197,249 research outputs found
Experimental Pharmacotherapy for Dry Eye Disease: A Review
No full textMonica Baiula, Santi Spampinato Department of Pharmacy and Biotechnology, University of Bologna, Bologna, ItalyCorrespondence: Santi SpampinatoDepartment of Pharmacy and Biotechnology, University of Bologna, via Irnerio 48, Bologna, ItalyTel +39 0512091851Email [email protected]: Dry eye disease (DED) is a complex multifactorial disease showing heterogenous symptoms, including dryness, photophobia, ocular discomfort, irritation and burning but also pain. These symptoms can affect visual function leading to restrictions in daily life activities and reduction in work productivity with a consequently high impact on quality of life. Several pathological mechanisms contribute to the disease: evaporative water loss leads to impairment and loss of tear homeostasis inducing either directly or indirectly to inflammation, in a self-perpetuating vicious cycle. Dysregulated ocular immune responses result in ocular surface damage, which further contributes to DED pathogenesis. Currently, DED treatment is based on a flexible stepwise approach to identify the most beneficial intervention. Although most of the available treatments may control to a certain extent some signs and symptoms of DED, they show significant limitations and do not completely address the needs of patients suffering from DED. This review provides an overview of the emerging experimental therapies for DED. Several promising therapeutic strategies are under development with the aim of dampening inflammation and restoring the homeostasis of the ocular surface microenvironment. Results from early phase clinical trials, testing the effects of EnaC blockers, TRPM8 agonist or mesenchymal stem cells in DED patients, are especially awaited to demonstrate their therapeutic value for the treatment of DED. Moreover, the most advanced experimental strategies in the pipeline for DED, tivanisiran, IL-1R antagonist EBI-005 and SkQ1, are being tested in Phase III clinical trials, still ongoing. Nevertheless, although promising results, further studies are still needed to confirm efficacy and safety of the new emerging therapies for DED.Keywords: dry eye disease, inflammation, tivanisiran, IL-1R antagonist, SkQ
Overview of genetic analysis of human opioid receptors
No full textThe human μ-opioid receptor gene (OPRM1), due to its genetic and structural variation, has been a target of interest in several pharmacogenetic studies. The μ-opioid receptor (MOR), encoded by OPRM1, contributes to regulate the analgesic response to pain and also controls the rewarding effects of many drugs of abuse, including opioids, nicotine, and alcohol. Genetic polymorphisms of opioid receptors are candidates for the variability of clinical opioid effects. The non-synonymous polymorphism A118G of the OPRM1 has been repeatedly associated with the efficacy of opioid treatments for pain and various types of dependence. Genetic analysis of human opioid receptors has evidenced the presence of numerous polymorphisms either in exonic or in intronic sequences as well as the presence of synonymous coding variants that may have important effects on transcription, mRNA stability, and splicing, thus affecting gene function despite not directly disrupting any specific residue. Genotyping of opioid receptors is still in its infancy and a relevant progress in this field can be achieved by using advanced gene sequencing techniques described in this review that allow the researchers to obtain vast quantities of data on human genomes and transcriptomes in a brief period of time and with affordable costs
PKC modulation of mu-opioid receptor gene (OPRM1) transcription is influenced by the transcription factor REST
No full textAndrea Bedini, Monica Baiula and Santi Spampinato. Department of Pharmacology, University of
Bologna, Irnerio 48 – 40126, Italy. RE1 Silencing Transcription Factor (REST) represses transcription
of neuronal genes in non-neuronal mature cells and in neuronal stem cells, where it is expressed at
high levels. REST plays a pivotal role in regulating neural cell fate determination by restricting
neuronal gene expression to post-mitotic neurons1
. Protein kinase C (PKC) induces different signaling
pathways, some of which are related to opioid receptors: PKC activation, in fact, down-regulates
endogenous human mu-opioid receptor (MOPr) mRNA in SH-SY5Y neuroblastoma cells2
whereas
enhances mu-opioid receptor gene (OPRM1) transcription in the same cells when OPRM1 promoter
fragment from nucleotide -2624 to nucleotide -165 is considered3
. These observations arise the
hypothesis that there might be transcription factors binding the OPRM1 promoter region closest to
ATG and down-regulating OPRM1 transcription: as REST has been shown to bind human OPRM1
promoter at nucleotides from -9 to +12 and we have previously observed that phorbol 12-myristate 13-
acetate (PMA) induces REST, we wondered whether REST is involved in PKC-mediated MOPr downregulation. Therefore, three reporter plasmids bearing OPRM1 promoter fragments -1672/+64, -1672/-
10 and -1672/-254, respectively, have been cloned and transfected in SH-SY5Y cells, which express
REST, and in PC-12, which lack of REST expression4
. In SH-SY5Y cells, PMA-induced PKC
determined transcriptional activation of the OPRM1 promoter fragments lacking the RE1 site for REST
(-1672/-10 and -1672/-254) and transcriptional repression of the OPRM1 promoter fragment bearing
the REST target sequence (-1672/+64). In PC-12 cells PMA-induced PKC determined the
transcriptional activation of all the OPRM1 promoter fragments. Then endogenous MOPr transcripts
were evaluated by real-time PCR: in SH-SY5Y cells, PMA down-regulated MOPr mRNA levels
whereas REST knockdown by specific antisense oligonucleotide prevented OPRM1 transcription
down-regulation by PMA. Similarly, retinoic acid differentiation of SH-SY5Y cells down-regulated
REST expression, thus preventing PMA-induced MOPr down-regulation. Taken together our results
show for the first time that PMA down-regulation of OPRM1 transcription in neurons involves REST,
which can be the critical switch that determines different PMA-mediated effects according to its
expression levels. Ref.: (1) Di Toro et al., European journal of neuroscience, 2005, 21, 46-58 ; (2) Gies
et al., Anestesiology, 1997, 87, 1127-38; (3) Börner et al., Molecular Pharmacology, 2002, 61, 800-5 ;
(4) Bedini et al., Journal of Neurochemistry, 200
Contribution of alpha(4)beta(1) integrin to the antiallergic effect of levocabastine.
No full textBiochem Pharmacol. 2008 Sep 15;76(6):751-62. Epub 2008 Jul 15.
Contribution of alpha4beta1 integrin to the antiallergic effect of levocabastine.
Qasem AR, Bucolo C, Baiula M, Spartà A, Govoni P, Bedini A, Fascì D, Spampinato S.
Source
Department of Medicine, Health Science Campus, University of Toledo, OH, USA.
Abstract
Levocabastine is an antiallergic drug acting as a histamine H1-receptor antagonist. In allergic conjunctivitis (AC), it may also antagonize up-regulation of the intercellular adhesion molecule-1 (ICAM-1) expressed on epithelial conjunctival cells. However, little is known about its effects on eosinophils, important effector cells in AC. The adhesion molecule integrin alpha(4)beta(1) is expressed in eosinophils; it interacts with the vascular cell adhesion molecule-1 (VCAM-1) and fibronectin (FN) in vascular endothelial cells and contributes to eosinophil activation and infiltration in AC. This study provides evidence that in a scintillation proximity assay levocabastine (IC(50) 406 microM), but not the first-generation antihistamine chlorpheniramine, displaced (125)I-FN binding to human integrin alpha(4)beta(1) and, in flow cytometry analysis, levocabastine antagonized the binding of a primary antibody to integrin alpha(4) expressed on the Jurkat cell surface. Levocabastine, but not chlorpheniramine, binds the alpha(4)beta(1) integrin and prevents eosinophil adhesion to VCAM-1, FN or human umbilical vascular endothelial cells (HUVEC) in vitro. Similarly, levocabastine affects alpha(L)beta(2)/ICAM-1-mediated adhesion of Jurkat cells. In a model of AC levocabastine eye drops reduced the clinical aspects of the late-phase reaction and the conjunctival expression of alpha(4)beta(1) integrin by reducing infiltrated eosinophils. We propose that blockade of integrin-mediated cell adhesion might be a target of the antiallergic action of levocabastine and may play a role in preventing eosinophil adhesion and infiltration in AC
Agonist-Regulated Internalization and desensitization of the Human Nociceptin Receptor Expressed in CHO Cells
No full textIn this study we examined agonist-induced internalization of the cloned human nociceptin receptor (hNOP) expressed in CHO-K1 cells. Internalization was proven by receptor binding assay on viable cells and confocal microscopy. The agonists nociceptin/orphanin FQ (NC), NC-NH2, NC(1-13)-NH2, [(pF)Phe4]NC-NH2 and RO 64-6198 promote a rapid, concentration-dependent internalization of the hNOP receptor. Under the same conditions, [Phe1,ψ(CH2NH)Gly2]NC(1-13)-NH2 and [Phe1, ψ(CH2NH)Gly2,Arg14,Lys15]NC(1-13)-NH2 failed to induce significant, concentration-dependent NOP receptor endocytosis; even when present at high concentrations (up to 1 mM) they promoted only an approximately 25-30% internalization of hNOP receptors. We also investigated hNOP receptor desensitization upon agonist challenge: ligand efficacy to inhibit forskolin-stimulated cAMP production. After 1 h exposure to NC, NC-NH2, NC(1-13)-NH2, [(pF)Phe4]NC-NH2 and RO 64-6198 (5 μM) ≈20 to 30% of receptor desensitization was observed. Moreover, we found that the blockade of hNOP receptor recycling by monensin would cause a more prolonged and relevant desensitization of this receptor. The non-internalizing agonists [Phe1,ψ(CH2NH)Gly2]NC(1-13)-NH2 and [Phe1, ψ(CH2NH)Gly2,Arg14,Lys15]NC(1-13)-NH2 (100 μM) resulted in a strong (67 and 74 %, respectively) receptor desensitization which was not influenced by monensin. Finally, CHO-hNOP cells exposed to the receptor-internalizing agonists for 24 h resulted in a significantly higher cAMP accumulation (defined supersensitization) compared with the non-internalizing agonists. In addition, blocking of receptor recycling by monensin led to a decrease of the cAMP accumulation only in cells exposed to internalizing agonists. These data show that prolonged receptor signaling mediated by receptor endocytosis and recycling/reactivation might reduce the development of tolerance but can enhance compensatory mechanisms that lead to supersensitivity of specific signaling pathways
Gesture and speech in maternal input to children with Down's syndrome
No full textBackground: Despite recent interest in relationships between maternal gesture and speech and communicative development in typically developing (M) children, little work has examined either speech or gesture in mothers of children with Down's syndrome (DS). Aims: To compare aspects of speech and gesture production by mothers of children with DS with that of mothers of TD children. Methods & Procedures: Participants were five mothers of children with DS (mean chronological age (CA)=47.6 months; mean mental age (MA,)=22.4 months) and five mothers of TD children. To equate for expressive language ability, children in the TD and DS groups were individually matched on the basis of: (1) gender; (2) correspondence between the TD child's chronological age and the DS child's language age; and (3) observed expressive vocabulary size. Each mother-child dyad was videotaped for approximately 30 min during free play. Data analyses focused on: (1) the number and types (speech only, gesture only, mixed) of maternal utterances; (2) the gesture types (deictic, iconic, conventional, emphatic); and (3) for mixed utterances, the structure and the temporal patterning of spoken and gestured components. Outcomes & Results: Relative to mothers of TD children, mothers of children with DS produced significantly fewer utterances overall, but the distribution of utterance types did not differ between the two groups. Relative to mothers of TD children, mothers of children with DS used proportionately more deictic gestures and made more frequent use of SHOWING. Mothers of TD children produced more POINTING gestures. Finally, mothers of children with DS produced a significantly higher proportion of utterances consisting of a single gesture and a single verbal utterance; in contrast to mothers of TD children, more complex structures (one gesture with multiple verbal utterances, one verbal utterance with multiple gestures) were never observed. Within the category of utterances consisting of a gesture and a single verbal utterance, mothers of children with DS tended to produce gestures that were held throughout the complete verbal utterance, while the gestures of mothers of TD children tended to co-occur with only a portion of the utterance. Conclusions: The findings suggest that mothers of children with DS adjust their communication to the developmental status of their child. Results are discussed in terms of the role of gesture in maternal communication and in the regulation of mother-child interaction
Phase II drugs under investigation for allergic conjunctivitis
No full textOcular allergies comprise a spectrum of conditions that are underreported and underdiagnosed, and are frequently associated with rhinoconjunctivitis. Although allergic conjunctivitis is often not a sight-threatening condition, it could have a significant impact on a person's quality of life, morbidity and productivity. A variety of agents are available for the treatment of allergic conjunctivitis, including antihistamines, mast-cell stabilizers, dual action agents, glucocorticoids, calcineurin inhibitors and immunotherapy
Conditioned place preference (CPP) in rats: From conditioning to reinstatement test
No full textOpioid addiction in humans is a chronically relapsing disorder characterized by discontinuous periods of drug use and abstinence resulting in dependence. With time, the probability of falling into renewed drug consumption becomes particularly high and constitutes a considerable problem in the management of opioid addicts. Opioid addiction represents an important health concern and animal models have been crucial in understanding the neurobiology and pathophysiology of this complex disease. Although animal models of addiction do not fully reproduce the human condition, they do permit investigation of specific elements of the process as well as identification of potential therapeutic targets. In this chapter, we provide a step-by-step description of the morphine-conditioned place preference (CPP) model that represents a useful preclinical animal model extensively used to study the rewarding/aversive effect of drugs
Nandrolone decreases mu opioid receptor expression in SH-SY5Y human neuroblastoma cells
No full textNandrolone and other anabolic androgenic steroids alter the expression and function of neurotransmitter systems and contribute to drug dependence. Nandrolone treatment (10-10 M) caused a time-dependent and concentration-dependent downregulation of mu opioid receptor (MOPr) transcripts in SH-SY5Y human neuroblastoma cells. This effect was prevented by the androgen receptor antagonist hydroxyflutamide. Receptor binding assays confirmed a decrease in MOPr of approximately 40% in nandrolone-treated cells. Treatment with actinomycin D (10 (-5)M), a transcription inhibitor, revealed that nandrolone might regulate MOPr mRNA stability. In SH-SY5Y cells transfected with a human MOPr luciferase promoter/reporter construct, nandrolone did not alter the rate of gene transcription. These results suggest that nandrolone may regulate MOPr expression through posttranscriptional mechanisms requiring the androgen receptor
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