Pathological signaling pathways in fibrosis Flashcards
Fibrosis
The excessive formation of connective tissue. Cells will behave differently in fibrotic tissue compared to healthy tissue. There will be less cells and the connective tissue takes over.
Wound healing in general
◦ Blood clotting: follows the coagulation cascade (hemostasis)
◦ Inflammation: inflammatory cells like neutrophils and macrophages neutralize the injury and material at the wound site
◦ Proliferation: granulation tissue is the foundation for regeneration
◦ Maturation: collagen fibers are deposited (by fibroblasts and myofibroblasts) to achieve remodeling and normal functioning of the tissue will be realized.
Wound healing when a biomaterial is implanted
injury – blood-material interaction – matrix formation – acute inflammation – chronic inflammation – granulation tissue formation – foreign body reactions – fibrosis. Due to the biomaterial, a chronic inflammation prolongs and almost always will a fibrotic layer forms around the biomaterial.
What four main steps can be identified in the response to healing?
- Physical response. Vasoconstriction and attraction of white blood cells initiates the inflammatory response. Inflammatory factors are release by damages cells (recruit cells).
- Coagulation cascade
- Inflammation. Inflammation is important to neutralize bacterial agents and initiate healing stages. Neutrophils (non-specific) are degraded by macrophages. Macrophages contribute to the proliferation stage by excretion of fibroblasts growth factors (FGFs). Fibroblasts will enter the wound site and induce:
◦ Angiogenesis
◦ Differentiation into myofibroblasts for wound contraction
◦ Excretion of ground substance and collagen fibers. This will form granulation tissue. The tissue can now be epithelialized.
At a certain point the granulation tissue ceases, the amount of fibroblasts decrease and the results will be a collagen rich tissue that can start to remodel/mature - The amount of collagen is constant but type III is replaced by type I and rearranged. The tensile strength of the wound increases over time and blood vessels and redness disappears. If the healing happens inappropriate, it can result in a chronic wound or (pathologic) scar formation.
Cytokines
Chemicals that are secreted by the immune system and signal other cells
Name 3 signaling pathways that are involved in fibrosis.
◦ TGF-β/Smad pathway = TGF-β is a ligand to an enzyme-coupled receptor (serine/threonine kinase receptor) which phosphorylates SMAD proteins
◦ Wnt/β-catenin pathway = Wnt is a ligand of the GPCR Frizzled. β-catenin is a transcription factor which can only bind to DNA when Wnt binds to its receptor.
◦ Yap/Taz pathway = YAP and TAZ are proteins that are involved in the Hippo pathway. Highly linked to the other two pathways.
These three pathways are connected and cross talk between these pathways leads to a complex cell signature for fibrosis. For example Wnt signaling can induce expression of TGF-β and TGF-β signaling can induce expression of Wnt/β-catenin.
Myofibroblasts
Myofibroblasts are cells with a phenotype in between fibroblasts and mooth muscle cells.
They:
◦ Can differentiate from a lot of different cell types.
◦ Are activated as a response to tissue injury
◦ Can sense, exert and withstand high stresses due to super mature focal adhesions (FAs)
◦ Able to contract (phenotype of fibroblasts and SMC) -> wound healing
◦ Able to excrete collagen -> stiffening of ECM
◦ Markers: α-smooth muscle actin (α-SMA), ED-A fibronectin (ED-A FN) and more?
◦ Functions
◦ Repair, but don’t regenerate defects (-> reduced tissue function and organ failure)
◦ Develop adhesion structures
◦ Formation of contractile bundles (-> remodel and contract ECM and adapt activity)
Differentiation into myofibroblasts
Myofibroblasts can differentiate from lots of different celltypes. First a cell wil differentiate into proto-myofibroblasts and later into fibroblasts. The main pathway to the differetiation of myofibroblast is though the tgf-beta pathway in fibroblasts.
The differentiation into myofibroblasts causes excessive ECM de position, which in turn causes more differtentiation into myofibroblasts.
TGF-β pathway, beginning
Normally, TGF-β1 is trapped in a large latent complex (TGF-β, LAP and LTBPs) in the ECM. Upon injury, mechanical stress or stiffness, TGF-β is released from this complex and can engage with its receptors.
SMAD pathway
When TGF-β is bound to its receptor. This causes the receptor to phosphorylate SMAD proteins. 2 R-SMADs and 1 co-SMAD can form one transcriptional factor which can be transported to the nucleus to bind to DNA promotor domains, which aid transcription. Inhibatory SMADs(I-SMADs) can prevent or disrupt binding or repress transcription. The degradation of SMADs happens through proteasomal degradation and is controlled through ubiquiting ligase(Smurf2) and I-SMADS.
SMAD protein
SMAD protein are the main signal transducers for the TGF-β pathway. There are 3 types:
- Regulatory (R-) SMADs (1,2,3,5,8)
- Co-stimulatory (Co-) SMADs (4)
- Inhibitory (I-) SMADs (6,7)
R/Co-SMADs are located in the cytoplasm but accumulate in the nucleus following TGF-β signaling.
What are the effects of the TGF- β pathway?
- Cell growth, differentiation, migration, apoptosis, immunity, embryonic development
- Fibrogenesis: excessive ECM production, activation of myofibroblasts, inhibition of ECM degradation
- Orientation, differentiation, development and proliferation of osteoblasts
Wnt/β-catenin pathway
Wnt is a ligand of the Frizzled receptor. When Wnt is not bound to Frizzled, a destruction complex inside the cell destructs β-catenin. When Wnt binds to Frizzled, that destruction complex is disassembled. β-catenin can now translocate to the nucleus and stimulates certain genes (transcription), for example fibrosis-related genes like FN. TGF-β recruits proteins that stabilize β-catenin in the cytoplasm (cannot be destructed and can translocate to the nucleus).
The Wnt/β-catenin signaling is important in fibrosis (induces transcription of fibrosis-related genes).
A Wnt inhibitor (e.g. DKK) -> decreased fibrosis (collagen deposition). Other inhibitors of the Wnt/β-catenin pathway also prevent, decrease and reverse fibrosis.
YAP/TAZ pathway
Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) are both proteins: transcriptional effects in the Hippo pathway. When YAP/TAZ are translocated to the nucleus, they bind to transcription factors and stimulate transcription. This induces proliferation and stiffening of the ECM. (fibroblast activation and fibrosis). YAP/TAZ are mechano-activated coordinators of the matrix-driven feedback loop that amplifies and sustains fibrosis.
YAP/TAZ localized in the nucleus and transcriptionally active
◦ Proliferation
◦ Osteoblasts differentiation (from MSCs)
◦ Large adhesive areas
◦ Stiff ECM
YAP/TAZ localized in the cytoplasm and are degraded (no transcription) ◦ Growth arrest, apoptosis ◦ Adipocyte differentiation (from MSCs) ◦ Small adhesive areas ◦ Soft ECM
