N-glycosylation of immunoglobulin A in children and adults with type 1 diabetes mellitus.

Aims: To identify N-glycan structures on immunoglobulin A related to type 1 diabetes mellitus among children at the disease onset and adults with type 1 diabetes mellitus. Methods: Human polyclonal IgA N-glycans were profiled using hydrophilic interaction ultra performance liquid chromatography in two cohorts. The first cohort consisted of 62 children at the onset of type 1 diabetes mellitus and 86 of their healthy siblings. The second cohort contained 84 adults with the disease and 84 controls. Associations between N-glycans and type 1 diabetes mellitus were tested using linear mixed model for the paediatric cohort, or general linear model for the adult cohort. False discovery rate was controlled by Benjamini-Hochberg method modified by Li and Ji. Results: In children, an increase in a single oligomannose N-glycan was associated with type 1 diabetes mellitus (B = 0.529, p = 0.0067). N-glycome of the adults displayed increased branching (B = 0.466, p = 0.0052), trigalactosylation (B = 0.466, p = 0.0052), trisialylation (B = 0.629, p < 0.001), and mannosylation (B = 0.604, p < 0.001). The strongest association with the disease was a decrease in immunoglobulin A core fucosylation (B = -0.900, p < 0.001). Conclusions: Changes in immunoglobulin N-glycosylation patterns in type 1 diabetes point to disruptions in immunoglobulin A catabolism and dysregulated inflammatory capabilities of the antibody, potentially impacting immune responses and inflammation.

Alterations in plasma protein N-glycosylation after caloric restriction and bariatric surgery.

BACKGROUND: Protein glycosylation is an enzymatic process known to reflect an individual's physiologic state and changes thereof. The impact of metabolic interventions on plasma protein N-glycosylation has only been sparsely investigated. OBJECTIVE: To examine alterations in plasma protein N-glycosylation following changes in caloric intake and bariatric surgery. SETTING: University of Texas Southwestern Medical Center, US and Oxford University Hospitals, UK. METHODS: This study included 2 independent patient cohorts that recruited 10 and 37 individuals with obesity undergoing a period of caloric restriction followed by bariatric surgery. In both cohorts, clinical data were collated, and the composition of plasma protein N-glycome was analyzed chromatographically. Linear mixed models adjusting for age, sex, and multiple testing (false discovery rate <.05) were used to investigate longitudinal changes in glycosylation features and metabolic clinical markers. RESULTS: A low-calorie diet resulted in a decrease in high-branched trigalactosylated and trisialylated plasma N-glycans and a concomitant increase in low-branched N-glycans in both cohorts. Participants from one cohort additionally underwent a washout period during which caloric intake and body weight increased, resulting in reversal of the initial low-calorie diet-related changes in the plasma N-glycome. Immediate postoperative follow-up revealed the same pattern of N-glycosylation changes in both cohorts-an increase in complex, high-branched, antennary fucosylated, extensively galactosylated and sialylated N-glycans and a substantial decline in simpler, low-branched, core fucosylated, bisected, agalactosylated, and asialylated glycans. A 12-month postoperative monitoring in one cohort showed that N-glycan complexity declines while low branching increases. CONCLUSIONS: Plasma protein N-glycosylation undergoes extensive alterations following caloric restriction and bariatric surgery. These comprehensive changes may reflect the varying inflammatory status of the individual following dietary and surgical interventions and subsequent weight loss.

Thallium Toxicity and its Interference with Potassium Pathways Tested on Various Cell Lines.

Thallium (Tl) is a highly toxic heavy metal whose mechanism of toxicity is still not completely understood. The aim of this study was to test Tl cytotoxicity on several cell lines of different tissue origin in order to clarify specific Tl toxicity to a particular organ. In addition, possible interference of Tl with cell potassium (K) transport was examined. Human keratinocytes (HaCaT), human hepatocellular carcinoma (HepG2), porcine kidney epithelial cells (PK15), human neuroblastoma (SH-SY5Y) and Chinese hamster lung fibroblast cells (V79) were treated with thallium (I) acetate in a wide concentration range (3.9-500 µg/mL) for 24 h, 48 and 72 h. To assess competitive interaction between Tl and K, the cells were treated with four Tl concentrations close to IC50 (15.63, 31.25, 62.50, 125 µg/mL) in combination with/or without potassium (I) acetate (500 µg/mL). The cells' morphology was monitored, and cytotoxic effect was assessed by 3-(4, 5-dimethylthiazole-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) test. The most sensitive to Tl exposure were SH-SY5Y cells, while HepG2 were the most resistant. The combined exposure to thallium (I) acetate and potassium (I) acetate for every cell line, except V79 cells, resulted in higher cell viability compared to thallium (I) acetate alone. The results of our study indicate that cell sensitivity to Tl treatment is largely affected by tissue culture origin, its function, and Na+/K+-ATPase activity.

Effects of epidurally administered dexmedetomidine and dexamethasone on postoperative pain, analgesic requirements, inflammation, and oxidative stress in thoracic surgery.

The aim of this study was to compare the effects of dexmedetomidine and dexamethasone as adjuvants to preoperative epidural administration of local anesthetic (ropivacaine) in thoracic surgery on the postoperative level of pain, use of analgesics, inflammation, and oxidative stress. The study enrolled 42 patients who underwent elective thoracic surgery in a one-year period at the University Hospital Dubrava (Zagreb, Croatia). Based on a computer-generated randomization list the patients were assigned to the dexmedetomidine (n = 18) or dexamethasone (n = 24) group. Postoperatively, patients of dexmedetomidine group reported lower pain (VAS value 1 h post surgery, 3.4 ± 2.7 vs. 5.4 ± 1.8, dexmedetomidine vs. dexamethasone, p < 0.01) and had lower anal-gesic requirements in comparison with dexamethasone group. Thus, dexmedetomidine in comparison with dexamethasone was more efficient in lowering pain and analgesia requirements 24 h after the surgery. On the contrary, dexamethasone had better anti-inflammatory properties (CRP level 24 h post surgery, 131.9 ± 90.7 vs. 26.0 ± 55.2 mg L-1, dexmedetomidine vs. dexamethasone, p < 0.01). Both dexmedetomidine and dexamethasone exhibited antioxidant effects, however, their antioxidant properties should be further explored. The results of this study improve current knowledge of pain control in thoracic surgery.

Predictive metabolites for incident myocardial infarction: a two-step meta-analysis of individual patient data from six cohorts comprising 7897 individuals from the COnsortium of METabolomics Studies.

AIMS: Myocardial infarction (MI) is a major cause of death and disability worldwide. Most metabolomics studies investigating metabolites predicting MI are limited by the participant number and/or the demographic diversity. We sought to identify biomarkers of incident MI in the COnsortium of METabolomics Studies. METHODS AND RESULTS: We included 7897 individuals aged on average 66 years from six intercontinental cohorts with blood metabolomic profiling (n = 1428 metabolites, of which 168 were present in at least three cohorts with over 80% prevalence) and MI information (1373 cases). We performed a two-stage individual patient data meta-analysis. We first assessed the associations between circulating metabolites and incident MI for each cohort adjusting for traditional risk factors and then performed a fixed effect inverse variance meta-analysis to pull the results together. Finally, we conducted a pathway enrichment analysis to identify potential pathways linked to MI. On meta-analysis, 56 metabolites including 21 lipids and 17 amino acids were associated with incident MI after adjusting for multiple testing (false discovery rate < 0.05), and 10 were novel. The largest increased risk was observed for the carbohydrate mannitol/sorbitol {hazard ratio [HR] [95% confidence interval (CI)] = 1.40 [1.26-1.56], P < 0.001}, whereas the largest decrease in risk was found for glutamine [HR (95% CI) = 0.74 (0.67-0.82), P < 0.001]. Moreover, the identified metabolites were significantly enriched (corrected P < 0.05) in pathways previously linked with cardiovascular diseases, including aminoacyl-tRNA biosynthesis. CONCLUSIONS: In the most comprehensive metabolomic study of incident MI to date, 10 novel metabolites were associated with MI. Metabolite profiles might help to identify high-risk individuals before disease onset. Further research is needed to fully understand the mechanisms of action and elaborate pathway findings.

Associations between plasma protein, IgG and IgA N-glycosylation and metabolic health markers in pregnancy and gestational diabetes.

BACKGROUND: Monitoring human circulating N-glycome could provide valuable insight into an individual's metabolic status. Therefore, we examined if aberrant carbohydrate metabolism in gestational diabetes mellitus (GDM) associates with alterations in plasma protein, immunoglobulin G (IgG) and immunoglobulin A (IgA) N-glycosylation. METHODS: Plasma protein, IgG and IgA N-glycans were enzymatically released, purified and chromatographically profiled in 48 pregnant women with normal glucose tolerance and 41 pregnant women with GDM, all sampled at 24-28 weeks of gestation. Linear mixed models adjusting for age and multiple testing (FDR<0.05) were used to investigate the associations between glycosylation features, metabolic markers and GDM status. RESULTS: Fasting insulin exhibited significant associations to numerous glycan traits, including plasma protein galactosylation, sialylation, branching, core fucosylation and bisection, to IgG core fucosylated, bisected (FA2B) and afucosylated disialylated (A2G2S2) glycan and to IgA trisialylated triantennary (A3G3S3) glycan (padj range: 4.37x10-05-4.94x10-02). Insulin resistance markers HOMA2-IR and HOMA2-%B were mostly associated to the same glycan structures as fasting insulin. Both markers showed positive association with high-branched plasma glycans (padj = 1.12x10-02 and 2.03x10-03) and negative association with low-branched plasma glycans (padj = 1.21x10-02 and 2.05x10-03). Additionally, HOMA2-%B index was significantly correlated with glycosylation features describing IgG sialylation. Multiple plasma protein IgG and IgA glycans showed significant associations with total cholesterol and triglyceride levels. None of the tested glycan traits showed a significant difference between GDM and normoglycemic pregnancies. CONCLUSION: Markers of glucose homeostasis and lipid metabolism in pregnancy show extensive associations to various N-glycosylation features. However, plasma protein, IgG and IgA N-glycans were not able to differentiate pregnant women with and without GDM, possibly due to numerous physiological changes accompanying pregnancy, which confound the impact of GDM on protein glycosylation.

Integrated glycomics and genetics analyses reveal a potential role for N-glycosylation of plasma proteins and IgGs, as well as the complement system, in the development of type 1 diabetes.

AIMS/HYPOTHESIS: We previously demonstrated that N-glycosylation of plasma proteins and IgGs is different in children with recent-onset type 1 diabetes compared with their healthy siblings. To search for genetic variants contributing to these changes, we undertook a genetic association study of the plasma protein and IgG N-glycome in type 1 diabetes. METHODS: A total of 1105 recent-onset type 1 diabetes patients from the Danish Registry of Childhood and Adolescent Diabetes were genotyped at 183,546 genetic markers, testing these for genetic association with variable levels of 24 IgG and 39 plasma protein N-glycan traits. In the follow-up study, significant associations were validated in 455 samples. RESULTS: This study confirmed previously known plasma protein and/or IgG N-glycosylation loci (candidate genes MGAT3, MGAT5 and ST6GAL1, encoding beta-1,4-mannosyl-glycoprotein 4-beta-N-acetylglucosaminyltransferase, alpha-1,6-mannosylglycoprotein 6-beta-N-acetylglucosaminyltransferase and ST6 beta-galactoside alpha-2,6-sialyltransferase 1 gene, respectively) and identified novel associations that were not previously reported for the general European population. First, novel genetic associations of IgG-bound glycans were found with SNPs on chromosome 22 residing in two genomic intervals close to candidate gene MGAT3; these include core fucosylated digalactosylated disialylated IgG N-glycan with bisecting N-acetylglucosamine (GlcNAc) (pdiscovery=7.65 × 10-12, preplication=8.33 × 10-6 for the top associated SNP rs5757680) and core fucosylated digalactosylated glycan with bisecting GlcNAc (pdiscovery=2.88 × 10-10, preplication=3.03 × 10-3 for the top associated SNP rs137702). The most significant genetic associations of IgG-bound glycans were those with MGAT3. Second, two SNPs in high linkage disequilibrium (missense rs1047286 and synonymous rs2230203) located on chromosome 19 within the protein coding region of the complement C3 gene (C3) showed association with the oligomannose plasma protein N-glycan (pdiscovery=2.43 × 10-11, preplication=8.66 × 10-4 for the top associated SNP rs1047286). CONCLUSIONS/INTERPRETATION: This study identified novel genetic associations driving the distinct N-glycosylation of plasma proteins and IgGs identified previously at type 1 diabetes onset. Our results highlight the importance of further exploring the potential role of N-glycosylation and its influence on complement activation and type 1 diabetes susceptibility.

Pleomorphic Variants of Borreliella (syn. Borrelia) burgdorferi Express Evolutionary Distinct Transcriptomes.

Borreliella (syn. Borrelia) burgdorferi is a spirochete bacterium that causes tick-borne Lyme disease. Along its lifecycle B. burgdorferi develops several pleomorphic forms with unclear biological and medical relevance. Surprisingly, these morphotypes have never been compared at the global transcriptome level. To fill this void, we grew B. burgdorferi spirochete, round body, bleb, and biofilm-dominated cultures and recovered their transcriptomes by RNAseq profiling. We found that round bodies share similar expression profiles with spirochetes, despite their morphological differences. This sharply contrasts to blebs and biofilms that showed unique transcriptomes, profoundly distinct from spirochetes and round bodies. To better characterize differentially expressed genes in non-spirochete morphotypes, we performed functional, positional, and evolutionary enrichment analyses. Our results suggest that spirochete to round body transition relies on the delicate regulation of a relatively small number of highly conserved genes, which are located on the main chromosome and involved in translation. In contrast, spirochete to bleb or biofilm transition includes substantial reshaping of transcription profiles towards plasmids-residing and evolutionary young genes, which originated in the ancestor of Borreliaceae. Despite their abundance the function of these Borreliaceae-specific genes is largely unknown. However, many known Lyme disease virulence genes implicated in immune evasion and tissue adhesion originated in this evolutionary period. Taken together, these regularities point to the possibility that bleb and biofilm morphotypes might be important in the dissemination and persistence of B. burgdorferi inside the mammalian host. On the other hand, they prioritize the large pool of unstudied Borreliaceae-specific genes for functional characterization because this subset likely contains undiscovered Lyme disease pathogenesis genes.

Semiquantitative classification (SQC) and Oxford classifications predict poor renal outcome better than The International Study of Kidney Disease in Children (ISKDC) and Haas in patients with IgAV nephritis: a multicenter study.

INTRODUCTION: Several histologic classifications are used in the evaluation of IgA vasculitis nephritis (IgAVN), however, to date, no studies have determined which one has the strongest association with the severity of IgAVN and, as a consequence, its outcomes. MATERIALS AND METHODS: Patients included in the study were diagnosed with IgAV and IgAVN in seven tertiary university medical centers in Croatia, Italy and Israel. The International Study of Kidney Disease in Children (ISKDC), Haas, Oxford, and Semiquantitative classification (SQC) classifications were used in the analysis and description of renal biopsy. Time from biopsy to outcome evaluation was a statistically significant factor in outcome prediction that was used to define the base model, and was a covariate in all the tested models. RESULTS: Sixty-seven patients were included in this study. The SQC classification proved to be the best one in outcome prediction, followed by the Oxford classification. The ISKDC and Haas classifications could not predict renal outcome. The Oxford parameters for mesangial hypercellularity and tubular atrophy, as well as the SQC parameters for cellular crescents showed an independent statistically significant contribution to outcome prediction. High level of twenty-four hour protein excretion was associated with a higher grade in the Oxford, SQC and ISKDC classifications. Endocapillary proliferation was positively associated with the Pediatric Vasculitis Activity Score (PVAS) at diagnosis, while tubular atrophy was negatively associated. CONCLUSION: The SQC, followed by the Oxford classification were found to provide the best classifications of renal biopsy analysis in patients to predict the outcome in patients with IgAVN. Cellular crescents, mesangial hypercellularity and tubular atrophy showed significant contributions, indicating that active and chronic variables should be included in the estimation.

High-Throughput Human Complement C3 N-Glycoprofiling Identifies Markers of Early Onset Type 1 Diabetes Mellitus in Children.

Recently, it was shown that children at the onset of type 1 diabetes (T1D) have a higher proportion of oligomannose glycans in their total plasma protein N-glycome compared to their healthy siblings. The most abundant complement component, glycoprotein C3, contains two N-glycosylation sites occupied exclusively by this type of glycans. Furthermore, complement system, as well as C3, was previously associated with T1D. It is also known that changes in glycosylation can modulate inflammatory responses, so our aim was to characterize the glycosylation profile of C3 in T1D. For this purpose, we developed a novel high-throughput workflow for human C3 concanavalin A lectin affinity enrichment and subsequent LC-MS glycopeptide analysis which enables protein-specific N-glycosylation profiling. From the Danish Childhood Diabetes Register, plasma samples of 61 children/adolescents newly diagnosed with T1D and 84 of their unaffected siblings were C3 N-glycoprofiled. Significant changes of C3 N-glycan profiles were found. T1D was associated with an increase in the proportion of unprocessed glycan structures with more mannose units. A regression model including C3 N-glycans showed notable discriminative power between children with early onset T1D and their healthy siblings with area under curve of 0.879. This study confirmed our previous findings of plasma high-mannose glycan changes in a cohort of recent onset T1D cases, suggesting the involvement of C3 N-glycome in T1D development. Our C3 glycan-based discriminative model could be valuable in assessment of T1D risk in children.

Cross-Sectional Blood Metabolite Markers of Hypertension: A Multicohort Analysis of 44,306 Individuals from the COnsortium of METabolomics Studies.

Hypertension is the main modifiable risk factor for cardiovascular morbidity and mortality but discovering molecular mechanisms for targeted treatment has been challenging. Here we investigate associations of blood metabolite markers with hypertension by integrating data from nine intercontinental cohorts from the COnsortium of METabolomics Studies. We included 44,306 individuals with circulating metabolites (up to 813). Metabolites were aligned and inverse normalised to allow intra-platform comparison. Logistic models adjusting for covariates were performed in each cohort and results were combined using random-effect inverse-variance meta-analyses adjusting for multiple testing. We further conducted canonical pathway analysis to investigate the pathways underlying the hypertension-associated metabolites. In 12,479 hypertensive cases and 31,827 controls without renal impairment, we identified 38 metabolites, associated with hypertension after adjusting for age, sex, body mass index, ethnicity, and multiple testing. Of these, 32 metabolite associations, predominantly lipid (steroids and fatty acyls) and organic acids (amino-, hydroxy-, and keto-acids) remained after further adjusting for comorbidities and dietary intake. Among the identified metabolites, 5 were novel, including 2 bile acids, 2 glycerophospholipids, and ketoleucine. Pathway analysis further implicates the role of the amino-acids, serine/glycine, and bile acids in hypertension regulation. In the largest cross-sectional hypertension-metabolomics study to date, we identify 32 circulating metabolites (of which 5 novel and 27 confirmed) that are potentially actionable targets for intervention. Further in-vivo studies are needed to identify their specific role in the aetiology or progression of hypertension.

Children at onset of type 1 diabetes show altered N-glycosylation of plasma proteins and IgG.

AIMS/HYPOTHESIS: Individual variation in plasma N-glycosylation has mainly been studied in the context of diabetes complications, and its role in type 1 diabetes onset is largely unknown. Our aims were to undertake a detailed characterisation of the plasma and IgG N-glycomes in patients with recent onset type 1 diabetes, and to evaluate their discriminative potential in risk assessment. METHODS: In the first part of the study, plasma and IgG N-glycans were chromatographically analysed in a study population from the DanDiabKids registry, comprising 1917 children and adolescents (0.6-19.1 years) who were newly diagnosed with type 1 diabetes. A follow-up study compared the results for 188 of these participants with those for their 244 unaffected siblings. Correlation of N-glycan abundance with the levels and number of various autoantibodies (against IA-2, GAD, ZnT8R, ZnT8W), as well as with sex and age at diagnosis, were estimated by using general linear modelling. A disease predictive model was built using logistic mixed-model elastic net regression, and evaluated using a 10-fold cross-validation. RESULTS: Our study showed that onset of type 1 diabetes was associated with an increase in the proportion of plasma and IgG high-mannose and bisecting GlcNAc structures, a decrease in monogalactosylation, and an increase in IgG disialylation. ZnT8R autoantibody levels were associated with higher IgG digalactosylated glycan with bisecting GlcNAc. Finally, an increase in the number of autoantibodies (which is a better predictor of progression to overt diabetes than the level of any individual antibody) was accompanied by a decrease in the proportions of some of the highly branched plasma N-glycans. Models including age, sex and N-glycans yielded notable discriminative power between children with type 1 diabetes and their healthy siblings, with AUCs of 0.915 and 0.869 for addition of plasma and IgG N-glycans, respectively. CONCLUSIONS/INTERPRETATION: We defined N-glycan changes accompanying onset of type 1 diabetes, and developed a predictive model based on N-glycan profiles that could have valuable potential in risk assessment. Increasing the power of tests to identify individuals at risk of disease development would be a considerable asset for type 1 diabetes prevention trials.

Changes in Specific Biomarkers Indicate Cardiac Adaptive and Anti-inflammatory Response of Repeated Recreational SCUBA Diving.

Objective: Recreational SCUBA (rSCUBA) diving has become a highly popular and widespread sport. Yet, information on molecular events underlying (patho)physiological events that follow exposure to the specific environmental conditions (hyperbaric conditions, coldness, immersion, and elevated breathing pressure), in which rSCUBA diving is performed, remain largely unknown. Our previous study suggested that repeated rSCUBA diving triggers an adaptive response of cardiovascular and immune system. To elucidate further molecular events underlying cardiac and immune system adaptation and to exclude possible adverse effects we measured blood levels of specific cardiac and inflammation markers. Methods: This longitudinal intervention study included fourteen recreational divers who performed five dives, one per week, on the depth 20-30 m that lasted 30 min, after the non-dive period of 5 months. Blood samples were taken immediately before and after the first, third, and fifth dives. Copeptin, immunoglobulins A, G and M, complement components C3 and C4, and differential blood count parameters, including neutrophil-to-lymphocyte ratio (NLR) were determined using standard laboratory methods. Cell-free DNA was measured by qPCR analysis and N-glycans released from IgG and total plasma proteins (TPP), were analyzed by hydrophilic interaction ultra-performance liquid chromatography. Results: Copeptin level increased after the first dive but decreased after the third and fifth dive. Increases in immunoglobulins level after every dive and during whole studied period were observed, but no changes in C3, C4, and cfDNA level were detected. NLR increased only after the first dive. IgG and TPP N-glycosylation alterations toward anti-inflammatory status over whole studied period were manifested as an increase in monogalyctosylated and core-fucosylated IgG N-glycans and decrease in agalactosylated TPP N-glycans. Conclusion: rSCUBA diving practiced on a regular basis promotes anti-inflammatory status thus contributing cardioprotection and conferring multiple health benefits.

2019 ACR/EULAR classification criteria and therapy in predicting organ damage accrual in patients with childhood-onset systemic lupus erythematosus: A retrospective study over the last 29 years.

OBJECTIVE: We explored damage occurrence in patients with childhood-onset SLE (cSLE) and aimed to predict the risk of organ damage occurrence in time. METHODS: The retrospective study included patients treated for cSLE at the Centre of Reference for Pediatric and Adolescent Rheumatology of the Republic Croatia over a 29-year period. RESULTS: The disease development of 97 patients (77 females) with cSLE was examined. The median (Q1, Q3) follow-up time was 6.5 (2.3, 12.0) years. SDI was determined at 5 time points (6, 12, 24, 36 months, and last follow-up). Thirty-eight patients (48%) had organ damage at the last follow-up. Prepubertal group of patients showed higher SLEDAI scores at the disease onset, while post-pubertal group had significantly lower proportion of patients with relapses. We estimated the time from the first symptom to the moment of damage and our findings suggest that it is unlikely that organ damage will occur in 50% of patients in the first 6 years since the diagnosis. The number of 2019 ACR/EULAR classification criteria at the time of diagnosis associated with SDI determined after 1 year of the follow-up period. The patients who received higher doses of glucocorticoids accumulated damage faster and mycophenolate mofetil was found to be a more frequent therapy in patients with SDI ≥3. CONCLUSION: Knowing that damage will most likely happen after the first 6 years after diagnosis in 50% of patients enables physicians to better predict damage occurrence. High number of 2019 ACR/EULAR criteria and treatment with glucocorticoids in childhood-onset SLE are associated with damage accrual and these findings could enable us to detect patients which should be closely monitored for higher risk of damage development.

Immunoglobulin G glycome composition in transition from premenopause to postmenopause.

Gonadal hormones affect immunoglobulin G (IgG) glycosylation, and the more proinflammatory IgG glycome composition might be one of the molecular mechanisms behind the increased proinflammatory phenotype in perimenopause. Using ultra-high-performance liquid chromatography, we analyzed IgG glycome composition in 5,080 samples from 1940 pre-, peri-, and postmenopausal women. Statistically significant decrease in galactosylation and sialylation was observed in postmenopausal women. Furthermore, during the transition from pre- to postmenopausal period, the rate of increase in agalactosylated structures (0.051/yr; 95%CI = 0.043-0.059, p < 0.001) and decrease in digalactosylated (-0.043/yr; 95%CI = -0.050 to -0.037, p < 0.001) and monosialylated glycans (-0.029/yr; 95%CI = -0.034 to -0.024, p < 0.001) were significantly higher than in either pre- or postmenopausal periods. The conversion to the more proinflammatory IgG glycome and the resulting decrease in the ability of IgG to suppress low-grade chronic inflammation may be an important molecular mechanism mediating the increased health risk in perimenopause and postmenopause.

Plasma N-glycome shows continuous deterioration as the diagnosis of insulin resistance approaches.

INTRODUCTION: Prediction of type 2 diabetes mellitus (T2DM) and its preceding factors, such as insulin resistance (IR), is of great importance as it may allow delay or prevention of onset of the disease. Plasma protein N-glycome has emerged as a promising predictive biomarker. In a prospective longitudinal study, we included patients with a first diagnosis of impaired glucose metabolism (IR or T2DM) to investigate the N-glycosylation's predictive value years before diabetes development. RESEARCH DESIGN AND METHODS: Plasma protein N-glycome was profiled by hydrophilic interaction ultra-performance liquid chromatography in 534 TwinsUK participants free from disease at baseline. This included 89 participants with incident diagnosis of IR or T2DM during the follow-up period (7.14±3.04 years) whose last sample prior to diagnosis was compared using general linear regression with 445 age-matched unrelated controls. Findings were replicated in an independent cohort. Changes in N-glycome have also been presented in connection with time to diagnosis. RESULTS: Eight groups of plasma N-glycans were different between incident IR or T2DM cases and controls (p<0.05) after adjusting for multiple testing using Benjamini-Hochberg correction. These differences were noticeable up to 10 years prior to diagnosis and are changing continuously as becoming more expressed toward the diagnosis. The prediction model was built using significant glycan traits, displaying a discriminative performance with an area under the receiver operating characteristic curve of 0.77. CONCLUSIONS: In addition to previous studies, we showed the diagnostic potential of plasma N-glycome in the prediction of both IR and T2DM development years before the clinical manifestation and indicated the continuous deterioration of N-glycome toward the diagnosis.

N-glycosylation of immunoglobulin G predicts incident hypertension.

OBJECTIVES: Glycosylation of immunoglobulin G (IgG) is an important regulator of the immune system and has been implicated in prevalent hypertension. The aim of this study is to investigate whether the IgG glycome begins to change prior to hypertension diagnosis by analysing the IgG glycome composition in a large population-based female cohort with two independent replication samples. METHODS: We included 989 unrelated cases with incident hypertension and 1628 controls from the TwinsUK cohort (mean follow-up time of 6.3 years) with IgG measured at baseline by ultra-performance liquid chromatography and longitudinal BP measurement available. We replicated our findings in 106 individuals from the 10 001 Dalmatians and 729 from KORA S4. Cox regression mixed models were applied to identify changes in glycan traits preincident hypertension, after adjusting for age, mean arterial pressure, BMI, family relatedness and multiple testing (FDR < 0.1). Significant IgG-incident hypertension associations were replicated in the two independent cohorts by leveraging Cox regression mixed models in the 10 001 Dalmatians and logistic regression models in the KORA cohort. RESULTS: We identified and replicated four glycan traits, incidence of bisecting GlcNAc, GP4, GP9 and GP21, that are predictive of incident hypertension after adjusting for confoundes and multiple testing [hazard ratio (95% CI) ranging from 0.45 (0.24-0.84) for GP21 to 2.9 (1.5-5.68) for GP4]. We then linearly combined the four replicated glycans and found that the glycan score correlated with incident hypertension, SBP and DBP. CONCLUSION: Our results suggest that the IgG glycome changes prior to the development of hypertension.

Sterigmatocystin, 5-Methoxysterigmatocistin, and Their Combinations Are Cytotoxic and Genotoxic to A549 and Hepg2 Cells and Provoke Phosphorylation of Chk2, but Not Fancd2 Checkpoint Proteins.

Sterigmatocystin (STC) and 5-methoxysterigmatocystin (5-M-STC) are structurally related mycotoxins with cytotoxic and genotoxic properties. In the present study, we hypothesized that DNA damage induced by non-cytotoxic concentrations of single and combined mycotoxins could alter the phosphorylation of the checkpoint proteins Chk2 and FANCD2 (ELISA) in HepG2 and A549 cells. The cytotoxic potential (MTT test) of single and combined STC and 5-M-STC, the nature of their interaction (additivity, antagonism, or synergy) and DNA damage level (alkaline comet assay) in HepG2 and A549 cells were also investigated. All experiments were performed after 24 h of mycotoxin treatment. 5-M-STC was 10-folds more cytotoxic than STC to both HepG2 and A549 cells. Both mycotoxins are genotoxic to HepG2 and A549 cells by inducing both double and single DNA strand breaks that activate Chk2 (especially in HepG2 cells) but not the FANCD2 protein. STC exerted higher genotoxic potential than 5-M-STC in HepG2 and A549 cells when both toxins were applied individually at the same concentration. Dual combinations of non-cytotoxic mycotoxin concentrations showed additive to antagonizing cytotoxic and genotoxic effects. The absence and low activation of checkpoint proteins during prolonged exposure to non-cytotoxic concentrations of STC and 5-M-STC could support cell proliferation and carcinogenesis.

Patients perspectives on drug shortages in six European hospital settings - a cross sectional study.

BACKGROUND: It is known that drug shortages represent a major challenge for all stakeholders involved in the process, but there is little evidence regarding insights into patients' awareness and perspectives. This study aimed to investigate the patients-perceived drug shortages experience and their view on outcomes in different European hospital settings. Furthermore, we wanted to explore information preferences on drug shortages. METHODS: A retrospective, cross sectional, a mixed method study was conducted in six European hospital settings. One hospital (H) from each of this country agreed to participate: Bosnia and Herzegovina (H-BiH), Croatia (H-CR), Germany (H-GE), Greece (H-GR), Serbia (H-SE) and Poland (H-PO). Recruitment and data collection was conducted over 27 months from November 2017 until January 2020. Overall, we surveyed 607 patients which completed paper-based questionnaire. Questions related to: general information (demographic data), basic knowledge on drug shortages, drug shortages experienced during hospitalization and information preferences on drug shortage. Differences between hospital settings were analyzed using Chi-squared test or Fisher's exact test. For more complex contingency tables, Monte Carlo simulations (N = 2000) were applied for Fisher's test. Post-hoc hospital-wise analyses were performed using Fisher's exact tests. False discovery rate was controlled using the Bonferroni method. Analyses were performed using R: a language and environment for statistical computing (v 3.6.3). RESULTS: 6 % of patients reported experiences with drug shortages while hospitalized which led to a deterioration of their health. The majority of affected patients were hospitalized at hematology and/or oncology wards in H-BiH, H-PO and H-GE. H-BiH had the highest number of affected patients (18.1 %, N = 19/105, p < 0.001) while the fewest patients were in H-SE (1 %, N = 1/100, p = 0.001). In addition, 82.5 %, (N = 501/607) of respondents wanted to be informed of alternative treatment options if there was a drug shortage without a generic substitute available. Majority of these patients (66.4 %, N = 386/501) prefer to be informed by a healthcare professional. CONCLUSIONS: Although drug shortages led to serious medical consequences, our findings show that most of the patients did not perceive shortages as a problem. One possible interpretation is that good hospital management practices by healthcare professionals helped to mitigate the perceived impact of shortages. Our study highlights the importance of a good communication especially between patients and healthcare professionals in whom our patients have the greatest trust.