Acute actions of thyroid hormone on blood vessel biochemistry and physiology

Purpose of review Description of the progress about the vascular effects promoted by thyroid hormones. Recent findings Over the past few years, a number of studies have shown that in addition to genomic effects on blood vessels, thyroid hormones exert extranuclear nongenomic effects on vascular smooth muscle cells and endothelium. These nongenomic effects occur rapidly and do not involve thyroid hormone response elements-mediated transcriptional events. In this context, the genomic and nongenomic events promoted by thyroid hormones act in concert to control the vascular hemodynamic and regulate the cardiovascular function. Summary Considering the antiatherogenic property of thyroid hormones and the rapid effects produced by this molecule as a vasodilator, including that in the coronary bed, a better understanding of the molecular mechanisms involved in its action may contribute to the development of drugs that can be clinically used to increase the known benefits promoted by thyroid hormones in cardiovascular physiology.

Increased Expression of GluR2-flip in the Hippocampus of the Wistar Audiogenic Rat Strain After Acute and Kindled Seizures

The Wistar Audiogenic Rat (WAR) is an epileptic-prone strain developed by genetic selection from a Wistar progenitor based on the pattern of behavioral response to sound stimulation. Chronic acoustic stimulation protocols of WARs (audiogenic kindling) generate limbic epileptogenesis, confirmed by ictal semiology, amygdale, and hippocampal EEG, accompanied by hippocampal and amygdala cell loss, as well as neurogenesis in the dentate gyrus (DG). In an effort to identify genes involved in molecular mechanisms underlying epileptic process, we used suppression-subtractive hybridization to construct normalized cDNA library enriched for transcripts expressed in the hippocampus of WARs. The most represented gene among the 133 clones sequenced was the ionotropic glutamate receptor subunit II (GluR2), a member of the a-amino-3-hydroxy-5-methyl-4-isoxazoleopropionic acid (AMPA) receptor. Although semiquantitative RT-PCR analysis shows that the hippocampal levels of the GluR2 subunits do not differ between naive WARs and their Wistar counterparts, we observed that the expression of the transcript encoding the splice-variant GluR2-flip is increased in the hippocampus of WARs submitted to both acute and kindled audiogenic seizures. Moreover, using in situ hybridization, we verified upregulation of GluR2-flip mainly in the CA1 region, among the hippocampal subfields of audiogenic kindled WARs. Our findings on differential upregulation of GluR2-flip isoform in the hippocampus of WARs displaying audiogenic seizures is original and agree with and extend previous immunohistochemical for GluR2 data obtained in the Chinese P77PMC audiogenic rat strain, reinforcing the association of limbic AMPA alterations with epileptic seizures. (C) 2009 Wiley-Liss, Inc.

Structural changes in the epithelium of the small intestine and immune cell infiltration of enteric ganglia following acute mucosal damage and local inflammation

An acute enteritis is commonly followed by intestinal neuromuscular dysfunction, including prolonged hyperexcitability of enteric neurons. Such motility disorders are associated with maintained increases in immune cells adjacent to enteric ganglia and in the mucosa. However, whether the commonly used animal model, trinitrobenzene sulphonate (TNBS)-induced enteritis, causes histological and immune cell changes similar to human enteric neuropathies is not clear. We have made a detailed study of the mucosal damage and repair and immune cell invasion following intralumenal administration of TNBS. Intestines from untreated, sham-operated and TNBS-treated animals were examined at 3 h to 56 days. At 3 h, the mucosal surface was completely ablated, by 6 h an epithelial covering was substantially restored and by 1 day there was full re-epithelialisation. The lumenal epithelium developed from a squamous cell covering to a fully differentiated columnar epithelium with mature villi at about 7 days. Prominent phagocytic activity of enterocytes occurred at 1-7 days. A surge of eosinophils and T lymphocytes associated with the enteric nerve ganglia occurred at 3 h to 3 days. However, elevated immune cell numbers occurred in the lamina propria of the mucosa until 56 days, when eosinophils were still three times normal. We conclude that the disruption of the mucosal surface that causes TNBS-induced ileitis is brief, a little more than 6 h, and causes a transient immune cell surge adjacent to enteric ganglia. This is much briefer than the enteric neuropathy that ensues. Ongoing mucosal inflammatory reaction may contribute to the persistence of enteric neuropathy.

Granulocyte Colony-stimulating Factor Reduces Mortality by Suppressing Ventricular Arrhythmias in Acute Phase of Myocardial Infarction in Rats

Our aim was to evaluate the effects of granulocyte colony-stimulating factor (G-CSF) on early cardiac arrhythmias after myocardial infarction (MI) and the impact on survival. Male Wistar rats received repeated doses of 50 mu g/kg G-CSF (MI-GCSF group) or vehicle (MI group) at 7, 3, and 1 days before surgery. MI was induced by permanent occlusion of left corollary artery. The electrocardiogram was obtained before occlusion and then for 30 minutes after surgery. Events and duration of ventricular arrhythmias were analyzed. The levels of connexin43 (Cx43) were measured by Western blot immediately before MI production. Survival was significantly increased in MI-GCSF pretreated group (74% versus 52.0% MI. P < 0.05). G-CSF pretreatment also significantly reduced the ventricular premature beats when compared with the untreated-MI group (201 +/- 47 versus 679 +/- 117, P < 0.05). The number and the duration of ventricular tachycardia were smaller in the MI-G-CSF group, as well as the number of ventricular fibrillation episodes (10% versus 69% in NIL P < 0.05). Cx43 levels were significantly increased by G-CSF treatment (1.27 +/- 0.13 versus 0.86 +/- 0.11; P < 0.05). The MI size 24 hours after occlusion was reduced by G-CSF pretreatment (36 +/- 3% versus 44 +/- 2% of left ventricle in MI group; P < 0.05). The increase of Cx43 expression in the heart may explain the reduced incidence in ventricular arrhythmias in the early phases after coronary artery occlusion in rats, thus increasing survival after MI.

Posttranscriptional regulation of sodium-iodide symporter mRNA expression in the rat thyroid gland by acute iodide administration

Serrano-Nascimento C, Calil-Silveira J, Nunes MT. Posttranscriptional regulation of sodium-iodide symporter mRNA expression in the rat thyroid gland by acute iodide administration. Am J Physiol Cell Physiol 298: C893-C899, 2010. First published January 27, 2010; doi:10.1152/ajpcell.00224.2009.-Iodide is an important regulator of thyroid activity. Its excess elicits the Wolff-Chaikoff effect, characterized by an acute suppression of thyroid hormone synthesis, which has been ascribed to serum TSH reduction or TGF-beta increase and production of iodolipids in the thyroid. These alterations take hours/days to occur, contrasting with the promptness of Wolff-Chaikoff effect. We investigated whether acute iodide administration could trigger events that precede those changes, such as reduction of sodium-iodide symporter (NIS) mRNA abundance and adenylation, and if perchlorate treatment could counteract them. Rats subjected or not to methylmercaptoimidazole treatment (0.03%) received NaI (2,000 mu g/0.5 ml saline) or saline intraperitoneally and were killed 30 min up to 24 h later. Another set of animals was treated with iodide and perchlorate, in equimolar doses. NIS mRNA content was evaluated by Northern blotting and real-time PCR, and NIS mRNA poly(A) tail length by rapid amplification of cDNA ends-poly(A) test (RACE-PAT). We observed that NIS mRNA abundance and poly(A) tail length were significantly reduced in all periods of iodide treatment. Perchlorate reversed these effects, indicating that iodide was the agent that triggered the modifications observed. Since the poly(A) tail length of mRNAs is directly associated with their stability and translation efficiency, we can assume that the rapid decay of NIS mRNA abundance observed was due to a reduction of its stability, a condition in which its translation could be impaired. Our data show for the first time that iodide regulates NIS mRNA expression at posttranscriptional level, providing a new mechanism by which iodide exerts its autoregulatory effect on thyroid.

Acute exhaustive exercise regulates IL-2, IL-4 and MyoD in skeletal muscle but not adipose tissue in rats

Background: The purpose of this study was to evaluate the effect of exhaustive exercise on proteins associated with muscle damage and regeneration, including IL-2, IL-4 and MyoD, in extensor digitorum longus (EDL) and soleus muscles and mesenteric (MEAT) and retroperitoneal adipose tissues (RPAT). Methods: Rats were killed by decapitation immediately (E0 group, n = 6), 2 (E2 group, n = 6) or 6 (E6 group, n = 6) hours after the exhaustion protocol, which consisted of running on a treadmill at approximately 70% of VO(2max) for fifty minutes and then at an elevated rate that increased at one m/min every minute, until exhaustion. Results: The control group (C group, n = 6) was not subjected to exercise. IL-2 protein expression increased at E0 in the soleus and EDL; at E2, this cytokine returned to control levels in both tissues. In the soleus, IL-2 protein expression was lower than that in the control at E6. IL-4 protein levels increased in EDL at E6, but the opposite result was observed in the soleus. MyoD expression increased at E6 in EDL. Conclusion: Exhaustive exercise was unable to modify IL-2 and IL-4 levels in MEAT and RPAT. The results show that exhaustive exercise has different effects depending on which muscle is analysed.

Reduced maximal oxygen consumption and overproduction of proinflammatory cytokines in athletes

Objective: It was the aim of this study to evaluate whether chronic pain in athletes is related to performance, measured by the maximum oxygen consumption and production of hormones and cytokines. Methods: Fifty-five athletes with a mean age of 31.9 +/- 4.2 years engaged in regular competition and showing no symptoms of acute inflammation, particularly fever, were studied. They were divided into 2 subgroups according to the occurrence of pain. Plasma concentrations of adrenaline, noradrenaline, cortisol, prolactin, growth hormone and dopamine were measured by radioimmunoassay, and the production of the cytokines interleukin (IL)-1, IL-2, IL-4, IL-6, tumor necrosis factor-alpha, interferon-alpha and prostaglandin E-2 by whole-blood culture. Maximal oxygen consumption was determined during an incremental treadmill test. Results: There was no change in the concentration of stress hormones, but the athletes with chronic pain showed a reduction in maximum oxygen consumption (22%) and total consumption at the anaerobic threshold (25%), as well as increased cytokine production. Increases of 2.7-, 8.1-, 1.7- and 3.7-fold were observed for IL-1, IL-2, tumor necrosis factor-alpha and interferon-alpha, respectively. Conclusions: Our data show that athletes with chronic pain have enhanced production of proinflammatory cytokines and lipid mediators and reduced performance in the ergospirometric test. Copyright (c) 2008 S. Karger AG, Basel.

Toxoplasma gondii in the peripheral blood of patients with acute and chronic toxoplasmosis

Background and aims Toxoplasmic retinochoroiditis may recur months or years after the primary infection. Rupture of dormant cysts in the retina is the accepted hypothesis to explain recurrence. Here, the authors present evidence supporting the presence of Toxoplasma gondii in the peripheral blood of immunocompetent patients. Methods Direct observation by light microscopy and by immunofluorescence assay was performed, and results were confirmed by PCR amplification of parasite DNA. Results The authors studied 20 patients from Erechim, Brazil, including acute infected patients, patients with recurrent active toxoplasmic retinochoroiditis, patients with old toxoplasmic retinal scars, and patients with circulating IgG antibodies against T gondii and absence of ocular lesions. Blood samples were analysed, and T gondii was found in the blood of acutely and chronically infected patients regardless of toxoplasmic retinochoroiditis. Conclusions The results indicate that the parasite may circulate in the blood of immunocompetent individuals and that parasitaemia could be associated with the reactivation of the ocular disease.

THE ROLE OF INNATE IMMUNITY IN SEPTIC ACUTE KIDNEY INJURIES

Acute kidney injury (AKI) is an important clinical syndrome characterized by abnormalities in the hydroelectrolytic balance. Because of high rates of morbidity and mortality (from 15% to 60%) associated with AKI, the study of its pathophysiology is critical in searching for clinical targets and therapeutic strategies. Severe sepsis is the major cause of AKI. The host response to sepsis involves an inflammatory response, whereby the pathogen is initially sensed by innate immune receptors (pattern recognition receptors [PRRs]). When it persists, this immune response leads to secretion of proinflammatory products that induce organ dysfunction such as renal failure and consequently increased mortality. Moreover, the injured tissue releases molecules resulting from extracellular matrix degradation or dying cells that function as alarmines, which are recognized by PRR in the absence of pathogens in a second wave of injury. Toll-like receptors (TLRs) and NOD-like receptors (NLRs) are the best characterized PRRs. They are expressed in many cell types and throughout the nephron. Their activation leads to translocation of nuclear factors and synthesis of proinflammatory cytokines and chemokines. TLRs` signaling primes the cells for a robust inflammatory response dependent on NLRs; the interaction of TLRs and NLRs gives rise to the multiprotein complex known as the inflammasome, which in turn activates secretion of mature interleukin 1 beta and interleukin 18. Experimental data show that innate immune receptors, the inflammasome components, and proinflammatory cytokines play crucial roles not only in sepsis, but also in organ-induced dysfunction, especially in the kidneys. In this review, we discuss the significance of the innate immune receptors in the development of acute renal injury secondary to sepsis.

Bone marrow mononuclear cells attenuate fibrosis development after severe acute kidney injury

One of the early phases that lead to fibrosis progression is inflammation. Once this stage is resolved, fibrosis might be prevented. Bone marrow mononuclear cells (BMMCs) are emerging as a new therapy for several pathologies, including autoimmune diseases, because they enact immunosuppression. In this study we aimed to evaluate the role of BMMC administration in a model of kidney fibrosis induced by an acute injury. C57Bl6 mice were subjected to unilateral severe ischemia by clamping the left renal pedicle for 1 h. BMMCs were isolated from femurs and tibia, and after 6 h of reperfusion, 1 x 10(6) cells were administrated intraperitoneally. At 24 h after surgery, treated animals showed a significant decrease in creatinine and urea levels when compared with untreated animals. Different administration routes were tested. Moreover, interferon (IFN) receptor knockout BMMCs were used, as this receptor is necessary for BMMC activation. Labeled BMMCs were found in ischemic kidney on FACS analysis. This improved outcome was associated with modulation of inflammation in the kidney and systemic modulation, as determined by cytokine expression profiling. Despite non-amelioration of functional parameters, kidney mRNA expression of interleukin (IL)-6 at 6 weeks was lower in BMMC-treated animals, as were levels of collagen 1, connective tissue growth factor (CTGF), transforming growth factor-beta (TGF-beta) and vimentin. Protective molecules, such as IL-10, heme oxygenase 1 (HO-1) and bone morphogenetic 7 (BMP-7), were increased in treated animals after 6 weeks. Moreover, Masson and Picrosirius red staining analyses showed less fibrotic areas in the kidneys of treated animals. Thus, early modulation of inflammation by BMMCs after an ischemic injury leads to reduced fibrosis through modulation of early inflammation.

Alboserpin, a Factor Xa Inhibitor from the Mosquito Vector of Yellow Fever, Binds Heparin and Membrane Phospholipids and Exhibits Antithrombotic Activity

The molecular mechanism of factor Xa (FXa) inhibition by Alboserpin, the major salivary gland anticoagulant from the mosquito and yellow fever vector Aedes albopictus, has been characterized. cDNA of Alboserpin predicts a 45-kDa protein that belongs to the serpin family of protease inhibitors. Recombinant Alboserpin displays stoichiometric, competitive, reversible and tight binding to FXa (picomolar range). Binding is highly specific and is not detectable for FX, catalytic site-blocked FXa, thrombin, and 12 other enzymes. Alboserpin displays high affinity binding to heparin (K(D) similar to 20 nM), but no change in FXa inhibition was observed in the presence of the cofactor, implying that bridging mechanisms did not take place. Notably, Alboserpin was also found to interact with phosphatidylcholine and phosphatidylethanolamine but not with phosphatidylserine. Further, annexin V (in the absence of Ca(2+)) or heparin outcompetes Alboserpin for binding to phospholipid vesicles, suggesting a common binding site. Consistent with its activity, Alboserpin blocks prothrombinase activity and increases both prothrombin time and activated partial thromboplastin time in vitro or ex vivo. Furthermore, Alboserpin prevents thrombus formation provoked by ferric chloride injury of the carotid artery and increases bleeding in a dose-dependent manner. Alboserpin emerges as an atypical serpin that targets FXa and displays unique phospholipid specificity. It conceivably uses heparin and phosphatidylcholine/phosphatidylethanolamine as anchors to increase protein localization and effective concentration at sites of injury, cell activation, or inflammation.

Modulation of inflammatory response by selective inhibition of cyclooxygenase-1 and cyclooxygenase-2 in acute kidney injury

This work explored the role of inhibition of cyclooxygenases (COXs) in modulating the inflammatory response triggered by acute kidney injury. C57Bl/6 mice were used. Animals were treated or not with indomethacin (IMT) prior to injury (days -1 and 0). Animals were subjected to 45 min of renal pedicle occlusion and sacrificed at 24 h after reperfusion. Serum creatinine and blood urea nitrogen, reactive oxygen species (ROS), kidney myeloperoxidase (MPO) activity, and prostaglandin E2 (PGE(2)) levels were analyzed. Tumor necrosis factor (TNF)-alpha, t-bet, interleukin (IL)-10, IL-1 beta, heme oxygenase (HO)-1, and prostaglandin E synthase (PGES) messenger RNA (mRNA) were studied. Cytokines were quantified in serum. IMT-treated animals presented better renal function with less acute tubular necrosis and reduced ROS and MPO production. Moreover, the treatment was associated with lower expression of TNF-alpha, PGE(2), PGES, and t-bet and upregulation of HO-1 and IL-10. This profile was mirrored in serum, where inhibition of COXs significantly decreased interferon (IFN)-gamma, TNF-alpha, and IL-12 p70 and upregulated IL-10. COXs seem to play an important role in renal ischemia and reperfusion injury, involving the secretion of pro-inflammatory cytokines, activation of neutrophils, and ROS production. Inhibition of COX pathway is intrinsically involved with cytoprotection.

Toll-like receptors-related genes in kidney transplant patients with chronic allograft nephropathy and acute rejection

Introduction: Toll-like receptors (TLR) comprehend an emerging family of receptors that recognize pathogen-associated molecular patterns and promote the activation of leukocytes. Surgical trauma and ischemia-reperfusion injury are likely to provide exposure to endogenous ligands for TLR in virtually all kidney transplant recipients. Methods: Macroarray (GEArray OHS-018.2 Series-Superarray) analyses of 128 genes involved in TLR signaling pathway were performed in nephrectomy samples of patients with chronic allograft nephropathy (CAN) and acute rejection (AR, vascular and non vascular). The analysis of each membrane was performed by GEArray Expression Analysis Suite 2.0. Results: Macroarray profile identified a gene expression signature that could discriminate CAN and AR. Three genes were significantly expressed between CAN and vascular AR: Pellino 2; IL 8 and UBE2V1. In relation to vascular and non-vascular AR, there were only two genes with statistical significance: IL-6 and IRAK-3. Conclusion: Vascular and non-vascular AR and CAN showed different expression of a few genes in TLR pathway. The analysis of nephrectomy showed that activation of TLR pathway is present in AR and CAN. (C) 2008 Elsevier B.V. All rights reserved.

Early modulation of inflammation by mesenchymal stem cell after acute kidney injury

Therapy with stem cells has showed to be promising for acute kidney injury (AKI), although how it works is still controversial. Modulation of the inflammatory response is one possible mechanism. Most of published data relies on early time and whether the protection is still maintained after that is not known. Here, we analyzed whether immune modulation continues after 24 h of reperfusion. MSC were obtained from male Wistar rats. After 3-5 passages, cells were screened for CD73, CD90, CD44, CD45, CD29 and CD 31. In addition, MSC were submitted to differentiation in adipocyte and in osteocyte. AKI was induced by bilaterally clamping of renal pedicles for 60 min. Six hours after injury, MSC (2 x 105 cells) were administered intravenously. MSC-treated animals presented the lowest serum creatinine compared to non-treated animals (24 h: 1.3 +/- 0.21 vs. 3.23 +/- 0.89 mg/dl, p<0.05). The improvement in renal function was followed by a lower expression of IL-1b, IL-6 and TNF-alpha and higher expression of IL-4 and IL-10. However, 48 h after reperfusion, this cytokine profile has changed. The decrease in Th1 cytokines was less evident and IL-6 was markedly up regulated. PCNA analysis showed that regeneration occurs faster in kidney tissues of MSC-treated animals than in controls at 24 h. And also ratio of Bcl-2/Bad was higher at treated animals after 24 and 48 h. Our data demonstrated that the immunomodulatory effects of MSC occur at very early time point, changing the inflammation profile toward a Th2 profile. (C) 2009 Elsevier B.V. All rights reserved.

A role for regulatory T cells in renal acute kidney injury

Ischemia reperfusion injury (IRI) is a potential contributor for the development of chronic allograft nephropathy. T cells are important mediators of injury, even in the absence of alloantigens. We performed a depletion of TCD4(+)CTLA4(+)Foxp3(+) cells with anti-CD25(PC61), a treatment with anti-GITR (DTA-1) and rat-IgG, followed by 45 min of ischemia and 24/72 h of reperfusion, and then analyzed blood urea, kidney histopathology and gene expression in kidneys by QReal Time PCR. After 24 h of reperfusion, depletion of TCD4(+)CTLA4(+)Foxp3(+) cells reached 30.3%(spleen) and 67.8%(lymph nodes). 72 h after reperfusion depletion reached 43.1%(spleen) and 90.22%(lymph nodes) and depleted animals presented with significantly poorer renal function, while DTA-1 (anti-GITR)-treated ones showed a significant protection, all compared to serum urea from control group (IgG: 150.10 +/- 50.04; PC61: 187.23 +/- 31.38; DTA-1: 64.53 +/- 25.65, mg/dL, p<0.05). These data were corroborated by histopathology. We observed an increase of HO-1 expression in animals treated with DTA-1 at 72 h of reperfusion with significant differences. Thus, our results suggest that PC61 (anti-CD25) mAb treatment is deleterious, while DTA-1 (anti-GITR) mAb treatment presents a protective role in the renal IRI, indicating that some regulatory populations of T cells might have a role in IRI. (C) 2009 Elsevier B.V. All rights reserved.