The balance between the production of tumor necrosis factor-alpha and interleukin-10 determines tissue injury and lethality during intestinal ischemia and reperfusion

A major goal in the treatment of acute ischemia of a vascular territory is to restore blood flow to normal values, i.e. to "reperfuse" the ischemic vascular bed. However, reperfusion of ischemic tissues is associated with local and systemic leukocyte activation and trafficking, endothelial barrier dysfunction in postcapillary venules, enhanced production of inflammatory mediators and great lethality. This phenomenon has been referred to as "reperfusion injury" and several studies demonstrated that injury is dependent on neutrophil recruitment. Furthermore, ischemia and reperfusion injury is associated with the coordinated activation of a series of cytokines and adhesion molecules. Among the mediators of the inflammatory cascade released, TNF-alpha appears to play an essential role for the reperfusion-associated injury. On the other hand, the release of IL-10 modulates pro-inflammatory cytokine production and reperfusion-associated tissue injury. IL-1beta, PAF and bradykinin are mediators involved in ischemia and reperfusion injury by regulating the balance between TNF-alpha and IL-10 production. Strategies that enhance IL-10 and/or prevent TNF-alpha concentration may be useful as therapeutic adjuvants in the treatment of the tissue injury that follows ischemia and reperfusion.

Hydrocortisone decreases apoptosis in jejunum of horses subjected to experimental ischemia and reperfusion

In order to evaluate the effect of hydrocortisone on apoptosis in the jejunum of horses subjected to ischemia and reperfusion, ten horses were paired and grouped into two groups - treated (n=5) and non treated (n=5). Segments of the jejunum were used as controls (C), or as venous ischemia (VIsc), which were subjected to 2h of ischemia followed by 2 or 12h of reperfusion. C samples were collected at time zero (prior to ischemia) and VIsc samples were collected at 2h of ischemia and at 2 and 12h of reperfusion. TUNEL positive apoptotic cells were counted in 10 microscopical fields in deep mucosa from each horse throughout the time course. After 12h of reperfusion, the number of apoptotic cells in treated group were significantly lower than in untreated animals, indicating that hydrocortisone inhibits apoptosis. These results indicate that hydrocortisone has a beneficial effects favoring the maintenance of jejunal integrity in horses with ischemia and reperfusion injuries by preventing apoptotic cell death.

Kallikrein-like amidase activity in renal ischemia and reperfusion

We assessed a kallikrein-like amidase activity probably related to the kallikrein-kinin system, as well as the participation of leukocyte infiltration in renal ischemia and reperfusion. Male C57BL/KSJmdb mice were subjected to 20 or 60 min of ischemia and to different periods of reperfusion. A control group consisted of sham-operated mice, under similar conditions, except for ischemia induction. Kallikrein-like amidase activity, Evans blue extravasation and myeloperoxidase activity were measured in kidney homogenates, previously perfused with 0.9% NaCl. Plasma creatinine concentration increased only in the 60-min ischemic group. After 20 min of ischemia and 1 or 24 h of reperfusion, no change in kallikrein-like amidase activity or Evans blue extravasation was observed. In the mice subjected to 20 min of ischemia, edema was evident at 1 h of reperfusion, but kidney water content returned to basal levels after 24 h of reperfusion. In the 60-min ischemic group, kallikrein-like amidase activity and Evans blue extravasation showed a similar significant increase along reperfusion time. Kallikrein-like amidase activity increased from 4 nmol PNA mg protein-1 min-1 in the basal condition to 15 nmol PNA mg protein-1 min-1 at 10 h of reperfusion. For dye extravasation the concentration measured was near 200 µg of Evans blue/g dry tissue in the basal condition and 1750 µg of Evans blue/g dry tissue at 10 h of reperfusion. No variation could be detected in the control group. A significant increase from 5 to 40 units of DAbs 655 nm g wet tissue-1 min-1 in the activity of the enzyme myeloperoxidase was observed in the 60-min ischemic group, when it was evaluated after 24 h of reperfusion. Histological analysis of the kidneys showed migration of polymorphonuclear leukocytes from the vascular bed to the interstitial tissue in the 60-min ischemic group after 24 h of reperfusion. We conclude that the duration of ischemia is critical for the development of damage during reperfusion and that the increase in renal cortex kallikrein-like amidase activity probably released from both the kidney and leukocytes may be responsible, at least in part, for the observed effects, probably through direct induction of increased vascular permeability.

Coronary endothelial dysfunction after ischemia and reperfusion: a new therapeutic target?

Although cardiac ischemia is usually characterized as a disease of the myocyte, it is clear that the vasculature, and especially endothelial cells, is also a major target of this pathology. Indeed, using a rat model of ischemia/reperfusion, we were able to detect severe endothelial dysfunction (assessed as a decreased response to acetylcholine) after acute or chronic reperfusion. Given the essential role of the endothelium in the regulation of vascular tone, as well as platelet and leukocyte function, such a severe dysfunction could lead to an increased risk of vasospasm, thrombosis and accelerated atherosclerosis. This dysfunction can be prevented by free radical scavengers and by exogenous nitric oxide. Endothelial dysfunction can also be prevented by preconditioning with brief periods of intermittent ischemia, thus extending to coronary endothelial cells the concept of endogenous protection previously described at the myocyte level. Experiments performed on cultured cells showed that the endothelial protection induced by free radical scavengers or by preconditioning was due to a lesser expression of endothelial adhesion molecules such as intercellular adhesion molecule-1, leading to a lesser adhesion of neutrophils to endothelial cells. Identification of the mechanisms of this protection may lead to the development of new strategies aimed at protecting the vasculature in ischemic heart diseases.

Hydrogen sulfide postconditioning protects isolated rat hearts against ischemia and reperfusion injury mediated by the JAK2/STAT3 survival pathway

The JAK2/STAT3 signal pathway is an important component of survivor activating factor enhancement (SAFE) pathway. The objective of the present study was to determine whether the JAK2/STAT3 signaling pathway participates in hydrogen sulfide (H2S) postconditioning, protecting isolated rat hearts from ischemic-reperfusion injury. Male Sprague-Dawley rats (230-270 g) were divided into 6 groups (N = 14 per group): time-matched perfusion (Sham) group, ischemia/reperfusion (I/R) group, NaHS postconditioning group, NaHS with AG-490 group, AG-490 (5 µM) group, and dimethyl sulfoxide (DMSO; <0.2%) group. Langendorff-perfused rat hearts, with the exception of the Sham group, were subjected to 30 min of ischemia followed by 90 min of reperfusion after 20 min of equilibrium. Heart rate, left ventricular developed pressure (LVDP), left ventricular end-diastolic pressure (LVEDP), and the maximum rate of increase or decrease of left ventricular pressure (± dp/dt max) were recorded. Infarct size was determined using triphenyltetrazolium chloride (TTC) staining. Myocardial TUNEL staining was used as the in situ cell death detection method and the percentage of TUNEL-positive nuclei to all nuclei counted was used as the apoptotic index. The expression of STAT3, bcl-2 and bax was determined by Western blotting. After reperfusion, compared to the I/R group, H2S significantly improved functional recovery and decreased infarct size (23.3 ± 3.8 vs 41.2 ± 4.7%, P < 0.05) and apoptotic index (22.1 ± 3.6 vs 43.0 ± 4.8%, P < 0.05). However, H2S-mediated protection was abolished by AG-490, the JAK2 inhibitor. In conclusion, H2S postconditioning effectively protects isolated I/R rat hearts via activation of the JAK2/STAT3 signaling pathway.

Putative role of ischemic postconditioning in a rat model of limb ischemia and reperfusion: involvement of hypoxia-inducible factor-1α expression

Hypoxia-inducible factor-1α (HIF-1α) is one of the most potent angiogenic growth factors. It improves angiogenesis and tissue perfusion in ischemic skeletal muscle. In the present study, we tested the hypothesis that ischemic postconditioning is effective for salvaging ischemic skeletal muscle resulting from limb ischemia-reperfusion injury, and that the mechanism involves expression of HIF-1α. Wistar rats were randomly divided into three groups (n=36 each): sham-operated (group S), hindlimb ischemia-reperfusion (group IR), and ischemic postconditioning (group IPO). Each group was divided into subgroups (n=6) according to reperfusion time: immediate (0 h, T0), 1 h (T1), 3 h (T3), 6 h (T6), 12 h (T12), and 24 h (T24). In the IPO group, three cycles of 30-s reperfusion and 30-s femoral aortic reocclusion were carried out before reperfusion. At all reperfusion times (T0-T24), serum creatine kinase (CK) and lactate dehydrogenase (LDH) activities, as well as interleukin (IL)-6, IL-10, and tumor necrosis factor-α (TNF-α) concentrations, were measured in rats after they were killed. Histological and immunohistochemical methods were used to assess the skeletal muscle damage and HIF-1α expression in skeletal muscle ischemia. In groups IR and IPO, serum LDH and CK activities and TNF-α, IL-6, and IL-10 concentrations were all significantly increased compared to group S, and HIF-1α expression was up-regulated (P<0.05 or P<0.01). In group IPO, serum LDH and CK activities and TNF-α and IL-6 concentrations were significantly decreased, IL-10 concentration was increased, HlF-1α expression was down-regulated (P<0.05 or P<0.01), and the pathological changes were reduced compared to group IR. The present study suggests that ischemic postconditioning can reduce skeletal muscle damage caused by limb ischemia-reperfusion and that its mechanisms may be related to the involvement of HlF-1α in the limb ischemia-reperfusion injury-triggered inflammatory response.

Epilepsy-induced electrocardiographic alterations following cardiac ischemia and reperfusion in rats

The present study evaluated electrocardiographic alterations in rats with epilepsy submitted to an acute myocardial infarction (AMI) model induced by cardiac ischemia and reperfusion. Rats were randomly divided into two groups: control (n=12) and epilepsy (n=14). It was found that rats with epilepsy presented a significant reduction in atrioventricular block incidence following the ischemia and reperfusion procedure. In addition, significant alterations were observed in electrocardiogram intervals during the stabilization, ischemia, and reperfusion periods of rats with epilepsy compared to control rats. It was noted that rats with epilepsy presented a significant increase in the QRS interval during the stabilization period in relation to control rats (P<0.01). During the ischemia period, there was an increase in the QRS interval (P<0.05) and a reduction in the P wave and QT intervals (P<0.05 for both) in rats with epilepsy compared to control rats. During the reperfusion period, a significant reduction in the QT interval (P<0.01) was verified in the epilepsy group in relation to the control group. Our results indicate that rats submitted to an epilepsy model induced by pilocarpine presented electrical conductivity alterations of cardiac tissue, mainly during an AMI episode.

Endothelium-dependent and -independent hepatic artery vasodilatation is not impaired in a canine model of liver ischemia-reperfusion injury

We investigated whether hepatic artery endothelium may be the earliest site of injury consequent to liver ischemia and reperfusion. Twenty-four heartworm-free mongrel dogs of either sex exposed to liver ischemia/reperfusion in vivo were randomized into four experimental groups (N = 6): a) control, sham-operated dogs, b) dogs subjected to 60 min of ischemia, c) dogs subjected to 30 min of ischemia and 60 min of reperfusion, and d) animals subjected to 45 min of ischemia and 120 min of reperfusion. The nitric oxide endothelium-dependent relaxation of hepatic artery rings contracted with prostaglandin F2a and exposed to increasing concentrations of acetylcholine, calcium ionophore A23187, sodium fluoride, phospholipase-C, poly-L-arginine, isoproterenol, and sodium nitroprusside was evaluated in organ-chamber experiments. Lipid peroxidation was estimated by malondialdehyde activity in liver tissue samples and by blood lactic dehydrogenase (LDH), serum aspartate aminotransferase (AST) and serum alanine aminotransferase (ALT) activities. No changes were observed in hepatic artery relaxation for any agonist tested. The group subjected to 45 min of ischemia and 120 min of reperfusion presented marked increases of serum aminotransferases (ALT = 2989 ± 1056 U/L and AST = 1268 ± 371 U/L; P < 0.01), LDH = 2887 ± 1213 IU/L; P < 0.01) and malondialdehyde in liver samples (0.360 ± 0.020 nmol/mgPT; P < 0.05). Under the experimental conditions utilized, no abnormal changes in hepatic arterial vasoreactivity were observed: endothelium-dependent and independent hepatic artery vasodilation were not impaired in this canine model of ischemia/reperfusion injury. In contrast to other vital organs and in the ischemia/reperfusion injury environment, dysfunction of the main artery endothelium is not the first site of reperfusion injury.

Quercetin postconditioning attenuates myocardial ischemia/reperfusion injury in rats through the PI3K/Akt pathway

Quercetin (Que), a plant-derived flavonoid, has multiple benefical actions on the cardiovascular system. The current study investigated whether Que postconditioning has any protective effects on myocardial ischemia/reperfusion (I/R) injury in vivo and its potential cardioprotective mechanisms. Male Sprague-Dawley rats were randomly allocated to 5 groups (20 animals/group): sham, I/R, Que postconditioning, Que+LY294002 [a phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway inhibitor], and LY294002+I/R. I/R was produced by 30-min coronary occlusion followed by 2-h reperfusion. At the end of reperfusion, myocardial infarct size and biochemical changes were compared. Apoptosis was evaluated by both TUNEL staining and measurement of activated caspase-3 immunoreactivity. The phosphorylation of Akt and protein expression of Bcl-2 and Bax were determined by Western blotting. Que postconditioning significantly reduced infarct size and serum levels of creatine kinase and lactate dehydrogenase compared with the I/R group (all P<0.05). Apoptotic cardiomyocytes and caspase-3 immunoreactivity were also suppressed in the Que postconditioning group compared with the I/R group (both P<0.05). Akt phosphorylation and Bcl-2 expression increased after Que postconditioning, but Bax expression decreased. These effects were inhibited by LY294002. The data indicate that Que postconditioning can induce cardioprotection by activating the PI3K/Akt signaling pathway and modulating the expression of Bcl-2 and Bax proteins.

Intestinal parasites and commensals among individuals from a landless camping in the rural area of Uberlândia, Minas Gerais, Brazil

We evaluated the occurrence of intestinal parasites and commensals among children and adults from a landless camping in the rural area of Uberlândia, State of Minas Gerais, Brazil, from October to November 2001. Stool samples from 78 individuals were examined by both the Baermann-Moraes and Lutz methods. Fifty-one (65.4%; CI 54.8 - 76.0) individuals were found to be infected, 23 (45.1%) children and 28 (54.9%) adults, of whom 34 (66.7%) were mono-infected, 9 (17.6%) bi-infected, and 8 (15.7%) poly-infected. In conclusion, the high prevalence of intestinal parasites and commensals suggests that parasitological exams should be periodically carried out in addition to the sanitation education and health special care in this population.

Intestinal parasitoses and environmental factors in a rural population of Argentina, 2002-2003

We evaluate the prevalence of intestinal parasites in 504 people and the degree of association between environmental variables and parasites found in population, soil and water in a rural area of Argentina during 2002-2003. A structured survey was used to evaluate the environmental variables and fecal-human, soil and water samples were analyzed. The prevalence of parasites was 45.4%. Most prevalent protozoa were Blastocystis hominis (27.2%) and Giardia lamblia (6.9%), while the most prevalent helminth was Ascaris lumbricoides (3.8%). The analyzed environmental variables showing association (p < 0.05) with presence of parasites in population were: cardboard-tin or wooden house, dirt floor, home or communal water pump, faucet outside the house or public faucet and cesspool or latrine. Parasite forms were found in 82.3% of the soil samples and in 84.2% of the water samples. In both samples we found parasites that were also found in people. In this study we have found deficient sanitary conditions associated with presence of parasites in population and we have evidenced that contaminated soil and water were the source of these parasites.

Intestinal parasitism and socio-environmental factors among Mbyá-Guarani indians, Porto Alegre, Rio Grande do Sul, Brazil

Disturbing data reveal the prevalence of intestinal parasites and their relationship with socio-environmental factors among Mbyá-Guarani Indians. The prevalence was determined by spontaneous sedimentation in water, centrifugation-floatation, and Kato-Katz. A socioeconomic questionnaire was submitted to each family. The overall prevalence of intestinal parasites was 88.7%, and 45.5% were polyparasitized. There was 90.5% prevalence of enteric parasites in children (1-12- year-old), and 85% among 13-65-year-old individuals, indicating that both age groups are extensively parasitized. The parasite load was low to moderate for geohelminths and 75% of the families did not have latrine, thus the practice of defecation occurred outdoors. These findings suggest that the multiple intestinal parasitism in the Mbyá-Guarani community is high to the point of being the rule, and that it relates essentially to the traditional lifestyle and health habits. It is urgently necessary to implement the association of anti-parasitic treatment with sanitation improvement. This should be done simultaneously with health education activities for this population.

PREVALENCE OF INTESTINAL PARASITISM AND ASSOCIATED SYMPTOMATOLOGY AMONG HEMODIALYSIS PATIENTS

Intestinal parasites are an important cause of morbidity and mortality. Immunocompromised individuals may develop more severe forms of these infections. Taking into account the immunity impairment in patients suffering from chronic renal failure (CRF), we will determine the prevalence and associated symptoms of intestinal parasites in these patients. Controls without CRF were used for comparison. Stool samples were collected and processed for microscopic identification of parasites using the Formalin-ether concentration method. For Cryptosporidium diagnosis, the ELISA technique was used. One hundred and ten fecal samples from hemodialysis patients were analyzed, as well as 86 from a community group used as control group. A result of 51.6% of intestinal parasites was observed in hemodialysis patients and 61.6% in the control group. Cryptosporidium and Blastocystis were the most common infections in patients with CRF (26.4% and 24.5%, respectively). Blastocystis was the most common infection in the control group (41.9%), however no individual was found positive for Cryptosporidium. Among the CRF patients, 73.6% were symptomatic, 54.3% of these tested positive for at least one parasite, in contrast to 44.8% in asymptomatic patients (p = 0.38). The most common symptoms in this group were flatulence (36.4%), asthenia (30.0%) and weight loss (30.0%). In the control group, 91.9% were symptomatic, 60.8% of these tested positive for at least one parasite, in contrast to 71.4% in asymptomatic patients (p = 0.703). A significant difference between the two groups was observed with regard to symptoms, with bloating, postprandial fullness, and abdominal pain being more frequent in the control group than in the hemodialysis group (all p < 0.05). Comparing symptomatic with asymptomatic, there was no association in either group between symptoms or the prevalence of parasitic infection, nor with the type of parasite or with multiple parasitic infections. Patients with chronic renal failure are frequent targets for renal transplantation, which as well as the inherent immunological impairment of the disease itself, results in immunosuppression by medication. For this reason, carriers of intestinal parasites with pathogenic potential can develop serious clinical complications influencing the success of transplantation. This fact, coupled with the high prevalence of intestinal parasites and the dissociation between symptoms and infection in CRF patients, suggests that the stool test should be incorporated in routine propedeutics. Furthermore, preventive measures for the acquisition of parasites through the fecal-oral contamination route should be introduced.

Intestinal nematodes and pulmonary tuberculosis

We report a significantly higher prevalence of intestinal nematodes in patients with pulmonary tuberculosis (TB) compared to a matched control group: 33/57 (57.8%) in patients with TB and 18/86 (20.9%) in the control group; OR=5.19; 95% CI= 2.33-11.69; p=0.000). When TB patients eosinophilia was also significantly higher among those with intestinal parasites (69.8%) compared to those without this condition (45.6%). We hypothesized that the immune modulation induced by nematodes is a factor that enhances TB infection/progression and that eosinophilia seen in TB patients is a consequence of helminth infection.

Intestinal spirochetosis and colon diverticulosis

A case of intestinal spirochetosis in a 62-year-old white male is reported. The condition was characterized by chronic flatulence and episodes of intestinal hemorrhage, in addition to the evidence of hypotonic diverticular disease, with a large number of slender organisms in the colon epithelium and cryptae. Spirochetes were demonstrated by Whartin-Starry stain. The serologic tests for syphilis and HIV were positive. Spirochetosis was treated with penicillin G, and the patient remains free of intestinal complaints 20 months later.