全部 标题 作者
关键词 摘要

OALib Journal期刊
ISSN: 2333-9721
费用:99美元

查看量下载量

相关文章

更多...
PLOS ONE  2012 

Leishmania donovani: Immunostimulatory Cellular Responses of Membrane and Soluble Protein Fractions of Splenic Amastigotes in Cured Patient and Hamsters

DOI: 10.1371/journal.pone.0030746

Full-Text   Cite this paper   Add to My Lib

Abstract:

Visceral leishmaniasis (VL), caused by the intracellular parasite Leishmania donovani, L. chagasi and L. infantum is characterized by defective cell-mediated immunity (CMI) and is usually fatal if not treated properly. An estimated 350 million people worldwide are at risk of acquiring infection with Leishmania parasites with approximately 500,000 cases of VL being reported each year. In the absence of an efficient and cost-effective antileishmanial drug, development of an appropriate long-lasting vaccine against VL is the need of the day. In VL, the development of a CMI, capable of mounting Th1-type of immune responses, play an important role as it correlate with recovery from and resistance to disease. Resolution of infection results in lifelong immunity against the disease which indicates towards the feasibility of a vaccine against the disease. Most of the vaccination studies in Leishmaniasis have been focused on promastigote- an infective stage of parasite with less exploration of pathogenic amastigote form, due to the cumbersome process of its purified isolation. In the present study, we have isolated and purified splenic amastigotes of L. donovani, following the traditional protocol with slight modification. These were fractionated into five membranous and soluble subfractions each i.e MAF1-5 and SAF1-5 and were subjected for evaluation of their ability to induce cellular responses. Out of five sub-fractions from each of membrane and soluble, only four viz. MAF2, MAF3, SAF2 and SAF3 were observed to stimulate remarkable lymphoproliferative, IFN-γ, IL-12 responses and Nitric Oxide production, in Leishmania-infected cured/exposed patients and hamsters. Results suggest the presence of Th-1 type immunostimulatory molecules in these sub-fractions which may further be exploited for developing a successful subunit vaccine from the less explored pathogenic stage against VL.

References

[1]  Guerin PJ, Olliaro P, Sundar S, Boelaert M, Croft SL, et al. (2002) Visceral leishmaniasis: current status of control, diagnosis, and treatment, and a proposed research and development agenda. Lancet Infect Dis 2: 494–501.
[2]  Desjeux P (2004) Leishmaniasis. Nat Rev Microbiol 2: 692.
[3]  Bhowmick S, Ravindran R, Ali N (2007) Leishmanial antigens in liposomes promote protective immunity and provide immunotherapy against visceral leishmaniasis via polarized Th1 response. Vaccine 25: 6544–6556.
[4]  Coler RN, Skeiky YA, Bernards K, Greeson K, Carter D, et al. (2002) Immunization with a polyprotein vaccine consisting of the T-Cell antigens thiol-specific antioxidant, Leishmania major stress-inducible protein 1, and Leishmania elongation initiation factor protects against leishmaniasis. Infect Immun 70: 4215–4225.
[5]  Pham NK, Mouriz J, Kima PE (2005) Leishmania pifanoi amastigotes avoid macrophage production of superoxide by inducing heme degradation. Infect Immun 73: 8322–8333.
[6]  Descoteaux A, Turco SJ (2002) Functional aspects of the Leishmania donovani lipophosphoglycan during macrophage infection. Microbes Infect 4: 975–981.
[7]  Garg R, Srivastava JK, Pal A, Naik S, Dube A (2005) Isolation of integral membrane proteins of Leishmania promastigotes and evaluation of their prophylactic potential in hamsters against experimental visceral leishmaniasis. Vaccine 23: 1189–1196.
[8]  Afrin F, Rajesh R, Anam K, Gopinath M, Pal S, et al. (2002) Characterization of Leishmania donovani antigens encapsulated in liposomes that induce protective immunity in BALB/c mice. Infect Immun 70: 6697–6706.
[9]  Reed SG, Scott P (1993) T-cell and cytokine responses in leishmaniasis. Curr Opin Immunol 5: 524–531.
[10]  Murray HW, Hariprashad J, Coffman RL (1997) Behavior of visceral Leishmania donovani in an experimentally induced T helper cell 2 (Th2)-associated response model. J Exp Med 185: 867–874.
[11]  Skeiky YA, Kennedy M, Kaufman D, Borges MM, Guderian JA, et al. (1998) LeIF: a recombinant Leishmania protein that induces an IL-12-mediated Th1 cytokine profile. J Immunol 161: 6171–6179.
[12]  Russo DM, Jardim A, Carvalho EM, Sleath PR, Armitage RJ, et al. (1993) Mapping human T cell epitopes in Leishmania gp63: identification of cross-reactive and species-specific epitopes. J Immunol 150: 4713.
[13]  Garg R, Gupta SK, Tripathi P, Hajela K, Sundar S, et al. (2006) Leishmania donovani: identification of stimulatory soluble antigenic proteins using cured human and hamster lymphocytes for their prophylactic potential against visceral leishmaniasis. Vaccine 24: 2900–2909.
[14]  Garg R, Dube A (2006) Animal models for vaccine studies for visceral leishmaniasis. Indian J Med Res 123: 439–454.
[15]  Bates PA (1993) Characterization of developmentally-regulated nucleases in promastigotes and amastigotes of Leishmania mexicana. FEMS Microbiol Lett 107: 53–58.
[16]  Eperon S, McMahon-Pratt D (1989) Extracellular cultivation and morphological characterization of amastigote-like forms of Leishmania panamensis and L. braziliensis. J Protozool 36: 502–510.
[17]  Pan AA (1984) Leishmania mexicana: serial cultivation of intracellular stages in a cell-free medium. Exp Parasitol 58: 72–80.
[18]  Burchmore RJ, Barrett MP (2001) Life in vacuoles–nutrient acquisition by Leishmania amastigotes. Int J Parasitol 31: 1311–1320.
[19]  Rochette A, Raymond F, Corbeil J, Ouellette M, Papadopoulou B (2009) Whole-genome comparative RNA expression profiling of axenic and intracellular amastigote forms of Leishmania infantum. Mol Biochem Parasitol 165: 32–47.
[20]  Garg R, Gupta SK, Tripathi P, Naik S, Sundar S, et al. (2005) Immunostimulatory cellular responses of cured Leishmania-infected patients and hamsters against the integral membrane proteins and non-membranous soluble proteins of a recent clinical isolate of Leishmania donovani. Clin Exp Immunol 140: 149–156.
[21]  Chang KP (1980) Human cutaneous lieshmania in a mouse macrophage line: propagation and isolation of intracellular parasites. Science 209: 1240–1242.
[22]  Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685.
[23]  Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A 76: 4350–4354.
[24]  Scott DA, Coombs GH, Sanderson BE (1987) Effects of methotrexate and other antifolates on the growth and dihydrofolate reductase activity of Leishmania promastigotes. Biochem Pharmacol 36: 2043–2045.
[25]  Choudhry A, Guru PY, Saxena RP, Tandon A, Saxena KC (1990) Enzyme-linked immunosorbent assay in the diagnosis of kala-azar in Bhadohi (Varanasi), India. Trans R Soc Trop Med Hyg 84: 363–366.
[26]  Molloy MP, Herbert BR, Walsh BJ, Tyler MI, Traini M, et al. (1998) Extraction of membrane proteins by differential solubilization for separation using two-dimensional gel electrophoresis. Electrophoresis 19: 837–844.
[27]  Santoni V, Kieffer S, Desclaux D, Masson F, Rabilloud T (2000) Membrane proteomics: use of additive main effects with multiplicative interaction model to classify plasma membrane proteins according to their solubility and electrophoretic properties. Electrophoresis 21: 3329–3344.
[28]  Ding AH, Nathan CF, Stuehr DJ (1988) Release of reactive nitrogen intermediates and reactive oxygen intermediates from mouse peritoneal macrophages. Comparison of activating cytokines and evidence for independent production. J Immunol 141: 2407–2412.
[29]  Samant M, Sahasrabuddhe AA, Singh N, Gupta SK, Sundar S, et al. (2007) Proteophosphoglycan is differentially expressed in sodium stibogluconate-sensitive and resistant Indian clinical isolates of Leishmania donovani. Parasitology 134: 1175–1184.
[30]  Glaser TA, Moody SF, Handman E, Bacic A, Spithill TW (1991) An antigenically distinct lipophosphoglycan on amastigotes of Leishmania major. Mol Biochem Parasitol 45: 337–344.
[31]  Charest H, Zhang WW, Matlashewski G (1996) The developmental expression of Leishmania donovani A2 amastigote-specific genes is post-transcriptionally mediated and involves elements located in the 3′-untranslated region. J Biol Chem 271: 17081–17090.
[32]  Kumari S, Samant M, Khare P, Sundar S, Sinha S, et al. (2008) Induction of Th1-type cellular responses in cured/exposed Leishmania-infected patients and hamsters against polyproteins of soluble Leishmania donovani promastigotes ranging from 89.9 to 97.1 kDa. Vaccine 26: 4813–4818.
[33]  Khalil EA, El Hassan AM, Zijlstra EE, Mukhtar MM, Ghalib HW, et al. (2000) Autoclaved Leishmania major vaccine for prevention of visceral leishmaniasis: a randomised, double-blind, BCG-controlled trial in Sudan. Lancet 356: 1565–1569.
[34]  Kahl LP, Scott CA, Lelchuk R, Gregoriadis G, Liew FY (1989) Vaccination against murine cutaneous leishmaniasis by using Leishmania major antigen/liposomes. Optimization and assessment of the requirement for intravenous immunization. J Immunol 142: 4441–4449.
[35]  Afonso LC, Scharton TM, Vieira LQ, Wysocka M, Trinchieri G, et al. (1994) The adjuvant effect of interleukin-12 in a vaccine against Leishmania major. Science 263: 235–237.
[36]  Champsi J, McMahon-Pratt D (1988) Membrane glycoprotein M-2 protects against Leishmania amazonensis infection. Infect Immun 56: 3272–3279.
[37]  Jaffe CL, Rachamim N, Sarfstein R (1990) Characterization of two proteins from Leishmania donovani and their use for vaccination against visceral leishmaniasis. J Immunol 144: 699–706.
[38]  White AC Jr, McMahon-Pratt D (1990) Prophylactic immunization against experimental Leishmania donovani infection by use of a purified protein vaccine. J Infect Dis 161: 1313–1314.
[39]  Melby PC (1991) Experimental leishmaniasis in humans: review. Rev Infect Dis 13: 1009–1017.
[40]  Mougneau E, Altare F, Wakil AE, Zheng S, Coppola T, et al. (1995) Expression cloning of a protective Leishmania antigen. Science 268: 563–566.
[41]  McMahon-Pratt D, Kima PE, Soong L (1998) Leishmania amastigote target antigens: the challenge of a stealthy intracellular parasite. Parasitol Today 14: 31–34.
[42]  Connell ND, Medina-Acosta E, McMaster WR, Bloom BR, Russell DG (1993) Effective immunization against cutaneous leishmaniasis with recombinant bacille Calmette-Guerin expressing the Leishmania surface proteinase gp63. Proc Natl Acad Sci U S A 90: 11473–11477.
[43]  Gicheru MM, Olobo JO (1994) Evaluation of recombinant gp63, the major Leishmania surface glycoprotein, as a diagnostic molecule for leishmaniasis in vervet monkeys. Acta Trop 58: 345–348.
[44]  Handman E (2001) Leishmaniasis: current status of vaccine development. Clin Microbiol Rev 14: 229–243.
[45]  Silvestre R, Cordeiro-da-Silva A, Tavares J, Sereno D, Ouaissi A (2006) Leishmania cytosolic silent information regulatory protein 2 deacetylase induces murine B-cell differentiation and in vivo production of specific antibodies. Immunology 119: 529–540.
[46]  Cabral SM, Silvestre RL, Santarem NM, Tavares JC, Silva AF, et al. (2008) A Leishmania infantum cytosolic tryparedoxin activates B cells to secrete interleukin-10 and specific immunoglobulin. Immunology 123: 555–565.
[47]  Trinchieri G (1995) Interleukin-12: a proinflammatory cytokine with immunoregulatory functions that bridge innate resistance and antigen-specific adaptive immunity. . Annu Rev Immunol 13: 251.
[48]  Sypek JP, Chung CL, Mayor SE, Subramanyam JM, Goldman SJ, et al. (1993) Resolution of cutaneous leishmaniasis: interleukin 12 initiates a protective T helper type 1 immune response. J Exp Med 177: 1797–1802.
[49]  Ghalib HW, Whittle JA, Kubin M, Hashim FA, el-Hassan AM, et al. (1995) IL-12 enhances Th1-type responses in human Leishmania donovani infections. J Immunol 154: 4623–4629.
[50]  Bloom S (2005) Parasite genome similarities offer hope for new drugs and vaccines. J Clin Invest 115: 2300–2301.
[51]  Tripathi P, Ray S, Sunder S, Dube A, Naik S (2006) Identification of Leishmania donovani antigens stimulating cellular immune responses in exposed immune individuals. Clin Exp Immunol 143: 380–388.
[52]  White AC Jr, Castes M, Garcia L, Trujillo D, Zambrano L (1992) Leishmania chagasi antigens recognized in cured visceral leishmaniasis and asymptomatic infection. Am J Trop Med Hyg 46: 123–131.
[53]  Costa SR, D'Oliveira A Jr, Bacellar O, Carvalho EM (1999) T cell response of asymptomatic Leishmania chagasi infected subjects to recombinant leishmania antigens. Mem Inst Oswaldo Cruz 94: 367–370.
[54]  Melby PC, Sacks DL (1989) Identification of antigens recognized by T cells in human leishmaniasis: analysis of T-cell clones by immunoblotting. Infect Immun 57: 2971–2976.
[55]  Gifawesen C, Farrell JP (1989) Comparison of T-cell responses in self-limiting versus progressive visceral Leishmania donovani infections in golden hamsters. Infect Immun 57: 3091–3096.
[56]  Rodrigues Junior V, Da Silva JS, Campos-Neto A (1992) Selective inability of spleen antigen presenting cells from Leishmania donovani infected hamsters to mediate specific T cell proliferation to parasite antigens. Parasite Immunol 14: 49–58.
[57]  Liew FY (1991) Role of cytokines in killing of intracellular pathogens. Immunol Lett 30: 193–197.
[58]  Assreuy J, Cunha FQ, Epperlein M, Noronha-Dutra A, O'Donnell CA, et al. (1994) Production of nitric oxide and superoxide by activated macrophages and killing of Leishmania major. Eur J Immunol 24: 672–676.
[59]  Diefenbach A, Schindler H, Rollinghoff M, Yokoyama WM, Bogdan C (1999) Requirement for type 2 NO synthase for IL-12 signaling in innate immunity. Science 284: 951–955.

Full-Text

Contact Us

service@oalib.com

QQ:3279437679

WhatsApp +8615387084133