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The article presents the theological interpretation of the phenomenon of spousal love in terms of examining its correlations with the call to holiness. This study belongs to the field of hagiological research aiming at developing a new concept that defines arguments in the Church’s strategy concerning the defence of every human life. The analysis concerns the statements and philosophical writings of Karol Wojtyła and then John Paul II on spousal love and the dependence of the person and his actions on the Truth and Good. The Christological-soteriological aspect of spousal love as conditioning the sanctification of the person has been indicated. The axiological conditions related to the Christological assumption have been termed as “the Splendour of Divinity,” identifying it with the space of the salvific influence on a person, sanctified by Christ’s spousal love and called to develop an ethos based on this love.

Dialogiczne myślenie w teologii odkrywa wewnętrzną harmonię rzeczywistości stworzonej uwarunkowaną dialogiką relacji osoby Boskiej z osobą ludzką.

Humanizm chrześcijański określa postawę poznawczą i praktyczną. Istotne jest tu zwrócenie uwagi na zrównoważenie twórczości kulturowej ze wspólnotową. Przeakcentowanie jednego wymiaru wprowadza dysproporcje w relacje społeczne i kulturowe, powodując zagubienie w nadmiarze jednego oraz szkody z powodu deficytu drugiego. W tym zakresie humanizm chrześcijański jawi się jako postawa badawcza w odniesieniu do dynamiki przemian w strukturach społecznych i gospodarczych.

Artykuł odpowiada na pytanie o specyfikę urzeczywistniania się egzystencji Jezusa w życiu małżonków sakramentalnych. Sakrament małżeństwa tworzy nową komórkę wspólnoty eklezjalnej. Małżonkowie sakramentalni wdrażają w swoje życie naukę Jezusa przez naśladowanie Jego etosu miłości. Dlatego zasadne jest stwierdzenie, że konkretyzacja egzystencji Jezusa w życiu małżonków sakramentalnych jest działaniem „in Persona Ecclesiae”.

Artykuł przedstawia kwestię sumienia eklezjalnego jako osobowej przestrzeni, w której człowiek wierzący dokonuje oceny swojej działalności eklezjotwórczej. Kształtowanie sumienia wiernych w formacie eklezjalnym jest zgodne z teandryczną i zarazem misyjną naturą Kościoła. Zagadnienie jest ukazane na bazie trzech dokumentów papieskich: Veritatis splendor, Caritas in veritate, Lumen fidei. Źródła te wyjaśniają związki między absolutną Prawdą, Dobrem i Miłością jako transcendentaliami warunkującymi indywidualny proces zbawienia człowieka.

Significance Cancer therapies often fail to cure patients because a proportion of tumor cells withstand the toxic effects of chemotherapy. How surviving cancer cells recover from sublethal drug-induced stress is not known, but given that cellular resources are finite, stress resolution may come at the expense of less essential systems. Here, we studied the global cellular events of stress buildup and resolution in the bone marrow cancer, multiple myeloma, after proteasome inhibition, a commonly used therapeutic approach. Using a temporal multiomics approach, we delineate the unexpectedly complex and protracted changes myeloma cells undergo during stress resolution and demonstrate that recovering cells are more vulnerable to specific insults than acutely stressed cells. Thus, the findings may provide avenues for optimizing cancer therapies., Cancer cells can survive chemotherapy-induced stress, but how they recover from it is not known. Using a temporal multiomics approach, we delineate the global mechanisms of proteotoxic stress resolution in multiple myeloma cells recovering from proteasome inhibition. Our observations define layered and protracted programs for stress resolution that encompass extensive changes across the transcriptome, proteome, and metabolome. Cellular recovery from proteasome inhibition involved protracted and dynamic changes of glucose and lipid metabolism and suppression of mitochondrial function. We demonstrate that recovering cells are more vulnerable to specific insults than acutely stressed cells and identify the general control nonderepressable 2 (GCN2)-driven cellular response to amino acid scarcity as a key recovery-associated vulnerability. Using a transcriptome analysis pipeline, we further show that GCN2 is also a stress-independent bona fide target in transcriptional signature-defined subsets of solid cancers that share molecular characteristics. Thus, identifying cellular trade-offs tied to the resolution of chemotherapy-induced stress in tumor cells may reveal new therapeutic targets and routes for cancer therapy optimization.

Cellular function depends on the maintenance of protein homeostasis (proteostasis) by regulated protein degradation. Chronic dysregulation of proteostasis is associated with neurodegenerative and age-related diseases, and drugs targeting components of the protein degradation apparatus are increasingly used in cancer therapies. However, as chronic imbalances rather than loss of function mediate their pathogenesis, research models that allow for the study of the complex effects of drugs on tissue properties in proteostasis-associated diseases are almost completely lacking. Here, to determine the functional effects of impaired proteostatic fine-tuning, we applied a combination of materials science characterisation techniques to a cell-derived, in vitro model of bone-like tissue formation in which we pharmacologically perturbed protein degradation. We show that low-level inhibition of VCP/p97 and the proteasome, two major components of the degradation machinery, have remarkably different effects on the bone-like material that human bone-marrow derived mesenchymal stromal cells (hMSC) form in vitro. Specifically, whilst proteasome inhibition mildly enhances tissue formation, Raman spectroscopic, atomic force microscopy-based indentation, and electron microscopy imaging reveal that VCP/p97 inhibition induces the formation of bone-like tissue that is softer, contains less protein, appears to have more crystalline mineral, and may involve aberrant micro- and ultra-structural tissue organisation. These observations contrast with findings from conventional osteogenic assays that failed to identify any effect on mineralisation. Taken together, these data suggest that mild proteostatic impairment in hMSC alters the bone-like material they form in ways that could explain some pathologies associated with VCP/p97-related diseases. They also demonstrate the utility of quantitative materials science approaches for tackling long-standing questions in biology and medicine, and could form the basis for preclinical drug testing platforms to develop therapies for diseases stemming from perturbed proteostasis or for cancer therapies targeting protein degradation. Our findings may also have important implications for the field of tissue engineering, as the manufacture of cell-derived biomaterial scaffolds may need to consider proteostasis to effectively replicate native tissues.

Background Cytosolic deacetylase histone deacetylase 6 (HDAC6) is involved in the autophagy degradation pathway of malformed proteins, an important survival mechanism in cancer cells. We evaluated modulation of autophagy-related proteins and cell death by the HDAC6-selective inhibitor C1A. Methods Autophagy substrates (light chain-3 (LC-3) and p62 proteins) and endoplasmic reticulum (ER) stress phenotype were determined. Caspase-3/7 activation and cellular proliferation assays were used to assess consequences of autophagy modulation. Results C1A potently resolved autophagy substrates induced by 3-methyladenine and chloroquine. The mechanism of autophagy inhibition by HDAC6 genetic knockout or C1A treatment was consistent with abrogation of autophagosome–lysosome fusion, and decrease of Myc protein. C1A alone or combined with the proteasome inhibitor, bortezomib, enhanced cell death in malignant cells, demonstrating the complementary roles of the proteasome and autophagy pathways for clearing malformed proteins. Myc-positive neuroblastoma, KRAS-positive colorectal cancer and multiple myeloma cells showed marked cell growth inhibition in response to HDAC6 inhibitors. Finally, growth of neuroblastoma xenografts was arrested in vivo by single agent C1A, while combination with bortezomib slowed the growth of colorectal cancer xenografts. Conclusions C1A resolves autophagy substrates in malignant cells and induces cell death, warranting its use for in vivo pre-clinical autophagy research.

Proteasome inhibitors (PIs) form the backbone of multiple myeloma (MM) treatment regimens used at diagnosis and relapse, but their clinical benefit is limited by varying degrees of resistance. The mechanisms that protect MM cells (MMCs) from PI-induced cell death are only partly understood, and experimental resistance studies often rely on the prolonged exposure of MMCs to relatively low levels of PIs. However, patients receive PI doses that result in high but short plasma peaks. Given that each PI dose reduces a patient's MMC load until a plateau is reached, one can assume that each PI dose kills some MMCs by triggering overwhelming stress. This proposition implies that a proportion of MMCs not only survive but also resolve PI-induced stress. We hypothesised that the mechanisms of PI-induced stress recovery require a redistribution of cellular resources, and that this process triggers specific recovery-associated and potentially druggable vulnerabilities. To test this hypothesis, we obtained a global systems-level view of the cellular response to proteasome inhibition by performing a multi-omics 10-day time-course experiment. To mimic in vivo pharmacokinetics and anti-MM effects, RPMI-8226 MMCs were treated with a 1h pulse of carfilzomib (CFZ; 750nmol/L), which reduced the number of viable MMCs to a nadir of approximately 50% on day +2, followed by a return to pretreatment baseline levels of viable cells by day +6. Levels of ubiquitinated proteins as a readout of effective proteasome inhibition peaked on day +1 and returned to baseline levels on day +6. Samples for transcriptome analysis by RNA-seq, proteome analysis by a multiplexed tandem mass tag (TMT)-based approach, and global metabolomic profiling by ultrahigh performance liquid chromatography-tandem mass spectroscopy (LC-MS) were collected 2h prior to the CFZ pulse (day 0) and on days +1, +2, +4, +6, +8, and +10. The results demonstrate extensive and kinetically complex changes of the MMC transcriptome, proteome and metabolome that did not fully return to baseline by day +10. Conventional and advanced computational analyses of RNA-seq data (including biological homogeneity score-validated graph-based clustering of temporal patterns combined with gene enrichment and KEGG/GO pathway analysis) revealed different patterns of involvement of multiple and functionally diverse pathways. Perhaps most notably, RNA-seq data revealed extensive metabolic pathway responses while cells were dying and throughout recovery. Consistently, metabolomic profiling by LC-MS showed wide-ranging metabolite changes. These included rapid intracellular depletion of glucose, which was paralleled by the accumulation of lactate and pyruvate and lasted until day +8. Consistently, the major glucose uptake regulator TXNIP stood out as upregulated during recovery, which was accompanied by the predicted downregulation of the glucose transporter GLUT1. These findings were confirmed by real-time PCR and immunoblotting in RPMI-8226 and other MMC lines. Moreover, pharmacological inhibition of glucose and lactate metabolising enzymes and transmembrane transport enhanced MMC killing by CFZ, confirming profound and druggable changes in glucose and energy metabolism during PI stress recovery. We also observed prolonged depletion of a number of amino acids, including glutamine, which was paralleled by depletion of glutamate and TCA cycle intermediates. Moreover, we found RNA-seq and real-time PCR/immunoblot-based evidence for an amino acid response (AAR) during stress recovery that was driven by the amino acid sensing kinase GCN2 (EIF2AK4). The preclinical GCN2 inhibitor GCN2iB was effective at blocking the AAR and enhanced MMC killing in a panel of CFZ-treated MMC lines. This effect was particularly pronounced during the intermediate stages of recovery, in line with the kinetics of amino acid depletion and the AAR. Thus, our observations define the complex and prolonged cellular responses that characterise the recovery of MMCs from proteasome inhibition. They reveal wide-ranging metabolic changes that persist far beyond immediate stress survival and highlight the dependence of MMCs on the GCN2-driven AAR to overcome PI stress. The findings demonstrate that the mechanisms of PI recovery trigger druggable vulnerabilities, providing the basis for ongoing investigations into sequential therapeutic interventions to enhance responses to PIs. Disclosures Caputo: GSK: Research Funding. Kaiser:Abbvie, Celgene, Takeda, Janssen, Amgen, Abbvie, Karyopharm: Consultancy; Celgene, Janssen: Research Funding; Takeda, Janssen, Celgene, Amgen: Honoraria, Other: Travel Expenses. Auner:Karyopharm: Consultancy; Takeda: Consultancy; Amgen: Other: Consultancy and Research Funding.

Pharmacologists have used pathogen-associated molecular patterns (PAMPs), such as lipopolysaccharide (LPS) for decades as a stimulus for studying mediators involved in inflammation and for the screening of anti-inflammatory compounds. However, in the view of immunologists, LPS was too non-specific for studying the mechanisms of immune signalling in infection and inflammation, as no receptors had been identified. This changed in the late 1990s with the discovery of the Toll-like receptors. These 'pattern recognition receptors' (PRRs) were able to recognise highly conserved sequences, the so called pathogen associated molecular patterns (PAMPs) present in or on pathogens. This specificity of particular PAMPs and their newly defined receptors provided a common ground between pharmacologists and immunologists for the study of inflammation. PRRs also recognise endogenous agonists, the so called danger-associated molecular patterns (DAMPs), which can result in sterile inflammation. The signalling pathways and ligands of many PRRs have now been characterised and there is no doubt that this rich vein of research will aid the discovery of new therapeutics for infectious conditions and chronic inflammatory disease.

The cellular mechanisms that control protein degradation may constitute a non-oncogenic cancer cell vulnerability and, therefore, a therapeutic target. Although this proposition is supported by the clinical success of proteasome inhibitors in some malignancies, most cancers are resistant to proteasome inhibition. The ATPase valosin-containing protein (VCP; p97) is an essential regulator of protein degradation in multiple pathways and has emerged as a target for cancer therapy. We found that pharmacological depletion of VCP enzymatic activity with mechanistically different inhibitors robustly induced proteotoxic stress in solid cancer and multiple myeloma cells, including cells that were insensitive, adapted, or clinically resistant to proteasome inhibition. VCP inhibition had an impact on two key regulators of protein synthesis, eukaryotic initiation factor 2α (eIF2α) and mechanistic target of rapamycin complex 1 (mTORC1), and attenuated global protein synthesis. However, a block on protein translation that was itself cytotoxic alleviated stress signaling and reduced cell death triggered by VCP inhibition. Some of the proteotoxic effects of VCP depletion depended on the eIF2α phosphatase, protein phosphatase 1 regulatory subunit 15A (PPP1R15A)/PP1c, but not on mTORC1, although there appeared to be cross-talk between them. Thus, cancer cell death following VCP inhibition was linked to inadequate fine-tuning of protein synthesis and activity of PPP1R15A/PP1c. VCP inhibitors also perturbed intracellular amino acid levels, activated eukaryotic translation initiation factor 2α kinase 4 (EIF2AK4), and enhanced cellular dependence on amino acid supplies, consistent with a failure of amino acid homeostasis. Many of the observed effects of VCP inhibition differed from the effects triggered by proteasome inhibition or by protein misfolding. Thus, depletion of VCP enzymatic activity triggers cancer cell death in part through inadequate regulation of protein synthesis and amino acid metabolism. The data provide novel insights into the maintenance of intracellular proteostasis by VCP and may have implications for the development of anti-cancer therapies.

Background Cigarette smoking is responsible for 5 million deaths worldwide each year, and is a major risk factor for cardiovascular and lung diseases. Cigarette smoke contains a complex mixture of over 4000 chemicals containing 1015 free radicals. Studies show smoke is perceived by cells as an inflammatory and xenobiotic stimulus, which activates an immune response. The specific cellular mechanisms driving cigarette smoke-induced inflammation and disease are not fully understood, although the innate immune system is involved in the pathology of smoking related diseases. Methodology/Principle findings To address the impact of smoke as an inflammagen on the innate immune system, THP-1 cells and Human PBMCs were stimulated with 3 and 10% (v/v) cigarette smoke extract (CSE) for 8 and 24 hours. Total RNA was extracted and the transcriptome analysed using Illumina BeadChip arrays. In THP-1 cells, 10% CSE resulted in 80 genes being upregulated and 37 downregulated by ≥1.5 fold after 8 hours. In PBMCs stimulated with 10% CSE for 8 hours, 199 genes were upregulated and 206 genes downregulated by ≥1.5 fold. After 24 hours, the number of genes activated and repressed by ≥1.5 fold had risen to 311 and 306 respectively. The major pathways that were altered are associated with cell survival, such as inducible antioxidants, protein chaperone and folding proteins, and the ubiquitin/proteosome pathway. Conclusions Our results suggest that cigarette smoke causes inflammation and has detrimental effects on the metabolism and function of innate immune cells. In addition, THP-1 cells provide a genetically stable alternative to primary cells for the study of the effects of cigarette smoke on human monocytes.

Introduction: Intracellular protein homeostasis requires a well-controlled protein degradation machinery to clear damaged and old proteins and to replenish intracellular amino acid pools. Cancer cells may be particularly susceptible to agents that disrupt protein degradation because of unbalanced protein levels related to gain or loss of genetic material, high protein turnover, or high-level production of specific proteins such as Ig in multiple myeloma (MM). The AAA ATPase VCP (p97) is a master regulator of protein degradation that has been implicated in oncogenesis. Small molecule VCP inhibition (VCP-i) rapidly activates caspases in cancer cells, induces endoplasmic reticulum (ER) stress, and has anti-tumor activity in murine xenograft models. A phase 1 trial of VCP-i in relapsed MM has recently opened. Methods: We investigated the cellular mechanisms that govern VCP-i mediated cancer cell death in MM cell lines including bortezomib-adapted AMO1 cells, MM cells from patients with bortezomib-resistant MM, as well as lung cancer (A549) and osteosarcoma (Saos2) cells. Results: The ATP-competitive inhibitor DBeQ and the allosteric inhibitor NMS873 induced cell line as well as primary MM cell death at similar low micromolar concentrations independently of bortezomib sensitivity. DBeQ and NMS873 caused phosphorylation of eIF2alpha in a time- and dose-dependent manner and resulted in strong transcriptional and translational up-regulation of ATF4, CHOP and GADD34. VCP-i also increased expression of ER chaperones BiP and p58IPK. Inhibition of eIF2alpha de-phosphorylation with guanabenz reduced S6 phosphorylation, a marker of protein translation, and increased A549 and OPM2 cell survival early (8h) after VCP-i. Direct inhibition of protein translation with cycloheximide also decreased early VCP-i mediated cell death. Using MEFs deficient in eIF2alpha kinases we show that eIF2alpha phosphorylation following VCP-i depends on both the unfolded protein response mediator PERK and the nutrient sensor GCN2. We found that DBeQ induces GCN2 phosphorylation in parallel with loss of mTORC1 signalling, induction of the key autophagy factor p62, and accumulation of LC3-II. DBeQ also induced a rapid decrease in free intracellular L-amino acids. Depletion of selected amino acids in the cell culture medium increased cell death and mRNA levels of CHOP and GADD34 following VCP-i with DBeQ or NMS873, but not following induction of ER stress with tunicamycin. Conclusion: Collectively, these data show that both ATP-competitive and allosteric VCP-i effectively kills cancer cells independently of bortezomib sensitivity. Disrupting VCP function induces early cell death via inappropriate eIF2alpha-regulated protein translation downstream of GADD34. VCP-i also depletes the intracellular free amino acid pool, resulting in cell death despite compensatory catabolic processes. Citation Format: Katarzyna Parzych, Sandra Loaiza, Tamara M. Chinn, Philippa May, Florentina Porsch, Anastasios Karadimitris, Christoph Driessen, Heather P. Harding, David Ron, Holger W. Auner. Comprehensive failure of intracellular protein homeostasis kills myeloma and solid cancer cells following VCP/p97 inhibition. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1261. doi:10.1158/1538-7445.AM2015-1261