mucosal homeostasis in the lungs Flashcards
Describe homeostasis in the alveolar space
the alveoli is a very fragile structure
the ends of the alveoli are made of 1 cell- type 1 or type 2 pneumocyte
the lungs have evolved to have site specific immune cells (AM) alveolar macrophages
they are on the luminal side of the lung, in contact with the atmosphere
they are there to survey, eliminate particles by phagocytosis and initiate an inflammatory response in the presence of a pathogen.
What are some general immune cell interactions in the healthy lung
The lung has specialised
tissue-resident immune
cells
Dendritic cells
Alveolar macrophages
Regulatory T cells
airways composed by barrier made of alveolar epithelial cells (the brick like cells)
thin layer of mucous (yellow outline)
specific microbiota as in all mucosal (green)
goblet cells produce mucous (shown in blue, dispersed in between epithelial cells)
cells are highly specialised to tissue- some are highly migratory such as dendritic cells which are depicted in purple. These are constantly travelling through the parenchyma (the light brown area or insides of the lung)
this helps to sample the environment- healthy surveillance of microbiota and any potential antigens from pathogens. This is what keeps tolerance in our lungs. If the dendritic cells encounter a pathogen, they are ready to capture it and activate T cells
in orange, we have highly specialised, tissue specific alveolar macrophages- they have a particular phenotype adapted to the lung
they have close interaction with epithelium through negative regulators- CD200, Il-10, TGF B
T cells can interact w macrophages via retinoic acid and regulatory T cells via prostaglandins
overall the healthy mucosal immune cells is where negative regulation is predominant
Describe alveolar macrophages
CD11c ++ SiglecF+ F4/80 int
Resident phagocytic patrolling cells
Pro and anti inflammatory
Remove apoptotic cells by efferocytosis
Induce regulatory T cells via retinoic acid
Mixed functional phenotype – highly phagocytic
and capable of producing TNF-⍺ and IL-1 but also
responsive to IL-4 and able to drive type 2 inflammation
as well as potentially wound healing – a response that is site specific
Important sentinels in asthma – depletion exacerbates disease
it is unique- highly specialised macrophage
evolved to develop characteristics adapted to the lung
identified using flow cytometry- high expression of CD11c, sigle cd, F4/80
phagocytic, survey and patrols tissue
has both pro and anti inflamatory phenotypes
phentoype of repair and wound healing m2- anti inflammatory and pro balance
can produce tnf alpha and IL 1- pro-inflammatory m1
can induce regulatory t cells via retinoic acid
type 2 inflammation is regulated by the lung microenvironment, macrophages are less responsive to IL4 (response against parasites). they become more responsible ti IL4 OUT of the lungs, showing the environment of the lungs exerts regulatory capacity
if you deplete these macrophages in mice you make asthma worse
Ontogeny and turnover of alveolar macrophages
origin of alveolar macrophages and how they replenish themselves/ change in numbers due to infection/chronic lung conditions:
steady state:
alveolar macrophages originate from foetal liver myeloid precursors in development
they differentiate and acquire their functions due to GM CSF (growth factor for myeloid cells)this helps them to proliferate and acquire characteristics
so if there is a depletion of alveolar macrophages eg due to toxin exposure, the original population can be recovered through in situ proliferation due to pre-existing alveolar monocytes
the lung itself can proliferate alveolar macrophages due to GM-CSF. In animals lacking this growth factor, you get lungs that are clumped with proteins, which is patholgoical because they don’t have enough alveolar macrophages to clear those proteins
if you deplete macrophages using radiation, moncytes are recruited from the bloodstream.
Describe how the alveolar macrophages are in a ‘balancing act’
Activation of alveolar macrophages
depends on a balance between signals
leading to activation and inhibition
In the absence of inflammation
regulation predominates through
negative signals
The epithelia raises the threshold above
which macrophages get activated
Negative signals are provided by the
airway epithelium and soluble
molecules like mucus
Regulation is linked to macrophage
function i.e. clearance of apoptotic cells
alveolar macrophages can have pro-infalmmatory activity, receiving signals from IL1, IFNG, TNF
but can also detect pathogens through toll like receptors, responding to TLR2, 4 and 6
can detect gram positive bacteria, gram negative bacteria through peptidoglycans and LPS
can detect lectins
provide activating and regulatory signals
they also express TGF B, IL 10, CD200, apoptotic cell sensing receptors like TAM receptors all provide negative signals
but it’s not just about pro-inflammatory cytokines or anti-inflammatory cytokines, but also the close contact with the spithelium
epithelium finds a threshold- when negative threshold overcome, macrophages can actually go into activity
Describe in situ negative regulation of homeostasis in the alveolar space
Regulation is highly dependent on contact with epithelia, setting a threshold above which macrophages activate
Antigen plus a disruption of the epithelia drives macrophage activity
After inflammation the tissue adapts to a novel state of regulation – we are never the same after inflammation
Processes that increased macrophage activation:
CD 200 expressed by epithelium, which provides signal from CD 200 RECEPTOR in alveolar macrophage
through adaptor molecules like DOC 2, blocks MRK and p38- this signalling normally produces cytokines
IL 10 is produced by epthelium, IL 10 receptor is on alveolar macrophages, this binding signalling process will act through SOCS3 to block cytokine production and promote regulatory micro RNAs
TGF B can act via SMAD and block their ability of macrophages to produce pro-inflammatory cytokines
soluble molcules like surfactant proteins can block TLRs, stop them from producing NFKB, activatingtors like serum alpha- blocking phagocytosis
Describe control of innate immunity in the airway
let’s focus on the interaction between alveolar macrophages and epithelium through the CD200, TGF B and IL 10 receptors
alveolar epithelium express high levels of CD200 on luminal side (CD200 recpetor in alveolar macrophages- nb. this receptor is expressed less in macrophages in spleen suggesting specialisation on lungs) , an integrin bound to TGF B (that can provide signal to macrophages) and IL 10 which can be released to the vicinity and activate the macrophage- providing them NEGATIVE regulation, steady state- defining the threshold over which the macrophage will get activated
Evidence:
animals that don’t have the integrin tethered to TGF B in their alveolar epithelium- they develop emphysema
in absence of IL 10 signal, when stimulating LPS, higher levels of IL 6 lead to higher levels of activation of macrophages
actual microbioita of lungs is altered without CD200 receptors on macrophage surface
infections change the bacteria
change immune responses
bacteria like acinteobacter can proliferate in the presence of infection
How do infections alter the state of regulation of homeostasis by affecting macrophage function
during infections
altered state of health
alveolar macrophages highly regulated by epithelial cells
in steady state (normal condiitons in healthy lung) there will be a threshold above which macrophages can get activated
there can be polymorphisms in these receptors, meaning in some people macrophages can be activated at different levels or have different thresholds
if we infect mice with influenza, levels of CD200 receptor increase and remain higher than before
ability of macrophages to activate is diminished due to greater level of regulation by CD 200 receptors
consequence is that if you then give the animal a bacterial infection like streptococcus pneumonia after, animals that have had the flu were more susceptible to the infection
flu changes levels of CD 200/ other regulatory signals, making them in a higher state of regulation and less able to fight the bacteria
How do we study immune mechanisms behind susceptibility to bacterial pneumonia during asthma?
look at how a chronic condition-asthma- effects the ability of the lung to fight bacteria
if mice exposed to HDM (house dust mites)
mice develop condition similar to human asthma
lungs recruit immune cells
mostly eosinophils recruited- same in humans with asthma
HDM allergen exposure in mice recapitulates a pathology that is comparable to human Asthma S.pneumoniae induces lethal pneumonia in asthmatic mice due to neutropenia
in bronchioles- strong purple staining
high perivascular infiltration of leukocytes
so we can give asthmatic mice strep pneumonia and investigate what happens at 4h, 48h and 96h
it is actually LETHAL due to neutropenia
explains why in the clinic asthmatic patients are susceptible to pneumonia
if we count the bacteria colonies in the mice who are asthmatic vs vehicle- larger colonies
only the asthmatic mice got pneumonia leading on to bacteraemia
associated with the lungs in asthmatic patients unable to gather cells needed to clear bacteria- neutrophils
blunted neutrophil activation-due to inability to produce appropriate cytokines and chemokines to recruit these neutrophils
How do macrophages sometimes mess with the apoptotic cells?
Apoptotic cells are present
in most inflammatory
conditions and their
clearance is required
apoptotic cells need to be cleared by phagocytosis- happens in healthy lungs
sometimes macrophages clear apoptotic epitheilal cells that are turning over
this diagram shows the TAM receptors (tyro3, AXL and MER)
translocation of phosophatidylserine to the outer plasma membrane is a signal for apoptosis
GAS 6 receptor binds to phosphatidylserine
this complex is detected by AXL
produces ‘eat me’ signal
detecting apoptotic cells provide negative signals
How does apoptotic cell clearance regulate alveolar macrophages
TAM receptors control cytokine
production and TLR signaling
altering macrophage function
Apoptotic cell clearance is
a hallmark of resolution in
inflammation
TLR3 and 9 can detect LPS or peptidoglycan (bacteria)
triggers adaptors downstream- TRIF and TRAM pathway
causes migration of NFKB pathway and translocation of NFKB (transcription factor)
activation of genes that code for cytokines like TNF alpha
give endocrine/autocrine signals to cells
via STAT pathway activate more pro-inflammatory cytokines causing NEUTROPHIL recruitment
signals provided by TAM receptors block both TLR signals and STAT pathway
resulting in reduction of production of pro inflammatory cytokines
Describe the role of sugars and mucus
Siglec-F is expressed by alveolar
macrophages extracted by lavage
mucus contains muc 5B
this protein contains specific sugar modifications, in this a2,3-SA
this binds specifically to receptors on macrophages (lectins)
lectins have a transmembrane domain (binding) and signalling domain
siglec F has an ITIM motif (tyrosine inhibitory protein) that can provide negative signals to control inflammation
Macrophages interact with mucins both in “health” and in inflammation via Siglecs
Siglec-F signals decrease the expression
of Trem2 and Axl on alveolar macrophages
we think that siglec F is highly expressed in alveolar macrophages and when they come in contact with epithelium or muc 5B
they produce more siglec F, regulating phagocytosis and producing cytokines
when macrophages are activated eg. through bacteria, this interaction is lost
macrophage activity is not regulated and is increased so they can interact with the mucus to interact with siglec F
in asthma, this production of siglec F can change regulation causing them to produce a lot of mucus
siglec F also regulates eosinophils
Alveolar macrophages are long-lived but do not return to the same state of regulation after inflammation
Key points for alveolar macrophages in chronic inflammation
Chronic conditions like asthma are not only or necessarily exacerbated by infection
Individuals with chronic respiratory conditions (or after acute inflammation) have an altered state of health that affects their response to infections
This altered state of health might be associated with a different level of regulation or activity of immune cells (alveolar macrophages) in the lungs
Alveolar macrophages interact with the environment in different ways in homeostasis or after inflammation
All of these principles seem to apply to multiple inflammatory conditions
The integrity and activity of the epithelia is key to maintain homeostasis
lecture summary
The lung mucosal epithelia is not just a barrier, it interacts with commensals and immune cells to maintain homeostasis and initiate regulated immune responses
The lung epithelia express a group of receptors that provide signals that control and harness macrophage function
Soluble factors also regulate macrophage function (environment)
The integrity of the epithelia is essential to maintain mucosal immune homeostasis
High expression of negative regulators, and negative feedback loops by TAM receptors, are predominant in resolution tipping the balance of macrophage function towards regulation
Alveolar macrophages change their responsiveness with every inflammatory response and phenotypic changes might be prolonged suggesting a process of adaptation
These data highlight the importance of understanding the mechanisms of mucosal regulation in health and the differed states of “altered health”
