Severe acute respiratory syndrome coronavirus 2 prevalence in 1170 asymptomatic Norwegian conscripts.

BACKGROUND: Accurate estimates of SARS-CoV-2 infection in different population groups are important for the health authorities. In Norway, public infection control measures have successfully curbed the pandemic. However, military training and service are incompatible with these measures; therefore extended infection control measures were implemented in the Norwegian Armed Forces. We aimed to describe these measures, discuss their value, and investigate the polymerase chain reaction (PCR) prevalence and seroprevalence of SARS-CoV-2, as well as changes in antibody titer levels over the 6-week military training period in a young, asymptomatic population of conscripts. METHODS: In April 2020, 1170 healthy conscripts (median age 20 years) enrolled in military training. Extended infection control measures included a pre-enrollment telephone interview, self-imposed quarantine, questionnaires, and serial SARS-CoV-2 testing. At enrollment, questionnaires were used to collect information on symptoms, and SARS-CoV-2 rapid antibody testing was conducted. Serial SARS-CoV-2 PCR and serology testing were used to estimate the prevalence of confirmed SARS-CoV-2 and monitor titer levels at enrollment, and 3 and 6 weeks thereafter. RESULTS: At enrollment, only 0.2% of conscripts were SARS-CoV-2 PCR-positive, and seroprevalence was 0.6%. Serological titer levels increased nearly 5-fold over the 6-week observation period. Eighteen conscripts reported mild respiratory symptoms during the 2 weeks prior to enrollment (all were PCR-negative; one was serology-positive), whereas 17 conscripts reported respiratory symptoms and nine had fever at enrollment (all were PCR- and serology-negative). CONCLUSIONS: The prevalence of SARS-CoV-2 was less than 1% in our sample of healthy Norwegian conscripts. Testing of asymptomatic conscripts seems of no value in times of low COVID-19 prevalence. SARS-CoV-2 antibody titer levels increased substantially over time in conscripts with mild symptoms.

Asymptomatic COVID-19 with ambiguous test results. / Asymptomatisk covid-19 med usikre testresultater.

BACKGROUND: Testing for SARS-CoV-2 using polymerase chain reaction (PCR) and SARS-CoV-2 antibody tests is a significant part of the effort to combat the COVID-19 pandemic. Mass testing of healthy individuals raises several issues, however, and the results can be challenging to interpret. CASE PRESENTATION: A healthy 19-year-old man entered the military after two weeks of quarantine. The recruit had no respiratory symptoms or fever before, during or after his enrolment, and no history of SARS-CoV-2 exposure. At enrolment, he had a positive rapid test and a venous blood sample showed antibodies against SARS-CoV-2. PCR tests of specimens obtained from the upper respiratory tract were negative at enrolment and at week three, but were positive at week six. INTERPRETATION: The overall assessment of all the tests indicates a probable asymptomatic infection. This case report illustrates the challenge of interpreting screening results in asymptomatic individuals.

Postoperative administration of tranexamic acid as approach to reduce blood loss after open-heart surgery.

BACKGROUND: Tranexamic acid (TXA) reduces perioperative bleeding among patients undergoing heart surgery. It is uncertain whether its postoperative administration, after prior administration before cardiopulmonary bypass (CPB), has an additional benefit. OBJECTIVE: Our study aimed to evaluate whether the postoperative administration of TXA reduces the blood loss after heart surgery. METHODS: In a retrospective cohort study at the University Heart Center Dresden, patients who underwent on-pump open-heart surgery and received 1 g TXA before CPB were included. Patients with postoperative administration of 1 g TXA were compared to patients without. Primary endpoint was the postoperative blood loss within 24 hours. RESULTS: Among 2,179 patients undergoing heart surgery between 1 July 2013 and 31 October 2014, 92 (4.2%) received TXA postoperatively. After matching, 71 patients with postoperative administration of TXA were compared to 71 without (n = 142). Postoperative administration of TXA did not result in decreased blood loss (MD 146.7 mL; p = 0.064). There was no evidence of an increased risk for thromboembolic complications. CONCLUSIONS: The postoperative administration of TXA did not reduce blood loss. The use of TXA was shown to be safe in terms of thromboembolic events and hospital mortality. Unless there is no clear evidence, the postoperative administration of TXA should be restricted to patients with massive blood loss and signs of hyperfibrinolysis only.

Subclinical Endocarditis Might be a Hidden Trigger of Early Prosthetic Valve Calcification: A Histological Study.

OBJECTIVE: Despite various improvements in valve prosthetics, early valve deterioration still occurs, leading to prosthetic failure. Studying the early phase of this deterioration is quite difficult, as the prosthesis to be examined is almost always explanted only after extensive deterioration. The objective of this research is to study the pathology of early valve deterioration in an early stage in order to reveal the possible trigger of the process. METHODS: Three cusps of the same type of bovine pericardium valve prosthesis underwent comparative examination. Two cusps (cusps 1 and 2) were retrieved from a valve prosthesis explanted three months post-implantation, and the third cusp was from a non-implanted valve prosthesis and used as a reference cusp (ref. cusp). The examination included macroscopic examination, Non-linear Optical Microscopy using a multiphoton microscope, and histological examination with staining, using Hematoxylin and Eosin, Movat Pentachrome stain, Von-Kossa stain, and Alizirin-Red stain. Parallel sections were decalcified using Osteosoft® solution prior to Von-Kossa and Alizirin-Red staining to exclude false positive results. RESULTS: Macroscopically, cusp 1 showed early deterioration, and cusp 2 showed endocarditic vegetations. Histologically, cusp 1 showed calcifications in acellular deposits on the surface of the cusp, with pathological signs of subacute/healed endocarditis and intact cusp tissue. The examination did not show calcifications of the cellular remnants within the valve tissue. Cusp 2 showed florid endocarditis, with microscopic destruction of the valve tissue. CONCLUSION: Early prosthetic valve deterioration can exist as early as three months post-implantation. Subacute or subclinical endocarditis can be the cause for early valve calcification and deterioration.

A randomized, controlled trial of Veriset™ hemostatic patch in halting cardiovascular bleeding.

BACKGROUND: Obtaining hemostasis during cardiovascular procedures can be a challenge, particularly around areas with a complex geometry or that are difficult to access. While several topical hemostats are currently on the market, most have caveats that limit their use in certain clinical scenarios such as pulsatile arterial bleeding. The aim of this study was to assess the effectiveness and safety of Veriset™ hemostatic patch in treating cardiovascular bleeding. METHODS: Patients (N=90) scheduled for cardiac or vascular surgery at 12 European institutions were randomized 1:1 to treatment with either Veriset™ hemostatic patch (investigational device) or TachoSil® (control). After application of the hemostat, according to manufacturer instructions for use, time to hemostasis was monitored. Follow-up occurred up to 90 days post-surgery. RESULTS: Median time to hemostasis was 1.5 min with Veriset™ hemostatic patch, compared to 3.0 min with TachoSil® (p<0.0001). Serious adverse events within 30 days post-surgery were experienced by 12/44 (27.3%) patients treated with Veriset™ hemostatic patch and 10/45 (22.2%) in the TachoSil® group (p=0.6295). None of these adverse events were device-related, and no reoperations for bleeding were required within 5 days post-surgery in either treatment group. CONCLUSION: This study reinforces the difference in minimum recommended application time between Veriset™ hemostatic patch and TachoSil® (30 s versus 3 min respectively). When compared directly at 3 min, Veriset™ displayed no significant difference, showing similar hemostasis and safety profiles on the cardiovascular bleeding sites included in this study.

Movat Pentachrom stain reveals unexpected high osteogenesis rate in aortic valves.

BACKGROUND AND AIM OF THE STUDY: Aortic valve (AV) stenosis is the most common valvular heart disease with an incidence of 3% for people ≥ 65years in the industrialized world with indication for a surgical or transcatheter valve replacement. Researchers suppose osteogenic processes as key mechanisms in calcific aortic valve stenosis. Recently, Torre et al. published impressive histological analyses and detected osseous and/or chondromatous metaplasia in 15.6% of 6685 native calcified aortic valves. Therefore one HE section per valve originated from the area with the greatest extent of calcification was analyzed. Aim of our experimental setup was to identify regions of neo-osteogenesis and to determine the rate of specimens with active mineralization in human aortic valve tissue by Movat Pentachrom staining of sections of lager tissue segments. METHODS: Operational replaced aortic valves of 35 patients, 15 female and 20 male with an average age of 66.2 years were formalin fixed and decalcified using Osteosoft®-solution. Tissue samples were cut and 2µm specimens were stained with Movat Pentachrom to visualize osteogenic regions. Instead of screening a large number of sections, tissue samples were cut up to five times with at least 100µm space each if no region of osseous and/or chondromatous metaplasia was visible. RESULTS/CONCLUSIONS: Using this setup, a region of osseous metaplasia was detected in 25 (71.4%) of 35 samples analyzed. In some cases, these regions were small sized and only visible due to the bright color of Movat Pentachrom stain. This leads to the suggestion that a higher rate of calcified aortic valve samples would be classified as cusps with areas of neo-osteogenesis after staining with Movat Pentachrom stain and by the systematic analysis of larger parts of the tissue blocks.

Reducing Mediastinitis after Sternotomy with Combined Chlorhexidine-Isopropyl Alcohol Skin Disinfection: Analysis of 3,000 Patients.

BACKGROUND: Reducing surgical site infections (SSI) following median sternotomy remains a challenge for cardiac surgeons. Standard prophylaxis of SSI at our institution includes pre-operative skin disinfection with isopropyl alcohol (IPA). The addition of chlorhexidine gluconate (CHG) has the theoretical advantage of longer antimicrobial activity (>48h), compared with 2 h for IPA alone. OBJECTIVES: This prospective registry study was conducted to evaluate the effect of combined CHG-IPA (ChloraPrep®) skin antiseptic on the incidence of sternal surgical incision infections after cardiac surgical procedures via median sternotomy. METHODS: Between September 2011 and November 2013, 3,942 consecutive patients underwent cardiac surgery with median sternotomy at our institution. Among them, 2,985 patients met inclusion criteria and were enrolled in the study. The complete cohort was prospectively divided into two registries. In registry 1 (1,523 patients), CHG-IPA was used to disinfect skin at the thoracic operative site prior to incision. In registry 2 (1,462 patients), single IPA disinfection was used. The primary endpoint was the incidence of post-sternotomy mediastinitis within 30 d of surgery. Secondary endpoints were SSI of any other kind, 30-d survival, and hospital length of stay. RESULTS: Both registries were well matched in baseline characteristics and main risk factors. Post-operative data analysis revealed reduction in the rate of post-sternotomy mediastinitis in registry 1 (29 patients, 1.9%) versus registry 2 (62 patients, 4.2%), p = 0.0002. No relevant difference in incidence of other surgical site infections, length of hospital stay, and 30-d mortality was found. CONCLUSIONS: Skin disinfection with combined chlorhexidine-isopropyl alcohol reduced the incidence of mediastinitis in elective adult cardiac surgery with median sternotomy but did not affect other types of surgical site infections.

Heart valve stenosis in laser spotlights: insights into a complex disease.

Degenerative heart valve disease is a life-threatening disease affecting about 3% of the population over 65 years. Up to date, cardiac surgery with heart valve replacement is the only available therapy. The disease is characterized by degenerative disorganization of the heart valve structure and alterations in the residing cell populations. Causes and mechanisms of disease genesis are still not fully understood and until now pharmacological therapies are not available. Thus there is enormous interest in new technologies that enable a better characterization of structure and composition of diseased valves. Currently most research techniques demand for extensive processing of extracted valve material. We present a novel approach combining coherent anti-Stokes Raman scattering, endogenous two-photon excited fluorescence and second harmonic generation. Cusp constituents can be examined simultaneously, three-dimensionally and without extensive manipulation of the sample enabling impressive insights into a complex disease.

A rare cause of recurrent cerebral emboli despite oral anticoagulation.

We report on a patient with a history of colon carcinoma and clinical presentation of recurrent cardiac emboli despite oral anticoagulation for atrial fibrillation. On delayed transoesophageal echocardiography, finally a left atrial myxoma was suspected. Surgery, however, revealed a left atrial metastatic tumour with histopathological features of a colon adenocarcinoma. Metastases of colorectal adenocarcinoma invading cardiac structures are rare. Isolated literature reports describe metastatic masses detected in the right atrium reflecting natural haematogenous spreading of cancer, but none in the left heart.

Reversal of myofibroblastic activation by polyunsaturated fatty acids in valvular interstitial cells from aortic valves. Role of RhoA/G-actin/MRTF signalling.

Valvular interstitial cells (VICs), the fibroblast-like cellular constituents of aortic heart valves, maintain structural integrity of valve tissue. Activation into contractile myofibroblasts occurs under pathological situations and under standard cell culture conditions of isolated VICs. Reversal of this phenotype switch would be of major importance in respect to fibrotic valve diseases. In this investigation, we found that exogenous polyunsaturated fatty acids (PUFAs) decreased contractility and expression of myofibroblastic markers like α-smooth muscle actin (αSMA) in cultured VICs from porcine aortic valves. The most active PUFAs, docosahexaenoic acid (DHA) and arachidonic acid (AA) reduced the level of active RhoA and increased the G/F-actin ratio. The G-actin-regulated nuclear translocation of myocardin-related transcription factors (MRTFs), co-activators of serum response factor, was also reduced by DHA and AA. The same effects were observed after blocking RhoA directly with C3 transferase. In addition, increased contractility after induction of actin polymerisation with jasplakinolide and concomitant expression of αSMA were ameliorated by active PUFAs. Furthermore, reduced αSMA expression under PUFA exposure was observed in valve tissue explants demonstrating physiological relevance. In conclusion, RhoA/G-actin/MRTF signalling is operative in VICs, and this pathway can be partially blocked by certain PUFAs whereby the activation into the myofibroblastic phenotype is reversed.

Omega-3 polyunsaturated Fatty acids reduce the incidence of postoperative atrial fibrillation in patients with history of prior myocardial infarction undergoing isolated coronary artery bypass grafting.

INTRODUCTION: The impact of omega-3 polyunsaturated fatty acids (PUFAs) for prevention of atrial fibrillation (AF) is still part of a lively debate. The present study evaluates the impact of orally administered omega-3 ethyl ester concentrate (omega-3 PUFA) on postoperative onset of AF in patients with recent myocardial infarction (≤ 3 months) undergoing isolated coronary artery bypass grafting (CABG). Patients and METHODS: The study included a total of 198 patients with recent (≤ 3 months) myocardial infarction. The treatment group consisted of 99 prospectively and randomly assigned patients. A matched control group was generated out of the entirety of patients undergoing isolated CABG during the same time period, being not treated with omega-3 PUFA. Primary endpoint was onset of postoperative AF. Patients of the treatment group received a daily dose of 2 g omega-3 PUFA, initiated 5 days before surgery. Effective serum levels were confirmed by laboratory testing. RESULTS: Patients of the treatment group had less frequently postoperative AF (treatment: 31.3% vs. control: 48.0%; p = 0.017). The reduction in relative risk was 34.8% in the treatment group, which conforms a number needed to treat (NNT) of 6.0 patients. A more pronounced effect with a NNT of 4.1 was observed in patients ≤ 70 years (p = 0.007). Besides, patients of the treatment group had a shorter intensive care unit stay (p = 0.001) and suffered less frequently from impaired wound healing by trend (p = 0.063). One patient out of treatment group and two out of control group died during hospital stay (p = 1.000). CONCLUSION: Preoperative administration of 2 g omega-3 PUFA reduces incidence of postoperative AF in patients with recent (≤ 3 months) myocardial infarction undergoing isolated CABG.

Ex vivo 4D visualization of aortic valve dynamics in a murine model with optical coherence tomography.

The heart and its mechanical components, especially the heart valves and leaflets, are under enormous strain and undergo fatigue, which impinge upon cardiac output. The knowledge about changes of the dynamic behavior and the possibility of early stage diagnosis could lead to the development of new treatment strategies. Animal models are suited for the development and evaluation of new experimental approaches and therefor innovative imaging techniques are necessary. In this study, we present the time resolved visualization of healthy and calcified aortic valves in an ex vivo artificially stimulated heart model with 4D optical coherence tomography and high-speed video microscopy.

Heparinized blood better preserves cellular energy charge and vascular functions of intraoperatively stored saphenous vein grafts in comparison to isotonic sodium-chloride-solution.

OBJECTIVE: Several studies have addressed the optimal storage conditions for vascular grafts during surgery. The results remain contradictionary. This may be attributed to the fact, that the various vascular beds have a different sensitivity to storage. We analyzed the impact of storage in isotonic saline solution (NaCl) or heparinized blood the vascular functions of human saphenous vein grafts. Special care was taken to choose storage conditions which are relevant for intraoperative storage of a saphenous vein graft in a setting of coronary artery bypass grafting with vein and internal mammary artery as grafts. METHODS: Intraoperatively isolated V. saphena-segments (n = 36) were stored in NaCl or heparinized blood for approximately 30 minutes at room temperature. Subsequently, the segments were examined in a Mulvany-myograph. Following preconstriction with norepinephrine, concentration-relaxation curves were assessed for bradykinin and sodium-nitroprusside to assess developed vessel-wall tension as well as endothelium- and smooth-muscle-cell dependent vasorelaxation. The availability of adenosintriphosphate (energy charge) was determined based on liquid chromatography measurements of nucleotide tissue levels. RESULTS: Mean storage time was 27.4 ± 2.4 min in NaCl- and 26.3 ± 2.7 min in blood-group, respectively. After this period, receptor-dependent and-independent maximum of developed vessel wall tension was significantly reduced in NaCl-group (p = 0.05 and p = 0.045, respectively). Furthermore, the energy charge was significantly (p = 0.046) better preserved after blood storage (74 ± 1%) in comparison to NaCl-group (68 ± 2%). Endothelium-induced vasodilatation in response to bradykinin reached only 12.3 ± 2.5% in NaCl-group, but 19.3 ± 5.2% in blood-group (p = 0.033). Alike, EC50-concentration of bradykinin for half-maximal relaxation was significantly lower in blood- than in NaCl-group (log EC50 -7.08 ± 0.3 and -5.91 ± 0.4; respectively; p = 0.046). Endothelium-independent smooth muscle relaxation in response to sodium-nitroprusside was not different between both groups. CONCLUSION: Heparinized blood better preserves vascular contractile and endothelial functions of the saphenous vein graft. Storage in NaCl rapidly compromises vascular functions and impaires cellular energy. NaCl should no longer be recommended for intraoperative storage of harvested V. saphena grafts.

Sphingosine-1-phosphate induces contraction of valvular interstitial cells from porcine aortic valves.

AIMS: Sphingosine-1-phosphate (S1P) has emerged as a potent bioactive lipid with multiple functions in cardiovascular pathophysiology. Potential roles of S1P in heart valve diseases and expression of relevant receptors (S1P1, S1P2, or S1P3) in valve tissue and in valvular interstitial cells (VICs), the major cell population with essential functions in maintenance of valvular structure, are currently unknown. METHODS AND RESULTS: Exposure to S1P (62-2000 nM) of cultured VICs from porcine aortic valves on cell culture polystyrene resulted in contraction and nodule formation. The S1P-dependent contraction was completely inhibited by blockers of S1P2, RhoA, and RhoA-associated protein kinase (ROCK). Activated RhoA was clearly increased after S1P treatment, whereas activated Rac1 was only slightly reduced. In addition, exposure to S1P induced a transient increase in cytosolic Ca(2+). Application of channel blockers and other effectors of Ca(2+) homeostasis showed that the S1P effect is largely caused by Ca(2+) release from internal stores. However, resistance to blocking S1P2, different kinetics, as well as concentration dependence exclude a major role of Ca(2+) influx in S1P-induced nodule formation. In order to verify the effects in situ, contractions of valve tissue slices were measured. The S1P-induced isometric contraction of valve leaflets was of similar force amplitude as observed with adrenaline. The effect was fully reversed by blocking S1P2. CONCLUSION: The results suggest that S1P induces contraction of VICs from porcine aortic valves by signalling via S1P2, RhoA, and ROCK. In this way, S1P may contribute to regulation of tissue tension in aortic valves.

Inhibition of glycogen synthase kinase-3ß prevents activation of focal adhesion kinase after ischemia/reperfusion of the rat lung.

Recent studies on the mechanisms of ischemic preconditioning in myocardial tissue have presented convincing evidence that multiple protective pathways converge on inhibition of glycogen synthase kinase-3ß (GSK-3ß). To directly address the role of GSK-3ß in ischemia and reperfusion (I/R) of the lung, a rat model of left lung in situ ischemia was used. The specific non-competitive inhibitor of GSK-3ß, TDZD-8, was injected (3 mg/kg, vehicle in controls) 5 min before the left lung hilum was occluded for 60 min. Animals in the ischemia group underwent the same treatment, but without administration of TDZD-8. Lung functional and biochemical parameters were determined at time points 15 min and 60 min reperfusion. Treatment with TDZD-8 improved gas exchange (arterial pO2), but I/R-induced inflammation (plasma interleukin-6, leukocyte invasion) was not affected. The I/R cycle induced a rapid (15 min reperfusion) increase of protein tyrosine phosphorylation, including the activating phosphorylation of focal adhesion kinase at Tyr397, Tyr407, Tyr577, and Tyr861, and the non-receptor kinase Src at Tyr416. The phosphorylation was blocked by the GSK inhibitor. This effect may be related to the reduced plasma level of the strong effector of focal adhesion kinase, transforming growth factor-ß1, in the TDZD group. The underlying mechanisms are elusive, but they deserve further investigation, especially in relation to the early increase of lung permeability in this rat model of I/R injury. In conclusion, the results suggest that inhibition of GSK-3ß improves rat lung function during an I/R cycle, but only during the early reperfusion phase.

Low dose aprotinin and low dose tranexamic acid in elective cardiac surgery with cardiopulmonary bypass.

The antifibrinolytic agents aprotinin and tranexamic acid have both been proven to be efficient in reducing postoperative blood loss and transfusion requirements in patients in cardiac surgery. In light of recent safety issues regarding aprotinin, this single-centre study compared efficacy and safety of low dose aprotinin (2 million KIU, pump-prime volume only) and low dose tranexamic acid (1 g, pump-prime volume) in 708 consecutive patients from two prospective registers undergoing elective cardiac procedures with cardiopulmonary bypass (CPB). Incidences of postoperative complications showed no significant differences between groups. Postoperative blood loss and transfusion requirements were significantly lower in aprotinin compared to tranexamic acid patients. Overall, both antifibrinolytic low dose regimens are safe components of perioperative patient management in elective cardiac surgery with CPB. Cardiac procedures requiring longer CPB times might benefit from the administration of low dose aprotinin.

Cell-cell junctions and vascular endothelial growth factor in rat lung as affected by ischemia/reperfusion and preconditioning with inhaled nitric oxide.

BACKGROUND: Previous investigations have shown that short term inhalation of nitric oxide (NO) before ischemia and reperfusion (I/R) prevents I/R-related consequences on lung function. Here we correlate effects of NO-induced preconditioning, especially on the lung permeability barrier, with analysis of cell junction proteins and the level of vascular endothelial growth factor (VEGF). METHODS: A rat model of left lung in situ I/R was used. After left lateral thoracotomy, left lung ischemia was maintained for 60 min, followed by 30 min or 4 h (h) reperfusion (I/R groups). In the NO groups, inhalation of NO (10 min, 15 ppm) preceded I/R. Animals in control groups underwent sham surgery without NO inhalation and ischemia. The extent of I/R injury was assessed in terms of oxygenation (arterial PO(2)) and lung permeability (Evans blue extravasation). Expression of junctional proteins and phosphorylation was determined in complete protein extracts from lung tissue, whereas the adherens junction (AJ) core complex was analyzed in Triton extracts by co-immunoprecipitation using antibodies against E-cadherin and VE-cadherin. RESULTS: The inhalation of NO prevented the I/R-induced increase of permeability at 30 min reperfusion, and the PO(2) increased from 27% of controls in the I/R group to 77% in the NO group. Left lung I/R correlated with a progressive loss of cadherins (VE-cadherin, E-cadherin, desmoglein 1) during reperfusion, whereas AJ catenins were largely preserved. Preconditioning with NO resulted in an increased ratio of catenins (alpha- and beta-catenin) to E-cadherin in immunoprecipitates and in reduced phosphorylation of beta-catenin. A reduction of VEGF in left lung lavage fluid was observed at 4 h but not at 30 min reperfusion. CONCLUSIONS: The NO-induced changes of the AJ complex may have contributed to the stabilization of the lung permeability barrier during reperfusion.

Prevention of ischemia/reperfusion-induced accumulation of matrix metalloproteinases in rat lung by preconditioning with nitric oxide.

BACKGROUND: Pulmonary ischemia/reperfusion (I/R) injury is associated with degradation of structural proteins. Preconditioning by short-term inhalation of nitric oxide (NO) ameliorates some of the severe consequences of an I/R cycle. The aim of this study was to evaluate the effects of NO preconditioning on I/R-induced changes of matrix metalloproteinase (MMP) activity. MATERIALS AND METHODS: Left lung in situ ischemia in rats was maintained for 1 h, followed by reperfusion for 30 min or 4 h. In the NO group, animals inhaled NO (15 ppm) for 10 min directly before ischemia. Changes of expression or activity of MMPs (MMP-2, MMP-7, MMP-9, MMP-14) and of neutrophil elastase (NE) in bronchoalveolar lavage fluid (BALF), lung tissue, and arterial plasma were analyzed by zymography and Western blotting. Western blotting was also used to detect tissue inhibitors of matrix proteases, the extracellular metalloproteinase inducer (EMMPRIN or CD147), and endostatin, a proteolytic collagen fragment. RESULTS: Ischemia resulted in an increase of lavagable MMP activity (12.3-fold MMP-2, 8.1-fold MMP-7) at 30 min reperfusion. The activity of MMP-9 and NE in lung tissue progressively increased with time, whereas MMP-14 and MMP-2 were constant. Inhalation of NO prevented the early increase of MMP-2 and MMP-7 in BALF, but the level of MMP-9 and NE in tissue was not affected. The expression of tissue inhibitors of matrix proteases and EMMPRIN did not respond to any treatment. The release of endostatin proceeded in parallel to the level of MMPs in BALF. Significant correlations between MMP-9 and myeloperoxidase in lung tissue and between MMP-2/MMP-7 and plasma protein extravasation were found. CONCLUSIONS: The early rise of MMP-2 and MMP-7 in BALF resulted from plasma protein extravasation, whereas MMP-9 and NE were imported into lung tissue via leukocyte invasion. The effect of NO inhalation on lavagable MMPs was secondary to the sealing of the permeability barrier.

Preconditioning by inhaled nitric oxide prevents hyperoxic and ischemia/reperfusion injury in rat lungs.

Since the generation of nitric oxide (NO) is an essential step in the trigger phase of ischemic preconditioning, short-term inhalation of NO before ischemia should ameliorate ischemia/reperfusion (I/R) injury of the lung. We tested this hypothesis in high oxygen (>99%) ventilated rats in order to additionally evaluate compatibility of NO and exposure to hyperoxia. Male adult Sprague-Dawley rats inhaled NO (15 ppm, 10 min) before the left lung hilum was clamped for 1 h, and the reperfusion phase was observed for 4 h (NO group). Animals in the I/R group underwent the same treatment, but without NO inhalation. A third group without I/R served as time-matched controls. Animals in the I/R group showed severe I/R injury in terms of arterial pO2 (apO2), which was reduced to 22% of surgical controls (SCs) at time point 30 min reperfusion, and increased endothelial permeability (Evans blue procedure). The pretreatment with NO attenuated these effects. The pO2 after 4 h reperfusion was still 3.0-fold higher in the NO group compared to I/R. In contrast, the I/R- and hyperoxia-induced invasion of leukocytes, as determined by measuring myeloperoxidase (MPO) activity, was not affected by NO. These data were correlated with the activity of major cellular signaling pathways by measuring the phosphorylation at activating and inhibitory sites of extracellular-signal regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38, protein kinase B (AKT), and glycogen synthase kinase 3beta (GSK-3beta), and by determination of cGMP in plasma and lung tissue. Inhalation of NO partly prevented the loss of activation by I/R and hyperoxic ventilation of ERK, JNK, and AKT, and it reduced the I/R-induced activation of GSK-3beta. The level of cGMP in plasma and lung tissue was increased in the NO group after 4 h reperfusion. In conclusion, application of inhaled NO in the preconditioning mode prevented I/R injury in the rat lung without interfering effects of hyperoxic ventilation. The effects of NO on cellular signaling pathways resemble mechanisms of ischemic preconditioning, but further studies have to evaluate the physiological relevance of these results.