Overview

1. D R A B H I J I T C H A U D H U R Y M D , D N B , D ( A B M M )
P R O F E S S O R O F M I C R O B I O L O G Y
S V I M S - S P M C ( W ) , T I R U P A T I , A P .
IMMUNOLOGY OF
LYMPHATIC FILARIASIS

2. AGENDA
1. Introduction
2. Global Burden of Lymphatic Filariasis.
3. Immunity to LF: Elimination Strategy
4. Hyper Reactive state
5. Hypo responsive stage of Chronic Infection
6. Immunopathology of Chronic LF
7. Conclusion.

3. The Three Comrades
 W. bancrofti has the widest distribution. It is
prevalent in Sub-Saharan Africa, south and
southeast Asia and is responsible for some 90% of all
cases of LF.
 B. malayi is found in tropical regions of South and
Southeast Asia, occasionally overlapping with the
range of W. Bancrofti.
 B. timori has the most restricted geographic range
found only in Indonesia and Timor-Leste.

4. Global Burden of LF
 947 million people worldwide remain threatened by
lymphatic filariasis.
 Presently, more than 90 million people in 73
countries are infected with Wuchereria bancrofti.
 Out of the 73 countries considered endemic at the
start of 2015, MDA was no longer required in 18
countries.

5. Global Burden of LF
 As many as 36 million cases worldwide are living
with hydrocoel and lymphoedema.
 LF accounts for at least 2.8 million Disability
Adjusted Life Years.
 MDA is still required among 501.1 million persons
worldwide.
 As of 2015, TAS ( Transmission assessment surveys )
has been completed and MDA stopped in 72 of 255
endemic districts in India and 20 of 61 endemic
districts in Nepal.

7. Clinical Phenotypes of Host Response
1. Endemic Normals: Infection and disease-free despite
continuous exposure to mosquito-transmitted infective
larvae.
2. Hyper-reactive Response/ Acute Infection:
Lymphadenitis and Lymphangitis.
3. Asymptomatic infection/ Hyporesponsive : Mf-ve
but are positive for circulating filarial antigens (Latent).
However in majority of the patients; few visible clinical
manifestations despite being Mf+ve (Patent).
4. Chronic LF: Lymphedema, elephantiasis and
hydrocele. Mf-ve.

8. IMMUNITY TO LF
 Granulocytes (eosinophils, neutrophils, and
basophils) are key effector cells at the frontline
against infections with filarial worms
 Granulocytes are rapidly activated and recruited to
sites of infection where they are important producers
of Th2 cytokines such as IL-4 and IL-13.

9. IMMUNITY TO LF
 Granulocytes can also attack filaria infections
through antibody- dependent cell mediated
cytotoxicity (ADCC) : The killing of antibody coated
parasites via the release of cytotoxic granules.
 Granulocytes are characterized by six major granule
proteins: major basic protein (MBP), eosinophil
peroxidase (EPO), eosinophil cationic protein (ECP),
eosinophil derived neurotoxin (EDN), neutrophil
elastase (NE) and histamine.
 MBP, EPO and ECP are potent helminth toxins.

10. IMMUNITY TO LF
 Major players responsible for early initiation of host
adaptive immune response include dendritic cells,
macrophages, basophils and innate lymphoid cells
(ILCs).
 ILC2 are expanded in both mouse models and
human filarial infections and produce copious
amount of IL-5 and IL-13, preceding the onset of
classical Th2 responses.
 The canonical immune response to lymphatic
filariasis is that of T-helper 2 (Th2) response coupled
with the production of key cytokines.

11. IMMUNITY TO LF
 The important ones are IL-4, IL-5, IL-9, IL-10, IL-13.
 Antibody response of IgG1, IgG4 and IgE .
 In uninfected individuals, the IgG4 isotype consists
of 5% of the total circulatory antibody but rises
sharply in active filarial infection to about 95%.
 IgG4 antibodies are upregulated in the presence of
the immunoregulatory cytokine such as IL-10.

12. IMMUNITY TO LF
 Most of the IgE produced is polyclonal IgE indicating
a non- antigen specific induction of IgE producing B
cells.
 IgE production both in mice and humans is
absolutely dependent on IL-4 or IL-13.
 In vivo data from mice deficient in IgE, showed
increased worm burdens with B. malayi indicating
an important role for IgE in host defense.

13. IMMUNITY TO LF
 IgG4 antibodies compete with IgE for binding sites
and thus obstruct the protective activity of IgE.
 Elevated levels of antigen specific IgG4 is directly
linked with parasite survival.
 A careful regulation of IgE is crucial as it promotes
pathology.
 This clearly suggests that the ratio of IgG4: IgE
immunoglobulin is crucial to infection phenotype.
 However, the regulators of the class-switching
mechanism between these two immunoglobulins are
yet to be defined.

14. IMMUNITY TO LF
 IgG1, IgG2 and IgE have been established to
negatively correlate with MF.
 IgG3 shows negative correlation with circulating
filarial antigen (CFA).
 IgG4 positively associates with CFA ,
 IgA negatively correlate with MF and CFA.
 These findings clearly suggest that a possible
combinatorial regulation of immunoglobulins may
significantly determine filarial infection outcome.

15. Hyper Reactive Response (Acute Infection)
 Recurrent attacks of fever associated with
lymphadenitis and lymphangitis.
 In Bancroftian filaria, fever is common without
other manifestations.
 The lymph nodes commonly involved are the
inguinal, axillary and epitrochlear nodes and the
lymphatic system of the male genitals.
 It is an immune-mediated inflammatory response to
dead or dying adult worms.

16. Hyper Reactive Response (Acute Infection)
 Over reactive T cell responses.
 This group exhibits increased IgE responses, and the
Treg compartment is greatly diminished.

17. The Hyporesponsive State
 The asymptomatic state confers a survival fitness to
the parasite.
 Filarial parasites exert profound immunoregulatory
effects on the host immune system with both
parasite-antigen specific and more generalized levels
of immune modulation.
 The main mechanisms of immune evasion include
immunosuppression, immunological tolerance and
modification of stereotypical Th2 responses.

18. The Hyporesponsive State
 Immunosuppression and immunological tolerance
are characterized typically by immunoregulatory
cytokine induced suppression of immune responses
and a state of immune tolerance in effector T cells,
respectively.
 In the modified Th2 response, antibody isotype
switching to the non-inflammatory isotype IgG4 and
induction of alternatively activated macrophages
occurs.
 The key cytokines involved are IL10 and TGF-β.

19. The Hyporesponsive State
 Alternatively Activated Macrophages: Macrophages
that are activated by the Th2-type cytokines IL-4 and
IL-13.
 They have been demonstrated to drive CD4+ Th2
responses, deviating the immune system from
inducing a pro-inflammatory Th1 response that
could be detrimental to parasite survival.
 Suppress the immune response against the parasite,
promoting anergy and/or tolerance.

20. The Hyporesponsive State
 The immunoregulatory parasite molecule is
Phosphorylcholine (PC) : A small hapten-like moiety
present in the excretory/secretory products.
 One particular PC containing molecule called ES-62
from filarial worms can :
1. Downregulate the proliferation of CD4+ T cells and
conventional B cells,
2. Decrease IL-4 and IFNγ production,
3. Upregulate proliferation and IL-10 production
4. Condition antigen presenting cells to drive Th2
differentiation with concomitant inhibition of Th1
responses.

21. The Hyporesponsive State
 In addition, a recent analysis of the filarial parasite
genome has identified a remarkable number of
human cytokine and chemokine mimics and/or
antagonists within the parasite genome.
 Other potential parasite derived
immunomodulators are serpins and cystatins,
indoleamine 2,3-dioxygenase etc.

22. The Hyporesponsive State
 Regulatory T cells (both natural and adaptive) have
been postulated to play a role in the immune
evasion.
 IL-10 and TGFβ, both factors associated with
regulatory T cells, are elicited in response to
helminth infections, and in vitro neutralization of IL-
10 and TGFβ has been demonstrated to partially
restore both T cell proliferation and cytokine
production in lymphatic filariasis.

23. The Hyporesponsive State
 Based on the expression of the canonical
transcription factor Foxp3, two Foxp3+ subsets have
been identified: thymus-derived (tTregs) and the
peripheral (pTregs) .
 In addition to the Foxp3 expressing Tregs, two other
subsets that do not express Foxp3 have been
described based on the regulatory cytokines
expressed by those cells.
 These include the type1 regulatory T cells (Tr1) that
express mainly IL-10 and the TGF-β expressing Th3
regulatory T cells.

24. The Hyporesponsive State
 Although IL-10 and TGF-β were originally thought
to be produced by Th2 cells and can be produced by
various cell types including regulatory T cells, it has
been shown that the major sources of IL-10 and
TGF- β are Tr1 and Th3 respectively.
 In addition to directly suppressing immune
responses IL-10 and TGF- β may indirectly regulate
not only the antibody response to filarial antigens
but also the function of antigen presenting cells .

25. The Chronic Stage
 In Bancroftian filariasis, the main clinical features
are hydrocele, lymphedema (LE), elephantiasis and
chyluria.
 The manifestations are hydrocele and swelling of the
testis and / or lymphedema of the entire lower limb,
the scrotum, the entire arm, the vulva, and the
breast.
 Lymph vessel dilation is probably the early event
following antigenic stimulation, which takes place
while the adult worms are still alive, when offspring
larvae are being released.

26. The Chronic Stage
 The enlarged lymph vessels become less efficient at
transporting lymph from the periphery making the
system more vulnerable to exogenous
microorganisms.
 Insufficient fluid transport will lead to fluid
extravasation, particularly in the lower limbs, and
eventually to LE.
 Thus, exogenous microorganisms, in particular
Streptococci, are important for the progression of the
condition from subclinical lymphatic aberrations to
LE.

27. The Chronic Stage
 Hydrocele : accumulation of fluid between the two
folia of the tunica vaginalis in the scrotum.
 In this condition, lymph vessel enlargement and
later obliteration, probably including a chronic
inflammatory stimulus for the tunica vaginalis to
secrete fluid between the two folia lead to the
pathology.

28. The Chronic Stage
 Many of these larvae are degenerate and will be
taken up by phagocytic cells,
 Exposure of phagocytes to filarial antigens is
accompanied by triggering of the innate immune
system with the release of not only pro-inflammatory
cytokines but also of molecules that promote
lymphangiogenesis.

29. The Chronic Stage
 In addition to antigens from the worm itself, those
from its Wolbachia endosymbionts appear to play a
major role in pathogenesis.
 Studies have shown that individuals with chronic
lymphatic pathology have elevated levels of pro-
inflammatory cytokines such as TNF-α, IL-6, soluble
TNF receptor, endothelin-1, IL-2, IL-8 in the
peripheral circulation.

30. The Chronic Stage
 Monocytes and granulocytes are thought to be the
predominant source of most of the above-mentioned
pro-inflammatory cytokines.
 The two major independent components of
lymphatic filarial disease are lymphangiectasia and
inflammatory reactions around the adult worms.
 Therefore, innate cytokines appear to play a
prominent role in the initiation of pathology.

31. The Chronic Stage
 Recent studies focusing on molecular mechanisms
regulating blood and lymphatic vessel growth have
shown that vascular endothelial growth factors
(VEGF) control angiogenesis and lymphangiogenesis
in humans.
 Over-expression of VEGF-A increases vascular
density, enhances WBC adhesion,and tissue
infiltration and also promotes lymphangiogenesis.

32. The Chronic Stage
 Over expression of VEGF-C results in lymphatic
endothelial proliferation and dilation of lymph
vessels.
 The expression of VEGF-A and VEGF-C has been
shown to be up-regulated by pro-inflammatory
cytokines, such as IL-1β, TNF and IL-17.

33. The Chronic Stage
ROLE OF WOLBACHIA:
 Filarial nematodes contain the endosymbiotic
bacteria Wolbachia of the order Rickettsiales .
 These endosymbiotic bacteria are found in the
hypodermis of male and female worms, in the
oocytes, embryos and larval stages.
 Play an essential role in worm fertility and survival.

34. The Chronic Stage
 Wolbachia antigens can stimulate host immune
responses that may be associated with development
and progression of pathogenesis of filarial diseases.
 Upon death of the MF or natural death of an adult
worm, the immune system would be exposed to a
large amount of pro-inflammatory stimuli, including
large numbers of Wolbachia.

35. The Chronic Stage
 Wolbachia have been shown to stimulate pro-
inflammatory cytokines, such as TNF, IL-1β and IL-
6, and nitric oxide, which are known to up-regulate
the expression of VEGF-C via TLR 2.
 Doxycycline treatment not only reduced Wolbachia
copy numbers by 95% in W. bancrofti-infected
patients but also reduced the plasma levels of VEGF-
C⁄ sVEGFR-3, resulting in amelioration of dilated
lymphatic vessels and improvement in the
conditions of LE patients.

36. The Chronic Stage
 Data suggest a similar phenomenon for hydrocele
being associated with lymphangiogenic markers,
specifically VEGF-A
 In hydrocele patients, targeting the Wolbachia
endosymbionts by doxycycline led to a reduction in
plasma levels of VEGF-A, with consequential
reduction in the size of hydrocele.

37. Tropical Pulmonary Eosinophilia
 Characterized by the immunological hypo-
responsiveness of human host to the parasite .
 Microfilariae trapped in the lungs degenerate and
release their antigenic constituents that trigger
intense local inflammatory and immune processes
that lead to ↑ ↑ concentration of eosinophils in the
peripheral blood, and increase the level of IgE.
 Mediated by IL-5 , probably alongwith IL-3 and GM-
CSF.

38. Conclusion
 Lymphatic filariasis is a disease whose
manifestations range from asymptomatic state to
gross pathologies seen in lymphedema and
hydrocele.
 Every event is maintained by a fine interplay of
innate immune response, a few key cytokines, and
adaptive response by important immunoglobulin
isotypes.

39. Conclusion
1. Natural immunity or elimination of the parasite is
mainly effected by granulocytes, innate lymphoid
cells, IgE, ADCC, and cytokines like IL-4. IL-13.
2. The asymptomatic state of infection is marked by
immune evasion and immunosuppression.
 Th2 response modified, IgG4 expression ↑ , IL10
and TGF-β production ↑,
 Alternatively Activated Macrophages and T
Regulatory cells (Tr1, Th3) play important role.
 Parasite derived Phosphorylcholine containing
molecule ES-62 acts as important antigen.

40. Conclusion
3. Chronic State:
 Largely mediated by innate immune response with ↑
pro-inflammatory cytokines.
 Overexpression of VEGF A and C play crucial roles
in the pathogenesis of lymphedema and hydrocele.
 Filarial as well as Wolbachia antigens are
responsible for the immunopathology.

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