Difficulties in early-stage diagnosis are among the factors contributing to the high mortality of nonsmall cell lung carcinoma (NSCLC) patients. Unfortunately, diagnostic biomarkers are currently lacking, limiting options in the clinic. To discover proteins that have potential for biomarker applications, we performed an in-depth quantitative proteomic analysis on a cohort of Filipino early-stage NSCLC lung adenocarcinoma (LUAD) patients. Differentially expressed proteins (DEPs) were obtained by using tandem mass tag (TMT) labeling and mass spectrometry (MS)-based quantitative proteomics. A total of 6240 quantified proteins were identified with 3155 significantly upregulated and 1248 significantly downregulated. Integration of the proteomic result with curated transcriptome data allowed the identification of 33 proteins with biomarker potential. This study also provided insights into relevant pathways in NSCLC LUAD, such as protein translation and metabolic pathways. Interestingly, all of the enzymes in the hexosamine biosynthetic pathway (HBP) are found to be upregulated, suggesting its important role in NSCLC LUAD. It is worthwhile to look at the potential of targeting the metabolic vulnerability of NSCLC LUAD as a new strategy in drug development. All MS data were deposited into ProteomeXchange with the identifier PXD050598.

Archaeal species encode a diversity of chromatin proteins that play distinct roles in genome compaction. Although our understanding of the individual proteins has been growing, their contributions to higher‐order folding in the archaeal chromosome remain to be explored. In this study, we investigated the chromatin structure of Thermoplasma volcanium, a euryarchaeon that encodes a heat‐unstable (HU) family protein (HUTvo) instead of histones. Atomic force microscopy revealed that the T. volcanium chromatin contains nucleoprotein structures of progressively increasing sizes, from 20 nm filaments to ~50 nm globules, indicative of higher‐order DNA folding. In vitro reconstitution showed that HUTvo wraps DNA similar to histones and forms highly compact yet disorganized nucleoprotein clusters on its own. Remarkably, HUTvo only dampens the formation of higher‐order structures by Alba. Our results provide an insight into how the interplay of chromatin proteins regulates genome compaction in histone‐free archaea.

Atomic force microscopy is a high-resolution imaging technique useful for observing the structures of biomolecular complexes. This approach provides a straightforward method to characterize the binding behavior of different chromatin architectural proteins and to analyze the increasingly complex structural units assembled on the DNA. The protocol describes the preparation, AFM imaging, and structural analysis of chromatin that is reconstituted in vitro using purified proteins and DNA. Here, we describe the successful application of the method on the chromatin architectural proteins of the archaeon Sulfolobus solfataricus.

Chitosan–acrylic acid (Cs–AA) hydrogels were prepared by blending different Cs concentrations and AA:Cs ratios. The Cs–AA hydrogel samples were analyzed using synchrotron radiation-induced near edge X-ray absorption fine structure (NEXAFS) and Fourier transform infrared (FTIR) spectroscopies. The characteristics of Cs were observed through the C, O, and N K edges in NEXAFS. Significant shifts in the Cs characteristic features were induced by the addition of AA. The NEXAFS findings confirmed the new covalent and ionic bridges to form the network of the Cs–AA hydrogel. Raw FTIR spectra were used to investigate the different functional groups of each of the macromolecules. The study attempted to discriminate the Cs–AA hydrogels prepared with different Cs wt% concentration and AA:Cs ratio using multivariate approaches. The preprocessed dataset was evaluated using principal component analysis (PCA) and hierarchical cluster analysis (HCA). The PCA and HCA methods showed the possibility of discriminating the Cs–AA hydrogel samples from the control samples and with each other. The approaches further confirmed that the CO groups found in the range of 1700–1800 cm−1 can be used as an identifier for Cs–AA hydrogel samples prepared using different solution parameters. The use of FTIR coupled with multivariate approaches provided a simple and reliable way to inspect the possible discrimination of Cs–AA hydrogels.

This study explores the effect of atmospheric pressure plasma (APP) treatment on chitosan-acrylic acid (Cs-AA) blends. The experiment involves a custom-built APP setup used to modify a polymeric blend composed of natural (Cs) and synthetic (AA) polymers. Even with a low process temperature (<40 °C), reactive oxygen and nitrogen species (RONS) were observed in an air plasma plume. The interaction of plasma with the liquid surface was also observed. With de-ionized water as the test liquid, the operating parameters such as the treatment time and flow rate were shown to influence the pH and absorption spectrum of the liquid. The presence of RONS was investigated using optical emission spectroscopy coupled with principal component analysis. The analysis revealed that the air plasma contains the different N systems, NO, OH, H α, monatomic N, and monatomic O species. Varying the gas flow rate influences the production of NO and OH radicals while measuring plasma discharge in different conditions (ambient air, DI H 2O, Cs, and Cs-AA blend) affects the concentration of the N positive and negative systems. The effect of these RONS on the Cs-AA blends was also investigated by assessing the chemical structure, pH, and viscosity of the solution. In correlation with all of the findings, it was observed that plasma treatment could degrade pure Cs solution by dehydrogenation and glycosidic bond cleaving. However, the addition of AA reduces the degradation so that the AA radicals created by plasma form a complex with the Cs that reduces Cs fragmentation and chain entanglement, as observed in the reduction of viscosity. In summary, the rich reactive species created by plasma in the Cs-AA solution not only provided stable species but also introduced more Cs-AA complexes.

TKIs targeting EGFR MT are standard for mNSCLC. Reports suggest co-expression of PD-L1 may play a role in acquiring TKIS resistance. This PDL1 prognostic role remains largely debatable. This study reports the influence of PD-L1 among EGFR MT mNSCLC Filipino cohort.

Chromatin compaction and regulation are essential processes for the normal function of all organisms, yet knowledge on how archaeal chromosomes are packed into higher-order structures inside the cell remains elusive. In this study, we investigated the role of archaeal architectural proteins Alba and Cren7 in chromatin folding and dynamics. Atomic force microscopy revealed that Sulfolobus solfataricus chromatin is composed of 28 nm fibers and 60 nm globular structures. In vitro reconstitution showed that Alba can mediate the formation of folded DNA structures in a concentration-dependent manner. Notably, it was demonstrated that Alba on its own can form higher-order structures with DNA. Meanwhile, Cren7 was observed to affect the formation of Alba-mediated higher-order chromatin structures. Overall, the results suggest an interplay between Alba and Cren7 in regulating chromatin compaction in archaea.

The use of materials to restore or replace the functions of damaged body parts has been proven historically. Any material can be considered as a biomaterial as long as it performs its biological function and does not cause adverse effects to the host. With the increasing demands for biofunctionality, biomaterials nowadays may not only encompass inertness but also specialized utility towards the target biological application. A hydrogel is a biomaterial with a 3D network made of hydrophilic polymers. It is regarded as one of the earliest biomaterials developed for human use. The preparation of hydrogel is often attributed to the polymerization of monomers or crosslinking of hydrophilic polymers to achieve the desired ability to hold large amounts of aqueous solvents and biological fluids. The generation of hydrogels, however, is shifting towards developing hydrogels through the aid of enabling technologies. This review provides the evolution of hydrogels and the different approaches considered for hydrogel preparation. Further, this review presents the plasma process as an enabling technology for tailoring hydrogel properties. The mechanism of plasma-assisted treatment during hydrogel synthesis and the current use of the plasma-treated hydrogels are also discussed.

This study explored the use of atmospheric pressure plasma in preparing a chitosan-acrylic acid (Cs-AA) hydrogel. Cs, the backbone of the hydrogel, has its structure confirmed using spectroscopic techniques such as Xray photoelectron spectroscopy, near edge X-ray fine structure spectroscopy, and Fourier transform infrared spectroscopy. The presence of C--O confirmed the presence of AA. AA promoted the formation of protonated amine. This creates the ionic linkage of the hydrogel network. Plasma treatment using compressed air created an inductive effect on the C-O species of the Cs backbone. Using N2 as working gas, there was an increase in the protonated amines in the Cs backbone, specially at high Cs concentration, aside from the inductive effect in the C-O species. This was confirmed from the shifts in the binding energy and photon energy values between the Cs and AA polymers. A combination of ionic and covalent linkages established the network of the Cs-AA hydrogel. The study showed that atmospheric pressure plasma treatment can be used not only for surface functionalization but also facilitating crosslinking of Cs and AA.

Despite advances in cancer treatment, breast cancer remains the second foremost cause of cancer mortality among women, with a high rate of relapse after initial treatment success. A subpopulation of highly malignant cancer cells, known as cancer stem cells (CSCs), is suspected to be linked to metastasis and relapse. Targeting of CSCs may therefore provide a means of addressing cancer-related mortality. However, due to their low population in vivo and a lack of proper culture platform for their propagation, much of the CSC biology remains unknown. Since maintenance of CSCs is heavily influenced by the tumor microenvironment, this study developed a 3D culture platform that mimics the metastatic tumor extracellular matrix (ECM) to effectively increase CSC population in vitro and allow CSC analysis. Through electrospinning, nanofibers that were aligned, porous, and collagen-coated were fabricated from polycaprolactone to recreate the metastatic tumor ECM assemblage. Breast cancer cells seeded onto the nanofiber scaffolds exhibited gross morphology and cytoskeletal phenotype similar to invasive cancer cells. Moreover, the population of breast cancer stem cells increased in nanofiber scaffolds. Analysis of breast cancer cells grown on the nanofiber scaffolds demonstrated an upregulation of mesenchymal markers and an increase in cell invasiveness suggesting the cells have undergone epithelial-mesenchymal transition. These results indicate that the fabricated nanofiber scaffolds effectively mimicked the tumor microenvironment that maintains the cancer stem cell population, offering a platform to enrich and analyze CSCs in vitro .

NSCLC remains to be the leading cause of cancer incidence and mortality in the Philippines. Early diagnosis, a better understanding of drug resistance mechanisms, and proper treatment monitoring are necessary to lower the incidence and mortality rate of NSCLC. As a proof of concept to investigate the proteomic profile of Filipino NSCLC, this study performed quantitative proteomic analysis to identify aberrant expressions in the tumor tissue relative to the adjacent normal tissue specimen of one Filipino NSCLC patient. A total of 4518 proteins were identified and from this, 1855 proteins were found to be significantly differentially expressed. Functional and pathway analyses were done to the 860 upregulated and downregulated proteins with at least four-fold expression change. The analysis revealed the possible activation of the upregulated protein MUC1, a known oncogenic driver involved in the stimulation of pathways that promote angiogenesis in NSCLC tumors and prevent apoptosis. Signal transducers and activators of transcription (STAT1 and STAT3), also found upregulated in the tumor specimen, are known to interact with MUC1, which results in the expression of proteins for cell proliferation. The analysis also suggests the stabilization of HIF, which may allow cell growth in the hypoxic tumor environment and aid in metabolic adaptation in tumor cells. The results also suggest the activation of EIF2 signaling that may lead to continuous translation of oncogenic proteins. Our data demonstrate the rich proteomic information that can be obtained from profiling of the NSCLC proteome and applications for a better understanding of NSCLC tumorigenicity. Further study should be performed on a larger cohort of Filipino NSCLC patients to provide the proteomic profile of Filipino NSCLC and extract information for biomarker and drug discovery. Data are available via ProteomeXchange with identifier PXD027710.

Archaeal species encode a variety of distinct lineage-specific chromosomal proteins. We have previously shown that inThermococcus kodakarensis, histone, Alba, and TrmBL2 play distinct roles in chromosome organization. Although our understanding of individual archaeal chromosomal proteins has been advancing, how archaeal chromosomes are folded into higher-order structures and how they are regulated are largely unknown. Here, we investigated the primary and higher-order structures of archaeal chromosomes from different archaeal lineages. Atomic force microscopy of chromosome spreads out ofThermoplasma acidophilumandPyrobaculum calidifontiscells revealed 10-nm fibers and 30-40-nm globular structures, suggesting the occurrence of higher-order chromosomal folding. Our results also indicated that chromosome compaction occurs toward the stationary phase. Micrococcal nuclease digestion indicated that fundamental structural units of the chromosome exist inT. acidophilumandT. kodakarensisbut not inP. calidifontisorSulfolobus solfataricus. In vitroreconstitution showed that, inT. acidophilum, the bacterial HU protein homolog HTa formed a 6-nm fiber by wrapping DNA, and that Alba was responsible for the formation of the 10-nm fiber by binding along the DNA without wrapping. Remarkably, Alba could form different higher-order complexes with histone or HTa on DNAin vitro. Mass spectrometry detected HTa and Rad50 in theT. acidophilumchromosome but not in other species. A putative transcriptional regulator of the AsnC/Lrp family (Pcal_1183) was detected on theP. calidifontischromosome, but not on that of other species studied. Putative membrane-associated proteins were detected in the chromosomes of the three archaeal species studied, includingT. acidophilum,P. calidifontis, andT. kodakarensis. Collectively, our data show that Archaea use different combinations of proteins to achieve chromosomal architecture and functional regulation.

Archaeal species encode a variety of distinct lineage-specific chromosomal proteins. We have previously shown that in Thermococcus kodakarensis, histone, Alba, and TrmBL2 play distinct roles in chromosome organization. Although our understanding of individual archaeal chromosomal proteins has been advancing, how archaeal chromosomes are folded into higher-order structures and how they are regulated are largely unknown. Here, we investigated the primary and higher-order structures of archaeal chromosomes from different archaeal lineages. Atomic force microscopy of chromosome spreads out of Thermoplasma acidophilum and Pyrobaculum calidifontis cells revealed 10-nm fibers and 30–40-nm globular structures, suggesting the occurrence of higher-order chromosomal folding. Our results also indicated that chromosome compaction occurs toward the stationary phase. Micrococcal nuclease digestion indicated that fundamental structural units of the chromosome exist in T. acidophilum and T. kodakarensis but not in P. calidifontis or Sulfolobus solfataricus. In vitro reconstitution showed that, in T. acidophilum, the bacterial HU protein homolog HTa formed a 6-nm fiber by wrapping DNA, and that Alba was responsible for the formation of the 10-nm fiber by binding along the DNA without wrapping. Remarkably, Alba could form different higher-order complexes with histone or HTa on DNA in vitro. Mass spectrometry detected HTa and Rad50 in the T. acidophilum chromosome but not in other species. A putative transcriptional regulator of the AsnC/Lrp family (Pcal_1183) was detected on the P. calidifontis chromosome, but not on that of other species studied. Putative membrane-associated proteins were detected in the chromosomes of the three archaeal species studied, including T. acidophilum, P. calidifontis, and T. kodakarensis. Collectively, our data show that Archaea use different combinations of proteins to achieve chromosomal architecture and functional regulation.

The organization and regulation of genomic DNA as nuclear chromatin is necessary for proper DNA function inside living eukaryotic cells. While this has been extensively explored, no true consensus is currently reached regarding the exact mechanism of chromatin organization. The traditional view has assumed that the DNA is packaged into a hierarchy of structures inside the nucleus based on the regular 30-nm chromatin fiber. This is currently being challenged by the fluid-like model of the chromatin which views the chromatin as a dynamic structure based on the irregular 10-nm fiber. In this review, we focus on the recent progress in chromatin structure elucidation highlighting the paradigm shift in chromatin folding mechanism from the classical textbook perspective of the regularly folded chromatin to the more dynamic fluid-like perspective.

Surfaces of polyimide (PI) sheets were modified using 13.56-MHz radio-frequency discharges to enhance their affinity with fibroblast cells. Physico-chemical analysis of pristine and plasma-treated PI sheets showed different responses against argon (Ar), oxygen (O2), and nitrogen (N2) as the plasma process gases. Overall, hydrophilicity of treated PI surfaces was enhanced, and its surface free energy increased from 53 dyn/cm to at least 73 dyn/ cm. Surface roughness values also increased from 1.3 to 40.8 nm, as demonstrated by atomic force microscopy analyses. Infrared spectral analyses showed a decrease of imide functional group peak intensities on plasma exposure, corresponding to chemical surface structural changes. In addition, plasma-treated surfaces significantly increased cell adhesion and proliferation compared to pristine samples. N2-containing plasma exhibited the greatest increase among the test gases, due to the possible inclusion of N2-based functional groups that enhance biochemical affinity of fibroblast cells.

Histones are highly conserved proteins among eukaryotes. However, yeast histones are more divergent in their sequences. In particular, the histone tail regions of the fission yeast, Schizosaccharomyces pombe, have fewer lysine residues, making their charges less positive than those of higher eukaryotes. In addition, the S. pombe chromatin lacks linker histones. How these factors affected yeast chromatin folding was analysed by biochemical reconstitution in combination with atomic force microscopy. Reconstitution of a nucleosome array showed that S. pombe chromatin has a more open structure similar to reconstituted human acetylated chromatin. The S. pombe nucleosomal array formed thinner fibers than those of the human nucleosomal array in the presence of mammalian linker histone H1. Such S. pombe fibers were more comparable to human acetylated fibers. These findings suggest that the core histone charges would determine the intrinsic characteristics of S. pombe chromatin and affect inter-nucleosomal interactions.