Extracellular Matrix Compartment Research Articles

Validation of an organ mapping antibody panel for cyclical immunofluorescence microscopy on normal human kidneys.

The lack of standardization in antibody validation remains a major contributor to irreproducibility of human research. To address this, we have applied a standardized approach to validate a panel of antibodies to identify 18 major cell types and 5 extracellular matrix compartments in the human kidney by immunofluorescence (IF) microscopy. We have used these to generate an organ mapping antibody panel for two-dimensional (2-D) and three-dimensional (3-D) cyclical IF (CyCIF) to provide a more detailed method for evaluating tissue segmentation and volumes using a larger panel of markers than would normally be possible using standard fluorescence microscopy. CyCIF also makes it possible to perform multiplexed IF microscopy of whole slide images, which is a distinct advantage over other multiplexed imaging technologies that are applicable to limited fields of view. This enables a broader view of cell distributions across larger anatomical regions, allowing a better chance to capture localized regions of dysfunction in diseased tissues. These methods are broadly accessible to any laboratory with a fluorescence microscope, enabling spatial cellular phenotyping in normal and disease states. We also provide a detailed solution for image alignment between CyCIF cycles that can be used by investigators to perform these studies without programming experience using open-sourced software. This ability to perform multiplexed imaging without specialized instrumentation or computational skills opens the door to integration with more highly dimensional molecular imaging modalities such as spatial transcriptomics and imaging mass spectrometry, enabling the discovery of molecular markers of specific cell types, and how these are altered in disease.NEW & NOTEWORTHY We describe here validation criteria used to define on organ mapping panel of antibodies that can be used to define 18 cell types and five extracellular matrix compartments using cyclical immunofluorescence (CyCIF) microscopy. As CyCIF does not require specialized instrumentation, and image registration required to assemble CyCIF images can be performed by any laboratory without specialized computational skills, this technology is accessible to any laboratory with access to a fluorescence microscope and digital scanner.

Engineering Tumor Stroma Morphogenesis Using Dynamic Cell-Matrix Spheroid Assembly.

The tumor microenvironment consists of resident tumor cells organized within a compositionally diverse, three-dimensional (3D) extracellular matrix (ECM) network that cannot be replicated in vitro using bottom-up synthesis. We report a new self-assembly system to engineer ECM-rich 3D MatriSpheres wherein tumor cells actively organize and concentrate microgram quantities of decellularized ECM dispersions which modulate cell phenotype. 3D colorectal cancer (CRC) MatriSpheres were created using decellularized small intestine submucosa (SIS) as an orthotopic ECM source that had greater proteomic homology to CRC tumor ECM than traditional ECM formulations such as Matrigel. SIS ECM was rapidly concentrated from its environment and assembled into ECM-rich 3D stroma-like regions by mouse and human CRC cell lines within 4-5 days via a mechanism that was rheologically distinct from bulk hydrogel formation. Both ECM organization and transcriptional regulation by 3D ECM cues affected programs of malignancy, lipid metabolism, and immunoregulation that corresponded with an in vivo MC38 tumor cell subpopulation identified via single cell RNA sequencing. This 3D modeling approach stimulates tumor specific tissue morphogenesis that incorporates the complexities of both cancer cell and ECM compartments in a scalable, spontaneous assembly process that may further facilitate precision medicine.

Abstract B095: Tenascin C promotes ovarian cancer stem cells interactions with the niche

Abstract Background: Effective treatment of ovarian cancer (OC) is greatly impacted by advanced, late-stage diagnosis which usually presents with intra-peritoneal metastases, chemoresistance and high recurrence. The development of intra-peritoneal metastases is correlated with the formation of multicellular spheroids disseminated in the peritoneal fluid. Spheroid formation is promoted by a change of cell adhesion properties, enhanced secretion of extracellular matrix (ECM) components that stiffen the tumor stroma and promote the aberrant activation of oncogenic processes, including survival, invasion, and stemness. Tenascin C (TNC) is one of the molecules secreted in the ECM and correlates with migration and metastatic progression in a variety of carcinomas. However, the functional role of TNC in spheroid formation and in the maintenance of the platinum resistant ovarian cancer stem phenotype has not been investigated yet. We tested the hypothesis that the increased expression of TNC and its secretion in the ECM compartment promotes survival of OCSCs, supports OC spheroid formation and tumor progression. Methods: We compared TNC expression in OC cells grown as spheroids and monolayers and in chemoresistant vs sensitive OC cells by realtime-PCR and Western Blot (WB). TNC blockade by siRNA knockdown (KD) was evaluated in combination with carboplatin on spheroid assay, colony formation, and by qPCR of stemness related gene expression. Chromatin immunoprecipitation (ChIP) assay detected the interaction between the β-catenin and the TNC promoter. Results: OC cells grown as spheroids showed a significant increase in TNC expression compared to monolayers. Our results also demonstrated an increase in TNC expression levels in platinum resistant OC cell lines when compared to the platinum sensitive counterpart. A comparison of normal adjacent ovarian epithelium and primary OC tumors on a multitissue array revealed that TNC expression coincided with a pro-tumorigenic phenotype (n=88, P<0.001), correlating with poor outcome. TNC-KD in combination with carboplatin led to a significant decrease in the expression levels of stemness-related genes, spheroid and colony formation. Mechanistically, b-catenin-KD decreased TNC expression levels and β-catenin coimmunoprecipitated with the TNC promoter, suggesting that TNC is a direct β-catenin target. Additionally, OC spheroids treated with the Wnt ligand WNT-3A showed increased TNC expression compared to control untreated cells, as demonstrated by WB. Importantly, the enhanced expression of TNC was followed by its secretion in the ECM compartment, as observed via immunofluorescence imaging, implying a direct role of TNC in the ECM-rearrangement which is responsible for CSC maintenance and spheroid formation. Conclusions: TNC is strongly linked to chemoresistance and is regulated by the oncogenic β-catenin pathway. Our data suggest that strategies aimed at interfering with TNC expression and/or secretion along with carboplatin treatment could be a potential combination therapy in OC. Citation Format: Amber Rogers, Maya Shebeshi, Morganna Gordon, Rula Atwani, Virginie Lazar, Salvatore Condello. Tenascin C promotes ovarian cancer stem cells interactions with the niche [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr B095.

Disrupted extracellular matrix and cell cycle genes in autism-associated Shank3 deficiency are targeted by lithium

The Shank3 gene encodes the major postsynaptic scaffolding protein SHANK3. Its mutation causes a syndromic form of autism spectrum disorder (ASD): Phelan-McDermid Syndrome (PMDS). It is characterized by global developmental delay, intellectual disorders (ID), ASD behavior, affective symptoms, as well as extra-cerebral symptoms. Although Shank3 deficiency causes a variety of molecular alterations, they do not suffice to explain all clinical aspects of this heterogenic syndrome. Since global gene expression alterations in Shank3 deficiency remain inadequately studied, we explored the transcriptome in vitro in primary hippocampal cells from Shank3∆11(−/−) mice, under control and lithium (Li) treatment conditions, and confirmed the findings in vivo. The Shank3∆11(−/−) genotype affected the overall transcriptome. Remarkably, extracellular matrix (ECM) and cell cycle transcriptional programs were disrupted. Accordingly, in the hippocampi of adolescent Shank3∆11(−/−) mice we found proteins of the collagen family and core cell cycle proteins downregulated. In vitro Li treatment of Shank3∆11(−/−) cells had a rescue-like effect on the ECM and cell cycle gene sets. Reversed ECM gene sets were part of a network, regulated by common transcription factors (TF) such as cAMP responsive element binding protein 1 (CREB1) and β-Catenin (CTNNB1), which are known downstream effectors of synaptic activity and targets of Li. These TFs were less abundant and/or hypo-phosphorylated in hippocampi of Shank3∆11(−/−) mice and could be rescued with Li in vitro and in vivo. Our investigations suggest the ECM compartment and cell cycle genes as new players in the pathophysiology of Shank3 deficiency, and imply involvement of transcriptional regulators, which can be modulated by Li. This work supports Li as potential drug in the management of PMDS symptoms, where a Phase III study is ongoing.

Decellularized diseased tissues: current state-of-the-art and future directions.

Decellularized matrices derived from diseased tissues/organs have evolved in the most recent years, providing novel research perspectives for understanding disease occurrence and progression and providing accurate pseudo models for developing new disease treatments. Although decellularized matrix maintaining the native composition, ultrastructure, and biomechanical characteristics of extracellular matrix (ECM), alongside intact and perfusable vascular compartments, facilitates the construction of bioengineered organ explants in vitro and promotes angiogenesis and tissue/organ regeneration in vivo, the availability of healthy tissues and organs for the preparation of decellularized ECM materials is limited. In this paper, we review the research advancements in decellularized diseased matrices. Considering that current research focuses on the matrices derived from cancers and fibrotic organs (mainly fibrotic kidney, lungs, and liver), the pathological characterizations and the applications of these diseased matrices are mainly discussed. Additionally, a contrastive analysis between the decellularized diseased matrices and decellularized healthy matrices, along with the development in vitro 3D models, is discussed in this paper. And last, we have provided the challenges and future directions in this review. Deep and comprehensive research on decellularized diseased tissues and organs will promote in-depth exploration of source materials in tissue engineering field, thus providing new ideas for clinical transformation.

Mechanobiological implications of age-related remodelling in the outer retina

The outer retina consists of the light-sensitive photoreceptors, the pigmented epithelium, and the choroid, which interact in a complex manner to sustain homeostasis. The organisation and function of these cellular layers are mediated by the extracellular matrix compartment named Bruch's membrane, situated between the retinal epithelium and the choroid. Like many tissues, the retina experiences age-related structural and metabolic changes, which are relevant for understanding major blinding diseases of the elderly, such as age-related macular degeneration. Compared with other tissues, the retina mainly comprises postmitotic cells, making it less able to maintain its mechanical homeostasis over the years functionally. Aspects of retinal ageing, like the structural and morphometric changes of the pigment epithelium and the heterogenous remodelling of the Bruch's membrane, imply changes in tissue mechanics and may affect functional integrity. In recent years, findings in the field of mechanobiology and bioengineering highlighted the importance of mechanical changes in tissues for understanding physiological and pathological processes. Here, we review the current knowledge of age-related changes in the outer retina from a mechanobiological perspective, aiming to generate food for thought for future mechanobiology studies in the outer retina.

Multi-compartment tumor organoids

Organoid cultures are widely used for tumor modeling because they preserve many phenotypic features of cancer cells in vivo. However, current organoids present issues of consistency, efficiency, mimicry, and cell-seeding control. More importantly, they can only contain only one extracellular matrix (ECM) compartment at a time, while solid tumors feature two main ECM compartments: the basement membrane and the stromal matrix. Here, we develop, test, and validate a high-throughput oil-in-water droplet microtechnology to generate highly uniform, small-volume, multi-compartment organoids. Each organoid culture features microenvironmental architectures that mimic both the basement membrane and stromal barriers. This matrix architecture, which allows us to simultaneously take into account and assess the proliferative and invasive properties of cancer cells in a single platform, has profound effect on observed drug responsiveness and tumor progression that correlate well with in vivo and clinical outcomes. Our method was tested on multiple types of cells including primary breast and ovarian cancer cells and immortalized cell lines, and we determined our platform is suitable even for cancer cells of poor standard organoid-forming ability such as primary patient samples. These new organoids also allow for direct orthotopic mouse implantation of cancer cells with unprecedented success.

Matrix remodeling controls a nuclear lamin A/C-emerin network that directs Wnt-regulated stem cell fate.

Skeletal stem cells (SSCs) reside within a three-dimensional extracellular matrix (ECM) compartment and differentiate into multiple cell lineages, thereby controlling tissue maintenance and regeneration. Within this environment, SSCs can proteolytically remodel the surrounding ECM in response to growth factors that direct lineage commitment via undefined mechanisms. Here, we report that Mmp14-dependent ECM remodeling coordinates canonical Wnt signaling and guides stem cell fate by triggering an integrin-activated reorganization of the SCC cytoskeleton that controls nuclear lamin A/C levels via the linker of nucleoskeleton and cytoskeleton (LINC) complexes. In turn, SSC lamin A/C levels dictate the localization of emerin, an inner nuclear membrane protein whose ability to regulate β-catenin activity modulates Wnt signaling while directing lineage commitment invitro and invivo. These findings define a previously undescribed axis wherein SSCs use Mmp14-dependent ECM remodeling to control cytoskeletal and nucleoskeletal organization, thereby governing Wnt-dependent stem cell fate decisions.

Retinoic Acid Receptors and the Control of Positional Information in the Regenerating Axolotl Limb.

We know little about the control of positional information (PI) during axolotl limb regeneration, which ensures that the limb regenerates exactly what was amputated, and the work reported here investigates this phenomenon. Retinoic acid administration changes the PI in a proximal direction so that a complete limb can be regenerated from a hand. Rather than identifying all the genes altered by RA treatment of the limb, we have eliminated many off-target effects by using retinoic acid receptor selective agonists. We firstly identify the receptor involved in this respecification process as RARα and secondly, identify the genes involved by RNA sequencing of the RARα-treated blastemal mesenchyme. We find 1177 upregulated genes and 1403 downregulated genes, which could be identified using the axolotl genome. These include several genes known to be involved in retinoic acid metabolism and in patterning. Since positional information is thought to be a property of the cell surface of blastemal cells when we examine our dataset with an emphasis on this aspect, we find the top canonical pathway is integrin signaling. In the extracellular matrix compartment, we find a MMP and several collagens are upregulated; several cell membrane genes and secretory factors are also upregulated. This provides data for future testing of the function of these candidates in the control of PI during limb regeneration.

Potential Therapeutic Significance of Laminin in Head and Neck Squamous Carcinomas.

Simple SummaryHead and neck cancers (HNC) account for approximately 500,000 new cases of tumors annually worldwide and are represented by upper aerodigestive tract malignant neoplasms, which particularly arise in oral cavity, larynx, and pharynx tissues. Thus, due to the biological diversity between the upper aerodigestive organs, and to the heterogeneity of risk factors associated with their malignant transformation, HNC behavior, and prognosis seem to strongly vary according to the tumor site. However, despite to the heterogeneity which characterizes head and neck tumors, squamous cell carcinomas (SCC) represent the predominant histopathologic HNC subtype. In this sense, it has been reported that SCC tumor biology is strongly associated with deregulations within the extracellular matrix compartment. Accordingly, it has been shown that laminin plays a remarkable role in the regulation of crucial events associated with head and neck squamous cell carcinomas (HNSCC) progression, which opens the possibility that laminin may represent a convergence point in HNSCC natural history.Head and neck squamous cell carcinomas (HNSCC) are among the most common and lethal tumors worldwide, occurring mostly in oral cavity, pharynx, and larynx tissues. The squamous epithelia homeostasis is supported by the extracellular matrix (ECM), and alterations in this compartment are crucial for cancer development and progression. Laminin is a fundamental component of ECM, where it represents one of the main components of basement membrane (BM), and data supporting its contribution to HNSCC genesis and progression has been vastly explored in oral cavity squamous cell carcinoma. Laminin subtypes 111 (LN-111) and 332 (LN-332) are the main isoforms associated with malignant transformation, contributing to proliferation, adhesion, migration, invasion, and metastasis, due to its involvement in the regulation of several pathways associated with HNSCC carcinogenesis, including the activation of the EGFR/MAPK signaling pathway. Therefore, it draws attention to the possibility that laminin may represent a convergence point in HNSCC natural history, and an attractive potential therapeutic target for these tumors.

Expression Changes and Impact of the Extracellular Matrix on Etoposide Resistant Human Retinoblastoma Cell Lines.

Retinoblastoma (RB) represents the most common malignant childhood eye tumor worldwide. Several studies indicate that the extracellular matrix (ECM) plays a crucial role in tumor growth and metastasis. Moreover, recent studies indicate that the ECM composition might influence the development of resistance to chemotherapy drugs. The objective of this study was to evaluate possible expression differences in the ECM compartment of the parental human cell lines WERI-RB1 (retinoblastoma 1) and Y79 and their Etoposide resistant subclones via polymerase chain reaction (PCR). Western blot analyses were performed to analyze protein levels. To explore the influence of ECM molecules on RB cell proliferation, death, and cluster formation, WERI-RB1 and resistant WERI-ETOR cells were cultivated on Fibronectin, Laminin, Tenascin-C, and Collagen IV and analyzed via time-lapse video microscopy as well as immunocytochemistry. We revealed a significantly reduced mRNA expression of the proteoglycans Brevican, Neurocan, and Versican in resistant WERI-ETOR compared to sensitive WERI-RB1 cells. Also, for the glycoproteins α1-Laminin, Fibronectin, Tenascin-C, and Tenascin-R as well as Collagen IV, reduced expression levels were observed in WERI-ETOR. Furthermore, a downregulation was detected for the matrix metalloproteinases MMP2, MMP7, MMP9, the tissue-inhibitor of metalloproteinase TIMP2, the Integrin receptor subunits ITGA4, ITGA5 and ITGB1, and all receptor protein tyrosine phosphatase β/ζ isoforms. Downregulation of Brevican, Collagen IV, Tenascin-R, MMP2, TIMP2, and ITGA5 was also verified in Etoposide resistant Y79 cells compared to sensitive ones. Protein levels of Tenascin-C and MMP-2 were comparable in both WERI cell lines. Interestingly, Fibronectin displayed an apoptosis-inducing effect on WERI-RB1 cells, whereas an anti-apoptotic influence was observed for Tenascin-C. Conversely, proliferation of WERI-ETOR cells was enhanced on Tenascin-C, while an anti-proliferative effect was observed on Fibronectin. In WERI-ETOR, cluster formation was decreased on the substrates Collagen IV, Fibronectin, and Tenascin-C. Collectively, we noted a different ECM mRNA expression and behavior of Etoposide resistant compared to sensitive RB cells. These findings may indicate a key role of ECM components in chemotherapy resistance formation of RB.

Pancreatic islets seeded in a novel bioscaffold forms an organoid to rescue insulin production and reverse hyperglycemia in models of type 1 diabetes

Therapeutic approaches to combat type 1 diabetes (T1D) include donor pancreas transplantation, exogenous insulin administration and immunosuppressive therapies. However, these clinical applications are limited due to insufficient tissue compatible donors, side effects of exogenous insulin administration and/or increased onset of opportunistic infections attributable to induced global immunosuppression. An alternative approach to alleviate disease states is to utilize insulin-producing pancreatic islets seeded in a bioscaffold for implantation into diabetic recipients. The present studies now report that a newly developed cationic polymer biomaterial serves as an efficient bioscaffold for delivery of donor syngeneic pancreatic islet cells to reverse hyperglycemia in murine streptozotocin induced- or non-obese diabetic mouse models of T1D. Intraperitoneal implantation of pancreatic islets seeded within the copolymer bioscaffold supports long-term cell viability, response to extracellular signaling cues and ability to produce soluble factors into the microenvironment. Elevated insulin levels were measured in recipient diabetic mice upon implantation of the islet-seeded biomaterial coupled with reduced blood glucose levels, collectively resulting in increased survival and stabilization of metabolic indices. Importantly, the implanted islet-seeded biomaterial assembled into a solid organoid substructure that reorganized the extracellular matrix compartment and recruited endothelial progenitors for neovascularization. This allowed survival of the graft long-term in vivo and access to the blood for monitoring glucose levels. These results highlight the novelty, simplicity and effectiveness of this biomaterial for tissue regeneration and in vivo restoration of organ functions.

Corelattions Between CD31, CD68, MMP-2 and MMP-9 Expression in Allograft Cardiac Rejection – Immunohistochemical Study

Abstract Introduction. The cardiac allograft rejections from the post-transplant period are attributable to the acute cellular rejection monitored by multiple endomyocardial biopsies. Compared to this, humoral rejection remains a matter of debate, with multiple therapeutic strategies, poor prognosis, and persisting uncertainty about diagnostic criteria. Acute allograft rejection is associated with significant modifications of the extracellular matrix compartment mainly regulated by matrix metalloproteinases (MMPs). In this context, the aim of this study was to evaluate the expression of MMP-2 and -9 and CD31, CD68 (endothelial and histiocytic markers) and the correlations between them using immunohistochemistry, in patients with cardiac allografts. Materials and methods. Tissue fragments were obtained by endomyocardial biopsy from 5 patients with allograft heart transplant, 2 in the medium post-transplant phase and 2 in late phase. After identifying and characterizing the morphological context the probes were processed by standard immunohistochemical technique using anti-MMP-2 and anti-MMP-9 antibodies (Santa Cruz Biotechnology, Inc.) and anti-CD31, anti-CD68 antibodies (Sigma). The samples were examined using the Olympus BX40 microscope with an Olympus E330 camera attached. Results and discussions. Sample examination revealed in all 4 cases the lack of IR (-) for CD31 and weak IR (+) for CD68 compared to MMPs, where we found moderate IR (++) for MMP-9 and weak IR (+) for MMP-2. These aspects complets the histological lesional aspects of these cases, indicating the lack of acute rejection. In conclusion, CD31 and CD68 IR correlated with MMPs IR (especially MMP-9) appear to represent predictive markers for cardiac allograft rejection and require further studies.

Lobular architecture of human adipose tissue defines the niche and fate of progenitor cells

Human adipose tissue (hAT) is constituted of structural units termed lobules, the organization of which remains to be defined. Here we report that lobules are composed of two extracellular matrix compartments, i.e., septa and stroma, delineating niches of CD45−/CD34+/CD31− progenitor subsets characterized by MSCA1 (ALPL) and CD271 (NGFR) expression. MSCA1+ adipogenic subset is enriched in stroma while septa contains mainly MSCA1−/CD271− and MSCA1−/CD271high progenitors. CD271 marks myofibroblast precursors and NGF ligand activation is a molecular relay of TGFβ-induced myofibroblast conversion. In human subcutaneous (SC) and visceral (VS) AT, the progenitor subset repartition is different, modulated by obesity and in favor of adipocyte and myofibroblast fate, respectively. Lobules exhibit depot-specific architecture with marked fibrous septa containing mesothelial-like progenitor cells in VSAT. Thus, the human AT lobule organization in specific progenitor subset domains defines the fat depot intrinsic capacity to remodel and may contribute to obesity-associated cardiometabolic risks.

Microbial volatile communication in human organotypic lung models

We inhale respiratory pathogens continuously, and the subsequent signaling events between host and microbe are complex, ultimately resulting in clearance of the microbe, stable colonization of the host, or active disease. Traditional in vitro methods are ill-equipped to study these critical events in the context of the lung microenvironment. Here we introduce a microscale organotypic model of the human bronchiole for studying pulmonary infection. By leveraging microscale techniques, the model is designed to approximate the structure of the human bronchiole, containing airway, vascular, and extracellular matrix compartments. To complement direct infection of the organotypic bronchiole, we present a clickable extension that facilitates volatile compound communication between microbial populations and the host model. Using Aspergillus fumigatus, a respiratory pathogen, we characterize the inflammatory response of the organotypic bronchiole to infection. Finally, we demonstrate multikingdom, volatile-mediated communication between the organotypic bronchiole and cultures of Aspergillus fumigatus and Pseudomonas aeruginosa.

Histologic features of stromogenic carcinoma of the prostate (carcinomas with reactive stroma grade 3)

Prostatic carcinoma, like many other carcinomas, generates a stromal reaction. This phenomenon is well established in the scientific literature. The normal parenchymal smooth muscle phenotype switches to a myofibroblastic phenotype in response to the presence of cancer cells, with an expansion of the extracellular matrix compartment. The amount of reactive stroma is a predictor of biochemical recurrence in both radical prostatectomies and biopsies. It is a predictor of prostate cancer-specific death in prostatectomies. The aim of this study is to improve our histologic understanding of reactive stroma in prostate cancer and to determine the histologic features of the malignant epithelium found in stromogenic carcinomas or carcinomas with reactive stromal grade 3. Tissue microarrays of 800 patients and hematoxylin and eosin-stained sections of 120 radical prostatectomies, previously determined to contain a high proportion of areas with stromogenic carcinoma, were evaluated and findings systematically recorded. We identified 3 histologic patterns of reactive stroma: extracellular matrix-rich, cellular variant and edematous/myxoid variant. The most common pattern of carcinoma in stromogenic areas is of the acinar type with angulated glands and periglandular halos. The nuclei are enlarged, opened, with prominent nucleoli. Luminal borders are undulated, and amorphous pink secretion is often seen. Perineural invasion is frequently identified. Because of the clinical relevance, identification and quantification of areas with high reactive stromal grade by pathologists and reproducibility of our findings by others become essential. We believe that with the previously proposed grading system and the present morphologic description, both can be achieved.

Adapting extracellular matrix proteomics for clinical studies on cardiac remodeling post-myocardial infarction.

Following myocardial infarction (MI), the left ventricle (LV) undergoes a series of cardiac wound healing responses that involve stimulation of robust inflammation to clear necrotic myocytes and tissue debris and induction of extracellular matrix (ECM) protein synthesis to generate a scar. Proteomic strategies provide us with a means to index the ECM proteins expressed in the LV, quantify amounts, determine functions, and explore interactions. This review will focus on the efforts taken in the proteomics research field that have expanded our understanding of post-MI LV remodeling, concentrating on the strengths and limitations of different proteomic approaches to glean information that is specific to ECM turnover in the post-MI setting. We will discuss how recent advances in sample preparation and labeling protocols increase our successes at detecting components of the cardiac ECM proteome. We will summarize how proteomic approaches, focusing on the ECM compartment, have progressed over time to current gel-free methods using decellularized fractions or labeling strategies that will be useful for clinical applications. This review will provide an overview of how cardiac ECM proteomics has evolved over the last decade and will provide insight into future directions that will drive forward our understanding of cardiac ECM turnover in the post-MI LV.

Weaponizing human EGF-containing fibulin-like extracellular matrix protein 1 (EFEMP1) for 21st century cancer therapeutics.

De-regulated EFEMP1 gene expression in solid tumors has been widely reported with conflicting roles. We dissected EFEMP1 to identify domains responsible for its cell context-dependent dual functions, with the goal being to construct an EFEMP1-derived tumor-suppressor protein (ETSP) that lacked tumor-promoting function. Exon/intron boundaries of EFEMP1 were used as boundaries of functional modules in constructing EFEMP1 variants, with removal of various module(s), and/or mutating an amino acid residue to convert a weak integrin binding-site into a strong one. A series of in vitro assays on cancerous features, and subcutaneous and intracranial xenograft-formation assays, were carried out for effects from overexpression of wild-type and variant forms of EFEMP1 in two glioma subpopulations characterized as tumor mass-forming cells (TMCs) or stem-like tumor initiating cells (STICs), where EFEMP1 showed cellcontext- dependent dual functions. One of the EFEMP1 variants was identified as the sought-after ETSP, which had a stronger tumor-suppression function in TMCs by targeting EGFR and angiogenesis, and a new tumor-suppression function in STICs by targeting NOTCH signaling and MMP2-mediated cell invasion. Therefore, ETSP may form the basis for further important research to develop a novel cancer therapy to treat many types of cancer by its tumor suppressor effect in the extracellular matrix compartment.

MMP-9 signaling in the left ventricle following myocardial infarction.

Following myocardial infarction (MI), the left ventricle (LV) undergoes a series of cardiac wound healing responses that involve both the stimulation of robust inflammation to clear necrotic myocytes and tissue debris and the induction of extracellular matrix (ECM) protein synthesis to generate an infarct scar. The collective changes in myocardial structure and function are termed LV remodeling, and matrix metalloproteinase-9 (MMP-9) is a key instigator of post-MI LV remodeling. Through direct molecular effects on ECM and inflammatory protein turnover as well as indirect effects on major cell types that coordinate cardiac wound healing, namely the infiltrating leukocytes and the cardiac fibroblasts, MMP-9 coordinates multiple aspects of LV remodeling. In this review, we will discuss recent research that has expanded our understanding of post-MI LV remodeling, including recent proteomic advances focused on the ECM compartment to provide novel functional and translational insights. This overview will summarize how our understanding of MMP-9 has evolved over the last decade and will provide insight into future directions that will drive our understanding of MMP-9-directed cardiac ECM turnover in the post-MI LV.

The extracellular matrix compartment of neural stem and glial progenitor cells.

Neuroepithelial and radial GLIA stem cells generate the majority of the cellular constituents of the central nervous system. Following precisely timed phases of neurogenesis and gliogenesis the stem cells recede, with the exception of adult neural stem cells that persist in two generally accepted canonical neurogenic regions, the subventricular zone of the lateral ventricle and the subgranular zone in the dentate gyrus of the hippocampus. It is believed that adult stem cells reside in privileged stem cell niche environments that provide favorable conditions for self-renewal and maintenance of this cellular compartment. Factors such as morphogens, cytokines, and growth factors influence the developmental pathway of neural stem/progenitor cells. By comparison, less is known about the regulatory roles of glycoproteins and proteoglycans of the extracellular matrix (ECM) and their receptors, although they represent important constituents of the micromolecular environment of the niche. Here, we summarize studies that indicate pivotal roles of the ECM micromilieu for the biology and instrumental use of glial stem and progenitor cells of the CNS. Advancing our understanding of structure-function relationships, signaling motifs and complementary receptors and their signal transduction pathways will be of central importance for the application of these cell types in regenerative medicine.