The combined action of glycoinositolphospholipid from Trypanosoma cruzi and macrophage migration inhibitory factor increases proinflammatory mediator production by cardiomyocytes and vascular endothelial cells.

Cardiomyopathy is the most serious complication of chronic Chagas disease, caused by infection with the protozoan Trypanosoma cruzi. Exacerbated inflammation of the myocardium constitutes a major pathologic component of the disease. In the myocardial microenvironment, parasite antigens and host inflammatory mediators may aggravate tissue damage. The glycoinositolphospholipid (GIPL) from T. cruzi is an inflammation-eliciting antigen recognized by Toll-like receptor 4 (TLR4), whereas the proinflammatory cytokine macrophage migration inhibitory factor (MIF) promotes progression of chronic Chagas cardiomyopathy. We herein aimed to examine the involvement of GIPL and MIF in molecular mechanisms leading to a pathogenic inflammatory response in HL-1 cardiomyocytes and HMEC microvascular endothelial cells. Immunofluorescence analysis revealed that GIPL enhanced TLR4 expression in both cell types. We found that TLR4/GIPL interaction and MIF activity modulated the arachidonic acid pathway implicated in persistent inflammation. The combination of GIPL at 50 µg/ml and MIF at 50 ng/ml upregulated type 2 cyclooxygenase (COX-2) levels in HL-1 and HMEC cells, in a stronger way than each molecule acting independently. Moreover, increased expression of prostanoid synthases and release of prostaglandin E2 (PGE2) and thromboxane B2 (TxB2) were detected in stimulated cells. Transfection experiments in HL-1 and HMEC cells showed that COX-2 induction was transcriptionally regulated through GIPL-TLR4 engagement and NFκB signaling cascade. (GIPL + MIF)-triggered NFκB activation was markedly attenuated by treatment with 100 µM Fenofibrate, a PPAR-α ligand. Fenofibrate reduced COX-2-dependent generation of bioactive lipids in HL-1 and HMEC cells. In addition, Fenofibrate abolished (GIPL + MIF)-fostered release of NO, IL-1ß, IL-6, TNF-α, and CCL2. The combined actions of GIPL and MIF display potential for amplifying the inflammatory response in myocardium of parasite-infected hosts. Our current findings might help develop more effective measures to ameliorate cardiovascular abnormalities associated with Chagas heart disease.

Long-term effectiveness and adverse effects of ketogenic diet therapy in infants with drug-resistant epilepsy treated at a single center in Argentina.

INTRODUCTION: Ketogenic diet therapy (KDT) is a metabolic treatment with proven effectiveness for the treatment of drug-resistant epilepsy in children. Although previously not used in infants under 2 years of age, recent studies have shown KDT to be highly effective and well tolerated in infants with epilepsy, especially those with epileptic encephalopathies. Here, we describe the effectiveness and tolerability of the diet in infants up to 2 years of age. MATERIAL AND METHODS: A prospective study was conducted in a cohort of infants younger than 2 years of age with drug-resistant epilepsy who received the classic ketogenic diet using a specific protocol at a single center in Argentina. RESULTS: 56 infants with treatment-refractory epilepsy were evaluated. The etiology was genetic in 21.4%, structural in 28.6%, unknown in 44.7%, and metabolic in 5.4%. At 3 months, a > 50% decrease in seizure frequency was observed in 35 patients (62.4%), of whom 11 (19.6%) became seizure free. At 6 months, 34 patients (60.7%) had a decrease in seizure frequency of > 50%, of whom 10 (17.8%) were seizure free. At the one-year follow-up, 27 patients (48.2%) had a > 50% decrease in seizure frequency, of whom six (10.7%) were seizure free. At two years, 14 patients (25%) had a > 50% seizure control, of whom four (7.1%) were seizure free. The most common early adverse effects were hypoglycemia and vomiting, while after 1 month and beyond metabolic acidosis, vomiting, and constipation more commonly found. A trend towards a higher rate of acute adverse events in infants younger than 1 year was observed. CONCLUSIONS: CKD showed to be a useful option in infants with treatment-resistant epilepsy. Adverse effects were common, but not a reason to discontinue the diet. Further studies are necessary to evaluate in which epilepsy syndromes and etiologies KDT is most effective.

Dual chemotherapy with benznidazole at suboptimal dose plus curcumin nanoparticles mitigates Trypanosoma cruzi-elicited chronic cardiomyopathy.

Curcumin (Cur) is a natural polyphenolic flavonoid isolated from the rhizomes of Curcuma longa. Its anti-inflammatory and cardioprotective properties are increasingly considered to have beneficial effects on the progression of cardiomyopathy associated with Chagas disease, caused by Trypanosoma cruzi. However, the Cur therapeutic limitation is its bioavailability and new Cur nanomedicine formulations are developed to overcome this obstacle. In this research, we provide evidence showing that oral therapy with a suboptimal dose of the standard parasiticidal drug benznidazole (BZ) in combination with Cur-loaded nanoparticles is capable of reducing myocardial parasite load, cardiac hypertrophy, inflammation and fibrosis in mice with long-term infection by T. cruzi. Treatment with BZ plus Cur was highly effective in downregulating myocardial expression of proinflammatory cytokines/chemokines (IL-1ß, TNF-α, IL-6, CCL5), and the level/activity of matrix metalloproteinases (MMP-2, MMP-9) and inducible enzymes (cyclooxygenase, nitric oxide synthase) implicated in leukocyte recruitment and cardiac remodeling. Oral administration of a Cur-based nanoformulation displays potential as a complementary strategy to the conventional BZ chemotherapy in the treatment of chronic Chagas heart disease.

Short-term changes in ocular surface signs and symptoms after phacoemulsification.

PURPOSE: The aim of this study is to evaluate the incidence, natural course, and distribution pattern of superficial punctate keratopathy and describe the changes in signs and symptoms of dry eye after cataract surgery. SETTING: The setting of this study is University Hospital Rio Hortega and Instituto Universitario de Oftalmobiología Aplicada, Valladolid, Spain. DESIGN: This is a prospective interventional study. MATERIALS AND METHODS: In total, 55 eyes of 55 different patients with no history of dry eye underwent standard phacoemulsification through a 2.75-mm-wide corneal incision. We measured tear break-up time, Schirmer test I, and tear meniscus height, and recorded the Ocular Surface Disease Index score, fluorescein staining patterns, and photo documentation of the ocular surface before and 1 day, 1 week, and 1 month postoperatively. Patients were divided into two groups (with and without superficial punctate keratopathy development, 1 day postoperatively). RESULTS: Patients (mean age: 75.75 ± 7.27 years) showed an incidence of 76.3% of superficial punctate keratopathy at 24 h. Location predominated in the center of the cornea until a week (32.7%) and then began to prevail in the inferior quadrant (21.8%) at 1 month. All dry eye tests were significantly worse after surgery. Ocular Surface Disease Index increased from 10.98 ± 5.05 to 15.87 ± 6.57 at 24 h (p < .001), to 12.80 ± 5.77 at 7 days (p < .001), and to 11.09 ± 4.63 at 1 month (p = .90). Fluorescein staining patterns got worse 24 h postoperatively with a score of 2.12 using the National Eye Institute/Industry-recommended guidelines staining grid. Average break-up time values were significantly lower at 1 day (6.61 ± 2.68),1 week (6.98 ± 2.79), and 1 month (7.05 ± 2.86) postoperatively than preoperatively (8.78 ± 2.97) (p < .001). The mean postoperative first month Schirmer test I value (8.32 ± 3.58) was significantly lower than preoperative value (9.05 ± 3.63) (p < .001). CONCLUSION: Phacoemulsification tends to induce short-term transitory ocular surface impairment manifesting as both signs and symptoms. Superficial punctate keratopathy distribution has a characteristic pattern evolution according to the postoperative time. Those patients with altered preoperative values are more likely to develop ocular surface disease and for longer time.

Endogenous osteopontin induces myocardial CCL5 and MMP-2 activation that contributes to inflammation and cardiac remodeling in a mouse model of chronic Chagas heart disease.

Cardiac dysfunction with progressive inflammation and fibrosis is a hallmark of Chagas disease caused by persistent Trypanosoma cruzi infection. Osteopontin (OPN) is a pro-inflammatory cytokine that orchestrates mechanisms controlling cell recruitment and cardiac architecture. Our main goal was to study the role of endogenous OPN as a modulator of myocardial CCL5 chemokine and MMP-2 metalloproteinase, and its pathological impact in a murine model of Chagas heart disease. Wild-type (WT) and OPN-deficient (spp1 -/-) mice were parasite-infected (Brazil strain) for 100days. Both groups developed chronic myocarditis with similar parasite burden and survival rates. However, spp1 -/- infection showed lower heart-to-body ratio (P<0.01) as well as reduced inflammatory pathology (P<0.05), CCL5 expression (P<0.05), myocyte size (P<0.05) and fibrosis (P<0.01) in cardiac tissues. Intense OPN labeling was observed in inflammatory cells recruited to infected heart (P<0.05). Plasma concentration of MMP-2 was higher (P<0.05) in infected WT than in spp1 -/- mice. Coincidently, specific immunostaining revealed increased gelatinase expression (P<0.01) and activity (P<0.05) in the inflamed hearts from T. cruzi WT mice, but not in their spp1 -/- littermates. CCL5 and MMP-2 induction occurred preferentially (P<0.01) in WT heart-invading CD8+ T cells and was mediated via phospho-JNK MAPK signaling. Heart levels of OPN, CCL5 and MMP-2 correlated (P<0.01) with collagen accumulation in the infected WT group only. Endogenous OPN emerges as a key player in the pathogenesis of chronic Chagas heart disease, through the upregulation of myocardial CCL5/MMP-2 expression and activities resulting in pro-inflammatory and pro-hypertrophic events, cardiac remodeling and interstitial fibrosis.

Unmethylated CpG motifs in Toxoplasma gondii DNA induce TLR9- and IFN-ß-dependent expression of α-defensin-5 in intestinal epithelial cells.

The gut epithelial barrier is a strategic place to prevent, or at least to limit, parasite dissemination upon oral infection with Toxoplasma gondii. Innate immunity to this pathogen results from delicate interactions involving different components of the infecting agent and the host. We herein aimed to examine the molecular mechanism by which protozoan DNA boosts the production of α-defensin-5 (DEFA-5), the main antimicrobial peptide at the target site of infection. The present study shows that DEFA-5 is rapidly upregulated in intestinal epithelial cells following intracellular Toll-like receptor 9 (TLR9) activation by unmethylated CpG motifs in DNA from T. gondii (CpG-DNA). Concomitantly, CpG-DNA purified from the pathogen markedly increased TLR9 mRNA expression levels in the Caco-2 cell line. We further verified that DEFA-5 production was dependent on interferon-ß released from these cells upon treatment with CpG-DNA prepared from tachyzoites. Our results suggest that, in protozoan DNA-stimulated intestinal epithelial cells, the TLR9/interferon-ß/DEFA-5 pathway may initiate an innate anti-T. gondii response without the need of parasite invasion. These findings highlight the key role of the gut epithelium in Toxoplasma recognition and amplification of local host defence against this microbe, thereby contributing to gain insight into immunoprotective mechanisms and to improve therapeutic strategies.