Glycosylation and Characterization of Human Transferrin in an End-Stage Kidney Disease.

Chronic kidney disease (CKD) is a global health concern affecting approximately one billion individuals worldwide. End-stage kidney disease (ESKD), the most severe form of CKD, is often accompanied by anemia. Peritoneal dialysis (PD), a common treatment for ESKD, utilizes the peritoneum for solute transfer but is associated with complications including protein loss, including transferrin (Tf) a key protein involved in iron transport. This study investigated Tf characteristics in ESKD patients compared to healthy individuals using lectin microarray, spectroscopic techniques and immunocytochemical analysis to assess Tf interaction with transferrin receptors (TfRs). ESKD patients exhibited altered Tf glycosylation patterns, evidenced by significant changes in lectin reactivity compared to healthy controls. However, structural analyses revealed no significant differences in the Tf secondary or tertiary structures between the two groups. A functional analysis demonstrated comparable Tf-TfR interaction in both PD and healthy samples. Despite significant alterations in Tf glycosylation, structural integrity and Tf-TfR interaction remained preserved in PD patients. These findings suggest that while glycosylation changes may influence iron metabolism, they do not impair Tf function. The study highlights the importance of a glucose-free dialysis solutions in managing anemia exacerbation in PD patients with poorly controlled anemia, potentially offering a targeted therapeutic approach to improve patient outcomes.

Association between nitric oxide synthase (NOS3) gene polymorphisms and diabetic nephropathy: An updated meta-analysis.

Polymorphisms located within NOS3 gene have been investigated as susceptibility variants for diabetic nephropathy (DN) in type 2 diabetes mellitus (T2DM) in a large number of studies. However, these previous articles yielded inconsistent results and we aimed at elucidating the impact of NOS3 variants on DN risk in T2DM by conducting an updated systematic data synthesis. A total of 36 studies (12,807 participants) were selected for qualitative data synthesis, while 33 records with 11,649 subjects were included in the meta-analysis. The pooled analysis demonstrated the association of minor alleles of rs2070744 and rs1799983 with an increased susceptibility to DN (P < 0.001 and P = 0.015 for allelic model, respectively). For both of these variants, a significant effect of subgrouping according to ethnicity was found. Rs869109213 displayed an association with DN susceptibility, with pooled effect measures indicating a predisposing effect of the minor allele a (Prec = 0.002, ORrec = 1.960, 95%CI 1.288-2.983; Paavs. bb = 0.001, ORaavs. bb = 2.014, 95%CI 1.316-3.083). These findings support the effects of NOS3 variants on the risk of developing DN in T2DM.

Exosomal Prostate-Specific Membrane Antigen (PSMA) and Caveolin-1 as Potential Biomarkers of Prostate Cancer-Evidence from Serbian Population.

Prostate-specific membrane antigen (PSMA) and caveolin-1 are membrane proteins that are overexpressed in prostate cancer (PCa) and are involved in tumor growth and increase in aggressiveness. The aim of the present study is therefore to evaluate PSMA and caveolin-1 proteins from plasma exosomes as effective liquid biopsy biomarkers for PCa. This study included 39 patients with PCa and 33 with benign prostatic hyperplasia (BPH). The shape and size of the exosomes were confirmed by transmission electron microscopy (TEM) and scanning electron microscopy (SEM) analysis. Immunogold analysis showed that PSMA is localized to the membrane of exosomes isolated from the plasma of both groups of participants. The relative protein levels of PSMA and caveolin-1 in the plasma exosomes of PCa and BPH patients were determined by Western blot analysis. The relative level of the analyzed plasma exosomal proteins was compared between PCa and BPH patients and the relevance of the exosomal PSMA and caveoin-1 level to the clinicopathological parameters in PCa was investigated. The analysis performed showed an enrichment of exosomal PSMA in the plasma of PCa patients compared to the exosomes of men with BPH. The level of exosomal caveolin-1 in plasma was significantly higher in PCa patients with high PSA levels, clinical-stage T3 or T4 and in the group of PCa patients with aggressive PCa compared to favorable clinicopathological features or tumor aggressiveness. Plasma exosomes may serve as a suitable object for the identification of potential biomarkers for the early diagnosis and prognosis of PCa as well as carriers of therapeutic agents in precision medicine of PCa treatment.

Serum Glycome as a Diagnostic and Prognostic Factor in Gestational Diabetes Mellitus.

Gestational diabetes mellitus (GDM) is a risk factor for both mother and fetus/neonate during and after the pregnancy. Inconsistent protocols and cumbersome screening procedures warrant the search for new and easily accessible biomarkers. We investigated a potential of serum N-glycome to differentiate between healthy pregnant women (n = 49) and women with GDM (n = 53) using a lectin-based microarray and studied the correlation between the obtained data and parameters of glucose and lipid metabolism. Four out of 15 lectins used were able to detect the differences between the control and GDM groups in fucosylation, terminal galactose/N-acetylglucosamine (Gal/GlcNAc), presence of Galα1,4Galß1,4Glc (Gb3 antigen), and terminal α2,3-sialylation with AUC values above 60%. An increase in the Gb3 antigen and α2,3-sialylation correlated positively with GDM, whereas the amount of fucosylated glycans correlated negatively with the content of terminal Gal/GlcNAc. The content of GlcNAc oligomers correlated with the highest number of blood analytes, indices, and demographic characteristics, but failed to discriminate between the groups. The presence of terminal Gal residues correlated positively with the glucose levels and negatively with the LDL levels in the non-GDM group only. The results suggest fucosylation, terminal galactosylation, and the presence of Gb3 antigen as prediction markers of GDM.

Significance of 1,25-Dihydroxyvitamin D3 on Overall Mortality in Peritoneal Dialysis Patients with COVID-19.

In previous publications, we pointed out the importance of mannosylation of fibrinogen for the development of cardiovascular complications and fucosylation as a predictor of peritoneal membrane dysfunction in patients on peritoneal dialysis (PD). After a follow-up period of 30 months from the onset of the COVID-19 pandemic, we evaluated the significance of 1,25-dihydroxyvitamin D3 (calcitriol) therapy, primary disease, biochemical and hematologic analyzes, and previously performed glycan analysis by lectin-based microarray as predictors of mortality in this patient group. After univariate Cox regression analysis, diabetes mellitus (DM) and calcitriol therapy were found to be potential predictors of mortality. Additional multivariate Cox regression analysis confirmed that only DM was a predictor of mortality. Nevertheless, the use of calcitriol in therapy significantly reduced mortality in this patient group, as shown by the Kaplan-Meier survival curve. The presence of DM as a concomitant disease proved to be a strong predictor of fatal outcome in PD patients infected with SARS-CoV-2. This is the first study to indicate the importance and beneficial effect of calcitriol therapy on survival in PD patients with COVID-19 infection. In addition, this study points to the possibility that adverse thrombogenic events observed in PD patients during the pandemic may be caused by aberrant fibrinogen glycosylation.

The association of human leucocyte antigen (HLA) alleles with COVID-19 severity: A systematic review and meta-analysis.

Due to their pivotal role in orchestrating the immune response, HLA loci were recognized as candidates for genetic association studies related to the severity of COVID-19. Since the findings on the effects of HLA alleles on the outcome of SARS-CoV-2 infection remain inconclusive, we aimed to elucidate the potential involvement of genetic variability within HLA loci in the molecular genetics of COVID-19 by classifying the articles according to different disease severity/outcomes and by conducting a systematic review with meta-analysis. Potentially eligible studies were identified by searching PubMed, Scopus and Web of Science literature databases. A total of 28 studies with 13,073 participants were included in qualitative synthesis, while the results of 19 studies with 10,551 SARS-CoV-2-positive participants were pooled in the meta-analysis. According to the results of quantitative data synthesis, association with COVID-19 severity or with the lethal outcome was determined for the following alleles and allele families: HLA-A*01, HLA-A*03, HLA-A*11, HLA-A*23, HLA-A*31, HLA-A*68, HLA-A*68:02, HLA-B*07:02, HLA-B*14, HLA-B*15, HLA-B*40:02, HLA-B*51:01, HLA-B*53, HLA-B*54, HLA-B*54:01, HLA-C*04, HLA-C*04:01, HLA-C*06, HLA-C*07:02, HLA-DRB1*11, HLA-DRB1*15, HLA-DQB1*03 and HLA-DQB1*06 (assuming either allelic or dominant genetic model). We conclude that alleles of HLA-A, -B, -C, -DRB1 and -DQB1 loci may represent potential biomarkers of COVID-19 severity and/or mortality, which needs to be confirmed in a larger set of studies.

MiR-203a-3p, miR-204-3p, miR-222-3p as useful diagnostic and prognostic tool for thyroid neoplasia spectrum.

PURPOSE: The challenge in the diagnosis and treatment of thyroid carcinoma is to correctly classify neoplasias with overlapping features and to identify the high-risk patients among those with a less aggressive form, in order to personalize the treatment of thyroid carcinoma patients accordingly. METHODS: MiR-203a-3p, miR-204-3p, and miR-222-3p levels were determined in 99 cases of thyroid neoplasias (77 papillary thyroid carcinomas (PTC) of diverse variants, 12 follicular thyroid adenomas (FTA) and 10 nodular goiters (NG)) along with 99 adjacent non-malignant thyroid tissues using quantitative RT-PCR. The results were evaluated in comparison with the clinicopathological features of the patients and available TCGA data. RESULTS: Down-regulated miR-203a-3p indicates the presence of thyroid tumor (PTC or FTA) with high sensitivity (75%) and specificity (73%), while its up-regulation indicates NG. If miR-203a-3p is down-regulated, up-regulated miR-204-3p with high sensitivity (83.3%) and specificity (74.4%) indicates FTA presence, while up-regulated miR-222-3p, with high sensitivity (76.6%) and specificity (75.0%), points to PTC. The expression of miR-204-3p and miR-222-3p depends on the PTC subtype (P < 0.05). While the deregulated expression of tested miRs is associated with a long-range of unfavorable clinicopathological parameters of PTC, only abundant expression of miR-222-3p may be used as an independent predictive factor for the presence of extrathyroid invasion and advanced pTNM stage of PTC (P < 0.05). CONCLUSION: Successive evaluation of miR-203a-3p, miR-204-3p, and miR-222-3p expression can help in the differential diagnosis of thyroid neoplasias. A high relative value of miR-222-3p expression is an independent predictive factor for the presence of extrathyroid invasion and advanced pTNM stage of PTC. The panel consisting of miR-203a-3p, miR-204-3p, and miR-222-3p could be used as a diagnostic and prognostic tool for personalizing the treatment of thyroid cancer patients.

MiR-375 and miR-21 as Potential Biomarkers of Prostate Cancer: Comparison of Matching Samples of Plasma and Exosomes.

MiR-21 and miR-375 have been reported as dysregulated in prostate cancer (PCa) in multiple previous studies. Still, variable or even opposing data for the expression of these microRNAs in PCa were found, and their potential biomarker properties remain elusive. In an attempt to clarify their significance as PCa biomarkers, as well as to compare different types of specimens as a source of relevant microRNAs, we used plasma and matching plasma-derived exosomes from patients with PCa and patients with benign prostatic hyperplasia (BPH). Plasma and exosomes were obtained from 34 patients with PCa and 34 patients with BPH, and their levels of expression of miR-21 and miR-375 were determined by RT-qPCR. We found no significant difference in the level of expression of these microRNAs in plasma and exosomes between patients with PCa and BPH. The level of exosomal miR-21 was elevated in PCa patients with high serum PSA values, as well as in patients with aggressive PCa, while for plasma samples, the results remained insignificant. For miR-375, we did not find an association with the values of standard prognostic parameters of PCa, nor with cancer aggressiveness. Therefore, our results support the potential prognostic role of exosomal miR-21 expression levels in PCa.

The association of ACE1, ACE2, TMPRSS2, IFITM3 and VDR polymorphisms with COVID-19 severity: A systematic review and meta-analysis.

Genes involved in the regulation of viral recognition and its entry into a host cell have been identified as candidates for genetic association studies on COVID-19 severity. Published findings on the effects of polymorphisms within ACE1, ACE2, TMPRSS2, IFITM3 and VDR genes remained inconclusive, so we conducted a systematic review and meta-analysis in order to elucidate their potential involvement in the genetic basis underlying the severity of COVID-19 and/or an outcome of SARS-CoV-2 infection. Identification of potentially eligible studies was based on PubMed, Scopus and Web of Science database search. Relevant studies (n=29) with a total number of 8247 SARS-CoV-2-positive participants were included in qualitative synthesis, while results of 21 studies involving 5939 were pooled in meta-analysis. Minor allele I of rs1799752 located within ACE1 was identified as a protective variant against severe COVID-19, while its effect on mortality rate was opposite. Similarly, minor allele A of ACE2 polymorphism, rs2285666, was found to associate with a decreased risk of severe COVID-19 (P = 0.003, OR = 0.512, 95 % CI = 0.331-0.793). Statistical significance was also seen for the association between COVID-19 severity and rs12329760 located within TMPRSS2. Our results did not support the supposed association of rs12252 in IFITM3 and polymorphisms within VDR with disease severity. We conclude that genetic variants within ACE1, ACE2 and TMPRSS2 may be potential biomarkers of COVID-19 severity, which needs to be further confirmed in a larger set of studies.

The Insulin-like Growth Factor System and Colorectal Cancer.

Insulin-like growth factors (IGFs) are peptides which exert mitogenic, endocrine and cytokine activities. Together with their receptors, binding proteins and associated molecules, they participate in numerous pathophysiological processes, including cancer development. Colorectal cancer (CRC) is a disease with high incidence and mortality rates worldwide, whose etiology usually represents a combination of the environmental and genetic factors. IGFs are most often increased in CRC, enabling excessive autocrine/paracrine stimulation of the cell growth. Overexpression or increased activation/accessibility of IGF receptors is a coinciding step which transmits IGF-related signals. A number of molecules and biochemical mechanisms exert modulatory effects shaping the final outcome of the IGF-stimulated processes, frequently leading to neoplastic transformation in the case of irreparable disbalance. The IGF system and related molecules and pathways which participate in the development of CRC are the focus of this review.

Albumin at the intersection between antioxidant and pro-oxidant in patients on peritoneal dialysis.

Albumin (HSA) is a multifunctional protein and due to its free Cys34 thiol group, represents a main source of free thiols in the circulation. This property of HSA, combined with its ability to sequester redox active Cu(II) ions, makes HSA a dominant circulatory antioxidant. End stage kidney disease (ESRD) is a condition accompanied by elevated oxidative stress. The aim of the present study was to examine changes in the antioxidative capacity of HSA and Cu(II) binding affinity in patients on peritoneal dialysis (PD), and relate it to the Cys34 thiol group content and other structural changes of this molecule. HSA molecules are modified in ESRD patients subjected to PD, having significantly lower thiol group and bound Cu(II) content, reduced antioxidant capacity, an increased content of advanced glycation end-products and altered conformation. Also, Cu(II) binding capacity of HSA in these patients is impaired, since a significant portion of the high-affinity metal-binding site is unable to interact with Cu(II). Taking into account that the concentration of Cu(II) in the circulation of ESRD patients is much higher than in healthy persons and that Cu(II) binding capacity of HSA in these patients is significantly impaired, HSA may be considered as a novel circulatory pro-oxidant, thus exacerbating oxidative stress.

Gestational Diabetes is Associated with an Increased Expression of miR-27a in Peripheral Blood Mononuclear Cells.

BACKGROUND: Dysregulation of microRNA-based mechanisms is associated with various human pathologies, including gestational diabetes mellitus (GDM), suggesting they may be  potential diagnostic and/or prognostic biomarkers of GDM. METHODS: The expression of miR-340-5p, miR-27a-3p and miR-222-3p in peripheral blood mononuclear cells (PBMCs) obtained from patients with GDM (n = 42) and healthy controls (n = 34) were evaluated, together with their correlation to the clinical parameters of participants and their newborns. Expression of the selected microRNAs was quantified by quantitative real-time polymerase chain reaction (qPCR), after reverse transcription with microRNA-specific stem-loop primers. RESULTS: The expression of miR-27a-3p was significantly higher in patients with GDM than in controls (p = 0.036), whereas no significant difference between groups was found for the other two tested microRNAs. The expression level of miR-27a-3p in GDM patients was found to negatively correlate with the number of erythrocytes, concentration of haemoglobin, haematocrit, and low- and high-density lipoprotein (LDL/HDL) ratio, and positively with the concentration of glycated haemoglobin (HbA1c). In the case of miR-222-3p, a negative correlation between its expression and the concentration of cholesterol, LDL and LDL/HDL ratio was found only in healthy pregnant women. The expression level of miR-340-5p negatively correlated with erythrocyte count, haemoglobin concentration and haematocrit in GDM patients, as well as with the concentration of cholesterol, LDL and LDL/HDL ratio in healthy women. CONCLUSIONS: The results obtained illustrate the potential of PBMC-derived microRNA miR-27a-3p to serve as a diagnostic biomarker of GDM. On the other hand, MiR-27a and miR-340 may help in assessing the metabolic status relevant for pregnancy.

Association between genetic variants in DICER1 and cancer risk: An updated meta-analysis.

Dysfunctions in mechanisms of gene regulation based on RNA interference are recognized as a common feature of the molecular basis of cancer pathogenesis. Therefore, as one of the crucial components of the machinery involved in the biogenesis of both siRNAs and microRNA molecules, DICER was recognized as one of the candidates for the research in the field of carcinogenesis. Due to their potential functional properties, several genetic variants located within DICER1 gene were analyzed for their possible association with the susceptibility to cancer through case-control studies. In order to elucidate their effect on the overall cancer risk, we conducted an updated meta-analysis of all eligible association studies. The publications were selected based on PubMed database search, while OpenMeta-analyst and MetaGenyo software were used for quantitative data synthesis. Statistically significant results were found for the association of rs1057035 with the overall cancer risk under multiple genetic models (PCT vs. TT < 0.001, ORCT vs. TT = 0.870, 95% CI = 0.812-0.933; Pallelic = 0.009, ORallelic = 0.896, 95% CI = 0.825-0.973; Pdom < 0.001, ORdom = 0.874, 95% CI = 0.817-0.934; Poverdom = 0.004, ORoverdom = 0.858, 95% CI = 0.773-0.953). Other selected genetic variants within DICER1, rs13078, rs1209904 and rs3742330, did not show the association with the overall susceptibility to malignant diseases. We conclude that rs1057035 may represent a potential biomarker associated with the risk of developing cancer, which requires a confirmation in a larger set of studies.

Fibrinogen Fucosylation as a Prognostic Marker of End-Stage Renal Disease in Patients on Peritoneal Dialysis.

Glycosylation may strongly affect protein structure and functions. A high risk of cardiovascular complications seen in patients with end-stage renal disease (ESRD) is, at least partly associated with delayed clot formation, increased clot strength, and delayed cloth lysis. Taking into consideration that fibrinogen mediates these processes, we isolated fibrinogen from the plasma from patients with ESRD on peritoneal dialysis (ESRD-PD), and examined glycosylation of native fibrinogen and its subunits by lectin-based microarray and lectin blotting. Compared to healthy controls, fibrinogen from patients had increased levels of A2BG2 and decreased levels of FA2 glycan. The distribution of glycans on individual chains was also affected, with the γ chain, responsible for physiological functions of fibrinogen (such as coagulation and platelet aggregation), being most prone to these alterations. Increased levels of multi-antennary N-glycans in ESRD-PD patients were also associated with the type of dialysis solutions, whereas an increase in the fucosylation levels was strongly related to the peritoneal membrane damage. Consequently, investigation of fibrinogen glycans can offer better insight into fibrinogen-related complications observed in ESRD-PD patients and, additionally, contribute to prognosis, choice of personalised therapy, determination of peritoneal membrane damage, and the length of utilization of peritoneum for dialysis.

Association of KLK3, VAMP8 and MDM4 Genetic Variants within microRNA Binding Sites with Prostate Cancer: Evidence from Serbian Population.

A growing number of studies have suggested that genetic variants affecting the micro-RNA- binding mechanisms (miRSNPs) constitute a promising novel class of biomarkers for prostate cancer (PCa) biology. Among the most extensively studied miRSNPs in the context of cancer is the variation rs4245739 in the MDM4 gene, while a recent large-scale analysis revealed significant differences in genotype distributions between aggressive and non-aggressive disease for rs1058205 in KLK3 and rs1010 in VAMP8. In this study, we examined a total of 1083 subjects for these three variants using Taqman® SNP Genotyping Assays. Three hundred and fifty-five samples of peripheral blood were obtained from patients with PCa and 358 samples from patients with benign prostatic hyperplasia (BPH). The control group consisted of 370 healthy volunteers. Comparisons of genotype distributions among PCa and BPH patients, as well as between PCa patients and healthy controls, yielded no evidence of association between the analyzed genetic variants and the risk of developing PCa. However, all three tested genetic variants have shown the association with the parameters of PCa progression. For KLK3 variant rs1058205, minor allele C was found to associate with the lower serum PSA score in PCa patients (PSA > 20 ng/ml vs. PSA < 10 ng/ml comparison, Prec = 0.038; ORrec = 0.20, 95%CI 0.04-1.05). The obtained results point out the potential relevance of the tested genetic variants for the disease aggressiveness assessment.

Association between genetic variants in genes encoding Argonaute proteins and cancer risk: A meta-analysis.

With the accumulation of evidence of the involvement of small-RNA-based regulatory mechanisms in carcinogenesis, genes encoding Ago proteins emerged as candidates for case-control studies on cancer. Since the data from association studies on various cancer types was not previously meta-analyzed, the potential effect of these variants on cancer risk in general was not previously evaluated. Therefore, we conducted a meta-analysis of all eligible studies, testing multiple genetic models of association. The identification of publication was based on PubMed database search, while OpenMeta-analyst, as well as MetaGenyo software, were used for quantitative data synthesis. AGO1 genetic variant rs636832 was found to associate with the overall cancer risk, assuming the overdominant genetic model (P =  0.030; ORoverdom = 0.865, 95%CI 0.759-0.986). For the same genetic variant, statistical significance was reached for the association with solid tumors, as well as with lung cancer susceptibility. Similar results were found in the Asians cohort for another AGO1 variant, rs595961. For rs4961280, none of the meta-analyses yielded statistically significant results. We conclude that genetic variants rs636832 and rs595961 located within AGO1 may represent susceptibility variants for specific types of cancer, while the association with malignant diseases was not determined for AGO2 variant rs4961280.

Analysis of association of potentially functional genetic variants within genes encoding miR-34b/c, miR-378 and miR-143/145 with prostate cancer in Serbian population.

MiRNA-associated genetic variants occurring in regulatory regions can affect the efficiency of transcription and potentially modify pri-miRNA or pre-miRNA processing. Since miRNA-based mechanisms are shown to be involved in the pathogenesis of prostate cancer (PCa), the aim of the present study was to evaluate the effect of rs4938723, rs1076064 and rs4705343 occurring in regulatory regions of miR-34b/c, miR-143/145 and miR-378, respectively, on PCa risk and progression in Serbian population. We examined a total of 1060 subjects, of which 350 were patients with PCa, 354 were patients with benign prostatic hyperplasia (BPH), while 356 healthy volunteers were included in the control group. Genotyping of rs4938723, rs1076064 and rs4705343 was performed by using Taqman® SNP Genotyping Assays. Allele C of rs4705342 was found to increase the risk of PCa (P=0.031 for codominant model, P=0.0088 for recessive model). Rs1076064 minor allele G was found to associate with serum PSA score, as well as with PCa T category and disease aggressiveness. For rs4938723 minor allele C was shown to be associated with the lower PCa T category (Pdom=0.0046; OR=0.36, 95 % CI 0.17-0.76) in T2 vs. T1 comparison. Rs4705342 was identified as PCa susceptibility variant in Serbian population, while for rs1076064 and rs4938723 association with PCa progression parameters was found.