Pathogenesis & Malarial Anaemia Flashcards
Broadly speaking, why doe some individuals, but not others, suffer from severe malaria?
- access to healthcare
- the person
-> age, immune status, genetic differences eg sickle cells, duffy negative)
the parasite (virulence factors)
Why is P. vivax only found in East Africa (Somalia, Ethiopia, Kenya)?
95% of the population lack the Duffy antigen on their RBCs
Vivax uses Duffy to enter RBCs
recent evidence in Madagascar of P. vivax infections in Duffy negative people
– exact mechanisms unclear
– seems inefficient and parasite load is low
– parasite will continue to evolve
Give some information on the Duffy antigen.
aka Duffy Antigen Receptor for Chemokines (DARC)
a glycoprotein expressed on the surface of RBCs, endothelial cells, and epithelial cells
acts as a receptor for chemokines like IL-8 and regulates inflammation by binding and clearing chemokines from the bloodstream
commonly caused by GATA-1 TF binding site mutation in the FY gene promoter
– prevents Duffy expression on RBCs
What is sickle cell disease?
Sickle cell anemia is caused by a mutation in the HBB gene, which encodes the beta-globin subunit of hemoglobin
This mutation leads to the production of hemoglobin S (HbS) instead of normal hemoglobin A (HbA)
When oxygen levels drop, HbS polymerizes, causing RBCs to become rigid, deformed, and “sickle-shaped”
What is the difference between those who carry one or two copies of the HbS gene?
Sickle Cell Trait (heterozygous):
offers significant protection against severe malaria without the severe health complications of sickle cell anaemia
Sickle Cell Anaemia (homozygous):
associated with complications like chronic anemia, vaso-occlusive crises, and organ damage
may still experience some protection from severe malaria, but the health risks associated with sickle cell anemia often outweigh this benefit
How does sickle cell anaemia confer resistance to malaria?
haem is present in a free form in the blood of mice with sickle cell trait
– injecting haem into the blood of normal mice before infecting them with malaria, researchers found mice did not develop disease
humans/animals with the sickle-cell trait have to break down and detoxify misshapen red blood cells their entire lives, phagocytes already in place when the malarial parasite attacks them
Sickled RBCs have altered membrane properties, reducing their ability to adhere to blood vessels and form rosettes, decreasing the risk of vascular blockages and severe complications
Why do those who have sickle cell disease have immunity to P. falciparum specifically?
Other malaria species, like P. vivax, rarely cause severe disease and do not rely on similar cytoadherence mechanisms
– P. vivax primarily infects reticulocytes (young RBCs) and uses the Duffy antigen for entry, processes less affected by HbS
P. falciparum has a high metabolic demand and is more sensitive to environmental stress compared to other malaria species
How do cyclical fevers occur in malaria?
in an RBC, trophozoites mature into schizonts which replicate until the cell burst and schizonts are released into the blood
erythrocyte death is what causes fever
– release of parasite nucelic acid, hemozoin, cell membranes into bloodstream
– recognised by innate immune system
cyclical as replication within and subsequent release from RBCs takes time:
– P. malariae = every 72h
– P. vivax, P. falciparum = every 48h
What cytokine is the main cause of fever in malaria infection?
treatment of cerebral malaria w an anti-TNF mAb reduces fever
but does not reduce malaria mortality and results in greater frequency of long-term neurological defects
fever = useful
How was a CHMI study used to look at differences is gene expression between individuals after infection with malaria?
14 volunteers injected w P. falciparum-infected blood
followed for 7-11 days, blood collected every 2 days
3 sets of people found:
– inflammatory = increased gene expression
– suppressive = decreased gene expression
– unresponsive = no change in gene expression
clinical symptoms also varied and matched transcriptional responses
who would go on to develop severe malaria?
What blood stage does sequestration of iRBCs occur?
mature iRBC stages (pigmented trophozoites –> schizonts)
Where does sequestration of iRBCs occur?
in microvascular beds (small blood vessels) throughout the body
brain, gut, skin, adipose tissue (fat), muscle, heart, lung, liver, kidney, spleen, and placenta
this is a cardinal feature of P. falciparum
How does sequestration relate to severe malaria in P. falciparum infection?
all P. falciparum isolates sequester, yet not all infections end in severe disease
severe malaria only develops in infections w a high parasite burden
– necessary but not sufficient
What is acidosis?
obstruction or anaemia leads to tissue hypoxia and a switch from aerobic to anaerobic glycolysis = build up of lactic acid = reduction in pH of blood
acidosis causes impaired consciousness/coma, respiratory distress, and organ failure
best prognostic factor for fatal outcome
– severe malaria is a metabolic disease
reversal of acidosis is v difficult
– only way is to get rid of the parasites
Why does P. falciparum sequester?
remodelling of RBC membrane = increased cell rigidity
the spleen removes rigid cells (usually old or damaged), so sequestration protects iRBCs from being removed
also, low oxygen environment in post-capillary venules is beneficial to parasite growth
What is cerebral malaria?
sequestration causes brain swelling
swelling has nowhere to go, usually goes down back of neck –> squashes brain stem –> stop breathing
Do host immune cells contribute to severe malaria pathology?
no widespread presence of inflammatory cells such as neutrophils and monocytes at sites of sequestration
limited evidence for monocytes or T cells at sites of brain sequestration in fatal human CM
– pronounced feature in experimental CM mouse model
Do cytokines contribute to severe malaria pathology?
high levels of host inflammatory cytokines may contribute to severe malarial anaemia (SMA)
no evidence they are causal, but host cytokines increase expression of cytoadherence receptors on endothelial cells
– bad situation made worse
What is rosetting?
a process in which P. falciparum-iRBCs bind to uninfected red blood cells (uRBCs) as well as endothelial cells, forming clusters or aggregates
mediated by binding of PfEMP-1 to different receptors
What receptors on endothelial cells does PfEMP1 bind to?
– CD36 -> tissue microvascular endothelium (ME) = mild malaria
– ICAM-1 -> cerebral ME = high mortality
– CSA -> placental ME = prematurity/death of foetus
What receptors on uRBCs does PfEMP1 bind to?
–complement receptor 1 (on platelets too)
– ABO blood group antigens (e.g., rosetting is stronger in individuals with blood group A than in those with blood group O)
– GAGs
Give some info on the variation of PfEMP1?
encoded by var genes, located on the end of chromosomes
– these regions more likely to recombine
only 1 var gene transcribed per cell, but daughter schizonts can switch what gene is transcribed
– there often tends to be a dominant var gene
What is the structure of PfEMP1?
extracellular domain, variable
made up of:
– Duffy-Binding-Like (DBL)
- Cysteine-rich Inter-Domain Regions (CIDR)
conserved TM domain and cytoplasmic tail - acidic terminal sequence (ATS)
What are the 3 well-conserved, “strain-transcending” var genes?
found in all P. falciparum isolates
var1 and var3 of unknown function
var2CSA encodes VAR2CSA PfEMP1 variant that mediates iRBC binding to chondroitin Sulfate A in placenta during pregnancy-associated malaria
What is Chondroitin Sulfate A?
ECM proteoglycan
consists of a long carbohydrate chain made up of repeating disaccharide unis attached to a core protein
a unique form of 4-sulfated CSA in placenta
Why is pregnancy associated malaria (PAM) an issue?
most adults are asymptomatic during malaria infection, an exception is pregnant women
PAM causes maternal anaemia, intra-uterine growth retardation, and perinatal deaths
How does sequestration in the placenta cause issues?
adhesion of P. falciparum iRBC on placental syncitiotrophoblast cells (CSA expressing)
– inflammation and fibrosis due to infiltration by neutrophils and monocytes impairs placental function
= low birth weight babies
Why is PAM only a concern during the first and second pregnancies?
VAR2CSA is highly conserved protein
women develop antibodies against this surface protein
vaccine currently in development
Why have PAMVAC trials been unsuccessful so far?
lack of antibody cross-reactivity against heterologous parasite isolates
whole protein is large and would be difficult to put into a vaccine, so only a part is being used
difficult to have trials on pregnant women -> unethical
The bone marrow as a malarial niche.
In the early stages of infection,P. falciparumgametocytes have been found in the bone marrow, where their maturation takes place
the BM has also been recognized as a development niche or reservoir forP. vivaxgametocytes
What is sensescence?
deterioration of cells with age
RBC lifespan = ~120 days
What is the process of RBC senescence?
RBCs experience chemical/mechanical stresses, causing damage
– now way to produce new proteins
lipid and protein oxidation –> inactive haemoglobin aggregates + clustering of Band 3 (anion exchanger)
auto-antibodies are abundant in serum
– clustering triggers binding by IgG
deposition of complement (C3b-IgG complex)
– target for opsonisation by macrophages of the reticuloendothelial system (RES)
What is the function of transferrin and ferroportin in erythrophagocytosis?
transferrin
–iron-binding glycoprotein in blood
– transports iron to tissues, including BM –> for erythropoiesis.
ferroportin
– macrophages digest haemoglobin from engulfed RBCs, releasing iron
– stored temporarily as ferritin or hemosiderin in macrophages
– Ferroportin exports this iron into the bloodstream, where it binds to transferrin for systemic transport
What is the function of transferrin and ferroportin in erythropoiesis?
transferrin
– Erythroid cells express transferrin receptors (TfR1) -> mediate uptake of transferrin-bound iron
– inside the cell, iron is released in the endosome and delivered to the mitochondria for incorporation into heme
ferroportin
– facilitates mobilization of iron from storage sites (e.g., macrophages and hepatocytes)
– maintains supply of transferrin-bound iron for erythropoiesis
What type of RBCs do different species of Plasmodium infect?
P. vivax - reticulocytes (young RBCs)
P. falciparum - mature RBCs
What is the function of erythropoietin in erythropoiesis?
EPO
hormone produced by kidneys in response to hypoxia to induce erythropoiesis
also binds tissue protective receptor (TPR) to regulate both adaptive and innate immune cells
How does hepcidin inhibit ferroportin?
liver derived hormone which degrades ferroportin
produced in response to inflammation
– prevents giving iron to pathogens
What is the pathogenesis for malarial anaemia?
- lysis of parasitised (p)RBCs
- removal of RBCs in the RES (spleen)
- decreased and/or suppression of erythropoiesis
- removal of uRBCs
– for every 1 pRBC, 12 uRBCs are lost
– rosetting, PfEMP1 deposition
What is hemozoin?
Hz
– crystalline w a brown pigment
– parasites use up to 80% of RBC Hb
– digestion of Hb creates free heme –> highly toxic
– heme aggregated to insoluble crystal
immunostimulatory - macrophages that take up Hz produce large amounts of cytokines and chemokines
–> Hz acts through TLR9, presenting parasite DNA
What are the arguments against giving blood transfusions to treat severe malaria?
- takes time to set up, 2-8h
- transfusion-related blood borne infections (in underdeveloped countries may be more tru)
– transfusion-transmitted malaria known - typical complications
– fluid overload, allergic reaction, incompatibility of transfused RBCs - more RBCs for parasites to invade or spleen to clear –> worse?
What are the arguments for giving blood transfusions to treat severe malaria?
– already widely used
– shorten recovery time
– Blood transfusions can quickly restore oxygen-carrying capacity, preventing hypoxia and organ failure
– despite uncertainty, important to add this to supportive care for those critically ill
Why does induction by EPO not reverse dyserythropoiesis?
Excessive levels of EPO can overstimulate the proliferation of erythroid progenitors, prioritizing rapid cell division over differentiation
erythroid progenitors to accumulate without progressing to terminally mature stages
– no increase in haemoglobin levels
– adds more opportunities for P. vivax infection
Why is RBC deformability important?
spleen is the largest filter of RBCs in the body, w v small openings (splenic slits)
surface to volume ratio is a predictor of deformability
IRBCs and uRBCs have reduced deformability = stuck = phagocytosis
What is eryptosis?
apoptosis of damaged eythrocytes
- shrinkage
- membrane blebbing
- activation of caspase 3 and 8
- phosphatidylserine (PS) on outside of cell
eryptotic RBCs are resistant to P. falciparum culture
How do external factors contribute to eryptosis?
antimalarial drugs
- primaquine can damage RBCs (mostly in G6PD deficient patients)
- artesunate can induce anaemia (20-25% of travellers)
iron deficiency (ID)
- does reduce severe malaria
- controversy around whether iron supplementation can worsen disease and outcome
How much of a problem is malaria co-infection?
12% children w severe malaria present w bacteraemia
NTS most prevalent –> 20% fatality rate
- decreases intestinal polymorphonuclear neutrophils
- reduced inflammation but increased systemic burden
How does Heme-Oxygenase-1 inhibit neutrophil function during malaria + NTS co-infection?
HO-1
- catabolises heme
- limits intracellular iron availability (in-vitro)
- limits ROS and RNS –> susceptible to infection
