Transcriptome Profile of Thyroid Glands in Bile Duct Ligation Mouse Model

Thyroid hormone (TH) contributes to multiple cellular mechanisms in the liver, muscle cells, adipose tissue, and brain, etc. In particular, the liver is an important organ in TH metabolism for the conversion of thyronine (T4) into triiodothyronine (T3) by the deiodinase enzyme. TH levels were significantly decreased and thyroid-stimulating hormone (TSH) levels were significantly increased in patients with liver failure compared with normal subjects. Among liver failure diseases, hepatic encephalopathy (HE) deserves more attention because liver damage and neuropathologies occur simultaneously. Although there is numerous evidence of TH dysregulation in the HE model, specific mechanisms and genetic features of the thyroid glands in the HE model are not fully understood. Here, we investigated the significantly different genes in the thyroid glands of a bile duct ligation (BDL) mouse model as the HE model, compared to the thyroid glands of the control mouse using RNA sequencing. We also confirmed the alteration in mRNA levels of thyroid gland function-related genes in the BDL mouse model. Furthermore, we evaluated the increased level of free T4 and TSH in the BDL mouse blood. Thus, we emphasize the potential roles of TH in liver metabolism and suggest that thyroid dysfunction-related genes in the HE model should be highlighted for finding the appropriate solution for an impaired thyroid system in HE.     

Bioorthogonal Probes for Analyzing Non-Enzymatic Post-Translational Modifications

Non-enzymatic post-translational modifications (nPTMs) have been proposed as indicators of cellular stresses and diseases. Unfortunately, direct assessment of nPTMs in native environment is extremely challenging due to the heterogeneity of adducts and the lack of tagging tools. Given these challenges, bioorthogonal probes (BPs) have been developed for the analysis of nPTMs. The rationality is that BPs could selectively install azides or alkynes into nPTMs as a biorthogonal handle for the following enrichment or tracking. Herein, we review the state-of-art of BPs used for nPTMs studies, clarify their working principles, and highlight how they advance our understanding of the biological functions of nPTMs.     

Nonswelling,Ultralow Content Inverse Electron‐Demand Diels–Alder Hyaluronan Hydrogels with Tunable Gelation Time: Synthesis and In Vitro Evaluation

Hyaluronan (HA) is a major component of the extracellular matrix and is particularly attractive for cell‐based assays; yet, common crosslinking strategies of HA hydrogels are not fully tunable and bioorthogonal, and result in gels subject to swelling, which affects their physicochemical properties. To overcome these limitations, HA hydrogels based on the inverse electron‐demand Diels–Alder (IEDDA) “click” reaction are designed. By crosslinking two modified HA components together, as opposed to using telechelic components, tunable gelation times as fast as 4.4 ± 0.4 min and as slow as 46.2 ± 1.8 min are achieved for facile use. By optimizing HA molar mass, ultralow polymer content hydrogels of 0.5% (w/v), resulting in minimal (<3–5% mass variation) to nonswelling (<1%), transparent and biodegradable hydrogels are synthesized. To demonstrate their versatility, the newly designed hydrogels are tested as matrices for 3D cell culture and retinal explant imaging where transparency is important. IEDDA hydrogels are cytocompatible with primary photoreceptors and enable multiphoton imaging of embedded retinal explants for double the time (>38 h) than agarose thermogels (<20 h). IEDDA HA hydrogels constitute a new hydrogel platform. They have low polymer content, tunable gelation time, and are stable, thereby making them suitable for a diversity of applications.     

Synthesis and biological characterisation of targeted pro-apoptotic peptide

We report herein the synthesis and in vitro assay of new, multimeric RGD-peptide conjugates for cell-targeted drug delivery. We generated a peptide scaffold comprising two functional domains, one a tumour blood vessel "homing" motif and the other a programmed cell-death-inducing peptide sequence. RGD peptides were selected to direct the molecular conjugate to alpha(V)beta(3) integrin-containing tumour cells. The pro-apoptotic (Lys-Leu-Ala-Lys-Leu-Ala-Lys)(2) peptide was found to be nontoxic outside cells, but toxic when internalized into targeted cells as it disrupted the mitochondrial membrane. The synthesis of these targeted pro-apoptotic conjugates was carried out by assembling three different units (that is, scaffold, RGD units and pro-apoptotic peptide) through chemoselective ligations. We show that one compound displays significant biological effect in alpha(V)beta(3) integrin-containing tumour cells.     

Efficient and Chemoselective Surface Immobilization of Proteins by Using Aniline‐Catalyzed Oxime Chemistry

Site‐specific immobilization of peptides and proteins is crucial to ensure their functionality in surface‐based assays. We report the use of aniline‐catalyzed oxime ligations as a very efficient and broadly applicable method to covalently attach the N terminus of proteins and peptides to a surface functionalized with alkoxy‐amine groups.

     

Chemical Synthesis and Structure of the Prokineticin Bv8

Bv8, a 77‐residue protein isolated from frogs, is the prototypic member of the prokineticin family of cytokines. Prokineticins (PKs) have only recently been identified in vertebrates (including humans), and they are believed to be involved in a number of key physiological processes, such as angiogenesis, neurogenesis, nociception, and tissue development. We used a combination of Boc solid‐phase peptide synthesis, native chemical ligation, and in vitro protein folding to establish robust chemical access to this molecule. Synthetic Bv8 was obtained in good yield and exhibited full activity in a human neuroblastoma cell line and rat dorsal root ganglion (DRG) neurons. The 3D structure of the synthetic protein was determined by using NMR spectroscopy and it was found to be homologous with that of mamba intestinal toxin 1, which is the only other known prokineticin structure. Analysis of a truncated mutant lacking five residues at the N terminus that are critical for receptor binding and activation showed no perturbation to the core protein structure. Together with the functional data, this suggests that receptor binding is likely to be a highly cooperative process possibly involving major allosterically driven structural rearrangements. The facile and efficient synthesis presented here will enable preparation of unique chemical analogues of prokineticins, which should be powerful tools for modulating the structure and function of prokineticins and their receptors, and studying the many physiological processes that have been linked to them.