15 research outputs found
Adsorption of proteins to thin-films of PDMS and its effect on the adhesion of human endothelial cells
This paper describes a simple and inexpensive procedure to produce thin-films of poly(dimethylsiloxane). Such films were characterized by a variety of techniques (ellipsometry, nuclear magnetic resonance, atomic force microscopy, and goniometry) and used to investigate the adsorption kinetics of three model proteins (fibrinogen, collagen type-I, and bovine serum albumin) under different conditions. The information collected from the protein adsorption studies was then used to investigate the adhesion of human dermal microvascular endothelial cells. The results of these studies suggest that these films can be used to model the surface properties of microdevices fabricated with commercial PDMS. Moreover, the paper provides guidelines to efficiently attach cells in BioMEMS devices.Fil: Chumbimuni Torres, Karin Y.. The University of Texas at San Antonio; Estados UnidosFil: Coronado, Ramon E.. The University of Texas at San Antonio; Estados UnidosFil: Mfuh, Adelphe M.. The University of Texas at San Antonio; Estados UnidosFil: Castro Guerrero, Carlos. The University of Texas at San Antonio; Estados UnidosFil: Silva, MarĂa Fernanda. Consejo Nacional de Investigaciones CientĂficas y TĂŠcnicas. Centro CientĂfico TecnolĂłgico Conicet - Mendoza. Instituto de BiologĂa AgrĂcola de Mendoza. Universidad Nacional de Cuyo. Facultad de Ciencias Agrarias. Instituto de BiologĂa AgrĂcola de Mendoza; ArgentinaFil: Negrete, George R.. The University of Texas at San Antonio; Estados UnidosFil: Bizios, Rena. The University of Texas at San Antonio; Estados UnidosFil: Garcia, Carlos D.. The University of Texas at San Antonio; Estados Unido
Small Molecule Interactome Mapping by Photoaffinity Labeling Reveals Binding Site Hotspots for the NSAIDs
Many
therapeutics elicit cell-type specific polypharmacology that
is executed by a network of molecular recognition events between a
small molecule and the whole proteome. However, measurement of the
structures that underpin the molecular associations between the proteome
and even common therapeutics, such as the nonsteroidal anti-inflammatory
drugs (NSAIDs), is limited by the inability to map the small molecule
interactome. To address this gap, we developed a platform termed small
molecule interactome mapping by photoaffinity labeling (SIM-PAL) and
applied it to the in cellulo direct characterization of specific NSAID
binding sites. SIM-PAL uses (1) photochemical conjugation of NSAID
derivatives in the whole proteome and (2) enrichment and isotope-recoding
of the conjugated peptides for (3) targeted mass spectrometry-based
assignment. Using SIM-PAL, we identified the NSAID interactome consisting
of over 1000 significantly enriched proteins and directly characterized
nearly 200 conjugated peptides representing direct binding sites of
the photo-NSAIDs with proteins from Jurkat and K562 cells. The enriched
proteins were often identified as parts of complexes, including known
targets of NSAID activity (e.g., NF-ÎşB) and novel interactions
(e.g., AP-2, proteasome). The conjugated peptides revealed direct
NSAID binding sites from the cell surface to the nucleus and a specific
binding site hotspot for the three photo-NSAIDs on histones H2A and
H2B. NSAID binding stabilized COX-2 and histone H2A by cellular thermal
shift assay. Since small molecule stabilization of protein complexes
is a gain of function regulatory mechanism, it is conceivable that
NSAIDs affect biological processes through these broader proteomic
interactions. SIM-PAL enabled characterization of NSAID binding site
hotspots and is amenable to map global binding sites for virtually
any molecule of interest
Scalable, Metal- and Additive-Free, Photoinduced Borylation of Haloarenes and Quaternary Arylammonium Salts
We
report herein a simple, metal- and additive-free, photoinduced
borylation of haloarenes, including electron-rich fluoroarenes, as
well as arylammonium salts directly to boronic acids. This borylation
method has a broad scope and functional group tolerance. We show that
it can be further extended to boronic esters and carried out on gram
scale as well as under flow conditions
Concise Total Synthesis of Trichodermamides A, B, and C Enabled by an Efficient Construction of the 1,2-Oxazadecaline Core
We
report herein a facile and efficient method of the construction
of the <i>cis</i>-1,2-oxazadecaline system, distinctive
of (pre)Âtrichodermamides, aspergillazine A, gliovirin, and FA-2097.
The formation of the 1,2-oxazadecaline core was accomplished by a
1,2-addition of an Îą<i>C</i>-lithiated <i>O</i>-silyl ethyl pyruvate oxime to benzoquinone, which is followed by
an oxa-Michael ring-closure. The method was successfully applied to
the concise total synthesis of trichodermamide A (in gram quantities)
and trichodermamide B, as well as the first synthesis of trichodermamide
C
Cytotoxicity and Mechanism of Action of the Marine-Derived Fungal Metabolite Trichodermamide B and Synthetic Analogues
The trichodermamides are modified
dipeptides isolated from a wide
variety of fungi, including <i>Trichoderma virens</i>. Previous
studies reported that trichodermamide B (<b>2</b>) initiated
cytotoxicity in HCT-116 colorectal cancer cells, while trichodermamide
A (<b>1</b>) was devoid of activity. We recently developed an
efficient total synthesis for the trichodermamides AâC (<b>1</b>â<b>3</b>). Multiple intermediates and analogues
were produced, and they were evaluated for biological effects to identify
additional structureâactivity relationships and the possibility
that a simplified analogue would retain the biological effects of <b>2</b>. The antiproliferative effects of 18 compounds were evaluated,
and the results show that <b>2</b> and four other compounds
are active in HeLa cells, with IC<sub>50</sub> values in the range
of 1.4â21 ÎźM. Mechanism of action studies of <b>2</b> and the other active analogues revealed different spectra of activity.
At the IC<sub>85</sub> concentration, <b>2</b> caused S-phase
accumulation and cell death in HeLa cells, suggesting response to
DNA double-strand breaks. The analogues did not cause S-phase accumulation
or induction of DNA damage repair pathways, consistent with an alternate
mode of action. The mechanistic differences are hypothesized to be
due to the chlorohydrin moiety in <b>2</b>, which is lacking
in the analogues, which could form a DNA-reactive epoxide
Additive- and Metal-Free, Predictably 1,2- and 1,3-Regioselective, Photoinduced Dual CâH/CâX Borylation of Haloarenes
We
report herein a simple, additive- and metal-free, photoinduced,
dual CâH/CâX borylation of chloro-, bromo-, and iodoarenes.
The reaction produces 1,2- and 1,3-diborylarenes on gram scales under
batch and continuous flow conditions. The regioselectivity of the
dual CâH/CâX borylation is determined by the solvent
and the substituents in the parent haloarenes
Additive- and Metal-Free, Predictably 1,2- and 1,3-Regioselective, Photoinduced Dual CâH/CâX Borylation of Haloarenes
We
report herein a simple, additive- and metal-free, photoinduced,
dual CâH/CâX borylation of chloro-, bromo-, and iodoarenes.
The reaction produces 1,2- and 1,3-diborylarenes on gram scales under
batch and continuous flow conditions. The regioselectivity of the
dual CâH/CâX borylation is determined by the solvent
and the substituents in the parent haloarenes
Identification of Inhibitors of CD36-Amyloid Beta Binding as Potential Agents for Alzheimerâs Disease
Neuroinflammation
is one of the hallmarks of Alzheimerâs
disease pathology. Amyloid β has a central role in microglia
activation and the subsequent secretion of inflammatory mediators
that are associated with neuronal toxicity. The recognition of amyloid
β by microglia depends on the expression of several receptors
implicated in the clearance of amyloid and in cell activation. CD36
receptor expressed on microglia interacts with fibrils of amyloid
inducing the release of proinflammatory cytokines and amyloid internalization.
The interruption of the interaction CD36-amyloid β compromises
the activation of microglia cells. We have developed and validated
a new colorimetric assay to identify potential inhibitors of the binding
of amyloid β to CD36. We have found seven molecules, structural
analogues of the Trichodermamide family of natural products that interfere
with the interaction CD36-amyloid β. By combining molecular
docking and dynamics simulations, we suggested the second fatty acids
binding site within the large luminal hydrophobic tunnel, present
in the extracellular domain of CD36, as the binding pocket of these
compounds. Free energy calculations predicted the nonpolar component
as the driving force for the binding of these inhibitors. These molecules
also inhibited the production of TNF-ι, IL-6, and IL-1β
by peritoneal macrophages stimulated with fibrils of amyloid β.
This work serves as a platform
for the identification of new potential anti-inflammatory agents for
the treatment of Alzheimerâs disease