Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 30
Filtrar
Más filtros











Base de datos
Intervalo de año de publicación
1.
Res Sq ; 2024 Jul 12.
Artículo en Inglés | MEDLINE | ID: mdl-39041040

RESUMEN

Current clinical strategies for the delivery of pulmonary therapeutics to the lung are primarily targeted to the upper portions of the airways. However, targeted delivery to the lower regions of the lung is necessary for the treatment of parenchymal lung injury and disease. Here, we have developed an mRNA therapeutic for the lower lung using one-component Ionizable Amphiphilic Janus Dendrimers (IAJDs) as a delivery vehicle. We deliver an anti-inflammatory cytokine mRNA, transforming growth factor-beta (TGF-ß), to produce transient protein expression in the lower regions of the lung. This study highlights IAJD's potential for precise, effective, and safe delivery of TGF-ß mRNA to the lung. This delivery system offers a promising approach for targeting therapeutics to the specific tissues, a strategy necessary to fill the current clinical gap in treating parenchymal lung injury and disease.

2.
J Am Chem Soc ; 146(6): 3627-3634, 2024 02 14.
Artículo en Inglés | MEDLINE | ID: mdl-38306714

RESUMEN

Constitutional isomerism has been previously demonstrated by one of our laboratories to represent a powerful design strategy for the elaboration of complex functional self-organizations. Here we report the design, synthesis, and characterization of 14 positional, skeletal, and functional constitutional isomeric one-component, multifunctional, sequence-defined, amphiphilic ionizable Janus dendrimers (IAJDs). Their coassembly by simple injection with luciferase mRNA (Luc-mRNA) to form dendrimersome nanoparticles (DNPs) was studied. Subsequently, the resulting DNPs were employed to investigate, with screening experiments, the delivery of Luc-mRNA in vivo. Constitutional isomerism was shown to produce changes of up to two orders of magnitude of the total-body luciferase activity and targeted luciferase activity to the spleen and liver, of up to three orders of magnitude difference in targeted luciferase activity to the lungs and up to six orders of magnitude to lymph nodes. These results indicate that constitutional isomerism may represent not only a simple but also an important synthetic strategy that most probably may impact the activity of all components of synthetic vectors used in RNA-based nanomedicine, including in mRNA vaccines and therapeutics.


Asunto(s)
Dendrímeros , Nanopartículas , Isomerismo , Dendrímeros/química , ARN Mensajero/genética , Luciferasas
3.
Sensors (Basel) ; 23(19)2023 Sep 30.
Artículo en Inglés | MEDLINE | ID: mdl-37837022

RESUMEN

In the last several decades, metal oxide thin films have attracted significant attention for the development of various existing and emerging technological applications, including pH sensors. The mandate for consistent and precise pH sensing techniques has been increasing across various fields, including environmental monitoring, biotechnology, food and agricultural industries, and medical diagnostics. Metal oxide thin films grown using physical vapor deposition (PVD) with precise control over film thickness, composition, and morphology are beneficial for pH sensing applications such as enhancing pH sensitivity and stability, quicker response, repeatability, and compatibility with miniaturization. Various PVD techniques, including sputtering, evaporation, and ion beam deposition, used to fabricate thin films for tailoring materials' properties for the advanced design and development of high-performing pH sensors, have been explored worldwide by many research groups. In addition, various thin film materials have also been investigated, including metal oxides, nitrides, and nanostructured films, to make very robust pH sensing electrodes with higher pH sensing performance. The development of novel materials and structures has enabled higher sensitivity, improved selectivity, and enhanced durability in harsh pH environments. The last decade has witnessed significant advancements in PVD thin films for pH sensing applications. The combination of precise film deposition techniques, novel materials, and surface functionalization strategies has led to improved pH sensing performance, making PVD thin films a promising choice for future pH sensing technologies.

4.
J Am Chem Soc ; 145(34): 18760-18766, 2023 08 30.
Artículo en Inglés | MEDLINE | ID: mdl-37606244

RESUMEN

Delivery of nucleic acids with viral and synthetic vectors has pioneered genetic nanomedicine. Four-component lipid nanoparticles (LNPs) consisting of ionizable lipids, phospholipids, cholesterol, and PEG-conjugated lipids, assembled by microfluidic or T-tube, are the benchmark synthetic vector for delivery of mRNA. One-component multifunctional sequence-defined ionizable amphiphilic Janus dendrimer (IAJD) delivery systems for mRNA were developed by us to complement LNPs. IAJDs consist of multifunctional hydrophilic low-generation dendrons or minidendrons conjugated to hydrophobic dendrons. They were inspired by amphiphilic Janus dendrimers and glycodendrimers. IAJDs coassemble with mRNA into predictable-size vesicles, named dendrimersome nanoparticles (DNPs), by simple injection in acetate buffer, rather than by the complex technology required by LNPs. Assembly of DNPs by simple injection together with sequence design in the hydrophilic and hydrophobic modules of IAJDs endowed rapid screening to access discovery. Molecular design principles for targeted delivery were elaborated when the branching points of IAJDs were constructed from symmetrically and nonsymmetrically substituted plant phenolic acids interconnected by pentaerythritol (PE). Here, we report the first library containing simplified IAJDs constructed in only three steps from symmetrically trialkylated PE in the hydrophobic domain and four different piperazine-based ionizable amines in the hydrophilic part. Rapid coassembly with mRNA and in vivo screening led to the discovery of the two most active IAJDs targeting the spleen, liver, and lymph nodes, one predominantly to the spleen and liver and six delivering equally to the spleen, liver, lung, and lymph nodes. These IAJDs represent the simplest synthetic vectors and the first viral or synthetic system delivering equally to multiple organs.


Asunto(s)
Dendrímeros , ARN Mensajero/genética , Hígado , Lípidos
5.
Pharmaceutics ; 15(6)2023 May 23.
Artículo en Inglés | MEDLINE | ID: mdl-37376020

RESUMEN

Viral and synthetic vectors to deliver nucleic acids were key to the rapid development of extraordinarily efficient COVID-19 vaccines. The four-component lipid nanoparticles (LNPs), containing phospholipids, PEG-conjugated lipids, cholesterol, and ionizable lipids, co-assembled with mRNA via a microfluidic technology, are the leading nonviral delivery vector used by BioNTech/Pfizer and Moderna to access COVID-19 mRNA vaccines. LNPs exhibit a statistical distribution of their four components when delivering mRNA. Here, we report a methodology that involves screening libraries to discover the molecular design principles required to realize organ-targeted mRNA delivery and mediate activity with a one-component ionizable multifunctional amphiphilic Janus dendrimer (IAJD) derived from plant phenolic acids. IAJDs co-assemble with mRNA into monodisperse dendrimersome nanoparticles (DNPs) with predictable dimensions, via the simple injection of their ethanol solution in a buffer. The precise location of the functional groups in one-component IAJDs demonstrated that the targeted organs, including the liver, spleen, lymph nodes, and lung, are selected based on the hydrophilic region, while activity is associated with the hydrophobic domain of IAJDs. These principles, and a mechanistic hypothesis to explain activity, simplify the synthesis of IAJDs, the assembly of DNPs, handling, and storage of vaccines, and reduce price, despite employing renewable plant starting materials. Using simple molecular design principles will lead to increased accessibility to a large diversity of mRNA-based vaccines and nanotherapeutics.

6.
Polymers (Basel) ; 15(5)2023 Feb 21.
Artículo en Inglés | MEDLINE | ID: mdl-36904317

RESUMEN

In 2022, the Nobel Prize in Chemistry was awarded to Bertozzi, Meldal, and Sharpless "for the development of click chemistry and biorthogonal chemistry". Since 2001, when the concept of click chemistry was advanced by Sharpless laboratory, synthetic chemists started to envision click reactions as the preferred choice of synthetic methodology employed to create new functions. This brief perspective will summarize research performed in our laboratories with the classic Cu(I)-catalyzed azide-alkyne click (CuAAC) reaction elaborated by Meldal and Sharpless, with the thio-bromo click (TBC) and with the less-used, irreversible TERminator Multifunctional INItiator (TERMINI) dual click (TBC) reactions, the last two elaborated in our laboratory. These click reactions will be used to assemble, by accelerated modular-orthogonal methodologies, complex macromolecules and self-organizations of biological relevance. Self-assembling amphiphilic Janus dendrimers and Janus glycodendrimers together with their biological membrane mimics known as dendrimersomes and glycodendrimersomes as well as simple methodologies to assemble macromolecules with perfect and complex architecture such as dendrimers from commercial monomers and building blocks will be discussed. This perspective is dedicated to the 75th anniversary of Professor Bogdan C. Simionescu, the son of my (VP) Ph.D. mentor, Professor Cristofor I. Simionescu, who as his father, took both science and science administration in his hands, and dedicated his life to handling them in a tandem way, to their best.

7.
Nat Commun ; 13(1): 6889, 2022 11 12.
Artículo en Inglés | MEDLINE | ID: mdl-36371422

RESUMEN

Stimulus transduction in cilia of olfactory sensory neurons is mediated by odorant receptors, Gαolf, adenylate cyclase-3, cyclic nucleotide-gated and chloride ion channels. Mechanisms regulating trafficking and localization of these proteins in the dendrite are unknown. By lectin/immunofluorescence staining and in vivo correlative light-electron microscopy (CLEM), we identify a retinitis pigmentosa-2 (RP2), ESCRT-0 and synaptophysin-containing multivesicular organelle that is not part of generic recycling/degradative/exosome pathways. The organelle's intraluminal vesicles contain the olfactory transduction proteins except for Golf subunits Gγ13 and Gß1. Instead, Gß1 colocalizes with RP2 on the organelle's outer membrane. The organelle accumulates in response to stimulus deprivation, while odor stimuli or adenylate cyclase activation cause outer membrane disintegration, release of intraluminal vesicles, and RP2/Gß1 translocation to the base of olfactory cilia. Together, these findings reveal the existence of a dendritic organelle that mediates both stimulus-regulated storage of olfactory ciliary transduction proteins and membrane-delimited sorting important for G protein heterotrimerization.


Asunto(s)
Neuronas Receptoras Olfatorias , Receptores Odorantes , Adenilil Ciclasas/metabolismo , Cuerpos Multivesiculares , Neuronas Receptoras Olfatorias/metabolismo , Receptores Odorantes/metabolismo , Olfato/fisiología , Cilios/metabolismo , Proteínas/metabolismo , Mucosa Olfatoria/metabolismo
8.
J Am Chem Soc ; 144(11): 4746-4753, 2022 03 23.
Artículo en Inglés | MEDLINE | ID: mdl-35263098

RESUMEN

Viral and synthetic vectors for delivery of nucleic acids impacted genetic nanomedicine by aiding the rapid development of the extraordinarily efficient Covid-19 vaccines. Access to targeted delivery of nucleic acids is expected to expand the field of nanomedicine beyond most expectations. Both viral and synthetic vectors have advantages and disadvantages. The major advantage of the synthetic vectors is their unlimited synthetic capability. The four-component lipid nanoparticles (LNPs) are the leading nonviral vector for mRNA used by Pfizer and Moderna in Covid-19 vaccines. Their synthetic capacity inspired us to develop a one-component multifunctional sequence-defined ionizable amphiphilic Janus dendrimer (IAJD) delivery system for mRNA. The first experiments on IAJDs provided, through a rational-library design combined with orthogonal-modular accelerated synthesis and sequence control in their hydrophilic part, some of the most active synthetic vectors for the delivery of mRNA to lung. The second experiments employed a similar strategy, generating, by a less complex hydrophilic structure, a library of IAJDs targeting spleen, liver, and lung. Here, we report preliminary studies designing the hydrophobic region of IAJDs by using dissimilar alkyl lengths and demonstrate the unexpectedly important role of the primary structure of the hydrophobic part of IAJDs by increasing up to 90.2-fold the activity of targeted delivery of mRNA to spleen, lymph nodes, liver, and lung. The principles of the design strategy reported here and in previous publications indicate that IAJDs could have a profound impact on the future of genetic nanomedicine.


Asunto(s)
COVID-19 , Dendrímeros , Nanopartículas , Vacunas contra la COVID-19 , Dendrímeros/química , Humanos , Liposomas , Nanopartículas/química , ARN Mensajero/química , ARN Mensajero/genética
9.
J Am Chem Soc ; 143(43): 17975-17982, 2021 11 03.
Artículo en Inglés | MEDLINE | ID: mdl-34672554

RESUMEN

Targeted and efficient delivery of nucleic acids with viral and synthetic vectors is the key step of genetic nanomedicine. The four-component lipid nanoparticle synthetic delivery systems consisting of ionizable lipids, phospholipids, cholesterol, and a PEG-conjugated lipid, assembled by microfluidic or T-tube technology, have been extraordinarily successful for delivery of mRNA to provide Covid-19 vaccines. Recently, we reported a one-component multifunctional sequence-defined ionizable amphiphilic Janus dendrimer (IAJD) synthetic delivery system for mRNA relying on amphiphilic Janus dendrimers and glycodendrimers developed in our laboratory. Amphiphilic Janus dendrimers consist of functional hydrophilic dendrons conjugated to hydrophobic dendrons. Co-assembly of IAJDs with mRNA into dendrimersome nanoparticles (DNPs) occurs by simple injection in acetate buffer, rather than by microfluidic devices, and provides a very efficient system for delivery of mRNA to lung. Here we report the replacement of most of the hydrophilic fragment of the dendron from IAJDs, maintaining only its ionizable amine, while changing its interconnecting group to the hydrophobic dendron from amide to ester. The resulting IAJDs demonstrated that protonated ionizable amines play dual roles of hydrophilic fragment and binding ligand for mRNA, changing delivery from lung to spleen and/or liver. Replacing the interconnecting ester with the amide switched the delivery back to lung. Delivery predominantly to liver is favored by pairs of odd and even alkyl groups in the hydrophobic dendron. This simple structural change transformed the targeted delivery of mRNA mediated with IAJDs, from lung to liver and spleen, and expands the utility of DNPs from therapeutics to vaccines.


Asunto(s)
Dendrímeros/química , ARN Mensajero/química , Aminas/química , Animales , Ésteres/química , Interacciones Hidrofóbicas e Hidrofílicas , Iones/química , Ratones , Nanopartículas/química , ARN Mensajero/inmunología , ARN Mensajero/metabolismo , Vacunas Sintéticas/química , Vacunas Sintéticas/inmunología , Vacunas Sintéticas/metabolismo
10.
J Am Chem Soc ; 143(31): 12315-12327, 2021 08 11.
Artículo en Inglés | MEDLINE | ID: mdl-34324336

RESUMEN

Efficient viral or nonviral delivery of nucleic acids is the key step of genetic nanomedicine. Both viral and synthetic vectors have been successfully employed for genetic delivery with recent examples being DNA, adenoviral, and mRNA-based Covid-19 vaccines. Viral vectors can be target specific and very efficient but can also mediate severe immune response, cell toxicity, and mutations. Four-component lipid nanoparticles (LNPs) containing ionizable lipids, phospholipids, cholesterol for mechanical properties, and PEG-conjugated lipid for stability represent the current leading nonviral vectors for mRNA. However, the segregation of the neutral ionizable lipid as droplets in the core of the LNP, the "PEG dilemma", and the stability at only very low temperatures limit their efficiency. Here, we report the development of a one-component multifunctional ionizable amphiphilic Janus dendrimer (IAJD) delivery system for mRNA that exhibits high activity at a low concentration of ionizable amines organized in a sequence-defined arrangement. Six libraries containing 54 sequence-defined IAJDs were synthesized by an accelerated modular-orthogonal methodology and coassembled with mRNA into dendrimersome nanoparticles (DNPs) by a simple injection method rather than by the complex microfluidic technology often used for LNPs. Forty four (81%) showed activity in vitro and 31 (57%) in vivo. Some, exhibiting organ specificity, are stable at 5 °C and demonstrated higher transfection efficiency than positive control experiments in vitro and in vivo. Aside from practical applications, this proof of concept will help elucidate the mechanisms of packaging and release of mRNA from DNPs as a function of ionizable amine concentration, their sequence, and constitutional isomerism of IAJDs.


Asunto(s)
Dendrímeros/química , Portadores de Fármacos/química , Nanopartículas/química , ARN Mensajero/metabolismo , Tensoactivos/química , Animales , Dendrímeros/síntesis química , Portadores de Fármacos/síntesis química , Liberación de Fármacos , Femenino , Células HEK293 , Humanos , Masculino , Ratones , Prueba de Estudio Conceptual , Tensoactivos/síntesis química
11.
eNeuro ; 8(2)2021.
Artículo en Inglés | MEDLINE | ID: mdl-33637538

RESUMEN

Stomatin-like protein-3 (STOML3) is an integral membrane protein expressed in the cilia of olfactory sensory neurons (OSNs), but its functional role in this cell type has never been addressed. STOML3 is also expressed in dorsal root ganglia neurons, where it has been shown to be required for normal touch sensation. Here, we extended previous results indicating that STOML3 is mainly expressed in the knob and proximal cilia of OSNs. We additionally showed that mice lacking STOML3 have a morphologically normal olfactory epithelium. Because of its presence in the cilia, together with known olfactory transduction components, we hypothesized that STOML3 could be involved in modulating odorant responses in OSNs. To investigate the functional role of STOML3, we performed loose patch recordings from wild-type (WT) and Stoml3 knock-out (KO) OSNs. We found that spontaneous mean firing activity was lower with additional shift in interspike intervals (ISIs) distributions in Stoml3 KOs compared with WT neurons. Moreover, the firing activity in response to stimuli was reduced both in spike number and duration in neurons lacking STOML3 compared with WT neurons. Control experiments suggested that the primary deficit in neurons lacking STOML3 was at the level of transduction and not at the level of action potential generation. We conclude that STOML3 has a physiological role in olfaction, being required for normal sensory encoding by OSNs.


Asunto(s)
Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Neuronas Receptoras Olfatorias , Olfato , Animales , Cilios , Ratones , Mucosa Olfatoria
12.
Biomacromolecules ; 21(5): 1902-1919, 2020 05 11.
Artículo en Inglés | MEDLINE | ID: mdl-31990544

RESUMEN

A mixed-ligand effect was observed for mixtures of tris(2-dimethylaminoethyl)amine (Me6-TREN) with tris(2-aminoethyl)amine (TREN) ligands during Cu(0) wire-catalyzed, single-electron transfer-living radical polymerization (SET-LRP) of methyl acrylate (MA) initiated with bis(2-bromopropionyl)ethane (BPE) in DMSO. The external order of reaction of SET-LRP both in the presence of Me6-TREN, TREN and of the mixed-ligand Me6-TREN/TREN, in DMSO, demonstrated a catalytic activity for DMSO similar to that reported in the presence of Cu(0) powder. The catalytic activity of DMSO, with close to 100% chain-end functionality, facilitates the much less expensive TREN to act as a very efficient ligand that is competitive with Me6-TREN and with the mixed-ligand and revitalizes TREN into an excellent ligand. The highest activity of the mixed-ligand at 1/1 ratio between ligands suggests that in addition to a fast exchange between these two ligands, a new single dynamic ligand stabilized by hydrogen-bonding, may generate these results.


Asunto(s)
Cobre , Dimetilsulfóxido , Transporte de Electrón , Ligandos , Polimerizacion
13.
Biomacromolecules ; 21(1): 250-261, 2020 01 13.
Artículo en Inglés | MEDLINE | ID: mdl-31566368

RESUMEN

The mixed-ligand system consisting of tris(2-aminoethyl)amine (TREN) and tris(2-dimethylaminoethyl)amine (Me6-TREN) during the Cu(0) wire-catalyzed single electron transfer-living radical polymerization (SET-LRP) of methyl acrylate (MA) in "programmed" biphasic mixtures of the dipolar aprotic solvents NMP, DMF, and DMAc with H2O is reported. Kinetic and chain end analysis studies by NMR and MALDI-TOF before and after thio-bromo "click" reaction demonstrated that Me6-TREN complements and makes the less expensive TREN a very efficient ligand in the absence of externally added Cu(II)Br2. Statistical analysis of the kinetic data together with control experiments demonstrated that this mixed-ligand effect enhanced the apparent rate constant of propagation, monomer conversion, and molecular weight control. The most efficient effect was observed at a 1/1 molar ratio between these two ligands, suggesting that in addition to a fast exchange between the two ligands, a new single dynamic ligand generated by hydrogen bonding may be responsible for the mixed ligand observed.


Asunto(s)
Cobre/química , Etilenodiaminas/química , Polimerizacion , Catálisis , Cinética , Ligandos , Espectroscopía de Resonancia Magnética , Polimetil Metacrilato/química , Solventes/química , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Agua/química
14.
J Gen Physiol ; 151(7): 954-966, 2019 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-31048412

RESUMEN

Glial-like supporting (or sustentacular) cells are important constituents of the olfactory epithelium that are involved in several physiological processes such as production of endocannabinoids, insulin, and ATP and regulation of the ionic composition of the mucus layer that covers the apical surface of the olfactory epithelium. Supporting cells express metabotropic P2Y purinergic receptors that generate ATP-induced Ca2+ signaling through the activation of a PLC-mediated cascade. Recently, we reported that a subpopulation of supporting cells expresses also the Ca2+-activated Cl- channel TMEM16A. Here, we sought to extend our understanding of a possible physiological role of this channel in the olfactory system by asking whether Ca2+ can activate Cl- currents mediated by TMEM16A. We use whole-cell patch-clamp analysis in slices of the olfactory epithelium to measure dose-response relations in the presence of various intracellular Ca2+ concentrations, ion selectivity, and blockage. We find that knockout of TMEM16A abolishes Ca2+-activated Cl- currents, demonstrating that TMEM16A is essential for these currents in supporting cells. Also, by using extracellular ATP as physiological stimuli, we found that the stimulation of purinergic receptors activates a large TMEM16A-dependent Cl- current, indicating a possible role of TMEM16A in ATP-mediated signaling. Altogether, our results establish that TMEM16A-mediated currents are functional in olfactory supporting cells and provide a foundation for future work investigating the precise physiological role of TMEM16A in the olfactory system.


Asunto(s)
Potenciales de Acción , Anoctamina-1/metabolismo , Mucosa Olfatoria/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Calcio/metabolismo , Cloruros/metabolismo , Ratones , Ratones Endogámicos C57BL , Mucosa Olfatoria/fisiología , Receptores Purinérgicos/metabolismo
15.
Proc Natl Acad Sci U S A ; 114(44): E9386-E9394, 2017 10 31.
Artículo en Inglés | MEDLINE | ID: mdl-29078327

RESUMEN

The ciliary localization of odorant receptors (ORs) is evolutionary conserved and essential for olfactory transduction. However, how the transport of ORs is regulated in mammalian olfactory sensory neurons is poorly understood. Here we demonstrate that odorant responsiveness and OR transport is regulated by the Hedgehog pathway. OR transport is inhibited by conditional gene inactivation of the Hedgehog signal mediator Smoothened (Smo) as well as by systemic administration of the Smo inhibitor vismodegib, a clinically used anticancer drug reported to distort smell perception in patients. The ciliary phenotype of Smo inhibition is haploinsufficient, cell autonomous, and correlates with the accumulation of OR-containing putative transport vesicles in the cytosol. The Smo-dependent OR transport route works in parallel with a low basal transport of vesicle containing both ORs and other olfactory transduction components. These findings both define a physiological function of Hedgehog signaling in olfaction and provide an important evolutionary link between olfaction and the requirement of a ciliary compartment for Hedgehog signaling.


Asunto(s)
Cilios/metabolismo , Proteínas Hedgehog/metabolismo , Receptores Odorantes/metabolismo , Anilidas/farmacología , Animales , Cilios/efectos de los fármacos , Citosol/efectos de los fármacos , Citosol/metabolismo , Ratones , Ratones Endogámicos C57BL , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/fisiología , Piridinas/farmacología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Receptor Smoothened/metabolismo
16.
PLoS One ; 10(6): e0129171, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26067252

RESUMEN

TMEM16A/ANO1 is a calcium-activated chloride channel expressed in several types of epithelia and involved in various physiological processes, including proliferation and development. During mouse embryonic development, the expression of TMEM16A in the olfactory epithelium is dynamic. TMEM16A is expressed at the apical surface of the entire olfactory epithelium at embryonic day E12.5 while from E16.5 its expression is restricted to a region near the transition zone with the respiratory epithelium. To investigate whether TMEM16A plays a role in the development of the mouse olfactory epithelium, we obtained the first immunohistochemistry study comparing the morphological properties of the olfactory epithelium and nasal glands in TMEM16A-/- and TMEM16A+/+ littermate mice. A comparison between the expression of the olfactory marker protein and adenylyl cyclase III shows that genetic ablation of TMEM16A did not seem to affect the maturation of olfactory sensory neurons and their ciliary layer. As TMEM16A is expressed at the apical part of supporting cells and in their microvilli, we used ezrin and cytokeratin 8 as markers of microvilli and cell body of supporting cells, respectively, and found that morphology and development of supporting cells were similar in TMEM16A-/- and TMEM16A+/+ littermate mice. The average number of supporting cells, olfactory sensory neurons, horizontal and globose basal cells were not significantly different in the two types of mice. Moreover, we also observed that the morphology of Bowman's glands, nasal septal glands and lateral nasal glands did not change in the absence of TMEM16A. Our results indicate that the development of mouse olfactory epithelium and nasal glands does not seem to be affected by the genetic ablation of TMEM16A.


Asunto(s)
Canales de Cloruro/metabolismo , Mucosa Nasal/metabolismo , Mucosa Olfatoria/metabolismo , Animales , Anoctamina-1 , Canales de Cloruro/deficiencia , Canales de Cloruro/genética , Embrión de Mamíferos/metabolismo , Femenino , Inmunohistoquímica , Queratina-8/metabolismo , Antígeno Ki-67/metabolismo , Ratones , Ratones Noqueados , Microscopía Confocal , Microvellosidades/metabolismo , Neuronas Receptoras Olfatorias/metabolismo , Embarazo
17.
J Gen Physiol ; 145(4): 285-301, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25779870

RESUMEN

Pheromones are substances released from animals that, when detected by the vomeronasal organ of other individuals of the same species, affect their physiology and behavior. Pheromone binding to receptors on microvilli on the dendritic knobs of vomeronasal sensory neurons activates a second messenger cascade to produce an increase in intracellular Ca(2+) concentration. Here, we used whole-cell and inside-out patch-clamp analysis to provide a functional characterization of currents activated by Ca(2+) in isolated mouse vomeronasal sensory neurons in the absence of intracellular K(+). In whole-cell recordings, the average current in 1.5 µM Ca(2+) and symmetrical Cl(-) was -382 pA at -100 mV. Ion substitution experiments and partial blockade by commonly used Cl(-) channel blockers indicated that Ca(2+) activates mainly anionic currents in these neurons. Recordings from inside-out patches from dendritic knobs of mouse vomeronasal sensory neurons confirmed the presence of Ca(2+)-activated Cl(-) channels in the knobs and/or microvilli. We compared the electrophysiological properties of the native currents with those mediated by heterologously expressed TMEM16A/anoctamin1 or TMEM16B/anoctamin2 Ca(2+)-activated Cl(-) channels, which are coexpressed in microvilli of mouse vomeronasal sensory neurons, and found a closer resemblance to those of TMEM16A. We used the Cre-loxP system to selectively knock out TMEM16A in cells expressing the olfactory marker protein, which is found in mature vomeronasal sensory neurons. Immunohistochemistry confirmed the specific ablation of TMEM16A in vomeronasal neurons. Ca(2+)-activated currents were abolished in vomeronasal sensory neurons of TMEM16A conditional knockout mice, demonstrating that TMEM16A is an essential component of Ca(2+)-activated Cl(-) currents in mouse vomeronasal sensory neurons.


Asunto(s)
Potenciales de Acción , Canales de Cloruro/metabolismo , Células Receptoras Sensoriales/metabolismo , Órgano Vomeronasal/metabolismo , Animales , Anoctamina-1 , Calcio/metabolismo , Células Cultivadas , Canales de Cloruro/genética , Eliminación de Gen , Ratones , Células Receptoras Sensoriales/fisiología , Órgano Vomeronasal/citología
18.
3 Biotech ; 5(5): 597-609, 2015 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-28324530

RESUMEN

Second-generation bioethanol can be produced from various lignocellulosic biomasses such as wood, agricultural or forest residues. Lignocellulosic biomass is inexpensive, renewable and abundant source for bioethanol production. The conversion of lignocellulosic biomass to bioethanol could be a promising technology though the process has several challenges and limitations such as biomass transport and handling, and efficient pretreatment methods for total delignification of lignocellulosics. Proper pretreatment methods can increase concentrations of fermentable sugars after enzymatic saccharification, thereby improving the efficiency of the whole process. Conversion of glucose as well as xylose to bioethanol needs some new fermentation technologies to make the whole process inexpensive. The main goal of pretreatment is to increase the digestibility of maximum available sugars. Each pretreatment process has a specific effect on the cellulose, hemicellulose and lignin fraction; thus, different pretreatment methods and conditions should be chosen according to the process configuration selected for the subsequent hydrolysis and fermentation steps. The cost of ethanol production from lignocellulosic biomass in current technologies is relatively high. Additionally, low yield still remains as one of the main challenges. This paper reviews the various technologies for maximum conversion of cellulose and hemicelluloses fraction to ethanol, and it point outs several key properties that should be targeted for low cost and maximum yield.

19.
Dev Neurobiol ; 74(7): 657-75, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24318978

RESUMEN

Calcium-activated chloride channels are involved in several physiological processes including olfactory perception. TMEM16A and TMEM16B, members of the transmembrane protein 16 family (TMEM16), are responsible for calcium-activated chloride currents in several cells. Both are present in the olfactory epithelium of adult mice, but little is known about their expression during embryonic development. Using immunohistochemistry we studied their expression in the mouse olfactory epithelium at various stages of prenatal development from embryonic day (E) 12.5 to E18.5 as well as in postnatal mice. At E12.5, TMEM16A immunoreactivity was present at the apical surface of the entire olfactory epithelium, but from E16.5 became restricted to a region near the transition zone with the respiratory epithelium, where localized at the apical part of supporting cells and in their microvilli. In contrast, TMEM16B immunoreactivity was present at E14.5 at the apical surface of the entire olfactory epithelium, increased in subsequent days, and localized to the cilia of mature olfactory sensory neurons. These data suggest different functional roles for TMEM16A and TMEM16B in the developing as well as in the postnatal olfactory epithelium. The presence of TMEM16A at the apical part and in microvilli of supporting cells is consistent with a role in the regulation of the chloride ionic composition of the mucus covering the apical surface of the olfactory epithelium, whereas the localization of TMEM16B to the cilia of mature olfactory sensory neurons is consistent with a role in olfactory signal transduction.


Asunto(s)
Canales de Cloruro/metabolismo , Mucosa Olfatoria/crecimiento & desarrollo , Mucosa Olfatoria/metabolismo , Animales , Anoctamina-1 , Anoctaminas , Desarrollo Embrionario , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Inmunohistoquímica , Ratones Endogámicos C57BL , Ratones Transgénicos , Microscopía Confocal , Microvellosidades/metabolismo , Mucosa Olfatoria/embriología , Neuronas Receptoras Olfatorias/embriología , Neuronas Receptoras Olfatorias/crecimiento & desarrollo , Neuronas Receptoras Olfatorias/metabolismo , Neuronas Receptoras Olfatorias/ultraestructura
20.
World J Microbiol Biotechnol ; 29(12): 2353-7, 2013 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-23764955

RESUMEN

In the current study, ferritic stainless grades AISI 439 and AISI 444 were investigated as possible construction materials for machinery and equipment in the cane-sugar industry. Their performance in corrosive cane-sugar juice environment was compared with the presently used low carbon steel AISI 1010 and austenitic stainless steel AISI 304. The Tafel plot electrochemical technique was used to evaluate general corrosion performance. Microbiologically influenced corrosion (MIC) behaviour in sugarcane juice environment was studied. Four microbial colonies were isolated from the biofilms on the metal coupon surfaces on the basis of their different morphology. These were characterized as Brevibacillus parabrevis, Bacillus azotoformans, Paenibacillus lautus and Micrococcus sp. The results of SEM micrographs showed that AISI 439 and AISI 304 grades had suffered maximum localized corrosion. MIC investigations revealed that AISI 444 steel had the best corrosion resistance among the tested materials. However from the Tafel plots it was evident that AISI 1010 had the least corrosion resistance and AISI 439 the best corrosion resistance.


Asunto(s)
Bacterias/clasificación , Bacterias/aislamiento & purificación , Saccharum/microbiología , Acero Inoxidable/química , Bacillus/aislamiento & purificación , Biopelículas , Brevibacillus/aislamiento & purificación , Corrosión , Técnicas Electroquímicas , Ensayo de Materiales , Micrococcus/aislamiento & purificación , Paenibacillus/aislamiento & purificación , Preparaciones de Plantas
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA