Crizotinib-based proteolysis targeting chimera suppresses gastric cancer by promoting MET degradation.

As one of the common malignant cancer types, gastric cancer (GC) is known for late-stage diagnosis and poor prognosis. Overexpression of the receptor tyrosine kinase MET is associated with poor prognosis among patients with advanced stage GC. However, no MET inhibitor has been used for GC treatment. Like other tyrosine kinase inhibitors that fit the "occupancy-driven" model, current MET inhibitors are prone to acquired resistance. The emerging proteolysis targeting chimera (PROTAC) strategy could overcome such limitations through direct degradation of the target proteins. In this study, we successfully transformed the MET-targeted inhibitor crizotinib into a series of PROTACs, recruiting cereblon/cullin 4A E3 ubiquitin ligase to degrade the MET proteins. The optimized lead PROTAC (PRO-6 E) effectively eliminated MET proteins in vitro and in vivo, inhibiting proliferation and motility of MET-positive GC cells. In the MKN-45 xenograft model, PRO-6 E showed pronounced antitumor efficacy with a well-tolerated dosage regimen. These results validated PRO-6 E as the first oral PROTAC for MET-dependent GC.

IL-22 suppresses HSV-2 replication in human cervical epithelial cells.

Interleukin (IL)-22, a member of the IL-10 family, plays a role in antiviral immune responses to a number of viral infections. However, it is unclear whether IL-22 is involved in the mucosal immunity against herpes simplex virus 2 (HSV-2) infection in the female reproductive tract (FRT). In this study, we studied whether IL-22 could inhibit HSV-2 infection of human cervical epithelial cells (End1/E6E7 cells). We showed that End1/E6E7 cells express the functional IL-22 receptor complex (IL-22R1 and IL-10R2). When treated with IL-22, End1/E6E7 cells expressed the higher levels of IFN-stimulated genes (ISGs: ISG15, ISG56, OAS-1, OAS-2, and Mx2) than untreated cells. In addition, IL-22-treated cells produced higher levels of the tight junction proteins (ZO-1 and Occludin) than untreated cells. Mechanistically, IL-22 could activate the JAK/STAT signaling pathway by inducing the phosphorylation of STAT1 and STAT3. These observations indicate the potential of IL-22 as an anti-HSV-2 agent in the FRT mucosal innate immunity against HSV-2 infection.

PHD3 Stabilizes the Tight Junction Protein Occludin and Protects Intestinal Epithelial Barrier Function.

Prolyl hydroxylase domain proteins (PHDs) control cellular adaptation to hypoxia. PHDs are found involved in inflammatory bowel disease (IBD); however, the exact role of PHD3, a member of the PHD family, in IBD remains unknown. We show here that PHD3 plays a critical role in maintaining intestinal epithelial barrier function. We found that genetic ablation of Phd3 in intestinal epithelial cells led to spontaneous colitis in mice. Deletion of PHD3 decreases the level of tight junction protein occludin, leading to a failure of intestinal epithelial barrier function. Further studies indicate that PHD3 stabilizes occludin by preventing the interaction between the E3 ligase Itch and occludin, in a hydroxylase-independent manner. Examination of biopsy of human ulcerative colitis patients indicates that PHD3 is decreased with disease severity, indicating that PHD3 down-regulation is associated with progression of this disease. We show that PHD3 protects intestinal epithelial barrier function and reveal a hydroxylase-independent function of PHD3 in stabilizing occludin. These findings may help open avenues for developing a therapeutic strategy for IBD.

The Endoplasmic Reticulum Stress Sensor IRE1α in Intestinal Epithelial Cells Is Essential for Protecting against Colitis.

Intestinal epithelial cells (IECs) have critical roles in maintaining homeostasis of intestinal epithelium. Endoplasmic reticulum (ER) stress is implicated in intestinal epithelium homeostasis and inflammatory bowel disease; however, it remains elusive whether IRE1α, a major sensor of ER stress, is directly involved in these processes. We demonstrate here that genetic ablation of Ire1α in IECs leads to spontaneous colitis in mice. Deletion of IRE1α in IECs results in loss of goblet cells and failure of intestinal epithelial barrier function. IRE1α deficiency induces cell apoptosis through induction of CHOP, the pro-apoptotic protein, and sensitizes cells to lipopolysaccharide, an endotoxin from bacteria. IRE1α deficiency confers upon mice higher susceptibility to chemical-induced colitis. These results suggest that IRE1α functions to maintain the intestinal epithelial homeostasis and plays an important role in defending against inflammation bowel diseases.

Substrate Stiffness Combined with Hepatocyte Growth Factor Modulates Endothelial Cell Behavior.

Endothelial cells (ECs) play a crucial role in regulating various physiological and pathological processes. The behavior of ECs is modulated by physical (e.g., substrate stiffness) and biochemical cues (e.g., growth factors). However, the synergistic influence of these cues on EC behavior has rarely been investigated. In this study, we constructed poly(l-lysine)/hyaluronan (PLL/HA) multilayer films with different stiffness and exposed ECs to these substrates with and without hepatocyte growth factor (HGF)-supplemented culture medium. We demonstrated that EC adhesion, migration, and proliferation were positively correlated with substrate stiffness and that these behaviors were further promoted by HGF. Interestingly, ECs on the lower stiffness substrates showed stronger responses to HGF in terms of migration and proliferation, suggesting that HGF can profoundly influence stiffness-dependent EC behavior correlated with EC growth. After the formation of an EC monolayer, EC behaviors correlated with endothelial function were evaluated by characterizing monolayer integrity, nitric oxide production, and gene expression of endothelial nitric oxide synthase. For the first time, we demonstrated that endothelial function displayed a negative correlation with substrate stiffness. Although HGF improved endothelial function, HGF was not able to change the stiffness-dependent manner of endothelial functions. Taken together, this study provides insights into the synergetic influence of physical and biochemical cues on EC behavior and offers great potential in the development of optimized biomaterials for EC-based regenerative medicine.

The Effect of Silencing HIF-1α Gene in BxPC-3 Cell Line on Glycolysis-Related Gene Expression, Cell Growth, Invasion, and Apoptosis.

Hypoxia has been proved to be a typical character of solid tumors. Tumor cells prefer to use glucose through the glycolysis pathway instead of aerobic respiration. However, the precise molecular mechanism underlying this so-called Warburg effect remains elusive. In the current study, siRNA was synthesized and transfected into BxPC-3 cell line to silence the expression of HIF-1α gene. It was found that hypoxia induced hypoxia-inducible factor 1α (HIF-1α) overexpression in BxPC-3 cells, enhanced the expression of pyruvate dehydrogenase kinase 1 and lactate dehydrogenase A, thus facilitating glycolysis and making tumor cells more tolerant to hypoxic stress. The silencing of HIF-1α gene significantly attenuated glycolysis under hypoxic conditions, inhibited the growth and invasion ability of BxPC-3 cells, and enhanced hypoxia-induced cell apoptosis.

Endoscopic treatment of bleeding gastric ulcer causing gastric wall necrosis: A case report.

BACKGROUND: Gastric wall necrosis is a rare complication of endoscopic treatment for bleeding gastric ulcer, which may exacerbate the patient's condition once it occurs and may even require surgical intervention for treatment. CASE SUMMARY: A 59-year-old man was admitted to our department with melena. Endoscopy revealed a giant ulcer in the gastric antrum with a visible vessel in its center, which was treated with sclerosants and tissue glue injection and resulted in necrosis of the gastric wall. CONCLUSION: Injection of sclerosants and tissue glue may lead to gastric wall necrosis, which is a serious complication. Therefore, before administering this treatment to patients, we should consider other more effective methods of hemostasis to avoid gastric wall necrosis.

Comprehensive review of drug-mediated ICD inhibition of breast cancer: mechanism, status, and prospects.

The escalating incidence of breast cancer (BC) in women underscores its grave health threat. Current molecular insights into BC's post-adjuvant therapy cure remain elusive, necessitating active treatment explorations. Immunotherapy, notably chemotherapy-induced immunogenic cell death (ICD), has emerged as a promising BC therapy. ICD harnesses chemotherapeutics to activate anti-tumor immunity via DAMPs, fostering long-term T-cell memory and primary BC cure. Besides chemotherapy drugs, Nanodrugs, traditional Chinese medicine (TCM) and ICIs also induce ICD, boosting immune response. ICIs, like PD-1/PD-L1 inhibitors, revolutionize cancer treatment but face limited success in cold tumors. Thus, ICD induction combined with ICIs is studied extensively for BC immunotherapy. This article reviews the mechanism of ICD related drugs in BC and provides reference for the research and development of BC treatment, in order to explore more effective clinical treatment of BC, we hope to explore more ICD inducers and make ICIs more effective vaccines.

The perilous consequences of bowel preparation: a case study with literature review of Boerhaave syndrome.

Colonoscopy is widely acknowledged as a prevalent and efficacious approach for the diagnosis and treatment of gastrointestinal disorders. In order to guarantee an effective colonoscopy, it is imperative for patients to undergo an optimal bowel preparation regimen. This entails the consumption of a substantial volume of a non-absorbable solution to comprehensively purge the colon of any fecal residue. Nevertheless, it is noteworthy to acknowledge that the bowel preparation procedure may occasionally elicit adverse symptoms such as nausea and vomiting. In exceptional instances, the occurrence of excessive vomiting may lead to the rupture of the distal esophagus, a grave medical condition referred to as Boerhaave syndrome (BS). Timely identification and efficient intervention are imperative for the management of this infrequent yet potentially perilous ailment. This investigation presents a case study of a patient who developed BS subsequent to the ingestion of mannitol during bowel preparation. Furthermore, an exhaustive examination of extant case reports and pertinent literature on esophageal perforation linked to colonoscopy has been conducted. This analysis provides valuable insights into the prevention, reduction, and treatment of such serious complications.

Fabrication of a Coculture Organoid Model in the Biomimetic Matrix of Alginate to Investigate Breast Cancer Progression in a TAMs-Leading Immune Microenvironment.

The current research models of breast cancer are usually limited in their capacity to recapitulate the tumor microenvironment in vitro. The lack of an extracellular matrix (ECM) oversimplifies cell-cell or cell-ECM cross-talks. Moreover, the lack of tumor-associated macrophages (TAMs), that can comprise up to 50% of some solid neoplasms, poses a major problem for recognizing various hallmarks of cancer. To address these concerns, a type of direct breast cancer cells (BCCs)-TAMs coculture organoid model was well developed by a sequential culture method in this study. Alginate cryogels were fabricated with appropriate physical and mechanical properties to serve as an alternative ECM. Then, our previous experience was leveraged to polarize TAMs inside of the cryogels for creating an in vitro immune microenvironment. The direct coculture significantly enhanced BCCs organoid growth and cancer aggressive phenotypes, including the stemness, migration, ECM remodeling, and cytokine secretion. Furthermore, transcriptomic analysis and protein-protein interaction networks implied certain pathways (PI3K-Akt pathway, MAPK signaling pathway, etc.) and targets (TNF, PPARG, TLR2, etc.) during breast cancer progression in a TAM-leading immune microenvironment. Future studies to advance treatment strategies for BCC patients may benefit from using this facile model to reveal and target the interactions between cancer signaling and the immune microenvironment.

Cytosolic DNA sensors activation of human astrocytes inhibits herpes simplex virus through IRF1 induction.

Introduction: While astrocytes participate in the CNS innate immunity against herpes simplex virus type 1 (HSV-1) infection, they are the major target for the virus. Therefore, it is of importance to understand the interplay between the astrocyte-mediated immunity and HSV-1 infection. Methods: Both primary human astrocytes and the astrocyte line (U373) were used in this study. RT-qPCR and Western blot assay were used to measure IFNs, the antiviral IFN-stimulated genes (ISGs), IFN regulatory factors (IRFs) and HSV-1 DNA. IRF1 knockout or knockdown was performed with CRISPR/Cas9 and siRNA transfection techniques. Results: Poly(dA:dT) could inhibit HSV-1 replication and induce IFN-ß/IFN-λs production in human astrocytes. Poly(dA:dT) treatment of astrocytes also induced the expression of the antiviral ISGs (Viperin, ISG56 and MxA). Among IRFs members examined, poly(dA:dT) selectively unregulated IRF1 and IRF9, particularly IRF1 in human astrocytes. The inductive effects of poly(dA:dT) on IFNs and ISGs were diminished in the IRF1 knockout cells. In addition, IRF1 knockout attenuated poly(dA:dT)-mediated HSV-1 inhibition in the cells. Conclusion: The DNA sensors activation induces astrocyte intracellular innate immunity against HSV-1. Therefore, targeting the DNA sensors has potential for immune activation-based HSV-1 therapy.

Recent Developments in Layer-by-Layer Assembly for Drug Delivery and Tissue Engineering Applications.

Surfaces with biological functionalities are of great interest for biomaterials, tissue engineering, biophysics, and for controlling biological processes. The layer-by-layer (LbL) assembly is a highly versatile methodology introduced 30 years ago, which consists of assembling complementary polyelectrolytes or biomolecules in a stepwise manner to form thin self-assembled films. In view of its simplicity, compatibility with biological molecules, and adaptability to any kind of supporting material carrier, this technology has undergone major developments over the past decades. Specific applications have emerged in different biomedical fields owing to the possibility to load or immobilize biomolecules with preserved bioactivity, to use an extremely broad range of biomolecules and supporting carriers, and to modify the film's mechanical properties via crosslinking. In this review, the focus is on the recent developments regarding LbL films formed as 2D or 3D objects for applications in drug delivery and tissue engineering. Possible applications in the fields of vaccinology, 3D biomimetic tissue models, as well as bone and cardiovascular tissue engineering are highlighted. In addition, the most recent technological developments in the field of film construction, such as high-content liquid handling or machine learning, which are expected to open new perspectives in the future developments of LbL, are presented.

Biophysical cues of in vitro biomaterials-based artificial extracellular matrix guide cancer cell plasticity.

Clinical evidence supports a role for the extracellular matrix (ECM) in cancer plasticity across multiple tumor types. The lack of in vitro models that represent the native ECMs is a significant challenge for cancer research and drug discovery. Therefore, a major motivation for developing new tumor models is to create the artificial ECM in vitro. Engineered biomaterials can closely mimic the architectural and mechanical properties of ECM to investigate their specific effects on cancer progression, offering an alternative to animal models for the testing of cancer cell behaviors. In this review, we focused on the biomaterials from different sources applied in the fabrication of the artificial ECM and their biophysical cues to recapitulate key features of tumor niche. Furthermore, we summarized how the distinct biophysical cues guided cell behaviors of cancer plasticity, including morphology, epithelial-to-mesenchymal transition (EMT), enrichment of cancer stem cells (CSCs), proliferation, migration/invasion and drug resistance. We also discuss the future opportunities in using the artificial ECM for applications of tumorigenesis research and precision medicine, as well as provide useful messages of principles for designing suitable biomaterial scaffolds.

Herpes Simplex Virus 1-Induced Ferroptosis Contributes to Viral Encephalitis.

Herpes simplex virus 1 (HSV-1) is a DNA virus belonging to the family Herpesviridae. HSV-1 infection causes severe neurological disease in the central nervous system (CNS), including encephalitis. Ferroptosis is a nonapoptotic form of programmed cell death that contributes to different neurological inflammatory diseases. However, whether HSV-1 induces ferroptosis in the CNS and the role of ferroptosis in viral pathogenesis remain unclear. Here, we demonstrate that HSV-1 induces ferroptosis, as hallmarks of ferroptosis, including Fe2+ overload, reactive oxygen species (ROS) accumulation, glutathione (GSH) depletion, lipid peroxidation, and mitochondrion shrinkage, are observed in HSV-1-infected cultured human astrocytes, microglia cells, and murine brains. Moreover, HSV-1 infection enhances the E3 ubiquitin ligase Keap1 (Kelch-like ECH-related protein 1)-mediated ubiquitination and degradation of nuclear factor E2-related factor 2 (Nrf2), a transcription factor that regulates the expression of antioxidative genes, thereby disturbing cellular redox homeostasis and promoting ferroptosis. Furthermore, HSV-1-induced ferroptosis is tightly associated with the process of viral encephalitis in a mouse model, and the ferroptosis-activated upregulation of prostaglandin-endoperoxide synthase 2 (PTGS2) and prostaglandin E2 (PGE2) plays an important role in HSV-1-caused inflammation and encephalitis. Importantly, the inhibition of ferroptosis by a ferroptosis inhibitor or a proteasome inhibitor to suppress Nrf2 degradation effectively alleviated HSV-1 encephalitis. Together, our findings demonstrate the interaction between HSV-1 infection and ferroptosis and provide novel insights into the pathogenesis of HSV-1 encephalitis. IMPORTANCE Ferroptosis is a nonapoptotic form of programmed cell death that contributes to different neurological inflammatory diseases. However, whether HSV-1 induces ferroptosis in the CNS and the role of ferroptosis in viral pathogenesis remain unclear. In the current study, we demonstrate that HSV-1 infection induces ferroptosis, as Fe2+ overload, ROS accumulation, GSH depletion, lipid peroxidation, and mitochondrion shrinkage, all of which are hallmarks of ferroptosis, are observed in human cultured astrocytes, microglia cells, and murine brains infected with HSV-1. Moreover, HSV-1 infection enhances Keap1-dependent Nrf2 ubiquitination and degradation, which results in substantial reductions in the expression levels of antiferroptotic genes downstream of Nrf2, thereby disturbing cellular redox homeostasis and promoting ferroptosis. Furthermore, HSV-1-induced ferroptosis is tightly associated with the process of viral encephalitis in a mouse model, and the ferroptosis-activated upregulation of PTGS2 and PGE2 plays an important role in HSV-1-caused inflammation and encephalitis. Importantly, the inhibition of ferroptosis by either a ferroptosis inhibitor or a proteasome inhibitor to suppress HSV-1-induced Nrf2 degradation effectively alleviates HSV-1-caused neuro-damage and inflammation in infected mice. Overall, our findings uncover the interaction between HSV-1 infection and ferroptosis, shed novel light on the physiological impacts of ferroptosis on the pathogenesis of HSV-1 infection and encephalitis, and provide a promising therapeutic strategy to treat this important infectious disease with a worldwide distribution.

Therapeutic delivery of siRNA silencing HIF-1 alpha with micellar nanoparticles inhibits hypoxic tumor growth.

The particular characteristics of the tumor microenvironment have the potential to strongly promote tumor growth, metastasis and angiogenesis and induce drug resistance. Therefore, the development of effective, systemic therapeutic approaches specifically based on the tumor microenvironment is highly desirable. Hypoxia-inducible factor-1α (HIF-1α) is an attractive therapeutic target because it is a key transcription factor in tumor development and only accumulates in hypoxic tumors. We report here that a cationic mixed micellar nanoparticle (MNP) consisting of amphiphilic block copolymers poly(ε-caprolactone)-block-poly(2-aminoethylethylene phosphate) (PCL(29)-b-PPEEA(21)) and poly(ε-caprolactone)-block-poly(ethylene glycol) (PCL(40)-b-PEG(45)) was a suitable carrier for HIF-1α siRNA to treat hypoxic tumors, which showed an average diameter of 58.0 ± 3.4 nm. The complex MNP(siRNA), formed by the interaction of MNP and siRNA, was transfected into PC3 prostate cancer cells efficiently, while the inhibition of HIF-1α expression by MNP loaded with HIF-1α siRNA (MNP(siHIF)) blocked PC3 cell proliferation, suppressed cell migration and disturbed angiogenesis under in vitro hypoxic mimicking conditions. It was further demonstrated that systemic delivery of MNP(siHIF) effectively inhibited tumor growth in a PC3 prostate cancer xenograft murine model without activating innate immune responses. Moreover, delivery of MNP(siHIF) sensitized PC3 tumor cells to doxorubicin chemotherapy in vitro and in vivo by downregulating MDR1 gene expression which was induced by hypoxia. The underlying concept of use of MNP(siHIF) to block HIF-1α holds promise as an example of a clinical approach using specific siRNA therapy for cancer treatment aimed at the hypoxic tumor microenvironment.

Fluoxetine promotes remission in acute experimental autoimmune encephalomyelitis in rats.

OBJECTIVE: This study was carried out to clarify the effects of the antidepressant fluoxetine, a selective serotonin reuptake inhibitor, for its potential use in autoimmune diseases like multiple sclerosis in a rat model of experimental autoimmune encephalomyelitis (EAE). METHODS: The rat EAE model was induced by subcutaneous injection of guinea pig spinal cord homogenate. Rats received fluoxetine via daily intragastric administration, starting 2 weeks prior to immune induction (fluoxetine pretreatment). Clinical scores and pathological changes in EAE rats were analyzed. Changes in serum cytokine levels were assessed by ELISA. RESULTS: Fluoxetine pretreatment significantly promoted remission in EAE. Histologically, fluoxetine-induced neuroprotection was accompanied by reductions in inflammatory foci and in the degree of demyelination in the spinal cord of EAE rats. The increase in serum IFN-γ in the EAE model was also suppressed by fluoxetine administration. CONCLUSIONS: These findings suggest that the prophylactic use of fluoxetine can relieve symptoms during remission in the acute EAE model, and these neuroprotective effects are associated with its anti-inflammatory effects.

A 3D-printed scaffold-based osteosarcoma model allows to investigate tumor phenotypes and pathogenesis in an in vitro bone-mimicking niche.

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Physical confinement in alginate cryogels determines macrophage polarization to a M2 phenotype by regulating a STAT-related mRNA transcription pathway.

The immunologic response is considered to play a pivotal role in the application of biomaterial implants, and intrinsic properties of biomaterials can significantly modulate the anti-inflammatory effects. However, how physical confinement influences M2 polarization of macrophages and the relevant mechanisms have not been clearly elucidated. In this study, pore size and porosity in cryogels can be mediated by utilizing alginates with different viscosities. Cryogels of small pore size and low porosity can restrict M2 polarization of macrophages in vitro, judging from cell morphology, secretion of cytokines and expression of key M2-related genes. In comparison, cryogels of large pore size or high porosity can induce M2 polarization in vivo, resulting in the anti-inflammation effects. High-throughput RNA-seq analysis demonstrates that the mRNA surveillance pathway is key in the polarization process, and four primary transcription factors (PPAR-γ, STAT6, NF-κB, and STAT1) participate probably by competition in DNA binding to regulate M2-related gene expression. This study confirms that enough physical space inside is necessary to promote M2 polarization for the anti-inflammatory performance, which can be applied widely in the fields of tissue engineering and regenerative medicine.

Targeted delivery of antisense inhibitor of miRNA for antiangiogenesis therapy using cRGD-functionalized nanoparticles.

MiRNAs are viable therapeutic targets for cancer therapy, but the targeted delivery of miRNA or its anti-miRNA antisense oligonucleotides (AMOs) remains a challenge. We report here a PEGylated LPH (liposome-polycation-hyaluronic acid) nanoparticle formulation modified with cyclic RGD peptide (cRGD) for specific and efficient delivery of AMO into endothelial cells, targeting α(v)ß3 integrin present on the tumor neovasculature. The nanoparticles effectively delivered anti-miR-296 AMO to the cytoplasm and downregulated the target miRNA in human umbilical vein endothelial cells (HUVECs), which further efficiently suppressed blood tube formulation and endothelial cell migration, owing to significant upregulation of hepatocyte growth factor-regulated tyrosine kinase substrate (HGS), whereas nanoparticles without cRGD modification showed only little AMO uptake and miRNA silencing activity. In vivo assessment of angiogenesis using Matrigel plug assay also demonstrated that cRGD modified LPH nanoparticles have potential for antiangiogenesis in miRNA therapeutics. With the delivery of anti-miR-296 AMO by targeted nanoparticles, significant decrease in microvessel formulation within Matrigel was achieved through suppressing the invasion of CD31-positive cells into Matrigel and prompting HGS expression in angiogenic endothelial cells.

Multiple functional hyperbranched poly(amido amine) nanoparticles: synthesis and application in cell imaging.

The hyperbranched poly(amido amine) nanoparticles (HPAMAM NPs) with multiple functions, such as biodegradability, autofluorescence, and specific affinity, were successfully prepared by Michael addition dispersion polymerization of CBA, AEPZ, and N-galactosamine hydrochloride (or N-glucosamine hydrochloride) in a mixture of methanol/water. The resultant NPs displayed strong photoluminescence, high photostability, broad absorption, and emission (from 430 to 620 nm) spectra. The fluorescence from HPAMAM NPs may be attributed to the tertiary amine chromophore. The incubation results of the liver cancer cells, HepG2, with the NPs showed that the NPs are nontoxic and can be recognized by asialoglycoprotein receptors on the surface of HepG2 and then can be internalized. Therefore, they have potential applications in bioimaging and drug or gene delivery.