Rapid LC-MS/MS Evaluation of Collagen and Elastin Crosslinks in Human and Mouse Lung Tissue with a Novel Bioanalytical Surrogate Matrix Approach.

Alterations to post-translational crosslinking modifications in the extracellular matrix (ECM) are known to drive the pathogenesis of fibrotic diseases, including idiopathic pulmonary fibrosis (IPF). Thus, the methodology for measuring crosslinking dynamics is valuable for understanding disease progression. The existing crosslinking analysis sample preparation and liquid chromatography tandem mass spectrometry (LC-MS/MS) methods are typically labor-intensive and time-consuming which limits throughput.

Selective inhibition of integrin αvβ6 leads to rapid induction of urinary bladder tumors in cynomolgus macaques.

Administration of a novel and selective small molecule integrin αvβ6 inhibitor, MORF-627, to young cynomolgus monkeys for 28 days resulted in the rapid induction of epithelial proliferative changes in the urinary bladder of 2 animals, in the absence of test agent genotoxicity. Microscopic findings included suburothelial infiltration by irregular nests and/or trabeculae of epithelial cells, variable cytologic atypia, and high mitotic rate, without invasion into the tunica muscularis. Morphologic features and patterns of tumor growth were consistent with a diagnosis of early-stage invasive urothelial carcinoma.

Collagen-producing lung cell atlas identifies multiple subsets with distinct localization and relevance to fibrosis.

Collagen-producing cells maintain the complex architecture of the lung and drive pathologic scarring in pulmonary fibrosis. Here we perform single-cell RNA-sequencing to identify all collagen-producing cells in normal and fibrotic lungs. We characterize multiple collagen-producing subpopulations with distinct anatomical localizations in different compartments of murine lungs.

Identification of Positive Allosteric Modulators of the D Dopamine Receptor That Act at Diverse Binding Sites.

The D dopamine receptor is linked to a variety of neuropsychiatric disorders and represents an attractive drug target for the enhancement of cognition in schizophrenia, Alzheimer disease, and other disorders. Positive allosteric modulators (PAMs), with their potential for greater selectivity and larger therapeutic windows, may represent a viable drug development strategy, as orthosteric D receptor agonists possess known clinical liabilities. We discovered two structurally distinct D receptor PAMs, MLS6585 and MLS1082, via a high-throughput screen of the NIH Molecular Libraries program small-molecule library.

Dopamine D2 receptor overexpression alters behavior and physiology in Drd2-EGFP mice.

Bacteria artificial chromosome (BAC) transgenic mice expressing the reporter protein enhanced green fluorescent protein (EGFP) under the control of the D1 and D2 dopamine receptor promoters (Drd1-EGFP and Drd2-EGFP) have been widely used to study striatal function and have contributed to our understanding of the physiological and pathological functions of the basal ganglia. These tools were produced and promptly made available to address questions in a cell-specific manner that has transformed the way we frame hypotheses in neuroscience. However, these mice have not been fully characterized until now.

Identifying novel protein-protein interactions using co-immunoprecipitation and mass spectroscopy.

Proteomics has evolved from genomic science due to the convergence of advances in protein chemistry, separations, mass spectroscopy, and peptide and protein databases. Where identifying protein-protein interactions was once limited to yeast two-hybrid analyses or empirical data, protein-protein interactions can now be examined in both cells and native tissues by precipitation of the protein complex of interest. Coupling this field to receptor pharmacology has recently allowed for the identification of proteins that differentially and selectively interact with receptors and are integral to their biological effects.

Reciprocal modulation of function between the D1 and D2 dopamine receptors and the Na+,K+-ATPase.

It is well documented that dopamine can increase or decrease the activity of the Na+,K+-ATPase (NKA, sodium pump) in an organ-specific fashion. This regulation can occur, at least partially, via receptor-mediated second messenger activation and can promote NKA insertion or removal from the plasma membrane. Using co-immunoprecipitation and mass spectrometry, we now show that, in both brain and HEK293T cells, D1 and D2 dopamine receptors (DARs) can exist in a complex with the sodium pump.

D1 and D2 dopamine receptor expression is regulated by direct interaction with the chaperone protein calnexin.

As for all proteins, G protein-coupled receptors (GPCRs) undergo synthesis and maturation within the endoplasmic reticulum (ER). The mechanisms involved in the biogenesis and trafficking of GPCRs from the ER to the cell surface are poorly understood, but they may involve interactions with other proteins. We have now identified the ER chaperone protein calnexin as an interacting protein for both D(1) and D(2) dopamine receptors.

His452Tyr polymorphism in the human 5-HT2A receptor destabilizes the signaling conformation.

Naturally occurring variation within the human 5-HT(2A) receptor results in an amino acid substitution in the carboxyl terminus of the receptor. This single nucleotide polymorphism (SNP), encoding a His452Tyr substitution, occurs at a frequency of 9% in the general population. It is noteworthy that this SNP has been linked to attention deficit hyperactivity disorder and has been associated with schizophrenic patients that do not respond to treatment with clozapine.