Cell therapy for ARDS: efficacy of endobronchial versus intravenous administration and biodistribution of MAPCs in a large animal model.

Introduction: Bone marrow-derived multipotent adult progenitor cells (MAPCs) are adult allogeneic adherent stem cells currently investigated clinically for use in acute respiratory distress syndrome (ARDS). To date, there is no agreement on which is the best method for stem cells delivery in ARDS. Here, we compared the efficacy of two different methods of administration and biodistribution of MAPC for the treatment of ARDS in a sheep model. Methods: MAPC were labelled with [18F] fluoro-29-deoxy-D-glucose and delivered by endobronchial (EB) or intravenous route 1 hour after lipopolysaccharide infusion in sheep mechanically ventilated. PET/CT images were acquired to determine the biodistribution and retention of the cells at 1 and 5 hours of administration. Results: The distribution and retention of the MAPC was dependent on the method of cell administration. By EB route, PET images showed that MAPC remained at the site of administration and no changes were observed after 5 hours, whereas with intravenous route, the cells had broad biodistribution to different organs, being the lung the main organ of retention at 1 and 5 hours. MAPC demonstrated an equal effect on arterial oxygenation recovery by either route of administration. Conclusion: The EB or intravenous routes of administration of MAPC are both effective for the treatment of ARDS in an acute sheep model, and the effect of MAPC therapy is not dependent of parenchymal integration or systemic biodistribution.

Effects of propofol on intraocular pressure in premedicated and nonpremedicated dogs with and without glaucoma.

OBJECTIVE To establish a study cutoff for evidence of glaucoma on the basis of IOP measurements from a large population of healthy dogs and to assess the effects of IV propofol administration on IOPs in premedicated and nonpremedicated dogs with and without glaucoma defined by this method. DESIGN Prospective, descriptive study. ANIMALS 234 client-owned dogs. PROCEDURES IOPs measured in 113 healthy dogs (226 eyes) were used to calculate an IOP value indicative of glaucoma. The IOPs were measured in an additional 121 dogs (237 eyes) undergoing ophthalmic surgery. Midazolam-butorphanol was administered IV as preanesthetic medication to 15 and 87 dogs with and without glaucoma, respectively. A placebo (lactated Ringer solution) was administered IV to 8 and 11 dogs with and without glaucoma, respectively. Anesthesia of surgical patients was induced with propofol IV to effect. The IOPs and physiologic variables of interest were recorded before (baseline) and after preanesthetic medication or placebo administration and after propofol administration. RESULTS An IOP > 25 mm Hg was deemed indicative of glaucoma. Compared with baseline measurements, mean IOP was increased after propofol administration in nonpremedicated dogs without glaucoma and unchanged in nonpremedicated dogs with glaucoma. Propofol-associated increases in IOP were blunted in premedicated dogs without glaucoma; IOP in affected eyes of premedicated dogs with glaucoma was decreased after preanesthetic medication and after propofol administration. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that preexisting IOP influences the response to anesthetic drugs, and administration of preanesthetic medication with muscle-relaxing properties may blunt or reduce propofol-induced increases in IOP. Further research with a larger number of dogs is needed to confirm our results in dogs with glaucoma.

Chemical signals of elephant musth: temporal aspects of microbially-mediated modifications.

Mature male African (Loxodonta africana) and Asian (Elephas maximus) elephants exhibit periodic episodes of musth, a state in which serum androgens are elevated, food intake typically decreases, aggressiveness often increases, and breeding success is enhanced. Urine is a common source of chemical signals in a variety of mammals. Elephants in musth dribble urine almost continuously for lengthy periods, suggesting that the chemicals in their urine may reveal their physiological condition to conspecifics. We investigated the volatile urinary chemicals in captive male elephants using automated solid phase dynamic extraction (SPDE) and gas chromatography-mass spectrometry (GC-MS). We found higher levels of alkan-2-ones, alkan-2-ols, and some aromatic compounds in urine from males in musth than in urine from non-musth males or from females. Levels of ketones and alcohols increased as the urine aged, likely due to microbial metabolism of fatty acids. Protein-derived aromatic metabolites also increased in abundance after urination, likely due to microbial hydrolysis of hydrophilic conjugates. We suggest that microbes may play an important role in timed release of urinary semiochemicals during elephant musth.

Hazards, safety, and anesthetic considerations for magnetic resonance imaging.

The increase in the use of magnetic resonance imaging (MRI) for diagnostic use in companion animals has increased the demand for anesthesia support in a strong magnetic environment. In many instances, this may necessitate anesthesia being provided by individuals that are unfamiliar with MRI and the hazards associated with it. The objective of this article is to describe the conditions and hazards associated with a strong magnetic field, review considerations for safe and effective anesthetic management of patients during an MRI, and promote close collaboration and communication between personnel in an effort to insure staff awareness and safety. This report describes conditions that exist for the superconducting high field strength magnets, 1.0, 1.5, and 3.0 Tesla, that are commonly used for clinical imaging of companion animals. Many of these same safety and anesthesia considerations can be applied to any MRI facility.

Fps/Fes protein-tyrosine kinase regulates mast cell adhesion and migration downstream of Kit and beta1 integrin receptors.

Activation of Kit receptor protein-tyrosine kinase (PTK) by its ligand Stem Cell Factor (SCF) is required for the development of mast cells, and for the regulation of mast cell proliferation, migration and modulation of inflammatory mediator release. Recent studies have implicated the non-receptor PTK Fps/Fes (hereafter referred to as Fes) in signaling downstream of oncogenic Kit, however, the potential role of Fes in regulating Kit signaling is not well defined. In this study, we show that SCF induces transient tyrosine phosphorylation of wild-type Fes as well as kinase-dead Fes in bone marrow-derived mast cells (BMMCs). The latter finding implicates an upstream kinase acting on Fes, which we identified as Fyn PTK. SCF treatment of BMMCs promoted recruitment of Fes to Kit, potentially via direct interaction of the Fes SH2 domain with phosphorylated Kit. While Fes was not required for SCF-induced signaling to Akt and Erk kinases, Fes-deficient (fes-/-) BMMCs displayed a defect in sustained p38 kinase activation, compared to control cells. SCF-treated Fes-deficient BMMCs also displayed elevated beta1 integrin-mediated cell adhesion and spreading on fibronectin, compared to control cells, and a reduction in cell polarization at later times of SCF treatment. Restoring Fes expression in fes-/- BMMCs by retroviral transduction was sufficient to rescue cell spreading and polarization defects. Interestingly, SCF-induced chemotaxis of BMMCs was also defective in Fes-deficient BMMCs, and restored in Fes-rescue BMMCs. Overall, these results implicate Fes in regulating cross-talk between Kit and beta1 integrins to promote cytoskeletal reorganization and motility of mast cells.

Contributions of F-BAR and SH2 domains of Fes protein tyrosine kinase for coupling to the FcepsilonRI pathway in mast cells.

This study investigates the roles of Fer-CIP4 homology (FCH)-Bin/amphiphysin/Rvs (F-BAR) and SH2 domains of Fes protein tyrosine kinase in regulating its activation and signaling downstream of the high-affinity immunoglobulin G (IgE) receptor (FcepsilonRI) in mast cells. Homology modeling of the Fes F-BAR domain revealed conservation of some basic residues implicated in phosphoinositide binding (R113/K114). The Fes F-BAR can bind phosphoinositides and induce tubulation of liposomes in vitro. Mutation of R113/K114 to uncharged residues (RK/QQ) caused a significant reduction in phosphoinositide binding in vitro and a more diffuse cytoplasmic localization in transfected COS-7 cells. RBL-2H3 mast cells expressing full-length Fes carrying the RK/QQ mutation show defects in FcepsilonRI-induced Fes tyrosine phosphorylation and degranulation compared to cells expressing wild-type Fes. This correlated with reduced localization to Lyn kinase-containing membrane fractions for the RK/QQ mutant compared to wild-type Fes in mast cells. The Fes SH2 domain also contributes to Fes signaling in mast cells, via interactions with the phosphorylated FcepsilonRI beta chain and the actin regulatory protein HS1. We show that Fes phosphorylates C-terminal tyrosine residues in HS1 implicated in actin stabilization. Thus, coordinated actions of the F-BAR and SH2 domains of Fes allow for coupling to FcepsilonRI signaling and potential regulation the actin reorganization in mast cells.

Intermittent bradyarrhythmia in a Hispaniolan Amazon parrot (Amazona ventralis).

A clinically normal 2-year-old Hispaniolan Amazon parrot (Amazona ventralis) was found to have periodic second-degree atrioventricular (AV) block with variable nodal conductions while anesthetized with isoflurane during a thermal-support research project. Arrhythmias were observed on 5 successive weekly electrocardiograms. A complete cardiac evaluation, including a diagnostic electrocardiogram, revealed intermittent bradyarrhythmias ranging from a 2:1 to a 7:1 second-degree AV block, with concurrent hypotensive episodes during the nodal blocks. Results of a complete blood cell count, plasma biochemical profile, blood gas analysis, and atropine-response test, as well as radiography and auscultation, revealed no obvious cause for the arrhythmias. Echocardiography demonstrated cardiac wall thickness, chamber size, and systolic function similar to other psittacine birds. On return to the colony, the parrot continued to be outwardly asymptomatic despite the dramatic conduction disturbances. Although cardiac arrhythmias, including second-degree AV block, have been widely reported in birds, the wide variation of nodal conductions, the intermittent nature, and an arrhythmia with a 7:1 second-degree AV block that spontaneously reverts to normal as seen in this case have not been well documented in parrots.

Fine-scale population genetic structure in a fission-fusion society.

Nonrandom patterns of mating and dispersal create fine-scale genetic structure in natural populations - especially of social mammals - with important evolutionary and conservation genetic consequences. Such structure is well-characterized for typical mammalian societies; that is, societies where social group composition is stable, dispersal is male-biased, and males form permanent breeding associations in just one or a few social groups over the course of their lives. However, genetic structure is not well understood for social mammals that differ from this pattern, including elephants. In elephant societies, social groups fission and fuse, and males never form permanent breeding associations with female groups. Here, we combine 33 years of behavioural observations with genetic information for 545 African elephants (Loxodonta africana), to investigate how mating and dispersal behaviours structure genetic variation between social groups and across age classes. We found that, like most social mammals, female matrilocality in elephants creates co-ancestry within core social groups and significant genetic differentiation between groups (Phi(ST) = 0.058). However, unlike typical social mammals, male elephants do not bias reproduction towards a limited subset of social groups, and instead breed randomly across the population. As a result, reproductively dominant males mediate gene flow between core groups, which creates cohorts of similar-aged paternal relatives across the population. Because poaching tends to eliminate the oldest elephants from populations, illegal hunting and poaching are likely to erode fine-scale genetic structure. We discuss our results and their evolutionary and conservation genetic implications in the context of other social mammals.

Behavioural inbreeding avoidance in wild African elephants.

The costs of inbreeding depression, as well as the opportunity costs of inbreeding avoidance, determine whether and which mechanisms of inbreeding avoidance evolve. In African elephants, sex-biased dispersal does not lead to the complete separation of male and female relatives, and so individuals may experience selection to recognize kin and avoid inbreeding. However, because estrous females are rare and male-male competition for mates is intense, the opportunity costs of inbreeding avoidance may be high, particularly for males. Here we combine 28 years of behavioural and demographic data on wild elephants with genotypes from 545 adult females, adult males, and calves in Amboseli National Park, Kenya, to test the hypothesis that elephants engage in sexual behaviour and reproduction with relatives less often than expected by chance. We found support for this hypothesis: males engaged in proportionally fewer sexual behaviours and sired proportionally fewer offspring with females that were natal family members or close genetic relatives (both maternal and paternal) than they did with nonkin. We discuss the relevance of these results for understanding the evolution of inbreeding avoidance and for elephant conservation.

Consuming broccoli does not induce genes associated with xenobiotic metabolism and cell cycle control in human gastric mucosa.

Epidemiological studies suggest that a diet rich in broccoli can reduce the risk of cancer at several sites. The anticarcinogenic activity has been largely attributed to the biological activity of sulforaphane (SF), the isothiocyanate derived from 4-methylsulphinylbutyl glucosinolate, which accumulates in broccoli. SF induces xenobiotic metabolizing genes in both cell cultures and animal models and induces genes associated with cell cycle arrest and apoptosis. However, it is not known whether these genes are induced in humans after consumption of broccoli. Sixteen subjects were recruited into a randomized, 3-phase crossover dietary trial of standard broccoli, high glucosinolate broccoli, and water. Global changes in gene expression that occurred 6 h after consuming broccoli soups or water were quantified in gastric mucosal tissue, using Affymetrix whole genome microarrays (n = 4), and in selected genes by real-time RT-PCR in the other individuals. Consumption of high glucosinolate broccoli resulted in up-regulation of several xenobiotic metabolizing genes, including thioredoxin reductase, aldoketoreductases, and glutamate cysteine ligase modifier subunit, which have previously been reported to be induced in cell and animal models after exposure to SF. Only 1 such gene was significantly up-regulated after consumption of standard broccoli. The consequences of these results in relation to the potential anticarcinogenic action of broccoli are discussed.

Forecasting Sclerotinia Disease on Lettuce: A Predictive Model for Carpogenic Germination of Sclerotinia sclerotiorum Sclerotia.

ABSTRACT A predictive model for production of apothecia by carpogenic germination of sclerotia is presented for Sclerotinia sclerotiorum. The model is based on the assumption that a conditioning phase must be completed before a subsequent germination phase can occur. Experiments involving transfer of sclerotia from one temperature regime to another allowed temperature-dependent rates to be derived for conditioning and germination for two S. sclerotiorum isolates. Although the response of each isolate to temperature was slightly different, sclerotia were fully conditioned after 2 to 6 days at 5 degrees C in soil but took up to 80 days at 15 degrees C. Subsequent germination took more than 200 days at 5 degrees C and 33 to 52 days at 20 degrees C. Upper temperature thresholds for conditioning and germination were 20 and 25 degrees C, respectively. A predictive model for production of apothecia derived from these data was successful in simulating the germination of multiple burials of sclerotia in the field when a soil water potential threshold of between -4.0 and -12.25 kilopascals (kPa) was imposed. The use of a germination model as part of a disease forecasting system for Sclerotinia disease in lettuce is discussed.

The Kansas Health Education Training Center: caring for the underserved.

The State of Kansas was awarded Health Education Training Centers (HETC) DHHS/HRSA, BrHPr Federal funds for the first time in 2002 through 2005. The three-year grant award was under the leadership of the Nurse-Midwifery Education faculty of the University of Kansas (KU) School of Nursing. Federal objectives included: 1. To facilitate interdisciplinary clinical training for health professions students from nursing, medicine, and allied health at targeted underserved communities. 2. To deliver culturally appropriate health care information to underprivileged populations at targeted underserved communities. 3. To conduct continuing education activities for community health workers and health professionals serving vulnerable urban and rural populations. 4. To recruit and mentor high school students from disadvantaged populations to pursue health professions. This three-year state-wide program addressed access to quality health care for underprivileged and underserved populations in select communities to improve health status, through community-based care that is culturally competent, multidisciplinary, sensitive to needs of special populations and augmented by health professions students. This article provides a review of two selected community-based interventions in Sedgwick and Wyandotte counties (addressing BrHPr federal objectives one and two) implemented to enhance care for immigrant pregnant women and underserved patients diagnosed with diabetes, respectively. Also included is a review of literature related to the efficacy and utilization of community based health workers (CHWs) in caring for vulnerable populations.

Glutathione S-transferase M1 polymorphism and metabolism of sulforaphane from standard and high-glucosinolate broccoli.

BACKGROUND: Broccoli consumption is associated with a reduction in the risk of cancer, particularly in persons with a functional glutathione S-transferase M1 allele, as opposed rotrose whose GSTM1 gene has been deleted. Sulforaphane, the major isothiocyanate derived from 4-methylsulfinylbutyl glucosinolate, is thought to be the main agent conferring protection. OBJECTIVE: We compared sulforaphane metabolism in GSTM1-null and GSTM1-positive subjects after they consumed standard broccoli and high-glucosinolate broccoli (super broccoli). DESIGN: Sixteen subjects were recruited into a randomized, 3-phase crossover dietary trial of standard broccoli, super broccoli, and water. Liquid chromatography linked to tandem mass spectrometry was used to quantify sulforaphane and its thiol conjugates in plasma and urine. RESULTS: GSTM1-null subjects had slightly higher, but statistically significant, areas under the curve for sulforaphane metabolite concentrations in plasma, a greater rate of urinary excretion of sulforaphane metabolites during the first 6 h after broccoli consumption, and a higher percentage of sulforaphane excretion 24 h after ingestion than did GSTM1-positive subjects. Consumption of high-glucosinolate broccoli led to a 3-fold greater increase in the areas under the curve and maximum concentrations of sulforaphane metabolites in plasma, a greater rate of urinary excretion of sulforaphane metabolites during the first 6 h after consumption, and a lower percentage of sulforaphane excretion after its ingestion than did the consumption of standard broccoli. CONCLUSIONS: GSTM1 genotypes have a significant effect on the metabolism of sulforaphane derived from standard or high-glucosinolate broccoli. It is possible that the difference in metabolism may explain the greater protection that GSTM1-positive persons gain from consuming broccoli. The potential consequences of consuming glucosinolate-enriched broccoli for GSTM1-null and -positive persons are discussed.

Transcriptome analysis of human colon Caco-2 cells exposed to sulforaphane.

Sulforaphane (SF), a dietary phytochemical obtained from broccoli, has been implicated in several physiological processes consistent with anticarcinogenic activity, including enhanced xenobiotic metabolism, cell cycle arrest, and apoptosis. In this study, we report changes in global gene expression in Caco-2 cells exposed to physiologically appropriate concentrations of SF, through the use of replicated Affymetrix array and RT-PCR experiments. After exposure to 50 micromol/L SF, 106 genes exhibited a >2-fold increase in expression and 63 genes exhibited a >2-fold decrease in expression. There were fewer changes in gene expression at lower SF concentrations. The majority of these genes had not previously been shown to be modulated by SF, suggesting novel mechanisms of possible anticarcinogenic activity, including induction of differentiation and modulation of fatty acid metabolism. The changes in the expression of 10 of these genes, together with 4 additional genes of biological interest, were further quantified in independent studies with RT-PCR. These genes include several that have recently become associated with carcinogenesis, such as Krüppel-like factor (KLF)4, a gut-enriched transcription factor associated with induction of differentiation and reduction in cellular proliferation; DNA (cytosine-5-)-methyltransferase 1, associated with methylation; and alpha-methylacyl-CoA racemase (AMACR), a marker associated with the development of colon and prostate cancer. The expression of 5 of these genes [caudal type homeo box transcription factor 2 (CDX-2), KLF4, KLF5, cyclin-dependent kinase inhibitor 1A (p21), and AMACR] was additionally studied after in vitro exposure to SF of surgically resected healthy and cancerous colon tissue from each of 3 patients. The study suggests the complex effects that SF has on gene expression and highlights several potential mechanisms by which the consumption of broccoli may reduce the risk of carcinogenesis.

Phosphate enema toxicosis in a pygmy goat wether.

Phosphate enema toxicity was diagnosed in a 7-month-old, castrated male, pygmy goat. On presentation, clinical findings included mild depression, tachycardia, tachypnea, rumen stasis, muscle tremors, hypocalcemia, hypokalemia, hypochloremia, hyperphosphatemia, azotemia, and metabolic acidosis. Fluid diuresis and parenteral antimicrobial therapy resulted in recovery after 3 d of treatment.

Forecasting sclerotinia disease on lettuce: toward developing a prediction model for carpogenic germination of sclerotia.

ABSTRACT The feasibility of developing a forecasting system for carpogenic germination of Sclerotinia sclerotiorum sclerotia was investigated in the laboratory by determining key relationships among temperature, soil water potential, and carpogenic germination for sclerotia of two S. sclerotiorum isolates. Germination of multiple burials of sclerotia to produce apothecia also was assessed in the field with concurrent recording of environmental data to examine patterns of germination under different fluctuating conditions. Carpogenic germination of sclerotia occurred between 5 and 25 degrees C but only for soil water potentials of >/=-100 kPa for both S. sclerotiorum isolates. Little or no germination occurred at 26 or 29 degrees C. At optimum temperatures of 15 to 20 degrees C, sclerotia buried in soil and placed in illuminated growth cabinets produced stipes after 20 to 27 days and apothecia after 27 to 34 days. Temperature, therefore, had a significant effect on both the rate of germination of sclerotia and the final number germinated. Rate of germination was correlated positively with temperature and final number of sclerotia germinated was related to temperature according to a probit model. Thermal time analysis of field data with constraints for temperature and water potential showed that the mean degree days to 10% germination of sclerotia in 2000 and 2001 was 285 and 279, respecttively, and generally was a good predictor of the observed appearance of apothecia. Neither thermal time nor relationships established in the laboratory could account for a decline in final percentage of germination for sclerotia buried from mid-May compared with earlier burials. Exposure to high temperatures may explain this effect. This, and other factors, require investigation before relationships derived in the laboratory or thermal time can be incorporated into a forecasting system for carpogenic germination.

Contrasting emotional approach coping with passive coping for chronic myofascial pain.

Passive or emotion-focused coping strategies are typically related to worse pain and adjustment among chronic pain patients. Emotional approach coping (EAC), however, is a type of emotion-focused coping that appears to be adaptive in some nonpain populations but has not yet been examined in a chronic pain population. In a sample of 80 patients (75% women, M = 48.67 years of age) with chronic myofascial pain, we contrasted how EAC (assessed with the Emotional Approach Coping Scale) and 5 passive pain-coping strategies (assessed with the Vanderbilt Multidimensional Pain Coping Inventory (VMPCI)) were related to sensory and affective pain, physical impairment, and depression. Passive coping strategies were positively correlated with one another, but EAC was inversely correlated with most of them. The VMPCI passive strategies were substantially positively related to negative affect, whereas EAC was inversely related to negative affect. Controlling for potentially confounding demographics, higher EAC was related to less affective pain and depression, even after controlling for negative affect. Using passive coping strategies, in contrast, was associated with more pain, impairment, and depression, although these relations were greatly attenuated after controlling for negative affect. When considered simultaneously, EAC, but not passive coping, was related to affective pain, and both EAC and passive coping were significant correlates of depression, although in opposite directions. In secondary analyses, we found that EAC was related to less pain (particularly sensory) among men and to less depression among women. Unlike the use of passive pain-coping strategies, which are associated with worse pain and adjustment, the use of EAC (emotional processing and emotional expression) with chronic pain is associated with less pain and depression. This suggests that some emotion-focused types of pain coping may be adaptive, and it highlights the need to assess emotional coping processes that are not confounded with distress or dysfunction.

The relationship of alexithymia to pain severity and impairment among patients with chronic myofascial pain: comparisons with self-efficacy, catastrophizing, and depression.

OBJECTIVE: Alexithymia is elevated among patients with chronic pain, but the relationship of alexithymia to the severity of pain among chronic pain patients is unclear. Also, studies have rarely examined whether alexithymia is unique from other, more widely studied constructs in the chronic pain literature (i.e., self-efficacy, catastrophizing, and depression), and research has not examined how alexithymia relates to the sensory versus affective dimensions of pain. METHODS: Among 80 patients with chronic myofascial pain, we tested how alexithymia (Toronto Alexithymia Scale-20) was related to three competing constructs--self-efficacy, catastrophizing, and depression--and to the sensory and affective dimensions of pain as well as physical impairment. We then determined whether alexithymia remained correlated with pain and impairment when tested simultaneously with each of the three competing constructs. RESULTS: Analyses controlled for patients' sex, age, marital status, and duration of pain. Alexithymia was moderately correlated with less self-efficacy and greater catastrophizing, and substantially correlated with greater depression. Alexithymia was positively related to both affective pain and physical impairment, but was unrelated to sensory pain, whereas all three of the competing constructs were related to both types of pain as well as physical impairment. Regression analyses indicated that alexithymia remained a significant and independent correlate of affective pain severity while controlling for either self-efficacy or catastrophizing, but depression accounted for alexithymia's relationship with affective pain. Also, alexithymia was no longer related to physical impairment, after controlling for any of the other three constructs. CONCLUSION: Although alexithymia is not related to the sensory component of pain, it is correlated positively with the affective or unpleasantness component of pain, independent of self-efficacy or catastrophizing. The emotional regulation deficits of alexithymia may lead to depression, which appears to mediate alexithymia's relationship to affective pain. Alexithymia's relationship with physical impairment appears to be better accounted for by self-efficacy or catastrophizing.

Increased activity and expression of iNOS in human duodenal enterocytes from patients with celiac disease.

The activity of nitric oxide synthase (NOS) was assayed in enterocytes isolated from human duodenal biopsies to determine its role in celiac disease. Patients were categorized into groups with irritable bowel syndrome, iron-deficiency anemia, B(12)/folate deficiency, and treated and untreated celiac disease. Enterocytes isolated from all groups showed 1400W-inhibitable Ca2+-independent NOS activity with a pH level and temperature optimum of 9.4 and 37 degrees C, respectively. Western blotting showed that enterocytes expressed the inducible NOS protein and proteins with nitrated tyrosine residues, the latter being indicative of nitric oxide-driven peroxynitrite and/or free-radical damage. Endothelial NOS was seen only in the lamina propria. Patients with celiac disease had higher NOS activity than other patient groups. Treatment of the condition led to a fall in activity. Enzyme-linked immunosorbent assay demonstrated cGMP production by the enterocyte fraction, but cGMP levels did not correlate with NOS activity. These results suggest that inducible NOS is constitutively expressed in human duodenal enterocytes, is increased in patients with untreated celiac disease, and is partially corrected when such patients are treated. We found no evidence to support a role for nitric oxide in the formation of cGMP within the small intestine. Furthermore, we were unable to demonstrate a role for peroxynitrite/free radical damage in the pathophysiology of celiac disease.