dont know (5) Flashcards
L16- Immunometabolism: Moonlight in T cells
Glucose metabolism elements…. as transcriptional and post-transcriptional regulators
of the adaptive immune response.
In a resting cd4 or cd8 t cel,GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) iis bound to interferon gamma mRNA. Stop from being translated into protein as GAPDH blocks so ribosome cannot bind. No translation. In naive t cell so cannot produce cytokines.
Tcr signals, activated, cell performs glycolysis. GAPDH is called to glycolysis and transforms glucose to pyruvate. So mrna of interferon gamma is free and ribosome can be activated.
Chatgpt:
The “moonlighting function” in this context refers to GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) having a dual role:
Primary (metabolic) role: In glycolysis, GAPDH converts glyceraldehyde-3-phosphate to 1,3-bisphosphoglycerate, a critical step in energy production.
Moonlighting (non-metabolic) role: In resting T cells, GAPDH binds to interferon-gamma (IFN-γ) mRNA and blocks its translation by preventing the ribosome from attaching to the mRNA.
L16- immune cells in tumour?
LINK WITH L20
Immune cells, such as CD8+ T cells, can become exhausted in the tumour microenvironment due to metabolic stress.
Normal Glucose Environment:
When the T cell receptor (TCR) is engaged, glycolysis is activated to meet the energy demands of the cell.
A byproduct of glycolysis, phosphoenolpyruvate (PEP), plays an important role in supporting the immune response.
PEP can modulate signaling pathways (such as NFAT, nuclear factor of activated T cells) critical for T cell function and antitumour activity.
Tumour Microenvironment:
In tumours, glucose availability is often restricted because tumour cells consume glucose rapidly (a phenomenon known as the Warburg Effect in cancer).
Without sufficient glucose, T cells cannot sustain glycolysis, leading to reduced PEP production.
Low PEP levels impair NFAT activation, which is essential for the transcription of genes involved in the immune response.
As a result, CD8+ T cells are less effective at mounting an antitumour response.
L16- immunometabolism: macrophages (M1 and M2)
M1 Macrophages (“Pro-inflammatory”):
M1 macrophages differentiate in response to bacterial infections or inflammatory signals (like LPS, a bacterial component, IFN-Y and Bacterial PAMPS).
They are responsible for secreting pro-inflammatory cytokines (e.g., IL-1β, TNF-α) and producing reactive oxygen species (ROS).
These macrophages are associated with pro-inflammatory responses and are key in fighting infections.
Their metabolic state is reprogrammed to support this function.
Activation of mTOR and HIF-1α:
LPS activates mTOR (mechanistic target of rapamycin) and HIF-1α (hypoxia-inducible factor 1-alpha).
HIF-1α is typically activated under low oxygen, but in macrophages, its activation can occur independently of oxygen levels due to LPS signaling.
Glycolysis Dominance (Warburg Effect):
Like effector T cells, M1 macrophages rely heavily on glycolysis (breaking down glucose for energy) instead of the full TCA cycle.
This shift (similar to the Warburg Effect in cancer cells) allows for rapid energy production and supports inflammatory functions.
Disruption of the TCA Cycle
When M1 macrophages are activated:
The TCA cycle (Krebs cycle) is broken into two parts.
The cycle is interrupted between citrate and aconitate, leading to the accumulation of citrate and succinate.
Key Metabolic Effects:
Citrate Accumulation:
Citrate is used for the biosynthesis of molecules needed for inflammation (e.g., fatty acids, prostaglandins).
It is also converted to itaconate, a molecule with antibacterial properties. This is an M1-specific defense mechanism.
Succinate Accumulation:
Succinate stabilizes and activates HIF-1α, which enhances the production of IL-1β, a key pro-inflammatory cytokine. Citrate facilitates biosynthesis. Citrate transforms in itaconate. Antibacterial. M1 specific mechanism.
M2 Macrophages (“Anti-inflammatory”):
M2 macrophages are associated with immune regulation, tissue repair, and resolving inflammation.
They help in wound healing by promoting granulation tissue formation and repairing damaged tissue.
Metabolic Reprogramming in M1 Macrophages
Metabolic Shift:
When macrophages receive a signal (like LPS binding to TLR4), they undergo a process called metabolic reprogramming to support their pro-inflammatory functions.
Cytokine Signals: IL-4 and IL-13
These anti-inflammatory cytokines activate M2 macrophages.
IL-4 and IL-13 bind to their receptors on macrophages, triggering intracellular signaling pathways.
STAT6 Activation:
The binding of IL-4 or IL-13 activates the STAT6 (Signal Transducer and Activator of Transcription 6) pathway.
STAT6 drives the transcription of genes that support anti-inflammatory responses and tissue repair.
PPAR and RXR Activation:
PPAR (Peroxisome Proliferator-Activated Receptor) and RXR (Retinoid X Receptor) are nuclear receptors that modulate lipid metabolism and gene expression.
These receptors work together to drive metabolic changes necessary for M2 macrophage functions, including fatty acid metabolism.
M2 macrophages rely on oxidative metabolism rather than glycolysis, in contrast to M1 macrophages. This includes:
Triglyceride (TG) Breakdown to Fatty Acids:
M2 macrophages metabolize triglycerides into fatty acids as their main energy source.
Fatty Acid Oxidation (FAO):
The fatty acids undergo β-oxidation in mitochondria, generating energy in a slow but sustained manner.
This metabolic pathway supports their prolonged anti-inflammatory and tissue-repair functions.
Intact TCA Cycle and Oxidative Phosphorylation (OXPHOS):
Unlike M1 macrophages, M2 macrophages have a fully functional TCA cycle (Krebs cycle) and oxidative phosphorylation (OXPHOS).
These processes provide efficient energy production and reduce oxidative stress, aligning with their roles in anti-inflammatory responses.
Functional Outcomes
Prolonged Responses:
M2 macrophages are metabolically optimized for long-term activity, supporting sustained anti-inflammatory and tissue repair functions.
Granulation Tissue Repair:
The metabolic pathways drive the production of molecules needed for wound healing and granulation tissue formation, like collagen and extracellular matrix proteins.
Anti-Inflammatory Gene Expression:
M2 macrophages upregulate genes involved in immune regulation, reducing inflammation and resolving tissue damage.
M2 feed on lipids like memory t cells. Tca not broken. Perform oxphos. Have lots of mitochondria. Make sure m2 are long lived
Purified antigen (subunit) vaccines? + adjuvent?
Purified antigen (subunit) vaccines: eliminate safety concerns with live vaccines, antigen purified from pathogen. Normally given together with an adjuvent e.g: tetanus
2nd -generation vaccines produced to eliminate the
safety concerns associated with attenuated microbe
vaccines
Subunit vaccines are composed of antigens purified
from microbes or inactivated toxins and are usually
administered with an adjuvant*
Purified antigen vaccines are used for the prevention of
diseases caused by bacterial toxins
Toxins can be rendered harmless without loss of
immunogenicity, and such toxoids induce strong
antibody responses
Diphtheria and tetanus are two infections whose life-
threatening consequences have been largely
controlled because of immunization of children with
toxoid preparations
*adjuvant is a chemical
substance that can be
added to a vaccine in
order to enhance the
immune response (via
DC cell activation)
Adjuvants
The initiation of T cell–dependent immune
responses against protein antigens requires
that the antigens be administered with
adjuvants
Adjuvants elicit innate immune responses, with
increased expression of co-stimulators and
production of cytokines, such as IL-12, that
stimulate T cell growth and differentiation
Only two are approved for patients—
aluminium hydroxide gel (which appears to
promote mostly B cell responses)
and a lipid formulation called Squalene that
may activate phagocytes
L19- Bacterial polysaccharide
a
Bacterial polysaccharide antigen vaccines
are used against pneumococcus
and Haemophilus influenzae
However, polysaccharides are T-independent
antigens, they tend to elicit low-affinity
antibody responses and are poorly
immunogenic in infants (who do not mount strong T
cell-independent antibody responses)
High-affinity antibody responses may be
generated against polysaccharide antigens
even in infants by coupling the
polysaccharides to proteins to
form conjugate vaccines: weak prufiied antigen given in combination with strong antigen to help immune response like a carrier protein. Used when bacterial polysaccharide antigen. Help t cell response, enable th to enable antibody production. These types do not gen cd8 responses but cd4. So not good for viruses as they often presented through mhc class 1 pathway so mhc class 1 aka cd8 needed.
These conjugate vaccines elicit helper T cells.
to simulate germinal centre GC reactions,
which would not occur with simple
polysaccharide vaccine
Purified protein-based vaccines?
stimulate helper CD4+ T cells and
antibody responses, but they do not
generate potent cytotoxic T cells (CD8+ T
cells or CTLs)
The reason for poor CD8+ CTL
development is that exogenous proteins
(and peptides) are inefficient at entering
the class I MHC pathway of antigen
presentation. Thus, protein vaccines are
not recognised efficiently by class I MHC-
restricted CD8+ T cells?
L19- Synthetic antigen vaccines
Made of recombinant dna derived from virus or pathogen like hepaptisis or hpv vaccine. Antigen cloned and incorporated into vaccine that generates immune response from antigens in vaccine. Also good at promoting immune responses.
A goal of vaccine research - identify the most
immunogenic microbial antigens or epitopes, to
synthesize these in the laboratory, and to use the
synthetic antigens as vaccines
- deduce the protein sequences of microbial antigens
from nucleotide sequence data and to prepare large
quantities of proteins by recombinant DNA
technology
Vaccines made of recombinant DNA-derived
antigens are now in use for hepatitis B virus and
human papilloma virus (HPV)
In the case of the most widely used HPV vaccine,
which was developed to prevent cancers caused by
the virus, recombinant viral proteins from four strains
(HPV 6, 11, 16, and 18) are made in yeast and
combined with an adjuvant
L19- Viral vectors - live viral vaccines – recombinant viruses?
Gene based approac (3rd generation) viral vector is source of antigen. Infect cells and induce t cell response.
Alternative gene based approach - introduce genes
encoding microbial antigens into a non-cytopathic virus and to
infect individuals with this virus
Thus, the virus serves as a source of the antigen
The great advantage of viral vectors is that they, like other
live viruses, induce the full complement of immune
responses, including strong CD8+ T cell/CTL responses
This technique uses vaccinia virus vectors, and canarypox
viral vectors, which are not pathogenic in humans
Recombinant viruses induces both humoral and T cell-
mediated immunity against the antigen produced by the
foreign gene
A potential problem - the recombinant viruses may infect host
cells, although they are not pathogenic, they may produce
antigens that stimulate CTL responses that kill the infected
host cells. These and other safety concerns have limited
widespread use of viral vectors for vaccine delivery
Gene-based vaccines carry the genetic
instructions for the host’s cells to make the
antigen, which more closely mimics a natural
infection
The viral vector technique transports genetic
information in a less harmful virus — often a
common cold–causing adenovirus —
sometimes engineered so it can’t replicate in
the host
ChAdOx1nCoV-19 a non–replicating viral
vector candidate in phase 3 trials from
AstraZeneca and the University of Oxford,
uses an adenovirus that infects chimpanzees
instead of humans. But, it’s possible that
cross-reacting pre-existing immunity to human
adenoviruses could still diminish the
response.
DNA vaccines (gene based)
Instead of using live virus, innoculating patient with plasmid encoding antigen in the case of dna? Very good at activating dendritic cells through toll-like receptors.
Inoculation of a plasmid containing complementary DNA
(cDNA) ‘naked DNA’ encoding a protein antigen leads to
humoral and CD4+ and CD8+ T cell responses to the antigen
It is likely that APCs, such as DCs, are transfected by the
plasmid and the cDNA is transcribed and translated into
immunogenic protein that elicits specific responses
Bacterial plasmids are rich in unmethylated CpG nucleotides
and are recognized by DCs (TLRs), thereby eliciting an innate
immune response that enhances adaptive immunity
Therefore, plasmid DNA vaccines could be effective even
when administered without adjuvants
The ability to store DNA without refrigeration for use in the field
also makes this technique promising + cost-effective
However, early DNA vaccines did not produce adequate
amounts of the immunogen (plus reduced generation of
immune responses when compared to ‘viral’-based vaccines)
Studies with newer vectors are currently in progress
DNA vaccine: the genetic material must first
enter the host cell’s nucleus. From there,
messenger mRNA is created, which travels
out of the nucleus into the cytoplasm, where
protein is formed from it. However, genetic
information can only enter the nucleus when
the cell is dividing, making the process
inefficient.
Recently shown not to generate a very robust immune response.
COVID-19 and mRNA vaccines (gene-based)
a
Chinese researchers posted the novel coronavirus’
RNA sequence CoV-19 on a preprint server
Rapid response genetic platforms have catapulted
development of vaccines, crucial during a pandemic
July 2020, mRNA-1273 and BNT162b2 from BioNTech
and Pfizer, both entered phase 3 trials with mRNA
vaccines, which together will enrol an estimated 60 000
volunteers
With COVID-19 - experts say that if the technology pans
out, the pandemic could help to usher in a new plug-and-
play approach to vaccinology
DNA and mRNA vaccine designs
deliver naked nucleic acids or, more
recently, encapsulate them in carrier
nanoparticles (e.g. BNT162b2).
The COVID-19 mRNA is taken up by
cells and translated intro viral protein
by host cells
Designed mrna so it is safe + stable + clean (free of contaminants). Have to avoid degradation when given as a vaccine, protected mrna within nanoparticles. Common with viral-based have advantage of activating cd8 and cd4 t cells.
For COVID-19 mRNA vaccine design, scientists have
focused on:
* mRNA design - the novel coronavirus’ spike protein
with genetic modifications that stabilise the spike —
important for a robust and safe antibody response
* making the mRNA less inflammatory and therefore
safer
* Purifying mRNA to rid it of contaminants
* Protecting the mRNA from degrading too quickly in the
body by encasing it in lipid carrier molecules –
nanoparticles (delivery vehicle)
* These delivery vehicles, already in use with
therapeutic small interfering RNAs (siRNA), help
mRNA cross the cell membrane and may even have
an immune-stimulating adjuvant effect
Gene-based vaccines have a potential
immunological advantage. In addition to
eliciting antibodies and CD4+ helper T cells,
they recruit CD8+ cytotoxic T cells/CTLs (‘killer
T cells’), through the major histocompatibility
class I (MHC-I) pathway
The body’s cells only display viral proteins on
their surface through this MHC-I pathway if
those cells themselves have produced the
viral proteins
(most protein-based vaccines don’t do this –
CD8+ T cells not stimulated)
In cutting out the viral vector, both DNA and
mRNA vaccines eliminate the risk of pre-
existing immunity against it, which can limit
effectiveness (immune system clears a vector
before entering cells)
Effectiveness?
mRNA platform – 95% of cells that meet the
RNA take it up and make protein, so it is an
incredibly efficient process
Safety?
mRNA can’t cause an infection. It also doesn’t
enter the cell’s nucleus, so the chance of its
integration into human DNA is believed to be
very low
In addition, the body breaks down mRNA and its
lipid carrier within a matter of hours, lessening
some concerns about long-term risks
Tolerability, Durability?
Do vaccines cause local injection pain?
Make host feel under the weather for a
day or two?
Antibodies may reduce over time to
Covid-19?
2-dose vaccine regimens could help to
overcome this and strengthen immunity
