Unraveling the Role of RNA-Binding Proteins, with a Focus on RPS5, in the Malignant Progression of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) represents a major global health concern, demanding a thorough understanding of its molecular mechanisms for effective therapeutic strategies. RNA-binding proteins (RBPs) play critical roles in post-transcriptional gene regulation, with their dysregulation increasingly recognized as a hallmark of various cancers. However, the specific contributions of RBPs to HCC pathogenesis and prevention remain incompletely understood. In this study, we systematically conducted an examination of the expression profiles and clinical relevance of RBPs in 556 clinical samples from well-established cohorts. Through comprehensive analyses, a subset of RBPs exhibiting significant overexpression in HCC was identified, establishing a noteworthy correlation between their aberrant expression and HCC progression. Furthermore, 40S ribosomal protein S5 (RPS5), a ribosomal protein, emerged as a potential key contributor in HCC progression. Rigorous analyses established a correlation between elevated RPS5 expression and advanced clinicopathological features, suggesting its potential as a prognostic biomarker. Experiments further confirmed the impact of RPS5 on pivotal cellular processes implicated in cancer progression, including cell proliferation and metastasis. Further mechanistic studies unveiled the potential of RPS5 to activate the cell cycle by binding to key molecules involved in the pathway, thereby promoting the malignant progression of HCC. Additionally, our analysis of the etiology behind RPS5 overexpression in HCC posited it as an outcome of transcriptional regulation by the transcription factors Nuclear Respiratory Factor 1 (NRF1) and MYC-associated zinc finger protein (MAZ). In conclusion, our study contributes to the growing evidence elucidating the intricate involvement of RBPs, exemplified by RPS5, in the malignant progression of HCC. The integration of genomic, transcriptomic, and functional analyses provides a comprehensive understanding of the regulatory mechanisms associated with RPS5 in HCC. This comprehensive analysis not only advances our knowledge of the molecular drivers behind HCC but also highlights the potential therapeutic relevance of targeting RBPs and their regulatory network for the development of more effective treatment strategies.

Proteomic Analysis of Subchronic Furan Exposure in the Liver of Male Fischer F344 Rats.

Furan is a volatile compound formed during the thermal processing of foods. Chronic exposure has been shown to cause cholangiocarcinoma and hepatocellular tumors in rodent models. We conducted a 90 day subchronic study in Fisher 344 rats exposed to various doses by gavage to determine the NOAEL. Previous reports have outlined changes in the liver using gross necropsy examination, histopathology, clinical biochemistry, hematology, immunohistochemistry, and toxicogenomics. The data revealed that males were more sensitive than females. The focus of this study was to evaluate the toxicoproteomic changes by 2-dimensional differential in gel electrophoresis followed by mass spectrometry analysis. To compliment previous studies, protein expression changes were evaluated of male animals after 90 days of exposure to doses of 0, 0.03, 0.5, and 8.0 mg/kg bw/d. Significant statistical treatment-related changes compared to the controls identified 45 protein spots containing 38 unique proteins. Proteins identified are implicated in metabolism, redox regulation, protein folding/proteolysis as well as structural and transport proteins. At lower doses, multiple cytoprotective pathways are activated to maintain a homeostasis but ultimately the loss of protein function and impairment of several pathways could lead to adverse health effects at higher doses of furan administration.

Entrapment of functionalized silica microspheres with photo-initiated acrylate-based polymers.

The entrapment of silica-based microspheres, commonly used as stationary phases in chromatography, with an organic porous polymer based on poly(butyl acrylate-co-1,3-butanediol diacrylate) was explored. The spheres were immobilized by photopolymerization leading to entrapped beds within 75 microm i.d. fused silica capillaries, and were mechanically stable, resisting pressure drops of over 5600 psi (38.6 MPa) for only 1 cm of material. The morphology of the polymer formation around the spheres was investigated by SEM and corroborated with back pressure measurements, which indicated that the spheres were held together by encapsulating polymer. The entrapped material was extruded from the capillary in some cases to facilitate imaging. The entrapment conditions were explored, varying the polarity of the sphere surface, the solvent, and the monomers, revealing that polymer formation is based on partitioning of the monomers between the surface and solvent. The resulting polymer morphology is discussed with respect to the effects of confinement, supported by experiments with varying microsphere diameters. The columns described here have favourable properties for use in capillary chromatography and supported catalysis among other applications, and is suitable for lab-on-a-chip devices.

Microsphere entrapped emitters for sample preconcentration and electrospray ionization mass spectrometry.

In this study a nano-electrospray emitter is constructed by precisely positioning entrapped octadecylsilane (ODS) particles within a photoinitiated polymer at the exit aperture of a capillary. Following poly-merization, the microsphere/polymer hybrid material is able to withstand pressures greater than 4000 psi for 1 cm length of material. Smaller microspheres (3 microm) patterned at the exit aperture of a capillary generated the most sensitive/stable electrospray from 100 to 1000 nL/min and moderately stable signal under 100 nL/min. Constant infusion of a standard PPG solution from a batch of eleven emitters resulted in a relatively small variance in total ion current (TIC) counts (8%). The entrapped microsphere emitter design yields an emitter that minimizes clogging and eliminates dead volume between the chromatographic bed and the electrospray emitter. The entrapped ODS microspheres can also be used for sample preparation prior to mass spectrometry (MS) analysis. We show the solid-phase extraction and preconcentration of 20-700 fmol of a peptide (leucine enkephalin) prior to MS analysis on an emitter with 1 cm of entrapped microspheres.

Photoinduced polymerization for entrapping of octadecylsilane microsphere columns for capillary electrochromatography.

A novel fritless capillary column for capillary electrochromatography (CEC) has been developed. The ODS microspheres were packed into a capillary and were then immobilized within an organic polymer prepared in situ through a photopolymerization process. The entrapment conditions were investigated to minimize the effect of the polymer matrix on the chromatographic properties of the packing material. The organic polymer matrix in the microsphere-packed column functions to link microspheres at specific sphere-sphere and sphere-capillary contact points. CEC separations of a PAH test mixture using entrapped columns with different UV illumination times were compared in terms of retention factor and separation efficiency. The optimized entrapped column demonstrated better chromatographic performance than similarly packed columns with conventional inlet and outlet frits. The electrochromatographic separations of hormones and peptides were also demonstrated on entrapped ODS columns.

Capillary-based solid-phase extraction with columns prepared using different bead trapping methods.

Three approaches of bead immobilization for CEC column preparation in a capillary were examined for SPE. The three approaches included a packed column with a single frit, a packed column with an inlet and outlet frit, and an entrapped column where beads were immobilized within an organic polymer. A direct comparison of SPE/preconcentration of 4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza-s-indacene and 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-propionic acid with a 2 cm long bed showed that the entrapped column yielded the best performance in terms of reproducibility and robustness. The room temperature chemistry utilized to form the entrapped column enables the column to be photopatterned anywhere within the capillary without loss in bead functionality, and effectively links individual beads to one another at specific bead-bead and bead-capillary contact points. A 0.5 cm long entrapped bed exhibits high mechanical strength and is able to withstand >4400 psi. The entrapped bed was used to preconcentrate progesterone and beta-estradiol providing signal enhancements of >600. Following preconcentration, the hormones could be separated using CEC. With the current availability of numerous well-characterized chromatographic packing materials and the relative simplicity of the fabrication method, this methodology can be readily adapted to HPLC, CEC, and micro total analysis system.