Modulation of goat monocyte cystatinfrom...

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Page 1: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory
Page 2: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

Modulation of goat monocyte function by HCcyst-2, a secreted

cystatin from Haemonchus contortus

By: Rezaei Riabi Roya

Page 3: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory
Page 4: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

Helminth Immunomodulation in Autoimmune Disease

Helminth

IL-4, IL-5, IL-10, and IL-13

(TGF) β

IL-10 IgE

mastcells, eosinophils, and basophils

promotes Th2 polarization

Page 5: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

• Failure of the immune system

o Inflammatory bowel disease (IBD)

o Multiple sclerosis (MS)

o Rheumatoid arthritis (RA)

o Type 1 diabetes (T1D)

Page 6: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

Haemonchus contortus

Scientific classification

Kingdom: Animalia

Phylum: Nematoda

Class: Secernentea

Subclass: Rhabditia

Order: Strongylida

Family: Trichostrongylidae

Genus: Haemonchus

Species: H. contortus

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• Excretory/secretory (ES)

products are the primary interface between parasitic worms and their hosts

• Cysteine protease inhibitor (cystatin)

• Cysteine protease inhibitor (cystatin)

• Migration inhibitory factor (MIF)

protease activity

monocyte

phagocytosis

Page 8: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

Life cycle

Page 9: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory
Page 10: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

Haemonchus contortus

Sheep, goats, deer, exotic ruminants

Blood-sucking parasite

– highly pathogenic

– causes severe anemia

– causes low blood protein -- “bottle jaw”

Most important parasite in sheep/goats raised in warm/wet environments

Page 11: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

Haemonchus contortus

• Haemonchus contortus – barberpole worm/ wire worm

– 1” (25 mm)

– Abomasum of

small ruminants

– feeds on blood

– Clinical signs

• anemia, bottle jaw, chronic weight loss

• death

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larvae (L3) cultured

2500 larvae/ml at 4 °C.

male goats were fed

fecal samples infected goats

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• Expression and purification of rHCcyst-2 in

Escherichia coli (RT PCR, pMD19-T vector )

• Cloning of HCcyst-2 and bioinformatics

analyses(cleaved by EcoRI)

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purified rHCcyst-2 with adjuvant

subcutaneously

Localization of HCcyst-2 by immunohistochemical study (bySPECTRAFLUOR )

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cystatins

• Belongs to superfamily papain-like cysteine proteases

• The cystatins family find in invertebrates, vertebrates, plants and even in

protozoa

• four subfamilies

Type 1 cystatins or stefins are cytoplasmic proteins

lacking of signal peptides

Type 2 cystatins are secretory-type proteins(excretory/secretory (ES)

Type 3 called kininogens (circulating plasma protein, blood

coagulation )

Type 4 fetuins and histidine-rich proteins

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cystatin gene of H. contortus (rHCcyst-2)143 amino acid & size of 14 kDa

+

goat monocyte

immune modulatory activity

Page 17: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory
Page 18: Modulation of goat monocyte cystatinfrom Haemonchuscontortusparasitology.sbmu.ac.ir/uploads/Presentation1.rigld.pdf · 143 amino acid &size of 14 kDa + goat monocyte immune modulatory

rHCcyst-2 inhibits MHC-II expression on goat monocytes. Monocytes were cultured in the presence of control buffer (PBS/DTT) or different concentrations of rHCcyst-2 for 24 h. MHC-II expression was measured by flow cytometric analysis and calculated as the percentage of mean fluorescence intensity (MFI) of controls. Bars represent the MFI ± SD of controls. (A) MHC-II; (B) MHC-I. The data are representative of three independent experiments

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• Detection of cytokine secretion (ELISA)

decreased the production of TNF-α, IL-1β and IL-12

increased the secretion of IL-10 in goat monocytes

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• Results showed:

I. rHCcyst-2 immunomodulatory effects on goat monocytes

II. Induced the production of IL-10 in goat monocytes

III. Induce an anti-inflammatory environment

IV. Decreased the production of TNF-α, IL-1β and IL-12

V. Increased the secretion of IL-10 in goat monocytes

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thanks foryour

attention