Breast cancer is the most frequently diagnosed cancer in women worldwide. Although dramatically increased survival rates of early diagnosed cases have been observed, late diagnosed patients and metastatic cancer may still be considered fatal. The present study's main focus was on cancer‐associated fibroblasts (CAFs) which is an active component of the tumor microenvironment (TME) regulating the breast cancer ecosystem. Transcriptomic profiling and analysis of CAFs isolated from breast cancer skin metastasis, cutaneous basal cell carcinoma, and squamous cell carcinoma unravelled major gene candidates such as IL6, VEGFA and MFGE8 that induced co‐expression of keratins‐8/‐14 in the EM‐G3 cell line derived from infiltrating ductal breast carcinoma. Western blot analysis of selected keratins (keratin‐8, ‐14, ‐18, ‐19) and epithelial‐mesenchymal transition‐associated markers (SLUG, SNAIL, ZEB1, E‐/N‐cadherin, vimentin) revealed specific responses pointing to certain heterogeneity of the studied CAF populations. Experimental in vitro treatment using neutralizing antibodies against IL-6, VEGF‐A and MFGE8 attenuated the modulatory effect of CAFs on EM‐G3 cells. The present study provided novel data in characterizing and understanding the interactions between CAFs and EM‐G3 cells in vitro. CAFs of different origins support the pro‐inflammatory microenvironment and influence the biology of breast cancer cells. This observation potentially holds significant interest for the development of novel, clinically relevant approaches targeting the TME in breast cancer. Furthermore, its implications extend beyond breast cancer and have the potential to impact a wide range of other cancer types.
- MeSH
- Antigens, Surface MeSH
- Cancer-Associated Fibroblasts * metabolism MeSH
- Fibroblasts metabolism MeSH
- Keratins genetics metabolism MeSH
- Humans MeSH
- Melanoma, Cutaneous Malignant MeSH
- MCF-7 Cells MeSH
- Milk Proteins genetics metabolism MeSH
- Cell Line, Tumor MeSH
- Tumor Microenvironment genetics MeSH
- Breast Neoplasms * drug therapy genetics metabolism MeSH
- Prognosis MeSH
- Transcriptome MeSH
- Check Tag
- Humans MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
Pericardial fluid (PF) has been suggested as a reservoir of molecular targets that can be modulated for efficient repair after myocardial infarction (MI). Here, we set out to address the content of this biofluid after MI, namely in terms of microRNAs (miRs) that are important modulators of the cardiac pathological response. PF was collected during coronary artery bypass grafting (CABG) from two MI cohorts, patients with non-ST-segment elevation MI (NSTEMI) and patients with ST-segment elevation MI (STEMI), and a control group composed of patients with stable angina and without previous history of MI. The PF miR content was analyzed by small RNA sequencing, and its biological effect was assessed on human cardiac fibroblasts. PF accumulates fibrotic and inflammatory molecules in STEMI patients, namely causing the soluble suppression of tumorigenicity 2 (ST-2), which inversely correlates with the left ventricle ejection fraction. Although the PF of the three patient groups induce similar levels of fibroblast-to-myofibroblast activation in vitro, RNA sequencing revealed that PF from STEMI patients is particularly enriched not only in pro-fibrotic miRs but also anti-fibrotic miRs. Among those, miR-22-3p was herein found to inhibit TGF-β-induced human cardiac fibroblast activation in vitro. PF constitutes an attractive source for screening diagnostic/prognostic miRs and for unveiling novel therapeutic targets in cardiac fibrosis.
- MeSH
- Fibroblasts metabolism MeSH
- Fibrosis * MeSH
- ST Elevation Myocardial Infarction metabolism pathology genetics MeSH
- Myocardial Infarction * metabolism genetics pathology MeSH
- Interleukin-1 Receptor-Like 1 Protein metabolism genetics MeSH
- Middle Aged MeSH
- Humans MeSH
- MicroRNAs * genetics metabolism MeSH
- Myocardium metabolism pathology MeSH
- Pericardial Fluid * metabolism MeSH
- Aged MeSH
- Transforming Growth Factor beta metabolism MeSH
- Check Tag
- Middle Aged MeSH
- Humans MeSH
- Male MeSH
- Aged MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
Crohn's disease (CD) is marked by recurring intestinal inflammation and tissue injury, often resulting in fibrostenosis and bowel obstruction, necessitating surgical intervention with high recurrence rates. To elucidate the mechanisms underlying fibrostenosis in CD, we analyzed the transcriptome of cells isolated from the transmural ileum of patients with CD, including a trio of lesions from each patient: non-affected, inflamed, and stenotic ileum samples, and compared them with samples from patients without CD. Our computational analysis revealed that profibrotic signals from a subset of monocyte-derived cells expressing CD150 induced a disease-specific fibroblast population, resulting in chronic inflammation and tissue fibrosis. The transcription factor TWIST1 was identified as a key modulator of fibroblast activation and extracellular matrix (ECM) deposition. Genetic and pharmacological inhibition of TWIST1 prevents fibroblast activation, reducing ECM production and collagen deposition. Our findings suggest that the myeloid-stromal axis may offer a promising therapeutic target to prevent fibrostenosis in CD.
- MeSH
- Crohn Disease * metabolism pathology immunology MeSH
- Adult MeSH
- Endopeptidases metabolism genetics MeSH
- Extracellular Matrix metabolism pathology MeSH
- Fibroblasts * metabolism pathology MeSH
- Fibrosis * MeSH
- Ileum pathology metabolism immunology MeSH
- Nuclear Proteins metabolism genetics MeSH
- Humans MeSH
- Cell Communication MeSH
- Monocytes * metabolism pathology immunology MeSH
- Mice MeSH
- Receptors, Cell Surface metabolism genetics MeSH
- Twist-Related Protein 1 * metabolism genetics MeSH
- Animals MeSH
- Check Tag
- Adult MeSH
- Humans MeSH
- Male MeSH
- Mice MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Pulmonary hypertension is a cardiovascular disease with a low survival rate. The protein galectin-3 (Gal-3) binding β-galactosides of cellular glycoproteins plays an important role in the onset and development of this disease. Carbohydrate-based drugs that target Gal-3 represent a new therapeutic strategy in the treatment of pulmonary hypertension. Here, we present the synthesis of novel hydrophilic glycopolymer inhibitors of Gal-3 based on a polyoxazoline chain decorated with carbohydrate ligands. Biolayer interferometry revealed a high binding affinity of these glycopolymers to Gal-3 in the subnanomolar range. In the cell cultures of cardiac fibroblasts and pulmonary artery smooth muscle cells, the most potent glycopolymer 18 (Lac-high) caused a decrease in the expression of markers of tissue remodeling in pulmonary hypertension. The glycopolymers were shown to penetrate into the cells. In a biodistribution and pharmacokinetics study in rats, the glycopolymers accumulated in heart and lung tissues, which are most affected by pulmonary hypertension.
- MeSH
- Pulmonary Artery drug effects metabolism MeSH
- Biomarkers MeSH
- Fibroblasts drug effects metabolism MeSH
- Galectin 3 * antagonists & inhibitors metabolism MeSH
- Rats MeSH
- Cells, Cultured MeSH
- Humans MeSH
- Hypertension, Pulmonary * drug therapy metabolism MeSH
- Polymers chemistry pharmacology MeSH
- Tissue Distribution MeSH
- Animals MeSH
- Check Tag
- Rats MeSH
- Humans MeSH
- Male MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Hyperthermia along with hydrocortisone (HC) are proven teratogens that can negatively influence embryo development during early pregnancy. Proliferation of cells is one of the main developmental processes during the early embryogenesis. This study was focused on testing the effect of elevated temperature and HC addition on proliferation of cells in in vitro cultures. The V79-4 cell line was treated with HC and cultured in vitro at 37 °C or 39 °C, respectively. To reveal the effect of both factors, the proliferation of cells cultured under different conditions was evaluated using various approaches (colony formation assay, generation of growth curves, computation of doubling times, and mitotic index estimation). Our results indicate that a short-term exposure to elevated temperature slightly stimulates and a long-term exposure suppresses cell proliferation. However, HC (0.1 mg/ml) acts as a stimulator of cell proliferation. Interestingly, the interaction of HC and long-term elevated temperature (39 °C) exposure results in at least partial compensation of the negative impact of elevated temperature by HC addition and in higher proliferation if compared with cells cultured at 39 °C without addition of HC.
- MeSH
- Cricetulus MeSH
- Fibroblasts * drug effects cytology metabolism MeSH
- Hydrocortisone * pharmacology MeSH
- Cells, Cultured MeSH
- Cell Proliferation * drug effects MeSH
- Temperature MeSH
- Hot Temperature MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
Natural products have many healing effects on the skin with minimal or no adverse effects. In this study, we analyzed the regenerative properties of a waste product (hydrolate) derived from Helichrysum italicum (HH) on scratch-tested skin cell populations seeded on a fluidic culture system. Helichrysum italicum has always been recognized in the traditional medicine of Mediterranean countries for its wide pharmacological activities. We recreated skin physiology with a bioreactor that mimics skin stem cell (SSCs) and fibroblast (HFF1) communication as in vivo skin layers. Dynamic culture models represent an essential instrument for recreating and preserving the complex multicellular organization and interactions of the cellular microenvironment. Both cell types were exposed to two different concentrations of HH after the scratch assay and were compared to untreated control cells. Collagen is the constituent of many wound care products that act directly on the damaged wound environment. We analyzed the role played by HH in stimulating collagen production during tissue repair, both in static and dynamic culture conditions, by a confocal microscopic analysis. In addition, we performed a gene expression analysis that revealed the activation of a molecular program of stemness in treated skin stem cells. Altogether, our results indicate a future translational application of this natural extract to support skin regeneration and define a new protocol to recreate a dynamic process of healing.
- MeSH
- Fibroblasts metabolism drug effects MeSH
- Helichrysum * chemistry MeSH
- Wound Healing * drug effects MeSH
- Stem Cells metabolism drug effects cytology MeSH
- Collagen * metabolism MeSH
- Cells, Cultured MeSH
- Skin * metabolism drug effects MeSH
- Humans MeSH
- Regeneration * drug effects MeSH
- Plant Extracts * pharmacology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
ANTXR1 is one of two cell surface receptors mediating the uptake of the anthrax toxin into cells. Despite substantial research on its role in anthrax poisoning and a proposed function as a collagen receptor, ANTXR1's physiological functions remain largely undefined. Pathogenic variants in ANTXR1 lead to the rare GAPO syndrome, named for its four primary features: Growth retardation, Alopecia, Pseudoanodontia, and Optic atrophy. The disease is also associated with a complex range of other phenotypes impacting the cardiovascular, skeletal, pulmonary and nervous systems. Aberrant accumulation of extracellular matrix components and fibrosis are considered to be crucial components in the pathogenesis of GAPO syndrome, contributing to the shortened life expectancy of affected individuals. Nonetheless, the specific mechanisms connecting ANTXR1 deficiency to the clinical manifestations of GAPO syndrome are largely unexplored. In this study, we present evidence that ANTXR1 deficiency initiates a senescent phenotype in human fibroblasts, correlating with defects in nuclear architecture and actin dynamics. We provide novel insights into ANTXR1's physiological functions and propose GAPO syndrome to be reconsidered as a progeroid disorder highlighting an unexpected role for an integrin-like extracellular matrix receptor in human aging.
- MeSH
- Actins metabolism MeSH
- Alopecia * metabolism pathology genetics MeSH
- Anodontia * MeSH
- Optic Atrophies, Hereditary genetics metabolism MeSH
- Fibroblasts * metabolism MeSH
- Humans MeSH
- Microfilament Proteins * MeSH
- Growth Disorders * MeSH
- Progeria genetics pathology metabolism MeSH
- Receptors, Cell Surface metabolism genetics deficiency MeSH
- Cellular Senescence * genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Fibroblasts are an integral cell type of mammary gland stroma, which plays crucial roles in development, homeostasis, and tumorigenesis of mammary epithelium. Fibroblasts produce and remodel extracellular matrix proteins and secrete a plethora of paracrine signals, which instruct both epithelial and other stromal cells of the mammary gland through mechanisms, which have not been fully understood. To enable deciphering of the intricate fibroblast-epithelial interactions, we developed several 3D co-culture methods. In this chapter, we describe methods for establishment of various types of embedded 3D co-cultures of mammary fibroblasts with mammary epithelial organoids, mammary tumor organoids, or breast cancer spheroids to investigate the role of fibroblasts in mammary epithelial development, morphogenesis, and tumorigenesis. The co-culture types include dispersed, aggregated, and transwell cultures.
- MeSH
- Cell Line MeSH
- Epithelium metabolism MeSH
- Epithelial Cells * MeSH
- Fibroblasts metabolism MeSH
- Carcinogenesis pathology MeSH
- Coculture Techniques MeSH
- Humans MeSH
- Mammary Glands, Animal * MeSH
- Organoids MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
Epithelial branching morphogenesis is an essential process in living organisms, through which organ-specific epithelial shapes are created. Interactions between epithelial cells and their stromal microenvironment instruct branching morphogenesis but remain incompletely understood. Here, we employed fibroblast-organoid or fibroblast-spheroid co-culture systems and time-lapse imaging to reveal that physical contact between fibroblasts and epithelial cells and fibroblast contractility are required to induce mammary epithelial branching. Pharmacological inhibition of ROCK or non-muscle myosin II, or fibroblast-specific knock-out of Myh9 abrogate fibroblast-induced epithelial branching. The process of fibroblast-induced branching requires epithelial proliferation and is associated with distinctive epithelial patterning of yes associated protein (YAP) activity along organoid branches, which is dependent on fibroblast contractility. Moreover, we provide evidence for the in vivo existence of contractile fibroblasts specifically surrounding terminal end buds (TEBs) of pubertal murine mammary glands, advocating for an important role of fibroblast contractility in branching in vivo. Together, we identify fibroblast contractility as a novel stromal factor driving mammary epithelial morphogenesis. Our study contributes to comprehensive understanding of overlapping but divergent employment of mechanically active fibroblasts in developmental versus tumorigenic programs.
- MeSH
- Epithelial Cells * metabolism MeSH
- Fibroblasts metabolism MeSH
- Coculture Techniques MeSH
- Mammary Glands, Animal * metabolism MeSH
- Morphogenesis physiology MeSH
- Mice MeSH
- Animals MeSH
- Check Tag
- Mice MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
AIMS: The effect of polyphenolic fraction of Lonicera caerulea (PFLC) and alkaloid fraction of Macleaya cordata (AFMC) mix on the production of inflammatory mediators in human gingival fibroblasts pretreated with lipopolysaccharide (LPS) was investigated. In addition, protective effects of mucoadhesive paste containing combination of PFLC and AFMC (0.05% and 0.01%, respectively; n=15, Group A) and placebo (n=15, Group B) were evaluated in patients after surgical extraction of lower third molars. METHODS: Gingival fibroblasts were pre-treated with LPS (10 μg/mL; 24 h) and PFLC/AFMC (25/0.25; 50/0.25; 100/0.25; 25/0.5; 50/0.5; 100/0.5 μg/mL) in serum-free medium was applied for 4 h. Then the interleukin-6 (IL-6), reactive oxygen species (ROS) generation, level of intracellular glutathione (GSH) and expression of cyclooxygenase-2 (COX-2) were evaluated. The study was a 6-day, single-center, randomized, double-blind and placebo-controlled trial consisting of two parallel treatment arms. A modified Oral health impact profile questionnaire including both general oral condition and extraction related questions, was used to evaluate the oral condition and other changes before (day 0) and on the days 1, 3 and 6 after surgical extraction. RESULTS AND CONCLUSION: The combination of PFLC with AFMC caused a reduction of ROS generation, reduced IL-6 production and suppressed the expression of COX-2. In group A the paste treatment contributed to improvement of oral health-related quality of life. Topical application of PFLC and AFMC into the extraction wound improved post-extraction site wound healing probably by antioxidant and anti-inflammatory mechanisms.
- MeSH
- Alkaloids * pharmacology MeSH
- Cyclooxygenase 2 pharmacology MeSH
- Phenols pharmacology MeSH
- Fibroblasts metabolism MeSH
- Wound Healing MeSH
- Interleukin-6 MeSH
- Quality of Life MeSH
- Humans MeSH
- Lipopolysaccharides pharmacology MeSH
- Molar, Third * surgery metabolism MeSH
- Reactive Oxygen Species metabolism pharmacology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Randomized Controlled Trial MeSH
