MicroRNA Regulation of Airway Inflammation and Airway Smooth Muscle Function: Relevance to Asthma.

Genetic and environmental factors contribute to the onset and severity of asthma. Molecular pathogenesis of asthma involves changes in gene expression by a variety of inflammatory mediators acting in autocrine and paracrine fashion on effector cells of the airways. Transcriptional regulation of gene expression in resident airway cells has been studied extensively. However, protein function in a target cell can be regulated at multiple levels starting from transcription followed by post-transcription, translation, and post-translation steps. In this context, small noncoding RNAs known as microRNAs (miRNAs) have evolved as one of the key regulators of gene expression post-transcriptionally. Most importantly, miRNA expression is dynamic in nature and can be regulated by a variety of external stimuli. Altered expression of individual or a group of miRNAs is thought to contribute to human diseases. Recent studies have implicated differential expression of miRNAs in the lungs during inflammation. Most importantly, advanced biochemical and molecular tools could be used to manipulate miRNA expression thereby effecting functional changes in target cells and organ systems. This review summarizes the current understanding of miRNA in the regulation of airway function in health and disease, and highlights the potential clinical utility of mRNAs as biomarkers of airway diseases and as potential therapeutic targets.

Magnetic resonance imaging assessment of articular disc position in temporomandibular disorder subjects with various bite registrations.

Objective: This study aimed to evaluate the validity and reliability of three bite registrations on articular disc position in temporomandibular disorder patients using magnetic resonance imaging (MRI). Materials and Methods: Fifteen clinically symptomatic and orthodontically untreated temporomandibular disorder patients within the age range of 17-40 years (mean age: 28.5 years) were examined. Each patient was subjected to three bite registrations, namely maximum intercuspation, initial contact bite and Roth power centric bite, and evaluated with MRI. Results: On the right side, the mean vertical and horizontal measurement values of the point in the most posterior aspect of the posterior band of the articular disc in relation to horizontal reference line (HRL) and vertical reference line (VRL) in the sagittal view in the Roth power centric bite were lesser (2.720 ± 1.239 mm and 2.380 ± 1.185 mm, respectively), in comparison with the other two bites, and on the left side too, it was lesser in the Roth power centric bite (2.293 ± 0.979 mm and 2.360 ± 1.078 mm, respectively), when compared to the other two bites. Statistical analysis also showed the significance of Roth power centric bite over the other two bites. Conclusions: Favourable articular disc positional changes were observed in the Roth power centric bite followed by the initial contact bite and that maximum disc recapture was observed in most patients with the Roth power centric bite rather than in initial contact bite and maximum intercuspation positions. The Roth power centric bite could be assumed to be the ideal method for articulation and fabrication of gnathological splints for treating patients with temporomandibular disorders.

Comparison of essential drug list in a rural secondary care hospital in south India with Indian & World Health Organization list 2011.

OBJECTIVE: Fixed drug combinations are a major marketing strategy in India but it can compromise the rational use of medicines. In this study we compared the fixed drug combinations and dosage forms in the hospital pharmacy before and after introducing the essential drug list. We also compared the Hospital Essential Drug List (HEDL) 2011 with the World Health Organization (WHO) Essential Drug List (EDL) 2011 and the National Essential Drug List of India (NEDL) 2011. METHODS: The study was done in a secondary level care charity hospital at Anantapur, AP with a bed size of 315 and an average OP per day of 1200-1700 visits. We compared the three essential drug lists (HEDL, WHOEDL and NEDL) and the hospital drug list before introducing EDL. Drugs which were present in NEDL and not present in the HEDL were also screened. Microsoft excel was used to tabulate the results and for graphs. RESULTS: The number of medicines used in the hospital before and after the introduction of the HEDL was 1627 and 424 respectively. On comparison, WHOEDL 2011 have 350 and NEDL of India have 348 medicines. While preparing the HEDL, 46 double drug combinations decreased to 15 and 9 triple drug combinations decreased to 1. In the case of injections, 20 double drug combinations decreased to 6 and 1 triple drug combination increased to 2. The number of tablets, capsules, injections, syrups, powders and inhalers was reduced to almost half. The great reductions were in 51 ointments to 9, 69 drops to 5, 11 paste to 0, 21 solutions to 3 and 14 creams to 1. The dosage forms removed included elixirs, insulin pens, gums, paste, paints, gargles and mouthwashes. CONCLUSIONS: There was drastic reduction in the number of medicines and dosage forms when the HEDL was implemented. Many of the fixed drug combinations were also removed for improving the rational use of medicines. The WHO essential drug list 2011, national essential drug list of India 2011 and the hospital essential drug list 2011 were comparable with few exceptions.

Formation of gap junctions by treatment in vitro with potassium conductance blockers.

Gap junctions were regularly seen in thin sections of canine tracheal smooth muscle incubated in vitro. Their number was increased in tissued exposed in vitro to either of two potassium conductance blockers, tetraethylammonium (TEA) and 4-aminopyridine (4-AP), and at the same time the muscles became mechanically active, with spontaneous contractions. The presence of gap junctions in this smooth muscle may provide one basis for cell-to-cell coupling, and their increase after TEA- and 4-AP-treatment could account for a decreased junctional resistance between cells, contributing to a longer space constant. However, an increase in gap junctions was not sufficient to change the behavior of trachealis smooth muscle from multiunit to single-unit type. Gap junctions in increased numbers persisted after washout of 4-AP, which caused inhibition of spontaneous contractions, and despite inhibition of the contractile effects of 4-AP by atropine. The rapid induction of gap junction formation was not dependent on de novo synthesis of protein. The fact that the number of gap junctions can be increased by chemical agents has important implications for control of their formation and provides a tool for analysis fo their role in cell-to-cell coupling.

Subcellular localization of cyclic ADP-ribosyl cyclase and cyclic ADP-ribose hydrolase activities in porcine airway smooth muscle.

Recent studies have provided evidence for a role of cyclic ADP-ribose (cADPR) in the regulation of intracellular calcium in smooth muscles of the intestine, blood vessels and airways. We investigated the presence and subcellular localization of ADP-ribosyl cyclase, the enzyme that catalyzes the conversion of beta-NAD(+) to cADPR, and cADPR hydrolase, the enzyme that degrades cADPR to ADPR, in tracheal smooth muscle (TSM). Sucrose density fractionation of TSM crude membranes provided evidence that ADP-ribosyl cyclase and cADPR hydrolase activities were associated with a fraction enriched in 5'-nucleotidase activity, a plasma membrane marker enzyme, but not in a fraction enriched in either sarcoplasmic endoplasmic reticulum calcium ATPase or ryanodine receptor channels, both sarcoplasmic reticulum markers. The ADP-ribosyl cyclase and cADPR hydrolase activities comigrated at a molecular weight of approximately 40 kDa on SDS-PAGE. This comigration was confirmed by gel filtration chromatography. Investigation of kinetics yielded K(m) values of 30.4+/-1.5 and 695. 3+/-171.2 microM and V(max) values of 330.4+/-90 and 102.8+/-17.1 nmol/mg/h for ADP-ribosyl cyclase and cADPR hydrolase, respectively. These results suggest a possible role for cADPR as an endogenous modulator of [Ca(2+)](i) in porcine TSM cells.

Comparison of frictional resistance of esthetic and semi-esthetic self-ligating brackets.

AIM: The frictional resistance encountered during sliding mechanics has been well established in the orthodontic literature, and it consists of complex interactions between the bracket, archwire, and method of ligation the claim of reduced friction with self-ligating brackets is often cited as a primary advantage over conventional brackets. This study was done to compare and evaluate the frictional forces generated between fully esthetic brackets and semi-aesthetic self-ligating brackets, which are of passive form and SEM (scanning electron microscope) study of the Brackets after Frictional evaluation. MATERIALS AND METHODS: Two types of self-ligating esthetic brackets, Damon clear (Ormco) made of fully ceramic and Opal (Ultradent Products, USA) and, Two types of self-ligating semi-esthetic brackets, Clarity SL (3M Unitek) and Damon 3 (Ormco) both of which are made of ceramic with metal slot. Arch wires with different dimensions and quality 17 × 25, 19 × 25 Titanium Molybdenum Alloy (TMA) and 17 × 25, 19 × 25 stainless steel that came from plain strands of wire were used for frictional comparison test. The brackets used in this study had 0.022 × 0.028 inch slot. RESULTS: The statistical tests showed significantly smaller amount of kinetic frictional forces is generated by Damon 3 (semi-esthetic self-ligating brackets). For each wire used, Damon 3 displayed significantly lower frictional forces (P ≤ 0.05) than any of the self-ligating system, followed by Opal (fully esthetic self-ligating brackets) which generated smaller amount of frictional forces but relatively on the higher side when compared with Damon 3. Damon clear (fully esthetic self-ligating brackets) generated the maximum amount of kinetic forces with all types of wire dimensions and properties when compared to the other three types of self-ligating system. Clarity SL (semi-esthetic self-ligating brackets) generated smaller amount of frictional forces when compared with Damon clear and relatively higher amount of frictional forces when compared to Opal and Damon 3.

A new approach for evaluation of canine dento alveolar distraction using cone-beam computed tomography.

OBJECTIVE: The aim was to evaluate and plan the canine dento alveolar distractions (DADs) with the use of cone-beam computed tomography (CBCT). MATERIALS AND METHODS: 5 patients are requiring 10 canine DADs were selected for the study. A custom-made DAD distractor was fabricated for the study. CBCT scans were taken prior to and post thedistraction. DAD parameters such as Canine retraction, canine and molar rotation, molar anchor loss and level of the osteotomy cut above the canine was evaluated. RESULTS: Average canine retraction was 7.5 mm in 17 days, molar anchor loss was 0.5 mm, canine and molar rotations were 8° and 0.40° and thedistance of the osteotomy cut to the canine was1.93 mm. CONCLUSION: The CBCT can be used to accurately evaluate the canine DADtechnique.

Spatial and temporal aspects of ACh-induced [Ca2+]i oscillations in porcine tracheal smooth muscle.

This study evaluated the relationship between regional elevation in intracellular calcium concentration ([Ca2+]i) induced by acetylcholine (ACh) and the global cellular responses in porcine tracheal smooth muscle (TSM) cells. Regional (approximately 1.5 microm3) and global (whole cell) changes in [Ca2+]i were measured in fluo-3 loaded TSM cells using real-time confocal microscopy. Regional responses appeared as propagating [Ca2+]i oscillations whereas global responses reflected the spatiotemporal integration of these regional responses. Within a region, [Ca2+]i oscillations were 'biphasic' with initial higher frequencies, followed by slower steady-state oscillations. With increasing ACh concentration, the peak (maximum value relative to 0 nM) of regional [Ca2+]i oscillations remained relatively constant, whereas both frequency and propagation velocity increased. In contrast, the global spatiotemporal integration of the regional oscillatory responses appeared as a concentration-dependent increase in peak as well as mean cellular [Ca2+]i. We conclude that the significance of ACh-induced [Ca2+]i oscillations lies in the establishment of mean [Ca2+]i level for slower Ca2+-dependent physiological processes via modulation of oscillation frequency and propagation velocity.

Effects of salbutamol on intracellular calcium oscillations in porcine airway smooth muscle.

Relaxation of airway smooth muscle (ASM) by beta-adrenoceptor agonists involves reduction of intracellular Ca2+ concentration ([Ca2+]i). In porcine ASM cells, acetylcholine induces [Ca2+]i oscillations that display frequency modulation by agonist concentration and basal [Ca2+]i. We used real-time confocal microscopy to examine the effect of salbutamol (1 nM to 1 microM), a beta 2-adrenoceptor agonist, on [Ca2+]i oscillations in freshly dissociated porcine ASM cells. Salbutamol decreased the frequency of [Ca2+]i oscillations in a concentration-dependent fashion, completely inhibiting the oscillations at 1 microM. These effects were mimicked by a cell-permeant analog of adenosine 3',5'-cyclic monophosphate. The inhibitory effect of salbutamol was partially reversed by BAY K 8644. Salbutamol reduced [Ca2+]i even when sarcoplasmic reticulum (SR) Ca2+ reuptake and Ca2+ influx were blocked. Lanthanum blockade of Ca2+ efflux attenuated the inhibitory effect of salbutamol on [Ca2+]i. The [Ca2+]i response to caffeine was unaffected by salbutamol. On the basis of these results, we conclude that beta 2-adrenoceptor agonists have little effect on SR Ca2+ release in ASM cells but reduce [Ca2+]i by inhibiting Ca2+ influx through voltage-gated channels and by enhancing Ca2+ efflux.

Modulation of nitric oxide-dependent relaxation of pig tracheal smooth muscle by inhibitors of guanylyl cyclase and calcium activated potassium channels.

Nitric oxide is released from intrinsic nonadrenergic, noncholinergic (NANC) nerves of pig tracheal smooth muscle (TSM) in response to electrical field stimulation (EFS). In this study, we investigated the role of guanylyl cyclase in the NANC relaxation by using guanylyl cyclase inhibitors, LY83583 and methylene blue (MB). The role of large conductance calcium-activated potassium (KCa) channels in mediating NANC relaxation was studied by using inhibitors of this channel, charybdotoxin and iberiotoxin. In carbachol-contracted TSM strips, LY83583 (10-20 microM) and MB (10-100 microM) resulted in inhibition of EFS-induced relaxations at all frequencies studied. Relaxations induced by exogenous 8-Bromo-cyclic 3',5'-guanosine monophosphate (8-Br-cGMP) were unaffected by LY83583. The concentration-relaxation curves to isoproterenol, which acts by elevating adenosine-3',5'-cyclic monophosphate (cAMP), and the nitric oxide donors sodium nitroprusside (SNP) or S-nitroso-n-acetylpenicillamine (SNAP) were unaffected by LY83583. Both charybdotoxin (240 nM) and iberiotoxin (180 nM) attenuated relaxations induced by EFS and SNAP. The role of guanylyl cyclase activation in the relaxation to EFS of pig TSM is suggested by the sensitivity of the responses to MB. The selective inhibitory effects of LY83583 on relaxation to neurally released, but not to the nitric oxide donors, suggests that it acts by inhibiting nitric oxide release. The lack of any effect of LY83583 on isoproterenol- or guanosine, 3'5'-cyclic monophosphate (cGMP)-mediated relaxation suggests a mechanism that does not involve elevation of cAMP but lies proximal to the generation of cGMP. The susceptibility of the relaxations to EFS and SNAP to charybdotoxin and iberiotoxin suggests a mechanism that involves the selective activation of KCa channels in airway smooth muscle cells.

Isolation of nitric oxide synthase in the rat uterus.

Nitric oxide synthase (NOS) was isolated from the uterus of adult female rats by diethylaminoethyl (DEAE) column and further purified by 2',5'-ADP agarose. The chromatographic properties revealed two isoenzymes, NOS1 and NOS2. The molecular weights of both isoenzymes was approximately 155 Kd by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS PAGE) which was similar to NOS1 and NOS2 from rat brain. The enzymes required nicotinamide adenine dinucleotide phosphate (NADPH), Ca+2 and calmodulin as cofactors. However, in the absence of calmodulin and/or calcium NOS1 was reduced by approximately 96%, while NOS2 was reduced by approximately 70%. This maximal enzyme activity was similar for brain. These results demonstrate that two isoforms of NOS are present in the rat uterus.

Effects of Pasteurella haemolytica A1 leukotoxin on bovine neutrophils: degranulation and generation of oxygen-derived free radicals.

To further define the role of Pasteurella haemolytica A1 leukotoxin in the pathogenesis of bovine pneumonic pasteurellosis, its in vitro effects on bovine neutrophils were investigated. Leukotoxin-containing culture supernatant, from P. haemolytica, stimulated a neutrophil respiratory burst as measured by the generation of oxygen-derived free radicals O2- and H2O2. This effect was immediate because preincubation of neutrophils with the culture supernatant for 5 min or longer substantially suppressed this respiratory burst. This suppression was due to cytolysis of the neutrophils. Prolonged incubation of neutrophils with the same culture supernatant caused further cytolysis and degranulation. Heat-inactivated P. haemolytica culture supernatant that had lost its cytotoxic properties failed to stimulate respiratory burst by neutrophils. Furthermore, the respiratory burst, cytolysis and degranulation were abrogated only by leukotoxin-neutralizing monoclonal and polyclonal antibodies, but not by antibodies against the lipopolysaccharide. These studies show that the leukotoxin component in the culture supernatant was responsible for the generation of oxygen-derived free radicals and proteolytic enzymes from neutrophils which may participate in direct lung injury.

Pasteurella haemolytica A1 and bovine respiratory disease: pathogenesis.

The severe fibrinonecrotic pneumonia associated with pneumonic pasteurellosis usually results from colonization of the lower respiratory tract by Pasteurella haemolytica biotype A, serotype 1(A1). Despite recent research efforts, the authors lack a detailed understanding of the interactions and host response to P. haemolytica in the respiratory tract. The authors hypothesize that management and environmental stress factors or viral infection alters the upper respiratory tract (URT) epithelium allowing P. haemolytica to colonize the epithelium. Once the URT is colonized, large numbers of organisms enter the lung where they interact with alveolar macrophages. Endotoxin, released from the bacteria, crosses the alveolar wall where it activates pulmonary intravascular macrophages, endothelium, neutrophils, lymphocytes, platelets, complement, and Hageman factor leading to complex interactions of cells and mediators. It is the progression of this inflammatory response with neutrophil influx that is ultimately responsible for the pulmonary injury. Leukotoxin is a major virulence factor of P. haemolytica that allows it to survive by destroying phagocytic cells. At subcytolytic concentrations it may also enhance the inflammatory response by activating cells to produce mediators and release reactive oxygen metabolites and proteases.

Effects of intestinal ischemia on in vitro activity of adjacent jejunum in samples obtained from ponies.

OBJECTIVE: To determine whether intestinal ischemia would alter activity of the jejunum in vitro or alter staining characteristics for certain types of enteric neurotransmitters. SAMPLE POPULATION: Jejunal samples obtained from 10 ponies. PROCEDURE: Jejunal samples were obtained from locations proximal and distal to an area of small intestine made ischemic for 60 minutes. A portion of each sample was stained to detect substance P-like immunoreactivity, cholinergic and adrenergic neurons, and nitric oxide synthase. Portions of the remaining samples were suspended in muscle baths. General activity patterns (frequency and amplitude of contraction), responses to neuronal depolarization induced by electrical field stimulation (EFS), and responses to 1 microM norepinephrine (NE) were compared with responses of a normal section of small intestine obtained prior to ischemic insult. RESULTS: Staining patterns were not altered. Proximal and distal sections had evidence of decreased contractility, compared with the normal section. Contraction frequency also was decreased, and distal sections had lower contraction frequency than proximal sections. Relaxation responses were decreased in distal sections. Responses to NE differed significantly for distal and proximal sections, compared with normal sections. CONCLUSIONS AND CLINICAL RELEVANCE: Short-term ischemia can significantly affect adjacent bowel. Contractile and relaxation responses are impaired. Discrepancies in intestinal motility patterns and alterations in response to NE for sections proximal and distal to ischemic intestine could lead to clinical ileus or slowed transit of ingesta.

Evaluation of substance P as a neurotransmitter in equine jejunum.

OBJECTIVE: To determine whether substance P (SP) functions as a neurotransmitter in equine jejunum. SAMPLE POPULATION: Samples of jejunum obtained from horses that did not have lesions in the gastrointestinal tract. PROCEDURE: Jejunal smooth muscle strips, oriented in the plane of the circular or longitudinal muscle, were suspended isometrically in muscle baths. Neurotransmitter release was induced by electrical field stimulation (EFS) delivered at 2 intensities (30 and 70 V) and various frequencies on muscle strips that were maintained at low tension or were under contraction. A neurokinin-1 receptor blocker (CP-96,345) was added to baths prior to EFS to interrupt SP neurotransmission. Additionally, direct effects of SP on muscle strips were evaluated, and SP-like immunoreactivity was localized in intestinal tissues, using indirect immunofluorescence testing. RESULTS: Substance P contracted circularly and longitudinally oriented muscle strips. Prior treatment with CP-96,345 altered muscle responses to SP and EFS, suggesting that SP was released from depolarized myenteric neurons. Depending on orientation of muscle strips and stimulation variables used, CP-96,345 increased or decreased the contractile response to EFS. Substance P-like immunoreactivity was detected in the myenteric plexus and circular muscle layers. CONCLUSIONS AND CLINICAL RELEVANCE: Substance P appears to function as a neurotransmitter in equine jejunum. It apparently modulates smooth muscle contractility, depending on preexisting conditions. Effects of SP may be altered in some forms of intestinal dysfunction. Altering SP neurotransmission in the jejunum may provide a therapeutic option for motility disorders of horses that are unresponsive to adrenergic and cholinergic drugs.

Adrenergic, cholinergic, and nonadrenergic-noncholinergic intrinsic innervation of the jejunum in horses.

OBJECTIVE: To determine the major neurotransmitters that regulate contractile activity in the jejunum of horses. SAMPLE POPULATION: Jejunal specimens from 65 horses without gastrointestinal tract lesions. PROCEDURE: Jejunal smooth muscle strips, oriented in the plane of the circular or longitudinal muscular layer, were suspended isometrically in muscle baths. Neurotransmitter release was induced by electrical field stimulation (EFS) delivered at 30 and 70 V intensities and at various frequencies on muscle strips maintained at low or high muscle tone. To detect residual nonadrenergic-noncholinergic neurotransmission, the response of muscle to EFS in the presence of adrenergic and cholinergic blockade was compared with the response in the presence of tetrodotoxin. RESULTS: Atropine (ATR) decreased the contractile response of muscle strips to EFS under most conditions. However, ATR increased the contractile response of high-tone circular muscle. Adrenergic blockade generally increased the muscle responses to 30 V EFS and in high-tone longitudinal muscle but decreased contractile responses in high-tone circular muscle. Tetrodotoxin significantly altered the responses to EFS, compared with adrenergic and cholinergic receptor blockade. CONCLUSIONS: Acetylcholine and norepinephrine appear to be important neurotransmitters regulating smooth muscle contractility in the equine jejunum. They induce contraction and relaxation, respectively, in most muscle preparations, although they may cause opposite effects under certain conditions. In addition, nonadrenergic-noncholinergic excitatory and inhibitory influences were detected. CLINICAL RELEVANCE: Acetylcholine or norepinephrine release within the myenteric plexus of horses may alter gastrointestinal motility.

Evaluation of pulmonary function and analgesia in dogs after intercostal thoracotomy and use of morphine administered intramuscularly or intrapleurally and bupivacaine administered intrapleurally.

Eighteen dogs undergoing lateral thoracotomy at the left fifth intercostal space were randomly assigned to 1 of 3 postoperative analgesic treatment groups of 6 dogs each as follows: group A, morphine, 1.0 mg/kg of body weight, IM; group B, 0.5% bupivacaine, 1.5 mg/kg given interpleurally; and group C, morphine, 1.0 mg/kg given interpleurally. Heart rate, respiratory rate, arterial blood pressure, arterial blood gas tensions, alveolar-arterial oxygen differences, rectal temperature, pain score, and pulmonary mechanics were recorded hourly for the first 8 hours after surgery, and at postoperative hours 12, 24, and 48. These values were compared with preoperative (control) values for each dog. Serum morphine and cortisol concentrations were measured at 10, 20, and 30 minutes, hours 1 to 8, and 12 hours after treatment administration. All dogs had significant decreases in pHa, PaO2, and oxygen saturation of hemoglobin, and significant increases in PaCO2 and alveolar-arterial oxygen differences in the postoperative period, but these changes were less severe in group-B dogs. Decreases of 50% in lung compliance, and increases of 100 to 200% in work of breathing and of 185 to 383% in pulmonary resistance were observed in all dogs after surgery. Increases in work of breathing were lower, and returned to preoperative values earlier in group-B dogs. The inspiratory time-to-total respiratory time ratio was significantly higher in group-B dogs during postoperative hours 5 to 8, suggesting improved analgesia. Blood pressure was significantly lower in group-A dogs for the postoperative hour. Significant decreases in rectal temperature were observed in all dogs after surgery, and hypothermia was prolonged in dogs of groups A and C. Significant differences in pain score were not observed between treatment groups. Cortisol concentration was high in all dogs after anesthesia and surgery, and was significantly increased in group-B dogs at hours 4 and 8. Significant differences in serum morphine concentration between groups A and C were only observed 10 minutes after treatment administration. In general, significant differences in physiologic variables between groups A and C were not observed. Results of the study indicate that the anesthesia and thoracotomy are associated with significant alterations in pulmonary function and lung mechanics. Interpleurally administered bupivacaine appears to be associated with fewer blood gas alterations and earlier return to normal of certain pulmonary function values. Interpleural administration of morphine does not appear to provide any advantages, in terms of analgesia or pulmonary function, compared with its IM administration.

Nitric oxide: biological role and clinical uses.

Nitric oxide is a product of the conversion of L-arginine by the enzyme nitric oxide synthase. Nitric oxide is involved in a variety of physiological situations and is produced by many different cell types. It is involved in neurotransmission, maintenance of vascular smooth muscle tone, and cytotoxicity. Nitric oxide has been suggested to play an anti-inflammatory role by inhibiting the expression of the genes for inflammatory cytokines. The pathophysiological role of nitric oxide is also evident in a variety of diseases, including septic shock, asthma, reperfusion injury, etc. Nitric oxide, by stimulating the production of cyclic GMP, relaxes smooth muscles of the cardiovascular, respiratory, gastrointestinal, and genito-urinary systems. Recent studies have provided important information on the use of inhaled nitric oxide for the management of several diseases characterized by the presence of abnormal pulmonary vascular tone, such as persistent pulmonary hypertension of the newborn. This review addresses the biology and clinical uses of inhaled nitric oxide.

TNF-alpha induced CD38 expression in human airway smooth muscle cells: role of MAP kinases and transcription factors NF-kappaB and AP-1.

In human airway smooth muscle (HASM) cells, the expression of CD38, which synthesizes the calcium-mobilizing molecule cyclic ADP-ribose, is augmented by TNF-alpha, a cytokine implicated in asthma. We determined the role of mitogen-activated protein kinase (MAPK) in the activation of NF-kappaB and AP-1 in the regulation of CD38 expression in HASM cells. In HASM cells exposed to TNF-alpha (40 ng/ml), the inhibitors of extracellular signal-regulated kinase (ERK), p38, or c-Jun NH(2)-terminal kinase (JNK) decreased CD38 expression and ADP-ribosyl cyclase activity. Transfection of HASM cells with a dominant negative MEK decreased while a wild-type ERK increased TNF-alpha-induced CD38 expression. Electrophoretic mobility shift assays (EMSAs) were performed using nuclear proteins and consensus sequences to detect the effect of the MAPKs on NF-kappaB and AP-1 activation. EMSAs confirmed the role of p38 and JNK in mediating NF-kappaB and AP-1 activation. Transfection of a dominant negative c-Jun decreased TNF-alpha-induced CD38 expression indicating involvement of AP-1. Stability of TNF-alpha-induced CD38 transcripts were determined in the presence of MAPK inhibitors after arresting the transcription with actinomycin D. Transcript stability decreased in the presence of ERK and p38 MAPK, but not the JNK, inhibitors. These results indicate that regulation of CD38 expression through p38 and JNK MAPKs involves NF-kappaB and AP-1 activation, and ERK and p38 MAPKs also regulate expression posttranscriptionally through message stability.

Mapping of the binding site for Mannheimia haemolytica leukotoxin within bovine CD18.

To map the site involved in Mannheimia haemolytica leukotoxin (LktA) binding and biological activity within bovine CD18, bovine x human CD18 chimeric constructs were generated and coexpressed with bovine CD11a in K562 cells. Studies with the chimeric leukocyte function-associated antigen 1 transductants demonstrate that the site required for LktA binding and biological effects resides within amino acid residues 500 and 600 of the extracellular region of bovine CD18.