Influence of gender on ischemia-reperfusion injury in lungs in an animal model.

As with other organ transplants even lung transplantation raises the question of the possibility of the influence of gender on ischemia-reperfusion injury. This is a current topic especially for increasingly utilized method of lung transplantation from non-heart-beating donors, where reperfusion preceded by a period of warm and cold ischemia with subsequent treatment options for lung graft reperfusion. For measurements we used our laboratory previously created and validated animal model for ex vivo lung transplantation. As with other organ systems of our monitoring resulted protective effect of female sex on ischemia reperfusion lung injury. In two of the three parameters that were monitored, we found a significant difference. In females, higher oxygen transfer ability after reperfusion was manifested as well as lower perfusion pressure (vascular compliance). Conversely, weight gain (the development of pulmonary edema) in males was not significant difference from the females. These conclusions could cause further studies leading to influence the selection of appropriate donor grafts.

Depletion of alveolar macrophages attenuates hypoxic pulmonary hypertension but not hypoxia-induced increase in serum concentration of MCP-1.

Exposure to hypoxia, leading to hypoxic pulmonary hypertension (HPH), is associated with activation of alveolar macrophages (AM). However, it remains unclear how AM participate in this process. There are studies which imply that the AM product monocyte chemoattractant protein-1 (MCP-1) plays an important role. Thus we tested: 1. if the selective elimination of AM attenuates HPH in rats, 2. the correlation of MCP-1 plasmatic concentrations with the presence and absence of AM during exposure to hypoxia, 3. the direct influence of hypoxia on MCP-1 production in isolated AM. We found that experimental depletion of AM attenuated the chronic hypoxia-induced increase in mean pulmonary arterial pressure, but did not affect the serum MCP-1 concentrations. Furthermore, the MCP-1 production by AM in vitro was unaffected by hypoxia. Thus we conclude that AM play a significant role in the mechanism of HPH, but MCP-1 release from these cells is most likely not involved in this process. The increase of MCP-1 accompanying the development of HPH probably originates from other sources than AM.

ROS scavenger decreases basal perfusion pressure, vasoconstriction and NO synthase activity in pulmonary circulation during pulmonary microembolism.

Two mechanisms contribute in the development of pulmonary hypertension in pulmonary embolism (PE) - obstruction of pulmonary blood vessels and vasoconstriction. We hypothesize that hypoxia, increased shear stress and/or activation of gathered leukocytes in the PE may cause a release of reactive oxygen species (ROS). Therefore our aim was to determine the influence of the ROS scavenger Tempol on pulmonary hypertension and to describe NO synthase activity and production of NO oxidative products (NOx) after PE. In general anesthesia sephadex microspheres suspended in PSS were applied in right jugular vein as the pulmonary microembolism. Than we measured in isolated salt solution-perfused lungs the changes in perfusion pressure, activity of NO synthase and NOx plasma concentration in 7 groups of rats: C: control group (n=5), CN: C + sodium nitroprusside (SN) (n=5), EN: PE + SN (n=5), ETN: Tempol + PE + SN (n=5), CL: C + L-NAME (n=5), EL: PE + L-NAME (n=5), ETL: Tempol + PE + L-NAME (n=5). Tempol was applied intraperitoneally before PE. Animals that received Tempol (groups TN, TL) had significantly lower basal perfusion pressure than those which did not receive Tempol (EN, EL). Overall we measured a higher decrease of perfusion pressure than in the control group (C) after application of SN. Administration of L-NAME after PE (EL) increased the pressure more than in the control group (NL). NOx concentration was higher after PE. We found that preventive administration of Tempol decreases the increase in perfusion pressure after PE. PE increased NO release and concentration of NOx.

Combined suppression of the intrarenal and circulating vasoconstrictor renin-ACE-ANG II axis and augmentation of the vasodilator ACE2-ANG 1-7-Mas axis attenuates the systemic hypertension in Ren-2 transgenic rats exposed to chronic hypoxia.

The aim of the present study was to test the hypothesis that chronic hypoxia would aggravate hypertension in Ren-2 transgenic rats (TGR), a well-defined monogenetic model of hypertension with increased activity of endogenous renin-angiotensin system (RAS). Systolic blood pressure (SBP) in conscious rats and mean arterial pressure (MAP) in anesthetized TGR and normotensive Hannover Sprague-Dawley (HanSD) rats were determined under normoxia that was either continuous or interrupted by two weeks´ hypoxia. Expression, activities and concentrations of individual components of RAS were studied in plasma and kidney of TGR and HanSD rats under normoxic conditions and after exposure to chronic hypoxia. In HanSD rats two weeks´ exposure to chronic hypoxia did not alter SBP and MAP. Surprisingly, in TGR it decreased markedly SBP and MAP; this was associated with substantial reduction in plasma and kidney renin activities and also of angiotensin II (ANG II) levels, without altering angiotensin-converting enzyme (ACE) activities. Simultaneously, in TGR the exposure to hypoxia increased kidney ACE type 2 (ACE2) activity and angiotensin 1-7 (ANG 1-7) concentrations as compared with TGR under continuous normoxia. Based on these results, we propose that suppression of the hypertensiogenic ACE-ANG II axis in the circulation and kidney tissue, combined with augmentation of the intrarenal vasodilator ACE2-ANG 1-7 axis, is the main mechanism responsible for the blood pressure-lowering effects of chronic hypoxia in TGR.

Intrapulmonary activation of the angiotensin-converting enzyme type 2/angiotensin 1-7/G-protein-coupled Mas receptor axis attenuates pulmonary hypertension in Ren-2 transgenic rats exposed to chronic hypoxia.

The present study was performed to evaluate the role of intrapulmonary activity of the two axes of the renin-angiotensin system (RAS): vasoconstrictor angiotensin-converting enzyme (ACE)/angiotensin II (ANG II)/ANG II type 1 receptor (AT1) axis, and vasodilator ACE type 2 (ACE2)/angiotensin 1-7 (ANG 1-7)/Mas receptor axis, in the development of hypoxic pulmonary hypertension in Ren-2 transgenic rats (TGR). Transgene-negative Hannover Sprague-Dawley (HanSD) rats served as controls. Both TGR and HanSD rats responded to two weeks´ exposure to hypoxia with a significant increase in mean pulmonary arterial pressure (MPAP), however, the increase was much less pronounced in the former. The attenuation of hypoxic pulmonary hypertension in TGR as compared to HanSD rats was associated with inhibition of ACE gene expression and activity, inhibition of AT1receptor gene expression and suppression of ANG II levels in lung tissue. Simultaneously, there was an increase in lung ACE2 gene expression and activity and, in particular, ANG 1-7 concentrations and Mas receptor gene expression. We propose that a combination of suppression of ACE/ANG II/AT1receptor axis and activation of ACE2/ANG 1-7/Mas receptor axis of the RAS in the lung tissue is the main mechanism explaining attenuation of hypoxic pulmonary hypertension in TGR as compared with HanSD rats.

The contractile response of isolated small pulmonary arteries induced by activated macrophages.

To test whether macrophages can play any role in hypoxic pulmonary vasoconstriction, we tested the in vitro response of rings from small pulmonary arteries to the activation of macrophages by FMLP, a substance stimulating predominantly membrane-bound NADPH oxidase. A small vessel myograph was used to measure the responses of rings from small pulmonary arteries (300-400 microm) isolated from rat lungs. Rings from 5 rats were placed into both chambers of the myograph. The vessels were stabilized for 40 min and then normalized by automatic stretching to a wall tension equivalent to the intravascular pressure 30 mm Hg. At the start of each experiment, vessels were exposed to 80 mM K+ to obtain maximal contractile response, which was used to normalize subsequent contractile responses. 2x10(6) viable macrophages, obtained by peritoneal lavage, were added into one chamber, then 5 microM FMLP was administrated to both chambers and the tension measurement was started. The hydrogen peroxide concentration produced by stimulated macrophages was measured luminometrically. The concentrations of H2O2 in specimens from chambers containing activated macrophages rose from 3.5+/-1.5 nM to 110+/-28 nM within 25 min of stimulation, while FMLP itself didn't increase the H2O2 concentration from the baseline value (4.5+/-3 nM) in samples from control chambers. After FMLP administration, the tension of the vessel rings in the presence of macrophages reached 0.23+/-0.07 of maximal contractile response, it did not change in controls. The addition of ROS scavenger 4-hydroxy-TEMPO blocked the contractile response to the activation of macrophages. We conclude that the activation of macrophages stimulates the contraction of small pulmonary arteries and that this contraction is probably mediated by reactive oxygen species.

L-arginine in combination with sildenafil potentiates the attenuation of hypoxic pulmonary hypertension in rats.

Chronic hypoxia induces an increased production of nitric oxide (NO) in pulmonary prealveolar arterioles. Bioavailability of the NO in the pulmonary vessels correlates with concentration of L-arginine as well as activity of phosphodiesterase-5 enzyme (PDE-5). We tested a hypothesis whether a combination of L-arginine and PDE-5 inhibitor sildenafil has an additive effect in reduction of the hypoxic pulmonary hypertension (HPH) in rats. Animals were exposed to chronic normobaric hypoxia for 3 weeks. In the AH group, rats were administered L-arginine during chronic hypoxic exposure. In the SH group, rats were administered sildenafil during chronic hypoxic exposure. In the SAH group, rats were treated by the combination of L-arginine as well as sildenafil during exposure to chronic hypoxia. Mean PAP, structural remodeling of peripheral pulmonary arterioles (%DL) and RV/LV+S ratio was significantly decreased in the SAH group compared to hypoxic controls even decreased compared to the AH and the SH groups in first two measured parameters. Plasmatic concentration of cGMP and NOx were significantly lower in the SAH group compared to hypoxic controls. We demonstrate that NO synthase substrate L-arginine and phosphodiesterase-5 inhibitor sildenafil administered in combination are more potent in attenuation of the HPH compared to a treatment by substances given alone.

Hypercapnia attenuates the hypoxia-induced blunting of the reactivity in chronically hypoxic rats.

Chronic hypoxia causes oxidative injury of pulmonary vessels and attenuates their reactivity to different stimuli. When combined with hypercapnia, biochemical markers of this injury are reduced but the effect of concomitant hypoxia and hypercapnia on vascular reactivity is not fully understood. This study was therefore designed to test whether hypercapnia can prevent also the hypoxia-induced loss of reactivity of pulmonary vessels. The reactivity of vessels from rats exposed either to hypoxia or hypoxia combined with hypercapnia was tested using a small vessel myograph (M 500A, Linton, Norfolk, GB). The second and third intrapulmonary branches of pulmonary arteries were isolated under a dissecting microscope from lungs of 8 control rats (group N), 6 rats exposed to hypoxia for 5 days (isobaric, 10 % O(2), group H) and 7 rats exposed to hypoxia combined with hypercapnia for 5 days (10 % O(2), 5 % CO(2), group H+CO(2)). The transmural pressure was set by automatic normalization to 30 mm Hg. The vessel size did not vary among the groups. After stabilization we challenged the vessels twice with KCl (80 mM) and once with PGF(2alpha) (0.1 mM). There were no significant differences in KCl induced contractions among the groups. The responses to PGF(2alpha) were expressed as a ratio to the maximal tension obtained by the exposure to 80 mM KCl. Contractions induced by PGF(2alpha) were markedly reduced in group H (0.07+/-0.02) and in group H+CO(2) (0.26+/-0.03) in comparison with group N (0.83+/-0.07). The vessels of group H responded to PGF(2alpha) less than those of group H+CO(2). However we observed the attenuated reactivity also in group H+CO(2) in comparison with N. Hypercapnia therefore partially blunted the hypoxia-induced loss of reactivity in pulmonary arteries. This finding supports the hypothesis that hypercapnia significantly alters the nature of lung injury induced by chronic hypoxia.

Rho-kinase inhibition attenuates acute hypoxic fetoplacental vasoconstriction in the rat.

The vessels on the fetal side of the placenta differ from most other vascular beds except the lungs in that they respond to acute hypoxia by vasoconstriction. An essential role of calcium influx in the mechanism of this hypoxic fetoplacental vasoconstriction (HFPV) has been shown previously. That finding does not, however, exclude the possible involvement of other mechanisms of vascular tone regulation. In this study we tested the hypothesis that Rho-kinase-mediated calcium sensitization is involved in HFPV. We used a model of isolated rat placenta dually perfused (from both the maternal and fetal side) with Krebs salt solution saturated with normoxic and hypoxic gas mixture respectively at constant flow rate. Rho-kinase pathway was inhibited by fasudil (10 microM). We found that fasudil reduced basal normoxic fetoplacental vascular resistance and completely prevented HFPV. This suggests that the activity of Rho-kinase signaling pathway is essential for HFPV.

Short-term fasting reduces the extent of myocardial infarction and incidence of reperfusion arrhythmias in rats.

The effect of three-day fasting on cardiac ischemic tolerance was investigated in adult male Wistar rats. Anesthetized open-chest animals (pentobarbitone 60 mg/kg, i.p.) were subjected to 20-min left anterior descending coronary artery occlusion and 3-h reperfusion for infarct size determination. Ventricular arrhythmias were monitored during ischemia and at the beginning (3 min) of reperfusion. Myocardial concentrations of beta-hydroxybutyrate and acetoacetate were measured to assess mitochondrial redox state. Short-term fasting limited the infarct size (48.5+/-3.3 % of the area at risk) compared to controls (74.3+/-2.2 %) and reduced the total number of premature ventricular complexes (12.5+/-5.8) compared to controls (194.9+/-21.9) as well as the duration of ventricular tachycardia (0.6+/-0.4 s vs. 18.8+/-2.5 s) occurring at early reperfusion. Additionally, fasting increased the concentration of beta-hydroxybutyrate and beta-hydroxybutyrate/acetoacetate ratio (87.8+/-27.0) compared to controls (7.9+/-1.7), reflecting altered mitochondrial redox state. It is concluded that three-day fasting effectively protected rat hearts against major endpoints of acute I/R injury. Further studies are needed to find out whether these beneficial effects can be linked to altered mitochondrial redox state resulting from increased ketogenesis.

Nitrotyrosine and nitrate/nitrite levels in cardiac arrest survivors treated with endovascular hypothermia.

The protective effect of therapeutic hypothermia in cardiac arrest survivors (CAS) has been previously well documented. Animal studies have indicated that attenuation of tissue oxidative stress (OS) may be involved in the mechanisms that lead to the beneficial effect of hypothermia. The extent of OS and nitric oxide (NO) production in adult CAS treated with endovascular hypothermia is, however, unknown. A total of 11 adult patients who experienced cardiac arrest out of hospital were included in the present study, and all were treated with mild hypothermia using the Thermogard XP (Alsius, USA) endovascular system. A target core temperature of 33 °C was maintained for 24 hours, with a subsequent rewarming rate of 0.15 °C per hour, followed by normothermia at 36.8 °C. Blood samples for the measurement of nitrotyrosine and nitrate/nitrite levels were drawn at admission and every 6 hours thereafter for two days. During the hypothermic period, the levels of nitrotyrosine and nitrates/nitrites were comparable with baseline values. During the rewarming period, serum levels of both parameters gradually increased and, during the normothermic period, the levels were significantly higher compared with hypothermic levels (nitrotyrosine, P<0.001; nitrates/nitrites, P<0.05). In our study, significantly lower levels of nitrotyrosine and nitrates/nitrites were demonstrated during hypothermia compared with levels during the normothermic period in adult CAS. These data suggest that attenuation of OS and NO production may be involved in the protective effect of hypothermia in adult CAS.

Reactive oxygen species production in the early and later stage of chronic ventilatory hypoxia.

Pulmonary hypertension resulting from chronic hypoxia is at least partly caused by the increased production of reactive oxygen species (ROS). The goal of the presented study was to investigate the dynamics and the site of production of ROS during chronic hypoxia. In our study Wistar rats were kept for 1, 4 and 21 days in an isobaric hypoxic chamber (F(iO2)=0.1), while controls stayed in normoxia. We compared NO production in expired air, plasma and perfusate drained from isolated rat lungs and measured superoxide concentration in the perfusate. We also detected the presence of superoxide products (hydrogen peroxide and peroxynitrite) and the level of ROS-induced damage expressed as the concentration of lipid peroxydation end products. We found that the production and release of ROS and NO during early phase of chronic hypoxia has specific timing and differs in various compartments, suggesting the crucial role of ROS interaction for development of hypoxic pulmonary hypertension.

Lung mast cells and hypoxic pulmonary hypertension.

Hypoxic pulmonary hypertension (HPH) is a syndrome characterized by the increase of pulmonary vascular tone and the structural remodeling of peripheral pulmonary arteries. Mast cells have an important role in many inflammatory diseases and they are also involved in tissue remodeling. Tissue hypoxia is associated with mast cell activation and the release of proteolytic enzymes, angiogenic and growth factors which mediate tissue destruction and remodeling in a variety of physiological and pathological conditions. Here we focused on the role of mast cells in the pathogenesis of hypoxic pulmonary hypertension from the past to the present.

Production of proteolytic enzymes in mast cells, fibroblasts, vascular smooth muscle and endothelial cells cultivated under normoxic or hypoxic conditions.

Matrix metalloproteinases (MMPs) is a family of proteolytic enzymes involved in remodeling of extracellular matrix. Although proteolytic enzymes are produced by many cell types, mast cells seem to be more important than other types in remodeling of pulmonary arteries during hypoxia. Therefore, we tested in vitro production of MMPs and serine proteases in four cell types (mast cells, fibroblasts, vascular smooth muscle cells and endothelial cells) cultivated for 48 h under normoxic or hypoxic (3% O2) conditions. MMP-13 was visualized by immunohistochemistry, MMP-2 and MMP-9 were detected by zymography in cell lysates. Enzymatic activities (MMPs, tryptase and chymase) were estimated in the cultivation media. Hypoxia had a minimal effect on total MMP activity in the cultivation media of all types of cells, but immunofluorescence revealed higher intensity of MMP-13 in the cells exposed to hypoxia except of fibroblasts. Tryptase activity was three times higher and chymase activity twice higher in mast cells cultivated in hypoxia than in those cultured in normoxia. Among all cell types studied here, mast cells are the most abundant source of proteolytic enzymes under normoxic and hypoxic conditions. Moreover, in these cells hypoxia increases the production of both specific serine proteases tryptase and chymase, which can act as MMPs activators.

Parameters of healing in approximative intestinal anastomosis.

INTRODUCTION: The restoration of bowel continuity using multiple classic anastomoses is mostly impossible in unstable critically ill extremely low birth weight neonates. The parameters of healing of approximative anastomoses in which integrity and continuity of bowel is achieved with limited number of stitches were evaluated in an experimental study. MATERIAL AND METHODS: Small bowel anastomoses were performed in twenty-two adult male rats. An approximative ileo-ileal anastomosis was performed with five seromuscular-interrupted sutures only; in the control group the anastomosis was performed with the conventional technique of interrupted sutures. The mechanical and biochemical parameters were compared. RESULTS: All anastomoses in both groups healed well without obstruction. The mean operating time needed for an approximative anastomosis was shorter (16 +/- 7.1 min versus 23.6 +/- 6.2 min, p = 0.016). The strength of the approximative anastomoses on the 1st day after surgery was 55 +/- 15 torr; the strength of the conventional anastomoses was 55 +/- 42 torr. The strength of the approximative anastomoses after 7 days was 249 +/- 39 torr; the strength of the conventional anastomoses was 218 +/- 23 torr (p = 0.118). The activity of the collagenolytic enzymes matrix metalloproteinase-2 and matrix metalloproteinase-9 in the anastomotic area was significantly increased compared with the activity in samples of non-operated bowel. There was no significant difference in collagenolytic activity between both types of anastomoses. CONCLUSION: The approximative anastomosis is a time-saving alternative to conventional anastomoses with a comparable course of anastomotic healing, anastomotic strength, and changes in collagen metabolism.

Hypercapnia attenuates hypoxic pulmonary hypertension by inhibiting lung radical injury.

Chronic lung hypoxia results in hypoxic pulmonary hypertension. Concomitant chronic hypercapnia partly inhibits the effect of hypoxia on pulmonary vasculature. Adult male rats exposed to 3 weeks hypoxia (Fi(02)=0.1) combined with hypercapnia (Fi(C02)=0.04-0.05) had lower pulmonary arterial blood pressure, increased weight of the right heart ventricle, and less pronounced structural remodeling of the peripheral pulmonary arteries compared with rats exposed only to chronic hypoxia (Fi(02)=0.1). According to our hypothesis, hypoxic pulmonary hypertension is triggered by hypoxic injury to the walls of the peripheral pulmonary arteries. Hypercapnia inhibits release of both oxygen radicals and nitric oxide at the beginning of exposure to the hypoxic environment. The plasma concentration of nitrotyrosine, the marker of peroxynitrite activity, is lower in hypoxic rats exposed to hypercapnia than in those exposed to hypoxia alone. Hypercapnia blunts hypoxia-induced collagenolysis in the walls of prealveolar pulmonary arteries. We conclude that hypercapnia inhibits the development of hypoxic pulmonary hypertension by the inhibition of radical injury to the walls of peripheral pulmonary arteries.

Hyperoxia blunts acute hypoxia- and PGF2alpha-induced pulmonary vasoconstriction in chronically hypoxic rats.

We investigated the influence of oxygenation of in vitro lung preparation on the pulmonary vascular reactivity. Small pulmonary vessels isolated from adult male Wistar rats exposed for 4 days to hypoxia (F(iO2) = 0.1, group CH) were compared with those of normoxic controls (group N). The bath in the chamber of small vessel myograph was saturated with gas mixture containing either 21% or 95% of O(2) with 5% CO(2) and we measured the reactions of vessels to acute hypoxic challenge with 0% O(2) or to PGF(2alpha). We did not observe any difference of the contractile responses between both groups when the normoxic conditions were set in the bath. When the bath oxygenation was increased to 95% O(2), the contractions induced by hypoxic challenge and PGF(2alpha) decreased in chronically hypoxic rats and did not change in normoxic controls. We hypothesize that reduced reactivity of vessels from hypoxic rats in hyperoxia results from the effect of chronic hypoxia on Ca(2+) signaling in the vascular smooth muscle, which is modulated by increased free radical production during the exposure to chronic hypoxia and further hyperoxia.

In vitro hypoxia increases production of matrix metalloproteinases and tryptase in isolated rat lung mast cells.

Chronic hypoxia results in hypoxic pulmonary hypertension characterized by fibrotization and muscularization of the walls of peripheral pulmonary arteries. This vessel remodeling is accompanied by an increase in the amount of lung mast cells (LMC) and the presence of small collagen cleavage products in the vessel walls. We hypothesize that hypoxia activates LMC, which release matrix metalloproteinases (MMPs) cleaving collagen and starting increased turnover of connective tissue proteins. This study was designed to determine whether in vitro hypoxia stimulates production of MMPs in rat LMC and increases their collagenolytic activity. The LMC were separated on the Percoll gradient and then were divided into two groups and cultivated for 24 h in 21 % O(2) + 5 % CO(2) or in 10 % O(2) + 5 % CO(2). Presence of the rat interstitial tissue collagenase (MMP-13) in LMC was visualized by immunohistological staining and confirmed by Western blot analysis. Total MMPs activity and tryptase activity were measured in both cultivation media and cellular extracts. Exposure to hypoxia in vitro increased the amount of cells positively labeled by anti-MMP-13 antibody as well as activities of all measured enzymes. The results therefore support the concept that LMC are an important source of increased collagenolytic activity in chronic hypoxia.

[Method of approximative intestinal anastomosis in experimental model]. / Metoda aproximacní strevní anastomózy v experimentálním modelu.

AIM: A technique of approximative anastomosis in witch integrity and continuity of bowel is achieved with limited number of interrupted seromuscular stitches was evaluated in experimental study. MATERIAL AND METHODS: Small bowel anastomosis were performed in twelve rats (Wistar, male) with weight range 197-242 g. An approximative anastomosis in the ileum of six rats was performed with five seromuscular-interrupted sutures only; in the second study group anastomosis was performed with conventional technique of interrupted sutures. The approximative anastomosis was evaluated concerning operating time, anastomotic healing, bursting pressure and adhesions in comparison to the conventional anastomosis. Statistics was calculated with Anova test. RESULTS: All anastomosis in both group healed well without obstruction. The median operating time needed for approximative anastomosis was shorter (31.7 +/- 1.6 minutes versus 35.2 +/- 1.5 minutes, p = 0.002). The strength of approximative anastomosis after 7 days was 249 +/- 39 torr; strength of standard anastomosis was 218 +/- 23 torr (p = 0.118). There were no significant differences in the others evaluated parameters between two study groups. CONCLUSION: In the animal model presented, the approximative anastomosis shows time saving alternative to standard anastomosis, with the same parameters of anastomotic strength, healing, and adhesions.

Evidence of secondary neuronal intestinal dysplasia in a rat model of chronic intestinal obstruction.

The etiology of neuronal intestinal dysplasia remains largely unknown. There is, however, supporting evidence of the existence of Hirschprung's disease or chronic intestinal obstruction associated with neuronal intestinal dysplasia. With the aim of investigating the possible development of neuronal intestinal dysplasia linked to chronic intestinal obstruction, we have examined the enteric nervous system response to long-term obstruction in a rat model. Three different surgical techniques were tested in Wistar male rats. In animals that survived longer than the cutoff chronic intestinal obstruction point (6 weeks), full-thickness biopsies and acetylcholinesterase (AChE), NADH, hematoxylin-eosin, and anti-S100 protein stainings were performed. The results of our model indicate that chronic intestinal obstruction induced different degrees of enteric nervous system dysplasia, including histological features of neuronal intestinal dysplasia. The relationship between chronic intestinal obstruction and anomalies of the enteric nervous system, including neuronal intestinal dysplasia, needs to be further studied.