Int. J. Mol. Sci., Volume 24, Issue 19 (October-1 2023) – 628 articles

Breast cancer is a significant global concern, with tamoxifen, the standard treatment, raising long-term safety issues due to side effects. In this study, we evaluated the potential of five onoceranoid triterpenes from Lansium domesticum Corr. cv. kokosan against estrogen receptor alpha (ERα) using in silico techniques. Utilizing molecular docking, Lipinski’s rule of five, in silico ADMET, and molecular dynamics simulations, we assessed the potency of five onoceranoid triterpenes against ERα. Molecular docking indicated competitive binding energies for these triterpenes relative to the active form of tamoxifen (4OHT) and estradiol, an ERα native ligand. Three triterpenes met drug-likeness criteria with favorable ADMET profiles. Notably, 2 demonstrated superior binding affinity in molecular dynamics simulations, outperforming estradiol, closely followed by 3 and 4. Hierarchical clustering on principal components (HCPC) and the spatial distribution of contact surface area (CSA) analyses suggest that these triterpenes, especially 2, may act as antagonist ligands akin to 4OHT. These findings highlight the potential of onoceranoid triterpenes in treating ERα-related breast cancer. Full article

Major adverse cardiovascular events occurring upon coronary artery bypass graft surgery are typically accompanied by endothelial dysfunction. Total arterial revascularisation, which employs both left and right internal thoracic arteries instead of the saphenous vein to create a bypass, is associated with better mid- and long-term outcomes. We suggested that molecular profiles of human coronary artery endothelial cells (HCAECs) and human internal mammary artery endothelial cells (HITAECs) are coherent in terms of transcriptomic and proteomic signatures, which were then investigated by RNA sequencing and ultra-high performance liquid chromatography-mass spectrometry, respectively. Both HCAECs and HITAECs overexpressed molecules responsible for the synthesis of extracellular matrix (ECM) components, basement membrane assembly, cell-ECM adhesion, organisation of intercellular junctions, and secretion of extracellular vesicles. HCAECs were characterised by higher enrichment with molecular signatures of basement membrane construction, collagen biosynthesis and folding, and formation of intercellular junctions, whilst HITAECs were notable for augmented pro-inflammatory signaling, intensive synthesis of proteins and nitrogen compounds, and enhanced ribosome biogenesis. Despite HCAECs and HITAECs showing a certain degree of molecular heterogeneity, no specific markers at the protein level have been identified. Coherence of differentially expressed molecular categories in HCAECs and HITAECs suggests synergistic interactions between these ECs in a bypass surgery scenario. Full article

Zingiberales includes eight families and more than 2600 species, with many species having important economic and ecological value. However, the backbone phylogenetic relationships of Zingiberales still remain controversial, as demonstrated in previous studies, and molecular dating based on chloroplast genomes has not been comprehensively studied for the whole order. Herein, 22 complete chloroplast genomes from 21 species in Zingiberales were sequenced, assembled, and analyzed. These 22 genomes displayed typical quadripartite structures, which ranged from 161,303 bp to 163,979 bp in length and contained 111–112 different genes. The genome structures, gene contents, simple sequence repeats, long repeats, and codon usage were highly conserved, with slight differences among these genomes. Further comparative analysis of the 111 complete chloroplast genomes of Zingiberales, including 22 newly sequenced ones and the remaining ones from the national center for biotechnology information (NCBI) database, identified three highly divergent regions comprising ccsA, psaC, and psaC-ndhE. Maximum likelihood and Bayesian inference phylogenetic analyses based on chloroplast genome sequences found identical topological structures and identified a strongly supported backbone of phylogenetic relationships. Cannaceae was sister to Marantaceae, forming a clade that was collectively sister to the clade of (Costaceae, Zingiberaceae) with strong support (bootstrap (BS) = 100%, and posterior probability (PP) = 0.99–1.0); Heliconiaceae was sister to the clade of (Lowiaceae, Strelitziaceae), then collectively sister to Musaceae with strong support (BS = 94–100%, and PP = 0.93–1.0); the clade of ((Cannaceae, Marantaceae), (Costaceae, Zingiberaceae)) was sister to the clade of (Musaceae, (Heliconiaceae, (Lowiaceae, Strelitziaceae))) with robust support (BS = 100%, and PP = 1.0). The results of divergence time estimation of Zingiberales indicated that the crown node of Zingiberales occurred approximately 85.0 Mya (95% highest posterior density (HPD) = 81.6–89.3 million years ago (Mya)), with major family-level lineages becoming from 46.8 to 80.5 Mya. These findings proved that chloroplast genomes could contribute to the study of phylogenetic relationships and molecular dating in Zingiberales, as well as provide potential molecular markers for further taxonomic and phylogenetic studies of Zingiberales. Full article

The circadian clock is a regulatory system, with a periodicity of approximately 24 h, which generates rhythmic changes in many physiological processes, including mitochondrial activity. Increasing evidence links chronodisruption with aberrant functionality in clock gene expression, resulting in multiple diseases such as cancer. Melatonin, whose production and secretion oscillates according to the light–dark cycle, is the principal regulator of clock gene expression. In addition, the oncostatic effects of melatonin correlate with an increase in mitochondrial activity. However, the direct links between circadian clock gene expression, mitochondrial activity, and the antiproliferative effects of melatonin in cancers, including head and neck squamous cell carcinoma (HNSCC), remain largely unknown. In this study, we analyzed the effects of melatonin on HNSCC cell lines (Cal-27 and SCC9), which were treated with 500 and 1000 µM melatonin. We found that the antiproliferative effect of melatonin is not mediated by the Bmal1 clock gene. Additionally, high doses of melatonin were observed to result in resynchronization of oscillatory circadian rhythm genes (Per2 and Sirt1). Surprisingly, the resynchronizing effect of melatonin on Per2 and Sirt1 did not produce alterations in the oscillation of mitochondrial respiratory activity. These results increase our understanding of the possible antiproliferative mechanisms in melatonin in the treatment of head and neck squamous cell carcinoma and suggest that its antiproliferative effects are independent of clock genes but are directly related to mitochondrial activity. Full article

Drosophila CP190 and CP60 are transcription factors that are associated with centrosomes during mitosis. CP190 is an essential transcription factor and preferentially binds to housekeeping gene promoters and insulators through interactions with architectural proteins, including Su(Hw) and dCTCF. CP60 belongs to a family of transcription factors that contain the N-terminal MADF domain and the C-terminal BESS domain, which is characterized by the ability to homodimerize. In this study, we show that the conserved CP60 region adjacent to MADF is responsible for interacting with CP190. In contrast to the well-characterized MADF-BESS transcriptional activator Adf-1, CP60 is recruited to most chromatin sites through its interaction with CP190, and the MADF domain is likely involved in protein–protein interactions but not in DNA binding. The deletion of the Map60 gene showed that CP60 is not an essential protein, despite the strong and ubiquitous expression of CP60 at all stages of Drosophila development. Although CP60 is a stable component of the Su(Hw) insulator complex, the inactivation of CP60 does not affect the enhancer-blocking activity of the Su(Hw)-dependent gypsy insulator. Overall, our results indicate that CP60 has an important but redundant function in transcriptional regulation as a partner of the CP190 protein. Full article

ADP-ribosylation is a post-translational modification of proteins that plays a key role in various cellular processes, including DNA repair. Recently, significant progress has been made in understanding the mechanism and function of ADP-ribosylation in DNA repair. ADP-ribosylation can regulate the recruitment and activity of DNA repair proteins by facilitating protein–protein interactions and regulating protein conformations. Moreover, ADP-ribosylation can influence additional post-translational modifications (PTMs) of proteins involved in DNA repair, such as ubiquitination, methylation, acetylation, phosphorylation, and SUMOylation. The interaction between ADP-ribosylation and these additional PTMs can fine-tune the activity of DNA repair proteins and ensure the proper execution of the DNA repair process. In addition, PARP inhibitors have been developed as a promising cancer therapeutic strategy by exploiting the dependence of certain cancer types on the PARP-mediated DNA repair pathway. In this paper, we review the progress of ADP-ribosylation in DNA repair, discuss the crosstalk of ADP-ribosylation with additional PTMs in DNA repair, and summarize the progress of PARP inhibitors in cancer therapy. Full article

Neuroblastoma (NB) is a childhood cancer, commonly treated with drugs, such as etoposide (ETO), whose efficacy is limited by the onset of resistance. Here, aiming at identifying new treatments for chemo-resistant NB, the effects of two synthesized imidazo-pyrazoles (IMPs) (4G and 4I) were investigated on ETO-sensitive (HTLA-230) and ETO-resistant (HTLA-ER) NB cells, detecting 4I as the more promising compound, that demonstrated IC₅₀ values lower than those of ETO on HTLA ER. Therefore, to further improve the activity of 4I, we developed 4I-loaded palmitic acid (PA) and polystyrene-based (P5) cationic nanoparticles (P5PA-4I NPs) with high drug loading (21%) and encapsulation efficiency (97%), by a single oil-in-water emulsification technique. Biocompatible PA was adopted as an emulsion stabilizer, while synthesized P5 acted as an encapsulating agent, solubilizer and hydrophilic–lipophilic balance (HLB) improver. Optic microscopy and cytofluorimetric analyses were performed to investigate the micromorphology, size and complexity distributions of P5PA-4I NPs, which were also structurally characterized by chemometric-assisted Fourier transform infrared spectroscopy (FTIR). Potentiometric titrations allowed us to estimate the milliequivalents of PA and basic nitrogen atoms present in NPs. P5PA-4I NPs afforded dispersions in water with excellent buffer capacity, essential to escape lysosomal degradation and promote long residence time inside cells. They were chemically stable in an aqueous medium for at least 40 days, while in dynamic light scattering (DLS) analyses, P5PA-4I showed a mean hydrodynamic diameter of 541 nm, small polydispersity (0.194), and low positive zeta potentials (+8.39 mV), assuring low haemolytic toxicity. Biological experiments on NB cells, demonstrated that P5PA-4I NPs induced ROS-dependent cytotoxic effects significantly higher than those of pristine 4I, showing a major efficacy compared to ETO in reducing cell viability in HTLA-ER cells. Collectively, this 4I-based nano-formulation could represent a new promising macromolecular platform to develop a new delivery system able to increase the cytotoxicity of the anticancer drugs. Full article

Phenylpyrazolo[3,4-d]pyrimidine is considered a milestone scaffold known to possess various biological activities such as antiparasitic, antifungal, antimicrobial, and antiproliferative activities. In addition, the urgent need for selective and potent novel anticancer agents represents a major route in the drug discovery process. Herein, new aryl analogs were synthesized and evaluated for their anticancer effects on a panel of cancer cell lines: MCF-7, HCT116, and HePG-2. Some of these compounds showed potent cytotoxicity, with variable degrees of potency and cell line selectivity in antiproliferative assays with low resistance. As the analogs carry the pyrazolopyrimidine scaffold, which looks structurally very similar to tyrosine and receptor kinase inhibitors, the potent compounds were evaluated for their inhibitory effects on three essential cancer targets: EGFR^WT, EGFR^T790M, VGFR2, and Top-II. The data obtained revealed that most of these compounds were potent, with variable degrees of target selectivity and dual EGFR/VGFR2 inhibitors at the IC₅₀ value range, i.e., 0.3–24 µM. Among these, compound 5i was the most potent non-selective dual EGFR/VGFR2 inhibitor, with inhibitory concentrations of 0.3 and 7.60 µM, respectively. When 5i was tested in an MCF-7 model, it effectively inhibited tumor growth, strongly induced cancer cell apoptosis, inhibited cell migration, and suppressed cell cycle progression leading to DNA fragmentation. Molecular docking studies were performed to explore the binding mode and mechanism of such compounds on protein targets and mapped with reference ligands. The results of our studies indicate that the newly discovered phenylpyrazolo[3,4-d]pyrimidine-based multitarget inhibitors have significant potential for anticancer treatment. Full article

The amygdala has large populations of neurons utilizing specific calcium-binding proteins such as parvalbumin (PV), calbindin (CB), or calretinin (CR). They are considered specialized subsets of γ-aminobutyric acid (GABA) interneurons; however, many of these cells are devoid of GABA or glutamate decarboxylase. The neurotransmitters used by GABA-immunonegative cells are still unknown, but it is suggested that a part may use glutamate. Thus, this study investigates in the amygdala of the guinea pig relationships between PV, CB, or CR-containing cells and GABA transporter (VGAT) or glutamate transporter type 2 (VGLUT2), markers of GABAergic and glutamatergic neurons, respectively. The results show that although most neurons using PV, CB, and CR co-expressed VGAT, each of these populations also had a fraction of VGLUT2 co-expressing cells. For almost all neurons using PV (~90%) co-expressed VGAT, while ~1.5% of them had VGLUT2. The proportion of neurons using CB and VGAT was smaller than that for PV (~80%), while the percentage of cells with VGLUT2 was larger (~4.5%). Finally, only half of the neurons using CR (~53%) co-expressed VGAT, while ~3.5% of them had VGLUT2. In conclusion, the populations of neurons co-expressing PV, CB, and CR are in the amygdala, primarily GABAergic. However, at least a fraction of neurons in each of them co-express VGLUT2, suggesting that these cells may use glutamate. Moreover, the number of PV-, CB-, and CR-containing neurons that may use glutamate is probably larger as they can utilize VGLUT1 or VGLUT3, which are also present in the amygdala. Full article

Microbiome dysbiosis is increasingly being recognized as implicated in immune-mediated disorders including multiple sclerosis (MS). The microbiome is modulated by genetic and environmental factors including lifestyle, diet, and drug intake. This study aimed to characterize the MS-associated gut microbiome in the Israeli populations and to identify associations with demographic, dietary, and clinical features. The microbiota from 57 treatment-naive patients with MS (PwMS) and 43 age- and gender-matched healthy controls (HCs) was sequenced and abundance compared. Associations between differential microbes with demographic or clinical characteristics, as well as diet and nutrient intake, were assessed. While there was no difference in α- or β-diversity of the microbiome, we identified 40 microbes from different taxonomic levels that differ in abundance between PwMS and HCs, including Barnesiella, Collinsella, Egerthella, Mitsuokella, Olsenella Romboutsia, and Succinivibrio, all enhanced in PwMS, while several members of Lacnospira were reduced. Additional MS-differential microbes specific to ethnicity were identified. Several MS-specific microbial patterns were associated with gender, vitamin D level, Mediterranean diet, nutrient intake, or disability status. Thus, PwMS have altered microbiota composition, with distinctive patterns related to geographic locations and population. Microbiome dysbiosis seem to be implicated in disease progression, gender-related differences, and vitamin D-mediated immunological effects recognized in MS. Dietary interventions may be beneficial in restoring a “healthy microbiota” as part of applying comprehensive personalized therapeutic strategies for PwMS. Full article

The microtubule-associated protein tau is an intrinsically disordered protein containing a few short and transient secondary structures. Tau physiologically associates with microtubules (MTs) for its stabilization and detaches from MTs to regulate its dynamics. Under pathological conditions, tau is abnormally modified, detaches from MTs, and forms protein aggregates in neuronal and glial cells. Tau protein aggregates can be found in a number of devastating neurodegenerative diseases known as “tauopathies”, such as Alzheimer’s disease (AD), frontotemporal dementia (FTD), corticobasal degeneration (CBD), etc. However, it is still unclear how the tau protein is compacted into ordered protein aggregates, and the toxicity of the aggregates is still debated. Fortunately, there has been considerable progress in the study of tau in recent years, particularly in the understanding of the intercellular transmission of pathological tau species, the structure of tau aggregates, and the conformational change events in the tau polymerization process. In this review, we summarize the concepts of tau protein aggregation and discuss the views on tau protein transmission and toxicity. Full article

S-glutathionylation is an oxidative post-translational modification, which is involved in the regulation of many cell signaling pathways. Increasing amounts of studies show that it is crucial in cell homeostasis and deregulated in several pathologies. However, the effect of S-glutathionylation on proteins’ structure and activity is poorly understood, and a drastic lack of structural information at the atomic scale remains. Studies based on the use of molecular dynamics simulations, which can provide important information about modification-induced modulation of proteins’ structure and function, are also sparse, and there is no benchmarked force field parameters for this modified cysteine. In this contribution, we provide robust AMBER parameters for S-glutathionylation, which we tested extensively against experimental data through a total of 33 $\mathsf{\mu }$s molecular dynamics simulations. We show that our parameter set efficiently describes the global and local structural properties of S-glutathionylated proteins. These data provide the community with an important tool to foster new investigations into the effect of S-glutathionylation on protein dynamics and function, in a common effort to unravel the structural mechanisms underlying its critical role in cellular processes. Full article

Graphitic carbon nitride (g-C₃N₄), a metal-free polymer semiconductor, has been recognized as an attractive photocatalytic material for environmental remediation because of its low band gap, high thermal and photostability, chemical inertness, non-toxicity, low cost, biocompatibility, and optical and electrical efficiency. However, g-C₃N₄ has been reported to suffer from many difficulties in photocatalytic applications, such as a low specific surface area, inadequate visible-light utilization, and a high charge recombination rate. To overcome these difficulties, the formation of g-C₃N₄ heterojunctions by coupling with metal oxides has triggered tremendous interest in recent years. In this regard, zinc oxide (ZnO) is being largely explored as a self-driven semiconductor photocatalyst to form heterojunctions with g-C₃N₄, as ZnO possesses unique and fascinating properties, including high quantum efficiency, high electron mobility, cost-effectiveness, environmental friendliness, and a simple synthetic procedure. The synergistic effect of its properties, such as adsorption and photogenerated charge separation, was found to enhance the photocatalytic activity of heterojunctions. Hence, this review aims to compile the strategies for fabricating g-C₃N₄/ZnO-based Z-scheme and S-scheme heterojunction photocatalytic systems with enhanced performance and overall stability for the photodegradation of organic pollutants. Furthermore, with reference to the reported system, the photocatalytic mechanism of g-C₃N₄/ZnO-based heterojunction photocatalysts and their charge-transfer pathways on the interface surface are highlighted. Full article

3β-hydroxy-12-oleanen-27-oic acid (ATA), a cytotoxic oleanane triterpenoid with C14-COOH isolated from the rhizome of Astilbe chinensis, has been previously proven to possess antitumor activity and may be a promising antitumor agent. However, its molecular mechanisms of antitumor action were still unclear. This study explored the underlying mechanisms of cytotoxicity and potential target of ATA against human colorectal cancer HCT116 cells via integrative analysis of transcriptomics and network pharmacology in combination with in vitro and in vivo experimental validations. ATA significantly inhibited the proliferation of HCT116 cells in a concentration- and time-dependent manner and induced the cell cycle arrest at the G0/G1 phase, apoptosis, autophagy, and ferroptosis. Transcriptomic analysis manifested that ATA regulated mRNA expression of the genes related to cell proliferation, cell cycle, and cell death in HCT116 cells. The integrated analysis of transcriptomics, network pharmacology, and molecular docking revealed that ATA exerted cytotoxic activity via interactions with FDFT1, PPARA, and PPARG. Furthermore, FDFT1 was verified to be an upstream key target mediating the antiproliferative effect of ATA against HCT116 cells. Of note, ATA remarkably suppressed the growth of HCT116 xenografts in nude mice and displayed an apparent attenuation of FDFT1 in tumor tissues accompanied by the alteration of the biomarkers of autophagy, cell cycle, apoptosis, and ferroptosis. These results demonstrate that ATA exerted in vitro and in vivo antiproliferative effects against HCT116 cells through inducing cell apoptosis, autophagy, and ferroptosis via targeting FDFT1. Full article

The management of patients with acute myeloid leukemia (AML) relapsed post allogeneic hematopoietic stem cell transplantation (HSCT) remains a clinical challenge. Intensive treatment approaches are limited by severe toxicities in the early post-transplantation period. Therefore, hypomethylating agents (HMAs) have become the standard therapeutic approach due to favorable tolerability. Moreover, HMAs serve as a backbone for additional anti-leukemic agents. Despite discordant results, the addition of donor lymphocytes infusions (DLI) generally granted improved outcomes with manageable GvHD incidence. The recent introduction of novel targeted drugs in AML gives the opportunity to add a third element to salvage regimens. Those patients harboring targetable mutations might benefit from IDH1/2 inhibitors Ivosidenib and Enasidenib as well as FLT3 inhibitors Sorafenib and Gilteritinib in combination with HMA and DLI. Conversely, patients lacking targetable mutations actually benefit from the addition of Venetoclax. A second HSCT remains a valid option, especially for fit patients and for those who achieve a complete disease response with salvage regimens. Overall, across studies, higher response rates and longer survival were observed in cases of pre-emptive intervention for molecular relapse. Future perspectives currently rely on the development of adoptive immunotherapeutic strategies mainly represented by CAR-T cells. Full article

The present study investigates the impact of two endocrine disruptors, namely Bisphenols (BPs) and Perfluoroalkyls (PFs), on human stem cells. These chemicals leach from plastic, and when ingested through contaminated food and water, they interfere with endogenous hormone signaling, causing various diseases. While the ability of BPs and PFs to cross the placental barrier and accumulate in fetal serum has been documented, the exact consequences for human development require further elucidation. The present research work explored the effects of combined exposure to BPs (BPA or BPS) and PFs (PFOS and PFOA) on human placenta (fetal membrane mesenchymal stromal cells, hFM-MSCs) and amniotic fluid (hAFSCs)-derived stem cells. The effects of the xenobiotics were assessed by analyzing cell proliferation, mitochondrial functionality, and the expression of genes involved in pluripotency and epigenetic regulation, which are crucial for early human development. Our findings demonstrate that antenatal exposure to BPs and/or PFs may alter the biological characteristics of perinatal stem cells and fetal epigenome, with potential implications for health outcomes at birth and in adulthood. Further research is necessary to comprehend the full extent of these effects and their long-term consequences. Full article

Although Pichia pastoris was successfully used for heterologous gene expression for more than twenty years, many factors influencing protein expression remain unclear. Here, we optimized the expression of a thermophilic endoglucanase from Thermothielavioides terrestris (TtCel45A) for cost-effective production in Pichia pastoris. To achieve this, we established a multifactorial regulation strategy that involved selecting a genome-editing system, utilizing neutral loci, incorporating multiple copies of the heterologous expression cassette, and optimizing high-density fermentation for the co-production of single-cell protein (SCP). Notably, even though all neutral sites were used, there was still a slight difference in the enzymatic activity of heterologously expressed TtCel45A. Interestingly, the optimal gene copy number for the chromosomal expression of TtCel45A was found to be three, indicating limitations in translational capacity, post-translational processing, and secretion, ultimately impacting protein yields in P. pastoris. We suggest that multiple parameters might influence a kinetic competition between protein elongation and mRNA degradation. During high-density fermentation, the highest protein concentration and endoglucanase activity of TtCel45A with three copies reached 15.8 g/L and 9640 IU/mL, respectively. At the same time, the remaining SCP of P. pastoris exhibited a crude protein and amino acid content of up to 59.32% and 46.98%, respectively. These findings suggested that SCP from P. pastoris holds great promise as a sustainable and cost-effective alternative for meeting the global protein demand, while also enabling the production of thermophilic TtCel45A in a single industrial process. Full article

Autophagy is a catabolic process that is essential to the maintenance of homeostasis through the cellular recycling of damaged organelles or misfolded proteins, which sustains energy balance. Additionally, autophagy plays a dual role in modulating the development and progression of cancer and inducing a survival strategy in tumoral cells. Programmed cell death-ligand 1 (PD-L1) modulates the immune response and is responsible for maintaining self-tolerance. Because tumor cells exploit the PD-L1–PD-1 interaction to subvert the immune response, immunotherapy has been developed based on the use of PD-L1-blocking antibodies. Recent evidence has suggested a bidirectional regulation between autophagy and PD-L1 molecule expression in tumor cells. Moreover, the research into the intrinsic properties of PD-L1 has highlighted new functions that are advantageous to tumor cells. The relationship between autophagy and PD-L1 is complex and still not fully understood; its effects can be context-dependent and might differ between tumoral cells. This review refines our understanding of the non-immune intrinsic functions of PD-L1 and its potential influence on autophagy, how these could allow the survival of tumor cells, and what this means for the efficacy of anti-PD-L1 therapeutic strategies. Full article

Recently, proenkephalin A (PENK A) has been shown to reflect glomerular dysfunction and to predict new-onset acute kidney injury and heart failure. While previous studies have investigated PENK A as a biomarker in individuals with preserved renal function, PENK A concentration in patients with end-stage kidney disease (ESKD) was not investigated. Plasma PENK A concentration was assessed in 88 patients with ESKD treated with hemodialysis (HD) or peritoneal dialysis (PD), and its associations with kidney function and heart failure indicators were investigated. In HD patients, the difference in PENK A levels before and after hemodialysis, was measured and further assessed for an association with the type of HD membrane used. PENK A levels did not differ significantly between HD and PD patients. In HD patients, the median PENK A concentration was significantly higher before than after hemodialysis (1.368 vs. 2.061, p = 0.003). No correlation was found between PENK A level and urea (p = 0.192), eGFR (p = 0.922), dialysis vintage (p = 0.637), and residual urine output (p = 0.784). Heart failure (p = 0.961), EF (p = 0.361), and NT-proBNP (p = 0.949) were not associated with increased PENK A concentration. PENK A does not reflect renal function and cardiac status in patients with ESKD. Further research is required to establish the clinical utility of the new biomarker in patients with impaired kidney function. Full article

Autophagy is an evolutionarily conserved mechanism for degrading and recycling various cellular components, functioning in both normal development and stress conditions. This process is tightly regulated by a set of autophagy-related (ATG) proteins, including ATG2 in the ATG9 cycling system and ATG5 in the ATG12 conjugation system. Our recent research demonstrated that autophagy-mediated compartmental cytoplasmic deletion is essential for pollen germination. However, the precise mechanisms through which autophagy regulates pollen germination, ensuring its fertility, remain largely unknown. Here, we applied multi-omics analyses, including transcriptomic and metabolomic approaches, to investigate the downstream pathways of autophagy in the process of pollen germination. Although ATG2 and ATG5 play similar roles in regulating pollen germination, high-throughput transcriptomic analysis reveals that silencing ATG5 has a greater impact on the transcriptome than silencing ATG2. Cross-comparisons of transcriptome and proteome analysis reveal that gene expression at the mRNA level and protein level is differentially affected by autophagy. Furthermore, high-throughput metabolomics analysis demonstrates that pathways related to amino acid metabolism and aminoacyl-tRNA biosynthesis were affected by both ATG2 and ATG5 silencing. Collectively, our multi-omics analyses reveal the central role of autophagy in cellular metabolism, which is critical for initiating pollen germination and ensuring pollen fertility. Full article

Sugar beet is susceptible to Beet curly top virus (BCTV), which significantly reduces yield and sugar production in the semi-arid growing regions worldwide. Sources of genetic resistance to BCTV is limited and control depends upon insecticide seed treatments with neonicotinoids. Through double haploid production and genetic selection, BCTV resistant breeding lines have been developed. Using BCTV resistant (R) [KDH13; Line 13 and KDH4-9; Line 4] and susceptible (S) [KDH19-17; Line 19] lines, beet leafhopper mediated natural infection, mRNA/sRNA sequencing, and metabolite analyses, potential mechanisms of resistance against the virus and vector were identified. At early infection stages (2- and 6-days post inoculation), examples of differentially expressed genes highly up-regulated in the ‘R’ lines (vs. ‘S’) included EL10Ac5g10437 (inhibitor of trypsin and hageman factor), EL10Ac6g14635 (jasmonate-induced protein), EL10Ac3g06016 (ribosome related), EL10Ac2g02812 (probable prolyl 4-hydroxylase 10), etc. Pathway enrichment analysis showed differentially expressed genes were predominantly involved with peroxisome, amino acids metabolism, fatty acid degradation, amino/nucleotide sugar metabolism, etc. Metabolite analysis revealed significantly higher amounts of specific isoflavonoid O-glycosides, flavonoid 8-C glycosides, triterpenoid, and iridoid-O-glycosides in the leaves of the ‘R’ lines (vs. ‘S’). These data suggest that a combination of transcriptional regulation and production of putative antiviral metabolites might contribute to BCTV resistance. In addition, genome divergence among BCTV strains differentially affects the production of small non-coding RNAs (sncRNAs) and small peptides which may potentially affect pathogenicity and disease symptom development. Full article

A series of novel derivatives of 18β-glycyrrhetinic acid (GA) were synthesized by introducing aromatic or heterocyclic structures to extend the side chain, thereby enhancing their interaction with amino acid residues in the active pocket of the target protein. These compounds were structurally characterized using ¹H NMR, ¹³C NMR, and HRMS. The compounds were subsequently evaluated for their inhibitory effects on HIV-1 protease and cell viability in the human cancer cell lines K562 and HeLa and the mouse cancer cell line CT26. Towards HIV-1 protease, compounds 28 and 32, which featured the introduction of heterocyclic moieties at the C3 position of GA, exhibited the highest inhibition, with inhibition rates of 76% and 70.5%, respectively, at 1 mg/mL concentration. Further molecular docking suggests that a 3-substituted polar moiety would be likely to enhance the inhibitory activity against HIV-1 protease. As for the anti-proliferative activities of the GA derivatives, incorporation of a thiazole heterocycle at the C3- position in compound 29 significantly enhanced the effect against K562 cells with an IC₅₀ value of 8.86 ± 0.93 µM. The introduction of electron-withdrawing substituents on the C3-substituted phenyl ring augmented the anti-proliferative activity against Hela and CT26 cells. Compound 13 exhibited the highest inhibitory activity against Hela cells with an IC₅₀ value of 9.89 ± 0.86 µM, whereas compound 7 exerted the strongest inhibition against CT26 cells with an IC₅₀ value of 4.54 ± 0.37 µM. These findings suggest that further modification of GA is a promising path for developing potent novel anti-HIV and anticancer therapeutics. Full article

Alzheimer’s disease (AD) is a growing global health crisis affecting millions and incurring substantial economic costs. However, clinical diagnosis remains challenging, with misdiagnoses and underdiagnoses being prevalent. There is an increased focus on putative, blood-based biomarkers that may be useful for the diagnosis as well as early detection of AD. In the present study, we used an unbiased combination of machine learning and functional network analyses to identify blood gene biomarker candidates in AD. Using supervised machine learning, we also determined whether these candidates were indeed unique to AD or whether they were indicative of other neurodegenerative diseases, such as Parkinson’s disease (PD) and amyotrophic lateral sclerosis (ALS). Our analyses showed that genes involved in spliceosome assembly, RNA binding, transcription, protein synthesis, mitoribosomes, and NADH dehydrogenase were the best-performing genes for identifying AD patients relative to cognitively healthy controls. This transcriptomic signature, however, was not unique to AD, and subsequent machine learning showed that this signature could also predict PD and ALS relative to controls without neurodegenerative disease. Combined, our results suggest that mRNA from whole blood can indeed be used to screen for patients with neurodegeneration but may be less effective in diagnosing the specific neurodegenerative disease. Full article

Saliva houses over 2000 proteins and peptides with poorly clarified functions, including proline-rich proteins, statherin, P-B peptides, histatins, cystatins, and amylases. Their genes are poorly conserved across related species, reflecting an evolutionary adaptation. We searched the nucleotide substitutions fixed in these salivary proteins’ gene loci in modern humans compared with ancient hominins. We mapped 3472 sequence variants/nucleotide substitutions in coding, noncoding, and 5′-3′ untranslated regions. Despite most of the detected variations being within noncoding regions, the frequency of coding variations was far higher than the general rate found throughout the genome. Among the various missense substitutions, specific substitutions detected in PRB1 and PRB2 genes were responsible for the introduction/abrogation of consensus sequences recognized by convertase enzymes that cleave the protein precursors. Overall, these changes that occurred during the recent human evolution might have generated novel functional features and/or different expression ratios among the various components of the salivary proteome. This may have influenced the homeostasis of the oral cavity environment, possibly conditioning the eating habits of modern humans. However, fixed nucleotide changes in modern humans represented only 7.3% of all the substitutions reported in this study, and no signs of evolutionary pressure or adaptative introgression from archaic hominins were found on the tested genes. Full article

Homeostasis of the host immune system is regulated by white blood cells with a variety of cell surface receptors for cytokines. Chemotactic cytokines (chemokines) activate their receptors to evoke the chemotaxis of immune cells in homeostatic migrations or inflammatory conditions towards inflamed tissue or pathogens. Dysregulation of the immune system leading to disorders such as allergies, autoimmune diseases, or cancer requires efficient, fast-acting drugs to minimize the long-term effects of chronic inflammation. Here, we performed structure-based virtual screening (SBVS) assisted by the Keras/TensorFlow neural network (NN) to find novel compound scaffolds acting on three chemokine receptors: CCR2, CCR3, and one CXC receptor, CXCR3. Keras/TensorFlow NN was used here not as a typically used binary classifier but as an efficient multi-class classifier that can discard not only inactive compounds but also low- or medium-activity compounds. Several compounds proposed by SBVS and NN were tested in 100 ns all-atom molecular dynamics simulations to confirm their binding affinity. To improve the basic binding affinity of the compounds, new chemical modifications were proposed. The modified compounds were compared with known antagonists of these three chemokine receptors. Known CXCR3 compounds were among the top predicted compounds; thus, the benefits of using Keras/TensorFlow in drug discovery have been shown in addition to structure-based approaches. Furthermore, we showed that Keras/TensorFlow NN can accurately predict the receptor subtype selectivity of compounds, for which SBVS often fails. We cross-tested chemokine receptor datasets retrieved from ChEMBL and curated datasets for cannabinoid receptors. The NN model trained on the cannabinoid receptor datasets retrieved from ChEMBL was the most accurate in the receptor subtype selectivity prediction. Among NN models trained on the chemokine receptor datasets, the CXCR3 model showed the highest accuracy in differentiating the receptor subtype for a given compound dataset. Full article

The color of the chili fruit is an important factor that determines the quality of the chili, as red chilies are more popular among consumers. The accumulation of capsanthin is the main cause of reddening of the chili fruit. Capsanthin is an important metabolite in carotenoid metabolism, and its production level is closely linked to the expression of the genes for capsanthin/capsorubin synthase (CCS) and carotenoid hydroxylase (CrtZ). We reported for the first time that the synthesis of capsanthin in chili was enhanced by using a geminivirus (Bean Yellow Dwarf Virus). By expressing heterologous β-carotenoid hydroxylase (CrtZ) and β-carotenoid ketolase (CrtW) using codon optimization, the transcription level of the CCS gene and endogenous CrtZ was directly increased. This leads to the accumulation of a huge amount of capsanthin in a very short period of time. Our results provide a platform for the rapid enhancement of endogenous CCS activity and capsanthin production using geminivirus in plants. Full article

Pain is the most significant impairment and debilitating challenge for patients with bone metastasis. Therefore, the primary objective of current therapy is to mitigate and prevent the persistence of pain. Thus, cancer-induced bone pain is described as a multifaceted form of discomfort encompassing both inflammatory and neuropathic elements. We have developed a novel non-addictive pain therapeutic, PNA6, that is a derivative of the peptide Angiotensin-(1-7) and binds the Mas receptor to decrease inflammation-related cancer pain. In the present study, we provide evidence that PNA6 attenuates inflammatory, chemotherapy-induced peripheral neuropathy (CIPN) and cancer pain confined to the long bones, exhibiting longer-lasting efficacious therapeutic effects. PNA6, Asp-Arg-Val-Tyr-Ile-His-Ser-(O-β-Lact)-amide, was successfully synthesized using solid phase peptide synthesis (SPPS). PNA6 significantly reversed inflammatory pain induced by 2% carrageenan in mice. A second murine model of platinum drug-induced painful peripheral neuropathy was established using oxaliplatin. Mice in the oxaliplatin-vehicle treatment groups demonstrated significant mechanical allodynia compared to the oxaliplatin-PNA6 treatment group mice. In a third study modeling a complex pain state, E0771 breast adenocarcinoma cells were implanted into the femur of female C57BL/6J wild-type mice to induce cancer-induced bone pain (CIBP). Both acute and chronic dosing of PNA6 significantly reduced the spontaneous pain behaviors associated with CIBP. These data suggest that PNA6 is a viable lead candidate for treating chronic inflammatory and complex neuropathic pain. Full article

This comprehensive review thoroughly explores the intricate involvement of insulin receptor (IR) isoforms and insulin-like growth factor receptors (IGFRs) in the context of the insulin and insulin-like growth factor (IGF) signaling (IIS) pathway. This elaborate system encompasses ligands, receptors, and binding proteins, giving rise to a wide array of functions, including aspects such as carcinogenesis and chemoresistance. Detailed genetic analysis of IR and IGFR structures highlights their distinct isoforms, which arise from alternative splicing and exhibit diverse affinities for ligands. Notably, the overexpression of the IR-A isoform is linked to cancer stemness, tumor development, and resistance to targeted therapies. Similarly, elevated IGFR expression accelerates tumor progression and fosters chemoresistance. The review underscores the intricate interplay between IRs and IGFRs, contributing to resistance against anti-IGFR drugs. Consequently, the dual targeting of both receptors could present a more effective strategy for surmounting chemoresistance. To conclude, this review brings to light the pivotal roles played by IRs and IGFRs in cellular signaling, carcinogenesis, and therapy resistance. By precisely modulating these receptors and their complex signaling pathways, the potential emerges for developing enhanced anti-cancer interventions, ultimately leading to improved patient outcomes. Full article

Major latex proteins (MLPs) play a key role in plant response to abiotic and biotic stresses. However, little is known about this gene family in tomatoes (Solanum lycopersicum). In this paper, we perform a genome-wide evolutionary characterization and gene expression analysis of the MLP family in tomatoes. We found a total of 34 SlMLP members in the tomato genome, which are heterogeneously distributed on eight chromosomes. The phylogenetic analysis of the SlMLP family unveiled their evolutionary relationships and possible functions. Furthermore, the tissue-specific expression analysis revealed that the tomato MLP members possess distinct biological functions. Crucially, multiple cis-regulatory elements associated with stress, hormone, light, and growth responses were identified in the promoter regions of these SlMLP genes, suggesting that SlMLPs are potentially involved in plant growth, development, and various stress responses. Subcellular localization demonstrated that SlMLP1, SlMLP3, and SlMLP17 are localized in the cytoplasm. In conclusion, these findings lay a foundation for further dissecting the functions of tomato SlMLP genes and exploring the evolutionary relationships of MLP homologs in different plants. Full article

β-Elemene (C₁₅H₂₄), a sesquiterpenoid compound isolated from the volatile oil of Curcuma wenyujin, has been proven to be effective for multiple cancers and is widely used in clinical treatment. Unfortunately, the β-elemene content in C. wenyujin is very low, which cannot meet market demands. Our previous research showed that methyl jasmonate (MeJA) induced the accumulation of β-elemene in C. wenyujin. However, the regulatory mechanism is unclear. In this study, 20 jasmonate ZIM-domain (JAZ) proteins in C. wenyujin were identified, which are the core regulatory factors of the JA signaling pathway. Then, the conservative domains, motifs composition, and evolutionary relationships of CwJAZs were analyzed comprehensively and systematically. The interaction analysis indicated that CwJAZs can form homodimers or heterodimers. Fifteen out of twenty CwJAZs were significantly induced via MeJA treatment. As the master switch of the JA signaling pathway, the CwMYC2-like protein has also been identified and demonstrated to interact with CwJAZ2/3/4/5/7/15/17/20. Further research found that the overexpression of the CwMYC2-like gene increased the accumulation of β-elemene in C. wenyujin leaves. Simultaneously, the expressions of HMGR, HMGS, DXS, DXR, MCT, HDS, HDR, and FPPS related to β-elemene biosynthesis were also up-regulated by the CwMYC2-like protein. These results indicate that CwJAZs and the CwMYC2-like protein respond to the JA signal to regulate the biosynthesis of β-elemene in C. wenyujin. Full article