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Stem Cell Research & Therapy Sep 2023To explore whether local transplantation of mesenchymal stem cells (MSCs) in temporal muscle can promote collateral angiogenesis and to analyze its main mechanisms of...
BACKGROUND
To explore whether local transplantation of mesenchymal stem cells (MSCs) in temporal muscle can promote collateral angiogenesis and to analyze its main mechanisms of promoting angiogenesis.
METHODS
Bilateral carotid artery stenosis (BCAS) treated mice were administrated with encephalo-myo-synangiosis (EMS), and bone marrow mesenchymal stem cells (BMSCs) were transplanted into the temporal muscle near the cerebral cortex. On the 30th day after EMS, the Morris water maze, immunofluorescence, laser speckle imaging, and light sheet microscopy were performed to evaluate angiogenesis; In addition, rats with bilateral common carotid artery occlusion were also followed by EMS surgery, and BMSCs from GFP reporter rats were transplanted into the temporal muscle to observe the survival time of BMSCs. Then, the concentrated BMSC-derived conditioned medium (BMSC-CM) was used to stimulate HUVECs and BMECs for ki-67 immunocytochemistry, CCK-8, transwell and chick chorioallantoic membrane assays. Finally, the cortical tissue near the temporal muscle was extracted after EMS, and proteome profiler (angiogenesis array) as well as RT-qPCR of mRNA or miRNA was performed.
RESULTS
The results of the Morris water maze 30 days after BMSC transplantation in BCAS mice during the EMS operation, showed that the cognitive impairment in the BCAS + EMS + BMSC group was alleviated (P < 0.05). The results of immunofluorescence, laser speckle imaging, and light sheet microscopy showed that the number of blood vessels, blood flow and astrocytes increased in the BCAS + EMS + BMSC group (P < 0.05). The BMSCs of GFP reporter rats were applied to EMS and showed that the transplanted BMSCs could survive for up to 14 days. Then, the results of ki-67 immunocytochemistry, CCK-8 and transwell assays showed that the concentrated BMSC-CM could promote the proliferation and migration of HUVECs and BMECs (P < 0.05). Finally, the results of proteome profiler (angiogenesis array) in the cerebral cortex showed that the several pro-angiogenesis factors (such as MMP-3, MMP-9, IGFBP-2 or IGFBP-3) were notably highly expressed in MSC transplantation group compared to others.
CONCLUSIONS
Local MSCs transplantation together with EMS surgery can promote angiogenesis and cognitive behavior in chronic brain ischemia mice. Our study illustrated that MSC local transplantation can be the potential therapeutical option for improving EMS treatment efficiency which might be translated into clinical application.
Topics: Mice; Rats; Animals; Ki-67 Antigen; Proteome; Sincalide; Neovascularization, Pathologic; Mesenchymal Stem Cells; Brain Ischemia
PubMed: 37667370
DOI: 10.1186/s13287-023-03465-7 -
Science Advances Aug 2023amplification () is a defining feature of high-risk neuroblastoma (NB) and predicts poor prognosis. However, whether genes within or in close proximity to the amplicon...
amplification () is a defining feature of high-risk neuroblastoma (NB) and predicts poor prognosis. However, whether genes within or in close proximity to the amplicon also contribute to NB remains poorly understood. Here, we identify that , a transcription factor encoding gene neighboring the locus, is frequently coexpressed with and promotes cell survival in NB. GREB1 controls gene expression independently of MYCN, among which we uncover myosin 1B () as being highly expressed in NB and, using a chick chorioallantoic membrane (CAM) model, as a crucial regulator of invasion and metastasis. Global secretome and proteome profiling further delineates MYO1B in regulating secretome reprogramming in NB cells, and the cytokine MIF as an important pro-invasive and pro-metastatic mediator of MYO1B activity. Together, we have identified a putative GREB1-MYO1B-MIF axis as an unconventional mechanism promoting aggressive behavior in NB and independently of MYCN.
Topics: Humans; N-Myc Proto-Oncogene Protein; Secretome; Neuroblastoma; Aggression; Cell Survival
PubMed: 37611092
DOI: 10.1126/sciadv.adg6693 -
Journal of Cell Science Nov 2023Centrosome amplification (CA) is a prominent feature of human cancers linked to tumorigenesis in vivo. Here, we report mechanistic contributions of CA induction alone to...
Centrosome amplification (CA) is a prominent feature of human cancers linked to tumorigenesis in vivo. Here, we report mechanistic contributions of CA induction alone to tumour architecture and extracellular matrix (ECM) remodelling. CA induction in non-tumorigenic breast cells MCF10A causes cell migration and invasion, with underlying disruption of epithelial cell-cell junction integrity and dysregulation of expression and subcellular localisation of cell junction proteins. CA also elevates expression of integrin β-3, its binding partner fibronectin-1 and matrix metalloproteinase enzymes, promoting cell-ECM attachment, ECM degradation, and a migratory and invasive cell phenotype. Using a chicken embryo xenograft model for in vivo validation, we show that CA-induced (+CA) MCF10A cells invade into the chick mesodermal layer, with inflammatory cell infiltration and marked focal reactions between chorioallantoic membrane and cell graft. We also demonstrate a key role of small GTPase Rap-1 signalling through inhibition using GGTI-298, which blocked various CA-induced effects. These insights reveal that in normal cells, CA induction alone (without additional oncogenic alterations) is sufficient to confer early pro-tumorigenic changes within days, acting through Rap-1-dependent signalling to alter cell-cell contacts and ECM disruption.
Topics: Chick Embryo; Humans; Animals; Female; Chickens; Neoplasms; Signal Transduction; Cell Movement; Centrosome; Cell Line, Tumor; Breast Neoplasms
PubMed: 37772773
DOI: 10.1242/jcs.261150 -
International Journal of Molecular... Sep 2023The chorioallantoic membrane (CAM) can be used as a valuable research tool to examine tumors. The CAM can be used to investigate processes such as migration, invasion,... (Review)
Review
The chorioallantoic membrane (CAM) can be used as a valuable research tool to examine tumors. The CAM can be used to investigate processes such as migration, invasion, and angiogenesis and to assess novel antitumor drugs. The CAM can be used to establish tumors in a straightforward, rapid, and cost-effective manner via xenotransplantation of cells or tumor tissues with reproducible results; furthermore, the use of the CAM adheres to the three "R" principle, i.e., replace, reduce, and refine. To achieve successful tumor establishment and survival, several technical aspects should be taken into consideration. The complexity and heterogeneity of diseases including neuroblastoma and cancers in general and their impact on human health highlight the importance of preclinical models that help us describe tumor-specific biological processes. These models will not only help in understanding tumor biology, but also allow clinicians to explore therapeutic alternatives that will improve current treatment strategies. In this review, we summarize the technical characteristics as well as the main findings regarding the use of this model to study neuroblastoma for angiogenesis, metastasis, drug sensitivity, and drug resistance.
Topics: Animals; Chick Embryo; Humans; Chickens; Chorioallantoic Membrane; Neuroblastoma; Antineoplastic Agents; Biology
PubMed: 37834193
DOI: 10.3390/ijms241914744 -
Cell Death & Disease Jan 2024GRB2 is an adaptor protein of HER2 (and several other tyrosine kinases), which we identified as a novel BECN1 (Beclin 1) interacting partner. GRB2 co-immunoprecipitated...
GRB2 is an adaptor protein of HER2 (and several other tyrosine kinases), which we identified as a novel BECN1 (Beclin 1) interacting partner. GRB2 co-immunoprecipitated with BECN1 in several breast cancer cell lines and regulates autophagy through a mechanism involving the modulation of the class III PI3Kinase VPS34 activity. In ovo studies in a CAM (Chicken Chorioallantoic Membrane) model indicated that GRB2 knockdown, as well as overexpression of GRB2 loss-of-function mutants (Y52A and S86A-R88A) compromised tumor growth. These differences in tumor growth correlated with differential autophagy activity, indicating that autophagy effects might be related to the effects on tumorigenesis. Our data highlight a novel function of GRB2 as a BECN1 binding protein and a regulator of autophagy.
Topics: Animals; Adaptor Proteins, Signal Transducing; Autophagy; Beclin-1; Carcinogenesis; Cell Transformation, Neoplastic; Humans; GRB2 Adaptor Protein
PubMed: 38182563
DOI: 10.1038/s41419-023-06387-7 -
European Radiology Experimental Sep 2023We evaluated the feasibility of a chick chorioallantoic membrane (CAM) tumor model for preclinical research on tumor radiofrequency ablation (RFA).
BACKGROUND
We evaluated the feasibility of a chick chorioallantoic membrane (CAM) tumor model for preclinical research on tumor radiofrequency ablation (RFA).
METHODS
Fertilized chicken eggs were incubated and divided into five cohorts: RFA for 30 s (n = 5), RFA for 60 s (n = 5), RFA for 120 s (n = 4), sham (n = 8), and controls (n = 6). Xenografting using pancreatic neuroendocrine tumor cells of the BON-1 cell line was performed on embryonic day (ED) 8. The RFA was performed on ED 12. Survival, stereomicroscopic observations, and histological observations using hematoxylin-eosin (H&E) and Ki67 staining were evaluated.
RESULTS
The survival rates in the 30-s, 60-s, and 120-s, sham and control cohort were 60%, 60%, 0%, 100%, and 50%, respectively. Signs of bleeding and heat damage were common findings in the evaluation of stereomicroscopic observations. Histological examination could be performed in all but one embryo. Heat damage, bleeding, thrombosis, and leukocyte infiltration and hyperemia were regular findings in H&E-stained cuts. A complete absence of Ki67 staining was recorded in 33.3% and 50% of embryos in the 30-s and 60-s cohorts that survived until ED 14, respectively.
CONCLUSIONS
The CAM model is a feasible and suiting research model for tumor RFA with many advantages over other animal models. It offers the opportunity to conduct in vivo research under standardized conditions. Further studies are needed to optimize this model for tumor ablations in order to explore promising but unrefined strategies like the combination of RFA and immunotherapy.
RELEVANCE STATEMENT
The chick chorioallantoic membrane model allows in vivo research on tumor radiofrequency ablation under standardized conditions that may enable enhanced understanding on combined therapies while ensuring animal welfare in concordance with the "Three Rs."
KEY POINTS
• The chorioallantoic membrane model is feasible and suiting for tumor radiofrequency ablation. • Radiofrequency ablation regularly achieved reduction but not eradication of Ki67 staining. • Histological evaluation showed findings comparable to changes in humans after RFA. • The chorioallantoic membrane model can enable studies on combined therapies after optimization.
Topics: Humans; Animals; Chickens; Chorioallantoic Membrane; Feasibility Studies; Ki-67 Antigen; Pancreatic Neoplasms; Eosine Yellowish-(YS)
PubMed: 37749303
DOI: 10.1186/s41747-023-00368-3 -
Genomics Jan 2024Dimorphism between male and female embryos has been demonstrated in many animal species, including chicken species. Likewise, extraembryonic membranes such as the...
Dimorphism between male and female embryos has been demonstrated in many animal species, including chicken species. Likewise, extraembryonic membranes such as the chorioallantoic membrane (CAM) are likely to exhibit a sex-specific profile. Analysis of the previously published RNA-seq data of the chicken CAM sampled at two incubation times, revealed 783 differentially expressed genes between the CAM of male and female embryos. The expression of some of these genes is sex-dependant only at one or other stage of development, while 415 genes are sex-dependant at both developmental stages. These genes include well-known sex-determining and sex-differentiation genes (DMRT1, HEGM, etc.), and are mainly located on sex chromosomes. This study provides evidence that gene expression of extra-embryonic membranes is differentially regulated between male and female embryos. As such, a better characterisation of associated mechanisms should facilitate the identification of new sex-specific biomarkers.
Topics: Animals; Male; Female; Chickens; Transcriptome; Chorioallantoic Membrane; Sex Differentiation; Gene Expression Regulation, Developmental
PubMed: 38061480
DOI: 10.1016/j.ygeno.2023.110754 -
Biophysics Reports Oct 2023The development of the vascular system is essential for embryonic development, including processes such as angiogenesis. Angiogenesis plays a critical role in many...
The development of the vascular system is essential for embryonic development, including processes such as angiogenesis. Angiogenesis plays a critical role in many normal physiological and pathological processes. It is driven by a set of angiogenic proteins, including angiogenic growth factors, chemokines, and extracellular matrix proteins. Among various animal model systems, the chorioallantoic membrane (CAM), a specialized and highly vascularized tissue of the avian embryo, has proven to be a valuable tool for analyzing the angiogenic potential of candidate cells or factors. In this protocol, we provide detailed procedures for establishing the CAM model to evaluate the function and mechanism of migrasomes in embryonic angiogenesis. This includes the CAM nylon mesh assay and CAM sprouting assay to assess CAM angiogenesis, as well as the observation, purification, and delivery of migrasomes. Additionally, we describe the generation of T4-KO-mCherry-KI embryos using the CRISPR system within the CAM tissue to investigate the role of migrasomes in angiogenesis.
PubMed: 38516298
DOI: 10.52601/bpr.2023.230021 -
Neuro-oncology Sep 2023High-grade gliomas (HGG) are aggressive brain tumors associated with short median patient survival and limited response to therapies, driving the need to develop tools...
BACKGROUND
High-grade gliomas (HGG) are aggressive brain tumors associated with short median patient survival and limited response to therapies, driving the need to develop tools to improve patient outcomes. Patient-derived xenograft (PDX) models, such as mouse PDX, have emerged as potential Avatar platforms for personalized oncology approaches, but the difficulty for some human grafts to grow successfully and the long time required for mice to develop tumors preclude their use for HGG.
METHODS
We used a rapid and efficient ex-ovo chicken embryo chorioallantoic membrane (CAM) culture system to evaluate the efficacy of oncologic drug options for HGG patients.
RESULTS
Implantation of fresh glioma tissue fragments from 59 of 60 patients, that include difficult-to-grow IDH-mutated samples, successfully established CAM tumor xenografts within 7 days, with a tumor take rate of 98.3%. These xenografts faithfully recapitulate the histological and molecular characteristics of the primary tumor, and the ability of individual fragments to form tumors was predictive of poor patient prognosis. Treatment of drug-sensitive or drug-resistant xenografts indicates that the CAM-glioma assay enables testing tumor sensitivity to temozolomide and carboplatin at doses consistent with those administered to patients. In a proof-of-concept study involving 14 HGG patients, we observed a correlation of 100% between the CAM xenograft response to temozolomide or carboplatin and the clinical response of patients.
CONCLUSION
The CAM-glioma model is a fast and reliable assay that has the potential to serve as a complementary model to drug discovery and a real-time Avatar platform to predict the best treatment for HGG patients.
Topics: Humans; Chick Embryo; Mice; Animals; Temozolomide; Heterografts; Carboplatin; Glioma; Brain Neoplasms; Disease Models, Animal; Xenograft Model Antitumor Assays
PubMed: 36821432
DOI: 10.1093/neuonc/noad047 -
Biomaterials Advances Nov 2023Triple negative breast cancer (TNBC) is a highly heterogenous disease not sensitive to endocrine or HER2 therapy and standardized treatment regimens are still missing....
Triple negative breast cancer (TNBC) is a highly heterogenous disease not sensitive to endocrine or HER2 therapy and standardized treatment regimens are still missing. Therefore, development of novel TNBC treatment approaches is of utmost relevance. Herein, the potential of MAPK/ERK downregulation by RNAi-based therapeutics in a panel of mesenchymal stem-like TNBC cell lines was uncovered. Our data revealed that suppression of one of the central nodes of this signaling pathway, MEK1, affects proliferation, migration, and invasion of TNBC cells, that may be explained by the reversion of the epithelial-mesenchymal transition phenotype, which is facilitated by the MMP-2/MMP-9 downregulation. Moreover, an exosome-based system was successfully generated for the siRNA loading (iExoMEK1). Our data suggested absence of modification of the physical properties and general integrity of the iExoMEK1 comparatively to the unmodified counterparts. Such exosome-mediated downregulation of MEK1 led to a tumor regression accompanied by a decrease of angiogenesis using the chick chorioallantoic-membrane model. Our results highlight the potential of the targeting of MAPK/ERK cascade as a promising therapeutic approach against TNBC.
Topics: Humans; Cell Proliferation; Cell Line, Tumor; RNA, Small Interfering; Triple Negative Breast Neoplasms; Exosomes
PubMed: 37778291
DOI: 10.1016/j.bioadv.2023.213643