Moisture-cured silicone-urethanes-candidate materials for tissue engineering: a biocompatibility study in vitro.

This study was performed to verify the response of human bone-derived cells (HBDCs) to moisture-cured silicone-urethanes (mcSUUs) in vitro, as the first step toward using them as scaffolds for bone tissue engineering. Good surgical handling, tissue cavity filling, stable mechanical properties, and potentially improved oxygen supply to cells after implantation justify the investigation of these nondegradable elastomers. A set of various mcSUUs were obtained by moisture-curing NCO-terminated prepolymers, synthesized from oligomeric siloxane diols of two different oligosiloxane chain lengths, and two different diisocyanates (MDI and IPDI), using two different NCO/OH molar ratios. Dibutyltindilaurate (DBTL) or N-dimethylethanolamine (N-met) served as catalysts. After 7 days of culture, cell number, viability, and alkaline phosphatase (ALP) activity were determined, and after 21 days, cell viability and collagen production were determined. Material characteristics significantly influenced the cell response. The mcSUUs prepared with DBTL (widely used in the syntheses of biomaterials) were cytotoxic. The MDI-based mcSUUs were significantly more favored by HBDCs than the IPDI-based ones in all performed tests. MDI-based material with low 2/1 NCO/OH and short chain length was the best support for cells, comparable with tissue-culture polystyrene (with ALP activity even higher). HBDCs cultured on porous scaffolds from this mcSUU produced a tissue-like structure in culture. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res, 2010.

Cardiocyte nuclear chromatin density correlates with transplanted heart left ventricular mass.

INTRODUCTION: Cardiocyte hypertrophy is accompanied by polyploidy, seen as a decrease in chromatin density in the enlarged nucleus. Repeated biopsies of a transplanted heart offer the possibility of a dynamic evaluation of these phenomena. The aim of this work was an evaluation of cardiocyte nuclear chromatin density in transplanted hearts during long-term follow-up. MATERIALS AND METHODS: The material encompassed myocardial biopsy specimens taken during the first week, first month, and then on an annual basis up to 10 years after surgery. Only biopsy specimens with no rejection were considered (grade "0" International Society for Heart and Lung Transplantation [ISHLT] 122 biopsy specimens). The control group consisted of 7 donor heart specimens. We evaluated the optical density-mean gray level-of cardiomyocyte nuclear chromatin. We determined correlations of this index with the nuclear area, and with left ventricle ultrasound measurements, using correlation analysis. RESULTS: The chromatin mean gray level decreased with time, correlating positively with interventricular septum thickness, left ventricle posterior wall diameter, and left ventricular mass. Analysis of individual periods showed a significant positive correlation of the mean grey level with the cardiocyte nuclear surface in year 3, 4, and 9 after transplantation, thereby suggesting the occurrence of polyploidy at those times. The significant negative correlation of these values (1 week and 1 year) indicated normalization of early cardiocyte hypertrophy. CONCLUSIONS: With the passage of time chromatin condenses, leading to pyknosis. The activity of cardiocyte chromatin correlated with left ventricular hypertrophy. Compensatory cardiomyocyte polyploidy is a periodical phenomenon.

The effect of acute and prolonged endurance exercise on transforming growth factor-beta1 generation in rat skeletal and heart muscle.

The serum level of the transforming growth factor-beta1 (TGF-beta1) is elevated after acute bouts of exercise and prolonged training, as well as after myocardial infarction. However, the source of this increase remains unclear. Contracting skeletal muscles are known to be the source of many cytokines. To determine whether skeletal or heart muscles produce TGF-beta1 during exercise, we investigated the effect of a single bout of acute exercise on TGF-beta1 generation in skeletal and heart muscles in untrained rats (UT, n=30) and in rats subjected to prolonged (6-week) endurance training (T, n=29). The UT and T (a day after final training) groups were subjected to an acute bout of exercise with the same work load. Rats from both groups were sacrificed and skeletal and heart muscle samples were collected before (pre), immediately after (0 h), or 3 hours (3 h) after acute exercise. TGF-beta1 mRNA was quantified by RT-PCR in these samples, and basal TGF-beta1 protein levels were determined in skeletal muscle in the UTpre and Tpre subgroups by ELISA. Acute exercise caused a non-significant increase in TGF-beta1 mRNA in skeletal muscle in UT0h rats, in compare to UTpre rats. There was a significant decrease of TGF-beta1 mRNA in the T0h group (p=0.0013) in compare to Tpre rats. Prolonged training caused a significant increase in TGF-beta1 mRNA (p=0.02); however, the TGF-beta1 protein level decreased (p=0.02). In heart muscle, there was a significant decrease of TGF-beta1 mRNA in UT0h (p=0.01) and UT3h (p=0.04) compared to UTpre rats. TGF-beta1 mRNA levels were unchanged in T0h and T3h compared to Tpre; basal TGF-beta1 mRNA expression after training was also unchanged (UTpre vs. Tpre). We conclude that physical exercise is a potent stimulus for inducing TGF-beta1 gene expression in skeletal muscle, but does not increase the protein level. Thus, skeletal and heart muscle do not contribute to increased serum levels of TGF-beta1 after physical exercise.

Remodeling of human transplanted myocardium in ten-year follow-up: a clinical pathology study.

UNLABELLED: Remodeling taking place in transplanted myocardium leads to a change in the number of cardiocytes. Ultrasound measurements and biopsy evaluation should reflect their loss and compensation. We sought to evaluate the morphology of the transplanted heart upon long-term follow-up. MATERIAL AND METHODS: Myocardial biopsies were obtained in the first week, first month, and then annually for 10 years from transplantation that did not show rejection (grade "0" ISHLT, 122 biopsies). The control group encompassed 7 donor heart fragments. Proliferation in biopsies was evaluated with Ki67 (M7240, DAKO), cardiocyte hypertrophy by measuring their diameter, the surface area of the nuclei, nuclear-sarcoplasmic index, and stromal fibrosis evaluated as the surface area fraction. Ultrasound measurements included diastolic thickness of the interventricular septum, posterior wall of the left ventricle, and left ventricular mass. The correlation of measurements with time from transplantation was evaluated using Spearman's test. RESULTS: A positive Ki67 reaction was observed in fibroblasts and endothelial cells. The increased cardiocyte nuclear area correlated with the time elapsed since transplantation (r = 0.2; P < .05) with a simultaneous decrease in cardiocyte thickness (r = -0.3; P < .05), without changes in the nuclear-cytoplasmic index (r = 0.02; P > .05). Stromal fibrosis also increased (r = 0.1; P < .05). Ultrasound measurements of the left ventricle showed a decreased tendency with the passage of time (r = -0.2 to -0.3; P < .05). CONCLUSION: A transplanted heart does not undergo hypertrophy but rather fibrous atrophy with apparent compensatory hypertrophy of the cardiomyocytes.

Cardiocyte nucleus shape as an indicator of heart graft aging.

UNLABELLED: Morphometric publications based on the measurement of cardiocyte nuclei indicated their progressive hypertrophy ignoring, however, their shape, which is a deciding factor for the microscopic-based diagnosis of hypertrophy. We sought was to demonstrate how the shapes of cardiocyte nuclei change over time and correlate them with the thickness of the interventricular septum, (IVS) the biopsy site. MATERIAL: We evaluated myocardial biopsies taken in the first week, first month, and then annually until posttransplant year 10. Only biopsies with no rejection were considered: grade "0" ISHLT (122 biopsies). The control group encompassed fragments from seven donor hearts. METHODS: Cardiomyocyte nuclei were evaluated morphometrically. We calculated the length, breadth, perimeter, roundness, elongation, and fullness factors for correlation with the IVS thickness, and selected indices. The relationships between karyometry and IVS thickness (measured by ultrasound) as well as time were calculated by Spearman's correlation test. RESULTS: Among the examined indices, only nuclear length did not correlate significantly with follow-up time. Among the remaining indices, the strongest correlations with time were observed with regard to breadth (r = 0.214), perimeter (r = 0.150), roundness (r = -0.06) and fullness (r = 0.06), and finally elongation (r = 0.02). The decreasing thickness of the interventricular septum (r = -0.31) showed a weak correlation only with the cardiocyte nuclear length (r = -0.05). CONCLUSION: Graft aging imitates hypertrophy inasmuch as cardiocyte nuclei become wider despite the decreased thickness of the interventricular septum. Therefore, karyometric measurements do not reflect myocardial morphology.

The serum levels of growth factors: PDGF, TGF-beta and VEGF are increased after strenuous physical exercise.

Strenuous physical exercise induces muscle fibers damage and non-specific inflammatory response. Activated by inflammatory process cells may serve as the source of wide spectrum of inflammatory mediators and growth factors. Namely Platelet Derived Growth Factor (PDGF), Transforming Growth Factor-beta (TGF-beta) and Vascular Endothelial Growth Factor (VEGF) could be released. The aim of present study was to assess the impact of physical exercise on growth factors generation in healthy young people. 14 young sportsmen were enrolled into the study. They performed strenuous physical exercise. Blood samples were drawn before, immediately after, and 2 hours after the exercise bout. Serum PDGF, TGF-beta and VEGF concentrations were measured using commercially available ELISA kit based on immunoenzimatic method. Serum level of PDGF increased significantly from 1.7 ng/ml before to 4.64 ng/ml (2.73-fold) immediately after, and to 3.3 ng/ml (1.94-fold) 2 hours after exertion. Serum level of TGF-beta increased significantly from 20.58 ng/ml before to 55.37 ng/ml (2.7-fold) immediately after, and to 40.03 ng/ml (1.95-fold) 2 hours after exertion. Serum level of VEGF increased significantly from 91.83 pg/ml before to 165.61 pg/ml (1.8-fold) immediately after the exercise. Two hours after the exertion serum level of VEGF was 137.22 pg/ml, what is 1.49-fold above the basal level; however not being significantly different. In summery, observed increased level of growth factors could be involved in the process of adaptation of human organism to physical training. In addition, in the context of the role of inflammation in the pathogenesis of various diseases, our results point to the potentially deleterious effect of strenuous physical exercise.

Proteolytic enzymes activities in patients after myocardial infarction correlate with serum concentration of TGF-beta.

Myocardial infarction (MI) is caused by occlusion of coronary artery and insufficient oxygen supply to a certain area of myocardium. Its necrosis appears as a result of MI. The process of tissue repair after MI is very complicated and it is influenced by numerous factors, including growth factors and proteolytic enzymes. The aim of the study was to determine serum transforming growth factor beta (TGF-beta) concentration on day 2 and 7 after MI and to asses the relationship of this growth factor with serum proteolytic activity of collagenase and elastase. In addition, the effect of fibrynolytic treatment on these factors was evaluated. About 100 patients with MI were enrolled to the study. The control group consisted of 50 healthy individuals. We observed that TGF-beta1 concentration correlated positively with collagenase activity on the second day after MI and that it also correlated positively with elastase activity on day 2 and 7 after MI. Moreover, treatment with streptokinase (SK) caused a significant increase of TGF-beta serum concentration. Our data indicate that TGF-beta1 may be one of the factors involved in tissue repair process after MI. Its effect seems to be mediated by collagenase and elastase and may change with the time that elapsed after MI.

Influence of the stimulation of central chemoreceptors on the gastric mucosal blood flow in artificially ventilated and spontaneously breathing rats.

Respiratory failure coincides frequently with the occurrence of gastric ulceration. In advanced respiratory insufficiency hypoxemia is often accompanied by hypercapnia, which is the stimulus for central chemoreceptors as well as for carotid body chemoreceptors. The purpose of the work was to investigate the reflex effect of stimulation of central chemoreceptors on gastric mucosal blood flow (GMBF) in the rat. Central chemoreceptors were stimulated by a gas mixture composed of 10% carbon dioxide, 50% oxide and 40% nitrogen. In artificially ventilated and spontaneously breathing animals, the stimulation of central chemoreceptors caused a significant increase in gastric mucosal vascular resistance, accompanied by a marked decline in blood flow. We hypothesize that in patients with respiratory insufficiency accompanied by hypercapnia, the reflex impairment of GMBF may contribute to gastric ulceration.

Influence of the stimulation of carotid body chemoreceptors on the gastric mucosal blood flow in artificially ventilated and spontaneously breathing rats.

The cardiovascular effects of the stimulation of arterial chemoreceptors are different in spontaneously breathing and artificially ventilated animals. Respiratory failure and long term sojourn at high altitude coincide frequently with the occurrence of gastric ulceration. In both these situations a profound stimulation of arterial chemoreceptors is present. The purpose of the paper was to investigate the reflex effect of stimulation carotid chemoreceptors on gastric mucosal blood flow in the rat. Arterial chemoreceptors were stimulated by two methods (I) substitution gas mixture of 10% oxygen in nitrogen for room air and (II) direct injection of acid saline ( 0.05 ml, pH = 6.8) into the distal part of left common carotid artery. In artificially ventilated rats stimulation of arterial chemoreceptors caused significant increase in gastric mucosal vascular resistance, accompanied by marked decline in blood flow. This effect was mediated by adrenergic mechanism. On the contrary to artificially ventilated rats, decline of gastric mucosal vascular resistance with concomitant increase in blood flow was found in spontaneously breathing animals. This effect was not abolished either by phentolamine or atropine. As vasodilatatory effect of arterial chemoreceptors stimulation was abolished by bilateral vagotomy, we postulate that non adrenergic and non cholinergic vagal fibers mediate observed vascular changes in gastric mucosa in spontaneously breathing rats. We hypothesize that in artificially ventilated patients with respiratory failure stimulation of arterial chemoreceptors by hypoxemia and or acidosis may contribute to the development of gastric mucosal lesions.

Fucoidan improves the renal blood flow in the early stage of renal ischemia/reperfusion injury in the rat.

It has been shown that monoclonal anti-P-selectin antibody administration protects renal function in an ischemic model of acute renal failure. This study was designed to evaluate the effect of administration of fucoidan, P-selectin inhibitor, on reduction in renal blood flow induced by ischemia/reperfusion injury in the rat. Experiments were performed on male Wistar rats weighting 35-400 g. The systemic blood pressure (mm Hg) (BP) and renal blood flow (RBF) were monitored continuously and renal vascular resistance (RVR) was calculated. After 20 min period of stabilization animals (6 rats in each group) received one of the following agents administered by continuous i.v. infusion during 165 min: 1 mg/kg of body weight of fucoidan (F1), 10 mg/kg of fucoidan (F10), 100 mg/kg of fucoidan (F100), 10 mg/kg of heparin (H), or 0.9% NaCl solution (control). After 15 min of drug administration the renal vessels of the both kidney were occluded with vascular clamps for 60 min. There were no significant changes in the initial values of RBF, RVR and BP between groups. None procedure affected significantly BP during all experiments. In F10 RBF returned to the initial values in 70th min of reperfusion and did not change up to 90th min. This value was significantly higher than respective value in the control group. In F1 group RBF in 90th min was also higher than in the control group, but it was not statistically significant. The dose of heparine and fucoidan used in the H and F100 groups failed to preserve RBF during reperfusion. In the present study we found that administration of fucoidan--P-selectin inhibitor, increases significantly postischemic renal blood flow and may have renoprotective activity.

Functional studies on sciatic nerve blood flow in respect to its vascular supply and tonic neural activity.

An introduction of laser flow meters for a continuous measurements of a tissue blood flow has opened new avenues for an accurate assessment blood flow in peripheral nerves. The aim of our study was: 1) to carry out a functional verification of anatomical sources of a sciatic nerve blood supply in the rat; 2) develop a measurement technique to facilitate standardisation of results; 3) to determine the role of nerve fibres tonic activity in the maintenance of a resting blood flow in the sciatic nerve. Based on results of the present study the following conclusions have been drawn out: 1) in order to obtain a real values of the blood flow through the sciatic nerve it is necessary to remove its muscular fascia; 2) an uninjured epineurium plays a crucial role in maintaining the resting blood flow; 3) major blood supply of sciatic nerve comes from inferior gluteal and popliteal arteries; 4) the tonic neural activity plays a role in the maintenance of the resting sciatic nerve blood flow in anaesthetised rats.

A role of network arrangements of microcirculation vessels in cardiovascular function.

The objective of the work was to explain the mechanisms for the appearance or creation of the so-called preferential channels for blood flow in tissues. We postulated that the addressed preferential blood flow occurs at the level of network structure of vascular system and results directly from its heterogeneity. The studies have been done in regular, two-dimensional network model of microvessels. Due to poorly defined dimensions, shapes and tortuousity of real vessels, the values of network elements were random. We examined how the heterogeneity of network elements influences flow redistribution within the network and the global network resistance to flow. The results indicated that the higher segmental resistance scatters the greater probability of the appearance within the network preferential flow paths passing through the whole network or being its local singularity. These channels significantly reduce the effect the global network resistance increases.

The adrenal renal vascular connection plays an essential role in the pathogenesis of renovascular hypertension in the rat.

The adrenal renal portal circulation (ARPC) contributes to decrease in renal blood flow occurring after renal artery clipping. The aim of present study was to determine the role of the ARPC in the development of the renovascular hypertension in 1-kidney 1-clip model in the rat. Experiments were performed on male Wistar rats. In the control group (A) the right nephrectomy and adrenalectomy were done. In the experimental groups renovascular hypertension was produced by clipping the left renal artery (silver clip ID 0.40 mm). In the first of the experimental groups (B) the right nephrectomy and adrenalectomy were done. In the second experimental group (C), for elimination of the ARPC, the right kidney and the left adrenal gland were removed. In the half number of rats from each group plasma renin activity was measured 48 hours after surgery. An increase in SBP was significantly higher in the group B (ARPC intact) than in the group C (ARPC eliminated) (172 +/- 4 vs 144 +/- 2 mmHg, p.<0.01). PRA was significantly higher in the group C than in the group B (39.0 +/- 1.4 vs 31.2 +/- 2.0 mmol/l/min, p.<0.05). In the control group (A) PRA was significantly lower as compared to the both experimental group (2.0 +/- 1.6 mmol/l/min, p.<0.05).

Kidney denervation combined with elimination of adrenal-renal portal circulation prevents the development of hypertension in spontaneously hypertensive rats.

1. Kidney denervation in spontaneously hypertensive rats (SHR) during the prehypertensive stage delays and attenuates the development of hypertension. The same results have been obtained after elimination of the adrenal-renal portal circulation (ARPC). The aim of the present study was to investigate the influence of concomitant kidney denervation and elimination of ARPC on hypertension in SHR. 2. Experiments were performed on 6-week-old male SHR and Wistar-Kyoto (WKY) rats. In the first group of animals (group I), the ARPC was eliminated by removing the left adrenal gland and the right kidney. In the second group of rats (group II), the right kidney and the right adrenal gland were removed and the left kidney was denervated. In the third group of rats (group III), the right adrenal gland and the left kidney were removed and the right kidney was denervated. In the fourth group of rats (group IV), the right adrenal gland and the right kidney were removed. Group IV served as the control group. Denervations were repeated every 3 weeks. Systolic blood pressure was measured indirectly. 3. Elimination of ARPC (group I) and kidney denervation (group II) delayed and attenuated hypertension to the same degree (163 +/- 5 and 157 +/- 4 mmHg, respectively). Application of the these two methods concomitantly (group III) prevented the development of hypertension (130 +/- 6 mmHg). 4. We conclude that both intact efferent sympathetic renal nerves and adrenal hormones reaching the kidney through the ARPC may be mandatory factors for the development of arterial hypertension in SHR.

Arterial hypertension due to occlusion of the adrenal vein in the rat is strain-dependent.

OBJECTIVE: To determine whether the development of arterial hypertension due to occlusion of the central adrenal vein in the rat is strain-dependent DESIGN AND METHODS: The experiments were performed on male rats weighing 300-400 g each, of the following strains: Wistar outbred, Wistar Glaxo, Lewis, Wistar-Kyoto (WKY) rats bred for high blood pressure (138 +/- 13.2 mmHg), WKY rats bred without the control of blood pressure (118 +/- 12.9 mmHg) and borderline hypertensive rats (BHR). BHR were the F1 spontaneously hypertensive rat and WKY rat crossbred rats. In order to increase blood flow through the adrenal-renal portal circulation, both central adrenal veins of rats in the experimental group were occluded. The systolic blood pressure was measured indirectly by a photoelectric method. RESULTS: By the ninth day after surgery systolic blood pressure had increased significantly only in the WKY rats bred for high blood pressure and BHR, reaching maximal values on 12th day for WKY rats bred for high blood pressure (167 +/- 5 mmHg) and on the 18th day for BHR (170 +/- 14 mmHg). CONCLUSIONS: These data show that the development of arterial hypertension due to augmentation of adrenal blood flow through adrenal-renal portal circulation occurs in rats of strains with a genetic background of hypertension.

Non-iatrogenic perforation of the stomach by a chest tube in a patient with traumatic diaphragmatic hernia.

A 40-year-old Libyan male was admitted to the intensive therapy unit of Zliten Central Hospital, Libya after a road traffic accident in which he had been the driver. On physical examination he was irritable, dyspnoeic, cyanotic, had contusions and abrasions on his chest and abdomen (mainly on the left side), a lacerated wound on the forehead, a large haematoma over the left thigh and tenderness over the left side of the chest and abdomen.

Occlusion of the adrenal vein leads to an increase in renal vascular resistance in the ipsilateral kidney.

1. The aim of the present study was to investigate the effect of an acute increase in blood flow through the adrenal-renal vascular connection (ARVC), due to occlusion of the adrenal vein, on renal blood flow (RBF) and renal vascular resistance (RVR). 2. Experiments were performed in Wistar-Kyoto rats. Animals were divided into four groups. In the first group, the adrenal vein was clamped using a metal clip. In the second group, the tissue between the adrenal gland and the kidney was cut before clamping. In the third and fourth groups, alpha-adrenoceptor blockade was induced by intravenous infusion of phentolamine before clamping the adrenal vein. In addition, in the fourth group, tissue between the adrenal gland and the kidney was cut before clamping. 3. In the first group, occlusion of the adrenal vein caused an increase in RVR, followed by a decrease in RBF. In the second, third and fourth groups, the same procedure did not change either RBF or RVR. 4. We conclude that the augmented inflow of catecholamines to the kidney through the ARVC changes the haemodynamics of the kidney and may contribute to development of arterial hypertension due to occlusion of the adrenal vein.

Adrenal-renal portal circulation contributes to decrease in renal blood flow after renal artery stenosis in rats.

The aim of the present study was to investigate a role of adrenal-renal portal circulation (ARPC) in a decrease in renal blood flow due to acute stenosis of the renal artery in rats. Animals were divided into three groups. In the control group (I), in order to eliminate the ARPC tissue between the adrenal gland and the ipsilateral kidney was cut. In the second and the third group (II) (III), left renal artery was stenosed by a silver clip (ID 0.40 mm). Then, in the group II, ARPC was surgically eliminated. In the group II, prior to the elimination of ARPC, alpha-adrenergic receptors blockade was produced by phentolamine administration. In the control group, ARPC elimination did not influence either renal blood flow (RBF) or renal vascular resistance (RVR). In the group II, elimination of ARPC caused increase in RBF and decrease in RVR In the group III elimination of ARPC influenced neither RBF nor renal vascular resistance (RVR). Results of the present study provide the functional evidence that catecholamines reaching the kidney through ARPC, contribute to the decrease in RBF and increase in RVR during acute renal artery stenosis in the rat.

The adrenal-renal vascular connection contributes to increase in renal vascular resistance during an experimental hypotension in the rat.

The adrenal vascular connection (ARVC) was described for the first time in the cat by Cow (1914) and by other authors in the dog, rat, rabbit and humans. The aim of the present study is to investigate the role of above connection in regulation of renal vascular resistance (RVR), and renal blood flow (RBF) during decrease in blood pressure in the rat. Animals were divided into three groups. In the first group, mean arterial pressure (MAP) was unchanged. In the second and the third group MAP was maintained at 50 mmHg. In addition in the third group, an alpha adrenergic receptor blockade was produced with intravenous infusion of phentolamine. After stabilisation of RBF, in all groups, the tissue between the adrenal gland and the kidney was cut. RBF and MAP were measured and recorded. In the first and the third group, the elimination of ARVC neither influenced RBF nor RVR. In the second group the elimination of ARVC caused increase in RBF and decrease in RVR (p < 0.01). Results of the present study provide the evidence that catecholamines reaching the kidney, directly from the adrenal gland through ARVC, during the severe hypotension are responsible for reducing of renal blood flow and increase in renal vascular resistance in the rat.