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Liraglutide, a type2 diabetes mellitus (T2DM)-related treatment, improves glycemic control and reduces the risks of adverse cardiovascular events in T2DM patients. However, the underlying mechanisms of the above-mentioned beneficial effects of Liraglutide are not well understood. To have better understanding of these mechanisms, we aimed to study the metabolic impacts of Liraglutide on the metabolome and corresponding pathways in T2DM patients, especially metabolism plays a very fundamental role in health and diseases and is influenced by drugs. In this study, plasma samples collected from T2DM patients (n = 20) and taken pre- and post-Liraglutide treatment were used for untargeted metabolomics analyses, including metabolome profiling and metabolic pathway/network analyses. The metabolome profiling analyses identified 93 endogenous metabolites that were significantly affected by Liraglutide treatment where 49 and 44 metabolites were up and down regulated, respectively. Liraglutide caused metabolic alterations impacting metabolic pathways such as pentose and glucuronate interconversion and alanine, aspartate and glutamate metabolism in T2DM patients. Since the last-mentioned pathways are affected by Liraglutide, it could explain partially the overall beneficial effects of Liraglutide in T2DM, especially that glucuronate interconversion pathway is known by its important roles in eliminating toxic and undesirable substances from the human body to maintain good health status. In addition, the metabolism of amino acids induced by Liraglutide could improve the function of immune cells, strengthening the immunity of T2DM patients. Also, Liraglutide induced the level of other metabolites that help in the defense mechanism against oxidative events. Overall, the findings of this study provide a deeper understanding of the underlying mechanisms involved in the beneficial effects of Liraglutide in T2DM from the metabolic aspect.

Abstract Background Among gynaecological malignancies, endometrial cancer (EC) is the most prevalent type of uterine cancer affecting women. This study explored the proteomic profiles of plasma samples obtained from EC patients, those with hyperplasia (Hy), and a control group (CO). A combination of techniques, such as 2D-DIGE, mass spectrometry, and bioinformatics, including pathway analysis, was used to identify proteins with modified expression levels, biomarkers and their associated metabolic pathways in these groups. Methods Thirty-four patients, categorized into three groups—10 with EC, 12 with Hy, and 12 CO—between the ages of 46 and 75 years old were included in the study. Untargeted proteomic analysis was carried out using two-dimensional difference in gel electrophoresis (2D-DIGE) coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Results In all three groups, 114 proteins that were significantly (p ≤ 0.05 and fold change ≥ 1.5) altered were successfully identified using peptide mass fingerprints (PMFs). Compared with those in the control group (CO), the EC samples had 85 differentially expressed proteins (39 upregulated and 46 downregulated), and in the Hy group, 81 proteins were dysregulated (40 upregulated and 41 downregulated) compared to those in the CO group, while 33 proteins exhibited differential regulation (12 upregulated and 21 downregulated) in the EC plasma samples compared to those in the Hy group. Vitamin D binding protein and complement C3 distinguished Hy and EC from CO with the greatest changes in expression. Among the differentially expressed proteins identified, enzymes with catalytic activity represented the largest group (42.9%). In terms of biological processes, most of the proteins were involved in cellular processes (28.8%), followed by metabolic processes (16.7%). STRING analysis for protein interactions revealed that the significantly differentially abundant proteins in the three groups are involved in three main biological processes: signalling of complement and coagulation cascades, regulation of insulin-like growth factor (IGF) transport and uptake by insulin-like growth factor binding proteins (IGFBPs), and plasma lipoprotein assembly, remodelling, and clearance. Conclusion The identified plasma protein markers have the potential to serve as biomarkers for differentiating between EC and Hy, as well as for early diagnosis and monitoring of cancer progression.

IntroductionTreatment and management of obesity is clinically challenging. The inclusion of GLP-1 receptor agonists (GLP1RA) in the medical management of obesity has proven to be efficacious. However, mechanisms underlying the molecular changes arising from GLP1RA treatment in patients with obesity remain to be elucidated.MethodsA single-center, prospective study was undertaken to evaluate the changes in the plasma proteins after liraglutide 3 mg therapy in twenty patients (M/F: 7/13) with obesity (mean BMI 40.65 ± 3.7 kg/m2). Anthropometric and laboratory parameters were measured, and blood samples were collected at two time points: baseline, before initiating treatment (pretreatment group, PT), and after three months of receiving the full dose liraglutide 3 mg (posttreatment group, PoT). An untargeted label-free LC MSMS mass spectrometric approach combined with bioinformatics and network pathway analysis was used to determine changes in the proteomic profiles.ResultsThe mean age of the study participants was 36.0 ± 11.1 years. A statistically significant change was observed in weight, BMI and HbA1c levels between the PT and PoT groups (paired t-test, P < 0.001). A significant dysregulation was noted in the abundances of 151 proteins (31 up and 120 downregulated) between the two groups. The potential biomarkers were evaluated using receiver operating characteristic (ROC) curves. The top ten proteins (area under the curve (AUC) of 0.999 (95% CI)) were identified as potential biomarkers between PT and PoT groups and included Cystatin-B, major vault protein, and plastin-3, which were upregulated, whereas multimerin-2, large ribosomal P2, and proline–rich acidic protein 1 were downregulated in the PoT group compared with the PT group. The top network pathway identified using ingenuity pathway analysis (IPA), centered around dysregulation of MAPK, AKT, and PKc signaling pathways and related to cell-to-cell signaling and interaction, cellular assembly and organization, cellular compromise and a score of 46 with 25 focus proteins.DiscussionThrough label-free quantitative proteomic analysis, our study revealed significant dysregulation of plasma proteins after liraglutide 3 mg treatment in patients with obesity. The alterations in the proteomic profile between the PT and PoT groups demonstrated a decrease in levels of proteins involved in inflammation and oxidative stress pathways. On the other hand proteins involved in the glycolytic and lipolytic metabolic pathways as well as those participating in cytoskeletal and endothelial reorganization were observed to be increased. Understanding actions of liraglutide at a molecular and proteomic levels provides a holistic look into how liraglutide impacts metabolism, induces weight loss and improves overall metabolic health.

(1) Background: Chronic rhinosinusitis (CRS) is a common chronic inflammation of the nasal mucosa and the paranasal sinuses. The pathogenesis of chronic rhinosinusitis (CRS) is multifactorial and, as of yet, not well understood. (2) Methods: Nasal lavage fluid samples were collected from patients diagnosed with chronic sinusitis with nasal polyposis (CRSwNP) (n = 10) and individuals without sinusitis (control group) (n = 10) who had no nasal complaints. In the present study, we used an untargeted label-free LC-MS/MS mass spectrometric approach combined with bioinformatics and network pathway analysis to compare the changes in the proteomic profiles of the CRSwNP group and the control group. Data from LC-MS/MS underwent univariate and multivariate analyses. (3) Results: The proteomic analyses revealed distinct differences in the abundances of nasal lavage fluid proteins between the CRSwNP and control groups: a total of 234 proteins, 151 up- and 83 down-regulated in CRSwNP. Functional Gene Ontology (GO) analysis showed that dysregulated proteins were involved in airway inflammatory reaction, immune response, and oxidative stress. The biomarkers were evaluated using the Receiver Operating Characteristic (ROC) curve; an Area Under the Curve (AUC) of 0.999 (95% CI) identified potential biomarkers between the CRSwNP and control group. EMILIN-3 and RAB11-binding protein RELCH were down-regulated, and Macrophage migration inhibitory factor and deoxyribonuclease-1 were up-regulated, in CRSwNP compared to the control group. (4) Conclusions: These differentially expressed proteins identified in CRSwNP are involved in airway inflammatory reaction, immune response, and oxidative stress. In particular, the identification of increased interleukin-36 gamma (IL-36γ), which contributes to inflammatory response, and a decrease in SOD, in this group are notable findings. In the future, several of these proteins may prove useful for exploring the pathogenesis of nasal polyps and chronic sinusitis or as objective biomarkers for quantitatively monitoring disease progression or response to therapy.

Background: Liraglutide, a long-acting glucagon-like peptide-1 receptor agonist (GLP1RA), is a well-established anti-diabetic drug, has also been approved for the treatment of obesity at a dose of 3 mg. There are a limited number of studies in the literature that have looked at changes in metabolite levels before and after liraglutide treatment in patients with obesity. To this end, in the present study we aimed to explore the changes in the plasma metabolomic profile, using liquid chromatography-high resolution mass spectrometry (LC-HRMS) in patients with obesity. Methods: A single-center prospective study was undertaken to evaluate the effectiveness of 3 mg liraglutide therapy in twenty-three patients (M/F: 8/15) with obesity, mean BMI 40.81 ± 5.04 kg/m2, and mean age of 36 ± 10.9 years, in two groups: at baseline (pre-treatment) and after 12 weeks of treatment (post-treatment). An untargeted metabolomic profiling was conducted in plasma from the pre-treatment and post-treatment groups using LC-HRMS, along with bioinformatics analysis using ingenuity pathway analysis (IPA). Results: The metabolomics analysis revealed a significant (FDR p-value ≤ 0.05, FC 1.5) dysregulation of 161 endogenous metabolites (97 upregulated and 64 downregulated) with distinct separation between the two groups. Among the significantly dysregulated metabolites, the majority of them were identified as belonging to the class of oxidized lipids (oxylipins) that includes arachidonic acid and its derivatives, phosphorglycerophosphates, N-acylated amino acids, steroid hormones, and bile acids. The biomarker analysis conducted using MetaboAnalyst showed PGP (a21:0/PG/F1alpha), an oxidized lipid, as the first metabolite among the list of the top 15 biomarkers, followed by cysteine and estrone. The IPA analysis showed that the dysregulated metabolites impacted the pathway related to cell signaling, free radical scavenging, and molecular transport, and were focused around the dysregulation of NF-κB, ERK, MAPK, PKc, VEGF, insulin, and pro-inflammatory cytokine signaling pathways. Conclusions: The findings suggest that liraglutide treatment reduces inflammation and modulates lipid metabolism and oxidative stress. Our study contributes to a better understanding of the drug’s multifaceted impact on overall metabolism in patients with obesity.

Background: Hypothyroidism is clinically characterized by a decrease in levels of the circulating thyroid hormones namely thyroxine and triiodothyronine. The main treatment for hypothyroidism is thyroid hormone replacement using levothyroxine to normalize serum thyroid hormone levels. Objectives: In this study, we explored the metabolic changes in the plasma of patients with hypothyroidism after reaching a euthyroid state with levothyroxine treatment. Methods: Plasma samples from 18 patients diagnosed as overt hypothyroidism were collected before and after levothyroxine treatment upon reaching a euthyroid state and were analyzed by high-resolution mass spectrometry-based metabolomics. Multivariate and univariate analyses evaluated data to highlight potential metabolic biomarkers. Results: Liquid chromatography-mass spectrometry-based metabolomics revealed a significant decrease in the levels of ceramide, phosphatidylch oline, triglycerides, acylcarnitine, and peptides after levothyroxine treatment; this could indicate a change in the fatty acid transportation system and an enhanced β-oxidation, compared with a hypothyroid state. At the same time, the decrease in the peptides suggested a shift in protein synthesis. In addition, there was a considerable rise in glycocholic acid following therapy, suggesting the involvement of thyroid hormones in stimulating bile acid production and secretion. Conclusions: A metabolomic analysis of patients with hypothyroidism revealed significant changes in several metabolites and lipids after treatment. This study showed the value of the metabolomics technique in providing a complementary understanding of the pathophysiology of hypothyroidism and as a crucial tool for examining the molecular impact of levothyroxine treatment on hypothyroidism. It was an important tool for investigating the therapeutic effects of levothyroxine on hypothyroidism at the molecular level.

Diabetes mellitus is a chronic multisystem disease with a high global prevalence. The glucagon-like peptide-1 (GLP-1) receptor agonist liraglutide is known to lower glucose levels and reduce weight. However, the mechanisms underlying the benefits of liraglutide treatment in patients with type 2 diabetes mellitus (T2DM) remain unclear. Twelve male patients with T2DM (pre and post liraglutide treatment) and HbA1c between 8% and 11% were recruited. In the present study, a two-dimensional difference gel electrophoresis (2D-DIGE) matrix-assisted laser desorption/ionization-time of flight (MALDI TOF) mass spectrometric approach combined with bioinformatics and network pathway analysis was used to explore the urine proteomic profile. The mean age of the patients was 52.4 ± 7.5 years. After treatment with liraglutide, a statistically significant change (p < 0.006) was observed in HbA1c with no significant changes in body weight or markers of dyslipidemia. Two-dimensional difference gel electrophoresis identified significant changes (≥1.5-fold change, ANOVA, p ≤ 0.05) in 32 proteins (4 down- and 28 upregulated) in liraglutide post treatment compared to the pre-treatment state. Albumin, serotransferrin, metallothionein-2 (MT-2), and keratins K1 and K10 were found to be upregulated after liraglutide treatment. The patients showed significant improvement in glycemic control after the 12-week treatment with liraglutide. The renoprotective effect of liraglutide may be linked to the increased urinary abundance of MT-2 and the decreased abundance of zinc alpha 2-glycoprotein (ZAG) and Alpha-1 antitrypsin (α1-AT). More studies are needed to elucidate the molecular mechanisms behind the renoprotective effects of liraglutide.

Uterine cancer is the most prevalent gynecologic malignancy in women worldwide. Endometrial cancer (EC) has an 81% five-year survival rate, depending on disease stage and time of diagnosis. While endometrial cancer is largely treatable when detected early, no established screening techniques are available in clinical practice. As a result, one of the most significant issues in the medical field is the development of novel ways for early cancer identification, which could boost treatment success rates. Liquid chromatography–high-resolution mass spectrometry (LC-HRMS)-based metabolomics was employed to explore the metabolomic markers and pathways unique to this cancer type and link them to the benign endometrial hyperplasia that may progress to cancer in 5% to 25% of patients. The study involved 59 postmenopausal participants, 20 with EC type 1, 20 with benign hyperplasia, and 19 healthy participants. Metabolite distribution changes were analyzed, and 338 of these features were dysregulated and significant. The first two main components, PC1 and PC2, were responsible for 11.5% and 12.2% of the total metabolites, respectively. Compared with the control group (CO), EC samples had 203 differentially expressed metabolites (180 upregulated and 23 downregulated); in hyperplasia (HP), 157 metabolites were dysregulated (127 upregulated and 30 downregulated) compared to the CO group while 21 metabolites exhibited differential regulation (16 upregulated and 5 downregulated) in EC plasma samples compared to the HP group. Hyperplasia samples exhibited similar metabolic changes to those reported in cancer, except for alterations in triglyceride levels, 7a,12 b-dihydroxy-5b-Cholan-24-oic acid, and Hept-2-enedioyl carnitine levels. The metabolites N-heptanoyl glycine and -(Methylthio)-2,3-isopentyl phosphate and formimino glutamic acid can be specific markers for hyperplasia conditions and dimethyl phosphatidyl ethanolamine and 8-isoprostaglandin E2 can be specific markers for EC conditions. Metabolic activities rely on mitochondrial oxidative phosphorylation for energy generation. The changes in metabolites identified in our study indicate that endometrial cancer cells adopt alternative strategies to increase energy production to meet the energy demand, thereby supporting proliferation.

Metformin is the first-line oral medication for treating type 2 diabetes mellitus (T2DM). In the current study, an untargeted lipidomic analytical approach was used to investigate the alterations in the serum lipidome of a cohort of 89 participants, including healthy lean controls and obese diabetic patients, and to examine the alterations associated with metformin administration. A total of 115 lipid molecules were significantly dysregulated (64 up-regulated and 51 down-regulated) in the obese compared to lean controls. However, the levels of 224 lipid molecules were significantly dysregulated (125 up-regulated and 99 down-regulated) in obese diabetic patients compared to the obese group. Metformin administration in obese diabetic patients was associated with significant dysregulation of 54 lipid molecule levels (20 up-regulated and 34 down-regulated). Levels of six molecules belonging to five lipid subclasses were simultaneously dysregulated by the effects of obesity, T2DM, and metformin. These include two putatively annotated triacylglycerols (TGs), one plasmenyl phosphatidylcholine (PC), one phosphatidylglycerol (PGs), one sterol lipid (ST), and one Mannosyl-phosphoinositol ceramide (MIPC). This study provides new insights into our understanding of the lipidomics alterations associated with obesity, T2DM, and metformin and offers a new platform for potential biomarkers for the progression of diabetes and treatment response in obese patients.

BackgroundHyperthyroidism is characterized by increased thyroid hormone production, which impacts various processes, including metabolism and energy expenditure. Yet, the underlying mechanism and subsequent influence of these changes are unknown. Metabolomics is a broad analytical method that enables qualitative and quantitative examination of metabolite level changes in biological systems in response to various stimuli, pathologies, or treatments.ObjectivesThis study uses untargeted metabolomics to explore the potential pathways and metabolic patterns associated with hyperthyroidism treatment.MethodsThe study consisted of 20 patients newly diagnosed with hyperthyroidism who were assessed at baseline and followed up after starting antithyroid treatment. Two blood samples were taken from each patient, pre (hyperthyroid state) and post-treatment (euthyroid state). Hyperthyroid and euthyroid states were identified based on thyroxine and thyroid-stimulating hormone levels. The metabolic alteration associated with antithyroid therapy was investigated using liquid chromatography- high-resolution mass spectrometry. The untargeted metabolomics data was analyzed using both univariate and multivariate analyses using MetaboAnalyst v5.0. The significant metabolic pattern was identified using the lab standard pipeline, which included molecular annotation in the Human Metabolome Database, LipidMap, LipidBlast, and METLIN. The identified metabolites were examined using pathway and network analyses and linked to cellular metabolism.ResultsThe results revealed a strong group separation between the pre- and post-hyperthyroidism treatment (Q2 = 0.573, R2 = 0.995), indicating significant differences in the plasma metabolome after treatment. Eighty-three mass ions were significantly dysregulated, of which 53 and 30 characteristics were up and down-regulated in the post-treatment compared to the pre-treatment group, respectively. The medium-chain acylcarnitines, octanoylcarnitine, and decanoylcarnitine, previously found to rise in hyperthyroid patients, were among the down-regulated metabolites, suggesting that their reduction could be a possible biomarker for monitoring euthyroid restoration. Kynurenine is a downregulated tryptophan metabolite, indicating that the enzyme kynurenine 3-hydroxylase, inhibited in hyperthyroidism, is back functioning. L-cystine, a cysteine dimer produced from cysteine oxidation, was among the down-regulated metabolites, and its accumulation is considered a sign of oxidative stress, which was reported to accompany hyperthyroidism; L-cystine levels dropped, this suggests that the plasma level of L-cystine can be used to monitor the progress of euthyroid state restoration.ConclusionThe plasma metabolome of patients with hyperthyroidism before and after treatments revealed differences in the abundance of several small metabolites. Our findings add to our understanding of hyperthyroidism’s altered metabolome and associated metabolic processes and shed light on acylcarnitines as a new biomarker for treatment monitoring in conjunction with thyroxine and thyroid-stimulating hormone.

Background: Obesity and type 2 diabetes mellitus (T2DM) are characterized by underlying low-grade chronic inflammation. Metformin has been used as the first line of therapy in T2DM as it decreases hepatic glucose production and glucose intestinal absorption, enhances insulin sensitivity and weight loss, and is known to ameliorate inflammation. The mechanisms through which metformin exerts its effect remain unclear. Proteomics has emerged as a unique approach to explore the biological changes associated with diseases, including T2DM. It provides insight into the circulating biomarkers/mediators which could be utilized for disease screening, diagnosis, and prognosis. Methods: This study evaluated the proteomic changes in obese (Ob), obese diabetics (OD), and obese diabetic patients on metformin (ODM) using a 2D DIGE MALDI-TOF mass spectrometric approach. Results: Significant changes in sixteen plasma proteins (15 up and 1 down, ANOVA, p ≤ 0.05; fold change ≥ 1.5) were observed in the ODM group when compared to the Ob and OD groups. Bioinformatic network pathway analysis revealed that the majority of these altered plasma proteins are involved in distinct pathways involving acute-phase response, inflammation, and oxidative response and were centered around HNF4A, ERK, JNK, and insulin signaling pathways. Conclusions: Our study provides important information about the possible biomarkers altered by metformin treatment in obese patients with and without T2DM. These altered plasma proteins are involved in distinct pathways involving acute-phase response, inflammation, and oxidative response and were centered around HNF4A, ERK, JNK, and insulin signaling pathways. The presented proteomic profiling approach may help in identifying potential biomarkers/mediators affected by metformin treatment in T2DM and inform the understanding of metformin’s mechanisms of action.

Goiter is a term to describe the enlargement of the thyroid gland. The pathophysiology and molecular changes behind development of diffuse benign goiter remains unclear. The present study targeted to identify and describe the alterations in the thyroid tissue proteome from patients (obese euthyroid) with benign diffuse goiter (BDG) using proteomics approach. Thyroid tissue samples, from 7 age and sex matched, patients with BDG and 7 controls were obtained at the time of surgery. An untargeted proteomic analysis of the thyroid tissue was performed out utilizing two-dimensional difference (2D-DIGE) in gel electrophoresis followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) for identification of the proteins. Progenesis software was used to identify changes in expression of tissue proteins and found statistically significant differences in abundance in a total of 90 proteins, 46 up and 44 down (1.5-fold change, ANOVA, p ≤ 0.05) in BDG compared to the control group. Bioinformatic analysis using Ingenuity Pathway Analysis (IPA) identified dysregulation of signalling pathways linked to ERK1/2, Glutathione peroxidase and NADPH oxidase associated to organismal injury and abnormalities, endocrine system disorders and cancer. The thyroid tissue proteome in patients with BDG revealed a significant decrease in thyroglobulin along with dysregulation of glycolysis and an increase in prooxidant peroxidase enzymes. Dysregulation of metabolic pathways related to glycolysis, redox proteins, and the proteins associated with maintaining the cytoskeletal structure of the thyrocytes was also identified.

Galloway-Mowat syndrome is a rare autosomal recessive disease characterized by a unique combination of renal and neurological manifestations, including early-onset steroid-resistant nephrotic syndrome, microcephaly, psychomotor delay, and gyral abnormalities of the brain. Most patients die during early childhood. Here, we identified a novel homozygous O-sialoglycoprotein endopeptidase (OSGEP) variant, NM_017807.3:c.973C>G (p.Arg325Gly), in four affected individuals in an extended consanguineous family from Saudi Arabia. We have described the detailed clinical characterization, brain imaging results, and muscle biopsy findings. The described phenotype varied from embryonic lethality to early pregnancy loss or death at the age of 9. Renal disease is often the cause of death. Protein modeling of this OSGEP variant confirmed its pathogenicity. In addition, proteomic analysis of the affected patients proposed a link between the KEOPS complex function and human pathology and suggested potential pathogenic mechanisms.

Metformin is a widely prescribed medication for the treatment of type 2 diabetes mellitus (T2DM). It possesses effective roles in various disorders, including cancer, dyslipidemia, and obesity. However, the underlying mechanisms of metformin's multiple benefits are not fully understood. Herein, a mass spectrometry-based untargeted metabolomics approach was used to investigate the metabolic changes associated with the administration of a single dose of metformin in the plasma of 26 healthy subjects at five-time points; pre-dose, before the maximum concentration of metformin (Cmax), Cmax, after Cmax, and 36 h post-dose. A total of 111 metabolites involved in various biochemical processes were perturbed, with branched-chain amino acid (BCAA) being the most significantly altered pathway. Additionally, the Pearson similarity test revealed that 63 metabolites showed a change in their levels dependent on metformin level. Out of these 63, the level of 36 metabolites was significantly altered by metformin. Significantly altered metformin-dependent metabolites, including hydroxymethyl uracil, propionic acid, glycerophospholipids, and eicosanoids, pointed to fundamental biochemical processes such as lipid network signaling, energy homeostasis, DNA lesion repair mechanisms, and gut microbiota functions that could be linked to the multiple beneficial roles of metformin. Thus, the distinctive metabolic pattern linked to metformin administration can be used as a metabolic signature to predict the potential effect and mechanism of actions of new chemical entities during drug development.

Metformin is widely used in the treatment of Type 2 Diabetes Mellitus (T2DM). However, it is known to have beneficial effects in many other conditions, including obesity and cancer. In this study, we aimed to investigate the metabolic effect of metformin in T2DM and its impact on obesity. A mass spectrometry (MS)-based metabolomics approach was used to analyze samples from two cohorts, including healthy lean and obese control, and lean as well as obese T2DM patients on metformin regimen in the last 6 months. The results show a clear group separation and sample clustering between the study groups due to both T2DM and metformin administration. Seventy-one metabolites were dysregulated in diabetic obese patients (30 up-regulated and 41 down-regulated), and their levels were unchanged with metformin administration. However, 30 metabolites were dysregulated (21 were up-regulated and 9 were down-regulated) and then restored to obese control levels by metformin administration in obese diabetic patients. Furthermore, in obese diabetic patients, the level of 10 metabolites was dysregulated only after metformin administration. Most of these dysregulated metabolites were dipeptides, aliphatic amino acids, nucleic acid derivatives, and urea cycle components. The metabolic pattern of 62 metabolites was persistent, and their levels were affected by neither T2DM nor metformin in obesity. Interestingly, 9 metabolites were significantly dysregulated between lean and obese cohorts due to T2DM and metformin regardless of the obesity status. These include arginine, citrulline, guanidoacetic acid, proline, alanine, taurine, 5-hydroxyindoleacetic acid, and 5-hydroxymethyluracil. Understanding the metabolic alterations taking place upon metformin treatment would shed light on possible molecular targets of metformin, especially in conditions like T2DM and obesity.

Uterine cancers are among the most prevalent gynecological malignancies, and endometrial cancer (EC) is the most common in this group. This study used tissue-based proteomic profiling analysis in patients with endometrial cancer and hyperplasia, and control patients. Conventional 2D gel electrophoresis, followed by a mass spectrometry approach with bioinformatics, including a network pathway analysis pipeline, was used to identify differentially expressed proteins and associated metabolic pathways between the study groups. Thirty-six patients (twelve with endometrial cancer, twelve with hyperplasia, and twelve controls) were enrolled in this study. The mean age of the participants was 46–75 years. Eighty-seven proteins were significantly differentially expressed between the study groups, of which fifty-three were significantly differentially regulated (twenty-eight upregulated and twenty-five downregulated) in the tissue samples of EC patients compared to the control (Ctrl). Furthermore, 26 proteins were significantly dysregulated (8 upregulated and 18 downregulated) in tissue samples of hyperplasia (HY) patients compared to Ctrl. Thirty-two proteins (nineteen upregulated and thirteen downregulated) including desmin, peptidyl prolyl cis-trans isomerase A, and zinc finger protein 844 were downregulated in the EC group compared to the HY group. Additionally, fructose bisphosphate aldolase A, alpha enolase, and keratin type 1 cytoskeletal 10 were upregulated in the EC group compared to those in the HY group. The proteins identified in this study were known to regulate cellular processes (36%), followed by biological regulation (16%). Ingenuity pathway analysis found that proteins that are differentially expressed between EC and HY are linked to AKT, ACTA2, and other signaling pathways. The panels of protein markers identified in this study could be used as potential biomarkers for distinguishing between EC and HY and early diagnosis and progression of EC from hyperplasia and normal patients.

Obesity is associated with an increased risk of insulin resistance (IR) and type 2 diabetes mellitus (T2DM) which is a multi-factorial disease associated with a dysregulated metabolism and can be prevented in pre-diabetic individuals with impaired glucose tolerance. A metabolomic approach emphasizing metabolic pathways is critical to our understanding of this heterogeneous disease. This study aimed to characterize the serum metabolomic fingerprint and multi-metabolite signatures associated with IR and T2DM. Here, we have used untargeted high-performance chemical isotope labeling (CIL) liquid chromatography-mass spectrometry (LC-MS) to identify candidate biomarkers of IR and T2DM in sera from 30 adults of normal weight, 26 obese adults, and 16 adults newly diagnosed with T2DM. Among the 3633 peak pairs detected, 62% were either identified or matched. A group of 78 metabolites were up-regulated and 111 metabolites were down-regulated comparing obese to lean group while 459 metabolites were up-regulated and 166 metabolites were down-regulated comparing T2DM to obese groups. Several metabolites were identified as IR potential biomarkers, including amino acids (Asn, Gln, and His), methionine (Met) sulfoxide, 2-methyl-3-hydroxy-5-formylpyridine-4-carboxylate, serotonin, L-2-amino-3-oxobutanoic acid, and 4,6-dihydroxyquinoline. T2DM was associated with dysregulation of 42 metabolites, including amino acids, amino acids metabolites, and dipeptides. In conclusion, these pilot data have identified IR and T2DM metabolomics panels as potential novel biomarkers of IR and identified metabolites associated with T2DM, with possible diagnostic and therapeutic applications. Further studies to confirm these associations in prospective cohorts are warranted.

Endometrial cancer (EC) is the most common form of gynecological cancer. Type 2 diabetes mellitus is associated with an increased risk of EC. Currently, no proteomic studies have investigated the role of diabetes in endometrial cancers from clinical samples. The present study aims to elucidate the molecular link between diabetes and EC using a proteomic approach. Endometrial tissue samples were obtained from age-matched patients (EC Diabetic and EC Non-Diabetic) during surgery. Untargeted proteomic analysis of the endometrial tissues was carried out using a two-dimensional difference in gel electrophoresis (2D-DIGE) coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI TOF). A total of 53 proteins were identified, with a significant difference in abundance (analysis of variance (ANOVA) test, p ≤ 0.05; fold-change ≥ 1.5) between the two groups, among which 30 were upregulated and 23 downregulated in the EC Diabetic group compared to EC Non-Diabetic. The significantly upregulated proteins included peroxiredoxin-1, vinculin, endoplasmin, annexin A5, calreticulin, and serotransferrin. The significantly downregulated proteins were myosin regulatory light polypeptide 9, Retinol dehydrogenase 12, protein WWC3, intraflagellar transport protein 88 homolog, superoxide dismutase [Cu-Zn], and retinal dehydrogenase 1. The network pathway was related to connective tissue disorder, developmental disorder, and hereditary disorder, with the identified proteins centered around dysregulation of ERK1/2 and F Actin signaling pathways. Cancer-associated protein alterations such as upregulation of peroxiredoxin-1, annexin 5, and iNOS, and downregulation of RDH12, retinaldehyde dehydrogenase 1, SOD1, and MYL 9, were found in the EC tissues of the diabetic group. Differential expression of proteins linked to cancer metastasis, such as the upregulation of vinculin and endoplasmin and downregulation of WWC3 and IFT88, was seen in the patients with diabetes. Calreticulin and alpha-enolase, which might have a role in the interplay between diabetes and EC, need further investigation.

Background and Objectives: Cervical cancer (CC) is the eighth most common cancer among Saudi women of all ages. With limited national data, we aimed to evaluate the public awareness of cervical cancer, CC risk factors, HPV infection, and HPV vaccines in different regions of Saudi Arabia. Materials and Methods: This was a survey-based cross-sectional study that encompassed 564 Saudi women over a period of a month. A self-administrated questionnaire was distributed through different social media platforms. Results: The collected data included sociodemographic variables and questions assessing awareness of CC, and the attitudes toward CC screening and human papillomavirus (HPV) vaccination. Most respondents were aware of CC (84.0%), although their primary source of information was the internet. However, only 45 females (8.0%) had a history of cervical screening. Furthermore, most females did not know that HPV was transmitted sexually (78.9%), or that it caused genital warts (81.7%) and CC (81.9%). Regarding the HPV vaccine, 100 females (17.7%) had heard about it, but only 11 (2.0%) took the vaccine, although more than half of the respondents (54.1%) were willing to take the vaccine after being informed about it. Conclusions: We noticed a remarkable lack of awareness among the respondents regarding HPV’s clinical implications; and the HPV vaccine, and its importance and availability. The main source of information for most of the Saudi women in this study was the internet, which may be an unreliable source, or provide misleading information that may delay screening or discourage vaccination. Thus, organized campaigns by the Ministry of Health or other health-advocating agencies, in addition to screening and vaccination programs, are strongly encouraged.

Background: Bladder cancer is a life-threatening disease and a major cause of cancer-associated complications. The main challenges confronted during the clinical management of bladder cancer are associated with recurrence and disease progression to the muscle-invasive phenotype. Improved early detection of the disease is of paramount importance to prevent disease progression and improve survival. Hence, novel clinically applicable biomarkers for early detection are warranted. Methods: In the current study, a comparative proteomic approach was undertaken using plasma samples to identify protein biomarkers associated with the muscle-invasive phenotype of bladder carcinoma. Isolated plasma proteins were depleted, DIGE-labeled, then subjected to conventional 2D electrophoresis followed by mass spectrometry for identification of differentially expressed proteins. Western blot was used for data validation. Results: Fourteen differentially expressed proteins with statistically significant changes in abundance between the cancer group and control group were identified. Three differentially expressed proteins were selected for validation, among which apolipoprotein A1 exhibited high specificity and sensitivity (AUC = 0.906). Ingenuity pathway analysis identified IFN-γ and TNF-α as the main signaling hub for the differentially regulated proteins. Conclusion: Our findings provide additional insight into understanding bladder cancer pathogenesis. Our data identified potential non-invasive plasma-derived biomarker proteins that merit additional investigation to validate its clinical usefulness to prevent bladder cancer progression.

Cannabis use has been growing recently and it is legally consumed in many countries. Cannabis has a variety of phytochemicals including cannabinoids, which might impair the peripheral systems responses affecting inflammatory and immunological pathways. However, the exact signaling pathways that induce these effects need further understanding. The objective of this study is to investigate the serum proteomic profiling in patients diagnosed with cannabis use disorder (CUD) as compared with healthy control subjects. The novelty of our study is to highlight the differentially changes proteins in the serum of CUD patients. Certain proteins can be targeted in the future to attenuate the toxicological effects of cannabis. Blood samples were collected from 20 male individuals: 10 healthy controls and 10 CUD patients. An untargeted proteomic technique employing two-dimensional difference in gel electrophoresis coupled with mass spectrometry was employed in this study to assess the differentially expressed proteins. The proteomic analysis identified a total of 121 proteins that showed significant changes in protein expression between CUD patients (experimental group) and healthy individuals (control group). For instance, the serum expression of inactive tyrosine protein kinase PEAK1 and tumor necrosis factor alpha-induced protein 3 were increased in CUD group. In contrast, the serum expression of transthyretin and serotransferrin were reduced in CUD group. Among these proteins, 55 proteins were significantly upregulated and 66 proteins significantly downregulated in CUD patients as compared with healthy control group. Ingenuity pathway analysis (IPA) found that these differentially expressed proteins are linked to p38MAPK, interleukin 12 complex, nuclear factor-κB, and other signaling pathways. Our work indicates that the differentially expressed serum proteins between CUD and control groups are correlated to liver X receptor/retinoid X receptor (RXR), farnesoid X receptor/RXR activation, and acute phase response signaling.

Osteoporosis is a common progressive metabolic bone disease resulting in decreased bone mineral density (BMD) and a subsequent increase in fracture risk. The known bone markers are not sensitive and specific enough to reflect the balance in the bone metabolism. Finding a metabolomics-based biomarker specific for bone desorption or lack of bone formation is crucial for predicting bone health earlier. This study aimed to investigate patients’ metabolomic profiles with low BMD (LBMD), including those with osteopenia (ON) and osteoporosis (OP), compared to healthy controls. An untargeted mass spectrometry (MS)-based metabolomics approach was used to analyze serum samples. Results showed a clear separation between patients with LBMD and control (Q2 = 0.986, R2 = 0.994), reflecting a significant difference in the dynamic of metabolic processes between the study groups. A total of 116 putatively identified metabolites were significantly associated with LBMD. Ninety-four metabolites were dysregulated, with 52 up- and 42 downregulated in patients with LBMD compared to controls. Histidine metabolism, aminoacyl-tRNA biosynthesis, glyoxylate, dicarboxylate metabolism, and biosynthesis of unsaturated fatty acids were the most common metabolic pathways dysregulated in LBMD. Furthermore, 35 metabolites were significantly dysregulated between ON and OP groups, with 11 up- and 24 downregulated in ON compared to OP. Among the upregulated metabolites were 3-carboxy-4-methyl-5-propyl-2-2furanopropionic acid (CMPF) and carnitine derivatives (i.e., 3-hydroxy-11-octadecenoylcarnitine, and l-acetylcarnitine), whereas phosphatidylcholine (PC), sphingomyelin (SM), and palmitic acid (PA) were among the downregulated metabolites in ON compared to OP. This study would add a layer to understanding the possible metabolic alterations associated with ON and OP. Additionally, this identified metabolic panel would help develop a prediction model for bone health and OP progression.

Very-long-chain acyl-coenzyme A dehydrogenase (VLCAD) is a coenzyme encoded by ACADVL that converts very-long-chain fatty acids into energy. This process is disrupted by c.65C > A; p.Ser22∗ mutation. To clarify mechanisms by which this mutation leads to VLCAD deficiency, we evaluated differences in molecular and cellular functions between mesenchymal stem cells with normal and mutant VLCAD. Saudi Arabia have a high incidence of this form of mutation. Stem cells with mutant VLCAD were isolated from skin of two patients. Metabolic activity and proliferation were evaluated. The Same evaluation was repeated on normal stem cells introduced with same mutation by CRISPR. Mitochondrial depiction was done by electron microscope and proteomic analysis was done on patients’ cells. Metabolic activity and proliferation were significantly lower in patients’ cells. Introducing the same mutation into normal stem cells resulted in the same defects. We detected mitochondrial abnormalities by electron microscopy in addition to poor wound healing and migration processes in mutant cells. Furthermore, in a proteomic analysis, we identified several upregulated or downregulated proteins related to hypoglycemia, liver disorder, and cardiac and muscle involvement. We concluded experimental assays of mutant ACADVL (c.65C > A; p.Ser22∗) contribute to severe neonatal disorders with hypoglycemia, liver disorder, and cardiac and muscle involvement.

Hyperthyroidism, which is characterized by increased circulating thyroid hormone levels, alters the body’s metabolic and systemic hemodynamic balance and directly influences renal function. In this study, the urinary proteome of patients with hyperthyroidism was characterized using an untargeted proteomic approach with network analysis. Urine samples were collected from nine age-matched patients before and after carbimazole treatment. Differences in the abundance of urinary proteins between hyperthyroid and euthyroid states were determined using a 2D-DIGE coupled to MALDI-TOF mass spectrometry. Alterations in the abundance of urinary proteins, analyzed via Progenesis software, revealed a statistically significant difference in abundance in a total of 40 spots corresponding to 32 proteins, 25 up and 7 down (≥1.5-fold change, ANOVA, p ≤ 0.05). The proteins identified in the study are known to regulate processes associated with cellular metabolism, transport, and acute phase response. The notable upregulated urinary proteins were serotransferrin, transthyretin, serum albumin, ceruloplasmin, alpha-1B-glycoprotein, syntenin-1, and glutaminyl peptide cyclotransferase, whereas the three notable downregulated proteins were plasma kallikrein, protein glutamine gamma-glutamyl transferase, and serpin B3 (SERPINB3). Bioinformatic analysis using ingenuity pathway analysis (IPA) identified the dysregulation of pathways associated with cellular compromise, inflammatory response, cellular assembly, and organization and identified the involvement of the APP and AKT signaling pathways via their interactions with interleukins as the central nodes.

Thyroid hormones critically modulate body homeostasis and haemostasis by regulating energy and metabolism. Previous studies have focused on individual pathways or proteins that are affected by increases in thyroid hormone levels, while an overall plasma proteomic signature of this increased level is lacking. Herein, an integrated untargeted proteomic approach with network analysis was used to identify changes in circulating proteins in the plasma proteome between hyperthyroid and euthyroid states. Plasma from 10 age-matched subjects at baseline (hyperthyroid) and post treatment with carbimazole (euthyroid) was compared by difference gel electrophoresis (DIGE) and matrix-assisted laser desorption/ionization time of flight (MALDI TOF) mass spectrometry (MS). A total of 20 proteins were identified with significant difference in abundance (analysis of variance (ANOVA) test, p ≤ 0.05; fold-change ≥ 1.5) between the two states (12 increased and 8 decreased in abundance in the hyperthyroid state). Twelve protein spots corresponding to ten unique proteins were significantly more abundant in the hyperthyroid state compared with the euthyroid state. These increased proteins were haptoglobin (HP), hemopexin (HPX), clusterin (CLU), apolipoprotein L1 (APOL1), alpha-1-B glycoprotein (A1BG), fibrinogen gamma chain (FGG), Ig alpha-1 chain C region (IGHA1), complement C6 (C6), leucine rich alpha 2 glycoprotein (LRG1), and carboxypeptidase N catalytic chain (CPN1). Eight protein spots corresponding to six unique proteins were significantly decreased in abundance in the hyperthyroid samples compared with euthyroid samples. These decreased proteins were apolipoprotein A1 (APOA1), inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4), plasminogen (PLG), alpha-1 antitrypsin (SERPINA1), fibrinogen beta chain (FGB), and complement C1r subcomponent (C1R). The differentially abundant proteins were investigated by ingenuity pathway analysis (IPA). The network pathway identified related to infectious disease, inflammatory disease, organismal injury and abnormalities, and the connectivity map focused around two central nodes, namely the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and p38 mitogen-activated protein kinase (MAPK) pathways. The plasma proteome of patients with hyperthyroidism revealed differences in the abundance of proteins involved in acute phase response signaling, and development of a hypercoagulable and hypofibrinolytic state. Our findings enhance our existing knowledge of the altered proteins and associated biochemical pathways in hyperthyroidism.

Camel milk is traditionally known to have medicinal properties and many potential health benefits. Natural milk contains many soluble proteins and nanoparticles, such as a milk fat globule membrane (MFGM), a three-layered membrane covering of milk fat globule mainly composed of proteins and lipids, which plays an important role in human health. MFGM proteins account for 1%–4% of total milk proteins, and their nutritive value and distribution depends on the different breeds. The differential composition of these membrane proteins among different camel breeds has not been explored. The current study, therefore, aimed to quantitatively analyze and compare the MFGM proteome between the milk produced by the two most common Saudi camel breeds, Camelus dromedarius: Safra and Wadha. Two-dimensional difference in gel electrophoresis (2D-DIGE) and mass spectrometry analysis revealed a total of 44 MFGM proteins that were identified with a significant difference in abundance (p ≤ 0.05; fold change ≥ 1.5) between the two breeds. Thirty-one proteins were up-regulated and 13 proteins were down-regulated in the Safra breed compared to the Wadha breed. The proteins identified with an increased abundance included α-lactalbumin, lactadherin, and annexin a8, whereas the down-regulated proteins included butyrophilin subfamily 1 member a1, lactotransferrin, and vinculin. The differentially abundant proteins were analyzed by the UNIPROT system and gene ontology (GO) to reveal their associations with known biological functions and pathways. Enzyme-linked immunosorbent assay (ELISA) confirmed the 2D-DIGE findings of butyrophilin (BTN) and α-lactalbumin (α-LA) levels obtained from Safra and Wadha breeds.

The quality of olive oils is sensorially tested by accurate and well established methods. It enables the classification of the pressed oils into the classes of extra virgin oil, virgin oil and lampant oil. Nonetheless, it would be convenient to have analytical methods for screening oils or supporting sensorial analysis using a reliable independent approach based on exploitation of mass spectrometric methodologies. A number of methods have been proposed to evaluate deficiencies of extra virgin olive oils resulting from inappropriate technological treatments, such as high or low temperature deodoration, and home cooking processes. The quality and nutraceutical value of extra virgin olive oil (EVOO) can be related to the antioxidant property of its phenolic compounds. Olive oil is a source of at least 30 phenolic compounds, such as oleuropein, oleocanthal, hydroxytyrosol, and tyrosol, all acting as strong antioxidants, radical scavengers and NSAI-like drugs. We now report the efficacy of MRM tandem mass spectrometry, assisted by the isotope dilution assay, in the evaluation of the thermal stability of selected active principles of extra virgin olive oil.