Peripheral blood lymphocyte phenotypes in Alzheimer and Parkinson's diseases.

INTRODUCTION: Neuroinflammation is involved in the pathophysiology of various neurological disorders, in particular Alzheimer disease (AD) and Parkinson's disease (PD). Alterations in the blood-brain barrier may allow peripheral blood lymphocytes to enter the central nervous system; these may participate in disease pathogenesis. OBJECTIVE: To evaluate the peripheral blood lymphocyte profiles of patients with AD and PD and their association with the disease and its progression. METHODS: The study included 20 patients with AD, 20 with PD, and a group of healthy individuals. Ten of the patients with AD and 12 of those with PD were evaluated a second time 17 to 27 months after the start of the study. Lymphocyte subpopulations and their activation status were determined by flow cytometry. All patients underwent neurological examinations using internationally validated scales. RESULTS: Compared to healthy individuals, patients with AD and PD showed significantly higher levels of activated lymphocytes, lymphocytes susceptible to apoptosis, central memory T cells, and regulatory T and B cells. As the diseases progressed, there was a significant decrease in activated cells (CD4+ CD38+ and CD8+ CD38+ in PD and AD, CD4+ CD69+ and CD8+ CD69+ in PD), T cells susceptible to apoptosis, and some regulatory populations (CD19+ CD5+ IL10+ in PD and AD, CD19+ CD5+ IL10+ FoxP3+, CD4+ FoxP3+ CD25+ CD45RO+ in PD). In patients with AD, disease progression was associated with lower percentages of CD4+ CD38+ cells and higher percentages of effector CD4 cells at the beginning of the study. Significant differences were observed between both diseases. CONCLUSIONS: This study provides evidence of changes in peripheral blood lymphocyte phenotypes associated with AD and PD and their severity. Considering effective blood-brain communication, our results open new avenues of research into immunomodulation therapies to treat these diseases.

Peripheral blood lymphocyte phenotypes in Alzheimer and Parkinson's diseases. / Fenotipos de linfocitos periféricos en las enfermedades de Alzheimer y Parkinson.

INTRODUCTION: Neuroinflammation is involved in the pathophysiology of various neurological disorders, in particular Alzheimer disease (AD) and Parkinson's disease (PD). Alterations in the blood-brain barrier may allow peripheral blood lymphocytes to enter the central nervous system; these may participate in disease pathogenesis. OBJECTIVE: To evaluate the peripheral blood lymphocyte profiles of patients with AD and PD and their association with the disease and its progression. METHODS: The study included 20 patients with AD, 20 with PD, and a group of healthy individuals. Ten of the patients with AD and 12 of those with PD were evaluated a second time 17 to 27 months after the start of the study. Lymphocyte subpopulations and their activation status were determined by flow cytometry. All patients underwent neurological examinations using internationally validated scales. RESULTS: Compared to healthy individuals, patients with AD and PD showed significantly higher levels of activated lymphocytes, lymphocytes susceptible to apoptosis, central memory T cells, and regulatory T and B cells. As the diseases progressed, there was a significant decrease in activated cells (CD4+ CD38+ and CD8+ CD38 + in PD and AD, CD4+ CD69+ and CD8+ CD69+ in PD), T cells susceptible to apoptosis, and some regulatory populations (CD19+ CD5+ IL10+ in PD and AD, CD19+ CD5+ IL10+ FoxP3+, CD4+ FoxP3+ CD25+ CD45RO+ in PD). In patients with AD, disease progression was associated with lower percentages of CD4+ CD38+ cells and higher percentages of effector CD4 cells at the beginning of the study. Significant differences were observed between both diseases. CONCLUSIONS: This study provides evidence of changes in peripheral blood lymphocyte phenotypes associated with AD and PD and their severity. Considering effective blood-brain communication, our results open new avenues of research into immunomodulation therapies to treat these diseases.

Impact of the GK-1 adjuvant on peritoneal macrophages gene expression and phagocytosis.

PURPOSE: The synthetic peptide GK-1 potentiates protective immunity elicited by the influenza vaccine in mice. In order to understand its adjuvant properties, this study was designed to determine the impact of GK-1 on gene expression and phagocytosis of peritoneal macrophages (PMa). METHODS: Increased gene expression of chemokines involved in leukocyte recruitment and of pro-inflammatory mediators was detected by microarray analysis of control and GK-1 treated PMa macrophages. The expression profile was subsequently confirmed by Multiplex Immunoassays analysis to measure cytokines levels, flow cytometer to describe M1/M2 surface markers and an assay to evaluate their phagocytic activity. RESULTS: Treatment of PMa with GK-1 results in development to the classically activated M1 functional macrophage subpopulation with increased expression of the CCL3 and CXCLO2 chemokines, IL-6 and TNF-α proinflammatory cytokines with a concomitant increase in the levels of NO, accompanied by the expression of modulatory factors that downregulate the inflammatory phenotype. GK-1 treated PMa significantly increased their phagocytic activity. CONCLUSION: GK-1 classical activated with enhanced phagocitic capacity may underlie in the increased specific immunity induced when concomitant administered with other antigens.

No association of IL2, IL4, IL6, TNF, and IFNG gene polymorphisms was found with Taenia solium human infection or neurocysticercosis severity in a family-based study.

Neurocysticercosis (NC) is caused by the establishment of the metacestode stage of Taenia solium in the human central nervous system. A great heterogeneity in the susceptibility to the infection and to the disease has been reported. While the factors involved in this heterogeneity are not completely understood, clearly different immune-inflammatory profiles have been associated to each condition. This study evaluated the association of cytokine single nucleotide polymorphisms (SNPs) with susceptibility to infection and disease severity in NC patients. Blood samples from 92 NC cases and their parents (trios) were genotyped for SNPs in five cytokines relevant for the immune response: IL4 (-589C/T), IL6 (-174C/G), IFNG (+874T/A), TNF (-238G/A), and IL2 (-330G/T). Specific DNA fragments were amplified by the polymerase chain reaction, using the 5'-nuclease Taqman assay on a 7500 platform, allowing the detection of the polymorphism genotypes. No association between the polymorphisms evaluated neither with susceptibility to infection nor with disease severity was found, although previous studies reported variations in the levels of these cytokines among different NC clinical pictures. These results, nevertheless, add new elements to our understanding of the complex pathogenic mechanisms involved in susceptibility to infection by T. solium cysticerci and the severity of the ensuing disease.

GK-1 peptide reduces tumor growth, decreases metastatic burden, and increases survival in a murine breast cancer model.

GK-1 is a parasite-derived peptide adjuvant of 18 amino acid-length that enhances T-cell function and increases survival in B16-F10 melanoma tumor-bearing mice. This study was designed to evaluate in vivo the antitumor efficacy of GK-1 on 4T1 mouse mammary carcinoma. BALB/c mice with palpable primary tumors were weekly intravenously injected three times with saline solution or three different concentrations (10, 50, or 100µg per mouse) of GK-1. GK-1 significantly increased lifespan (p<0.0001) and reduced the primary tumor weight (p=0.014) and volume (p<0.0001) with respect to control mice, with no statistically significant differences among GK-1 doses. At the primary tumor, we found increased necrotic areas associated with a reduction in tumor mass, as well as an increase in the antitumor cytokine IL-12. Especially encouraging is the ability of GK-1 to reduce the number of lung metastasis (p=0.006) disregarding the dose used. The participation of IL-6 in metastasis development and the decreased levels of CCL-2, CCL-3, TNF-α, CXCL-9, GM-CSF, and b-FGF found in lungs of GK-1-treated mice is discussed. Our study supports the effectiveness of GK-1 as an antineoplastic agent that merits further exploration in combination with other therapeutic approaches in future translational studies.

Intranasal delivery of dexamethasone efficiently controls LPS-induced murine neuroinflammation.

Neuroinflammation is the hallmark of several infectious and neurodegenerative diseases. Synthetic glucocorticoids (GCs) are the first-line immunosuppressive drugs used for controlling neuroinflammation. A delayed diffusion of GCs molecules and the high systemic doses required for brain-specific targeting lead to severe undesirable effects, particularly when lifelong treatment is required. Therefore, there is an urgent need for improving this current therapeutic approach. The intranasal (i.n.) route is being employed increasingly for drug delivery to the brain via the olfactory system. In this study, the i.n. route is compared to the intravenous (i.v.) administration of GCs with respect to their effectiveness in controlling neuroinflammation induced experimentally by systemic lipopolysaccharide (LPS) injection. A statistically significant reduction in interleukin (IL)-6 levels in the central nervous system (CNS) in the percentage of CD45+ /CD11b+ /lymphocyte antigen 6 complex locus G6D [Ly6G+ and in glial fibrillary acidic protein (GFAP) immunostaining was observed in mice from the i.n.-dexamethasone (DX] group compared to control and i.v.-DX-treated animals. DX treatment did not modify the percentage of microglia and perivascular macrophages as determined by ionized calcium binding adaptor molecule 1 (Iba1) immunostaining of the cortex and hippocampus. The increased accumulation of DX in brain microvasculature in DX-i.n.-treated mice compared with controls and DX-IV-treated animals may underlie the higher effectiveness in controlling neuroinflammation. Altogether, these results indicate that IN-DX administration may offer a more efficient alternative than systemic administration to control neuroinflammation in different neuropathologies.

Electric stimulation of the vagus nerve reduced mouse neuroinflammation induced by lipopolysaccharide.

BACKGROUND: Neuroinflammation (NI) is a key feature in the pathogenesis and progression of infectious and non-infectious neuropathologies, and its amelioration usually improves the patient outcome. Peripheral inflammation may promote NI through microglia and astrocytes activation, an increased expression of inflammatory mediators and vascular permeability that may lead to neurodegeneration. Several anti-inflammatory strategies have been proposed to control peripheral inflammation. Among them, electrical stimulation of the vagus nerve (VNS) recently emerged as an alternative to effectively attenuate peripheral inflammation in a variety of pathological conditions with few side effects. Considering that NI underlies several neurologic pathologies we explored herein the possibility that electrically VNS can also exert anti-inflammatory effects in the brain. METHODS: NI was experimentally induced by intraperitoneal injection of bacterial lipopolysaccharide (LPS) in C57BL/6 male mice; VNS with constant voltage (5 Hz, 0.75 mA, 2 ms) was applied for 30 s, 48 or 72 h after lipopolysaccharide injection. Twenty four hours later, pro-inflammatory cytokines (IL-1ß, IL-6, TNFα) levels were measured by ELISA in brain and spleen extracts and total brain cells were isolated and microglia and macrophage proliferation and activation was assessed by flow cytometry. The level of ionized calcium binding adaptor molecule (Iba-1) and glial fibrillary acidic protein (GFAP) were estimated in whole brain extracts and in histologic slides by Western blot and immunohistochemistry, respectively. RESULTS: VNS significantly reduced the central levels of pro-inflammatory cytokines and the percentage of microglia (CD11b/CD45low) and macrophages (CD11b/CD45high), 24 h after the electrical stimulus in LPS stimulated mice. A significantly reduced level of Iba-1 expression was also observed in whole brain extracts and in the hippocampus, suggesting a reduction in activated microglia. CONCLUSIONS: VNS is a feasible therapeutic tool to attenuate the NI reaction. Considering that NI accompanies different neuropathologies VNS is a relevant alternative to modulate NI, of particular interest for chronic neurological diseases.

Interleukin 10 and dendritic cells are the main suppression mediators of regulatory T cells in human neurocysticercosis.

Neurocysticercosis is caused by the establishment of Taenia solium cysticerci in the central nervous system. It is considered that, during co-evolution, the parasite developed strategies to modulate the host's immune response. The action mechanisms of regulatory T cells in controlling the immune response in neurocysticercosis are studied in this work. Higher blood levels of regulatory T cells with CD4(+) CD45RO(+) forkhead box protein 3 (FoxP3)(high) and CD4(+) CD25(high) FoxP3(+) CD95(high) phenotype and of non-regulatory CD4(+) CD45RO(+) FoxP3(med) T cells were found in neurocysticercosis patients with respect to controls. Interestingly, regulatory T cells express higher levels of cytotoxic T lymphocyte antigen 4 (CTLA-4), lymphocyte-activation gene 3 (LAG-3), programmed death 1 (PD-1) and glucocorticoid-induced tumour necrosis factor receptor (GITR), suggesting a cell-to-cell contact mechanism with dendritic cells. Furthermore, higher IL-10 and regulatory T cell type 1 (Tr1) levels were found in neurocysticercosis patients' peripheral blood, suggesting that the action mechanism of regulatory T cells involves the release of immunomodulatory cytokines. No evidence was found of the regulatory T cell role in inhibiting the proliferative response. Suppressive regulatory T cells from neurocysticercosis patients correlated negatively with late activated lymphocytes (CD4(+) CD38(+) ). Our results suggest that, during neurocysticercosis, regulatory T cells could control the immune response, probably by a cell-to-cell contact with dendritic cells and interleukin (IL)-10 release by Tr1, to create an immunomodulatory environment that may favour the development of T. solium cysticerci and their permanence in the central nervous system.

Immunopathology in Taenia solium neurocysticercosis.

Neurocysticercosis is a clinically and radiologically heterogeneous disease, ranging from asymptomatic infection to a severe, potentially fatal clinical picture. The intensity and extension of the parasite-elicited inflammatory reaction is a key factor for such variability. The main features of the inflammatory process found in the brain and in the peripheral blood of neurocysticercosis patients will be discussed in this review, and the factors involved in its modulation will be herein presented.

Influenza vaccine: development of a novel intranasal and subcutaneous recombinant adjuvant.

The synthetic peptide GK-1, derived from Taenia crassiceps, enhances the protection induced by human influenza vaccine in both young and aged mice. Herein, the adjuvant properties of GK-1 fused to the pVIII protein of a heat-inactivated phagemid vector (FGK1) when co-administered with the influenza vaccine were assessed, to evaluate its feasibility as a low-cost adjuvant. In mice, FGK1 significantly increased the expected IgG and IgA anti-influenza antibody levels both in sera and in bronchoalveolar fluids when intranasally or subcutaneously co-administered with influenza vaccine. Single-dose pig co-immunization with FGK1 and influenza vaccine induced serum levels of IgG anti-influenza antibodies similar to those elicited by a two-dose immunization with the influenza vaccine alone. Preclinical evaluation of FGK1 with the influenza vaccine is currently in progress, in order to recommend its use for veterinary purposes.

Limits of the therapeutic properties of synthetic S3Pvac anti-cysticercosis vaccine.

The S3Pvac synthetic vaccine, composed of three peptides (GK1, KETc1 and KETc12) effectively protects against cysticercosis under experimental and field conditions. Additionally, S3Pvac vaccine can effectively damage early-established cysticerci in experimentally lightly infected young pigs. This study was designed to explore if also fully-developed cysticerci that eluded immunity induced by the infection can be damaged by S3Pvac-induced immunity in naturally, heavily infected adult pigs. Fourteen pigs identified as cysticercotic by tongue inspection from rural communities were purchased and moved to controlled conditions in the Faculty of Veterinary Medicine. Half of these pigs were treated once a month three times with S3Pvac plus saponin, and the other half received only saponin (controls). Twelve months later pigs were euthanized, and the number of cysticerci, their macro and microscopic status and their capacity to transform into tapeworms were determined. S3Pvac failed to damage fully-developed muscle cysticerci of naturally, heavily infected adult pigs. To explore possible factors involved in the failure of the therapeutic capacity pooled sera from control and treated cysticercotic pigs were added to mice mononuclear peripheral cells. Pooled sera from non-infected pigs were also tested. Sera from control and treated infected pigs almost completely suppressed the T cell proliferative responses, pointing to the presence of suppressor factors. In conclusion, S3Pvac vaccine failed to damage fully-developed cysticerci in pigs in which a host parasite relationship had evolved after months of infection with immunological implications.

Preferential growth of Taenia crassiceps cysticerci in female mice holds across several laboratory mice strains and parasite lines.

A retrospective study of our 14-yr records on experimental Taenia crassiceps (ORF(fast) line) cysticercosis (n = 1,198) shows that in 16 of 17 different mice strains, female mice are more frequently infected and carry larger individual parasite loads than males. However, sexual differences in parasite loads significantly varies between strains in relation to their different genetic backgrounds (BALB > C57Bl = OTHERS > C3H). The coefficient of variation in all female mice is significantly smaller than that of all males, an indication of males' more potent, but erratically effective, restraint of cysticercus growth. Similar positive growth bias for female mice is shown by other lines of cysticerci, i.e., HYG(slow) and WFU(slow). These results contravene the usual expectation of female hosts being more resistant than males to parasite infections, and they point to the multiple factors that combined determine sex related differences of mice to experimental cysticercosis infection.

Neurocysticercosis: detection of Taenia solium DNA in human cerebrospinal fluid using a semi-nested PCR based on HDP2.

Human neurocysticercosis (NC) is caused by Taenia solium larvae lodged in the central nervous system. This disease is usually diagnosed by radiology but the results are not always clear-cut and so immunological assays are often also used. A semi-nested PCR, based on the non-coding HDP2 sequence of T. saginata, has now been developed for detecting DNA from T. solium cysticerci and confirming NC. This PCR, which amplifies a 171-bp T. solium product, allowed the specific detection of just 174 attograms of T. solium DNA. The efficacy of the PCR was tested using cerebrospinal fluid (CSF) from neurological patients, including 46 confirmed Mexican cases of NC and 32 patients from non-endemic Spain. Eighteen of the confirmed cases [including 10 (71%) of the 14 with vesicular extraparenchymal cysticerci and four (17%) of the 24 with damaged cysticerci] and two (33%) of the six patients with 'uncertain' diagnosis (in whom a diagnosis of NC could not be established by radiological and immunological studies) were found PCR-positive. The 36 patients known to have neurological problems other than NC were found PCR-negative. The HDP2 PCR offers a new tool in the diagnosis of NC and in exploring the pathogenesis of this serious disease.

Human and porcine neurocysticercosis: differences in the distribution and developmental stages of cysticerci.

OBJECTIVE: To describe and compare the clinical impacts of neurocysticercosis (NC) caused by Taenia solium in humans and pigs. METHODS: Comparative study of the brains of 16 asymptomatic pigs and 35 human NC cases (15 asymptomatic and 20 symptomatic). RESULTS: In humans, cysticerci were more frequently located in the ventricles and subarachnoid space at the base of the brain (11.8%vs. 1.6%; P = 0.001 and 25.9%vs. 0%; P < 0.0001, respectively) while in pigs, cysticerci were more frequently found in the parenchyma (44.4%vs. 7.6%; P < 0.0001). In human brains, 75.9% of the cysticerci were calcified, while in pigs all cysticerci were in the vesicular stage. CONCLUSION: The duration of infection and the host-parasite relationship (such as immune reactivity and brain haemodynamics) differ between humans and pigs. This may account for the different distribution and stage of the cysticerci among humans and pigs.

Parasite contamination of soil in households of a Mexican rural community endemic for neurocysticercosis.

High neurocysticercosis (NC) prevalence was recently determined by a computed tomography (CT) scan study in the community of Tepetzitzintla, State of Puebla, Mexico. The aim of the present work was to evaluate the magnitude of fecal and parasite contamination by Taenia spp. in the soil of households of this community during the four seasons of the year. The toilet, backyard, kitchen, washboard, water containers and corrals of 14 to 26 households were sampled during each season. High Taenia spp. egg intensity was found in 24.2% of the sampled areas. The highest percentage was detected in Spring and the lowest in Summer. Significantly higher levels of Taenia spp. eggs were present in kitchen soil samples. A significant correlation was found between the presence of Taenia spp. eggs in household soil during the Summer, and NC diagnoses of the inhabitants by CT scan. Coproparasitological examinations and anti-cysticercal antibodies were determined in a cohort of inhabitants of the sampled households. Antibody levels and coproparasitological results were not associated with NC. Overall, these results illustrate the high degree of fecal contamination of potential risk to human health in rural communities and could be of use for control programmes.

The immune response in Taenia solium cysticercosis: protection and injury.

This article reviews current knowledge on the innate and acquired immune responses in human Taenia solium neurocysticercosis, highlighting the conditions that appear to be favourable for the survival or destruction of the parasite and for the benefit or injury to its host.

Impact of naturally acquired Taenia solium cysticercosis on the hormonal levels of free ranging boars.

In chronically infected BALBc/AnN male mice, Taenia crassiceps cysticercosis induces changes in the host's sex steroids hormone that lead to their estrogenization and deandrogenization, with possible repercussions on their susceptibility to infections. Here reported are the serum steroid levels in free range cysticercotic male boars. Therefore, the possible effects of Taenia solium cysticerci over the pig steroid levels were evaluated. Herein are described the sex steroids and cortisol levels of non-cysticercotic (n=25) and cysticercotic (n=22) adult boars, as diagnosed by tongue inspection, all free-ranging in a typical village of an endemic rural area in Mexico. A significant reduction of testosterone (P=0.022) and a likely one of 17beta-estradiol (P=0.08) levels were found in the cysticercotic boars in comparison with those non-cysticercotic, whilst no significant differences in the cortisol and DHEA levels were detected. Serum levels of specific antibodies did not correlate with infection nor with the levels of any of the hormones measured. Results suggest that T. solium cysticercosis significantly affects the hormonal status of its porcine host independently of their antibody response.

Detection of HP10 antigen in serum for diagnosis and follow-up of subarachnoidal and intraventricular human neurocysticercosis.

INTRODUCTION: Neurocysticercosis (NC), a parasitic disease caused by Taenia solium, may be either asymptomatic or show a mild to severe clinical picture with intracranial hypertension. The most severe form of the disease is caused when viable cysticerci are localised in the ventricles or in subarachnoidal cisterns at the base of the skull. Detection of the secreted metacestode antigen HP10 in cerebrospinal fluid is a sensitive and specific method for the diagnosis of these severe NC cases. OBJECTIVE AND METHODS: To evaluate the validity of HP10 antigen detection ELISA when applied to serum, using paired serum and cerebrospinal fluid samples from 116 radiologically and clinically characterised NC patients. RESULTS: The HP10 antigen assay exhibited a similarly high sensitivity in identifying severe NC cases from sera (84.8%) and CSF (91.3%). In contrast, HP10 antigen was rarely detected in asymptomatic or mild NC cases (3 of 57). Importantly, the HP10 antigen assay applied to serum showed high specificity (94%) when used in 126 serum samples of non-NC subjects from an endemic community with a confirmed coproparasitological diagnosis of intestinal parasitic infections. Finally, the HP10 assay also proved to be of value in the follow-up of treated patients. CONCLUSION: This study confirms that detection of the metacestode HP10 antigen in serum is a useful tool for diagnosis and follow-up of patients with severe forms of NC treated with cysticidal drugs.

Further evaluation of the synthetic peptide vaccine S3Pvac against Taenia solium cysticercosis in pigs in an endemic town of Mexico.

Taenia solium cysticercosis is a parasitic disease frequently affecting human health and the pig industry in many developing countries. A synthetic peptide vaccine (designated S3Pvac) against porcine cysticercosis has been developed previously as an aid to interrupt transmission and has been shown to be effective. The results of the present study support the effectiveness of the vaccine under endemic field conditions. However, given the time-frame of the vaccination trial, no changes in the local levels of transmission were detectable before and after vaccination using sentinel pigs. Thus, this investigation shows the limited usefulness of single vaccination as the sole means of interrupting Taenia solium transmission in an endemic region.