Integrated Transcriptome Sequencing Analysis Reveals Role of miR-138-5p/ TBL1X in Placenta from Gestational Diabetes Mellitus.

BACKGROUND/AIMS: The placenta has been suggested to play a crucial role in the pathology of gestational diabetes mellitus (GDM). Placenta-specific microRNAs (miRNAs) and the corresponding targeting genes involved in the pathology of GDM still remain to be elucidated. We aimed to identify the dysregulated miRNAs and the corresponding mRNA targets through an integrated miRNA and mRNA transcriptomic profiles analysis and investigate the role of differentially expressed miR-138-5p/TBL1X in GDM. METHODS: RNA sequencing (RNA-seq) was performed in 16 placentas from GDM and control group. Differentially expressed mRNAs and miRNAs in GDM were validated by quantitative PCR (qPCR). The wound healing assay and transwell migration assay were used to analyze cell migration ability. The cell proliferation was determined by CCK8 assay. Luciferase assay was used to confirm the direct binding of the targeted TBL1X with miR-138-5p. RESULTS: Totally, 281 mRNAs and 32 miRNAs were found to be differentially expressed in the GDM placentas. The biological relationships of the miRNA/mRNA pairs were related to cellular development and function and organ morphology. Among the aberrantly expressed molecules, we selected miR-138-5p from the bioinformatics analysis and found that miR-138-5p significantly inhibited the migration and proliferation of trophoblasts (HTR-8/SVneo) by targeting the 3'-UTR of TBL1X. Furthermore, the aberrant expression of miR-138-5p and TBL1X was significantly correlated with the weight of the placenta. CONCLUSION: We present the first integrative analysis of miRNA and mRNA expression profiles in GDM placenta and uncover a more detailed role for miR-138-5p, as well as its target TBL1X in the pathology of GDM.

Up-regulation of microRNA-367 promotes liver steatosis through repressing TBL1 in obese mice.

OBJECTIVE: Increasing evidence has demonstrated that microRNAs (miRNAs) play a critical role in the progression of metabolic disorders, including obesity-induced non-alcoholic fatty liver disease (NAFLD). In the present study, the expression and function of miR-367 were investigated. MATERIALS AND METHODS: C57BL/6 male mice aged 8 weeks were fed with a normal diet (ND) or high-fat-diet (HFD). The expression levels of miR-367 were analyzed in livers from two groups of mice by quantitative real-time PCR. Adenovirus containing miR-367 or negative control (NC) were constructed and administrated into C57BL/6 mice by tail vain injection. Potential targets of miR-367 were screened by miRWalk software and luciferase reporter assays. Mutagenesis analysis and Western blots were used to further determine the target of miR-367 in obese mice. RESULTS: We found that the expression of hepatic miR-367 was up-regulated in obese mice. In vitro and in vivo studies further demonstrated that overexpression of miR-367 mimics promoted triglyceride accumulation in cells and lean mice. At the molecular level, transducin beta-like 1 (TBL1), a coactivator of nuclear receptor peroxisome proliferator-activated receptor (PPAR) α, was identified as a direct target of miR-367. As a result, miR-367 overexpression resulted in an inhibition of fatty acid oxidation, leading to hepatosteatosis. CONCLUSIONS: Our data suggest miR-367/TBL1 regulatory pathway might have an important role for in the development of NAFLD.

betagamma-Transducin stimulates hydrolysis and synthesis of phosphatidylinositol 4,5-bisphosphate in bovine rod outer segment membranes.

T betagamma was shown to stimulate the hydrolysis and synthesis of PtdInsP2 in dark-adapted bovine retinal rod outer segments. In contrast, T alphaGDP blocked the effect of betagamma-transducin. It was also demonstrated that T betagamma was a stimulator of 32P incorporation into PtdInsP2 in ROS. These findings explain the modulating actions of GTP and light on PtdInsP2 hydrolysis and synthesis in ROS. The possible existence of cross-talk between the cGMP and phosphoinositide cascades in retinal rods was discussed.

The functional involvement of gut-expressed sweet taste receptors in glucose-stimulated secretion of glucagon-like peptide-1 (GLP-1) and peptide YY (PYY).

BACKGROUND & AIMS: Enteroendocrine cells are thought to directly sense nutrients via α-gustducin coupled taste receptors (originally identified in the oral epithelium) to modulate the secretion of glucagon-like peptide-1 (GLP-1) and peptide YY (PYY). METHODS: We measured mRNA expression of α-gustducin and T1R3 along the human gut; immunohistochemistry was used to confirm co-localization with GLP-1. Functional implication of sweet taste receptors in glucose-stimulated secretion of GLP-1 and PYY was determined by intragastric infusion of glucose with or without lactisole (a sweet taste receptor antagonist) in 16 healthy subjects. RESULTS: α-gustducin was expressed in a region-specific manner (predominantly in the proximal gut and less in ileum and colon, P < 0.05). Both, T1R3 and α-gustducin were co-localized with GLP-1. Glucose-stimulated secretions of GLP-1 (P = 0.026) and PYY (P = 0.034) were reduced by blocking sweet receptors with lactisole. CONCLUSION: Key proteins implicated in taste signaling are present in the human gut and co-localized with GLP-1 suggesting that these proteins are functionally linked to peptide secretion from enteroendocrine cells. Glucose-stimulated secretion of GLP-1 and PYY is reduced by a sweet taste antagonist, suggesting the functional involvement of gut-expressed sweet taste receptors in glucose-stimulated secretion of both peptides in humans.

Hepatic deficiency in transcriptional cofactor TBL1 promotes liver steatosis and hypertriglyceridemia.

The aberrant accumulation of lipids in the liver ("fatty liver") is tightly associated with several components of the metabolic syndrome, including type 2 diabetes, coronary heart disease, and atherosclerosis. Here we show that the impaired hepatic expression of transcriptional cofactor transducin beta-like (TBL) 1 represents a common feature of mono- and multigenic fatty liver mouse models. Indeed, the liver-specific ablation of TBL1 gene expression in healthy mice promoted hypertriglyceridemia and hepatic steatosis under both normal and high-fat dietary conditions. TBL1 deficiency resulted in inhibition of fatty acid oxidation due to impaired functional cooperation with its heterodimerization partner TBL-related (TBLR) 1 and the nuclear receptor peroxisome proliferator-activated receptor (PPAR) α. As TBL1 expression levels were found to also inversely correlate with liver fat content in human patients, the lack of hepatic TBL1/TBLR1 cofactor activity may represent a molecular rationale for hepatic steatosis in subjects with obesity and the metabolic syndrome.

[Inhibition of transducin by lithium: electrophysiological demonstration using the isolated retina]. / Inhibition de la transducine par le lithium: mise en évidence électrophysiologique sur rétine isolée.

Recently Avissar et al. have established that Li+ can inhibit G proteins implicated in brain function. In order to investigate the effect of Li+ on transducin, the evolution of the electroretinogram (ERG) recorded on isolated rat retina has been studied in presence of lithium. Results indicate that 10(-5) M Li+ had no effect on ERG while 10(-3) M Li+, which corresponds to therapeutic blood levels, significantly decreases ERG amplitude. This effect being nearly totally inhibited by cholera toxin (75 micrograms/l), it is concluded that Li+ acts on transducin and so inhibits the visual transduction process.

The phosphorylation state of phosducin determines its ability to block transducin subunit interactions and inhibit transducin binding to activated rhodopsin.

Heterotrimeric GTP-binding proteins (G-proteins) serve many different signal transduction pathways. Phosducin, a 28-kDa phosphoprotein, is expressed in a variety of mammalian cell types and blocks activation of several classes of G-proteins. Phosphorylation of phosducin by cyclic AMP-dependent protein kinase prevents phosducin-mediated inhibition of G-protein GTPase activity (Bauer, P. H., Müller, S., Puzicha, M., Pippig, S., Obermaier, B., Helmreich, E. J. M., and Lohse, M. J. (1992) Nature 358, 73-76). In retinal rods, phosducin inhibits transducin (Gt) activation by binding its beta gamma subunits. While rod phosducin is phosphorylated in the dark and dephosphorylated after illumination (Lee, R.-H., Brown, B. M., and Lolley, R. N. (1984) Biochemistry 23, 1972-1977), the significance of these reactions is still unclear. The data presented here permit a more precise characterization of phosducin function and the consequences of its phosphorylation. Dephosphophosducin blocked binding of the Gt alpha 1 subunit to activated rhodopsin in the presence of stoichiometric amounts of Gt beta gamma, whereas phosphophosducin did not. Surprisingly, the binding affinity of phosphophosducin for Gt beta gamma was not significantly reduced compared with the binding affinity of dephosphophosducin. However, the association of phosducin with Gt beta gamma in a size exclusion column matrix was dependent on the phosphorylation state of phosducin. Moreover, the ability of phosducin to compete with Gt alpha for binding to Gt beta gamma was also dependent on the phosphorylation state of phosducin. No interaction was found between phosducin and Gt alpha. These data indicate that phosducin decreases rod responsiveness by binding to the beta gamma subunits of Gt and preventing their interaction with Gt alpha, thereby inhibiting Gt alpha activation by the activated receptor. Moreover, phosphorylation of phosducin blocks its ability to compete with Gt alpha for binding to Gt beta gamma.

Inhibition of GTP-utilizing enzymes by tyrphostins.

Tyrphostins are a group of organic compounds which are widely used as a tool to specifically inhibit protein tyrosine kinases (Yaish, P., Gazit, A., Gilon, C., and levitzki A. (1988) Science 242, 933-935; Gazit, A., Yaish, P., Gilon, C., and Levitzki A. (1989) J. Med. Chem. 32, 2344-2352; Lyall, R. M., Zilberstein, A., Gazit, A., Gilon, C., Levitzki, A., and Schlessinger J. (1989) J. Biol. Chem. 264, 14503-14509; Osherov, N., Gazit, A., Gilon, C., and Levitzki, A. (1993) J. Biol. Chem. 268, 11134-11142). We report here that members of the tyrphostin family inhibit the GTPase activity of transducin and the enzymatic activities of other GTP-utilizing proteins in retinal rod outer segments, such as guanylyl cyclase or fructose-6-phosphate kinase. In contrast, ATP-utilizing enzymes such as hexokinase or rhodopsin kinase were not effected.

Active site-directed inactivation of constitutively active mutants of rhodopsin.

Recently, mutations of the active site Lys296 residue in rhodopsin (Lys296-->Glu and Lys296-->Met) have been found as the cause of disease in some patients with autosomal dominant retinitis pigmentosa. In vitro, these mutations result in constitutive activation of the protein. In an effort to develop a potential therapeutic agent for treatment of the disease, we have examined various amine derivatives of 11-cis- and 9-cis-retinal for ability to irreversibly inactivate a related constitutively active mutant, K296G. Three amines were prepared by reductive amination of retinal: 11-cis-retinylpropylamine, 11-cis-retinylamine, and 9-cis-retinylamine. All three compounds inactivated K296G, and the inactivation could not be reversed upon exposure to light. None of the compounds inactivated the wild-type protein. Although the amines were not effective on the naturally occurring retinitis pigmentosa mutants, presumably because of unfavorable steric interactions with the bulky Glu and Met side chains at position 296, the success with K296G makes it highly encouraging that this approach will evolve related compounds that are capable of inactivating the naturally occurring mutants as well.

The molecular origin of the inhibition of transducin activation in rhodopsin lacking the 9-methyl group of the retinal chromophore: a UV-Vis and FTIR spectroscopic study.

The formation of the active rhodopsin state metarhodopsin II (MII) is believed to be partially governed by specific steric constraints imposed onto the protein by the 9-methyl group of the retinal chromophore. We studied the properties of the synthetic pigment 9-demethyl rhodopsin (9dm-Rho), consisting of the rhodopsin apoprotein regenerated with synthetic retinal lacking the 9-methyl group, by UV-vis and Fourier transform infrared difference spectroscopy. Low activation rates of the visual G-protein transducin by the modified pigment reported in previous studies are shown to not be caused by the reduced activity of its MII state, but to be due to a dramatic equilibrium shift from MII to its immediate precursor, MI. The MII state of 9dm-Rho displays only a partial deprotonation of the retinal Schiff base, leading to the formation of two MII subspecies absorbing at 380 and 470 nm, both of which seem to be involved in transducin activation. The rate of MII formation is slowed by 2 orders of magnitude compared to rhodopsin. The dark state and the MI state of 9dm-Rho are distinctly different from their respective states in the native pigment, pointing to a more relaxed fit of the retinal chromophore in its binding pocket. The shifted equilibrium between MI and MII is therefore discussed in terms of an increased entropy of the 9dm-Rho MI state due to changed steric interactions.

Nucleotide exchange and cGMP phosphodiesterase activation by pertussis toxin inactivated transducin.

Transducin, the signal coupling protein of retinal rod photoreceptor cells, is one of a family of G proteins that can be inactivated by pertussis toxin. We have investigated the nature of this inactivation in order to determine (1) whether it requires the toxin-catalyzed transfer of ADP-ribose from NAD+ to cysteine-347 of the alpha subunit and (2) whether it involves locking the alpha subunit in the inactive conformation characteristic of its GDP-bound state, or is limited to disruption of binding to photoexcited rhodopsin (R*). Our results indicate that all observed effects of pertussis toxin treatment, including a shift in the electrophoretic mobility of transducin's alpha subunit and functional inactivation, require NAD+ and that the appearance of the shift parallels incorporation of ADP-ribose. We have also found that, apart from interactions with photoexcited rhodopsin, the functional properties of ADP-ribosylated transducin are essentially the same as those of unmodified transducin. Normal spontaneous nucleotide exchange kinetics and the ability to activate cGMP phosphodiesterase are preserved following quantitative ADP-ribosylation, as are the abilities to hydrolyze GTP, to bind to a dye affinity column, and to display enhanced fluorescence upon addition of Al3+ and F-. Thus, ADP-ribosylation merely blocks catalysis of transducin nucleotide exchange by R* and does not lock transducin in an inactive state.(ABSTRACT TRUNCATED AT 250 WORDS)

The first and second cytoplasmic loops of the G-protein receptor, rhodopsin, independently form beta-turns.

The cytoplasmic face of the transmembrane protein, rhodopsin, is made up of one carboxyl terminal and three cytoplasmic loops connecting six of the seven transmembrane helices. Neither the high-resolution, three-dimensional structure of this G-protein receptor nor any other cell surface receptor is known. In this work, the structures of peptides containing the amino acid sequence of the first and second cytoplasmic loops of rhodopsin have been determined. Both loops show ordered structures in solution. In both loops, the ends of the transmembrane helices unwind and form a beta-turn. The conformations of the two loops are remarkably similar, even though their sequences are not. These data suggest a structural motif for short loops in transmembrane proteins. The well-ordered structures of these loops, in the absence of the transmembrane helices, indicate that the primary sequences of these loops stabilize the beta-turn. These data further suggest that the loops may contribute to the folding of such membrane proteins during their synthesis and insertion into membranes.

Light inhibition of bovine retinal rod guanylyl cyclase mediated by beta gamma-transducin.

Photoreceptor guanylyl cyclase (ROS-GC), converting GTP into cGMP and pyrophosphate, is a key enzyme in the regulation of the visual transduction cascade. ROS-GC requires GC-activating proteins (GCAPs) and low free [Ca] for full activity. We found that when choline or potassium were the major cations present, light caused a 70% inhibition of stimulated ROS-GC in native unstripped membranes. In the presence of sodium ions, however, no inhibition was observed. ROS-GC activity of ROS membranes, stripped of transducin and other components, was not affected by light when reconstituted with GCAP1 only. However, when stripped ROS membranes were reconstituted with both GCAP1 and either transducin (T alpha beta gamma) or the T beta gamma-subunits, the inhibition of ROS-GC by light was restored. The T alpha-subunit alone was ineffective. These results suggest that under saturating light conditions, ROS-GC may be regulated by T beta gamma and cations, providing a possible mechanism of desensitization and light adaptation.

Inhibition of transducin activation and guanosine triphosphatase activity by aluminum ion.

Aluminum ion perturbs the activity of a number of physiologically important enzymes, including members of a family of guanine nucleotide-binding proteins (G-proteins). G-proteins couple cellular receptor proteins to a variety of effector enzymes (including adenylate cyclase, phospholipase C, and the rod photoreceptor phosphodiesterase). We show herein that subnanomolar concentrations of free aluminum ion, produced in a carefully defined and kinetically stable manner through the buffering of total aluminum at 0.1-1.0 mM with calculated ratios of chelating agents, inhibit both the receptor-mediated activation and the self-inactivating GTPase activity of the rod photoreceptor G-protein, Gv. In the presence of 4 X 10(-10) M free aluminum ion, GTPase activity is inhibited from about 25-60% as the magnesium ion concentration is reduced from 10(-3) to about 5 X 10(-5) M. The principal effect of aluminum ion upon Gv is to inhibit receptor catalyzed nucleotide exchange. Binding of the GTP analog 5'-guanylyl imidodiphosphate can be reduced by as much as 90% by aluminum ion following subsaturating rhodopsin stimulation. Aluminum ion can produce either competitive or mixed noncompetitive inhibition of rhodopsin-catalyzed Gv activation and GTPase activity, as a function of whether Gv undergoes single (competitive), or multiple (mixed noncompetitive) nucleotide exchanges. The rod photoreceptor phosphodiesterase is only slightly inhibited by similar aluminum ion activities. Light- and Gv-coupled phosphodiesterase activation exhibits both a lower maximum rate of cyclic guanosine monophosphate hydrolysis and a slower inactivation in the presence of aluminum ion activities from about 10(-12) - 10(-10) M. These data suggest that intracellular free aluminum ion concentrations in the subnanomolar range could markedly affect the ability of cells to transduce extracellular signals. Interestingly, the combination of Al3+ and F- to produce the fluoro-aluminate species (AlFx) also inhibits the GTPase of G-proteins, although the mechanism of inhibition (e.g. binding to the G-protein.Mg2+.GDP complex) is totally distinct from that observed for free Al3+ and the overall effect on signal transduction (e.g. enhanced signal amplification) is in complete opposition to that observed for free Al3+.

Mechanism of inhibition of transducin guanosine triphosphatase activity by vanadate.

The visual excitation system of the retinal rod outer segments and the hormone-sensitive adenylate cyclase complex are regulated through guanine nucleotide-binding proteins, transducin in the former and inhibitory and stimulatory regulatory components, Gi and Gs, in the latter. These proteins are functionally and structurally similar; all are heterotrimers composed of alpha, beta, and gamma subunits and exhibit guanosine triphosphatase activity stimulated by light-activated rhodopsin or the agonist-receptor complex. Adenylate cyclase can be stimulated by vanadate, which, like NaF, probably acts through Gs. Effects of vanadate on the function of a guanine nucleotide-binding protein were investigated in a reconstituted model system consisting of purified transducin subunits (T alpha, T beta gamma) and rhodopsin in phosphatidylcholine vesicles. Vanadate (decameric) inhibited [3H]GTP binding to T alpha and noncompetitively inhibited GTP hydrolysis in a concentration-dependent manner with maximal inhibition of approximately 90% at 3-5 mM. Vanadate also inhibited release of bound GDP but did not affect the rate of hydrolysis of bound GTP (single turnover rate), indicating that vanadate did not interfere with the intrinsic GTPase activity of T alpha. Binding of T alpha to rhodopsin and the ADP-ribosylation of T alpha by pertussis toxin, both of which are enhanced in the presence of T beta gamma, were inhibited by vanadate. These findings are consistent with the conclusion that vanadate can cause the dissociation of T alpha from T beta gamma, resulting in the inhibition of GDP-GTP exchange and thereby GTP hydrolysis. Adenylate cyclase activation could result from a similar effect of vanadate on Gs.

On the mechanism of the inhibition of transducin function by farnesylcysteine analogs.

The gamma subunits of heterotrimeric G proteins are isoprenylated/methylated on their carboxy termini. The photoreceptor G protein, transducin, is farnesylated/methylated at this position. Since the isoprenyl group is required for G protein function, it is of great interest to determine the mechanism by which the farnesyl group of Tgamma interacts with the other transducin subunits and/or the activated photoreceptor, rhodopsin. Farnesylcysteine derivatives (N-acetyl-S-farnesyl-L-cysteine and farnesylated peptides) have been previously shown to have effects on transducin activity at high concentrations. Here, an extensive survey is done of farnesylcysteine analogs and other lipid molecules, which are tested for their ability to inhibit GTP/GDP exchange in transducin catalyzed by photolyzed rhodopsin. These studies are carried out to determine the nature of the inhibition process. While it does not appear that these molecules exhibit the specificity which would characterize a ligand-receptor type mechanism, the results suggest that these compounds are not acting in a nonspecific detergent-like manner either. The most likely mode of action of famesylcysteine analogs is that they interfere with the lipid-lipid based association of Talpha and Tbetagamma through the lipid modifications present on each subunit.

Blocking taste receptor activation of gustducin inhibits gustatory responses to bitter compounds.

Gustducin, a transducin-like guanine nucleotide-binding regulatory protein (G protein), and transducin are expressed in taste receptor cells where they are thought to mediate taste transduction. Gustducin and transducin are activated in the presence of bovine taste membranes by several compounds that humans perceive to be bitter. We have monitored this activation with an in vitro assay to identify compounds that inhibited taste receptor activation of transducin by bitter tastants: AMP and chemically related compounds inhibited in vitro responses to several bitter compounds (e.g., denatonium, quinine, strychnine, and atropine). AMP also inhibited behavioral and electrophysiological responses of mice to bitter tastants, but not to NaCl, HCl, or sucrose. GMP, although chemically similar to AMP, inhibited neither the bitter-responsive taste receptor activation of transducin nor the gustatory responses of mice to bitter compounds. AMP and certain related compounds may bind to bitter-responsive taste receptors or interfere with receptor-G protein coupling to serve as naturally occurring taste modifiers.

Effect of light on phosphatidate phosphohydrolase activity of retina rod outer segments: the role of transducin.

The aim of the present paper is to evaluate the modulation of phosphatidate phosphohydrolase (PAPase) and diacylglyceride lipase (DGL) activities in bovine rod outer segment (ROS) under dark and light conditions and to evaluate the role of transducin (T) in this phenomenon. In dark-adapted ROS membranes exposed to light, PAPase activity is inhibited by 20% with respect to the activity found under dark conditions. To determine whether the retinal G protein, T, participates in the regulation of PAPase activity in these membranes, the effects of GTPgammaS and GDPbetaS on enzyme activity were examined. Under dark conditions in the presence of GTPgammaS, which stabilizes T in its active form (Talpha + Tbetagamma), enzyme activity was inhibited and approached control values under light conditions. GDPbetaS, on the other hand, which stabilizes the inactive state of T (Talphabetagamma), stimulated PAPase activity by 36% with respect to control light conditions. ADP-ribosylation by cholera and pertussis toxin was also studied. In ADP-rybosilated ROS membranes with pertussis toxin under dark conditions, PAPase activity was 36% higher than the activity found under control light conditions. ADP-ribosylation by CTx, on the other hand, inhibited PAPase activity by 22%, with respect to dark control conditions, mimicking light effect. The effects of GTPgammaS and GDPbetaS and conditions of ADP-ribosylation by PTx and CTx on DGL activity were similar to those of PAPase activities. Based on NEM sensitivity we have also demonstrated that the PAPase present in ROS is the PAP 2 isoform. Our findings therefore suggest that light inhibition of PAP 2 in ROS is a transducin-mediated mechanism.

The effect of anti-integrin monoclonal antibodies on antigen-induced pulmonary inflammation in allergic rabbits.

The integrin adhesion molecules are involved in the recruitment and activation of inflammatory cells at sites of inflammation in a variety of diseases. In the present study, we have investigated the effects of blocking monoclonal antibodies (mAbs) directed against CD49d (alpha(4) integrin), CD18 (beta(2) integrin) and the alpha sub-units of beta(2) integrin CD11a (LFA-1 integrin) and CD11b (Mac-1 integrin), on antigen (Ag)-induced acute bronchoconstriction and cellular recruitment in allergic rabbits in vivo. Inhaled Ag (Alternaria tenuis) challenge of neonatally sensitised rabbits caused an acute bronchoconstriction demonstrated by an increase in lung resistance (R(L)) and decrease in dynamic compliance (C(dyn)) and pulmonary inflammation characterised by an increase in bronchoalveolar lavage (BAL) inflammatory cells, particularly eosinophils, 24 h after challenge. Pre-treatment with the anti-CD49d mAb (Max-68P), significantly inhibited the Ag-induced acute bronchoconstriction in terms of R(L) and (C(dyn)). Treatment with the other anti-integrin mAbs had no effect on the acute bronchoconstriction after inhaled Ag challenge.Pre-treatment with the anti-integrin mAbs had differential effects in blocking the recruitment of inflammatory cells 24 h after inhaled Ag in the allergic rabbits. The data show that in the allergic rabbit model of asthma, VLA-4 (CD49d/CD29) only, is involved in the acute bronchoconstriction, suggesting an involvement of mast cell degranulation. Furthermore, eosinophil recruitment and activation appears to be mediated by a combination of VLA-4 (CD49d/CD29) and LFA-1 (CD18/CD11a). However in contrast, lymphocyte recruitment appears to be mediated by a combination of LFA-1 (CD18/CD11a) and Mac-1 (CD18/CD11b).