Biomedical subjects

Kristina M Reber

Publications and source records attributed to Kristina M Reber.

12 recordsLinked to original sources

PubMed2007

Endothelial nitric oxide synthase in human intestine resected for necrotizing enterocolitis.

OBJECTIVE: To determine the expression and function of endothelial nitric oxide synthase (eNOS) in submucosal arterioles harvested from human intestine resected for necrotizing enterocolitis (NEC) or congenital bowel disease. STUDY DESIGN: eNOS expression was determined by using immunohistochemistry. The arteriolar diameter was measured in vitro at pressures of 10 to 40 mm Hg and also in response to the eNOS agonist acetylcholine (ACh), the exogenous nitric oxide (NO) donor S-nitroso-N-acetylpenicillamine, and the smooth muscle relaxant papaverine. Arteriolar release of NO in response to ACh was determined with a Sievers NOAnalyzer. Hemodynamics were also determined at flow rates of 50 and 100 microL/min. RESULTS: eNOS was present in microvessels from both groups, but NEC arterioles failed to demonstrate physiological evidence of eNOS function: they constricted in response to pressure, failed to dilate or generate NO in response to ACh, and failed to dilate in response to flow. However, they dilated in response to exogenous NO and papaverine, indicating functional vascular smooth muscle and vasodilator reserve. CONCLUSION: eNOS-derived NO, a vasodilator in the newborn intestine, did not contribute to vasoregulation in arterioles harvested from intestine resected for NEC. These vessels were constricted; lack of eNOS-derived NO may contribute to this vasoconstriction.

Arterioles↗

PubMed2006

Effects of prenatal lipopolysaccharide exposure on epithelial development and function in newborn rat intestine.

BACKGROUND: Maternal infection during pregnancy is associated with several neonatal morbidities, including periventricular leukomalacia and lung maldevelopment and injury. OBJECTIVE: To test the hypothesis that responses to prenatal maternal exposure to lipopolysaccharide (LPS) alter intestinal epithelial development and function in newborn rats. DESIGN/METHODS: Timed-pregnancy female Sprague-Dawley rats were administered either 2 mg LPS or an equal volume of isotonic saline by intraperitoneal injection at E16 and allowed to deliver naturally. Pups were weighed and then killed at days of life (DOL) 0, 3, 7 and 14. Morphometric parameters were measured on standard hematoxylin and eosin-stained sections using ImagePro software. Immunohistochemistry was performed with antibody specific for inducible nitric oxide synthase (iNOS) and 3-nitrotyrosine on distal ileal intestinal samples analyzed at each time point. Optical density was determined and quantified for site-specific regions of intestinal sections. On DOL 14, in vivo mucosal permeability was measured by feeding rats fluorescein isothiocyanate (FITC) via orogastric tube; and then serum FITC was measured. RESULTS: There were no significant differences in pup weights. Mucosal thicknesses were significantly less in the distal ileum from pups born to LPS-exposed dams on DOL 0, 3 and 7 (P < 0.001). On DOL 0, iNOS protein concentrations in the prenatal LPS treatment group were significantly greater than iNOS protein concentrations in the distal villus (P < 0.001), proximal villus/crypts (P < 0.01), submucosa (P < 0.001) and muscularis (P < 0.01) in the distal small intestine of the control group. On DOL 3, 7 and 14, significant differences were observed in iNOS protein concentrations in the distal villus and submucosal regions between groups (P < 0.001). On DOL 0, 3, 7 and 14, 3-nitrotyrosine immunostaining was significantly elevated in the prenatal LPS-exposed pups in the distal villus on (P < 0.001) as well as in the submucosa on DOL 3 (P < 0.001). Serum FITC measurement was significantly greater in prenatal LPS exposure group at DOL 14 (P < 0.001). CONCLUSIONS: Maternal exposure to LPS during pregnancy alters intestinal growth and regulation of iNOS in the newborn rat intestine.

Animals↗

PubMed2006

Intestinal O2 consumption in necrotizing enterocolitis: role of nitric oxide.

We tested the hypothesis that inducible isoform of nitric oxide synthase (iNOS)-derived nitric oxide (NO) inhibits oxygen consumption (VO2) in human intestine resected for necrotizing enterocolitis (NEC). Each NEC resection specimen was divided into two sections based on histologic appearance: healthy or diseased. Intestine removed from infants for reasons other than NEC was used as control. The tissue injury score (0-6, with 6 indicating complete necrosis) was 0.4 +/- 0.2 in control tissue, 1.2 +/- 0.4 in NEC-healthy tissue, and 4.6 +/- 0.5 in NEC-diseased tissue. Prominent iNOS staining was present in villus enterocytes in NEC-healthy tissue but not in the other tissue types. Intestinal VO2 (per direct oximetry, in nM O2/min/g) was significantly greater in control tissue than in NEC-healthy or NEC-diseased tissues. Accumulation of NO into buffer bathing intestinal slices (in nM NO/microL/g) was greater in NEC-healthy tissue than control or NEC-diseased tissues. The specific iNOS antagonist L-Nomega-(1-iminoethyl)-lysine (L-NIL) reduced buffer NO concentration 76% and increased VO2 by 90% in NEC-healthy tissue; however, L-NIL had no effect on NO or VO2 in control or NEC-diseased tissue. Addition of exogenous NO via S-nitroso-N-acetylpenicillamine depressed VO2 in NEC-healthy and control tissues but not NEC-diseased tissue. A significant correlation was present between buffer NO concentration and VO2 in NEC-healthy tissue. We conclude that iNOS-derived NO suppresses VO2 in intestine resected for NEC that demonstrates a relatively normal histology on light microscopy.

Animals↗

PubMed2005

Gene transfer with inducible nitric oxide synthase decreases production of urea by arginase in pulmonary arterial endothelial cells.

Nitric oxide (NO) is a vasodilator produced from L-arginine (L-Arg) by NO synthase (NOS). Gene therapy for hypertensive disorders has been proposed using the inducible isoform of NOS (iNOS). L-Arg also can be metabolized to urea and L-ornithine (L-Orn) by arginase, and L-Orn can be metabolized to proline and/or polyamines, which are vital for cellular proliferation. To determine the effect of iNOS gene transfer on arginase, we transfected bovine pulmonary arterial endothelial cells (bPAEC) with an adenoviral vector containing the gene for iNOS (AdiNOS). As expected, NO production in AdiNOS bPAEC was substantially greater than in control bPAEC. Although urea production was significantly less in the AdiNOS bPAEC than in the control bPAEC, despite similar levels of arginase I protein, AdiNOS transfection of bPAEC had no effect on the uptake of L-Arg. Inhibiting NO production with Nomega-nitro-L-arginine methyl ester increased urea production, and inhibiting urea production with L-valine increased nitrite production, in AdiNOS bPAEC. The addition of L-Arg to the medium increased urea production by AdiNOS bPAEC in a concentration-dependent manner. Thus, in these iNOS-transfected bPAEC, the transfected iNOS and native arginase compete for a common intracellular pool of L-Arg. This competition for substrate resulted in impaired proliferation in the AdiNOS-transfected bPAEC. These findings suggest that the use of iNOS gene therapy for pulmonary hypertensive disorders may not only be beneficial through NO-mediated pulmonary vasodilation but also may decrease vascular remodeling by limiting L-Orn production by native arginase.

Adenoviridae↗

PubMed2005

Cytokine-induced endothelial arginase expression is dependent on epidermal growth factor receptor.

L-arginine is metabolized to nitric oxide (NO) by NO synthase (NOS), or to urea and L-ornithine by arginase. L-ornithine contributes to vascular remodeling in pulmonary hypertension via metabolism to polyamines and proline. Previously we found that cytokines upregulate both NOS and arginase in pulmonary arterial endothelial cells. We hypothesized that cytokine-induced arginase I and II expression depend on epidermal growth factor (EGF) receptor (EGFR) activity. Bovine pulmonary arterial endothelial cells were treated with lipopolysaccharide and tumor necrosis factor-alpha (L/T). L/T treatment resulted in a substantial increase in urea production, and this increase in urea production was potently inhibited by both genistein and AG1478, inhibitors of EGFR. Levels of arginase I protein and arginase II mRNA were increased in response to L/T treatment, and genistein prevented the L/T-induced elevations in both arginase I protein and arginase II mRNA levels. L/T treatment increased production of nitrites and inducible NOS mRNA accumulation, and genistein and AG1478 had little effect on these changes. EGF (50 ng/ml) treatment resulted in enhanced urea production. Finally, a 170-kD protein was phosphorylated upon treatment with either EGF or L/T. Our results indicate that arginase induction by L/T depends in part on EGFR activity. We speculate that EGFR inhibitors may attenuate vascular remodeling without affecting NO release, and thus may represent novel therapeutic modalities for pulmonary hypertensive disorders.

Animals↗

PubMed2005

Endothelin-1 in human intestine resected for necrotizing enterocolitis.

OBJECTIVES: We asked if the tissue concentration of the potent vasoconstrictor endothelin-1 (ET-1) is greater in areas of human preterm intestine that demonstrate histologic evidence of necrotizing enterocolitis (NEC) when compared with relatively healthy areas within the same resection specimen. We then evaluated if ET-1 participates in hemodynamic regulation within intestinal subserosal arterioles harvested from portions of human preterm intestine that demonstrate NEC. STUDY DESIGN: Human preterm intestine resected for NEC was divided into three zones based on proximity to the perforation (zone 1 most proximal, zone 3 most distal). Histologic evidence of NEC was determined in each zone (normal = 0, advanced necrosis = 6). The tissue concentration of ET-1 was determined by enzyme-linked immunosorbent assay within intestinal homogenates prepared from each zone. Arteriolar hemodynamics were determined in vitro on subserosal arterioles harvested from different zones. Arteriolar flow rate, diameter, and resistance were determined at pressure gradients (DeltaP) of 20 and 40 mmHg under control conditions and again after blockade of endothelin ET A receptors with BQ610 (10 -9 mol/L). RESULTS: The tissue concentration of ET-1 (pg/mg protein) and histologic score in the three zones were: zone 1: 84 +/- 14, 5.5 +/- 0.3; zone 2: 99 +/- 12, 4.7 +/- 0.4, and zone 3: 33 +/- 9, 0.8 +/- 0.6, respectively (M +/- SD, n = 10 resection specimens, P < .05, zone 3 vs zones 1 and 2). Zone 2 arterioles demonstrated significantly lower flow rate and diameter and increased resistance under control conditions than zone 3 arterioles when DeltaP was either 20 or 40 mmHg (n = 7, P < .05). Treatment with BQ610 had no effect on zone 3 arterioles but significantly vasodilated zone 2 arterioles, increasing flow rate and vessel diameter, and decreasing vascular resistance (n = 7, P < .05). CONCLUSIONS: The tissue concentration of ET-1 is greater in human preterm intestine that demonstrates histologic evidence of NEC. Arterioles harvested from intestine exhibiting histologic evidence of NEC demonstrate vasoconstriction when compared with arterioles from relatively healthy intestine in the same resection specimen. This vasoconstriction was reversed by blockade of endothelin ET A receptors.

Arterioles↗

PubMed2004

Developmental expression of endothelin receptors in postnatal swine mesenteric artery.

Studies were conducted to determine whether endothelin (ET) ETA and ETB receptor protein and mRNA expression is developmentally regulated in the postnatal swine mesenteric circulation. To this end, Western blotting and real-time reverse PCR were performed on protein and total RNA isolated from the mesenteric artery harvested from 3-, 10-, and 30-d-old swine. Western blot analysis revealed that ETA and ETB receptor protein expression in the swine mesenteric artery decreased over the age range studied; thus, ETA and ETB receptor protein expression was significantly greater in the 3-d-old group then progressively declined over the first postnatal month. Similar to the Western data, real-time PCR analysis revealed that ETA and ETB receptor mRNA expression also decreased over the age range studied; thus, ETA and ETB receptor mRNA expression was significantly greater in the 3-d-old group then progressively declined over the first postnatal month. Immunohistochemistry localized the ETA receptor to the vascular smooth muscle and the ETB receptor to the endothelial cell layer. Additionally, we report a partial cDNA sequence for the swine ETB receptor. We conclude that ETA and ETB receptor protein and mRNA expression is developmentally regulated in the postnatal swine mesenteric artery, being expressed to a greater degree in younger animals.

Amino Acid Sequence↗

PubMed2004

Necrotizing enterocolitis: preventative strategies.

Necrotizing enterocolitis (NEC) remains a major cause of morbidity and mortality in premature infants. Although the pathogenesis of NEC is unclear, prevention strategies have been developed based on clinical observations and epidemiologic and experimental data. Most current strategies have centered on feeding practices (eg, institution of feeds, advancement of feeds, composition of feeds, and standardization of feeding practices). Emerging strategies include amino acid supplementation, the use of platelet-activating factor(PAF) antagonists or PAF-acetylhydrolase administration, polyunsaturated fatty acid administration, epidermal growth factor administration, and the use of pre- and probiotics.

Administration, Oral↗

PubMed2004

The effect of common origin of the carotid arteries in neurologic outcome after neonatal ECMO.

BACKGROUND: Common origin of the carotid arteries (COCA) is a normal anatomic variant reported to occur in approximately 11% of the general population. The objective of this study was to determine whether this variant places venoarterial extracorporeal membrane oxygenation (ECMO) patients at a higher risk for adverse neurologic sequelae owing to potential occlusion of both carotid arteries by the arterial cannula. METHODS: The authors reviewed clinical records and echocardiograms of the initial 220 ECMO patients at their institution. Aortic arch morphology was determined by a pediatric cardiologist blinded to all other data. After exclusion of predetermined patients, 131 patients were divided into 2 groups: those with separate origin of the carotid arteries (n = 111) and those with COCA (n = 20). The neurologic outcome variables studied included the results of magnetic resonance imaging (MRI); computed tomography (CT); electroencephalogram (EEG); brainstem auditory-evoked response (BAER), head ultrasound scan, and Bayley Scales of Infant Development reported as Psychomotor Developmental Index (PDI) and Mental Developmental Index (MDI). RESULTS: COCA had no predictive value in determining PDI and MDI outcomes and no significance in predicting an increased risk of adverse neurologic sequelae based on MRI, CT, EEG, BAER, or head ultrasound scan. CONCLUSIONS: This study confirms that COCA is a common aortic arch variant (15%, n = 20 of 131) and that this variant does not appear to increase the risk of neurologic injury in infants undergoing venoarterial ECMO.

Brachiocephalic Trunk↗

PubMed2003

Development of the myogenic response in postnatal intestine: role of PKC.

Previous attempts to determine developmental changes in the vascular myogenic response have been confounded by the presence of competing vasoactive stimuli or the use of isolated vessels with markedly different baseline diameters. To circumvent these issues, small mesenteric arteries (diameter approximately 150 microm) from 1- and 10-day-old piglets were studied in vitro under no-flow conditions. In situ studies demonstrated that the intravascular pressure and diameter of these vessels were similar in both age groups, allowing an effective comparison of the myogenic response not obscured by differences in basal diameter. The pressure-diameter relationship was age specific. Thus, although small mesenteric arteries from both age groups demonstrated myogenic constriction in response to stepwise increases in pressure (0 to 100 mmHg, in 20-mmHg increments), the intensity of contraction was significantly greater in vessels from 1-day-old piglets particularly within the pressure range normally experienced by these vessels in situ. Attenuation or activation of PKC with calphostin C or indolactam, respectively, substantially altered the pressure-diameter relationship in 1-, but not 10-day-old arteries; thus calphostin C essentially eliminated the contractile response to pressure elevation in younger subjects, whereas indolactam significantly increased the intensity of the myogenic response and shifted its activation point to a lower pressure range. Immunoblots carried out on protein recovered from these arteries revealed the presence of alpha, beta, epsilon, iota, and lambda; notably, expression of the alpha- and epsilon-isoforms substantially decreased between postnatal days 1 and 10.

Animals↗

PubMed2002

Newborn intestinal circulation. Physiology and pathophysiology.

The physiologic characteristics of the newborn intestinal circulation are unique when compared with the adult condition. Most important, intestinal vascular resistance across newborn intestine is exceptionally low and this transient reduction is mediated by an increased constitutive and stimulated production of NO. The low vascular resistance characteristic of newborn intestine alters the capacity of this vasculature to respond to systemic circulatory perturbations, such as hypotension and arterial hypoxemia. The essential role of endothelial production of NO in maintaining newborn intestinal hemodynamics might be important in the pathogenesis of NEC, because endothelial dysfunction would limit, or possibly eliminate, NO production, leading to substantial intestinal ischemia.

Blood Circulation↗

PubMed2002

Developmental expression of eNOS in postnatal swine mesenteric artery.

Developmental changes in the expression of endothelial nitric oxide synthase (eNOS) within the mesenteric artery of swine were studied in fetal (110 days postconception/117 days total gestation) and on postnatal days 1, 3, 10, and 30. Subjects in the 1-day-old group were subdivided into fed and nonfed. Transcription of eNOS was determined by real-time PCR, protein expression was evaluated by Western blotting, and hemodynamic and oxygenation parameters were measured within in situ gut loops before and after the administration of N(G)-monomethyl-L-arginine (L-NMMA). The abundance of eNOS mRNA remained steady throughout all ages. In contrast, expression of eNOS protein was twofold greater in the 1-day-old fed subjects compared with fetal or 1-day-old nonfed subjects. eNOS protein expression remained elevated on day 3, increased on day 10, and then declined to a level similar to the day 1 nonfed group by postnatal day 30. Intestinal vascular resistance was 31% lower in the day 1 fed group when compared to the day 1 nonfed group; resistance continued to decline through day 10 but then significantly increased on day 30. We conclude that the expression of eNOS changes within the mesenteric artery during early postnatal development at a posttranscriptional level.

Aging↗