Chorioallantoic Membrane Model Research Articles

Genistein–Aspirin Combination Exerts Cytotoxic and Anti-Migratory Effects in Human Colorectal Cancer Cells

Colorectal cancer (CRC) is a heterogenous pathology with high incidence and mortality rates globally, but it is also preventable so finding the most promising candidates (natural compounds or repurposed drugs) to be chemopreventive alternatives has become a topic of interest in recent years. The present work aims to elucidate the potential effects of a combination between genistein (GEN), an isoflavone of natural origin, and aspirin (ASA) in CRC prevention/treatment by performing an in vitro evaluation in human colorectal cancer cells (HCT-116) and an in ovo analysis using the chick embryo chorioallantoic membrane (CAM) model. Cell viability was verified by an MTT (migratory potential by scratch) assay, and the expressions of MMP-2 and MMP-9 were analyzed using RT-qPCR. Our results indicated a dose-dependent cytotoxic effect of ASA (2.5 mM) + GEN (10–75 µM) combination characterized by reduced cell viability and morphological changes (actin skeleton reorganization and nuclei deterioration), an inhibition of HCT-116 cells’ migratory potential by down-regulating MMP-2 and MMP-9 mRNA expressions, and an antiangiogenic effect by modifying the vascular network. These promising results raise the possibility of future in-depth investigations regarding the chemopreventive/therapeutical potential of ASA+GEN combination.

Ultra high frequency ultrasound enables real-time visualization of blood supply from chorioallantoic membrane to human autosomal dominant polycystic kidney tissue

Ultra high frequency (UHF) ultrasound enables the visualization of very small structures that cannot be detected by conventional ultrasound. The utilization of UHF imaging as a new imaging technique for the 3D-in-vivo chorioallantoic membrane (CAM) model can facilitate new insights into tissue perfusion and survival. Therefore, human renal cystic tissue was grafted onto the CAM and examined using UHF ultrasound imaging. Due to the unprecedented resolution of UHF ultrasound, it was possible to visualize microvessels, their development, and the formation of anastomoses. This enabled the observation of anastomoses between human and chicken vessels only 12 h after transplantation. These observations were validated by 3D reconstructions from a light sheet microscopy image stack, indocyanine green angiography, and histological analysis. Contrary to the assumption that the nutrient supply of the human cystic tissue and the gas exchange happens through diffusion from CAM vessels, this study shows that the vasculature of the human cystic tissue is directly connected to the blood vessels of the CAM and perfusion is established within a short period. Therefore, this in-vivo model combined with UHF imaging appears to be the ideal platform for studying the effects of intravenously applied therapeutics to inhibit renal cyst growth.

Denosumab Induces Neoplastic Stromal Cell Apoptosis via p62 Downregulation Dependent on Autophagy Pathway in Giant Cell Tumour of Bone.

As the only humanized monoclonal antibody against receptor activator of nuclear factor-κB ligand (RANKL) for giant cell tumour of bone (GCTB) therapy, denosumab has limited antitumour effect on neoplastic stromal cells. Nevertheless, its mechanism of action has not yet been clarified. A previous study has revealed that p62 may play an important role in the antitumour activity of denosumab. The study aimed to investigate if the mechanism by which denosumab inhibits GCTB neoplastic stromal cells growth is via p62 modulation and other related mechanisms. p62 expression before and after denosumab therapy was analysed by RT‒qPCR, western blot, ELISA, and immunohistochemical assays. Two primary neoplastic stromal cells were isolated from fresh GCTB tumour tissue (L cell) and metastatic tissue (M cell). Cell proliferation, migration, apoptosis, and autophagy were investigated in p62 knockdown neoplastic stromal cells transfected by short hairpin RNA lentivirus in vitro. Tumor growth was evaluated in the chick chorioallantoic membrane model in vivo. p62 expression was found to be downregulated following denosumab therapy. The patients with a decrease in p62 expression had lower recurrence-free survival rates. The proliferation of M cells was not inhibited by denosumab therapy, but it was restored by p62 knockdown. Moreover, p62 knockdown inhibited tumour growth in vivo. Denosumab induced M cell apoptosis and arrested the cell cycle at the G1/G0 transition and these effects were also enhanced by p62 knockdown. Autophagic flux assays revealed p62 modulation to be dependent on autophagy following denosumab incubation. Denosumab induced neoplastic stromal cells apoptosis via p62 downregulation dependent on autophagy pathway. The combination of p62 and RANKL knockdown might be a better strategy than RANKL knockdown alone for GCTB targeted therapy.

Sevofluran Maruziyetinin Civciv Embriyo Gelişimi Üzerindeki Ex-Ovo Değerlendirilmesi: Anjiyogenez Etkilerinin Araştırılması

Objective: Angiogenesis, defined as the formation of new blood vessels from pre-existing ones, plays a vital role in both physiological and pathological conditions. Understanding agents that can influence angiogenesis is crucial in managing diseases where angiogenesis is dysregulated. This study focuses on sevoflurane, a commonly used inhalation anesthetic, whose effects on angiogenesis and embryonic development are not well understood. Our objective was to assess the impact of sevoflurane exposure on angiogenesis using the ex-ovo chick chorioallantoic membrane model. Methods: In this model, fertilized chicken eggs were exposed to sevoflurane at concentrations of 2% and 4%, for varying durations of 1, 2, and 4 hours. The embryos were then divided into control and experimental groups for quantitative angiogenesis assessment using Image J software, followed by statistical analysis with one-way analysis of variance. Results: The results indicate that sevoflurane exposure has a dose-dependent positive effect on angiogenesis, with significant increases in vascular density observed in embryos exposed to both concentrations compared to the control group. Additionally, the length of exposure was found to further enhance these angiogenic effects. Conclusion: Despite the dose and duration-dependent impact of sevoflurane on angiogenesis, the existing literature presents mixed findings, highlighting the need for additional research to elucidate sevoflurane’s role in angiogenesis. This is particularly important for understanding its implications in various medical conditions, such as cancer, wound healing, and fetal development. Future investigations into sevoflurane’s effects on placental angiogenesis could also provide valuable insights into its potential consequences on intrauterine growth. Keywords: Angiogenesis, sevoflurane, chicken embryo, model organisms, ex-ovo

Glycosaminoglycans Modulate the Angiogenic Ability of Type I Collagen-Based Scaffolds by Acting on Vascular Network Remodeling and Maturation

Type I collagen, prevalent in the extracellular matrix, is biocompatible and crucial for tissue engineering and wound healing, including angiogenesis and vascular maturation/stabilization as required processes of newly formed tissue constructs or regeneration. Sometimes, improper vascularization causes unexpected outcomes. Vascularization failure may be caused by extracellular matrix collagen and non-collagen components heterogeneously. This study compares the angiogenic potential of collagen type I-based scaffolds and collagen type I/glycosaminoglycans scaffolds by using the chick embryo chorioallantoic membrane (CAM) model and IKOSA digital image analysis. Two clinically used biomaterials, Xenoderm (containing type I collagen derived from decellularized porcine extracellular matrix) and a dual-layer collagen sponge (DLC, with a biphasic composition of type I collagen combined with glycosaminoglycans) were tested for their ability to induce new vascular network formation. The AI-based IKOSA app enhanced the research by calculating from stereomicroscopic images angiogenic parameters such as total vascular area, branching sites, vessel length, and vascular thickness. The study confirmed that Xenoderm caused a fast angiogenic response and substantial vascular growth, but was unable to mature the vascular network. DLC scaffold, in turn, produced a slower angiogenic response, but a more steady and organic vascular maturation and stabilization. This research can improve collagen-based knowledge by better assessing angiogenesis processes. DLC may be preferable to Xenoderm or other materials for functional neovascularization, according to the findings.

Enhancing bone tissue engineering using iron nanoparticles and magnetic fields: A focus on cytomechanics and angiogenesis in the chicken egg chorioallantoic membrane model.

To evaluate the potential of iron nanoparticles (FeNPs) in conjunction with magnetic fields (MFs) to enhance osteoblast cytomechanics, promote cell homing, bone development activity, and antibacterial capabilities, and to assess their in vivo angiogenic viability using the chicken egg chorioallantoic membrane (CAM) model. Experimental study conducted in a laboratory setting to investigate the effects of FeNPs and MFs on osteoblast cells and angiogenesis using a custom titanium (Ti) substrate coated with FeNPs. A custom titanium (Ti) was coated with FeNPs. Evaluations were conducted to analyze the antibacterial properties, cell adhesion, durability, physical characteristics, and nanoparticle absorption associated with FeNPs. Cell physical characteristics were assessed using protein markers, and microscopy, CAM model, was used to quantify blood vessel formation and morphology to assess the FeNP-coated Ti's angiogenic potential. This in vivo study provided critical insights into tissue response and regenerative properties for biomedical applications. Statistical analysis was performed using appropriate tests to compare experimental groups and controls. Significance was determined at P < 0.05. FeNPs and MFs notably improved osteoblast cell mechanical properties facilitated the growth and formation of new blood vessels and bone tissue and promoted cell migration to targeted sites. In the group treated with FeNPs and exposed to MFs, there was a significant increase in vessel percentage area (76.03%) compared to control groups (58.11%), along with enhanced mineralization and robust antibacterial effects (P < 0.05). The study highlights the promising potential of FeNPs in fostering the growth of new blood vessels, promoting the formation of bone tissue, and facilitating targeted cell migration. These findings underscore the importance of further investigating the mechanical traits of FeNPs, as they could significantly advance the development of effective bone tissue engineering techniques, ultimately enhancing clinical outcomes in the field.

Abstract 2738: XON7, a new Glyco-Humanized Polyclonal Antibody, first in class immunotherapy against several solid tumors

Abstract Introduction: Cancer immunotherapy has recently generated much excitement after the success of the immunomodulating anti-CTLA-4 and anti-PD-1 antibodies against various types of cancers. However, for many cancers, there is still a lack of effective treatment that can result in long-term cancer-free survival and lower metastatic and relapse risks. The emergence of cancer resistance could be minimized by drug combination or by multi-targeting of tumor cells. Polyclonal antibodies can target several tumor-associated antigens simultaneously and would be more efficient than a conventional mAbs. Here we evaluate the safety and efficacy of XON7, a first in class glyco-humanized polyclonal antibody (GH-pAb), in cancer preclinical models. Material and Methods: XON7 ability to induce Complement Dependent Cytotoxicity (CDC) and apoptosis was tested in a panel of cancer cell lines and PBMC. XON7 was also evaluated in sphere formation assay to predict its effects on cancer resistant. Specific binding to human tumors and healthy tissues was assessed by immunochemistry on tissue micro-array. Cross-cancer activity was evaluated on biopsies from patients with solid tumor such as CRC, NSCLC, osteosarcoma, and breast cancers. In vivo XON7 efficacy was evaluated in NMRI nude mice using A549: NSCLC, HCT-116: colon cancer, LNCaP: prostate cancer and MDA-MB-231: breast cancer cells. A human NSCLC chorio-allantoic membrane (CAM) model was used to evaluate the efficacy of XON7 + anti PD-1 on tumor growth and metastasis. Pharmacokinetics and safety of this drug were assessed in marmoset after single and repeated IV dosing up to 60mg/kg. Results: XON7 showed a potent in vitro antitumor activity in a panel of cancer cell lines, it induced specific tumor cell CDC (EC50=50ug/mL) and apoptosis (IC50= 100ug/mL). it was able to kill up to 100% of the cancer cells without affecting PBMC. In addition, it induced a striking decrease of tumor sphere formation in HCT116, A549 and MDA-MB-231 cancer cell lines. XON7 showed preferential recognition of tumor cells as compared to normal cells, it demonstrated cross-reaction to tumor patient biopsy without staining of the healthy tissues. In vitro XON7 potency was translated into in vivo efficacy in different mice xenograft models. XON7 induced a significant reduction of tumor growth ranging from 40 to 90% across several tested tumors. Furthermore, it showed, significant increase of anti-tumor response rate and decrease of metastasis when it is associated with anti PD-1 in vivo CAM model (&amp;gt;90%, p=0.001). XON7 was also characterized by high tolerance and satisfactory exposure in marmoset. Conclusion: Based on its safe toxicity profile and potent activity, XON7 appeared as a novel and promising cancer immunotherapy to fight against recurrent solid tumor, it is now aimed to reach clinical development, FIH in patients is planned to start before the end of 2023. Citation Format: Carine Ciron, Gwenaelle Evanno, Pierre Morice, Pierre-Joseph Royer, Françoise Shneiker, Odile Duvaux, Bernard Vanhove, Firas Bassissi. XON7, a new Glyco-Humanized Polyclonal Antibody, first in class immunotherapy against several solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2738.

Abstract 1274: Investigating PLOD2 as a therapeutic target to overcome metastasis in radiorecurrent prostate cancer

Abstract Prostate cancer (PCa) is the second most common cancer in males, with 1 in 8 men developing the disease in their lifetime. For PCa, a common treatment strategy is external beam radiation therapy to the prostate. However, when cancer recurs (radiorecurrent PCa), it often behaves aggressively by invading into surrounding organs or spreading distantly. Radiorecurrent PCa metastasis is therefore a significant cause of morbidity and mortality that must be overcome to improve survival of advanced PCa patients. Proteomic analysis revealed the procollagen enzyme Lysyl Hydroxylase 2 (PLOD2) to be upregulated in our radiorecurrent, and highly aggressive, conventionally fractionated DU145 (DU145-CF) PCa cell line. Given its established function as a mediator of invasion in various other cancers, we sought to characterize the role of PLOD2 in the aggressive phenotype of our radiorecurrent PCa cells. Bioinformatic analysis of clinical data revealed PLOD2 genomic amplification to be significantly associated with biochemical recurrence in PCa patients. Upon further examination in vitro, it was revealed that PLOD2 knockdown significantly reduces matrigel invasion and migration in our radiorecurrent DU145-CF cell line, in addition to numerous other PCa cell lines, including primary cells we derived directly from PCa patients. Using the in vivo chick Chorioallantoic Membrane (CAM) model, we confirmed that PLOD2 knockdown significantly reduces the ability of DU145-CF cells to extravasate from the CAM vasculature into the surrounding stroma, a critical step of the metastatic cascade. To explore mechanisms of metastatic cellular reprogramming downstream of PLOD2, RNA sequencing of DU145-CF cells was conducted; a total of 681 genes were discovered to be dysregulated by PLOD2 knockdown, with their functional analysis suggesting changes in cellular metabolism and respiration. Finally, since PLOD2 is known to be regulated by hypoxia-induced protein HIF1α, we explored whether PLOD2 expression could be inhibited by the HIF1α inhibitor, PX-478. Treatment with PX-478 reduced both HIF1α and PLOD2 protein expression, and significantly reduced invasion, migration, and in vivo extravasation in DU145-CF cells, thereby indicating its potential as a pharmacological inhibitor of HIF1α-associated PLOD2 in radiorecurrent PCa. Together, our results demonstrate for the first time the role of PLOD2 in radiorecurrent PCa invasiveness, and point towards its potential as a therapeutic target to reduce metastasis and improve survival outcomes in PCa patients. Citation Format: Gavin Frame, Hon Leong, Roni Haas, Xiaoyong Huang, Jessica Wright, Paul C. Boutros, Thomas Kislinger, Stanley K. Liu. Investigating PLOD2 as a therapeutic target to overcome metastasis in radiorecurrent prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1274.

In vitro and in ovo photodynamic efficacy of nebulized curcumin-loaded tetraether lipid liposomes prepared by DC as stable drug delivery system

Lung cancer is one of the most common causes of high mortality worldwide. Current treatment strategies, e.g., surgery, radiotherapy, chemotherapy, and immunotherapy, insufficiently affect the overall outcome. In this study, we used curcumin as a natural photosensitizer in photodynamic therapy and encapsulated it in liposomes consisting of stabilizing tetraether lipids aiming for a pulmonary drug delivery system against lung cancer. The liposomes with either hydrolyzed glycerol-dialkyl-glycerol tetraether (hGDGT) in different ratios or hydrolyzed glycerol-dialkyl-nonitol tetraether (hGDNT) were prepared by dual centrifugation (DC), an innovative method for liposome preparation. The liposomes’ physicochemical characteristics before and after nebulization and other nebulization characteristics confirmed their suitability. Morphological characterization using atomic force and transmission electron microscopy showed proper vesicular structures indicative of liposomes. Qualitative and quantitative uptake of the curcumin-loaded liposomes in lung adenocarcinoma (A549) cells was visualized and proven. Phototoxic effects of the liposomes were detected on A549 cells, showing decreased cell viability. The generation of reactive oxygen species required for PDT and disruption of mitochondrial membrane potential were confirmed. Moreover, the chorioallantoic membrane (CAM) model was used to further evaluate biocompatibility and photodynamic efficacy in a 3D cell culture context. Photodynamic efficacy was assessed by PET/CT after nebulization of the liposomes onto the xenografted tumors on the CAM with subsequent irradiation. The physicochemical properties and the efficacy of tetraether lipid liposomes encapsulating curcumin, especially liposomes containing hGDNT, in 2D and 3D cell cultures seem promising for future PDT usage against lung cancer.

Hydrogels of Alginate Derivative‐Encased Nanodots Featuring Carbon‐Coated Manganese Ferrite Cores with Gold Shells to Offer Antiangiogenesis with Multimodal Imaging‐Based Theranostics

AbstractThe investigation seeks to develop gold‐shelled carbon‐coated manganese ferrite nanodots enclosed within oxidized alginate polymeric hydrogels to improve encapsulation efficiency, biocompatibility, and multimodal imaging capabilities. These engineered particles, MNF@C‐Au@OSA, are crafted through a hydrothermal synthesis, succeeded by the application of gold nanoparticle shelling and subsequent encapsulation within hydrogels derived from synthesized alginate derivatives, enhancing their overall appeal. Diverse characterization methodologies are employed in order to validate the multiple steps of the synthesis process. The resulting engineered nanoparticles exhibit dual‐mode (T1 and T2) MRI contrast capabilities and compatibility with CT and fluorescence‐based imaging. To illustrate minimal toxicity, A375 cell lines are utilized for in vitro testing, whereas zebrafish embryos are employed for in vivo assessments. A Chorioallantoic Membrane Model (CAM) is used to establish the antiangiogenic characteristics of MNF@C‐Au@OSA, which is confirmed by histopathological observations. Taking these studies together, one can conclude that the engineered MNF@C‐Au@OSA possesses multimodal imaging capabilities and has demonstrated antiangiogenic properties, thereby establishing them as potential theranostic agents.This article is protected by copyright. All rights reserved

Analysis of Osteosarcoma Cell Lines and Patient Tissue Using a 3D In Vivo Tumor Model—Possible Effects of Punicalagin

Osteosarcomas are the most common primary malignant bone tumors and mostly affect children, adolescents, and young adults. Despite current treatment options such as surgery and polychemotherapy, the survival of patients with metastatic disease remains poor. In recent studies, punicalagin has reduced the cell viability, angiogenesis, and invasion in cell culture trials. The aim of this study was to examine the effects of punicalagin on osteosarcomas in a 3D in vivo tumor model. Human osteosarcoma biopsies and SaOs-2 and MG-63 cells, were grown in a 3D in vivo chorioallantoic membrane (CAM) model. After a cultivation period of up to 72 h, the tumors received daily treatment with punicalagin for 4 days. Weight measurements of the CAM tumors were performed, and laser speckle contrast imaging (LSCI) and a deep learning-based image analysis software (CAM Assay Application v.3.1.0) were used to measure angiogenesis. HE, Ki-67, and Caspase-3 staining was performed after explantation. The osteosarcoma cell lines SaOs-2 and MG-63 and osteosarcoma patient tissue displayed satisfactory growth patterns on the CAM. Treatment with punicalagin decreased tumor weight, proliferation, and tumor-induced angiogenesis, and the tumor tissue showed pro-apoptotic characteristics. These results provide a robust foundation for the implementation of further studies and show that punicalagin offers a promising supplementary treatment option for osteosarcoma patients. The 3D in vivo tumor model represents a beneficial model for the testing of anti-cancer therapies.

Inhibition of PRMT1 Suppresses the Growth of U87MG-Derived Glioblastoma Stem Cells by Blocking the STAT3 Signaling Pathway.

Glioblastoma stem cells (GSCs) play a pivotal role in the initiation, progression, resistance to treatment, and relapse of glioblastoma multiforme (GBM). Thus, identifying potential therapeutic targets and drugs that interfere with the growth of GSCs may contribute to improved treatment outcomes for GBM. In this study, we first demonstrated the functional role of protein arginine methyltransferase 1 (PRMT1) in GSC growth. Furamidine, a PRMT1 inhibitor, effectively inhibited the proliferation and tumorsphere formation of U87MG-derived GSCs by inducing cell cycle arrest at the G0/G1 phase and promoting the intrinsic apoptotic pathway. Moreover, furamidine potently suppressed the in vivo tumor growth of U87MG GSCs in a chick embryo chorioallantoic membrane model. In particular, the inhibitory effect of furamidine on U87MG GSC growth was associated with the downregulation of signal transducer and activator of transcription 3 (STAT3) and key GSC markers, including CD133, Sox2, Oct4, Nanog, aldehyde dehydrogenase 1, and integrin α6. Our results also showed that the knockdown of PRMT1 by small interfering RNA significantly inhibited the proliferation of U87MG GSCs in vitro and in vivo through a molecular mechanism similar to furamidine. In addition, combined treatment with furamidine and berbamine, a calcium/calmodulin-dependent protein kinase II gamma (CaMKIIγ) inhibitor, inhibited the growth of U87MG GSCs more strongly than single-compound treatment. The increased antiproliferative effect of combining the two compounds resulted from a stronger downregulation of STAT3-mediated downstream GBM stemness regulators through dual PRMT1 and CaMKIIγ function blockade. In conclusion, these findings suggest that PRMT1 and its inhibitor, furamidine, are potential novel therapeutic targets and drug candidates for effectively suppressing GSC growth.

Targeting HER2-positive breast cancer cells by a combination of dasatinib and BMS-202: Insight into the molecular pathways

BackgroundRecent investigations have reported the benefits of using a tyrosine kinase inhibitor, dasatinib (DA), as well as programmed death-ligand 1 (PD-L1) inhibitors in the management of several solid tumors, including breast cancer. Nevertheless, the outcome of the combination of these inhibitors on HER2-positive breast cancer is not explored yet.MethodsHerein, we investigated the impact of DA and PD-L1 inhibitor (BMS-202) combination on HER2-positive breast cancer cell lines, SKBR3 and ZR75.ResultsOur data reveal that the combination significantly inhibits cell viability of both cancer cell lines as compared to monotreatment. Moreover, the combination inhibits epithelial-mesenchymal transition (EMT) progression and reduces cancer cell invasion by restoring E-cadherin and β-catenin expressions and loss of vimentin, major biomarkers of EMT. Additionally, the combination reduces the colony formation of both cell lines in comparison with their matched control. Also, the combination considerably inhibits the angiogenesis of the chorioallantoic membrane model compared with monotreatment. Molecular pathway analysis of treated cells shows that this combination blocks HER2, AKT, β-catenin, and JNK1/2/3 activities.ConclusionOur findings implicate that a combination of DA and BMS-202 could have a significant impact on the management of HER2-positive breast cancer.

Biofabrication of nanocomposite-based scaffolds containing human bone extracellular matrix for the differentiation of skeletal stem and progenitor cells

Autograft or metal implants are routinely used in skeletal repair. However, they fail to provide long-term clinical resolution, necessitating a functional biomimetic tissue engineering alternative. The use of native human bone tissue for synthesizing a biomimetic material ink for three-dimensional (3D) bioprinting of skeletal tissue is an attractive strategy for tissue regeneration. Thus, human bone extracellular matrix (bone-ECM) offers an exciting potential for the development of an appropriate microenvironment for human bone marrow stromal cells (HBMSCs) to proliferate and differentiate along the osteogenic lineage. In this study, we engineered a novel material ink (LAB) by blending human bone-ECM (B) with nanoclay (L, Laponite®) and alginate (A) polymers using extrusion-based deposition. The inclusion of the nanofiller and polymeric material increased the rheology, printability, and drug retention properties and, critically, the preservation of HBMSCs viability upon printing. The composite of human bone-ECM-based 3D constructs containing vascular endothelial growth factor (VEGF) enhanced vascularization after implantation in an ex vivo chick chorioallantoic membrane (CAM) model. The inclusion of bone morphogenetic protein-2 (BMP-2) with the HBMSCs further enhanced vascularization and mineralization after only seven days. This study demonstrates the synergistic combination of nanoclay with biomimetic materials (alginate and bone-ECM) to support the formation of osteogenic tissue both in vitro and ex vivo and offers a promising novel 3D bioprinting approach to personalized skeletal tissue repair.Graphic abstract

Antiangiogenic Effects of Infliximab in Chick Chorioallantoic Membrane Model

Introduction: TNF-α plays a key role in initiating pathologic angiogenesis, however, literature on the effects of infliximab on angiogenesis is limited.&#x0D; Objective: The study aimed to investigate the effects of infliximab on angiogenesis in an in vivo chick chorioallantoic membrane (CAM) model.&#x0D; Methods: The study was approved by the local ethics committee on animal experimentation. Thirty fertilized specific pathogen-free eggs were incubated and kept under appropriate temperature and humidity control. On the third day of the incubation, infliximab (1 µmol) dissolved in phosphate-buffered saline in the first group, phosphate-buffered saline (negative control) (0.1 ml) in the second group, anti-VEGF (positive control) (1 µmol) in the third group were administered by injection. On the eighth day of incubation, the vascular structures of the CAMs were macroscopically examined, and standard digital photographs were taken. The digital images were analyzed, and data including mean vessel density, thickness, and number were compared between groups. P &lt; 0.05 was considered statistically significant.&#x0D; Results: Angiogenesis was significantly reduced in the anti-VEGF and infliximab groups compared to the saline-only group. Vessel thickness, vessel number, and vessel density were significantly less in the infliximab and anti-VEGF groups compared to the saline-only group (p=0.034, 0.029, 0.024, respectively).&#x0D; Conclusions: Infliximab showed promising antiangiogenic effects in the chick CAM model. Thus, infliximab could be a treatment agent in pathological processes in which angiogenesis is responsible. The antiangiogenic effect of infliximab could be due to the inhibition of various angiogenesis-related cytokines and adhesion molecules. Further studies are needed to clarify the exact mechanisms.

A bioconvergence study on platinum-free concurrent chemoradiotherapy for the treatment of HPV-negative head and neck carcinoma

Locally advanced head and neck squamous cell carcinoma (LA-HNSCC) is characterized by high rate of recurrence, resulting in a poor survival. Standard treatments are associated with significant toxicities that impact the patient’s quality of life, highlighting the urgent need for novel therapies to improve patient outcomes. On this regard, noble metal nanoparticles (NPs) are emerging as promising agents as both drug carriers and radiosensitizers. On the other hand, co-treatments based on NPs are still at the preclinical stage because of the associated metal-persistence. In this bioconvergence study, we introduce a novel strategy to exploit tumour chorioallantoic membrane models (CAMs) in radio-investigations within clinical equipment and evaluate the performance of non-persistent nanoarchitectures (NAs) in combination with radiotherapy with respect to the standard concurrent chemoradiotherapy for the treatment of HPV-negative HNSCCs. A comparable effect has been observed between the tested approaches, suggesting NAs as a potential platinum-free agent in concurrent chemoradiotherapy for HNSCCs. On a broader basis, our bioconvergence approach provides an advance for the translation of Pt-free radiosensitizer to the clinical practice, positively shifting the therapeutic vs. side effects equilibrium for the management of HNSCCs.

Development of a highly porous bioscaffold by the combination of bubble entrapping and freezing-thawing techniques to fabricate hyaluronic acid/gelatin tri-layer wound dressing

The electrospun nanofibers and porous scaffolds hold great promise in regenerative medicine. A novel nanofiber-hydrogel‑silicone tri-layer wound dressing has been designed and fabricated to address the limitations of each platform. The bottom nanofiber layer with a 110 mm diameter meets the wound surface and regulates cell attachment and migration. The middle hydrogel layer was fabricated through the optimization of chemical crosslink formation and freezing-thawing cycles (physical crosslink). The fabricated hydrogel with interconnected porous structure has optimized properties (gel fraction (89.45 %) and porosity (80 %)) for wound dressing application. The silicone layer on the outer surface was designed to fix the wound dressing on the skin and prevent the penetration of pathogens. The scanning electron microscope micrograph showed structural integrity in the tri-layer scaffold. In vivo data showed that the tri-layer scaffold accelerates wound healing in the mice model and angiogenesis in the chorioallantoic membrane model. Therefore, the designed scaffold inspired by the skin's structure can be used as a wound dressing to treat wounds.

A partial epithelial-mesenchymal transition signature for highly aggressive colorectal cancer cells that survive under nutrient restriction.

Partial epithelial-mesenchymal transition (p-EMT) has recently been identified as a hybrid state consisting of cells with both epithelial and mesenchymal characteristics and is associated with the migration, metastasis, and chemoresistance of cancer cells. Here, we describe the induction of p-EMT in starved colorectal cancer (CRC) cells and identify a p-EMT gene signature that can predict prognosis. Functional characterisation of starvation-induced p-EMT in HCT116, DLD1, and HT29 cells showed changes in proliferation, morphology, and drug sensitivity, supported by in vivo studies using the chorioallantoic membrane model. An EMT-specific quantitative polymerase chain reaction (qPCR) array was used to screen for deregulated genes, leading to the establishment of an in silico gene signature that was correlated with poor disease-free survival in CRC patients along with the CRC consensus molecular subtype CMS4. Among the significantly deregulated p-EMT genes, a triple-gene signature consisting of SERPINE1, SOX10, and epidermal growth factor receptor (EGFR) was identified. Starvation-induced p-EMT was characterised by increased migratory potential and chemoresistance, as well as E-cadherin processing and internalisation. Both gene signature and E-cadherin alterations could be reversed by the proteasomal inhibitor MG132. Spatially resolving EGFR expression with high-resolution immunofluorescence imaging identified a proliferation stop in starved CRC cells caused by EGFR internalisation. In conclusion, we have gained insight into a previously undiscovered EMT mechanism that may become relevant when tumour cells are under nutrient stress, as seen in early stages of metastasis. Targeting this process of tumour cell dissemination might help to prevent EMT and overcome drug resistance. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Evaluation of chicken chorioallantoic membrane model for tumor imaging and drug development: Promising findings.

The chicken chorioallantoic membrane (CAM) model is a potential alternative to the mouse model based on the 3R principles. However, its value for determination of the invivo behaviors of radiolabeled peptides through positron emission tomography (PET) imaging needed investigation. Herein, the chicken CAM tumor models were established, and their feasibility was evaluated for evaluating the imaging properties of radiolabeled peptides using a 68 Ga-labeled HER2 affibody. Two human breast cancer cell lines were inoculated into chicken CAM and mice, respectively. The tumor-targeting potential and pharmacokinetic profile of a 68 Ga-labeled affibody, 68 Ga-MZHER, in both tumor models were also determined. The tumor-formation time in chicken CAM model was shorter than that of mouse model. The uptake values of human epithelial growth factor receptor-2 (HER2)-positive Bcap37 tumors in chicken CAM and mouse models were 5.36 ± 0.26% ID/g and 5.26 ± 0.43% ID/g at 30 min postinjection of 68 Ga-MZHER, respectively. At the same time points, the uptake values of HER2-negative MDA-MB-231 tumors in the chicken CAM models and mouse models were 1.57 ± 0.15% ID/g and 1.67 ± 0.25% ID/g, respectively. Exvivo biodistribution confirmed that more radioactivity accumulated in Bcap37 tumors than in MDA-MD-231 tumors in both CAM and mouse models. In this study, the CAM tumor model was successfully prepared. The chicken CAM model is a novel tool for quickly determining the invivo properties of radiolabeled peptides targeting biomarkers. It may be beneficial for early monitoring of the therapeutic effect of a new drug through PET imaging with specific peptides.

New retinoblastoma (RB) drug delivery approaches: anti-tumor effect of atrial natriuretic peptide (ANP)-conjugated hyaluronic-acid-coated gold nanoparticles for intraocular treatment of chemoresistant RB.

Intraocular drug delivery is a promising approach for treatment of ocular diseases. Chemotherapeutic drugs used in retinoblastoma (RB) treatment often lead to side effects and drug resistances. Therefore, new adjuvant therapies are needed to treat chemoresistant RBs. Biocompatible gold nanoparticles (GNPs) have unique antiangiogenic properties and can inhibit cancer progression. The combination of gold and low-molecular-weight hyaluronan (HA) enhances the stability of GNPs and promotes the distribution across ocular barriers. Attached to HA-GNPs, the atrial natriuretic peptide (ANP), which diminishes neovascularization in the eye, is a promising new therapeutic agent for RB treatment. In the study presented, we established ANP-coupled HA-GNPs and investigated their effect on the tumor formation potential of chemoresistant RB cells in an in ovo chicken chorioallantoic membrane model and an orthotopic in vivo RB rat eye model. Treatment of etoposide-resistant RB cells with ANP-HA-GNPs in ovo resulted in significantly reduced tumor growth and angiogenesis compared with controls. The antitumorigenic effect could be verified in the rat eye model, including a noninvasive application form via eye drops. Our data suggest that ANP-HA-GNPs represent a new minimally invasive, adjuvant treatment option for RB.