Differential regulation of PGC-1 alpha expression in rat liver and skeletal muscle in response to voluntary running

Background: The beneficial actions of exercise training on lipid, glucose and energy metabolism and insulin sensitivity appear to be in part mediated by PGC-1 alpha. Previous studies have shown that spontaneously exercised rats show at rest enhanced responsiveness to exogenous insulin, lower plasma insulin levels and increased skeletal muscle insulin sensitivity. This study was initiated to examine the functional interaction between exercise-induced modulation of skeletal muscle and liver PGC-1 alpha protein expression, whole body insulin sensitivity, and circulating FFA levels as a measure of whole body fatty acid (lipid) metabolism. Methods: Two groups of male Wistar rats (2 Mo of age, 188.82 +/- 2.77 g BW) were used in this study. One group consisted of control rats placed in standard laboratory cages. Exercising rats were housed individually in cages equipped with running wheels and allowed to run at their own pace for 5 weeks. At the end of exercise training, insulin sensitivity was evaluated by comparing steady-state plasma glucose (SSPG) concentrations at constant plasma insulin levels attained during the continuous infusion of glucose and insulin to each experimental group. Subsequently, soleus and plantaris muscle and liver samples were collected and quantified for PGC-1 alpha protein expression by Western blotting. Collected blood samples were analyzed for glucose, insulin and FFA concentrations. Results: Rats housed in the exercise wheel cages demonstrated almost linear increases in running activity with advancing time reaching to maximum value around 4 weeks. On an average, the rats ran a mean (Mean +/- SE) of 4.102 +/- 0.747 km/day and consumed significantly more food as compared to sedentary controls (P < 0.001) in order to meet their increased caloric requirement. Mean plasma insulin (P < 0.001) and FFA (P < 0.006) concentrations were lower in the exercise-trained rats as compared to sedentary controls. Mean steady state plasma insulin (SSPI) and glucose (SSPG) concentrations were not significantly different in sedentary control rats as compared to exercise-trained animals. Plantaris PGC-1 alpha protein expression increased significantly from a 1.11 +/- 0.12 in the sedentary rats to 1.74 +/- 0.09 in exercising rats (P < 0.001). However, exercise had no effect on PGC-1 alpha protein content in either soleus muscle or liver tissue. These results indicate that exercise training selectively up regulates the PGC-1 alpha protein expression in high-oxidative fast skeletal muscle type such as plantaris muscle. Conclusion: These data suggest that PGC-1 alpha most likely plays a restricted role in exercise-mediated improvements in insulin resistance (sensitivity) and lowering of circulating FFA levels.

Prevalence, Incidence Density, and Genotype Distribution of GB Virus C Infection in a Cohort of Recently HIV-1-Infected Subjects in Sao Paulo, Brazil

Background: The results of previous studies elsewhere have indicated that GB virus C (GBV-C) infection is frequent in patients infected with the human immunodeficiency virus type 1 (HIV-1) due to similar transmission routes of both viruses. The aim of this study was to determine the prevalence, incidence density and genotypic characteristics of GBV-C in this population. Methodology/Principal Findings: The study population included 233 patients from a cohort primarily comprised of homosexual men recently infected with HIV-1 in Sao Paulo, Brazil. The presence of GBV-C RNA was determined in plasma samples by reverse transcriptase-nested polymerase chain reaction and quantified by real-time PCR. GBV-C genotypes were determined by direct sequencing. HIV viral load, CD4+ T lymphocyte and CD8+ T lymphocyte count were also tested in all patients. The overall prevalence of GBV-C infection was 0.23 (95% CI: 0.18 to 0.29) in the study group. There was no significant difference between patients with and without GBV-C infection and Glycoprotein E2 antibody presence regarding age, sex, HIV-1 viral load, CD4+ and CD8+ T cell counts and treatment with antiretroviral drugs. An inverse correlation was observed between GBV-C and HIV-1 loads at enrollment and after one year. Also, a positive but not significant correlation was observed between GBV-C load and CD4+ T lymphocyte. Phylogenetic analysis of the GBV-C isolates revealed the presence of genotype 1 and genotype 2, these sub classified into subtype 2a and 2b. Conclusion/Significance: GBV-C infection is common in recently HIV -1 infected patients in Sao Paulo, Brazil and the predominant genotype is 2b. This study provides the first report of the GBV-C prevalence at the time of diagnosis of HIV-1 and the incidence density of GBV-C infection in one year.

Rat Adipose Tissue-Derived Stem Cells Transplantation Attenuates Cardiac Dysfunction Post Infarction and Biopolymers Enhance Cell Retention

Background: Cardiac cell transplantation is compromised by low cell retention and poor graft viability. Here, the effects of co-injecting adipose tissue-derived stem cells (ASCs) with biopolymers on cell cardiac retention, ventricular morphometry and performance were evaluated in a rat model of myocardial infarction (MI). Methodology/Principal Findings: (99m)Tc-labeled ASCs (1 x 10(6) cells) isolated from isogenic Lewis rats were injected 24 hours post-MI using fibrin a, collagen (ASC/C), or culture medium (ASC/M) as vehicle, and cell body distribution was assessed 24 hours later by gamma-emission counting of harvested organs. ASC/F and ASC/C groups retained significantly more cells in the myocardium than ASC/M (13.8+/-2.0 and 26.8+/-2.4% vs. 4.8+/-0.7%, respectively). Then, morphometric and direct cardiac functional parameters were evaluated 4 weeks post-MI cell injection. Left ventricle (LV) perimeter and percentage of interstitial collagen in the spare myocardium were significantly attenuated in all ASC-treated groups compared to the non-treated (NT) and control groups (culture medium, fibrin, or collagen alone). Direct hemodynamic assessment under pharmacological stress showed that stroke volume (SV) and left ventricle end-diastolic pressure were preserved in ASC-treated groups regardless of the vehicle used to deliver ASCs. Stroke work (SW), a global index of cardiac function, improved in ASC/M while it normalized when biopolymers were co-injected with ASCs. A positive correlation was observed between cardiac ASCs retention and preservation of SV and improvement in SW post-MI under hemodynamic stress. Conclusions: We provided direct evidence that intramyocardial injection of ASCs mitigates the negative cardiac remodeling and preserves ventricular function post-MI in rats and these beneficial effects can be further enhanced by administrating co-injection of ASCs with biopolymers.

The Effects of a 785-nm AlGaInP Laser on the Regeneration of Rat Anterior Tibialis Muscle After Surgically-Induced Injury

Objective: This study aims to investigate the effects of low-level laser therapy (LLLT) on muscle regeneration. For this purpose, the anterior tibialis muscle of 48 male Wistar rats received AlGaInP laser treatment (785 nm) after surgically-induced injury. Background Data: Few studies have been conducted on the effects of LLLT on muscle regeneration at different irradiation doses. Materials and Methods: The animals were randomized into four groups: uninjured rats (UN); uninjured and laser-irradiated rats (ULI); injured rats (IN); and injured and laser-irradiated rats (ILI). The direct contact laser treatment was started 24 h after surgery. An AlGaInP diode laser emitting 75 mW of continuous power at 785 nm was used for irradiation. The laser probe was placed at three treatment points to deliver 0.9 J per point, for a total dose of 2.7 J per treatment session. The animals were euthanized after treatment sessions 1, 2, and 4. Mounted sections were stained with hematoxylin and eosin and used for quantitative morphological analysis, in which the number of leukocytes and fibroblasts were counted over an area of 4480 mu m(2). The data were statistically analyzed by analysis of variance (ANOVA) and the Bonferroni t-test. Results: Quantitative data showed that the number of both polymorphonuclear and mononuclear leukocytes in the inflammatory infiltrate at the injury site was smaller in the ILI(1), ILI(2), and ILI(4) subgroups compared with their respective control subgroups (IN(1), IN(2), and IN(4)) for sessions 1, 2, and 4, respectively (p < 0.05). On the other hand, the number of fibroblasts increased after the fourth treatment session (p < 0.05). With regard to the regeneration of muscle fibers following injury, only after the fourth treatment session was it possible to find muscle precursor cells such as myoblasts and some myotubes in the ILI(4) subgroup. Conclusion: During the acute inflammatory phase, the AlGaInP laser treatment was found to have anti-inflammatory effects, reducing the number of leukocytes at the injury site and accelerating the regeneration of connective tissue.

Methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism and high plasma homocysteine in chronic hepatitis C (CHC) infected patients from the Northeast of Brazil

Background/Aim: Hyperhomocysteinemia due to Methylenetetrahydrofolate Reductase (MTHFR) gene, in particular the C677T (Ala222Val) polymorphism were recently associated to steatosis and fibrosis. We analyzed the frequency of MTHFR gene in a cross-sectional study of patients affected by Chronic Hepatitis C (CHC) from Northeast of Brazil. Method: One hundred seven-four untreated patients with CHC were genotyped for the C677T MTHFR. Genomic DNA was extracted from peripheral blood cells and the C677T MTHFR polymorphism was identified by PCR-RFLP. The homocysteine (Hcy) levels were determined by chemiluminescence method. All patients were negative for markers of Wilson's disease, hemochromatosis and autoimmune diseases and have current and past daily alcohol intake less than 100 g/week. Results: Among subjects infected with CHC genotype non-1 the frequency of MTHFR genotypes TT was 9.8% versus 4.4% genotype 1 (p = 0.01). Nevertheless, association was found between the MTHFR genotype TT x CT/CC polymorphism and the degree of steatosis and fibrosis in both hepatitis C genotype (p < 0.05). A significant difference was found on plasma Hcy levels in patients with steatosis regardless of HCV genotype (p = 0.03). Conclusion: Our results indicate that plasma Hcy levels is highly prevalent in subjects with chronic hepatits C with steatosis regardless of HCV genotype and vitamin deficiency. The presence of genotype TT of MTHFR C677T polymorphism was more common in CHC genotype non-1 infected patient regardless of histopathological classification and genotype TT+CT frequencies were significant in the presence of fibrosis grade 1+2 and of steatosis in CHC infected patients from the northeast of Brazil regardless of HCV genotype. The genetic susceptibility of MTHFR C677T polymorphism should be confirmed in a large population.

Role of upper endoscopy in diagnosing opportunistic infections in human immunodeficiency virus-infected patients

Highly active antiretroviral therapy (HAART) has dramatically decreased opportunistic infections (OIs) in human immunodeficiency virus (HIV)-infected patients. However, gastrointestinal disease continues to account for a high proportion of presenting symptoms in these patients. Gastrointestinal symptoms in treated patients who respond to therapy are more likely to the result of drug-induced complications than OI. Endoscopi evaluation of the gastrointestinal tract remains a cornerstone of diagnosis, especially in patients with advanced immunodeficiency, who are at risk for OI. The peripheral blood CD4 lymphocyte count helps to predict the risk of an OI, with the highest risk seen in HIV-infected patients with low CD4 count (< 200 cells/mm(3)). This review provides an update of the role of endoscopy in diagnosing OI in the upper gastrointestinal tract in HIV-infected patients in the era of HAART. (C) 2009 The WJG Press and Baishideng. All rights reserved.

Vaccination in Brazilian HIV-infected adults: A cross-sectional study

HIV-infected patients are at risk for vaccine-preventable infections. The Brazilian National Immunization Program provided recommendations for this population. However, the vaccine coverage reached by this program is unknown. This study aimed at evaluating the vaccine coverage of HIV-infected adults followed at Hospital das Clinicas, University of Sao Paulo School of Medicine. Data were collected on age, gender, mode of HIV transmission, Centers for Disease Classification 1993 classification (CDC/93), antiretrovirals, CD4 count, HIV viral load, and immunization charts, from April 2003 to August 2004. We interviewed 144 randomly selected patients, 74% male; mean age, 39.95 years; CDC classification: A, 40.6%; B, 19.6%; and C, 39.9%. Most of patients were undergoing highly active antiretroviral therapy (HAART; 86.8%). Mean CD4 count 442.6 cells/mm(3). Viral load less than 400 copies per milliliter in 59.4% of patients. Only 36.1% of patients were adequately immunized for diphtheria/tetanus, 54.9% for pneumococcus, 24.3% for flu, and 76.9% for hepatitis B. In relation to live attenuated vaccines, 5 patients received measles, mumps, and rubella vaccine and 7 patients yellow fever vaccine. Two patients were vaccinated against yellow fever despite CD4 less than 200 cell/mm(3). We verified poor vaccine coverage in HIV-infected patients. Vaccination campaigns and incorporation of vaccine rooms in sexually transmitted disease (STD)/AIDS clinics could improve this situation.

Selective Decrease of Components of the Creatine Kinase System and ATP Synthase Complex in Chronic Chagas Disease Cardiomyopathy

Background: Chronic Chagas disease cardiomyopathy (CCC) is an inflammatory dilated cardiomyopathy with a worse prognosis than other cardiomyopathies. CCC occurs in 30 % of individuals infected with Trypanosoma cruzi, endemic in Latin America. Heart failure is associated with impaired energy metabolism, which may be correlated to contractile dysfunction. We thus analyzed the myocardial gene and protein expression, as well as activity, of key mitochondrial enzymes related to ATP production, in myocardial samples of end-stage CCC, idiopathic dilated (IDC) and ischemic (IC) cardiomyopathies. Methodology/Principal Findings: Myocardium homogenates from CCC (N = 5), IC (N = 5) and IDC (N = 5) patients, as well as from heart donors (N = 5) were analyzed for protein and mRNA expression of mitochondrial creatine kinase (CKMit) and muscular creatine kinase (CKM) and ATP synthase subunits aplha and beta by immunoblotting and by real-time RT-PCR. Total myocardial CK activity was also assessed. Protein levels of CKM and CK activity were reduced in all three cardiomyopathy groups. However, total CK activity, as well as ATP synthase alpha chain protein levels, were significantly lower in CCC samples than IC and IDC samples. CCC myocardium displayed selective reduction of protein levels and activity of enzymes crucial for maintaining cytoplasmic ATP levels. Conclusions/Significance: The selective impairment of the CK system may be associated to the loss of inotropic reserve observed in CCC. Reduction of ATP synthase alpha levels is consistent with a decrease in myocardial ATP generation through oxidative phosphorylation. Together, these results suggest that the energetic deficit is more intense in the myocardium of CCC patients than in the other tested dilated cardiomyopathies.

High levels of T lymphocyte activation in Leishmania-HIV-1 co-infected individuals despite low HIV viral load

Background: Concomitant infections may influence HIV progression by causing chronic activation leading to decline in T-cell function. In the Americas, visceral (AVL) and tegumentary leishmaniasis (ATL) have emerged as important opportunistic infections in HIV-AIDS patients and both of those diseases have been implicated as potentially important co-factors in disease progression. We investigated whether leishmaniasis increases lymphocyte activation in HIV-1 co-infected patients. This might contribute to impaired cellular immune function. Methods: To address this issue we analyzed CD4(+) T absolute counts and the proportion of CD8(+) T cells expressing CD38 in Leishmania/HIV co-infected patients that recovered after anti-leishmanial therapy. Results: We found that, despite clinical remission of leishmaniasis, AVL co-infected patients presented a more severe immunossupression as suggested by CD4(+) T cell counts under 200 cells/mm(3), differing from ATL/HIV-AIDS cases that tends to show higher lymphocytes levels (over 350 cells/mm(3)). Furthermore, five out of nine, AVL/HIV-AIDS presented low CD4(+) T cell counts in spite of low or undetectable viral load. Expression of CD38 on CD8(+) T lymphocytes was significantly higher in AVL or ATL/HIV-AIDS cases compared to HIV/AIDS patients without leishmaniasis or healthy subjects. Conclusions: Leishmania infection can increase the degree of immune system activation in individuals concomitantly infected with HIV. In addition, AVL/HIV-AIDS patients can present low CD4(+) T cell counts and higher proportion of activated T lymphocytes even when HIV viral load is suppressed under HAART. This fact can cause a misinterpretation of these laboratorial markers in co-infected patients.

Genetic Diversity on the Integrase Region of the pol Gene among HIV Type 1-Infected Patients Naive for Integrase Inhibitors in Sao Paulo City, Brazil

The presence of mutations associated with integrase inhibitor (INI) resistance among INI-naive patients may play an important clinical role in the use of those drugs Samples from 76 HIV-1-infected subjects naive to INIs were submitted to direct sequencing. No differences were found between naive (25%) subjects and subjects on HAART (75%). No primary mutation associated with raltegravir or elvitegravir resistance was found. However, 78% of sequences showed at least one accessory mutation associated with resistance. The analysis of the 76 IN sequences showed a high polymorphic level on this region among Brazilian HIV-1-infected subjects, including a high prevalence of aa substitutions related to INI resistance. The impact of these findings remains unclear and further studies are necessary to address these questions.

Predictors of HBeAg status and hepatitis B viraemia in HIV-infected patients with chronic hepatitis B in the HAART era in Brazil

Background: HBV-HIV co-infection is associated with an increased liver-related morbidity and mortality. However, little is known about the natural history of chronic hepatitis B in HIV-infected individuals under highly active antiretroviral therapy (HAART) receiving at least one of the two drugs that also affect HBV (TDF and LAM). Information about HBeAg status and HBV viremia in HIV/HBV co-infected patients is scarce. The objective of this study was to search for clinical and virological variables associated with HBeAg status and HBV viremia in patients of an HIV/HBV co-infected cohort. Methods: A retrospective cross-sectional study was performed, of HBsAg-positive HIV-infected patients in treatment between 1994 and 2007 in two AIDS outpatient clinics located in the Sao Paulo metropolitan area, Brazil. The baseline data were age, sex, CD4 T+ cell count, ALT level, HIV and HBV viral load, HBV genotype, and duration of antiretroviral use. The variables associated to HBeAg status and HBV viremia were assessed using logistic regression. Results: A total of 86 HBsAg patients were included in the study. Of these, 48 (56%) were using combination therapy that included lamivudine (LAM) and tenofovir (TDF), 31 (36%) were using LAM monotherapy, and 7 patients had no previous use of either one. Duration of use of TDF and LAM varied from 4 to 21 and 7 to 144 months, respectively. A total of 42 (48. 9%) patients were HBeAg positive and 44 (51. 1%) were HBeAg negative. The multivariate analysis revealed that the use of TDF for longer than 12 months was associated with undetectable HBV DNA viral load (serum HBV DNA level < 60 UI/ml) (p = 0. 047). HBeAg positivity was associated with HBV DNA > 60 UI/ml (p = 0. 001) and ALT levels above normality (p = 0. 038). Conclusion: Prolonged use of TDF containing HAART is associated with undetectable HBV DNA viral load. HBeAg positivity is associated with HBV viremia and increased ALT levels.

Incidence and predictors of severe liver fibrosis in HIV-infected patients with chronic hepatitis C in Brazil

The aim of this study was to examine the incidence and factors associated with the severity of liver fibrosis in 234 coinfected patients in Brazil. Patients were cared for in our clinic, from 1996 to 2004. Eligible patients were defined as patients with documented HIV and hepatitis C virus (HCV) infections and had previously undergone a liver biopsy. Patients with persistently normal alanine aminotransferase (ALT) were also included. The variables selected for study were age, gender, risk category, history of high alcohol consumption, CD4(+) T cell count, antiretroviral therapy usage, HCV genotype and duration of HCV infection. Stage of fibrosis was scored as follows: F0, no fibrosis; F1, portal fibrosis with no septa; F2, portal fibrosis with few septa; F3, bridging fibrosis with many septa; and F4, cirrhosis. The liver fibrosis stage was F3 in 39 (16.6%) and F4 in 20(8.5%) patients. Among patients with normal ALT, the liver fibrosis stage was F3-F4 in three patients (5.6%). Predictors of severe liver fibrosis (17344) by multivariate analysis were age (older patients) and genotype 3 (genotype I odds ratio [OR], 0.28; 95% confidence interval [0], 0.12 0.65). In summary, in the present study severe liver fibrosis was found in 25% of our patients and was associated with an age of more than 38 years at the time of liver biopsy as well as, HCV genotype 3. No differences were found with respect to CD4(+) T cell counts although patients with a CD4(+) T cell count greater than 50 were excluded.

BALB/c Mice Infected with Antimony Treatment Refractory Isolate of Leishmania braziliensis Present Severe Lesions due to IL-4 Production

Background: Leishmania braziliensis is the main causative agent of cutaneous leishmaniasis in Brazil. Protection against infection is related to development of Th1 responses, but the mechanisms that mediate susceptibility are still poorly understood. Murine models have been the most important tools in understanding the immunopathogenesis of L. major infection and have shown that Th2 responses favor parasite survival. In contrast, L. braziliensis-infected mice develop strong Th1 responses and easily resolve the infection, thus making the study of factors affecting susceptibility to this parasite difficult. Methodology/Principal Findings: Here, we describe an experimental model for the evaluation of the mechanisms mediating susceptibility to L. braziliensis infection. BALB/c mice were inoculated with stationary phase promastigotes of L. braziliensis, isolates LTCP393(R) and LTCP15171(S), which are resistant and susceptible to antimony and nitric oxide (NO), respectively. Mice inoculated with LTCP393(R) presented larger lesions that healed more slowly and contained higher parasite loads than lesions caused by LTCP15171(S). Inflammatory infiltrates in the lesions and production of IFN-gamma, TNF-alpha, IL-10 and TGF-beta were similar in mice inoculated with either isolate, indicating that these factors did not contribute to the different disease manifestations observed. In contrast, IL-4 production was strongly increased in LTCP393(R)-inoculated animals and also arginase I (Arg I) expression. Moreover, anti-IL-4 monoclonal antibody (mAb) treatment resulted in decreased lesion thickness and parasite burden in animals inoculated with LTCP393(R), but not in those inoculated with LTCP15171(S). Conclusion/Significance: We conclude that the ability of L. braziliensis isolates to induce Th2 responses affects the susceptibility to infection with these isolates and contributes to the increased virulence and severity of disease associated with them. Since these data reflect what happens in human infection, this model could be useful to study the pathogenesis of the L. braziliensis infection, as well as to design new strategies of therapeutic intervention.

The Lectin ArtinM Induces Recruitment of Rat Mast Cells from the Bone Marrow to the Peritoneal Cavity

Background: The D-mannose binding lectin ArtinM is known to recruit neutrophils, to degranulate mast cells and may have potential therapeutic applications. However, the effect of ArtinM on mast cell recruitment has not been investigated. Methodology: Male Wistar rats were injected i.p. with ArtinM or ConA (control). The ability of the lectin to degranulate peritoneal and mesenteric mast cells was examined. Recruitment of mast cells to the peritoneal cavity and mesentery after ArtinM injection was examined with or without depletion of peritoneal mast cells by distilled water. Results: ArtinM degranulated both peritoneal and mesentery mast cells in vitro. Three days after i.p. injection of the lectin there were reduced numbers of mast cells in the peritoneal lavage, while at 7 days post injection of ArtinM, the number of peritoneal mast cells was close to control values. Since immature mast cells are recruited from the bone marrow, the effect of the lectin on bone marrow mast cells was examined. Injection of ArtinM resulted in an increased number of mast cells in the bone marrow. To determine if degranulation of mast cells in the peritoneal cavity was required for the increase in bone marrow mast cells, the peritoneal cavity was depleted of mast cells with ultrapure water. Exposure to ArtinM increased the number of mast cells in the bone marrow of rats depleted of peritoneal mast cells. Conclusions: The ArtinM induced recruitment of mast cells from the bone marrow to the peritoneal cavity may partially explain the therapeutic actions of ArtinM.

Challenges and opportunities for primary, secondary, and tertiary prevention of Chagas' disease

A century after its discovery, Chagas' disease still represents a major public health challenge in Latin America. Moreover, because of growing population movements, an increasing number of cases of imported Chagas' disease have now been detected in non-endemic areas, such as North America and some European countries. This parasitic zoonosis, caused by Trypanosoma cruzi, is transmitted to humans by infected Triatominae insects, or occasionally by non-vectorial mechanisms, such as blood transfusion, mother to fetus, or oral ingestion of materials contaminated with parasites. Following the acute phase of the infection, untreated individuals enter a chronic phase that is initially asymptomatic or clinically unapparent. Usually, a few decades later, 40-50% of patients develop progressive cardiomyopathy and/or motility disturbances of the oesophagus and colon. In the last decades several interventions targeting primary, secondary and tertiary prevention of Chagas' disease have been attempted. While control of both vectorial and blood transfusion transmission of T cruzi (primary prevention) has been successful in many regions of Latin America, early detection and aetiological treatment of asymptomatic subjects with Chagas' disease (secondary prevention) have been largely underutilised. At the same time, in patients with established chronic disease, several pharmacological and non-pharmacological interventions are currently available and have been increasingly used with the intention of preventing or delaying complications of the disease (tertiary prevention). In this review we discuss in detail each of these issues.