Vaccination with recombinant Parasite Surface Antigen 2 from Leishmania major induces a Th1 type of immune response but does not protect against infection

Vaccination with the native Parasite Surface Antigen 2 of Leishmania major with Corynebacterium parvum as adjuvant protects mice from leishmaniasis through a Th1 mediated response. Here we show that vaccination with a recombinant form of this protein, purified from Escherichia coli and administered in iscoms or with C. parvum as adjuvant, does not induce protective immunity despite the induction of Th1 responses. The results suggest that protective immunity depends on the ability of the vaccinating antigen to induce Th1-like T cells with ability to be recalled by infection. Therefore, the conformation of antigens may play a more major role for the induction of T cell mediated immunity than originally considered.     

Leishmania major-infected C3H mice treated with anti-IL-12 mAb develop but do not maintain a Th2 response

Leishmania major-infected C3H mice develop a Th1 response, but studies have shown that treatment of C3H mice with anti-IL-12 or anti-IFN-gamma mAb promotes the development of a Th2 response and susceptibility. However, we discovered that C3H mice treated for 3 wk with either anti-IL-12 or anti-IFN-gamma mAb eventually resolved their lesions and switched from a Th2 to a Th1 response. No significant differences in IL-4, IL-10, or IFN-gamma levels or in the parasite burden could be detected between BALB/c and anti-IL-12-treated C3H mice early after infection, suggesting that the instability of the Th2 response in anti-IL-12-treated C3H mice was unrelated to levels of these cytokines and parasite numbers. However, anti-IL-12-treated C3H mice continued to produce IL-12 in spite of exhibiting a Th2 phenotype. To determine whether the production of IL-12 was associated with the healing observed in these animals, we treated C3H mice with anti-IL-12 continuously for 12 wk. In contrast to C3H mice given anti-IL-12 for 3 wk, C3H mice continuously treated with anti-IL-12 failed to heal. These results suggest that qualitative differences in Th2-type responses may influence their stability and that the presence of IL-4, IL-10, or high parasite numbers is not sufficient to maintain a Th2 response in mice with certain genetic backgrounds.     

Increased capacity for interleukin-2 synthesis parallels disease progression in mice infected with Leishmania major

Lymph node cells of BALB/c mice with progressive leishmaniasis produced sixfold more interleukin-2 (IL-2) in culture than those of healing C57BL/6 mice. IL-2 synthesis also increased in C57BL/6 mice made susceptible by IL-12 or gamma interferon deficiency. However, IL-2 mRNA levels in vivo did not reflect IL-2 production in vitro. Because IL-2 contributes to the pathogenesis of progressive leishmaniasis, the functional significance of these findings should be further explored.     

Fate of the promastigotes of Leishmania tropica major and L. gymnodactyli in the body of Phlebotomus papatasi under conditions of a mixed infection

We have shown that 8S and U2 fractions of low molecular weight nuclear RNA lmwn RNA) have tow stable conformational states with different electrophoretic mobilities. We have shown also that fractions UIa and UIb are conformational isomers. Fractions 8S and U2 are present only in one conformational state during electrophoresis, while UI RNA is always represented by both conformations. The conformation of lmwn RNAs is dependent upon conditions of isolation of nuclei and of extraction of RNA. It may change during storage of water solution of RNA at -20 degrees. Conformational changes discovered are not the result of degradation. Only one of two stable conformations of any lmwn RNA may be physiologically active, as it is the case of 5S rRNA and tRNAS. One ought to keep this in mind while studying the functions of low molecular weight nuclear RNAs.     

Leishmania mexicana cysteine proteinase-deficient mutants have attenuated virulence for mice and potentiate a Th1 response

Leishmania mexicana mutants lacking cysteine proteinase genes cpa (delta cpa), cpb (delta cpb), or both cpa and cpb (delta cpa/cpb) have been generated by targeted gene disruption. Delta cpa mutants produce a disease phenotype in BALB/c mice close to that of wild-type L. mexicana, but delta cpb mutants are much less infective, producing very slowly growing small lesions, and delta cpa/cpb double mutants do not induce lesion growth. Immunologic analysis of Ab isotype during infection and splenocyte IFN-gamma, IL-2, and IL-4 production following stimulation with Leishmania Ag or Con A indicates that there was a significant shift from a predominantly Th2-associated immune response in mice infected with wild-type L. mexicana to a Th1-associated response in mice inoculated with delta cpb or delta cpa/cpb. Significantly, delta cpa altered the balance of the immunologic response to a lesser extent than did the other mutants. Similar disease outcomes and switches in the Th1/Th2 balance were also observed when other L. mexicana-susceptible mouse strains were infected with the mutants. BALB/c and C57BL/6 mice vaccinated with delta cpa/cpb and CBA/Ca mice vaccinated with delta cpb or delta cpa/cpb were subsequently more resistant, to varying degrees, than were untreated mice to infection with wild-type parasites, as measured by development of lesions and parasite burden. These data implicate leishmanial cysteine proteinases not only as parasite virulence factors but also in modulation of the immune response and provide strong encouragement that cysteine proteinase-deficient L. mexicana mutants are candidate attenuated live vaccines.     

Parasite dose determines the Th1/Th2 nature of the response to Leishmania major independently of infection route and strain of host or parasite

Leishmania major causes cutaneous leishmaniasis in mice and man. Infection of mice with relatively low or high numbers of parasites leads respectively to parasite containment, associated with a Th1, cell-mediated response, or progressive disease, associated with a Th2, antibody response in all circumstances studied. These include different parasite strains, different routes of infection, and different hosts previously classified as susceptible, resistant or of intermediate susceptibility. This dose dependency appears to reflect a general rule. We argue that this rule may allow the design of a vaccination strategy that is effective among a genetically diverse population, and that it imposes severe constraints upon proposals for the nature of the "decision criterion" determining whether antigen induces a Th1 or Th2 response.     

Newborn mice develop balanced Th1/Th2 primary effector responses in vivo but are biased to Th2 secondary responses

Newborn mice are impaired in their abilities to mount protective immune responses. For decades, it was generally held that the poor responses of newborns were largely due to the developmentally immature state of the T cells. In vitro studies showing that neonatal T cells were deficient in Th1 cytokine production, proliferation, and secondary responsiveness strongly supported this idea. Recently, several studies have challenged this view; animals exposed to Ag as neonates were shown to have mature Th1 responses in adulthood. However, it is not clear whether the mature immune responses were actually mounted by T cells generated after the neonatal stage. We have reexamined this issue by analyzing the capabilities of neonatal lymph node T cells to develop into Ag-specific effector cells during the actual neonatal period. Our results demonstrate that the capacity to develop a balanced Th1/Th2 primary effector response is fully mature within the first week of life. However, while neonatal and adult primary cytokine profiles were very similar, Th2 secondary responses predominated in animals first immunized as newborns. Moreover, we have observed other differences between adults and neonatal responses, including 1) the kinetics of cytokine production and responsiveness to adjuvant during the primary response, and 2) the contribution of spleen and lymph node to secondary responses. We propose that these differences reflect developmental regulation of effector cell function that has important consequences to neonatal immune function.     

An in vitro model for infection with Leishmania major that mimics the immune response in mice

By using a primary in vitro response specific for Leishmania major, normal T cells from resistant CBA/CaH-T6J and susceptible BALB/c mice commit to a Th1 and a Th2 response, respectively. Since commitment occurred, we measured the production of gamma interferon (IFN-gamma), interleukin-1 (IL-1), IL-2, IL-4, IL-5, IL-10, and IL-12, prostaglandin E2 (PGE2), transforming growth factor beta (TGF-beta), and nitric oxide in the first 7 days of the response to identify factors that are critical for Th1 and Th2 development. While cells from resistant CBA mice produced more IFN-gamma, IL-10, and nitric oxide, cells from susceptible BALB/c mice produced more IL-1alpha, IL-5, PGE2, and TGF-beta. Although substantial amounts of IL-12 were detected, IL-12 did not associate with either Th1 or Th2 development. We did not anticipate that cells from resistant CBA mice would make more IL-10 in vitro. However, this also occurred in vivo since CBA mice produced substantial amounts of IL-10 following infection with L. major. Moreover, adding anti-IL-10 to primary in vitro responses enhanced production of IFN-gamma and nitric oxide by cells from CBA and BALB/c mice. Therefore, IL-10 cannot be regarded as a cytokine that associates with susceptibility to infection with L. major. Finally, the data presented here suggest that a collection of factors that can be produced by accessory cells influence Th commitment (e.g., IL-1, PGE2, and TGF-beta favor Th2 development).     

HIV-1 envelope protein gp120 triggers a Th2 response in mice that shifts to Th1 in the presence of human growth hormone

Immunization of mice with HIV-1-gp120 results in predominant activation of the Th2 lymphocyte subset, leading to enhanced IL-4 production. Administration of human growth hormone at the time of gp120 immunization provokes a change in the cytokine production pattern, with lower IL-4 and higher gamma-IFN and IL-2 synthesis levels, indicating a preferential switch in stimulation from Th2 to Th1 cells. A growth hormone would thus be of great use for pharmacological intervention in those cases in which an infectious microorganism evades immune defenses by provoking a Th2 response. In addition, the ability of growth hormone to induce a Th1-type response upon vaccination with an HIV-antigen should be examined in the development of new therapeutic strategies or in the design of novel vaccines against HIV infection.     

Th1 response in Salmonella typhimurium-infected mice with a high or low rate of bacterial clearance

Previous studies have shown that the capacity to clear an attenuated strain of Salmonella typhimurium after the second week of infection varies widely among mouse strains. Bacterial clearance is mediated by CD4+ T cells and is regulated in part by the H-2 complex. The aim of the present study was to compare the patterns of cytokine mRNA expression in the spleens of C57BL/6 (H-2b) and CBA (H-2k) mice, which exhibit a low and a high rate of bacterial clearance, respectively. A transient increase in interleukin-12 (IL-12) mRNA levels was found in both mouse strains. Gamma interferon (IFN-gamma) gene expression was higher and more sustained in C57BL/6 than in CBA mice. No increase in IL-4 mRNA was detected. A transient increase in IL-10 mRNA was found in C57BL/6 mice. Separation of spleen cells into CD4+ and CD4- fractions showed that CD4+ T cells produced the bulk of IFN-gamma in both mouse strains and of IL-10 in C57BL/6 mice. Infection of H-2 congenic mice induced a higher level of IFN-gamma mRNA expression by CD4+ T cells in mice with a low rate of clearance (H-2b) than in mice with a high rate of clearance (H-2q). Treatment of infected C57BL/6 mice with anti-IFN-gamma or anti-CD4 monoclonal antibodies indicated that IFN-gamma participates in resistance in the early phase of infection, but not in bacterial clearance, and that CD4+ T cells mediate bacterial clearance during the 3rd week of infection. Taken together, these results suggest that defective bacterial clearance in H-2b mice is not linked to defective IFN-gamma production and that CD4+ T cells mediate bacterial clearance by an IFN-gamma-independent mechanism.     

Unequal death in T helper cell (Th)1 and Th2 effectors: Th1, but not Th2, effectors undergo rapid Fas/FasL-mediated apoptosis

T helper cell (Th) 1, but not Th2, effectors undergo rapid Fas/Fas ligand (FasL)-mediated, activation-induced cell death upon restimulation with antigen. Unequal apoptosis is also observed without restimulation, after a longer lag period. Both effectors undergo delayed apoptosis induced by a non-Fas-mediated pathway. When Th1 and Th2 effectors are co-cultured, Th2 effectors survive preferentially, suggesting the responsible factor(s) is intrinsic to each population. Both Th1 and Th2 effectors express Fas and FasL, but only Th2 effectors express high levels of FAP-1, a Fas-associated phosphatase that may act to inhibit Fas signaling. The rapid death of Th1 effectors leading to selective Th2 survival provides a novel mechanism for differential regulation of the two subsets.     

AMPA receptor GluR2, but not GluR1, subunit deletion impairs emotional response conditioning in mice.

Deletions of gria1 or gria2 genes encoding alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic-acid-receptor subunits differ in their effects on appetitive conditioning. The authors investigated whether similar differences would occur in an aversive conditioning test. The ability of a discrete stimulus paired with footshock to subsequently inhibit food-maintained operant responding (conditioned emotional response) was examined in mice with deletions of gria1 or gria2 genes. Whereas gria1 knockout (KO) mice performed normally compared with wild-type (WT) controls, gria2 KO mice displayed no reduction in response rates when the shock-paired stimulus was presented. Nevertheless, gria2 KOs displayed evidence of freezing in a footshock-paired context, indicating that aversive learning could occur. In addition, gria1 KO mice showed some evidence of increased anxiety, and gria2 KOs showed reduced anxiety, in the elevated plus-maze. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Endogenous opioids modulate allograft rejection time in mice: possible relation with Th1/Th2 cytokines

Massive enlargement of an extracerebral cavernous malformation and extension across tissue planes is very uncommon. The authors present the case of a 49-year-old woman with a giant cavernous malformation in the left frontotemporal area. It progressively enlarged during several decades, extended through the calvaria to the extradural space, and was surgically treated. The lesion may have originated in the soft tissue or the skull. The locations of cavernous malformations in various parts of the body are reviewed and their mechanisms of growth are discussed. Surgical excision is the treatment of choice.     

The IL-4 rapidly produced in BALB/c mice after infection with Leishmania major down-regulates IL-12 receptor beta 2-chain expression on CD4+ T cells resulting in a state of unresponsiveness to IL-12

Within 1 day of infection with Leishmania major, susceptible BALB/c mice produce a burst of IL-4 in their draining lymph nodes, resulting in a state of unresponsiveness to IL-12 in parasite-specific CD4+ T cells within 48 h. In this report we examined the molecular mechanism underlying this IL-12 unresponsiveness. Extinction of IL-12 signaling in BALB/c mice is due to a rapid down-regulation of IL-12R beta2-chain mRNA expression in CD4+ T cells. In contrast, IL-12R beta2-chain mRNA expression was maintained on CD4+ T cells from resistant C57BL/6 mice. The down-regulation of the IL-12R beta2-chain mRNA expression in BALB/c CD4+ T cells is a consequence of the early IL-4 production. In this murine model of infection, a strict correlation is shown in vivo between expression of the IL-12R beta2-chain in CD4+ T cells and the development of a Th1 response and down-regulation of the mRNA beta2-chain expression and the maturation of a Th2 response. Treatment of BALB/c mice with IFN-gamma, even when IL-4 has been produced for 48 h, resulted in maintenance of IL-12R beta2-chain mRNA expression and IL-12 responsiveness. The data presented here support the hypothesis that the genetically determined susceptibility of BALB/c mice to infection with L. major is primarily based on an up-regulation of IL-4 production, which secondarily induces extinction of IL-12 signaling.     

The identity of Leishmania isolated from sand flies and vertebrate hosts in a major focus of cutaneous leishmaniasis in Baturite, northeastern Brazil

The oxidation states of intracellular myoglobin and cytochrome oxidase aa3 were monitored by reflectance spectrophotometry in isolated perfused rat hearts subjected to an acutely magnesium deficient environment. After exposure to low extracellular [Mg2+]o (i.e., 0.3 mM) for 30 min, more than 80% of the oxymyoglobin converted to its deoxygenated form. The level of reduced cytochrome oxidase aa3 also increased about 80% in low [Mg2+]o. The deoxymyoglobin was converted further to a species identified as ferrylmyoglobin by its reaction with Na2S to form ferrous sulfmyoglobin which was optically visible. This process, set into motion by acute Mg deficiency, resulted from a direct accessibility of the exogenous peroxide to the cytosolic protein. The results suggest that a pathway leading to cardiac tissue damage, induced by magnesium deficiency, is probably involved in the generation of a ferrylmyoglobin radical which could be prevented by addition of ascorbate, which is known to be a one-electron reductant of this hypervalent form of myoglobin. In further studies, we also investigated whether addition of different concentrations of ascorbic acid (AA) to the perfusate could enhance myocardial function after exposure to low [Mg2+]o perfusion. Four concentrations of AA (0.5, 1, 5, 10 mM) were tested, and the results indicate that they exert their effects in a concentration-dependent manner; 1 mM AA was the most effective dose in improving aortic output in a Mg-deficient heart. Ferrylmyoglobin formation was found to be formed considerably before intracellular release of either creatine phosphokinase or lactic dehydrogenase. These studies may have wide implications as a new mechanism by which low extracellular Mg2+ can induce myocardial injury and subsequent cardiac failure.     

Reconstitution of C.B-17 scid mice with BALB/c T cells initiates a T helper type-1 response and renders them capable of healing Leishmania major infection

C.B-17 scid mice, which were found to be very susceptible to infection with Leishmania major, were reconstituted with various doses of T cells, T plus B cells or unfractionated spleen cells from nonhealer BALB/c mice. All reconstitution protocols, except for the transfer of very high numbers of BALB/c spleen cells, led to a spontaneously healing infection and resistance to reinfection, rather than the lethal, nonhealing infection typical of BALB/c mice. These healing responses were associated with a strong T helper 1 (Th1)-like response characterized by delayed-type hypersensitivity (DTH) responsiveness, but no elevation of serum IgE, and by the production of high levels of interferon-gamma (IFN-gamma), but no interleukin-4 (IL-4) by lymph node and spleen cells after restimulation with antigen in vitro. The development of this Th1 response from BALB/c Th cells requires IFN-gamma during the initial infection period. Treatment of scid mice with a single injection of neutralizing anti-IFN-gamma antibody prior to infection and reconstitution prevented healing and permitted the development of a Th-2 like response as indicated by elevated serum IgE, but no DTH, and by the production of IL-4, but very little IFN-gamma, after antigen stimulation in vitro. As few as 10(4) transferred T cells led to a Th1-like response, suggesting that the IFN-gamma is of host rather than donor origin. The transfer of very high numbers (7.5 x 10(7)) of BALB/c spleen cells overcame the effects of the IFN-gamma and led to the nonhealing infection and cytokine pattern characteristic of BALB/c mice. The enrichment or depletion of B cells from the transferred T cells had no measurable effect upon the development of a healing response in reconstituted scid mice.     

Mice defective in Fas are highly susceptible to Leishmania major infection despite elevated IL-12 synthesis, strong Th1 responses, and enhanced nitric oxide production

MRL/MP-lpr/lpr (MRL/lpr) mice have a single mutation (lpr) of the fas apoptosis gene. The mutant mice developed significantly smaller lesions than the wild-type mice at the earlier stage of infection with the intracellular protozoan parasite Leishmania major. However, while all the wild-type mice achieved complete lesion resolution, the disease in the mutant mice progressed inexorably. The mutant mice had more IL-12 and nitrite/nitrate in the serum than wild-type mice following infection. Lymphoid cells from infected MRL/lpr mice produced more IFN-gamma but less IL-4 and IL-5 than cells from MRL-+/+ mice. Peritoneal macrophages from the mutant mice also produced more IL-12 and NO after stimulation with LPS. Thus, Fas expression is essential for resistance against leishmaniasis, and Fas-mediated apoptosis may form an integral part of the Th1-mediated microbicidal function.