Publications by authors named "Marco Tizzano"

26 Publications

  • Page 1 of 1

Equine rhinosporidiosis in Buenos Aires, Argentina.

Rev Argent Microbiol 2021 Apr 15. Epub 2021 Apr 15.

Cátedra de Micología, Facultad de Ciencias Veterinarias (UNLP), Buenos Aires, Argentina; CCT CONICET La Plata, Argentina.

Rhinosporidiosis is caused by Rhinosporidium seeberi, a parasitic organism of the family Rhinosporideacea family, class Micomycetozoa. The disease is endemic in India; however, some cases were reported in Europe, Africa, North America, and South America. The aim of the present study is to report three cases of rhinosporidiosis in wild horses in different cities of Buenos Aires province, Argentina. We confirm the presence of R. seeberi in the analyzed samples using histopathological and PCR sequencing techniques.
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http://dx.doi.org/10.1016/j.ram.2020.12.007DOI Listing
April 2021

Mouse mandibular retromolar taste buds associated with a mucus salivary gland.

Chem Senses 2021 Apr 15. Epub 2021 Apr 15.

Monell Chemical Senses Center, Philadelphia, PA, USA.

We have characterized a recently rediscovered chemosensory structure at the rear of the mandibular mucosa in the mouse oral cavity originally reported in the 1980s. This consists of unorganized taste buds, not contained within troughs, associated with the ducts of an underlying minor salivary gland. Using whole-mount preparations of transgenic mice expressing green fluorescent protein under the promoter of taste-signaling-specific genes, we determined that the structure contains taste bud clusters and salivary gland orifices at the rear of each mandible, distal to the last molar and anterior to the ascending ramus. Immunohistochemical analysis show in the retromolar taste buds expression of the taste receptors Tas2R131 and T1R3 and taste cascade molecules TrpM5, PLCβ2, and GNAT3, consistent with type II taste cells, and expression of GAD1, consistent with type III taste cells. Furthermore, the neuronal marker CGRP in retromolar mucosa tissue wrapping around TrpM5+ taste buds was observed. RT-PCR showed that retromolar taste buds express all three mouse tas1r genes, 28 of the 35 tas2r genes, and taste transduction signaling genes gnat3, plcb2, and trpm5, making the retromolar TBs similar to other lingual and palate taste buds. Finally, histochemistry demonstrated that the mandibular retromolar secretory gland is a minor salivary gland of mucous type. The mandibular retromolar taste structure may thus play a role in taste sensation and represent a potential novel pharmacological target for taste disorders.
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http://dx.doi.org/10.1093/chemse/bjab019DOI Listing
April 2021

Endometrial expression of key genes related to fertility in repeat breeder and non-repeat breeder cows.

Reprod Domest Anim 2020 Nov 27;55(11):1660-1664. Epub 2020 Oct 27.

Facultad de Ciencias Veterinarias, Instituto de Investigaciones en Reproducción Animal (INIRA), Universidad Nacional de La Plata (FCV-UNLP), La Plata, Argentina.

The aim of the present study was to compare the endometrial gene expression of epidermal growth factor receptor (EGFR), nodal growth differentiation factor (NODAL), prostaglandin-endoperoxide synthase 2 (PTGS2), oestrogen receptor 1 (ESR1) and progesterone receptor (PGR) in repeat breeder cows (RBC) and non-RBC during diestrus. Endometrial samples were collected by cytobrush technique and stored in RNA stabilizing solution at -20°C until RT-qPCR analysis. Differences in endometrial mRNA expression of selected genes were assessed by ANOVA and simple (r) and the partial correlations (rp) among selected genes were performed. Results demonstrated that mRNA expression of EGFR and NODAL were higher in RBC than in non-RBC (3 and 25-fold change, p < .01 and p < .01, respectively), while the mRNA expression of PTGS2 was lower (1.56-fold change, p < .01). Although there were no differences detected in the mRNA expression of ESR1 and PGR, there was a positive correlation between the expression of ESR1 and EGFR (0.84, p < .05) and a negative correlation between PGR and PTGS2 (-0.49, p < .05). In conclusion, the difference on the endometrial mRNA expression of the genes included in the study between RBC and non-RBC indicates a deregulation of important mechanisms that are vital to establish a successful pregnancy. Thus, the present study provides useful insight as a base for future studies to elucidate the causes of RBC.
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http://dx.doi.org/10.1111/rda.13841DOI Listing
November 2020

Gingival solitary chemosensory cells are immune sentinels for periodontitis.

Nat Commun 2019 10 3;10(1):4496. Epub 2019 Oct 3.

Monell Chemical Senses Center, Philadelphia, PA, 19104, USA.

Solitary chemosensory cells (SCCs) are epithelial sentinels that utilize bitter Tas2r receptors and coupled taste transduction elements to detect pathogenic bacterial metabolites, triggering host defenses to control the infection. Here we report that SCCs are present in mouse gingival junctional epithelium, where they express several Tas2rs and the taste signaling components α-gustducin (Gnat3), TrpM5, and Plcβ2. Gnat3 mice have altered commensal oral microbiota and accelerated naturally occurring alveolar bone loss. In ligature-induced periodontitis, knockout of taste signaling molecules or genetic absence of gingival SCCs (gSCCs) increases the bacterial load, reduces bacterial diversity, and renders the microbiota more pathogenic, leading to greater alveolar bone loss. Topical treatment with bitter denatonium to activate gSCCs upregulates the expression of antimicrobial peptides and ameliorates ligature-induced periodontitis in wild-type but not in Gnat3 mice. We conclude that gSCCs may provide a promising target for treating periodontitis by harnessing innate immunity to regulate the oral microbiome.
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http://dx.doi.org/10.1038/s41467-019-12505-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6776549PMC
October 2019

Microvillous cells in the olfactory epithelium express elements of the solitary chemosensory cell transduction signaling cascade.

PLoS One 2018 13;13(9):e0202754. Epub 2018 Sep 13.

Monell Chemical Senses Center, Philadelphia, Pennsylvania, United States of America.

The nasal cavity hosts an array of chemoresponsive cells, including the extended olfactory system and several other cells involved in detection of and responses to irritants. Solitary chemosensory cells (SCCs), which respond to irritants and bacteria, express the transient receptor potential channel TRPM5 an essential element of the taste transduction-signaling cascade. Microvillous cells (MVCs), non-neuronal cells situated in the apical layer of the main olfactory epithelium, also express TRPM5, but their function has not yet been clarified. TRPM5-positive MVCs, like SCCs, show a cholinergic phenotype expressing choline acetyl transferase (ChAT), but none of the other elements of the bitter taste transduction cascade could be detected. We reexamined TRPM5-positive MVCs with more sensitive gene expression and staining techniques to clarify whether they rely only on TRPM5 and ChAT or express other elements of the taste/SCC transduction cascade. Analyzing existing RNA sequencing data from whole olfactory mucosa and isolated olfactory sensory neurons, we determined that several elements of the taste/SCC transduction cascade, including taste receptors, are expressed in the olfactory mucosa in cells other than olfactory sensory neurons. Immunostaining confirmed the presence TRPM5 and ChAT in a subset of cells of the olfactory mucosa, which also showed the expression of PLCB2, gustducin, and T1R3. Specifically, these cells were identified as TRPM5-positive MVCs. Furthermore, we examined whether MVCs are innervated by trigeminal fibers, similarly to SCCs. Using antibodies against trigeminal nerve markers calcitonin gene-related peptide and substance P, we determined that, despite the cholinergic phenotype, most MVCs in the olfactory mucosa lacked consistent trigeminal innervation. Our findings indicate that MVCs, like SCCs, express all the elements of the bitter taste transduction cascade but that, unlike SCCs, they possess only sparse trigeminal innervation. The cholinergic phenotype of MVCs suggests a modulatory function of the surrounding olfactory epithelium, through the release of acetylcholine.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0202754PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6136699PMC
February 2019

Immunohistochemical Analysis of Human Vallate Taste Buds.

Chem Senses 2015 Nov 22;40(9):655-60. Epub 2015 Sep 22.

Rocky Mountain Taste & Smell Center, University of Colorado School of Medicine, 12801 E. 17th Avenue, Aurora, CO 80045, USA, Department of Cell & Developmental Biology, University of Colorado School of Medicine, 12801 E. 17th Avenue, Aurora, CO 80045, USA and Department of Otolaryngology, University of Colorado School of Medicine, 12700 E. 19th Avenue, Aurora, CO 80045, USA

The morphology of the vallate papillae from postmortem human samples was investigated with immunohistochemistry. Microscopically, taste buds were present along the inner wall of the papilla, and in some cases in the outer wall as well. The typical taste cell markers PLCβ2, GNAT3 (gustducin) and the T1R3 receptor stain elongated cells in human taste buds consistent with the Type II cells in rodents. In the human tissue, taste bud cells that stain with Type II cell markers, PLCβ2 and GNAT3, also stain with villin antibody. Two typical immunochemical markers for Type III taste cells in rodents, PGP9.5 and SNAP25, fail to stain any taste bud cells in the human postmortem tissue, although these antibodies do stain numerous nerve fibers throughout the specimen. Car4, another Type III cell marker, reacted with only a few taste cells in our samples. Finally, human vallate papillae have a general network of innervation similar to rodents and antibodies directed against SNAP25, PGP9.5, acetylated tubulin and P2X3 all stain free perigemmal nerve endings as well as intragemmal taste fibers. We conclude that with the exception of certain molecular features of Type III cells, human vallate papillae share the structural, morphological, and molecular features observed in rodents.
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http://dx.doi.org/10.1093/chemse/bjv048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4751242PMC
November 2015

Cholinergic neurotransmission links solitary chemosensory cells to nasal inflammation.

Proc Natl Acad Sci U S A 2014 Apr 7;111(16):6075-80. Epub 2014 Apr 7.

Rocky Mountain Taste and Smell Center, Department of Cellular and Developmental Biology and Neuroscience Program, University of Colorado School of Medicine, Anschutz Medical Center, Aurora, CO 80045.

Solitary chemosensory cells (SCCs) of the nasal cavity are specialized epithelial chemosensors that respond to irritants through the canonical taste transduction cascade involving Gα-gustducin and transient receptor potential melastatin 5. When stimulated, SCCs trigger peptidergic nociceptive (or pain) nerve fibers, causing an alteration of the respiratory rate indicative of trigeminal activation. Direct chemical excitation of trigeminal pain fibers by capsaicin evokes neurogenic inflammation in the surrounding epithelium. In the current study, we test whether activation of nasal SCCs can trigger similar local inflammatory responses, specifically mast cell degranulation and plasma leakage. The prototypical bitter compound, denatonium, a well-established activator of SCCs, caused significant inflammatory responses in WT mice but not mice with a genetic deletion of elements of the canonical taste transduction cascade, showing that activation of taste signaling components is sufficient to trigger local inflammation. Chemical ablation of peptidergic trigeminal fibers prevented the SCC-induced nasal inflammation, indicating that SCCs evoke inflammation only by neural activity and not by release of local inflammatory mediators. Additionally, blocking nicotinic, but not muscarinic, acetylcholine receptors prevents SCC-mediated neurogenic inflammation for both denatonium and the bacterial signaling molecule 3-oxo-C12-homoserine lactone, showing the necessity for cholinergic transmission. Finally, we show that the neurokinin 1 receptor for substance P is required for SCC-mediated inflammation, suggesting that release of substance P from nerve fibers triggers the inflammatory events. Taken together, these results show that SCCs use cholinergic neurotransmission to trigger peptidergic trigeminal nociceptors, which link SCCs to the neurogenic inflammatory pathway.
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http://dx.doi.org/10.1073/pnas.1402251111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4000837PMC
April 2014

Expression of the hemagglutinin HA1 subunit of the equine influenza virus using a baculovirus expression system.

Rev Argent Microbiol 2013 Oct-Dec;45(4):222-8

Department of Virology, Faculty of Veterinary Sciences, National University of La Plata, La Plata, Buenos Aires, Argentina.

Equine influenza virus is a leading cause of respiratory disease in horses worldwide. Disease prevention is by vaccination with inactivated whole virus vaccines. Most current influenza vaccines are generated in embryonated hens' eggs. Virions are harvested from allantoic fluid and chemically inactivated. Although this system has served well over the years, the use of eggs as the substrate for vaccine production has several well-recognized disadvantages (cost, egg supply, waste disposal and yield in eggs). The aim of this study was to evaluate a baculovirus system as a potential method for producing recombinant equine influenza hemagglutinin to be used as a vaccine. The hemagglutinin ectodomain (HA1 subunit) was cloned and expressed using a baculovirus expression vector. The expression was determined by SDS-PAGE and immunoblotting. A high yield, 20μg/ml of viral protein, was obtained from recombinant baculovirus-infected cells. The immune response in BALB/c mice was examined following rHA1 inoculation. Preliminary results show that recombinant hemagglutinin expressed from baculovirus elicits a strong antibody response in mice; therefore it could be used as an antigen for subunit vaccines and diagnostic tests.
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http://dx.doi.org/10.1016/S0325-7541(13)70028-2DOI Listing
May 2014

Solitary chemosensory cells and bitter taste receptor signaling in human sinonasal mucosa.

Int Forum Allergy Rhinol 2013 Jun 12;3(6):450-7. Epub 2013 Feb 12.

Department of Otolaryngology-Head and Neck Surgery, University of Colorado Denver, Aurora, CO 80045, USA.

Background: Solitary chemosensory cells (SCCs) are specialized cells in the respiratory epithelium that respond to noxious chemicals including bacterial signaling molecules. SCCs express components of bitter taste transduction including the taste receptor type 2 (TAS2R) bitter taste receptors and downstream signaling effectors: α-Gustducin, phospholipase Cβ2 (PLCβ2), and transient receptor potential cation channel subfamily M member 5 (TRPM5). When activated, SCCs evoke neurogenic reflexes, resulting in local inflammation. The purpose of this study was to test for the presence SCCs in human sinonasal epithelium, and to test for a correlation with inflammatory disease processes such as allergic rhinitis and chronic rhinosinusitis.

Methods: Patient demographics and biopsies of human sinonasal mucosa were obtained from control patients (n = 7) and those with allergic rhinitis and/or chronic rhinosinusitis (n = 15). Reverse transcription polymerase chain reaction (RT-PCR), quantitative PCR (qPCR), and immunohistochemistry were used to determine whether expression of signaling effectors was altered in diseased patients.

Results: RT-PCR demonstrated that bitter taste receptors TAS2R4, TAS2R14, and TAS2R46, and downstream signaling effectors α-Gustducin, PLCβ2, and TRPM5 are expressed in the inferior turbinate, middle turbinate, septum, and uncinate of both control and diseased patients. PLCβ2/TRPM5-immunoreactive SCCs were identified in the sinonasal mucosa of both control and diseased patients. qPCR showed similar expression of α-Gustducin and TRPM5 in the uncinate process of control and diseased groups, and there was no correlation between level of expression and 22-item Sino-Nasal Outcomes Test (SNOT-22) or pain scores.

Conclusion: SCCs are present in human sinonasal mucosa in functionally relevant areas. Expression level of signaling effectors was similar in control and diseased patients and did not correlate with measures of pain and inflammation. Further study into these pathways may provide insight into nasal inflammatory diseases and may offer potential therapeutic targets.
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http://dx.doi.org/10.1002/alr.21149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3655139PMC
June 2013

Chemosensors in the nose: guardians of the airways.

Physiology (Bethesda) 2013 Jan;28(1):51-60

Department of Cell & Developmental Biology, Rocky Mountain Taste & Smell Center, University of Colorado School of Medicine, Aurora, Colorado, USA.

The G-protein-coupled receptor molecules and downstream effectors that are used by taste buds to detect sweet, bitter, and savory tastes are also utilized by chemoresponsive cells of the airways to detect irritants. Here, we describe the different cell types in the airways that utilize taste-receptor signaling to trigger protective epithelial and neural responses to potentially dangerous toxins and bacterial infection.
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http://dx.doi.org/10.1152/physiol.00035.2012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3736818PMC
January 2013

Neuronal guidance molecule netrin-1 attenuates inflammatory cell trafficking during acute experimental colitis.

Gut 2012 May 3;61(5):695-705. Epub 2011 Aug 3.

Mucosal Inflammation Program, Department of Anesthesiology, University of Colorado Denver, Aurora, CO 80045, USA.

Background: Inflammatory bowel diseases, encompassing Crohn's disease and ulcerative colitis, are characterised by persistent leucocyte tissue infiltration leading to perpetuation of an inappropriate inflammatory cascade. The neuronal guidance molecule netrin-1 has recently been implicated in the orchestration of leucocyte trafficking during acute inflammation. We therefore hypothesised that netrin-1 could modulate leucocyte infiltration and disease activity in a model of inflammatory bowel disease.

Design: DSS-colitis was performed in mice with partial genetic netrin-1 deficiency (Ntn-1(+/-) mice) or wild-type mice treated with exogenous netrin-1 via osmotic pump to examine the role of endogenous and therapeutically administered netrin-1. These studies were supported by in vitro models of transepithelial migration and intestinal epithelial barrier function.

Results: Consistent with our hypothesis, we observed induction of netrin-1 during intestinal inflammation in vitro or in mice exposed to experimental colitis. Moreover, mice with partial netrin-1 deficiency demonstrated an exacerbated course of DSS-colitis compared to littermate controls, with enhanced weight loss and colonic shortening. Conversely, mice treated with exogenous mouse netrin-1 experienced attenuated disease severity. Importantly, permeability studies and quantitative assessment of apoptosis reveal that netrin-1 signalling events do not alter mucosal permeability or intestinal epithelial cell apoptosis. In vivo studies of leucocyte transmigration demonstrate suppression of neutrophil trafficking as a key function mediated by endogenous or exogenously administered netrin-1. Finally, genetic studies implicate the A2B adenosine receptor in netrin-1-mediated protection during DSS-colitis.

Conclusions: The present study identifies a previously unrecognised role for netrin-1 in attenuating experimental colitis through limitation of neutrophil trafficking.
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http://dx.doi.org/10.1136/gutjnl-2011-300012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3322588PMC
May 2012

First report of Israeli acute paralysis virus in asymptomatic hives of Argentina.

Rev Argent Microbiol 2011 Apr-May;43(2):84-6

CCT CONICET La Plata, Prov. Buenos Aires, Argentina.

Honey bee mortality has recently been associated with Israeli acute paralysis virus (IAPV), a proposed etiological agent for a new syndrome known as Colony Collapse Disorder. Bees infected with this virus show shivering wings, progress into paralysis, and finally die outside the hive. During the last years, honey bee mortality became a serious problem for Argentinean beekeepers. We herein report the preliminary results of a survey carried out to detect IAPV in samples taken from several Argentine provinces, by using a reverse transcription Polymerase Chain Reaction assay. Our data indicate the existence of high frequency of IAPV in asymptomatic hives of Argentina.
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http://dx.doi.org/10.1590/S0325-75412011000200003DOI Listing
August 2011

Expression of taste receptors in solitary chemosensory cells of rodent airways.

BMC Pulm Med 2011 Jan 13;11. Epub 2011 Jan 13.

Rocky Mountain Taste and Smell Center, Department of Cell and Developmental Biology, University of Colorado at Denver & Health Sciences Center, Aurora, CO, USA.

Background: Chemical irritation of airway mucosa elicits a variety of reflex responses such as coughing, apnea, and laryngeal closure. Inhaled irritants can activate either chemosensitive free nerve endings, laryngeal taste buds or solitary chemosensory cells (SCCs). The SCC population lies in the nasal respiratory epithelium, vomeronasal organ, and larynx, as well as deeper in the airway. The objective of this study is to map the distribution of SCCs within the airways and to determine the elements of the chemosensory transduction cascade expressed in these SCCs.

Methods: We utilized a combination of immunohistochemistry and molecular techniques (rtPCR and in situ hybridization) on rats and transgenic mice where the Tas1R3 or TRPM5 promoter drives expression of green fluorescent protein (GFP).

Results: Epithelial SCCs specialized for chemoreception are distributed throughout much of the respiratory tree of rodents. These cells express elements of the taste transduction cascade, including Tas1R and Tas2R receptor molecules, α-gustducin, PLCβ2 and TrpM5. The Tas2R bitter taste receptors are present throughout the entire respiratory tract. In contrast, the Tas1R sweet/umami taste receptors are expressed by numerous SCCs in the nasal cavity, but decrease in prevalence in the trachea, and are absent in the lower airways.

Conclusions: Elements of the taste transduction cascade including taste receptors are expressed by SCCs distributed throughout the airways. In the nasal cavity, SCCs, expressing Tas1R and Tas2R taste receptors, mediate detection of irritants and foreign substances which trigger trigeminally-mediated protective airway reflexes. Lower in the respiratory tract, similar chemosensory cells are not related to the trigeminal nerve but may still trigger local epithelial responses to irritants. In total, SCCs should be considered chemoreceptor cells that help in preventing damage to the respiratory tract caused by inhaled irritants and pathogens.
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http://dx.doi.org/10.1186/1471-2466-11-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3031280PMC
January 2011

First report of viral infections that affect argentine honeybees.

Environ Microbiol Rep 2010 Dec;2(6):749-51

Centro de Investigaciones de Fitopatología. Unidad de Bacteriología (CIDEFI). Facultad de Ciencias Agrarias y Forestales. Universidad Nacional de La Plata. 60 y 119, La Plata, 1900, Bs. As. Argentina. CCT La Plata - CONICET. Avenida Rivadavia 1917, Bs As - Argentina. Cátedra de Virología. Facultad de Ciencias Veterinarias. Universidad Nacional de La Plata. 60 y 118, La Plata, 1900, Bs As. Argentina. Comisión de Investigaciones Científicas, Provincia de Buenos Aires, Argentina. (CIC-PBA). 10 y 526, La Plata, Bs. As. Argentina.

Honey is one of the most important agricultural products for export in Argentina. In fact, more than 3.5 million beehives and 50 000 beekeepers are related with this production, mainly located in Buenos Aires province. Honeybee mortality is a serious problem that beekeepers in Argentina have had to face during the last 3 years. It is known that the consequence of the complex interactions between environmental and beekeeping parameters added to the effect of different disease agents such as viruses, bacteria, fungi and parasitic mites may result in a sudden collapse of the colony. In addition, multiple viral infections are frequently detected concomitantly in bee colonies. We describe here the preliminary results of a survey of three honeybee-pathogenic viruses, acute bee paralysis viruses (ABPV), chronic bee paralysis viruses (CBPV) and Sacbrood viruses (SBV) detected during a screening of 61 apiaries located in the main honey producer province using a RT-PCR assay. This is the first molecular report of the presence of these viruses in Argentine apiaries.
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http://dx.doi.org/10.1111/j.1758-2229.2010.00173.xDOI Listing
December 2010

Evidence for a role of glutamate as an efferent transmitter in taste buds.

BMC Neurosci 2010 Jun 21;11:77. Epub 2010 Jun 21.

Department of Otolaryngology, University of Colorado, Denver, CO, USA.

Background: Glutamate has been proposed as a transmitter in the peripheral taste system in addition to its well-documented role as an umami taste stimulus. Evidence for a role as a transmitter includes the presence of ionotropic glutamate receptors in nerve fibers and taste cells, as well as the expression of the glutamate transporter GLAST in Type I taste cells. However, the source and targets of glutamate in lingual tissue are unclear. In the present study, we used molecular, physiological and immunohistochemical methods to investigate the origin of glutamate as well as the targeted receptors in taste buds.

Results: Using molecular and immunohistochemical techniques, we show that the vesicular transporters for glutamate, VGLUT 1 and 2, but not VGLUT3, are expressed in the nerve fibers surrounding taste buds but likely not in taste cells themselves. Further, we show that P2X2, a specific marker for gustatory but not trigeminal fibers, co-localizes with VGLUT2, suggesting the VGLUT-expressing nerve fibers are of gustatory origin. Calcium imaging indicates that GAD67-GFP Type III taste cells, but not T1R3-GFP Type II cells, respond to glutamate at concentrations expected for a glutamate transmitter, and further, that these responses are partially blocked by NBQX, a specific AMPA/Kainate receptor antagonist. RT-PCR and immunohistochemistry confirm the presence of the Kainate receptor GluR7 in Type III taste cells, suggesting it may be a target of glutamate released from gustatory nerve fibers.

Conclusions: Taken together, the results suggest that glutamate may be released from gustatory nerve fibers using a vesicular mechanism to modulate Type III taste cells via GluR7.
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http://dx.doi.org/10.1186/1471-2202-11-77DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2898831PMC
June 2010

Nasal chemosensory cells use bitter taste signaling to detect irritants and bacterial signals.

Proc Natl Acad Sci U S A 2010 Feb 26;107(7):3210-5. Epub 2010 Jan 26.

Rocky Mountain Taste and Smell Center, Department of Cell and Developmental Biology, University of Colorado Denver, Aurora, CO 80045, USA.

The upper respiratory tract is continually assaulted with harmful dusts and xenobiotics carried on the incoming airstream. Detection of such irritants by the trigeminal nerve evokes protective reflexes, including sneezing, apnea, and local neurogenic inflammation of the mucosa. Although free intra-epithelial nerve endings can detect certain lipophilic irritants (e.g., mints, ammonia), the epithelium also houses a population of trigeminally innervated solitary chemosensory cells (SCCs) that express T2R bitter taste receptors along with their downstream signaling components. These SCCs have been postulated to enhance the chemoresponsive capabilities of the trigeminal irritant-detection system. Here we show that transduction by the intranasal solitary chemosensory cells is necessary to evoke trigeminally mediated reflex reactions to some irritants including acyl-homoserine lactone bacterial quorum-sensing molecules, which activate the downstream signaling effectors associated with bitter taste transduction. Isolated nasal chemosensory cells respond to the classic bitter ligand denatonium as well as to the bacterial signals by increasing intracellular Ca(2+). Furthermore, these same substances evoke changes in respiration indicative of trigeminal activation. Genetic ablation of either G alpha-gustducin or TrpM5, essential elements of the T2R transduction cascade, eliminates the trigeminal response. Because acyl-homoserine lactones serve as quorum-sensing molecules for gram-negative pathogenic bacteria, detection of these substances by airway chemoreceptors offers a means by which the airway epithelium may trigger an epithelial inflammatory response before the bacteria reach population densities capable of forming destructive biofilms.
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http://dx.doi.org/10.1073/pnas.0911934107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2840287PMC
February 2010

Acyl homoserine lactones induce early response in the airway.

Anat Rec (Hoboken) 2009 Mar;292(3):439-48

Department of Morphological-Biomedical Sciences, Section of Human Anatomy and Histology, University of Verona, Verona, Italy.

Acyl homoserine lactones (AHLs) are intercellular signaling molecules used in quorum sensing by Gram-negative bacteria. We studied the early effects on the rat airway of in vivo intratracheal administration of AHLs (i.e., P. aeruginosa and B. cepacia) to test the hypothesis that AHLs also act on the airway cells, modifying secretory mechanisms which are important in mucosal defense. One hour after treatment, N-butyryl-homoserine lactone (C4-HL) had caused dilated extracellular spaces, loss of cilia, reduction of secretory material, and the presence of pre-necrotic elements in the epithelium, while N-octanoyl-homoserine lactone (C8-HL) caused a mild lesion in the epithelium. After treatment with either C4- or C8-HL, reduced immunoreactivity was found using CC10 antibody. At ultrastructural examination, dilatation of the mitochondria was evident in ciliate and secretory cells, while solitary chemosensory cells appeared better preserved, showing aspects of nucleocytoplasmic activation. Using microarray analysis, we found down-regulation of early gene Fos and Egr1 in all AHL-treated specimens. In vivo pharmacological magnetic resonance imaging after C4- or C8-HL treatment showed a slight increase in tracheal secretion at a first evaluation 5 min after administration, with no increase in the following minutes. In conclusion, AHLs induce an early mucosal response, and the chondriomas of ciliate and secretory cells are the main cytological target of AHL action. Our results show that AHL action is not limited to activation of conspecific bacteria, but also modifies innate airway defense mechanisms.
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http://dx.doi.org/10.1002/ar.20866DOI Listing
March 2009

Expression of Galpha14 in sweet-transducing taste cells of the posterior tongue.

BMC Neurosci 2008 Nov 13;9:110. Epub 2008 Nov 13.

Rocky Mountain Taste & Smell Center, University of Colorado Denver School of Medicine, Aurora, CO 80045, USA.

Background: "Type II"/Receptor cells express G protein-coupled receptors (GPCRs) for sweet, umami (T1Rs and mGluRs) or bitter (T2Rs), as well as the proteins for downstream signalling cascades. Transduction downstream of T1Rs and T2Rs relies on G-protein and PLCbeta2-mediated release of stored Ca2+. Whereas Galphagus (gustducin) couples to the T2R (bitter) receptors, which Galpha-subunit couples to the sweet (T1R2 + T1R3) receptor is presently not known. We utilized RT-PCR, immunocytochemistry and single-cell gene expression profiling to examine the expression of the Galphaq family (q, 11, 14) in mouse taste buds.

Results: By RT-PCR, Galpha14 is expressed strongly and in a taste selective manner in posterior (vallate and foliate), but not anterior (fungiform and palate) taste fields. Galphaq and Galpha11, although detectable, are not expressed in a taste-selective fashion. Further, expression of Galpha14 mRNA is limited to Type II/Receptor cells in taste buds. Immunocytochemistry on vallate papillae using a broad Galphaq family antiserum reveals specific staining only in Type II taste cells (i.e. those expressing TrpM5 and PLCbeta2). This staining persists in Galphaq knockout mice and immunostaining with a Galpha11-specific antiserum shows no immunoreactivity in taste buds. Taken together, these data show that Galpha14 is the dominant Galphaq family member detected. Immunoreactivity for Galpha14 strongly correlates with expression of T1R3, the taste receptor subunit present in taste cells responsive to either umami or sweet. Single cell gene expression profiling confirms a tight correlation between the expression of Galpha14 and both T1R2 and T1R3, the receptor combination that forms sweet taste receptors.

Conclusion: Galpha14 is co-expressed with the sweet taste receptor in posterior tongue, although not in anterior tongue. Thus, sweet taste transduction may rely on different downstream transduction elements in posterior and anterior taste fields.
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http://dx.doi.org/10.1186/1471-2202-9-110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2596171PMC
November 2008

Is rat LRRP Ba1-651 a Delta-1-pyrroline-5-carboxylate dehydrogenase activated by changes in the concentration of sweet molecules?

Med Hypotheses 2007 23;68(4):864-7. Epub 2006 Oct 23.

Department of Morphological-Biomedical Sciences, Section of Anatomy and Histology, University of Verona, Verona, Italy.

The liver is one of the most complex organs in the body, which responds to hepatocellular damage with inflammatory, regenerative and repair processes designed to restore functional liver tissue mass. Rat LRRP Ba1-651, a liver regeneration related protein induced during partial hepatectomy, is classified as a member of the aldehyde dehydrogenase (ALDh) 4A1 superfamily. During a BLAST protein search, this protein basically showed three structural and functional domains: an intermediate filament-like protein, a Delta-1-pyrroline-5-carboxylate dehydrogenase (P5CDh) and an atrial natriuretic factor (ANF) receptor. We suggest that all amniotic mammals possess a Ba1-651 ortholog to that of rats. The ANF receptor domain of rat LRRP Ba1-651, which domain is part of the receptor family ligand binding region, shows a very high sequence homology (almost identity) to the extracellular amino-terminal domains of the mammalian sweet taste receptor T1R2. This receptor belongs to the type C family of G protein coupled receptors (GPCRs) and is characterized by the presence of large extracellular amino-terminal domains, a nine cysteine domain of family 3 GPCR and a 7tm_3 transmembrane type domain. We suggest that rat LRRP Ba1-651 protein is a liver P5CDh-ANF that is activated by changes in the concentration of sweet molecules. If the sugar concentration in the organ increases due to liver damage or the intake of carbohydrate-rich or protein-rich foods, the P5CDh-ANF enzyme is activated to help in P5C catabolism. The hormone insulin probably plays a key role in the regulation of this enzyme. In the model that we propose, the P5CDh-ANF enzyme is activated by a conformational change in protein structure in the P5C docking site due to sugars binding in the AFN receptor region of the LRRP Ba1-651 protein. Our research could be a further understanding of the biological significance of this P5CDh-ANF enzyme, with important potential applications in the treatment of HPII and liver diseases and in liver transplantation. Further studies of our P5CDh-ANF enzyme are needed to clarify its features and functions, and which substances are involved in its induction. These might use liver cell lines or purified LRRP Ba1-651 protein with sweet molecules in vitro. Other experiments may help to localize LRRP Bal-651 in the organ and to link its abnormal presence or absence to certain tumors like hepatocellular carcinoma.
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http://dx.doi.org/10.1016/j.mehy.2006.08.040DOI Listing
July 2007

Evidence of solitary chemosensory cells in a large mammal: the diffuse chemosensory system in Bos taurus airways.

J Anat 2006 Sep;209(3):333-7

Department of Morphological-Biomedical Sciences, Section of Anatomy and Histology, University of Verona, Verona, Italy.

The diffuse chemosensory system (DCS) of the respiratory apparatus is composed of solitary chemosensory cells (SCCs) that resemble taste cells but are not organized in end organs. The discovery of the DCS may open up new approaches to respiratory diseases. However, available data on mammalian SCCs have so far been collected from rodents, the airways of which display some differences from those of large mammals. Here we investigated the presence of the DCS and of SCCs in cows and bulls (Bos taurus), in which the airway cytology is similar to that in humans, focusing our attention on detection in the airways of molecules involved in the transduction cascade of taste [i.e. alpha-gustducin and phospholipase C of the beta2 subtype (PLCbeta2)]. The aim of the research was to extend our understanding of airway chemoreceptors and to compare the organization of the DCS in a large mammal with that in rodents. Using immunocytochemistry for alpha-gustducin, the taste buds of the tongue and arytenoid were visualized. In the trachea and bronchi, alpha-gustducin-immunoreactive SCCs were frequently found. Using immunocytochemistry for PLCbeta2, the staining pattern was generally similar to those seen for alpha-gustducin. Immunoblotting confirmed the expression of alpha-gustducin in the tongue and in all the airway regions tested. The study demonstrated the presence of SCCs in cows and bulls, suggesting that DCSs are present in many mammalian species. The description of areas with a high density of SCCs in bovine bronchi seems to indicate that the view of the DCS as made up of isolated cells totally devoid of ancillary elements is probably an oversimplification.
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http://dx.doi.org/10.1111/j.1469-7580.2006.00617.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2100327PMC
September 2006

Hormone fatty acid modifications: gram negative bacteria and vertebrates demonstrate common structure and function.

Med Hypotheses 2006 27;67(3):513-6. Epub 2006 Apr 27.

Department of Morphological-Biomedical Sciences, Section of Anatomy and Histology, University of Verona, Strade le Grazie, 8, 37134 Verona, Italy.

Bacteria are known to regulate diverse physiological processes through a mechanism called quorum sensing (QS). Prokaryotes communicate by extracellular signalling compounds, i.e. autoinducers (acyl homoserine lactone, AHL of Gram negative bacteria) or pheromones (post-translationally modified peptides of Gram positive bacteria), which activate genetic pathways when they reach a sufficient concentration (QS). A large number of Gram-negative quorum-sensing systems studied so far utilize N-acyl homoserine lactones as signal molecules. In vertebrates small synthetic molecules called growth hormone secretagogues (GHSs) stimulate the release of growth hormone (GH) from the pituitary. GH release is stimulated by hypothalamic GH-releasing hormone (GHRH) and ghrelin (endogenous ligand of the GHS-receptor, GHS-R). Ghrelin is a 28-amino acid peptide, in which the serine-3 (Ser3) is n-octanoylated, and this modification is essential for ghrelin's activity. Ghrelin is the first known case of a peptide hormone modified by a fatty acid. The major active form of ghrelin is a 28-amino acid peptide with octanoylated Ser3; one of the more represented bacterial autoinducers is the N-Octanoyl-DL-homoserine lactone (C8-HL) molecule. The authors hypothesize that Gram-negative bacteria and vertebrates have a functional similarity in the search of food and an important structural homology of AHL and ghrelin for the highly conserved Serine-acylated motive in both molecules. Our suggestions could help one to understand the convergent origin and the biologic meaning of the Serine-acylated group in these organisms, a biologic meaning very important due to the high conservation in two kingdoms which are so different.
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http://dx.doi.org/10.1016/j.mehy.2006.03.010DOI Listing
September 2006

Axon-like processes in type III cells of taste organs.

Anat Rec A Discov Mol Cell Evol Biol 2006 Mar;288(3):276-9

Department of Morphological-Biomedical Sciences, Section of Anatomy and Histology, University of Verona, Verona, Italy.

Type III cells of the taste organs are widely considered to be chemoreceptors. The present study was performed on the frog taste disk and describes an axon-like process in type III cells, which often contains a bundle of densely-packed parallel microfilaments. These processes pass through the basal membrane of the gustatory epithelium, running into the lamina propria (transbasal membrane processes, tBMPs). In their intraepithelial tract, tBMPs contain dense-cored vesicles revealing their origin from type III cells. Type III cells showing both classic nonrigid processes (with vesicles and nerve contacts) and tBMPs are present. The connective tract of a tBMP usually contains dense-cored vesicles only in its proximal portion. In some cases, the connective tract of tBMPs is almost perpendicular to the basal lamina. In other cases, it runs parallel to and below the basal lamina. Some tBMPs contact nerve fibers running in the subepithelial connective tissue; the contact area is rather wide but evident synapse-like junctions were never detected. Contacts between tBMPs and nerve fibers innervating basal cells are also found. In conclusion, the data demonstrate the existence of epithelial cells resembling primitive neurons that display an apical dendrite and axon-like basal processes. Until now, it was not considered possible that epithelial receptor cells extend processes out of the epithelium.
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http://dx.doi.org/10.1002/ar.a.20313DOI Listing
March 2006

alpha-Gustducin immunoreactivity in the airways.

Cell Tissue Res 2005 Feb 20;319(2):211-9. Epub 2004 Nov 20.

Department of Morphological-Biomedical Sciences, Human Anatomy and Histology Section, University of Verona, Verona, Italy.

The G-protein subunit alpha-gustducin is a marker of chemoreceptive cells. In the present study, we examined the immunohistochemical localization of alpha-gustducin in rat airway epithelium both by light and electron microscopy. alpha-Gustducin immunoreactivity was found in solitary cells that presented ultrastructural features of chemoreceptor cells, i.e. flask-shaped or pear-shaped, with an apical process with thin microvilli protruding into the lumen. The immunostaining was mainly concentrated in the apical process and along the basolateral cell surface. To investigate whether alpha-gustducin-immunoreactive cells represented a distinct cell subset in rat airways, we performed double-label immunocytochemistry with antibodies to protein gene groduct (PGP) 9.5, a marker of neuroendocrine cells, and to phospholipase C beta2 (PLCbeta2), a component of the bitter signalling pathway. alpha-Gustducin-immunoreactive cells were present in a subset of PGP-9.5-immunoreactive elements, although not all alpha-gustducin-positive cells expressed PGP 9.5 labelling. In addition, a subset of alpha-gustducin-expressing cells colocalized PLCbeta2. This work thus demonstrates that solitary alpha-gustducin-immunoreactive cells exist throughout the airways and represent a specialized cell type with morphological and immunohistochemical characteristics of chemoreceptor cells.
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http://dx.doi.org/10.1007/s00441-004-1007-2DOI Listing
February 2005

Laryngeal chemosensory clusters.

Chem Senses 2004 Oct;29(8):683-92

Department of Morphological-Biomedical Sciences, Human Anatomy and Histology Section, University of Verona, Italy.

The expression of molecules involved in the transductory cascade of the sense of taste (TRs, alpha-gustducin, PLCbeta2, IP3R3) has been described in lingual taste buds or in solitary chemoreceptor cells located in different organs. At the laryngeal inlet, immunocytochemical staining at the light and electron microscope levels revealed that alpha-gustducin and PLCbeta2 are mainly localized in chemosensory clusters (CCs), which are multicellular organizations differing from taste buds, being generally composed of two or three chemoreceptor cells. Compared with lingual taste buds, CCs are lower in height and smaller in diameter. In laryngeal CCs, immunocytochemistry using the two antibodies identified a similar cell type which appears rather unlike the alpha-gustducin-immunoreactive (IR) and PLCbeta2-IR cells visible in lingual taste buds. The laryngeal IR cells are shorter than the lingual ones, with poorly developed basal processes and their apical process is shorter and thicker. Some cells show a flask-like shape due to the presence of a large body and the absence of basal processes. CCs lack pores and their delimitation from the surrounding epithelium is poorly evident. The demonstration of the existence of CCs strengthens the hypothesis of a phylogenetic link between gustatory and solitary chemosensory cells.
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http://dx.doi.org/10.1093/chemse/bjh071DOI Listing
October 2004

Identification and characterization of a specific sensory epithelium in the rat larynx.

J Comp Neurol 2004 Jul;475(2):188-201

Department of Morphological-Biomedical Sciences, Human Anatomy and Histology Section, University of Verona, 37134 Verona, Italy.

A specific laryngeal sensory epithelium (SLSE), which includes arrays of solitary chemoreceptor cells, is described in the supraglottic region of the rat. Two plates of SLSE were found, one on each side of the larynx. The first plate was located in the ventrolateral wall of the larynx, and the second was located in the interarytenoidal region. In SLSE, immunoblotting showed the presence of alpha-gustducin and phospholipase C beta2 (PLCbeta2), which are two markers of chemoreceptor cells. At immunocytochemistry, laryngeal immunoreactivity for alpha-gustducin was localized mainly in solitary chemosensory cells. Double-label immunocytochemistry using confocal microscopy demonstrated that alpha-gustducin-expressing cells in large part colocalize type III IP3 receptor (IP3R3), another key molecule in bitter taste perception. However, some IP3R3-expressing cells do not colocalize alpha-gustducin. At ultrastructural immunocytochemistry, these cells showed packed apical microvilli, clear cytoplasmic vesicles, and cytoneural junctions. SLSE was characterized by high permeability to a tracer due to poorly developed junctional contacts between superficial cells. Junctions were short in length and showed little contact with the terminal web. Ultrastructural analysis showed deep pits among the superficial cells. In SLSE, high density of intraepithelial nerve fibers was found. The lamina propria of the SLSE appeared thicker than that in other supraglottic regions. It was characterized by the presence of a well-developed subepithelial nerve plexus. The immunocytochemical and ultrastructural data suggested that SLSE is a chemoreceptor located in an optimal position for detecting substances entering the larynx from the pharynx or the trachea.
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http://dx.doi.org/10.1002/cne.20172DOI Listing
July 2004

Single and double knockouts of the genes for photosystem I subunits G, K, and H of Arabidopsis. Effects on photosystem I composition, photosynthetic electron flow, and state transitions.

Plant Physiol 2002 Jun;129(2):616-24

Zentrum zur Identifikation von Genfunktionen durch Insertionsmutagenese bei Arabidopsis thaliana, Max-Planck-Institut für Züchtungsforschung, Carl-von-Linné Weg 10, 50829 Köln, Germany.

Photosystem I (PSI) of higher plants contains 18 subunits. Using Arabidopsis En insertion lines, we have isolated knockout alleles of the genes psaG, psaH2, and psaK, which code for PSI-G, -H, and -K. In the mutants psak-1 and psag-1.4, complete loss of PSI-K and -G, respectively, was confirmed, whereas the residual H level in psah2-1.4 is due to a second gene encoding PSI-H, psaH1. Double mutants, lacking PSI-G, and also -K, or a fraction of -H, together with the three single mutants were characterized for their growth phenotypes and PSI polypeptide composition. In general, the loss of each subunit has secondary, in some cases additive, effects on the abundance of other PSI polypeptides, such as D, E, H, L, N, and the light-harvesting complex I proteins Lhca2 and 3. In the G-less mutant psag-1.4, the variation in PSI composition suggests that PSI-G stabilizes the PSI-core. Levels of light-harvesting complex I proteins in plants, which lack simultaneously PSI-G and -K, indicate that PSI subunits other than G and K can also bind Lhca2 and 3. In the same single and double mutants, psag-1.4, psak-1, psah2-1.4, psag-1.4/psah2-1.4, and psag-1.4/psak-1 photosynthetic electron flow and excitation energy quenching were analyzed to address the roles of the various subunits in P700 reduction (mediated by PSI-F and -N) and oxidation (PSI-E), and state transitions (PSI-H). Based on the results, we also suggest for PSI-K a role in state transitions.
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http://dx.doi.org/10.1104/pp.002089DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC161688PMC
June 2002