Stress in the regulation of the immune response in Leishmaniasis
Felix J. TapiaInstituto de BiomedicinaFacultad de MedicinaUniversidad Central de Venezuela
Reuniäo Cyted 2009Projecto Mecanismo de controle e imunoprofilaxia na LeishmanioseFaculdade de Medicina da Bahia (FAMEB)
Neuroendocrine-immune axis
Diffuse neuroendocrine system
Peripheral Immune system
Bioactive Polypeptides
hormones neuropeptides cytokines
Molecules of communication
Murine models of American cutaneous Leishmaniasis
C57BL/6 BALB/c
Th1IFN-
Th2IL-4
L. mexicanaMHOM/BZ/82/BEL21
Sánchez et al. Acta Micros 1993, 2: 180-187Aguilar-Torrentera et al. Am J Trop Med Hyg 2002, 66: 273–279
Díaz et al. Clin Exp Dermatol 2003, 28: 288-293
Sensory peptides: Calcitonin gene-related peptide (CGRP) Substance (Sub P) mediators of neurogenic inflammation
Epidermal dendritic (Langerhans) cells:Skin take up and antigen processing Peripheral lymphoid organs initiate primary T cell responses
Leishmania infection of BALB/c and C57BL/6 mice:103 amastigotes L. mexicana strain MHOM/BZ/82/BEL21 Course of infection 12 weeks
Sites of evaluation
Background
Dendritic Langerhans cells are involved in the pathogenesis of cutaneous leishmaniasis.
Epidermal cell signaling is essential to determine the type of cytokine-related immune response to be generated against Leishmania parasites.
Dendritic Langerhans cells, SubP and CGRP are affected by acute stress in contact hypersensitivity models.
Phases of Cutaneous Regulation
Activation Inmunostimulation Effector Phase
ChallengingAntigen CaptureMigration
Antigen PresentationClonal Expansion Recruitment
RetentionProliferationSuppression
epidermis
dermis
Mo
ag
Mo
..
.
granuloma
gangliolinfático
endoteliovascular
IL-4
mastocyte
IFN-
NK
MΦ
IL-12
T memory
QCCL
GM-CSF
IL-1TNF-
chemokines
C
T
T
Objective
Analyze the effect of acute stress (immobilization and odorant) on Langerhans cells, Sub P, CGRP, and the natural course of infection in susceptible and resistant mouse models of cutaneous leishmaniasis
Acute stressor (2, 8 hrs):Immobilization by placement in 50 ml polypropylene centrifuge tubes, before inoculation of Leishmania parasites.
Materials and methodsStress procedure
BALB/cC57/BL6
Measurement footpad thickness every week for 12
weeks
Mice stressed for 2 hr and infected with L. mexicana
Mice stressed for 8 hr and infected with L. mexicana
Mice infected with L. mexicanaInfection Control
Mice stressed for 2 hr and non-infected with L. mexicanaStress Control
(n=20)
(n=20)
(n=20)
(n=20)
Footpad biopsies taken at weeks 0, 4 and 8 after
infection
Cryopreservation
Immunodetection of CD205+ Langerhans
cells, CGRP and Sub P
(n=20)
(n=20)
(n=20)
(n=20)
Experimental design
Progression of L. mexicana infection in experimental groups of BALB/c and C57BL/6 mice
0 1 2 3 4 5 6 7 8 9 10 11 12 131
2
3
4
5
6
Weeks of iinfection
Size
of
lesi
on in
mm
0 1 2 3 4 5 6 7 8 9 10 11 12 131
2
3
4
5
6
2hr stress healthy
2hr stress infected8hr stress infectedNon-stressed infected
Langerhans cell density in experimental groups of BALB/c and C57BL/6 at weeks O, 4 and 8
0.0 4.0 8.0 12.00
250
500
750
10002hr stress healthy2hr stress infected8hr stress infecedNon-stressed infected
0
1000
2000
0 sem 4 sem 8 sem
Weeks of infection
cells
/mm
2
Progression of infection with L. mexicana in BALB/c mice subjected to acute stress
Langerhans cells Iad+ in separated epidermis of BALB/c mice infected with L. mexicana
Non stressed Stressed for 2 hr
Substance P in skin lesions of BALB/c mice stressed and infected with L. mexicana
week 4 - stressed for 8 hr
Vascularedotheli
um
infiltrate
Conclusions
Acute immobilization stress affects the numbers and function of Langerhans cells and Substance P and CGRP innervations.
The skew of the skin immune response after stress and infection may be diverted by epidermal accessory signals to distinct Th1 or Th2 responses in resistant or susceptible mice, respectively.
The genetic background of the mouse strain determined the response to acute stress in murine leishmaniasis.
Odorant inhalation stress in resistant and susceptible mouse models of cutaneous leishmaniasis
Acute stressor (48 hrs):Inhalation of citral for 48 hours.
At 24 hrs of stress, mice were infected Leishmania mexicana
Materials and methodsStress procedure
Odorant inhalation stress alters the immune response in resistant and susceptible mice infected with L. mexicana
Odorant inhalation stress alters the immune response in resistant and susceptible mice infected with L. mexicana
Odorant inhalation stress alters the immune response in resistant and susceptible mice infected with L. mexicana
Conclusions
Citral inhalation stress induces clinical and immunological alterations in the natural course of infection in mice inoculated with L. mexicana.
These alterations include decreased numbers of dendritic Langerhans cells and their morphological, and downregulation of Th1 and Th2 cytokines in both resistant and susceptible mice. Only IL-12 and iNOS remained in normal values.
The results indicate that citral inhalation suppress the immune response against the Leishmania parasite regardless the genetic background of the host.
Macrophages treated by sera from mice sressed and infected with L. mexicana24 hours, Giemsa
1 3 5 9 110
100
200
BALB/c stressed & infect
BALB/c infect
Weeks of infection
cells
/mm
2
Density of DC-SIGN positive cells in lesions of stressed and L.mexicana-infected BALB/c mice
1 3 5 9 110
1000
2000
3000
BALB/c infectado yestresado BALB/c infectado
* *
*
*
Density of macrophages MOMA-2 positive cells in lesions of stressed and L.mexicana-infected BALB/c mice
* p 0.05
cells
/mm
2
Weeks of infection
TLR-2 positive cells in BALB/c mice stressed and infected with L. mexicana5 weeks
1 3 5 9 110
100
200
300
BALB/c stressed & infect
BALB/c infect
Weeks of infection
cells
/mm
2
1 3 5 9 110
100
200
BALB/c stressed & infect
BALB/c infect
*
Weeks of infection
cells
/mm
2
TLR-4 positive cells in BALB/c mice stressed and infected with L. mexicana5 weeks
* p 0.05
CD14 positive cells in BALB/c mice stressed and infected with L. mexicana5 weeks
1 3 5 9 110
100
200
300
BALB/c stressed & infect
BALB/c infect
Weeks of infectionce
lls/m
m2
1 3 5 9 110
100
200
BALB/c stressed & infect
BALB/c infect
TLR-9 positive cells in BALB/c mice stressed and infected with L. mexicana5 weeks
cells
/mm
2
Weeks of infection
Laboratorio de Biología MolecularInstituto de Biomedicina
Estudiantes:Ysamar Chirinos
Alba QuiñonesMaría del Rosario Ruiz
Fabiola CabreraMaría Alejandra Sarabia
Eliana FigueiraMónica Fernández
Adriana ArbeláezCelsy Hernández
Alejandra Da AlmeidaPedro SalazarMaría García
Izaskun Urdanibia
Laboratorio:Nilka L. Díaz
Iraima B. MonsalveMónica SuárezMiguel MarzalLuis González
Colaboradores:Martín A. Sánchez
Zelandia FermínLadys Sarmiento
ObrigadoThanksGracias
American cutaneous leishmaniasis
C57BL/6IFN-
BALB/cIL-4
LCL
Intermediate forms
MCL CCL
DCL
CBAIFN / IL-4