What are the phases of the cell cycle?
- 1) G0 (resting)
- 2) G1 (presynthetic)
- 3) S (DNA synthesis)
- 4) G2 (premitotic)
- 5) M (mitotic)
What is meant by assymetric replication of stem cells?
One cell is self-renewing and the other differentiates after division.
Name the GFs/cytokines involved in differentiation of specific cell lineages:
- Endo - VEGF, FGF
- Chondroblasts - Sox9
- Osteoblasts - CBFA1
- Fat - PPARg
- Muscle - Myo D, myogenin
Name the three germ layers and what they develop into.
- Endoderm - epith cells of lung, liver, GI
- Mesoderm - comprise both mesodermal and BM progenitor cells
- mesodermal progenitors develop into myocytes, osteoblasts, chondrocytes, adipocytes, endoth cells
- BM prog diff into hematopoietic cells but can also generate cells for tissues of endo and ectoderm origin
- Ectoderm - keratinocyte precursores, neurons, oligodendricytes, ependymal cells
What is the source and function of EGF?
- Source: Plts, macs, saliva, urine, milk, plasma
- Fxn: mitogenic for keratinocytes, stim migration and granulation tissue formation
What is the source and function of TGF-a?
- Source: Macs, T lymphs, keratinocytes, many tissues
- Fxn: similar to EGF, stim hepatic replication and some epi cells
What is the source and function of HGF?
- Also called scatter factor.
- Source: mesenchymal cells
- Function: prolif of epi cells and hepatocytes, increases cell motility
What is the source and function of VEGF?
- Source: Mesench cells
- Fxn: Incr vascular perm, mitogenic for endoth cells
What is the source and function of PDGF?
- Source: plts, macs, endo, keratinocytes, smooth mm
- 1) chemotactic for PMNs, macs, fibroblats, smooth mm cells
- 2) activates PMNs, macs, fibroblasts
- 3) mitogenic for fibroblasts, endoth, sm mm
- 4) stim prod of MMPs, fibronectin, and HA
- 5) stim angiogenesis, wound contraction
- 6) inh plt ag
- 7) regulates integrin expression
What is the source and function of FGF?
- Source: Macs, mast cells, T cells, endo cells, fibroblasts, many
- 1) chemotactic for fibroblasts
- 2) mitogenic for fibroblasts and keratinocytes
- 3) stim keratinocyte migration, angiogenesis, wound contraction and matric deposition
What is the source and function of TGF-b?
- Source: plts, T cells, macs, endo, keratinocytes, sm mm, fibro
- 1) chemotactic for PMNs, macs, lymphs, fibroblasts and sm mm
- 2) stim TIMP synthesis, keratinocyte migration, angiogenesis and fibroplasia
- 3) inh MMP prod and keratinocyte prolif
- 4) regulates integrin expression and other cytokines
- 5) induces TGF-b production
What is the source and function of KGF?
- Source: fibroblasts
- Fxn: stim keratinocyte migration, prolif, and diff
What is the source and function of IGF-1?
- Source: Macs, fibro, other
- 1) synthesis of sulfated proteoglycans, collagen, keratinocyte migration, fibroblast prolif
- 2) GH-like endocrine effects
What is the source and function of TNF?
- Source: macs, mast cell, Tcells
- Fxn: activates macs, regulates other cytokines
What is the source and function of ILs?
- Source: macs, mast cells, keratinocytes, lymphs, many tissues
- Fxn: many - ex.
- 1) IL1 - chemotactic for PMNs and stim MMP-1 synthesis
- 2) IL4 - chemotactic for fibroblasts
- 3) IL8 - angiogenesis
- 4) IL6 - TIMP synthesis
- 5) Regulates other cytokines
What is the source and function of IFNs?
- Source: lymphs and fibroblasts
- 1) activates macs
- 2) inh fibroblast prolif and synthesis of MMPs
- 3) cytokine regulation
Hints for the cytokines involved in tissue repair.
- VEGF and HGF - only prod by mes cells
- Macs produce them all except - VEGF, HGF, KGF and IFNs
- KGF - only prod by fibroblasts
- T Lymphs produce - TGF-a, FGF, TGF-b, TNF, ILs (T and B), IFNs (T and B)
- Plts produce: EGF, PDGF, TGF-b
Name the four types of receptors and signal transduction pathways and explain how they work. (look at figures)
- 1) Receptors with intrinsic tyrosine kinase activity: most growth factors - ex. EGF, VEGF, PDGF....
- - binding of ligand --- dimerization of receptor --- tyrosine phosphorylation --- activate tyrosine kinase
- - activates downstream effector molecules
- - acts as dock for other effector molecules (PI3K- activates akt - cell prol and inh apop, PLC-g - breaks down membrane PL into IP3 which incr. Ca and DAG which activate transcr factors, Src)
- - phosphorylates GRB2-SOS bridging protein - activates RAS - then RAF - then MEK - then ERK - synthesis and phosphorylation of c-jun and c-fos - prod of GF controling entry of cell into cycle
- 2) Receptors that recruit kinases - cytokines (ex. IL2, IL3), IFNs, CSF, GH, prolactin
- - transmit extra cell signals to nucleus via JAK STAT pathway
- 3) Seven TM G-coupled receptors - serotonin, hist, vasopressin, epi, norepi, calcitonin, gluc, PTH, ACTH
- - binding of ligand - conformational change - activation of G prot by exchange of GDP with GTP
- - also produces IP3 which incr. Ca release from ER (like TK receptors)
- 4) Steroid hormone receptors
- - ligands diffuse through cell membrane and bine to nuclear receptors
List the phases of the cell cycle. What is the rate limiting step? What controls progression through the cell cycle?
- G1-S is rate limiting (point of no return) - checks DNA before replication
- Cyclins and cyclin dependent kinases control progression
- G2/M checks DNA after replication
What is the role of RB protein in the cell cycle
Normally bound to E2F transcription factor but phos of RB releases it and E2F is activated and stimulates transcription
What triggers tissue regeneration (liver)? Describe the gene expression phases.
- Cytokines and GFs trigger cells to enter cell cycle. The two major restriction points are G0-G1 and G1-S.
- Early gene response --- cFOS and cJUN dimerize to form tf AP1, cMYC and other tfs such as NFkB, STAT3 and C-EBP
What primes hepatocytes to enter cell cycle during liver regeneration? What acts on primed hepatocytes to stim progression through cell cycle? What inhibits continued growth and prolif?
- Priming - IL6, TNF, others
- Proliferation - HGF, TGFa, others
- - adjuvants are: norepi, insulin, GF, T4
- Growth inhibitors: TGF-b, activin, others - basically get incr in cell cycle inh and decr in GF and/or metabolic demands of the liver
Name the three types of macromolecules in the ECM?
- 1) structural proteins (collagens, elastins)
- 2) adhesive glycoprot
- 3) proteoglycans and hyaluronic acid
Name the components of the basement membrane.
collagen IV, laminin, heparan sulfate, proteoglycan, other glycoproteins
Name the types of collagen and their tissue distribution.
- I - hard and soft tissue (defects - osteogenesis imperf and ehlers-danlos syndrome -arth type)
- II - cartilage, IV disk, vitreous
- III - hollow organs, soft tissues (ED - vascular)
- IV - BM (non-fibrillar - present in sheets)
- V - soft tissues, BV (classical ED)
- VI - microfibrils
- VII - anchoring fibrisl at dermal-epi jxn (dystrophic epidermolysis bullosa)
- IX - cartilage, vitreous (stickler syndrome)
- XVII - transmembrane collagen in epi cells (epidermolysis bullosa - benign atrophic)
- XV and XVIII - endostatin-forming collagens, endoth cells (knoblock syn)
How is collagen formed?
- 1) procollagen transcribed by collagen genes and hydroxylated to form a triple helix (Vit c needed)
- 2) procollagen secreted from cell and cleaved by proteases
- 3) fibrils formed by extracellular lysyl oxidase
- 4) fibrils are then cross-linked
What are the 4 main families of the cell adhesion molecules (CAMs)? What are their functions?
- 1) Ig family
- 2) Cadherins - Ca dep adhesion prot
- 3) integrins
- 4) Selectins
- Functions - cell-to-cell adhesion, adhesion of cell to ECM, integrins and cadherins link cell surface with to cytoskeleton by binding to actin and intermed filaments - signal transduction, motility
How are cadherins linked to the cytoskeleton?
b-catenin links cadherin to a-catenin which connects to actin -
Name the 7 phases of healing.
- 1) Inflammation
- 2) Prol and migration of CT cells
- 3) Angiogenesis
- 4) Synthesis of ECM proteins
- 5) Tissue remodeling
- 6) Wound contraction
- 7) Wound stregth
Name the steps involved in angiogenesis from pre-existing vessels.
- 1) Vasodilation d/t NO and incr. perm d/t VEGF
- 2) Degredation of BM and break cell to cell contact between endo
- 3) Migration of endo cells to angiogenic stim
- 4) Proliferation of endo cells behind leading edge
- 5) Recruit periendo cells for support - ex pericytes, sm mm
Name inducing agents for VEGF.
Hypoxia, TGF-a, TGF-b, PDGF
Receptors for VEGF and where located?
- VEGFR1, 2 and 3
- 2 - only on endo cells, binding of receptor stim mobilization of precursors from BM
- 3 - only on lymphatic endo cells
Functions of VEGF
- vascular perm
- endo migration and prol
- hyperplasia of lymphatics
- upreg expression of plasminogen activator, plasminogen activator inhibitor-1, TF, interstitial collagenase
- NOTE: Effects enhanced by FGF-2
What are the actions fo angiopoietins, PDGF and TGF-b in angiogenesis?
- Ang1 interacts with Tie2 (on endo) to recruits periendoth cells and matures vessels
- Ang2/Tie2 has oposite effect and loosens endo cells so more responsive to VEGF (if present) or angiogenic inhib if VEGF absent
- PDGF - recruits sm mm
- TGF-b - incr prod of ECM proteins to stabalize vessels
What stimulates fibroblast migration and proliferation?
- TGF-b, FGF, EGF, PDGF, IL1 and TNF
- TGF-b most NB GF in inflammatory fibrosis
What is the most NB GF in inflammatory fibrosis and what are its actions?
- 1) migration and proliferation of fibroblasts
- 2) incr synthesis of collagen/fibronectin
- 3) decr degredation of ECM
- 4) chemotactic for monos
What GF stimulates collagen synthesis?
TGF-b, PDGF, FGF, IL1, IL13
List some of the critical steps/processes in wound healing and the GF and cytokines involved.
- Mono chemotaxis: PDGF, FGF, TGF-b
- Fibroblast migration: PDGF, FGF, TGF-b, IL1, TNF
- Fibroblast prol: PDGF, FGF, EGF, TNF
- Angiogenesis: VEGF, Ang, FGF
- Collagen synthesis: PDGF, FGF, IL1, IL13, TGF-b
- Collagenase secretion: PDGF, FGF, EGF, TNF, TGF-b (inhibits)
- NOTE: FGF (all steps), PDGF (all except angiogenesis), ILs in coll syn and fib migration, TGF-b inh. collagenase
Name the factors that retard wound healing.
- Local: bld supply, denervation, infection, FB, hematoma, mech stress, necrosis, dressings, tissue type
- Systemic: age, anemia, drugs, DM, neoplasia, genetic dz, malnut, obesity, infection, temp, uremia, mineral def, hormones
Name the three phases of cutaneous wound healing.
- 1) Inflammation
- 2) Granulation tissue and reepithelialization
- 3) Wound contraction, ECM depo and remodeling
Detail the stages involved in first intention wound healing.
- 1) 1st 24-48 hrs: neuts at margin, fibrin clot formation, at 24-48 hrs epis move from edges and deposit BM - fuse at midline under scab
- 2) day 3: neuts replaced by macs, gran tissue, collagen at margins vertically oriented, epi layer thickens
- 3) day 5: fills with gran tissue, neovasc, collagen more numerous and bridge
- 4) 2 wk: accum of collagen and prol of fibrobasts, WBCs and edema gone, scar formation, new vessels disappear
- 5) 30d - scar made of cellular CT
How does 2nd intention healing differ from 1st?
- 1) larger clot, more inflammation
- 2) more gran tissue
- 3) wound contraction occurs
- 4) more scarring
What is the progression of wound strength over time?
approx. 10% at 1 wk, then incr to about 70-80% over 3 mos