes with a superior pharmacokinetic profile and great accumulation in the tumor versus traditionally reported native exosomes but also reduced premature hepatic sequestration and clearance of exosomes, findings that promise enhanced therapeutic delivery efficacy and safety in future studies. More importantly, this study provides important guidelines about surface engineering, radiochemistry, and molecular imaging in obtaining accurate and quantitative biodistribution information on exosomes, which may benefit future exploration in the realm of exosomes.

Nitropeptide Profiling and Identification Illustrated by Angiotensin II.

Protein nitration is one of the most important post-translational modifications (PTM) on tyrosine residues and it can be induced by chemical actions of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in eukaryotic cells. Precise identification of nitration sites on proteins is crucial for understanding the physiological and pathological processes related to protein nitration, such as inflammation, aging, and cancer. Since the nitrated proteins are of low abundance in cells even under induced conditions, no universal and efficient methods have been developed for the profiling and identification of protein nitration sites. Here we describe a protocol for nitropeptide enrichment by using a chemical reduction reaction and biotin labeling, followed by high resolution mass spectrometry. In our method, nitropeptide derivatives can be identified with high accuracy. Our method exhibits two advantages compared to the previously reported methods. First, dimethyl labeling is used to block the primary amine on nitropeptides, which can be used to generate quantitative results. Second, a disulfide bond containing NHS-biotin reagent is used for the enrichment, which can be further reduced and alkylated to enhance the detection signal on a mass spectrometer. This protocol has been successfully applied to the model peptide Angiotensin II in the current paper.

Irreversible electroporation reverses resistance to immune checkpoint blockade in pancreatic cancer.

Immunotherapy has only limited efficacy against pancreatic ductal adenocarcinoma (PDAC) due to the presence of an immunosuppressive tumor-associated stroma. Here, we demonstrate an effective modulation of that stroma by irreversible electroporation (IRE), a local ablation technique that has received regulatory approval in the United States. IRE induces immunogenic cell death, activates dendritic cells, and alleviates stroma-induced immunosuppression without depleting tumor-restraining collagen. The combination of IRE and anti-programmed cell death protein 1 (anti-PD1) immune checkpoint blockade promotes selective tumor infiltration by CD8+ T cells and significantly prolongs survival in a murine orthotopic PDAC model with a long-term memory immune response. Our results suggest that IRE is a promising approach to potentiate the efficacy of immune checkpoint blockade in PDAC.

Intelligent Photosensitive Mesenchymal Stem Cells and Cell-Derived Microvesicles for Photothermal Therapy of Prostate Cancer.

Targeted delivery of nanomedicines into the tumor site and improving the intratumoral distribution remain challenging in cancer treatment. Here, we report an effective transportation system utilizing both of mesenchymal stem cells (MSCs) and their secreted microvesicles containing assembled gold nanostars (GNS) for targeted photothermal therapy of prostate cancer. The stem cells act as a cell carrier to actively load and assemble GNS into the lysosomes. Accumulation of GNS in the lysosomes facilitates the close interaction of nanoparticles, which could result in a 20 nm red-shift of surface plasmon resonance of GNS with a broad absorption in the near infrared region. Moreover, the MSCs can behave like an engineering factory to pack and release the GNS clusters into microvesicles. The secretion of GNS can be stimulated via light irradiation, providing an external trigger-assisted approach to encapsulate nanoparticles into cell derived microvesicles. In vivo studies demonstrate that GNS-loaded MSCs have an extensive intratumoral distribution, as monitored via photoacoustic imaging, and efficient antitumor effect under light exposure in a prostate-cancer subcutaneous model by intratumoral and intravenous injection. Our work presents a light-responsive transportation approach for GNS in combination of MSCs and their extracellular microvesicles and holds the promise as an effective strategy for targeted cancer therapy including prostate cancer.