Markers Of Tissue Injury Research Articles

The Biologic Effects of Traction in Hip Arthroscopy (SS-41)

<h3>Introduction</h3> The purpose of this study was to evaluate the effects of traction on the lower extremity during hip arthroscopy by perioperative monitoring of venous blood flow, nerve conduction, and markers of soft tissue injury, blood clots, and patient pain. <h3>Methods</h3> (In progress at the time of this report.) In a prospective, nonrandomized design 30 subjects who had elected to undergo hip arthroscopy that required traction for access to the central compartment of the hip joint were analyzed. Measurements of visual analog pain scale (VAS), creatine phosphokinase (CPK-MM), and D-dimer were obtained preoperatively (pre), postoperatively (post), and 5 days postoperatively (5d). Perioperative documentation included operating room time, traction time, and force of traction. The sample population was divided into two cohorts, a Doppler ultrasound group (DUG, n=15) and a somatosensory evoked potentials group (SSEP, n=15). DUG were monitored for venous blood flow at the level of the femoral and popliteal veins prior to traction, and following application and removal of traction. SSEP of the posterior tibial nerve (PTN) and the superficial peroneal nerve (SPN) were monitored intra-operatively in 5-15 minute intervals beginning prior to traction and continued through the end of surgery. <h3>Results</h3> (Preliminary results at the time of this report, n=19.) Average operating room time was 114.9 minutes. Traction was applied for an average of 26.5 minutes at an average of 57.1 lbs. With the application of traction, 15 of 15 subjects in DUG experienced >50% decreased blood flow of the popliteal vein and 13% (2 subjects) experience decreased blood flow at the femoral vein. Blood flow returned to baseline following removal of traction in all subjects. Average CPK-MM levels were: pre=81.5 ± 35.8 milliunits/mL, post=173.8 ± 130.5 milliunits/mL, and 5d=120.3 ± 71.5 milliunits/mL. Two subjects were D-dimer positive at baseline which remained positive through 5d post. At the time of this report 4 SSEP subjects were complete. One of four showed >50% decrease in amplitude of the PTN in both the operative and non-operative legs, which returned to normal within 10 minutes of knee flexion of both legs. There were no significant correlations determined between VAS, BMI, CPK-MM, traction time, or operating room time. <h3>Conclusion</h3> The Doppler ultrasound group demonstrated greater than 50% decrease in blood flow at the popliteal vein with the application of traction, which returned to normal after the removal of traction. SSEP demonstrated changes both with and without traction on both operative and non-operative legs of the posterior peroneal nerve. Consideration should be given for knee flexion of the contralateral leg when traction is discontinued for the protection of the peroneal nerve. There is variability in the soft tissue damage with hip arthroscopy, which is independent of time (under 2 hours), BMI, or correlations of VAS. Traction in the present form affects vascular and neurologic structure of the operative and non-operative extremity independent of time. A better method to create space in the central compartment and prevent harm to the soft tissue, vascular, and neural structures is needed.

NADH as an Endogenous Marker of Cardiac Tissue Injury at the Site of Radiofrequency Ablation

Atrial fibrillation (AF) remains the most commonly occurring cardiac arrhythmia; its incidence is associated with a lower quality of life including a higher rate of morbidity & mortality. Percutaneous left atrial catheter ablation for the purpose of eliminating AF has become a common treatment option and is currently being investigated for superiority over medical therapy in a large clinical trial. RFA produces lesions that can block the spread of electrical activity from the sites of abnormal activity including pulmonary veins. Today, there are limited means for real time monitoring of tissue injury during the RFA procedure. To address this need, we explored the fluorescence of endogenous NADH as a possible live marker of tissue injury during the ablation procedure. Studies were conducted in blood-free and blood-perfused isolated rat hearts. Epicardial RFA lesions were seen as areas of low NADH fluorescence which encompassed both irreversible and reversibly damaged tissue. Their size significantly exceeded TTC negative staining (TTC - Triphenyl Tetrazolium Chloride, a redox indicator used to differentiate between metabolically active and inactive tissues). Real-time monitoring of NADH fluorescence allowed visualization of gaps of viable tissue between the RFA lesions. Dual recordings of NADH and epicardial electrical activity linked these gaps to the occurrence of post-ablation reentries. We conclude that fluorescence of endogenous NADH can assist visualization of injured epicardial tissue near the tip of the RFA catheter lesions.

Levosimendan reduces plasma cell-free DNA levels in patients with ischemic cardiomyopathy

Heart failure (HF) is a condition associated with the apoptosis and cell death of both cardiac myocytes and cardiac non-myocytes. DNA fragments released from programmed cell death or acute cellular injury are the main sources of disease-associated elevation of cell-free (cf) DNA. We hypothesized that cfDNA could be a relevant marker of cardiac apoptosis in HF patients that could be affected by the improvement of myocardial performance. To test our hypothesis, we measured plasma cfDNA in 19 patients with ischemic HF and severe left ventricular (LV) systolic dysfunction before and 12 h after completion of levosimendan infusion. Echocardiographic and biochemical markers of LV diastolic pressure and LV systolic function were also assessed. In accordance with previous observations levosimendan improved echocardiographic and biochemical indices of LV function. Plasma cfDNA was significantly reduced in HF patients post-levosimendan treatment (median: 89.4, interquartile range: 87.1 to median: 25.9, interquartile range: 12.3, P = 0.028). Notably, in 15/19 patients there was a reduction in cfDNA levels post-levosimendan infusion; while in 12/19 patients, a more than 50% reduction in plasma cfDNA was observed. Since cfDNA is a marker of tissue injury and apoptosis these results indicate that improvement of LV function has a potential impact on cell preservation and survival. Further studies are needed to substantiate our promising results regarding the role of plasma cfDNA as a marker of HF.

Inflammatory and Hemostatic Response to Cardiopulmonary Bypass in Pediatric Population: Feasibility of Seriological Testing of Multiple Biomarkers

Perioperative myocardial and cerebral damages are the major determinants of postoperative morbidity and mortality in pediatric cardiac surgery. Cardiopulmonary bypass (CPB) causes alterations in the levels of biomarkers related to inflammation, tissue damage, and other tissue pathologies. Early and accurate evaluation of inflammation and tissue damage would therefore be clinically useful. Our objective is to assess the suitability of using Multi-Analyte Profiling (MAP) (Rules Based Medicine, Austin, TX, USA) in pediatric cardiac surgery as a potential surrogate marker of clinical outcome. MAP technology platform allowed us to analyze 90 different biomarkers using only 100µL of plasma to detect any changes in the levels of 90 biomarkers. Plasma samples (100µL) were collected at five different time points: 1. before midline incision; 2. on CPB for 3-5min; 3. at the end of CPB; 4. 1h after CPB; and 5. 24h after CPB. After removing the outliers, the average and standard deviation of the values obtained from the 10 patients were calculated for each time point. The average values of each biomarker at each time point were then compared to each other and to the baseline. The pilot protocol included 10 patients (ages from 3 months to 4 years old) with similar Jenkins risks stratifications who underwent nonpulsatile CPB. We detected changes in the levels of 90 biomarkers. Biomarkers were assessed in groups. Myeloperoxidase (MPO) and pregnancy-associated plasma protein A (PAPP-A) were the earliest markers to rise with 49- and 18-fold increases 3-5min after the onset of CPB, respectively. The most striking increase was noted in the heart-type fatty acid-binding protein (FABP) levels. FABP increased 25, 193, 151, and 4-fold at time points 2, 3, 4, and 5, respectively. Surges in the novel markers of injury were followed by the markers of inflammation (i.e., C-reactive protein, interleukins) peaking at 24h after CPB. This pilot study shows that it is possible to measure 90 different biomarkers using only a very small sample of plasma to evaluate the effects of CPB. Novel markers of tissue injury (FABP, PAPP-A, or MPO) are the earliest markers to rise. Serial monitoring of multiple biomarkers may help to predict and improve outcomes after pediatric cardiac surgery.

Follistatin-like 1 regulates renal IL-1β expression in cisplatin nephrotoxicity

Follistatin-like 1 (FSTL1) is a secreted protein with homology to both Follistatin and the SPARC/BM40 family of matricellular proteins. In this study, we sought to determine the expression patterns of Fstl1 and its cognate receptor Dip2a in the adult, and to assess the consequences of Fstl1 inactivation on development and homeostasis of the kidney. We find that FSTL1 circulates at high levels in both the human and the mouse and that it is also locally expressed in the loop of Henle in the kidney. To begin to understand the in vivo functions of Fstl1, we generated a mouse mutant using a genetrap approach. The hypomorphic Fstl1 genetrap strain displays a strong reduction in FSTL1 expression at the protein level, but it does not show overt developmental defects. FSTL1 has previously been implicated in diverse disease processes as a regulator of inflammatory cytokine expression, and we therefore evaluated the response of the genetrap strain to cisplatin-mediated acute kidney injury, a disease model with highly cytokine-dependent pathology. We find that although TNF-α and Il6 levels are unchanged relative to wild-type, renal Il-1β expression is increased in genetrap mice following cisplatin treatment. Furthermore, histopatological analysis, expression of the tissue injury marker Havcr1, and measurement of serum creatinine demonstrate that reduction of Fstl1 expression sensitizes the kidney to acute cisplatin nephrotoxicity, suggesting a role for FSTL1-mediated Il-1β suppression in protection of the kidney from acute nephrotoxic injury.

Diarrhea during the Conditioning Regimen Is Correlated with the Occurrence of Severe Acute Graft-versus-Host Disease through Systemic Release of Inflammatory Cytokines

Graft-versus-host disease (GVHD) is a major obstacle to the success of allogeneic hematopoietic stem cell transplantation (allo-HSCT). Although many indicators have been used to predict GVHD, the prediction of GVHD remains very difficult. Determining how to predict the occurrence of acute GVHD (aGVHD) as exactly as possible has been a huge challenge so far. Systemic release of inflammatory cytokines during the conditioning regimen and early-phase post-HSCT plays a crucial role in the generation of aGVHD; additionally, the conditioning regimen causes the damage to the intestinal tract. Diarrhea is a common symptom of intestinal tract damage during the conditioning regimen. We therefore performed a prospective study to investigate the relationship between diarrhea related to the conditioning regimen, systemic release of inflammatory cytokines during the conditioning regimen and the early-phase post-HSCT, and the development of aGVHD. This study demonstrated that duration of diarrhea was >5.5 days, with a maximal volume >8.72 mL/kg and a mean volume of diarrhea for days -3 to 0 >7.94 mL/kg were risk factors of grade II to IV aGVHD. Diarrhea with any 1 risk factor was defined as severe diarrhea. Furthermore, this study first confirmed that the correlation between diarrhea and aGVHD was related to the serum levels of tumor necrosis factor (TNF)-α and interleukin (IL)-6 during the conditioning regimen and during the early phase after transplantation. Our study demonstrated that diarrhea related to the conditioning regimen could be used as a marker for the prediction of aGVHD and further explain the possible mechanism underlying the linkage described here. Therefore, for the patients with severe diarrhea related to the conditioning regimen, low-dose glucocorticoid may be used to reduce the levels of inflammatory cytokine release by a damaged intestinal tract, and possibly further reduce the occurrence of aGVHD. For these patients, prophylaxis of aGVHD may be need to be adjusted individually.

Partial bile duct ligation in mice: A novel model of acute cholestasis

The standard model for research in cholestasis is the total ligation of the bile duct (tBDL). Because this model causes severe hepatic injury in mice, we developed a novel model of cholestasis using a partial bile duct ligation (pBDL) and evaluate different mechanisms of injury. Male C57Bl/6 mice were subjected to sham operation, tBDL, or pBDL. Blood from tail veins was taken repeatedly until day 14 after surgery to assess markers of tissue injury (aspartate aminotransferase [AST]) and cholestasis (bilirubin, alkaline phosphatase [AP]). Also, liver samples were obtained at various time points to determine the histologic injury (hematoxylin and eosin) and tissue repair (Ki67). In addition, the biliary pressure and serum bile acids were evaluated as potential mechanisms of injury. Both models of cholestasis were equal in terms of bilirubin, AST, and AP serum levels during the first week of the experiment. Although these parameters remained constantly elevated thereafter in the tBDL model, all parameters normalized within the second week after pBDL. Moreover, pBDL resulted in significantly less necrosis formation (P = .001) and consequent hepatocyte proliferation (P= .01). Most important, serum bile acid levels (P = .04) and biliary pressures (P = .02) were significantly lower after pBDL than after tBDL and were the best predictors for hepatic necrosis formation. We established a model of acute cholestasis, which is ideal for research in resolved acute cholestasis (eg, surgery for Klatskin tumors). Moreover, biliary pressure and toxic bile acid serum levels may be better predictors of cholestatic liver injury than standard laboratory parameters.

IL-17 Is Necessary for Host Protection against Acute-Phase Trypanosoma cruzi Infection

IL-17A is a key cytokine that induces inflammatory responses through the organized production of inflammatory cytokines, such as IL-6, TNF-alpha, and GM-CSF, and induces neutrophil migration. The roles of IL-17A in infection of intracellular protozoan parasites have not been elucidated, although augmented immune responses by IL-17A are important for the resolution of some bacterial and fungal infections. Therefore, we experimentally infected IL-17A-deficient (IL-17A(-/-)) mice with Trypanosoma cruzi. IL-17A(-/-) mice had a lower survival rate and prolonged worse parasitemia compared with control C57BL/6 wild-type (WT) mice postinfection. In the infected IL-17A(-/-) mice, multiple organ failure was observed compared with WT mice, as reflected by the marked increase in serologic markers of tissue injury, such as aspartate aminotransferase, which resulted in increased mortality of IL-17A(-/-) mice. Expression of cytokines, such as IFN-gamma, IL-6, and TNF-alpha, was lower in liver-infiltrating cells from the IL-17A(-/-) mice compared with WT mice. A similar defect was observed in the expression of neutrophil enzymes, such as myeloperoxidase and lipoxygenase, whereas cellular infiltration into the infected tissues was not affected by IL-17A deficiency. These results suggested that the efficient activation of immune-related cells critical for the killing of T. cruzi was impaired in the absence of IL-17A, resulting in the greater susceptibility of those mice to T. cruzi infection. From these results, we conclude that IL-17A is important for the resolution of T. cruzi infection.

Effects of a Short-acting β1 Receptor Antagonist Landiolol on Hemodynamics and Tissue Injury Markers in Patients With Subarachnoid Hemorrhage Undergoing Intracranial Aneurysm Surgery

Sympathetic activation after subarachnoid hemorrhage (SAH) can induce tachycardia as well as cardiac and brain injury. We examined the effects of beta1 receptor antagonist landiolol on hemodynamics and the levels of tissue injury markers in patients with SAH. Fifty-six SAH patients undergoing intracranial aneurysm surgery with tachycardia (>or=90 beats per minute) randomly allocated to landiolol (L) or control (C) group were examined. In L group, landiolol was continuously administered during anesthesia. In C group, landiolol was not administered except bolus dose used in cases that exhibited uncontrolled tachycardia. Hemodynamics, the incidence of electrocardiographic abnormality, and levels of B-type natriuretic peptide, troponin T, S-100beta, 8-Hydroxy-2'-deoxyguanosine, interleukin-6 (IL-6), and IL-1 receptor antagonist were compared. Heart rate values from time of intubation to the end of anesthesia were significantly lower in L group than in C group, whereas blood pressure was similar between the groups. Although the incidence of bradycardia (<60 beats per minute) was significantly higher in L group than in C group (57% vs. 18%, respectively), bradycardia could be recovered without any adverse effects. The serum S-100beta levels 24 hours after operation were significantly lower in L group than in C group, whereas there were no significant differences in the incidence of electrocardiographic abnormality and levels of B-type natriuretic peptide, troponin T, 8-Hydroxy-2'-deoxyguanosine, IL-6, and IL-1 receptor antagonist between groups. We conclude that landiolol can be effectively used in the treatment of tachycardia in SAH patients and significantly reduced the serum S-100beta levels 24 hours after the operation.

Dilated intercellular spaces in subtypes of gastroesophagic reflux disease

Dilatation of the intercellular spaces by electron microscopy has been considered as an early morphological marker of tissue injury in gastroesophageal reflux disease. The degree of dilatation in Barret's esophagus in currently unknown. To determine the frequency of dilated intervellular spaces in Barrett's esophagus. Cross-sectional and prospective analysis of consecutive patients with gastroesophageal reflux disease. We selected symptomatic patients > 18 years and both sexes. Patients with recent PPI use (< 14 days), H-2 antagonists, NSAID's or previous upper GI tract surgery were excluded. VARIABLES INCLUDED: Clinical-demographic data, Carlsson-Dent score, conventional endoscopy findings, pH-metry results (in non-erosive) and normal mucosal biopsies at 2 and 5 cm above the squamocolumnar junction. Dilation of intercellular spaces was measured by electron microscopy. Chi square test with a significance level 0.05 was calculated. The following four groups were compared: a) non-erosive reflux disease (n = 14); b) erosive esophagitis (n = 5); c) Barrett's esophagus (n = 13); and d) healthy controls (n = 5). the dilation of intervellular spaces was increasingly greater form non-erosive revlux to Barrett's esophagus and higher in biopsies taken at 5 cm than at 2 cm of the squamous columnar junction (2.72 +/- 1.35 vs. 1.71 +/- 0.48 microg) (p = 0.001). There was no difference between biopsies at 2 and 5 cm in the order groups. dilation of intercellular spaces was greater in Barrett s esophagus than in the other groups and higher at 5 cm from the squamocolumnar junction.

Attenuation of Renal Ischemia and Reperfusion Injury by Human Adrenomedullin and its Binding Protein

Acute renal failure secondary to ischemia and reperfusion (I/R) injury poses a significant burden on both surgeons and patients. It carries a high morbidity and mortality rate and no specific treatment currently exists. Major causes of renal I/R injury include trauma, sepsis, hypoperfusion, and various surgical procedures. We have demonstrated that adrenomedullin (AM), a novel vasoactive peptide, combined with AM binding protein-1 (AMBP-1), which augments the activity of AM, is beneficial in various disease conditions. However, it remains unknown whether human AM/AMBP-1 provides any beneficial effects in renal I/R injury. The objective of our study therefore was to determine whether administration of human AM/AMBP-1 can prevent and/or minimize damage in a rat model of renal I/R injury. Male adult rats were subjected to renal I/R injury by bilateral renal pedicle clamping with microvascular clips for 60 min followed by reperfusion. Human AM (12 microg/kg BW) and human AMBP-1 (40 microg/kg BW) or vehicle (52 microg/kg BW human albumin) were given intravenously over 30 min immediately following the clip removal (i.e., reperfusion). Rats were allowed to recover for 24 h post-treatment, and blood and renal tissue samples were collected. Plasma levels of AM were measured using a radioimmunoassay specific for rat AM. Plasma AMBP-1 was measured by Western analysis. Renal water content and serum levels of systemic markers of tissue injury were measured. Serum and renal TNF-alpha levels were also assessed. At 24 h after renal I/R injury, plasma levels of AM were significantly increased while plasma AMBP-1 was markedly decreased. Renal water content and systemic markers of tissue injury (e.g., creatinine, BUN, AST, and ALT) were significantly increased following renal I/R injury. Serum and renal TNF-alpha levels were also increased post injury. Administration of human AM/AMBP-1 decreased renal water content, and plasma levels of creatinine, BUN, AST, and ALT. Serum and renal TNF-alpha levels were also significantly decreased after AM/AMBP-1 treatment. Treatment with human AM/AMBP-1 in renal I/R injury significantly attenuated organ injury and the inflammatory response. Thus, human AM combined with human AMBP-1 may be developed as a novel treatment for patients with acute renal I/R injury.

Super-short solid silicon microneedles for transdermal drug delivery applications

In this study, the super-short microneedles with a length of 70–80 μm were fabricated using silicon wet etching technology. As evident from the visual inspection of pierced human skin, appearance of blue spots array after Evans Blue (EB) application indicated that the super-short microneedles were able to pierce into skin by pressing and swaying against the microneedles backing layer continually with a finger. The micro-conduits created in skin were validated by histological examination. Transport studies revealed that: (i) skin pretreated with super-short microneedles resulted in a remarkable enhancement of the galanthamine (GAL) transport, and the permeated amount increased as the insertion force increased; (ii) the super-short microneedles with flat tips were better than that with sharp tips for enhancing skin permeability; (iii) the longer time of super-short microneedles detained in skin resulted in a higher increase of skin permeability; (iv) there was no linear correlation between the GAL permeated through skin and the number of microneedles. The EIIIA + (526 bp) segment, a very sensitive marker of tissue injury, did not expressed in the skin pierced by super-short microneedles. Meanwhile, the skin pretreated with super-short microneedles did not infected after incubation with the Staphylococcus aureus. These results suggest that super-short microneedles may be a safe and efficient alternative for transdermal drug delivery of hydrophilic molecules.

Inclusion of a Nitric Oxide Congener in the Insufflation Gas Repletes S‐Nitrosohemoglobin and Stabilizes Physiologic Status During Prolonged Carbon Dioxide Pneumoperitoneum

A method to maintain organ blood flow during laparoscopic surgery has not been developed. Here we determined if ethyl nitrite, an S-nitrosylating agent that would maintain nitric oxide bioactivity (the major regulator of tissue perfusion), might be an effective intervention to preserve physiologic status during prolonged pneumoperitoneum. The study was conducted on appropriately anesthetized adult swine; the period of pneumoperitoneum was 240 minutes. Cohorts consisted of an anesthesia control group and groups insufflated with CO2 alone or CO2 containing fixed amounts of ethyl nitrite (1-300 ppm). Insufflation with CO2 alone produced declines in splanchnic organ blood flows and it reduced circulating levels of S-nitrosohemoglobin (i.e., nitric oxide bioactivity); these reductions were obviated by ethyl nitrite. In a specific example, preservation of kidney blood flow with ethyl nitrite kept serum creatinine and blood urea nitrogen concentrations constant whereas in the CO2 alone group both increased as kidney blood flow declined. The data indicate ethyl nitrite can effectively attenuate insufflation-induced decreases in organ blood flow and nitric oxide bioactivity leading to reductions in markers of acute tissue injury. This simple intervention provides a method for controlling a major source of laparoscopic-related morbidity and mortality: tissue ischemia and altered postoperative organ function.

Early versus delayed restoration of flow with temporary vascular shunt reduces circulating markers of injury in a porcine model.

Temporary vascular shunting to restore flow after vascular injury has been advocated. The effectiveness of this adjunct in protecting against ischemic injury has not been established. This study will assess the temporal impact of shunts on ischemic injury and arterial flow. A porcine model of hind-limb ischemia via iliac artery occlusion was used (N = 36; weight [kg] +/- SD: 89 +/- 4.4). Animals were randomized into one control (Iscctrl) and four study groups (Isc0, Isc1, Isc3, and Isc6) according to ischemic time. Shunt placement followed ischemia, and flow and circulating injury markers were collected incrementally during 18 hours of reperfusion. Flow proportions and a calculated Ischemia Injury Index were used to characterize group differences. There were no intergroup differences concerning initial weight, hemodynamic, or laboratory values. Shunt patency was 92% in the absence of anticoagulation. The proportion of common femoral arterial flow to baseline flow in the Isc6 group was lower than the Iscctrl group (p = 0.02). There was a similar trend with the Isc1 and Isc3 groups. The Ischemia Injury Index demonstrated that there was a difference in the Isc3 and Isc6 groups (late shunt placement) compared with the Iscctrl, Isc0, and Isc1 groups (early shunt placement) (p < 0.001). This study provides physiologic insight into the benefit of shunts in a model of extremity ischemia. Early shunting protects the extremity from further ischemic insult and reduces circulating markers of tissue injury. Additionally, the presence of a shunt does not increase the Ischemic Injury Index and patency is maintained in the absence of heparinization.

Heat Shock Protein 70 Secretion by Neonatal Tracheal Tissue During Mechanical Ventilation: Association With Indices of Tissue Function and Modeling

Mechanical ventilation (MV) of the neonatal airway alters mechanical properties and activates tissue-modeling pathways. Heat shock protein (HSP70) is a marker of tissue injury and modulates inflammation, which may influence subsequent pulmonary tissue modeling by matrix metalloproteinases (MMPs). HSP70 secretion is up-regulated in MV airway tissues and associated with changes in airway elasticity and secretion of MMPs. Proximal tracheal segments were isolated in 13 newborn lambs and were either MV for 4 h or SHAM. At baseline and hourly, tracheal segments were flushed and tracheal elasticity was determined. Tracheal wash fluid was assayed for HSP70 by ELISA and for MMPs by substrate zymography. HSP70 secretion increased from baseline to a peak at 1 h in both groups (p < 0.01), greater in the MV group (p < 0.05), and returned to baseline values by 2 h. This response was in contrast to the progressive decrease in tracheal elasticity (p < 0.05). The HSP70 elevation pattern was noted in MMP-2, but beyond 1 h, MMP-2 returned to baseline values in MV group but remained elevated in SHAM (p < 0.05). HSP70 secretion is associated with the degree of biophysical tracheal injury as well as the time course of MMP-2 secretion by tracheal tissues.

High Levels of Proinflammatory Cytokines, but Not Markers of Tissue Injury, in Unaffected Intestinal Areas from Patients with IBD

Intestinal alterations in IBD are triggered and maintained by an overexpression of proinflammatory cytokines. Additionally, increased immune activation has been found in the adjacent intestinal areas without displaying any apparent histological alterations, however, the regulatory environment is not well established. Biopsy specimens from patients with ulcerative colitis (UC) and Crohn's disease (CD), from both affected and unaffected areas, and also from a group of colonic biopsies from healthy controls, were included in our study. Cytokines and markers of mucosal damage were analyzed by real-time PCR, and some of the results confirmed by western-blot and ELISA. Levels of IFNγ, TNFα, IL-6, IL-15, IL-18, and IL-23 were increased (above healthy controls) in both affected and unaffected areas from IBD. IL-1β, IL-6, IL-12, and IL-27 were higher in affected areas compared to unaffected ones in UC but not CD. In general, a correlation was observed between mRNA levels of these cytokines and both iNOS and Granzyme B. SOCS-2 and SOCS-3 were also increased in the affected areas. In conclusion, the unaffected areas from IBD show increased levels of a restricted set of cytokines that may exert immune activating roles in these areas without being able to trigger tissue damage.

Intercellular space diameters of the oesophageal epithelium in NERD patients: Head to head comparison between light and electron microscopy analysis

Dilation of intercellular space diameters of oesophageal epithelium detected at transmission electron microscopy morphometry is a marker of tissue injury in non-erosive reflux disease patients. Semi-quantitative evaluation of intercellular space diameters using light microscopy seems to provide promising results. To comparatively evaluate intercellular space diameters in the same patients, by means of morphometry and semi-quantitative analysis, both on light microscopy and transmission electron microscopy microphotographs, biopsies were taken in 29 non-erosive reflux disease patients at distal and proximal oesophagus. Twelve asymptomatic controls underwent the same protocol. Morphometric analysis on transmission electron microscopy microphotographs showed mean intercellular space diameter values of patients, at distal and proximal oesophagus, 3- and 2-fold, respectively, higher than those in controls (p<0.001). On light microscopy microphotographs, mean intercellular space diameter values of patients at distal oesophagus were higher than those in controls, an overlap between patients and controls being observed. The semi-quantitative score was positive in 79% of patients and in 25% of controls at distal esophagus. Intercellular space diameter morphometric analysis at light microscopy is widely available, allows intercellular space diameter to be quantitatively measured with good sensitivity and specificity and could represent a useful tool in non-erosive reflux disease diagnosis. Despite satisfactory sensitivity, the semi-quantitative score at light microscopy is hampered by much lower specificity than transmission electron microscopy- and light microscopy-morphometry.

Glatiramer acetate after mitoxantrone induction improves MRI markers of lesion volume and permanent tissue injury in MS

Glatiramer acetate (GA) therapy following brief, low-dose induction with mitoxantrone was safe and more effective than GA alone in suppressing inflammatory disease activity, as determined by a significant reduction in gadolinium (Gd)- enhancing MRI lesions, in a 15- month, randomized, single-blind study of relapsing-remitting multiple sclerosis (RRMS) patients. To determine whether effects on MRI markers of disease burden and tissue damage support and extend data on the benefits of mitoxantrone induction therapy before initiation of long-term GA therapy. 40 RRMS patients, aged 18 to 55 years, with 1-15 Gd-enhancing lesions on screening MRI and EDSS score 0-6.5 were randomized to receive GA (20 mg/d SC), starting 2 weeks after the last of 3 monthly mitoxantrone infusions (36 mg/m2 total; n = 21), or to GA alone (20 mg/d SC; n = 19), for a total of 15 months. MRIs were obtained at baseline and months 6, 9, 12, and 15. At baseline, mean (+/- SD) age was 37.2 +/- 9.7 years; disease duration, 3.5 +/- 4.8 years; EDSS score, 2.3 +/- 1.1; and number of Gd-enhancing lesions, 3.75 +/- 3.95. Reductions in Gd-enhancing lesions (RR = 0.30, 95 % CI, 0.11-0.86, p = 0.0147) and relapse activity favoring mitoxantrone- GA were accompanied by significant differences in changes in T2w lesion volume (p = 0.0139), T1w hypointense lesion volume (p = 0.0303), and proportion of Gdenhancing lesions that evolved into black holes (p = 0.0023) compared with GA alone. Longterm continuous GA after brief, low-dose mitoxantrone induction is safe and more effective than GA alone. A trend toward decreased clinical disease activity was accompanied by major effects on MRI measures of disease burden and severe tissue injury.

234 A novel in vivo marker for ischemic tissue injury early after coronary occlusion

234 A novel in vivo marker for ischemic tissue injury early after coronary occlusion

P38 MAPK-dependent eNOS upregulation is critical for 17β-estradiol-mediated cardioprotection following trauma-hemorrhage

Studies have shown that p38 MAPK and nitric oxide (NO), generated by endothelial NO synthase (eNOS), play key roles under physiological and pathophysiological conditions. Although administration of 17beta-estradiol (E2) protects cardiovascular injury from trauma-hemorrhage, the mechanism by which E2 produces those effects remains unknown. Our objective was to determine whether the E2-mediated activation of myocardial p38 MAPK and subsequent eNOS expression/phosphorylation would protect the heart following trauma-hemorrhage. To study this, male Sprague-Dawley rats underwent soft-tissue trauma (midline laparatomy) and hemorrhagic shock (mean blood pressure 35-40 mmHg for 90 min), followed by fluid resuscitation. Animals were pretreated with specific p38 MAPK inhibitor SB-203580 (SB; 2 mg/kg), and nonselective NO synthase inhibitor NG-nitro-l-arginine methyl ester (l-NAME; 30 mg/kg) 30 min before vehicle (cyclodextrin) or E2 (100 microg/kg) treatment, followed by resuscitation, and were killed 2 h thereafter. Cardiovascular performance and other parameters were measured. E2 administration following trauma-hemorrhage increased cardiac p38 MAPK activity, eNOS expression and phosphorylation at Ser(1177), and nitrate/nitrite levels in plasma and heart tissues; these were associated with normalized cardiac performance, which was reversed by SB administration. In addition, E2 also prevented trauma-hemorrhage-induced increase in cytokines (IL-6 and TNF-alpha), chemokines (macrophage inflammatory protein-2 and cytokine-induced neutrophil chemoattractant-1), and ICAM-1, which was reversed by l-NAME administration. Administration of E2 following trauma-hemorrhage attenuated cardiac tissue injury markers, myeloperoxidase activity, and nitrotyrosine level, which were reversed by treatment with SB and l-NAME. The salutary effects of E2 on cardiac functions and tissue protection following trauma-hemorrhage are mediated, in part, through activation of p38 MAPK and subsequent eNOS expression and phosphorylation.