Comprehensive characterization of protein modifications using mass spectrometry and dry blood spots.

PURPOSE: The main objective of this study is to characterize and analyze modified peptides in DBS samples. This includes deciphering their specific PTMs and understanding their potential impact on the population or disease cohort under study. EXPERIMENTAL DESIGN: Using mass spectrometry-based proteomic approaches, we performed a comprehensive analysis of DBS samples. Our focus was on the identification and quantification of modified peptides. We also took advantage of recent advances in DBS mass spectrometry to ensure accurate detection and quantification. RESULTS: A comprehensive analysis identified 972 modified peptides in DBS samples. Of these, a subset of 211 peptides was consistently present in all samples, highlighting their potential biological importance and relevance. This indicates a diverse spectrum of PTMs in the proteome of DBS samples. CONCLUSIONS AND CLINICAL RELEVANCE: Integration of mass spectrometry and proteomics has revealed a broad spectrum of modified peptides in DBS samples and highlighted their importance in biological processes and disease progression. Accurate detection of these PTMs may be critical for risk stratification and disease management. This study improves the understanding of molecular mechanisms underlying biological processes and disease development, providing important insights for clinical applications.

Gold nanoparticle probes for colorimetric detection of plasma galectin-3: a simple and rapid approach.

Galectin-3 (Gal-3) is a carbohydrate-binding protein associated with the development and progress of heart failure. Here, we report the first colorimetric and low-cost approach for detecting and quantifying Gal-3 using gold nanoparticles (AuNPs) bioconjugated with Gal-3 antibody. The interaction of Gal-3 with the resulting nanoprobes led to a linear response of the absorbance ratio A750nm/A526nm to Gal-3 concentration, accompanied by a change in color intensity. The assay showed a linear optical response even in complex samples, such as saliva and fetal bovine serum (FBS), up to a concentration of 200 µg L-1. The limit of detection (LOD) followed the trend LODPBS (10.0 µg L-1) < LODsaliva (22.6 µg L-1) < LODFBS (25.3 µg L-1). The potential applicability of this method to the analysis of human plasma samples was also demonstrated. Compared to conventional detection techniques, this colorimetric assay provides faster results (∼1 h) and is more cost-effective due to the use of simple and unexpensive equipment. This assay represents an exciting solution for the rapid screening of high risk for rapid progression of heart failure in patient samples (Gal-3 > 25.9 µg L-1).

Gal-3 Protein Expression and Localization in Prostate Tumours.

Gal-3 plays an important role in cell survival, mRNA splicing, and cell-cell and cell-matrix interactions. Depending on its cellular localization and cancer type, Gal-3 may have tumour-suppressive or tumour-promoting activities. Given the promising diagnostic role of Gal-3 in the urine of PCa patients found in our previous study, its concordant gene and protein expression levels, and its involvement in PCa-related biological processes (e.g., morphogenesis of the prostate gland epithelium), we aimed to investigate this protein immunohistochemically in tumour and normal prostate tissues. Gal-3 protein expression was evaluated in 48 tumour prostate tissues, eight normal prostate tissues and 14 adjacent-normal prostate tissues. Decreased Gal-3 staining was detected in tumour tissues compared with normal tissues. Although Gal-3 staining was decreased in tumour tissues with GS 5-8 and pT2 and pT3 stages compared with normal prostate tissue, no correlation was found between Gal-3 expression and PCa progression. In the present study, the pattern of cellular localization differed between groups, as Gal-3 was predominantly excluded from the nucleus in tumour tissues. Furthermore, Gal-3 had no significant effect on survival and relapse in these PCa patients. This work confirms Gal-3 as a promising marker for PCa diagnosis.

Feasibility of Brewer's Spent Yeast Microcapsules as Targeted Oral Carriers.

Brewer's spent yeast (BSY) microcapsules have a complex network of cell-wall polysaccharides that are induced by brewing when compared to the baker's yeast (Saccharomyces cerevisiae) microcapsules. These are rich in (ß1→3)-glucans and covalently linked to (α1→4)- and (ß1→4)-glucans in addition to residual mannoproteins. S. cerevisiae is often used as a drug delivery system due to its immunostimulatory potential conferred by the presence of (ß1→3)-glucans. Similarly, BSY microcapsules could also be used in the encapsulation of compounds or drug delivery systems with the advantage of resisting digestion conferred by (ß1→4)-glucans and promoting a broader immunomodulatory response. This work aims to study the feasibility of BSY microcapsules that are the result of alkali and subcritical water extraction processes, as oral carriers for food and biomedical applications by (1) evaluating the resistance of BSY microcapsules to in vitro digestion (IVD), (2) their recognition by the human Dectin-1 immune receptor after IVD, and (3) the recognition of IVD-solubilized material by different mammalian immune receptors. IVD digested 44-63% of the material, depending on the extraction process. The non-digested material, despite some visible agglutination and deformation of the microcapsules, preserved their spherical shape and was enriched in (ß1→3)-glucans. These microcapsules were all recognized by the human Dectin-1 immune receptor. The digested material was differentially recognized by a variety of lectins of the immune system related to (ß1→3)-glucans, glycogen, and mannans. These results show the potential of BSY microcapsules to be used as oral carriers for food and biomedical applications.

Innovative processes in smart packaging. A systematic review.

Smart packaging provides one possible solution that could reduce greenhouse gas emissions. In comparison with traditional packaging, which aims to extend the product's useful life and to facilitate transport and marketing, smart packaging allows increased efficiency, for example by ensuring authenticity and traceability from the product's origin, preventing fraud and theft, and improving security. Consequently, it may help to reduce pollution, food losses, and waste associated with the food supply chain. However, some questions must be answered to fully understand the advantages and limitations of its use. What are the most suitable smart packaging technologies for use in agro-industrial subsectors such as meat, dairy, fruits, and vegetables, bakery, and pastry? What are the opportunities from a perspective of life extension, process optimization, traceability, product quality, and safety? What are the future challenges? An up-to-date, systematic review was conducted of literature relevant to the application of indicator technologies, sensors, and data carriers in smart packaging, to answer these questions. © 2022 Society of Chemical Industry.

What is the role of active packaging in the future of food sustainability? A systematic review.

Nowadays, the strong increase in products consumption, the purchase of products on online platforms as well as the requirements for greater safety and food protection are a concern for food and packaging industries. Active packaging brings huge advances in the extension of product shelf-life and food degradation and losses reduction. This systematic work aims to collect and evaluate all existing strategies and technologies of active packaging that can be applied in food products, with a global view of new possibilities for food preservation. Oxygen scavengers, carbon dioxide emitters/absorbers, ethylene scavengers, antimicrobial and antioxidant active packaging, and other active systems and technologies are summarized including the products commercially available and the respective mechanisms of action. © 2022 Society of Chemical Industry.

Galectin-3 in prostate cancer and heart diseases: a biomarker for these two frightening pathologies?

Galectin-3 (Gal-3) belongs to galectin protein family, a type of ß-galactose-binding lectin having more than one evolutionarily conserved domain of carbohydrate recognition. Gal-3 is mainly located in the cytoplasm, but it also enters the nucleus and is secreted into the extracellular environment and biological fluids such as urine, saliva, and serum. It plays an important role in many biological functions, such as angiogenesis, apoptosis, cell differentiation, cell growth, fibrosis, inflammation, host defense, cellular modification, splicing of pre-mRNA, and transformation. Many previous studies have shown that Gal-3 can be used as a diagnostic or prognostic biomarker for heart ailments, kidney diseases, and other major illnesses including cancer. Moreover, it may also play a major role in risk stratification in different diseases, and in this review, we have summarized the potential roles and application of Gal-3 as diagnostic, prognostic, and risk stratifying biomarker from previously reported studies in heart diseases and cancer, with special emphasis on prostate cancer.

A peptide-centric approach to analyse quantitative proteomics data- an application to prostate cancer biomarker discovery.

Bottom-up proteomics is a popular approach in molecular biomarker research. However, protein analysts have realized the limitations of protein-based approaches for identifying and quantifying proteins in complex samples, such as the identification of peptides sequences shared by multiple proteins and the difficulty in identifying modified peptides. Thus, there are many exciting opportunities to improve analysis methods. Here, an alternative method focused on peptide analysis is proposed as a complement to the conventional proteomics data analysis. To investigate this hypothesis, a peptide-centric approach was applied to reanalyse a urine proteome dataset of samples from prostate cancer patients and controls. The results were compared with the conventional protein-centric approach. The relevant proteins/peptides to discriminate the groups were detected based on two approaches, p-value and VIP values obtained by a PLS-DA model. A comparison of the two strategies revealed high inconsistency between protein and peptide information and greater involvement of peptides in key PCa processes. This peptide analysis unveiled discriminative features that are lost when proteins are analyzed as homogeneous entities. This type of analysis is innovative in PCa and integrated with the widely used protein-centric approach might provide a more comprehensive view of this disease and revolutionize biomarker discovery. SIGNIFICANCE: In this study, the application of a protein and peptide-centric approaches to reanalyse a urine proteome dataset from prostate cancer (PCa) patients and controls showed that many relevant proteins/peptides are missed by the conservative nature of p-value in statistical tests, therefore, the inclusion of variable selection methods in the analysis of the dataset reported in this work is fruitful. Comparison of protein- and peptide-based approaches revealed a high inconsistency between protein and peptide information and a greater involvement of peptides in key PCa processes. These results provide a new perspective to analyse proteomics data and detect relevant targets based on the integration of peptide and protein information. This data integration allows to unravel discriminative features that normally go unnoticed, to have a more comprehensive view of the disease pathophysiology and to open new avenues for the discovery of biomarkers.

Interaction of Colloidal Gold Nanoparticles with Urine and Saliva Biofluids: An Exploratory Study.

The use of gold nanoparticles for drug delivery, photothermal or photodynamic therapy, and biosensing enhances the demand for knowledge about the protein corona formed on the surface of nanoparticles. In this study, gold nanospheres (AuNSs), gold nanorods (AuNRs), and gold nanoflowers (AuNFs) were incubated with saliva or urine. After the interaction, the surface of gold nanoparticles was investigated using UV-VIS spectroscopy, zeta potential, and dynamic light scattering. The shifting of the localized surface plasmon resonance (LSPR) band, the increase in hydrodynamic diameter, and the changes in the surface charge of nanoparticles indicated the presence of biomolecules on the surface of AuNSs, AuNRs, and AuNFs. The incubation of AuNFs with saliva led to nanoparticle aggregation and minimal protein adsorption. AuNSs and AuNRs incubated in saliva were analyzed through liquid chromatography with tandem mass spectrometry (LC-MS/MS) to identify the 96 proteins adsorbed on the surface of the gold nanoparticles. Among the 20 most abundant proteins identified, 14 proteins were common in both AuNSs and AuNRs. We hypothesize that the adsorption of these proteins was due to their high sulfur content, allowing for their interaction with gold nanoparticles via the Au-S bond. The presence of distinct proteins on the surface of AuNSs or AuNRs was also investigated and possibly related to the competition between proteins present on the external layers of corona and gold nanoparticle morphology.

Label-free dynamic light scattering assay for C-reactive protein detection using magnetic nanoparticles.

In this work, we present a simple method for label-free detection of C-reactive protein (CRP) in diluted saliva samples without the use of specific molecules against CRP. We use the dynamic light scattering (DLS) technique and silica-coated Fe3O4 nanoparticles (∼50 nm in diameter) functionalized with amino carboxylate moieties (Fe3O4@SiO2/COOH) as probes. After contact with the sample, the particles could be easily separated with a handy magnet and redispersed for DLS analysis simply by vortex shaking. The variation of the hydrodynamic diameter of the nanoparticles (Z-average size) could be correlated with the concentration of CRP up to concentrations of 10 mg L-1. The detection limit (LOD) in diluted saliva samples that were spiked with CRP was 0.205 mg L-1, which is below salivary levels of CRP detected in unhealthy individuals. The coefficient of variation was found to be less than 1.5% in the entire detection range. The variation of Z-average size of non-functionalized silica coated nanoparticles (Fe3O4@SiO2) also correlated well with CRP concentration. Nevertheless, the Fe3O4@SiO2/COOH nanoparticles were less susceptible to interference from other biomolecules present in saliva and adsorbed more CRP, indicating higher selectivity toward CRP than nonfunctionalized nanoparticles. This higher affinity was attributed to the chelating interaction between the aminocarboxylate groups of the organosilane N-[3-(trimethoxysilyl)propyl]ethylenediaminetriacetic acid trisodium salt (EDTA-TMS) grafted onto the surface of the Fe3O4@SiO2/COOH nanoparticles and the Ca2+ ions of CRP. LC-MS/MS analyses allowed identification of the proteins adsorbed on the nanoparticles and confirmation of the presence of CRP, which is involved in several biological processes, including immune response, response to stress and transport.

ß-Glucan-Functionalized Nanoparticles Down-Modulate the Proinflammatory Response of Mononuclear Phagocytes Challenged with Candida albicans.

Systemic fungal infections are associated with significant morbidity and mortality, and Candida albicans is the most common causative agent. Recognition of yeast cells by immune cell surface receptors can trigger phagocytosis of fungal pathogens and a pro-inflammatory response that may contribute to fungal elimination. Nevertheless, the elicited inflammatory response may be deleterious to the host by causing excessive tissue damage. We developed a nanoparticle-based approach to modulate the host deleterious inflammatory consequences of fungal infection by using ß1,3-glucan-functionalized polystyrene (ß-Glc-PS) nanoparticles. ß-Glc-PS nanoparticles decreased the levels of the proinflammatory cytokines TNF-α, IL-6, IL-1ß and IL-12p40 detected in in vitro culture supernatants of bone marrow-derived dendritic cells and macrophage challenged with C. albicans cells. Moreover, ß-Glc-PS nanoparticles impaired the production of reactive oxygen species by bone marrow-derived dendritic cells incubated with C. albicans. This immunomodulatory effect was dependent on the nanoparticle size. Overall, ß-Glc-PS nanoparticles reduced the proinflammatory response elicited by fungal cells in mononuclear phagocytes, setting the basis for a targeted therapy aimed at protecting the host by lowering the inflammatory cost of infection.

Do men and women have different musculoskeletal symptoms at the same musculoskeletal discomfort level?

Owing to biological and social factors, illness-related musculoskeletal symptoms tend to vary between men and women. However, in the past, conceptualised discomfort metrics were applied uniformly to both genders. This study aimed to develop a scale to measure musculoskeletal discomfort that compares the symptoms between men and women. The scale aimed to determine the gender-based response patterns related to symptoms. A total of 707 men and 1302 women reported their symptoms on a body map. Factor analysis and item response theory were used to differentiate the identified symptoms in the construction of a musculoskeletal discomfort scale. Differences in work exposure appeared to explain the symptom patterns between men and women. The scale had eight levels, and it was found that at the same level of discomfort, men and women reported symptoms in different body regions.Practitioner summary: On this discomfort scale, the response patterns of men and women were categorised into eight levels. Symptoms differed by gender at the same musculoskeletal discomfort level. This is in contrast to previous studies in which scales were devised without considering differences between the genders.Abbreviations: WMSDs: work-related musculoskeletal disorders; BMI: body mass index; FA: factor analysis; IRT: item response theory; KMO: Kaiser-Meyer-Olkin; BST: Bartlett's test of sphericity; F: factor loading; h2: communality; α: Cronbach's alpha; ωt: McDonald's omega; ai: parameters of discrimination of the items; bik: parameters of difficulty of response categories; θj: latent trait; RMSEA: root mean square error of approximation; CFI: comparative fit index; TLI: Tucker-Lewis index; odu: musculoskeletal discomfort units; RA: rarely; OF: often; AL: always.

Application of Proteogenomics to Urine Analysis towards the Identification of Novel Biomarkers of Prostate Cancer: An Exploratory Study.

To identify new protein targets for PCa detection, first, a shotgun discovery experiment was performed to characterize the urinary proteome of PCa patients. This revealed 18 differentially abundant urinary proteins in PCa patients. Second, selected targets were clinically tested by immunoblot, and the soluble E-cadherin fragment was detected for the first time in the urine of PCa patients. Third, the proteogenome landscape of these PCa patients was characterized, revealing 1665 mutant protein isoforms. Statistical analysis revealed 6 differentially abundant mutant protein isoforms in PCa patients. Analysis of the likely effects of mutations on protein function and PPIs involving the dysregulated mutant protein isoforms suggests a protective role of mutations HSPG2*Q1062H and VASN*R161Q and an adverse role of AMBP*A286G and CD55*S162L in PCa patients. This work originally characterized the urinary proteome, focusing on the proteogenome profile of PCa patients, which is usually overlooked in the analysis of PCa and body fluids. Combined analysis of mass spectrometry data using two different software packages was performed for the first time in the context of PCa, which increased the robustness of the data analysis. The application of proteogenomics to urine proteomic analysis can be very enriching in mutation-related diseases such as cancer.

Integration of Automatic Text Mining and Genomic and Proteomic Analysis to Unravel Prostate Cancer Biomarkers.

Prostate cancer (PCa) is the most prevalent noncutaneous cancer among men. The limited accuracy and/or invasive nature of the current diagnostic tools have driven the demand for new and noninvasive biomarkers. Urine as a noninvasive sample that contains prostatic secretions is a promising source of PCa markers. The automatic text-mining functionality of VOSviewer was used to retrieve and create co-occurrence networks of terms associated with PCa. These results were complemented with DisGENET data, a repository of PCa associations, and with a recent bioinformatic analysis integrating all differentially expressed proteins identified in tumor tissue and urine from PCa patients to address the limited term selection of VOSviewer. Afterward, the results were integrated with gene expression data from the Gene Expression Omnibus database to correlate gene and protein levels. This study suggests AXIN2, GSTM2, KLK3, LGALS3, MSMB, PRTFDC1, and SH3RF1 as important entities in PCa context. KLK, LGALS3, and MSMB proteins are common to a previous bioinformatic analysis, and a concordance was found between the levels of gene and protein expression. The applicability of the pipeline presented here was validated by showing altered urinary levels of galectin-3 protein in PCa patients compared to noncancer subjects.

Involvement of the Iron-Regulated Loci hts and fhuC in Biofilm Formation and Survival of Staphylococcus epidermidis within the Host.

Staphylococcus epidermidis is a major nosocomial pathogen with a remarkable ability to persist on indwelling medical devices through biofilm formation. Nevertheless, it remains intriguing how this process is efficiently achieved under the host's harsh conditions, where the availability of nutrients, such as essential metals, is scarce. Following our previous identification of two iron-regulated loci putatively involved in iron transport, hts and fhuC, we assessed here their individual contribution to both bacterial physiology and interaction with host immune cells. Single deletions of the hts and fhuC loci led to marked changes in the cell iron content, which were partly detrimental for planktonic growth and strongly affected biofilm formation under iron-restricted conditions. Deletion of each of these two loci did not lead to major changes in S. epidermidis survival within human macrophages or in an ex vivo human blood model of bloodstream infection. However, the lack of either hts or fhuC loci significantly impaired bacterial survival in vivo in a murine model of bacteremia. Collectively, this study establishes, for the first time, the pivotal role of the iron-regulated loci hts and fhuC in S. epidermidis biofilm formation and survival within the host, providing relevant information for the development of new targeted therapeutics against this pathogen. IMPORTANCE Staphylococcus epidermidis is one of the most important nosocomial pathogens and a major cause of central line-associated bloodstream infections. Once in the bloodstream, this bacterium must surpass severe iron restriction in order to survive and establish infection. Surprisingly, very little is known about the iron acquisition mechanisms in this species. This study represents the first report on the involvement of the S. epidermidis iron-regulated loci hts and fhuC in biofilm formation under host relevant conditions and, most importantly, in survival within the host. Ultimately, these findings highlight iron acquisition and these loci in particular, as potential targets for future therapeutic strategies against biofilm-associated S. epidermidis infections.

Gender differences in associating musculoskeletal complaints, housework, electronic device usage and physical exercise for administrative workers.

Objectives. This study assessed musculoskeletal complaints (MSCs) in administrative workers, associating MSCs with non-paid housework, home use of electronic devices and physical exercise, while keeping a distinctive gender approach. This may promote the development of more effective preventive measures, by meeting the specific strengths and weaknesses of each gender. Methods. Ninety-six administrative workers (58 women and 38 men) who used computers more than 50% of their working time participated in a cross-sectional study. A questionnaire concerning individual socio-demographic data, habits and lifestyle, and including the Nordic musculoskeletal questionnaire (NMQ), was deployed. Gender-based odds ratios for MSCs in body areas over the previous 12 months and correlation coefficients between habits and lifestyle variables and NMQ variables were computed. Results. Women did not incur a higher risk of MSCs than men. Analysis of the association did not yield meaningful associations for either gender. Results suggest giving future consideration to development of gender-specific preventive measures. Conclusion. Computerized work performed concomitantly with physical exposures outside the workplace showed mixed associations with MSCs, according to gender and depending on the kind of exposure. Results are indicative of the need for development of gender-specific preventive measures.

Multiple cardiac rhabdomyomas, prenatal diagnosis. Case report / Rabdomiomas cardiacos múltiples, diagnóstico prenatal. Reporte de casos.

Cardiac tumors are rare in children, having a maximum reported incidence of 0.027% in prenatal diagnosis, increasing the incidence in necropsy diagnosis; rhabdomyomas are the most frequent cardiac tumors, some cases are associated with tuberous sclerosis. We present the report of two cases in our unit that were diagnosed prenatal with follow-up after birth and one of them was associated with tuberous sclerosis. Both cases were admitted in a third level center, uncomplicated, without requiring surgical treatment, and could be discharged.

Los tumores cardiacos son raros en la población infantil, teniendo una incidencia máxima reportada del 0.027% en el diagnóstico prenatal, incrementándose la incidencia en el diagnóstico por necropsia. Los rabdomiomas son los tumores cardiacos más frecuentes, algunos casos pueden asociarse con esclerosis tuberosa. Presentamos el reporte de dos casos en nuestra unidad a los cuales se les realizó diagnóstico prenatal con seguimiento posterior al nacimiento y asociándose uno de ellos a esclerosis tuberosa. Ambos casos recibieron atención en un centro de tercer nivel, sin complicaciones, sin requerir tratamiento quirúrgico, pudiendo ser egresados.

Isolation and identification of an arabinogalactan extracted from pistachio external hull: Assessment of immunostimulatory activity.

This work studies the extraction and purification of a novel arabinogalactan from pistachio external hull. It was extracted with a simple method from pistachio hull which is considered as unexploited waste. Based on the results of sugar analysis by GC-FID, glycosidic linkage by GC-MS, NMR spectroscopy, and molecular weight by Size Exclusion Chromatography, pistachio hull water soluble polysaccharides (PHWSP) were identified as a type II arabinogalactan (AG), with characteristic terminally linked α-Araf, (α1 â†’ 5)-Araf, (α1 â†’ 3,5)-Araf, terminally linked ß-Galp, (ß1 â†’ 6)-Galp, and (ß1 â†’ 3,6)-Galp. DEPT-135, HSQC, HMBC and COSY NMR data suggested the presence of (ß1 â†’ 3)-Galp mainly branched at O-6 with (ß1 â†’ 6)-Galp chains, α-Araf chains, and terminally linked α-Araf. These AG from pistachio external hulls showed in vitro stimulatory activity for B cells, suggesting their possible use as an immunological stimulant in nutraceutical and biomedical applications.

Dectin-1-Mediated Production of Pro-Inflammatory Cytokines Induced by Yeast ß-Glucans in Bovine Monocytes.

Yeast-derived products containing ß-glucans have long been used as feed supplements in domesticated animals in an attempt to increase immunity. ß-glucans are mainly recognized by the cell surface receptor CLEC7A, also designated Dectin-1. Although the immune mechanisms elicited through Dectin-1 activation have been studied in detail in mice and humans, they are poorly understood in other species. Here, we evaluated the response of bovine monocytes to soluble and particulate purified ß-glucans, and also to Zymosan. Our results show that particulate, but not soluble ß-glucans, can upregulate the surface expression of costimulatory molecules CD80 and CD86 on bovine monocytes. In addition, stimulated cells increased production of IL-8 and of TNF, IL1B, and IL6 mRNA expression, in a dose-dependent manner, which correlated positively with CLEC7A gene expression. Production of IL-8 and TNF expression decreased significantly after CLEC7A knockdown using two different pairs of siRNAs. Overall, we demonstrated here that bovine monocytes respond to particulate ß-glucans, through Dectin-1, by increasing the expression of pro-inflammatory cytokines. Our data support further studies in cattle on the induction of trained immunity using dietary ß-glucans.

Mechanisms underlying the pathophysiology of heart failure with preserved ejection fraction: the tip of the iceberg.

Heart failure with preserved ejection fraction (HFpEF) is a multifaceted syndrome with a complex aetiology often associated with several comorbidities, such as left ventricle pressure overload, diabetes mellitus, obesity, and kidney disease. Its pathophysiology remains obscure mainly due to the complex phenotype induced by all these associated comorbidities and to the scarcity of animal models that adequately mimic HFpEF. Increased oxidative stress, inflammation, and endothelial dysfunction are currently accepted as key players in HFpEF pathophysiology. However, we have just started to unveil HFpEF complexity and the role of calcium handling, energetic metabolism, and mitochondrial function remain to clarify. Indeed, the enlightenment of such cellular and molecular mechanisms represents an opportunity to develop novel therapeutic approaches and thus to improve HFpEF treatment options. In the last decades, the number of research groups dedicated to studying HFpEF has increased, denoting the importance and the magnitude achieved by this syndrome. In the current technological and web world, the amount of information is overwhelming, driving us not only to compile the most relevant information about the theme but also to explore beyond the tip of the iceberg. Thus, this review aims to encompass the most recent knowledge related to HFpEF or HFpEF-associated comorbidities, focusing mainly on myocardial metabolism, oxidative stress, and energetic pathways.