Elastomeric Cardiowrap Scaffolds Functionalized with Mesenchymal Stem Cells-Derived Exosomes Induce a Positive Modulation in the Inflammatory and Wound Healing Response of Mesenchymal Stem Cell and Macrophage.

A challenge in contractile restoration of myocardial scars is one of the principal aims in cardiovascular surgery. Recently, a new potent biological tool used within healing processes is represented by exosomes derived from mesenchymal stem cells (MSCs). These cells are the well-known extracellular nanovesicles released from cells to facilitate cell function and communication. In this work, a combination of elastomeric membranes and exosomes was obtained and tested as a bioimplant. Mesenchymal stem cells (MSCs) and macrophages were seeded into the scaffold (polycaprolactone) and filled with exosomes derived from MSCs. Cells were tested for proliferation with an MTT test, and for wound healing properties and macrophage polarization by gene expression. Moreover, morphological analyses of their ability to colonize the scaffolds surfaces have been further evaluated. Results confirm that exosomes were easily entrapped onto the surface of the elastomeric scaffolds, increasing the wound healing properties and collagen type I and vitronectin of the MSC, and improving the M2 phenotype of the macrophages, mainly thanks to the increase in miRNA124 and decrease in miRNA 125. We can conclude that the enrichment of elastomeric scaffolds functionalized with exosomes is as an effective strategy to improve myocardial regeneration.

Possible Link Between the ABO Blood Group of Bioprosthesis Recipients and Specific Types of Structural Degeneration.

Background Pigs/bovines share common antigens with humans: α-Gal, present in all pigs/bovines close to the human B-antigen; and AH-histo-blood-group antigen, identical to human AH-antigen and present only in some animals. We investigate the possible impact of patients' ABO blood group on bioprosthesis structural valve degeneration (SVD) through calcification/pannus/tears/perforations for patients ≤60 years at implantation. Methods and Results This was a single-center study (Paris, France) that included all degenerative bioprostheses explanted between 1985 and 1998, mostly porcine bioprostheses (Carpentier-Edwards second/third porcine bioprostheses) and some bovine bioprostheses. For the period 1998 to 2014, only porcine bioprostheses with longevity ≥13 years were included (total follow-up ≥29 years). Except for blood groups, important predictive factors for SVD were prospectively collected (age at implantation/longevity/number/site/sex/SVD types) and analyzed using logistic regression. All variables were available for 500 explanted porcine bioprostheses. By multivariate analyses, the A group was associated with an increased risk of: tears (odds ratio[OR], 1.61; P=0.026); pannus (OR, 1.5; P=0.054), pannus with tears (OR, 1.73; P=0.037), and tendency for lower risk of: calcifications (OR, 0.63; P=0.087) or isolated calcification (OR, 0.67; P=0.17). A-antigen was associated with lower risk of perforations (OR 0.56; P=0.087). B-group patients had an increased risk of: perforations (OR, 1.73; P=0.043); having a pannus that was calcified (OR, 3.0, P=0.025). B-antigen was associated with a propensity for calcifications in general (OR, 1.34; P=0.25). Conclusions Patient's ABO blood group is associated with specific SVD types. We hypothesize that carbohydrate antigens, which may or may not be common to patient and animal bioprosthetic tissue, will determine a patient's specific immunoreactivity with respect to xenograft tissue and thus bioprosthesis outcome in terms of SVD.

Elastomeric cardiopatch scaffold for myocardial repair and ventricular support.

OBJECTIVES: Prevention of postischaemic ventricular dilatation progressing towards pathological remodelling is necessary to decrease ventricular wall deterioration. Myocardial tissue engineering may play a therapeutic role due to its capacity to replace the extracellular matrix, thereby creating niches for cell homing. In this experimental animal study, a biomimetic cardiopatch was created with elastomeric scaffolds and nanotechnologies. METHODS: In an experimental animal study in 18 sheep, a cardiopatch was created with adipose tissue-derived progenitor cells seeded into an engineered bioimplant consisting of 3-dimensional bioabsorbable polycaprolactone scaffolds filled with a peptide hydrogel (PuraMatrix™). This patch was then transplanted to cover infarcted myocardium. Non-absorbable poly(ethyl) acrylate polymer scaffolds were used as controls. RESULTS: Fifteen sheep were followed with ultrasound scans at 6 months, including echocardiography scans, tissue Doppler and spectral flow analysis and speckle-tracking imaging, which showed a reduction in longitudinal left ventricular deformation in the cardiopatch-treated group. Magnetic resonance imaging (late gadolinium enhancement) showed reduction of infarct size relative to left ventricular mass in the cardiopatch group versus the controls. Histopathological analysis at 6 months showed that the cardiopatch was fully anchored and integrated to the infarct area with minimal fibrosis interface, thereby promoting angiogenesis and migration of adipose tissue-derived progenitor cells to surrounding tissues. CONCLUSIONS: This study shows the feasibility and effectiveness of a cardiopatch grafted onto myocardial infarction scars in an experimental animal model. This treatment decreased fibrosis, limited infarct scar expansion and reduced postischaemic ventricular deformity. A capillary network developed between our scaffold and the heart. The elastomeric cardiopatch seems to have a positive impact on ventricular remodelling and performance in patients with heart failure.

Intrapulmonary shear stress enhancement: a new therapeutic approach in acute myocardial ischemia.

OBJECTIVE: Ischemic heart disease (IHD) is a leading cause of mortality with insufficient results of current therapies, most probably due to maintained endothelial dysfunction conditions. Alternatively, we propose a new treatment that promotes endothelial shear stress (ESS) enhancement using an intrapulmonary pulsatile catheter. METHODS: Twelve piglets, divided in equal groups of 6: pulsatile (P) and non-pulsatile (NP), underwent permanent left anterior descending coronary artery ligation through sternotomy. After 1 h of ischemia and heparin injection (150 IU/kg): in P group, a pulsatile catheter was introduced into the pulmonary trunk and pulsated intermittently over 1 h, and irrespective of heart rate (110 bpm). In NP group, nitrates were given (7 ± 2 mg/kg/min) for 1 h. RESULTS: In P group all 6 animals survived ischemia for 120 min, but in NP group only 2 animals survived. The 4 animals that died during the experiment in NP group survived for 93 ± 14 min. Hemodynamics and cardiac output (CO) were significantly improved in P group compared with NP group: CO was 0.92 ± 0.15 vs. 0.52 ± 0.08 in NP group (L/min; p < 0.05), respectively. Vascular resistances (dynes.s.cm(-5)/kg) were significantly (p < 0.05) lower in P group versus NP group: pulmonary resistance was 119 ± 13 vs. 400 ± 42 and systemic resistance was 319 ± 43 vs. 1857 ± 326, respectively. Myocardial apoptosis was significantly (p < 0.01) lower in P group (0.66 ± 0.07) vs. (4.18 ± 0.27) in NP group. Myocardial endothelial NO synthase mRNA expression was significantly (p < 0.01) greater in P group (0.90 ± 0.09) vs. (0.25 ± 0.04) in NP group. CONCLUSIONS: Intrapulmonary pulsatile catheter could improve hemodynamics and myocardial contractility in acute myocardial ischemia. This represents a cost-effective method, suitable for emergency setting as a first priority, regardless of classical coronary reperfusion.

Development of cardiac support bioprostheses for ventricular restoration and myocardial regeneration.

OBJECTIVES: Ventricular constraint devices made of polyester and nitinol have been used to treat heart failure patients. Long-term follow-up has not demonstrated significant benefits, probably due to the lack of effects on myocardial tissue and to the risk of diastolic dysfunction. The goal of this experimental study is to improve ventricular constraint therapy by associating stem cell intrainfarct implantation and a cell-seeded collagen scaffold as an interface between the constraint device and the epicardium. METHODS: In a sheep ischaemic model, three study groups were created: Group 1: coronary occlusion without treatment (control group). Group 2: postinfarct ventricular constraint using a polyester device (Acorn CorCap). Group 3: postinfarct treatment with stem cells associated with collagen matrix and the polyester device. Autologous adipose mesenchymal stem cells cultured in hypoxic conditions were injected into the infarct and seeded into the collagen matrix. RESULTS: At 3 months, echocardiography showed the limitation of left ventricular end-diastolic volume in animals both treated with constraint devices alone and associated with stem cells/collagen. In Group 3 (stem cell + collagen treatment), significant improvements were found in ejection fraction (EF) and diastolic function evaluated by Doppler-derived mitral deceleration time. In this group, histology showed a reduction of infarct size, with focuses of angiogenesis and minimal fibrosis interface between CorCap and the epicardium due to the interposition of the collagen matrix. CONCLUSIONS: Myocardial infarction treated with stem cells associated with a collagen matrix and ventricular constraint device improves systolic and diastolic function, reducing adverse remodelling and fibrosis. The application of bioactive molecules and the recent development of nanobiotechnologies should open the door for the creation of a new semi-degradable ventricular support bioprosthesis, capable of controlled stability or degradation in response to physiological conditions of the left or right heart.

Development of bioartificial myocardium by electrostimulation of 3D collagen scaffolds seeded with stem cells.

Electrostimulation (ES) can be defined as a safe physical method to induce stem cell differentiation. The aim of this study is to evaluate the effectiveness of ES on bone marrow mesenchymal stem cells (BMSCs) seeded in collagen scaffolds in terms of proliferation and differentiation into cardiomyocytes. BMSCs were isolated from Wistar rats and seeded into 3D collagen type 1 templates measuring 25 × 25 × 6 mm. Bipolar in vitro ES was performed during 21 days. Electrical impedance and cell proliferation were measured. Expression of cardiac markers was assessed by immunocytochemistry. Viscoelasticity of collagen matrix was evaluated. Electrical impedance assessments showed a low resistance of 234±41 Ohms which indicates good electrical conductivity of collagen matrix. Cell proliferation at 570 nm as significantly increased in ES groups after seven day (ES 0.129±0.03 vs non-stimulated control matrix 0.06±0.01, P=0.002) and after 21 days, (ES 0.22±0.04 vs control 0.13±0.01, P=0.01). Immunocytoche mistry of BMSCs after 21 days ES showed positive staining of cardiac markers, troponin I, connexin 43, sarcomeric alpha-actinin, slow myosin, fast myosin and desmin. Staining for BMSCs marker CD29 after 21 days was negative. Electrostimulation of cell-seeded collagen matrix changed stem cell morphology and biochemical characteristics, increasing the expression of cardiac markers. Thus, MSC-derived differentiated cells by electrostimulation grafted in biological scaffolds might result in a convenient tissue engineering source for myocardial diseases.

Intrapulmonary shear stress enhancement: a new therapeutic approach in pulmonary arterial hypertension.

OBJECTIVE: Pulmonary arterial hypertension (PAH) is a dysfunctional endothelium disease with increased pulmonary vascular resistance (PVR) and poor prognosis. Current therapies are still insufficient. Here we propose a new pulsatile device as a more effective tool for PAH management compared with traditional treatments. MATERIALS AND METHODS: Twelve piglets (10.3 ± 3.8 kg) were given either intrapulmonary pulsatile [P (n = 6)] or nonpulsatile [NP (n = 6)] tadalafil treatment. After median sternotomy and heparin injection (250 IU/kg), both groups underwent aorto-pulmonary surgical shunt for 1 h. During a second 1 h period in group P, a catheter prototype, driven by a small ventilator, was introduced into the pulmonary trunk and pulsated intermittently at 110 bpm irrespective of heart rate (90.6 ± 10.74 bpm). In group NP, tadalafil was given orally (1 mg/kg). RESULTS: Hemodynamics and cardiac output (CO) were significantly (p < 0.05) improved in group P compared with group NP: CO was 0.56 ± 0.0.26 versus 0.54 ± 0.11 (L/min), respectively. Mean pulmonary artery pressure (PAP) was decreased in group P compared with group NP: PAP was 9.6 ± 2.97 versus 32.2 ± 5.07, respectively. Vascular resistances (dynes.s.cm(-5)/kg) were significantly lower in group P versus group NP: pulmonary resistance was 85 ± 42.12 versus 478 ± 192.91 and systemic resistance was 298.8 ± 172.85 versus 1301 ± 615.79, respectively. Using Western blot analysis, endogenous NO synthase expression in PA segments was nonsignificantly (p > 0.05) greater in group P (0.81 ± 0.78) versus (0.62 ± 0.35) group NP. CONCLUSION: Induced with an appropriate device, intrapulmonary shear stress-mediated endothelial function enhancement provides a more effective nearly physiological therapy for PAH.

Comparison of the effects of adenosine, inosine, and their combination as an adjunct to reperfusion in the treatment of acute myocardial infarction.

Adenosine and inosine are both key intracellular energy substrates for nucleotide synthesis by salvage pathways, especially during ischemic stress conditions. Additionally they both possess cell protective and cell repair properties. The objective of this study is to detect potential advantages of the combination of adenosine and inosine versus each drug alone, in terms of ventricular function, infarct size reduction and angiogenesis. Myocardial ischemia was created in rodents and treated with adenosine, inosine or their combination. Results of experiments showed that the combination of both drugs significantly reduced infarct size and improved myocardial angiogenesis and ventricular function. The two compounds, while chemically similar, use different intracellular pathways, allowing for complementary biological activities without overlapping. The drug combination at specific 1 : 5 adenosine : inosine dose ratio demonstrated positive cardiologic effects, deserving further evaluation as an adjunct to reperfusion techniques during and after acute coronary syndrome. The association of adenosine and inosine may contribute to reduce myocardial infarction morbidity and mortality rates.

Cardiac xenotransplantation technology provides materials for improved bioprosthetic heart valves.

OBJECTIVES: Human subjects and Old World primates have high levels of antibody to galactose-α-1,3 galactose ß-1,4-N-acetylglucosamine (α-Gal). Commercially available bioprosthetic heart valves of porcine and bovine origin retain the Gal antigen despite current processing techniques. Gal-deficient pigs eliminate this xenoantigen. This study tests whether binding of human anti-Gal antibody effects calcification of wild-type and Gal-deficient glutaraldehyde-fixed porcine pericardium by using a standard subcutaneous implant model. METHODS: Expression of α-Gal was characterized by lectin Griffonia simplicifolia-IB4 staining. Glutaraldehyde-fixed pericardial disks from Gal-positive and Gal-deficient pigs were implanted into 12-day-old Wistar rats and 1.5-kg rabbits with and without prelabeling with affinity-purified human anti-Gal antibody. Calcification of the implants was determined after 3 weeks by using inductively coupled plasma spectroscopy. RESULTS: The α-Gal antigen was detected in wild-type but not Gal-deficient porcine pericardium. Wild-type disks prelabeled with human anti-Gal antibody exhibited significantly greater calcification compared with that seen in antibody-free wild-type samples (mean ± standard error of the mean: 111 ± 8.4 and 74 ± 9.6 mg/g, respectively; P = .01). In the presence of anti-Gal antibody, a significantly greater level of calcification was detected in wild-type compared with GTKO porcine pericardium (111 ± 8.4 and 55 ± 11.8 mg/g, respectively; P = .005). Calcification of Gal-deficient pericardium was not affected by the presence of anti-Gal antibody (51 ± 9.1 and 55 ± 11.8 mg/g). CONCLUSIONS: In this model anti-Gal antibody accelerates calcification of wild-type but not Gal-deficient glutaraldehyde-fixed pericardium. This study suggests that preformed anti-Gal antibody present in all patients might contribute to calcification of currently used bioprosthetic heart valves. Gal-deficient pigs might become the preferred source for new, potentially calcium-resistant bioprosthetic heart valves.

Genesis of myocardial repair with cardiac progenitor cells and tissue engineering.

BACKGROUND: There is mounting evidence to suggest that the heart has regenerative potential in the event of myocardial injury. Recent studies have shown that a resident population of cardiac progenitor cells (CPCs) in the heart contains both vasculogenic and myogenic lineages. CPCs are able to migrate to the site of injury in the heart for participation in the healing process. The resident CPCs in the heart may also be activated through outside pharmacological intervention to promote their participation in the intrinsic repair process. In the light of these characteristics, CPCs provide a logical source for the heart cell therapy. During the regenerative cardiac process, stem cell niches (a specialised environment surrounding stem cells) provide crucial support needed for their maintenance. DISCUSSION: Compromised niche function may lead to the selection of stem cells that no longer depend on self-renewal factors produced by its environment. The objective of stem cell transplantation associated with tissue-engineered approaches is to create a new modality in the treatment of heart failure. The use of efficient scaffolds will aid to re-establish a favourable microenvironment for stem cell survival, multiplication, differentiation and function. Cardiac tissue engineering using natural and/or synthetic materials in this regard provides a novel possibility in cardiovascular therapeutics.

Gal knockout pig pericardium: new source of material for heart valve bioprostheses.

BACKGROUND: Although glutaraldehyde fixation is known to reduce immunogenicity and degeneration of heart valve bioprostheses, some degree of immunogenicity persists, which may trigger calcification. The aims of this study were to: (1) define the role of alpha-1,3-galactosyltransferase (alpha-Gal) antigen in valve calcification by comparing alpha-Gal-positive and alpha-Gal-deficient (GT-KO) pig pericardium; and (2) elucidate the role of human anti-Gal antibodies in the process of calcification and to determine the potential influence of different tissue-fixation techniques. METHODS: Glutaraldehyde-treated pericardium from alpha-Gal-positive and GT-KO pigs, with or without pre-labeling with human anti-Gal antibodies, were implanted in rats during 1 month. RESULTS: In glutaraldehyde-fixed pericardium, calcification levels were significantly lower in GT-KO pig pericardium (132.8 +/- 5.8 microg/mg) as compared with alpha-Gal-positive pig pericardium (155.7 +/- 7.1 microg/mg) (p < 0.015). In glutaraldehyde-fixed pig pericardium followed by a mix of formaldehyde, ethanol and Tween 80 (FET), the calcification levels were lower in GT-KO pig pericardium (0.35 +/- 0.1 microg/mg) as compared with alpha-Gal-positive pig pericardium (4.6 +/- 4.2 microg/mg). In glutaraldehyde-fixed pig pericardium + FET pre-incubated with human anti-Gal antibodies, calcification levels were significantly greater in alpha-Gal-positive pig pericardium (43.8 +/- 8.5 microg/mg) as compared with GT-KO pig pericardium (5.7 +/- 2.9 microg/mg) (p < 0.0001). CONCLUSIONS: This study demonstrates the role of alpha-Gal antigen and human alpha-Gal antibodies in the calcification process of valvular bioprostheses. It is suggested that GT-KO pig pericardium could be beneficial as a new source of material for heart valve bioprostheses.

Carbon monoxide can prevent acute lung injury observed after ischemia reperfusion of the lower extremities.

BACKGROUND: Pulmonary expression of heme oxygenase has been observed in multiple studies. This expression has been found beneficial in decreasing the severity of acute lung injury (ALI) post ischemia-reperfusion (I/R). The aim of this study was to assess the role of exogenous administration of the end-products of heme oxygenase reaction, carbon monoxide, and bilirubin, in the severity of ALI. STUDY DESIGN: We compared five groups of rats (n = 7/group) including a sham group and four I/R of the lower extremities by clamping the abdominal aorta for 2 h followed by reperfusion for 2 h. The four I/R groups included a control group, one pretreated with bilirubin (50 micromol/kg IV), another with inhaled carbon monoxide (CO) (250 ppm), and the last pretreated with both. The severity of ALI has been evaluated by a histological assay grading neutrophilic infiltration, as well as a study of the microvascular permeability using the Evans blue. RESULTS: The administration of CO prevented pulmonary microvascular permeability alteration noted after I/R of the lower limbs (pulmonary content of Evans blue: 141 +/- 23 microg/g of tissue in the isolated I/R group versus 68 +/- 34 microg/g of tissue in CO group; P < 0.001). Histologically CO administration inhibited neutrophilic sequestration observed after I/R. On the other hand, treatment by bilirubin alone (50 micromol/kg IV) did not modify the extent of pulmonary injury. CONCLUSION: Exogenous administration of carbon monoxide by inhalation at low doses prevented ALI post-I/R in this model.

Pulmonary expression of inducible heme-oxygenase after ischemia/reperfusion of the lower extremities in rats.

BACKGROUND: Expression of inducible heme-oxygenase (HO-1) has been shown to be increased in various inflammatory disorders, which may confer a protective role. The aim of our study was to assess pulmonary expression of HO-1 after ischemia/reperfusion (I/R) of the lower limbs in rats. MATERIALS AND METHODS: We compared three groups of rats (n = 5/group): one Sham group, and two I/R groups (aorta cross-clamped for 2 h followed by 2 h of reperfusion), one of which pre-treated with Zn-protoporphyrin (Zn-PP), a competitive inhibitor of HO (50 micromol/kg, i.p.). At the end of experiment, lungs were harvested for determination of HO activity and HO-1 expression by Western blot and immunohistochemistry. Lung injury was assessed by bronchoalveolar lavage, histological study, and determination of the lung Evans Blue dye content, an index of microvascular permeability. RESULTS: I/R of the lower limbs was responsible for acute lung injury (ALI), characterized by neutrophilic infiltration (87 +/- 20 x 10(3) neutrophils/mm(3), Sham group versus 191 +/- 38 x 10(3) neutrophils/mm(3), I/R group; P < 0.002) and an increase in lung Evans blue dye content: (74 +/- 6 microg/g, Sham group versus 122 +/- 48 microg/g, I/R group; P < 0.05). Pre-treatment with Zn-PP further increases the Evans Blue content (122 +/- 48 microg/g, I/R group versus 179 +/- 23 microg/g Zn-PP group P < 0.05) and the neutrophilic infiltration. Pulmonary heme-oxygenase activity, and HO-1 content were increased after I/R. (10.5 +/- 12 pmol bilirubin/mg protein/h, Sham group versus 101.2 +/- 66 pmol bilirubin/mg protein/h, I/R group; P < 0.02). Immunohistochemistry revealed that the expression of HO-1 was mainly localized to inflammatory cells. CONCLUSIONS: ALI following I/R of the lower limbs in rats is associated with an increase of pulmonary expression of HO-1, inhibition of this expression increase the severity of ALI.

Angiogenic growth factors and/or cellular therapy for myocardial regeneration: a comparative study.

BACKGROUND: Locally delivered angiogenic growth factors and cell implantation have been proposed for patients with myocardial infarcts without a possibility of percutaneous or surgical revascularization. The goal of this study was to compare the effects of these techniques in an experimental model of myocardial infarct. METHODS: Left ventricular myocardial infarction was created in 27 sheep by ligation of 2 coronary arteries. Three weeks after creation of the infarct, animals were randomized into 4 groups. In group 1, sheep received a culture medium injection to the infarct area (control group); group 2 underwent autologous myoblast implantation; group 3 received vascular endothelial growth factor; and group 4 received injection of both vascular endothelial growth factor and myoblasts. Evaluation included serum troponin IC levels, echocardiography (2-dimensional and color kinesis), and immunohistologic studies for quantitative analysis of capillaries (3 months after surgery). RESULTS: Four animals died of refractory ventricular fibrillation during myocardial infarction; 2 died after surgery because of stroke and 2 because of infections. Serum troponin increased to 45.6 +/- 4.7 ng/mL at postinfarction day 2. Echocardiography at 3 months showed a significant limitation of left ventricular dilation in the cell group (57 +/- 11.1 mL) and in the cell plus vascular endothelial growth factor group (58.6 +/- 6.6 mL: control group, 74.4 +/- 11.2 mL; vascular endothelial growth factor group, 68.1 +/- 3.4 mL). Color kinesis echography showed important improvements of regional fractional area change in the cell group (from 13.6% +/- 0.8% to 21.1% +/- 1.5%) and in the cell plus vascular endothelial growth factor group (from 12.8% +/- 0.9% to 18.7% +/- 2.3%). The number of capillaries increased in the peri-infarct region of the vascular endothelial growth factor group (1036 +/- 75: control group, 785 +/- 31; cell group, 830 +/- 75; cell plus vascular endothelial growth factor group, 831 +/- 83). CONCLUSIONS: In the cell therapy groups, regional ventricular contractility improved and heart dilatation was limited compared with either vascular endothelial growth factor or control; thus, postischemic remodeling was reduced. Angiogenesis was demonstrated in the vascular endothelial growth factor group, without improvement of ventricular function and remodeling. To improve local conditions for cell survival, further studies are warranted on prevascularization of myocardial scars with angiogenic therapy.

Exhaled carbon monoxide and inducible heme oxygenase expression in a rat model of postperfusion acute lung injury.

OBJECTIVE: Expression of inducible heme oxygenase has been shown to be increased in various visceral inflammatory disorders, which may confer a protective role. The purpose of our study was to determine whether the expression of inducible heme oxygenase was up-regulated within lungs in a rat model of extracorporeal circulation. METHODS: Wistar rats underwent either a partial femoro-femoral extracorporeal circulation in normothermia for 3 hours (n = 5) or a sham procedure (n = 5). Exhaled carbon monoxide concentration was monitored with an infrared analyzer. After the rats were killed, lungs were harvested for determination of heme oxygenase activity and inducible heme oxygenase expression (by Western blot and immunohistochemistry). Lung injury was also assessed by arterial blood gas analysis and microscopic study. RESULTS: Extracorporeal circulation was responsible for a lung injury characterized by decreased arterial blood oxygen saturation and typical morphologic findings (marked alveolar neutrophil infiltration; interstitial edema). Exhaled carbon monoxide concentration remained stable throughout the experiment in all sham rats, whereas it increased after extracorporeal circulation (from 0.16 +/- 0.05 ppm at baseline to 0.7 +/- 0.2 ppm at end of experiment; P =.0001). Pulmonary heme oxygenase activity and inducible heme oxygenase content (assessed by Western blot) were increased within lungs of rats that underwent extracorporeal circulation. Immunohistochemistry revealed that the expression of inducible heme oxygenase was mainly localized to inflammatory cells. CONCLUSIONS: Post-extracorporeal circulation acute lung injury in rats was associated with an increased expression of inducible heme oxygenase, the functional significance of which remains to be determined.

Human leukocyte antigen-G expression after heart transplantation is associated with a reduced incidence of rejection.

BACKGROUND: Human leukocyte antigen (HLA)-G, a nonclassic major histocompatibility complex class I molecule expressed in the extravillous cytotrophoblast at the feto-maternal interface, is known to protect the fetus from maternal cellular immunity. In a preliminary study, we showed that HLA-G is expressed in the hearts of some patients after heart transplantation. METHODS AND RESULTS: In the present study, a larger number of patients was investigated to confirm this finding and to look for possible correlations between HLA-G expression and the number and types of rejection. Expression of HLA-G in endomyocardial biopsy specimens was investigated by immunohistochemical analysis, and detection of the soluble HLA-G in the serum was performed by immunoprecipitation followed by Western blot analysis. HLA-G was detected in the biopsy specimens and serum of 9 of 51 patients (18%). The number of episodes of acute rejection was significantly lower in HLA-G-positive patients (1.2+/-1.1) as compared with HLA-G-negative patients (4.5+/-2.8) (P<0.001). No chronic rejection was observed in HLA-G-positive patients, whereas 15 HLA-G-negative patients had chronic rejection (P<0.032). A longitudinal study of these patients reveals that the status of HLA-G expression was maintained after 6 months both in serum and in biopsy specimens. During this period, HLA-G-positive patients did not have chronic rejection. CONCLUSIONS: There is a significant correlation between rejection and HLA-G expression in the heart after transplantation. HLA-G expression and its effect in reducing the incidence and severity of rejection seem to be stable throughout the evolution.