Archivio Istituzionale della Ricerca - Università degli Studi della Campania "Luigi Vanvitelli"
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Intoduction. Inclusive Marketing and Value Creation Strategies
No full textIn recent years, growing attention from academics and marketers has been
devoted to “inclusive marketing” or “diversity marketing” as a means to
create value for the organisation, the market, and society in general by
reflecting the diversities of consumer groups with a “socially inclusive
approach”. Driven by changing consumer expectations and social
movements, brands have been increasingly required to meet Diversity,
Equity, and Inclusion (DEI) themes while maintaining business profitability. This chapter proposes a
model for inclusive marketing. This
model is intended to contribute to the emerging stream of inclusive
marketing literature and help marketers conform to a socially inclusive
approach, enhance the effectiveness of their marketing programs, and
create both marketing value and social value
Steel exoskeletons for low-impact and Integrated seismic retrofit of existing buildings
No full textA significant portion of Europe's building stock was constructed following World War II. The absence of updates and enhancements has left this constructed environment exceedingly vulnerable to seismic events and energy inefficient. Due to PNRR tax incentives, there is a distinct necessity to delineate and supply qualified professionals with integrated, efficient, prompt, and cost-effective retrofit solutions. These treatments ought to be formulated with a comprehensive approach and grounded in the principle of Life Cycle Thinking. Traditional seismic retrofitting is often hindered by high costs, long execution times requiring building halts, and environmental concerns stemming from excessive material usage. This thesis investigates modern, non-disruptive, and environmentally conscious alternatives, focusing on external steel frame structures, or exoskeletons, which facilitate an integrated approach to resilience and energy improvements. Exoskeletons leverage the simplicity of steel assembly to reduce costs, minimize operational disruptions, and improve structural resilience. They are also compatible with dissipative or self-centering devices for improved seismic performance. The integration of energy-dissipating devices allows for the absorption and dissipation of seismic energy via inelastic deformations, thereby decreasing stress on the existing reinforced concrete structure. Such a retrofit provides a precise strategy, preventing excessive strengthening and concentrating on seismic demands. This leads to lower loads for new foundations and minimal impact on the original building, potentially diminishing overall intervention costs. In this scenario, innovative devices have been selected that can dissipate energy resulting from seismic activity, as well as seek to improve the resilience of the structure through devices that allow the building to return to its plumb position. To achieve this objective, the devices analyzed within this thesis work are rectangular and hourglass-shaped Buckling-Restrained Aluminum Shear-Yielding Plates; Dumbbell-shaped Steel Slit Dampers; Shape-Memory-Alloy-based dampers. This thesis applies and sizes several exoskeleton systems for strategic RC structures. The investigated retrofit strategies, all characterized as steel exoskeletons, include Dissipative Eccentric Braced Frames, Self-Centering Concentrically Braced Frames, Self-Centering Dual-Rocking Frames, and Base-Rocking Dual-Core Frames. The design of these interventions utilizes the Displacement-based Design approach. This method features constraints that may emerge when employed to build a retrofit for an actual case study. This thesis addresses several issues arising from the retrofit system or the as-built structure. The considerations encompass the energy dissipation capacity of the installed device and the connection system between the structure and the exoskeleton, which must guarantee the complete transmission of stresses from seismic forces while preserving rigid behavior, including the validation of the initial rigid diaphragm assumption. The retrofit strategies investigated in the matter of this work were thought and designed for real-world strategic RC buildings case studies. Their capacity to handle seismic forces was assessed starting from the linear (eigenvalue) modal and nonlinear (pushover) static analyses by Eurocode 8. Based on collapse mechanisms of each structure, the target displacement was defined, which is useful for designing the various interventions. The effectiveness of the retrofit strategy and the design procedure have been assessed by nonlinear Time-History analyses under a set of earthquake-strong ground motions selected and scaled for the Collapse Prevention Limit State based on the spectrum-compatibility rules of the Italian Code (2018)
Optimizing electronic cooling: Harnessing potentials of impinging jet flows with metal foam heat sinks for superior thermal management
No full textEffective heat dissipation is essential for cooling power-intensive chips and densely packed electronics with significant heat output such as those used in data centers. This study explores the hybrid thermal management strategies that combine the use of impinging jet flow (IJF) and high-porosity metal foam (MF) heat sinks to enhance convective heat transfer. Unlike conventional jet cooling systems, the integration of metal foam introduces a highly conductive, porous medium that amplifies surface area, promotes flow mixing, and accelerates thermal diffusion — resulting in 1 - 2 times higher heat transfer compared to clear cases (CC). A 2D axisymmetric numerical model is developed in ANSYS Fluent using the Reynolds-Averaged Navier–Stokes (RANS) equations with the standard k-ε turbulence model. The setup features a single circular nozzle, aluminum target plate, aluminum metal foams with air as a cooling fluid. Key parameters including nozzle-to-plate spacing (IH = 2.8 – 8.5), Peclet number (Pe = 2200 – 17,000), and foam porosity (ε = 0.90 – 0.95) — are systematically varied. The findings indicate that minimizing porosity and internal heating (IH) substantially enhances heat transfer performance. A novel correlation for average Nusselt number (Nu ̅) has been developed, accompanied by an enhancement factor that quantifies the thermal improvement across the analyzed configurations. This work provides a robust framework for optimizing hybrid cooling systems and can be extended to explore alternative working fluids, foam geometries, and transient thermal loads in future studies
Circulating levels of chemokines and cardiometabolic profile in adults with type 1 diabetes: an observational study
No full textAims – People with type 1 diabetes (T1D) face an increased cardiovascular (CV) risk compared to the general population. While increasing evidence supports the role for chemokines, including RANTES and CXCL1, in the pathogenesis of T1D, their relationship with CV risk profile remains debated. We aimed to stratify a cohort of adults with T1D according to their cardiometabolic profile and investigate the association between these chemokines and established risk determinants.
Methods – We conducted an observational study enrolling adults of both sexes with type 1 diabetes on stable intensive insulin therapy. Participants were stratified based on three key cardio-metabolic determinants: 1) 10-years cardiovascular risk calculated by the Steno Type 1 Risk Engine (ST1RE), 2) insulin resistance defined by an estimated Glucose Disposal Rate (eGDR) < 8 and 3) body mass index (BMI) ≥ 25 kg/m2. Circulating levels of RANTES and CXCL1 were quantified using enzyme-linked immunosorbent assays (ELISA) kits.
Results – We included 67 adults, with median age of 28 years and diabetes duration of 17 years. Prevalence of insulin resistance and overweight/obesity was 31.3% and 44.8%, respectively. RANTES levels were significantly higher in overweight/obese participants compared with normal-weight peers (overweight/obese vs normal weight, 4010.4 vs 2684.2, pg/ml, P=0.044). Conversely, CXCL1 levels were significantly lower in insulin-resistant individuals (eGDR < 8 vs eGDR ≥ 8, 195.1 vs 319.0, pg/ml, P=0.048). Furthermore, CXCL1 levels were higher in people with shorter diabetes duration (<17 years) compared to those with longer diabetes duration (P=0.007). Regression analysis confirmed an inverse association between CXCL1 levels and diabetes duration (β=-0.320, P=0.008), while no other significant association was found between chemokines and the determinants of cardio-metabolic profile.
Conclusions – In adults with T1D, the circulating levels of RANTES reflect obesity or overweight status, whereas CXCL1 levels are inversely associated with disease duration. Neither chemokine correlated with the estimated cardiovascular risk score (ST1RE) or insulin resistance in regression models, suggesting that these inflammatory markers may not reflect the current cardio-metabolic burden in young adults with established disease and low-to-moderate risk
On Some Analytical Questions Concerning the Dynamics of Newtonian and non-Newtonian Incompressible Fluids
No full textThe purpose of this thesis is to present several results concerning different topics and problems
in Fluid Dynamics: more specifically, the well-posedness of problems involving Newtonian fluids
and a particular class of non-Newtonian fluids, the power-law fluids, is investigated. It is based
on the results presented in [25, 26, 46, 45] and is organized into four chapters:
• In Chapters 1 and 2, we study the well-posedness of a parabolic p-Laplacian system with
a convective term, derived from the power-law system in the subquadratic case (p < 2),
by replacing the symmetric gradient with the full gradient and eliminating the pressure
term. It should be noted that these modifications make the results less relevant from
a Fluid Dynamics perspective, since the corresponding constitutive law does not comply
with the principle of material invariance. Nevertheless, they are useful to better delimit
the expectations for possible results in the fluid dynamics context.
We estabilish existence and a maximum principle for regular solutions (for p∈ 3
2 ,2 ) and
weak solutions (for p∈(1,2)) for an initial datum v◦(x) ∈L∞(Ω); for regular solutions we
analyze the property of extinction in a finite time under suitable smallness assumptions
on the initial datum. Moreover, for v◦(x) ∈L∞(Ω) ∩W1,2
0 (Ω), we are able to prove the
uniqueness of regular solutions for p∈ 5
3 ,2.
• In Chapter 3, we focus on a different problem concerning power-law fluids: the initial
value problem in a spatially periodic domain, with the goal of constructing a weak solution
satisfying an energy equality. The result is a weak solution that satisfies a sort of energy
equality, i.e., an energy equality with additional dissipation.
• InChapter4, weinvestigateaprobleminthecontextofNewtonianfluidtheory, focusingon
theStokessysteminthehalf-spacewithintheframeworkofweightedLebesguespaces. This
analysis is motivated by the long-term goal of understanding the Navier–Stokes equations
in weighted settings, not only in the half-space but also in exterior domains. In particular,
we introduce a class of weights defined as the products of powers of distances from fixed
points, and we establish existence, uniqueness, and regularity results for strong solutions
to the Stokes problem in the half-space.
Moreover, in the Appendix we present some background material, and a comprehensive bibliog-
raphy of the relevant literature is provided
p57 phosphorylations: importance in protein stability, localization, and interactions
No full textp57Kip2 is a 316-amino-acid protein encoded by the CDKN1C gene. It is a cyclin-dependent kinase inhibitor belonging to the CIP/Kip family, which also includes p21CIP1/Waf1 and p27Kip1. It was originally identified by sequence homology with its siblings. P57 is the least studied CIP/Kip protein so far.
Structurally, p57 shares with the other CIP/Kip proteins a highly conserved domain in the N- terminal region called the Kinase Inhibitory Domain, which is necessary and sufficient for binding and inhibiting cyclin/CDK (Cyclin-Dependent Kinase) complexes. CIP/KIP proteins act on different cyclin/CDK complexes principally active in G1 phase and in G1/S by targeting the complex cyclin D/CDK4(6) and cyclin E(A)/CDK2), and they work in G2/M by inhibiting cyclin B/CDK2 (Besson et al., 2008). In the C-terminal region, p57 contains, similarly to p21, a PCNA (Proliferating Cell Nuclear Antigen) binding domain and, similarly to p27, the QT (Glutamine, Threonine) domain, involved mostly in CDK-independent interactions. Unlike the other members of the family and unlike the mouse protein, human p57 has a central region composed of a highly repeated sequence of proline and alanine residues (“PAPA region”), whose function, still undefined, has been associated with the possibility of interacting with different binding partners.
In a previous work from the research group where I carried out my thesis, an NLS was identified in the C-terminal of the protein, defining a bipartite NLS for human p57 (Stampone et al., 2021).
For a long time, the importance of p57 has been associated with its central role in embryogenesis. Indeed, p57 is the only CKI that is shown to be essential during embryogenesis (Yan et al., 1997; Zhang et al.,1997; Takahashi et al., 2000) and it is demonstrated by mice lacking Cdkn1c die immediately after birth due to dyspnea caused by severe developmental abnormalities (Yan et al., 1997). Furthermore, alterations in CDKN1C coding sequence are implicated in the pathogenesis of different growth syndromes associated with various developmental defects such as Beckwith-Wiedemann Syndrome (Brioude et al., 2015), Silver- Russell Syndrome (Binder et al., 2020) and IMAGe (Arboleda et al., 2012). These observations helped to define a key role for p57 in embryonic development, in cell commitment processes, in cellular differentiation, and in the maintenance of cellular homeostasis (Stampone et al., 2018).
Unlike the other CIP/Kip proteins, p57 is not ubiquitous, and its expression is restricted to
specific tissues and organs such as lung, kidney, pancreas, testes, placenta, nervous system, and skeletal muscle. p57 is downregulated in many tumors (Borriello et al., 2011) and given its function as an inhibitor of cyclin/CDK complexes, it has been defined as a tumor suppressor (Coley et al., 2012).
It should be noted that p57 is an intrinsically unstructured protein, a characteristic that allows it to adapt to numerous interactors (Creff et al., 2020). Being an IUP, its localization, stability, and binding to functional partners may be strongly modulated by post-translational modifications. The post-translational modifications contribute to this complexity, modulating its localization and stability.
Only limited data are currently available regarding the post-translational regulation of the protein. Literature data, together with those obtained in our laboratory, have shown that p57 undergoes several post-translational modifications. Indeed, in a paper published by the research group where the current study was conducted, it was shown that in a chronic myeloid leukemia cell line, K562, p57 is mostly phosphorylated, and it can be considered a phosphoprotein (Borriello et al., 2011). These observations suggest that identifying and functionally characterizing the phospho-isoforms of the protein is of central importance for understanding its functions.
Thus, this thesis focused on the characterization of the phosphorylation pattern of p57. It was
characterized in tumoral and no-tumoral cell lines as well as in asynchronous and synchronized cells in the different cell cycle phases and subsequently analyzed by bidimensional analysis (2D-SDS-PAGE/IB) and immunoblotting. The presence of similar two-dimensional patterns of p57 in different cell lines suggests the conservation of the main phosphorylated isoforms of the protein in different phenotypes. Moreover, the bidimensional analysis allowed us to also determine the presence of different forms of phosphorylation, from the monophosphorylated to the bis- o tri- phosphorylated isoforms, and it was shown that they differently distribute in the cellular compartments, nucleus and cytoplasm. Specifically, a clear separation of p57 isoforms emerged, with the unmodified form and a monophosphorylated form detected exclusively in the nucleus, while the more phosphorylated forms were particularly abundant in the cytoplasm. The absence of the unmodified form in the cytoplasm and the different distribution of p57 phosphoisoforms allowed us to hypothesize that the phosphorylation state of the protein may modulate its nuclear/cytoplasmic translocation and/or that the different phosphoisoforms specifically control distinct processes in the two cellular compartments. This implies that p57 may perform distinct roles based on its phosphorylation state and the cellular compartment in which it is found. Thus, the identification of the p57 phosphoisoforms that bind to the CDKs implicated in the G1/S and G2/M phase transitions was crucial. Previous mechanistic studies have shown that p57 inhibits the kinase activity of different cyclin–CDK complexes to different extents (Creff et al., 2020) and our finding that distinct p57 phosphoisoforms are associated with different cyclin/CDK complexes might help in explaining the heterogeneous effects of p57 on the kinase activity.
Additionally, phosphorylation determines the stability of the protein by determining its degradation and nucleus-cytoplasm shuttling, making it able to interact with recognized interactors, including LIMK1 and CRM1 (Stampone et al., 2024). All the CIP/Kip have a role in regulating the dynamics of actin–myosin stress fibers and inhibit at different levels the RhoA/ROCK/LIMK1 pathway. Specifically, the PAPA region of p57 is reported to interact with LIMK1 (Vlachos et al., 2009).
Furthermore, it has been proposed that the cytoplasmic delocalization of p57 may be due to binding with CRM1. Experiments of co-immunoprecipitation followed by 2D/WB analysis from the identification of the p57 phosphoisoforms interacting with the two proteins showed a specificity of binding.
As the last step of this study, we perform experiments aimed at identifying the specific residues of p57 that are potentially phosphorylated and their functional roles. Using in silico analysis on the hp57 coding sequence, the more probable phosphorylatable serine/threonine residues have been identified. Based on the in silico prediction, a series of site-directed experiments have been carried out to substitute specific serine/threonine with nonphosphorylatable residues. Among all analyzed residues, the T147A substitution produced the highest effect on the 2D isoform pattern of p57 fragments, and therefore we focused our analysis on this site.
Since the interaction of p57 with CDKs is regulated by its phosphorylation status and different p57 phosphoisoforms are associated with specific CDKs in a cell-cycle-dependent manner, we evaluate whether the absence of phosphorylation at Thr147 could influence CDK binding affinity. We demonstrated that the phosphorylation of T147 is involved in the binding to the CDK2. Notably, pT147A p57 interacts with CDK2 with a higher affinity compared to the p57 WT. This data enhanced the hypothesis that phosphorylations (including at least in part T147) have an essential role for controlling the binding of CKI with CDK2.p57Kip2 (p57) è una proteina di 316 aminoacidi codificata dal gene CDKN1C. È un inibitore delle chinasi ciclina-dipendente (CKI) appartenente alla famiglia CIP/Kip, che comprende anche p21CIP/Waf1 e p27Kip1. È stata identificata per omologia di sequenza con gli altri membri, ed è la proteina CIP/Kip meno studiata.
Strutturalmente, p57 condivide con le altre CIP/Kip un dominio molto conservato nella regione N-terminale chiamato Kinase Inhibitory Domain (KID), necessario e sufficiente per il legame e l’inibizione dei complessi ciclina/CDK principalmente nella transizione G1/S, quali ciclina D/CDK4(6) e ciclina E(A)/CDK2 (Besson et al., 2008). Nella regione C-terminale p57 presenta, in analogia con p21, un dominio di legame a PCNA (Proliferating Cell Nuclear Antigen) e, in analogia con p27, il dominio QT (Glutammina, Treonina) coinvolto in interazioni proteina-proteina per lo più CDK-indipendenti. Diversamente dagli altri membri della famiglia e dalla proteina di topo, p57 umana presenta una regione centrale che è costituita da una sequenza altamente ripetuta di residui di prolina e alanina ("PAPA region”), la cui funzione, non ancora definita, è stata associata alla possibilità di interazione con diversi partner.
In un precedente lavoro del gruppo di ricerca presso cui ho svolto la mia tesi, è stato identificato un NLS nella regione C-terminale della proteina, definendo un NLS bipartito per p57 umana (Stampone et al., 2021).
Per molto tempo l’importanza funzionale di p57 è stata associata ad un ruolo centrale nel processo di embriogenesi. Infatti, p57 è l’unico CKI che si è dimostrato essere essenziale durante l’embriogenesi (Yan et al., 1997; Zhang et al., 1997; Takahashi et al., 2000), come evidenziato dal fatto che i topi Cdkn1c -/- muoiono immediatamente dopo la nascita a causa di dispnea dovuta a gravi anomalie dello sviluppo (Yan et al., 1997). Inoltre, alterazioni nel gene umano CDKN1C hanno un’importanza funzionale nella patogenesi di diverse sindromi dell’accrescimento come la sindrome di Beckwith-Wiedemann (BWS) (Brioude et al., 2015), IMAGe (Arboleda et al., 2012) e Silver Russell (Binder et al., 2020). Queste osservazioni hanno contribuito a definire un ruolo chiave per p57 durante lo sviluppo embrionale, nei processi di “cell commitment”, di differenziamento cellulare e di mantenimento dell’omeostasi cellulare (Stampone et al., 2018).
A differenza delle altre proteine CIP/Kip, p57 non è ubiquitariamente espressa e la sua presenza è limitata a tessuti e organi specifici quali polmone, rene, pancreas, testicoli, placenta, sistema nervoso e muscolo scheletrico. Inoltre, p57 risulta ridotta in molti tumori (Borriello et al., 2011). Di conseguenza, dato il suo ruolo come inibitore dei complessi ciclina/CDK, è stata considerata una proteina con attività di oncosoppressore (Coley et al., 2012).
p57 è una proteina intrinsecamente non strutturata (IUP), una caratteristica che le permette di adattarsi a numerosi interattori (Creff et al., 2020). In quanto IUP, la sua localizzazione, stabilità e capacità di legare partner funzionali possono essere fortemente modulate dalle modifiche post-traduzionali. A generare questo tipo di complessità concorrono le modifiche post- traduzionali che ne modulano anche la localizzazione e la stabilità. Attualmente pochi dati sono disponibili sulla regolazione post-traduzionale della proteina. Dati di letteratura, insieme a quelli ottenuti nel nostro laboratorio, hanno evidenziano come p57 presenti diverse modifiche post-traduzionali.
In particolare, in un lavoro pubblicato dal gruppo di ricerca in cui è stato condotto il presente studio, è stato dimostrato che, nella linea cellulare di leucemia mieloide cronica, K562, p57 è prevalentemente fosforilata e può essere considerata una fosfoproteina (Borriello et al., 2011). Pertanto, l’identificazione e la caratterizzazione funzionale delle fosfoisoforme della proteina appaiono di fondamentale importanza per comprenderne le funzioni di p57.
A tal fine, in questo lavoro di tesi, è stato caratterizzato il pattern di fosforilazione di p57. È stato caratterizzato in modelli cellulari tumorali e non, nonché in cellule asincrone e sincronizzate nelle diverse fasi del ciclo, attraverso analisi di elettroforesi bidimensionale seguita da immunoblotting (2D-SDS-PAGE/IB).
La presenza di pattern bidimensionali di p57 simili in diverse linee cellulari suggerisce il mantenimento delle principali isoforme fosforilate della proteina in diversi fenotipi. Inoltre, numerose isoforme associate a mono, bi- e trifosforilazioni sono state evidenziate. L’analisi bidimensionale ha permesso anche di dimostrare chiare differenze nella distribuzione delle fosfoisoforme nei compartimenti citosolico e nucleare, sia in cellule asincrone sia sincronizzate. In particolare, la forma non modificata è rilevabile solo nel nucleo, mentre le forme più acide sono presenti nel citoplasma. L'assenza della forma non modificata nel citoplasma e la diversa distribuzione delle fosfoisoforme di p57 ci ha permesso di ipotizzare che lo stato di fosforilazione della proteina possa modularne la traslocazione nucleare/citoplasmatica e/o che le diverse fosfoisoforme controllino specificamente processi distinti nei due compartimenti cellulari.
Pertanto, appare probabile che p57 possa svolgere ruoli distinti in base al suo stato di fosforilazione e al compartimento cellulare in cui si trova. L’identificazione delle fosfoisoforme di p57 che legano le CDK coinvolte nelle transizioni G1/S e G2/M è risultata cruciale. Studi precedenti hanno mostrato che p57 inibisce l’attività chinasica di diversi complessi ciclina– CDK con differente efficacia (Creff et al., 2020) e considerando che distinte isoforme fosforilate di p57 sono associate a differenti complessi ciclina/CDK, questo potrebbe contribuire a spiegare gli effetti eterogenei di p57 sull’attività chinasica. È stato inoltre dimostrato che lo stato di fosforilazione di p57 influenza la stabilità della proteina regolandone la sua degradazione e lo shuttling nucleo-citoplasma, rendendo possibile la sua interazione con diversi interattori proteici noti, quali LIMK1 e CRM1 (Stampone et al., 2024).
Tutte le proteine CIP/Kip partecipano al controllo della dinamica dell'actina, inibendo a diversi livelli la via di segnalazione RhoA/ROCK/ LIMK1. In particolare, per la PAPA region di p57 è stata ipotizzata una funzione regolativa dell’interazione con diversi partner, come per LIMK1 (Vlachos et al., 2009). Inoltre, è stato proposto che la delocalizzazione citoplasmatica di p57 possa essere dovuta al legame con CRM1. Al fine di identificare le isoforme di p57 coinvolte in tali interazioni, esperimenti di co-trasfezione e successive immunoprecipitazioni di LIMK1 e CRM1 sono state eseguite.
Dagli esperimenti di co-immunoprecipitazione seguiti da analisi 2D/WB, per l’identificazione delle isoforme di p57 che interagiscono con le due proteine, è emerso che esiste una specificità nella fosforilazione della CKI. Infine, sono stati eseguiti esperimenti mirati all’identificazione dei residui amminoacidici potenzialmente fosforilabili e la loro caratterizzazione funzionale. Attraverso un’analisi di predizione in silico della sequenza codificante di p57 umano sono stati individuati residui di Serina, Treonina e Tirosina con alta probabilità di essere fosforilati. Sulla base dei dati ottenuti, per ottenere informazioni sul ruolo delle diverse fosfoisoforme, è stata effettuata una mutagenesi sito-specifica sostituendo i residui presumibilmente fosforilabili con alanina, al fine di generare varianti non fosforilabili. Tra i residui analizzati, la sostituzione T147A ha prodotto l’effetto più marcato sul pattern bidimensionale della proteina e, per questo motivo, le successive analisi sono state condotte su questo residuo.
Poiché l’interazione di p57 con le CDK è regolata dal suo stato di fosforilazione e diverse fosfoisoforme di p57 si associano a specifiche CDK in maniera dipendente dal ciclo cellulare, abbiamo valutato se l’assenza della fosforilazione in Thr147 potesse influenzare l’affinità di legame con le CDK. Abbiamo dimostrato che la fosforilazione di T147 è coinvolta nel legame con CDK2. In particolare, p57 T147A interagisce con CDK2 con un’affinità maggiore rispetto alla proteina wild-type. Questo dato rafforza l’ipotesi che le fosforilazioni (incluse almeno in parte quelle su T147) svolgano un ruolo essenziale nel controllo del legame tra il CKI e CDK2
Context-dependent responses of tomato (Solanum lycopersicum) to a Chlamydomonas reinhardtii biostimulant extract under saline and non-saline conditions
No full textDisentangling the complex interplay between biostimulants and environmental stress is a key frontier in sustainable agriculture. In particular, robustly distinguishing between a biostimulant’s intrinsic bioactivity and its specific stress-mitigating properties remains a challenge. We used a fully factorial design and high-throughput RNA-sequencing to examine the molecular interaction between an aqueous extract from the microalga Chlamydomonas reinhardtii (MA) and NaCl stress (75 mM) in tomato (Solanum lycopersicum). We assessed vegetative growth, leaf ion content, and performed transcriptomic analysis of leaf tissue. The application of MA significantly improved vegetative growth, increasing leaf area by 16% and leaf hydration (dry matter decreased from 13.49% to 11.47%) regardless of salinity. Factorial transcriptomic analysis revealed that MA’s molecular effects depend on the plant’s stress status, with 138 genes showing a significant Salt × MA interaction. Under salinity, MA suppressed typical osmotic and oxidative stress-response genes, suggesting it reduces stress perception and costly defenses. In non-saline conditions, MA triggered a “priming” effect, upregulating temperature-response genes while downregulating genes involved in energy-heavy ribosome biogenesis, highlighting an anticipatory mechanism that prepares the plant for future challenges while conserving resources. This study provides a conceptual framework for developing next-generation tools to enhance crop resilience through context-aware biostimulant application
Additive benefits of sacubitril/valsartan to dapagliflozin in cardiometabolic and age related cardiac dysfunction
No full textBackground. Pathophysiology of heart failure with preserved ejection fraction (HFpEF) remains incompletely understood due to the marked heterogeneity of affected patients, who are typically older, frequently female, and burdened by multiple comorbidities. HFpEF disproportionately affects postmenopausal women and is strongly associated with central obesity and metabolic dysfunction.
While current guidelines recommend SGLT2i for all HFpEF patients , combination treatments targeting complementary pathways may improve outcomes due to potential synergistic effects. Many HFpEF patients are already receiving treatments for underlying comorbidities or may require additional therapies. In this view, the identification of patient subgroups most likely to benefit from specific therapeutic combinations is crucial. Evidence from PARAGON-HF suggests enhanced responsiveness to angiotensin receptor neprilysin inhibitors (ARNi) in women. Because both aging and obesity amplify
renin-angiotensin-aldosterone system RAAS activation, dual ARNi-SGLT2i therapy may provide synergistic benefit in this subgroup.
Aim. To evaluate the long-term effects of sacubitril/valsartan (S/V) alone or combined with dapagliflozin (D) in a dysmetabolic aging heart failure model (DAHF) using female Fischer-344 rats that reproduce features of metabolic stress, adiposity, and cardiac aging typical of HFpEF.
Methods. 15-month-old Fischer 344 female rats were treated with S/V (60 mg/kg/day) alone DAHF(S/V) or with a combination of S/V and dapagliflozin (0.1mg/Kg/day) DAHF-(S/V+D) for 45 weeks.
Echocardiography and hemodynamic analysis were used to assess cardiac function. Heart tissue was used for molecular and histological analysis. All data were analyzed using one-way ANOVA, and Tukey’s post-test. A p-value <0.05 was considered statistically significant.
Results. Echocardiographic and hemodynamic analyses showed that both treatments effectively improved diastolic dysfunction. These benefits were accompanied by reduced plasma NT-proBNP and myocardial BNP levels, consistent with alleviated wall stress. Both treatments rebalanced RAAS signaling, downregulating ACE1/AT1R and upregulating the ACE2/Ang-(1-7)/Mas axis, thereby restoring a cardioprotective profile and improving metabolic homeostasis. the reductions in body weight and relative visceral and liver fat observed in DAHF-S/V group, with no further significant reduction,
when combined with dapagliflozin, indicating that integrated modulation of RAAS and natriuretic peptide pathways interrupts the “vicious cycle” ectopic fat. Both treatments attenuated fibrosis, with reduced expression of key pro-fibrotic markers (collagen I/III, MMPs, TGF-β/TGF-βR1, SMADs, and pro-fibrotic miRNAs). Notably, S/V and S/V+D reduced inflammation and endothelial dysfunction by decreasing inflammatory markers (NF-κB, MCP-1), cytokines (IL-1β, IL-6), and endothelial indicators (VCAM-1, ET-1, iNOS).
Only the combination therapy restored eNOS expression, in line with the marked reduction in oxidative stress observed exclusively in the S/V+D group, which exhibited lower reactive oxygen species and reactivation of the Nrf2/Keap1 antioxidant pathway. In parallel, the combined treatment strengthened the AMPK/NAMPT/Sirtuin axis, promoting mitochondrial biogenesis, enhanced antioxidant capacity, and improved metabolic resilience. It also reduced the pro-senescent miRNAs miR-34a and miR-146a
both negative regulators of the NAD+-biosynthetic enzyme NAMPT, thereby modulating key senescence-related metabolic pathways. Consistent with these effects, the combination restored NAMPT and SIRT1 expression and decreased acetyl-p53Lys381 and p21CIP, indicating a reversal of senescenceassociated signaling. Moreover, the longevity protein Klotho, typically diminished in aged and dysmetabolic myocardium, was reinstated only by combined therapy, suggesting synergistic reinforcement of AMPK/SIRT1 and Nrf2/Keap1 antioxidant pathways.
Conclusions. Long-term treatment with S/V, especially when combined with DAPA, improves cardiac function but also has a beneficial effects on deeper metabolic, oxidative-stress and ageing-related mechanisms, particularly relevant in older female rats who develop the cardiometabolic HFpEF phenotype. In conclusion, the combination of SGLT2i and ARNi provided enhanced cardiac protection in a model representative of postmenopausal HFpEF with central obesity, offering mechanistic evidence that supports sex-specific therapeutic strategies. These findings strengthen the rationale for a precision medicine approach tailored to elderly women with HFpEF
