Serum Clusterin Concentration and Its Glycosylation Changes as Potential New Diagnostic Markers of SARS-CoV-2 Infection and Recovery Process.

COVID-19 is an infectious disease caused by the SARS-CoV-2 virus. Glycoprotein clusterin (CLU) has many functions such as phagocyte recruitment, complement system inhibition, apoptosis inhibition, hormone and lipid transport, as well as in the immune response. The study aimed to assess the changes in CLU concentrations and the profile and degree of CLU glycosylation between patients with severe COVID-19, convalescents, and healthy subjects (control). The profile and degree of serum CLU N-glycosylation were analyzed using lectin-ELISA with specific lectins. CLU concentrations were significantly lower and relative reactivities of CLU glycans with SNA (Sambucus nigra agglutinin) were significantly higher in severe COVID-19 patients in comparison to convalescents and the control group. The relative reactivities of CLU glycans with MAA (Maackia amurensis agglutinin), together with relative reactivity with LCA (Lens culinaris agglutinin), were also significantly higher in patients with severe COVID-19 than in convalescents and the control group, but they also significantly differed between convalescents and control. The development of acute inflammation in the course of severe COVID-19 is associated with a decrease in CLU concentration, accompanied by an increase in the expression of α2,3-linked sialic acid, and core fucose. Both of these parameters can be included as useful glycomarkers differentiating patients with severe COVID-19 from convalescents and the control group, as well as convalescents and healthy subjects.

Evaluating the Neuroprotective Potential of Caffeinated Coffee in the Context of Aluminum-Induced Neurotoxicity: Insights from a PC12 Cell Culture Model.

Exposure to aluminum (Al) and its compounds is an environmental factor that induces neurotoxicity, partially through oxidative stress, potentially leading to the development of neurodegenerative diseases. Components of the diet, such as caffeinated coffee, may play a significant role in preventing these diseases. In the present study, an experimental model of PC12 cells (rat pheochromocytoma tumor cells) was developed to investigate the influence of caffeine and caffeinated coffee on neurotoxicity induced by Al compounds and/or oxidative stress. For the induction of neurotoxicity, aluminum maltolate (Almal) and H2O2 were used. The present study demonstrates that 100 µM Almal reduced cell survival, while caffeinated coffee with caffeine concentrations of 5 µg/mL and 80 µg/mL reversed this effect, resulting in a higher than fivefold increase in PC12 cell survival. However, despite the observed antioxidant properties typical for caffeine and caffeinated coffee, it is unlikely that they are the key factors contributing to cell protection against neurotoxicity induced by both oxidative stress and Al exposure. Moreover, the present study reveals that for coffee to exert its effects, it is possible that Al must first activate certain mechanisms within the cell. Therefore, various signaling pathways are discussed, and modifications of these pathways might significantly decrease the risk of Al-induced neurotoxicity.

The Influence of Clusterin Glycosylation Variability on Selected Pathophysiological Processes in the Human Body.

The present review gathers together the most important information about variability in clusterin molecular structure, its profile, and the degree of glycosylation occurring in human tissues and body fluids in the context of the utility of these characteristics as potential diagnostic biomarkers of selected pathophysiological conditions. The carbohydrate part of clusterin plays a crucial role in many biological processes such as endocytosis and apoptosis. Many pathologies associated with neurodegeneration, carcinogenesis, metabolic diseases, and civilizational diseases (e.g., cardiovascular incidents and male infertility) have been described as causes of homeostasis disturbance, in which the glycan part of clusterin plays a very important role. The results of the discussed studies suggest that glycoproteomic analysis of clusterin may help differentiate the severity of hippocampal atrophy, detect the causes of infertility with an immune background, and monitor the development of cancer. Understanding the mechanism of clusterin (CLU) action and its binding epitopes may enable to indicate new therapeutic goals. The carbohydrate part of clusterin is considered necessary to maintain its proper molecular conformation, structural stability, and proper systemic and/or local biological activity. Taking into account the wide spectrum of CLU action and its participation in many processes in the human body, further studies on clusterin glycosylation variability are needed to better understand the molecular mechanisms of many pathophysiological conditions. They can also provide the opportunity to find new biomarkers and enrich the panel of diagnostic parameters for diseases that still pose a challenge for modern medicine.

The Association between Clusterin Sialylation Degree and Levels of Oxidative-Antioxidant Balance Markers in Seminal Plasmas and Blood Sera of Male Partners with Abnormal Sperm Parameters.

Nearly 30% of infertility cases are caused by male factor. This study aimed at checking the associations between the sialylation degree of glycoprotein clusterin (CLU) and levels of oxidative-antioxidant balance markers in infertile men. Using lectin-ELISA with biotinylated lectins specific to α2,6-linked (Sambucus nigra agglutinin, SNA) and α2,3-linked (Maackia amurensis agglutinin, MAA) sialic acid (SA), the CLU sialylation in 132 seminal plasmas (SP) and 91 blood sera (BS) were analyzed. Oxidative-antioxidant status was measured by determining Sirtuin-3 (SIRT3), Sirtuin-5 (SIRT5), total antioxidant status (TAS), and ferric reducing antioxidant power (FRAP) levels. We indicate that multiple sperm disorders are associated with decreased expression of MAA-reactive SA in SP. Decreased SP SIRT3 concentrations may be associated with teratozoospermia and oligoasthenoteratozoospermia. ROC curve and cluster analysis revealed that SP relative reactivity of CLU glycans with MAA, the value of MAA/SNA ratio, and SIRT3 and SIRT5 concentrations may constitute an additional set of markers differentiating infertile oligoasthenoteratozoospermic patients (OAT) from normozoospermic (N), asthenoteratozoospermic (AT) and teratozoospermic (T). The multinomial logistic regression analysis confirmed the potential utility of SIRT3 determinations for differentiation between N and OAT groups as well as between N and T groups for SIRT3 and SIRT5. For BS, based on ROC curve and cluster analysis, relative reactivities of CLU glycans with SNA, MAA, SIRT3 and FRAP concentrations may be useful in the differentiation of normozoospermic patients from those with sperm disorders. The multinomial logistic regression analysis showed that the SNA relative reactivity with CLU glycans significantly differentiated the N group from AT, OAT and T groups, and FRAP concentrations significantly differed between N and AT groups, which additionally confirms the potential utility of these biomarkers in the differentiation of infertile patients with abnormal sperm parameters. The knowledge about associations between examined parameters may also influence future research aimed at seeking new male infertility therapies.

Caffeine as a Factor Influencing the Functioning of the Human Body-Friend or Foe?

Nowadays, caffeine is one of the most commonly consumed substances, which presents in many plants and products. It has both positive and negative effects on the human body, and its activity concerns a variety of systems including the central nervous system, immune system, digestive system, respiratory system, urinary tract, etc. These effects are dependent on quantity, the type of product in which caffeine is contained, and also on the individual differences among people (sex, age, diet etc.). The main aim of this review was to collect, present, and analyze the available information including the latest discoveries on the impact of caffeine on human health and the functioning of human body systems, taking into account the role of caffeine in individual disease entities. We present both the positive and negative sides of caffeine consumption and the healing properties of this purine alkaloid in diseases such as asthma, Parkinson's disease, and others, not forgetting about the negative effects of excess caffeine (e.g., in people with hypertension, children, adolescents, and the elderly). In summary, we can conclude, however, that caffeine has a multi-directional influence on various organs of the human body, and because of its anti-oxidative properties, it was, and still is, an interesting topic for research studies including those aimed at developing new therapeutic strategies.

The possible association of clusterin fucosylation changes with male fertility disorders.

In the seminal plasma (n = 118) and serum (n = 90) clusterin (CLU) the fucosylation and the expression of selected fucosyltransferases (FUTs) were analyzed. Samples from infertile men were divided into groups based on the results of the standard semen analysis: normozoospermic (N), teratozoospermic (T), asthenoteratozoospermic (AT) and oligoasthenoteratozoospermic (OAT). The CLU fucosylation was analyzed using lectin-ELISAs with biotinylated lectins specific to α1,3-, α1,2-linked antennary fucose, and α1,6-linked core fucose (LTA, UEA, and LCA, respectively). The concentrations of FUT3 and FUT4, reflecting the expression of Le oligosaccharide structures, were measured using ELISA tests. The differences in serum CLU and FUT4 concentrations, and in the expression of core fucose and antennary fucose α1,2-linked in CLU glycans between the N group and other groups examined suggest that the disturbances in sperm count, motility, and morphology are not the only cause of male infertility. Lack of similarities between levels of examined parameters in blood serum and seminal plasma may suggest the differences in mechanisms leading to glycoproteins glycosylation. It confirmed the observed differences in concentrations of seminal plasma CLU, FUT3, and FUT4 between the OAT group and N, T, AT groups, indicating that decreased sperm count may be related to these parameters expression. The serum CLU concentrations and expression of core fucose and fucose α1,2-linked in CLU, seem to be good markers differentiating normozoospermic men from those with abnormal sperm parameters, which was not observed for seminal plasma.

The Role of ApoE Expression and Variability of Its Glycosylation in Human Reproductive Health in the Light of Current Information.

Apolipoprotein E (ApoE), a 34-kDa glycoprotein, as part of the high-density lipoprotein (HDL), has antioxidant, anti-inflammatory and antiatherogenic properties. The variability of ApoE expression in the course of some female fertility disorders (endometriosis, POCS), and other gynecological pathologies such as breast cancer, choriocarcinoma, endometrial adenocarcinoma/hyperplasia and ovarian cancer confirm the multidirectional biological function of ApoE, but the mechanisms of its action are not fully understood. It is also worth taking a closer look at the associations between ApoE expression, the type of its genotype and male fertility disorders. Another important issue is the variability of ApoE glycosylation. It is documented that the profile and degree of ApoE glycosylation varies depending on where it occurs, the type of body fluid and the place of its synthesis in the human body. Alterations in ApoE glycosylation have been observed in the course of diseases such as preeclampsia or breast cancer, but little is known about the characteristics of ApoE glycans analyzed in human seminal and blood serum/plasma in the context of male reproductive health. A deeper analysis of ApoE glycosylation in the context of female and male fertility will both enable us to broaden our knowledge of the biochemical and cellular mechanisms in which glycans participate, having a direct or indirect relationship with the fertilization process, and also give us a chance of contributing to the enrichment of the diagnostic panel in infertile women and men, which is particularly important in procedures involved in assisted reproductive techniques. Moreover, understanding the mechanisms of glycoprotein glycosylation related to the course of various diseases and conditions, including infertility, and the interactions between glycans and their specific ligands may provide us with an opportunity to interfere with their course and thus develop new therapeutic strategies. This brief overview details some of the recent advances, mainly from the last decade, in understanding the associations between ApoE expression and some female and male fertility problems, as well as selected female gynecological diseases and male reproductive tract disorders. We were also interested in how ApoE glycosylation changes influence biological processes in the human body, with special attention to human fertility.

Diagnostic Significance of Selected Serum Inflammatory Markers in Women with Advanced Endometriosis.

Endometriosis is a gynecological disease, the pathogenesis of which seems to be directly associated with inflammatory processes. Serum concentrations of IL-1ß, IL-6, hs-CRP, IgG, YKL 40 and PRL, in comparison to the well-known CA 125 levels, were studied with the aim of identifying an additional noninvasive inflammatory marker or set of markers characteristic for endometriosis. The study group included 43 women with endometriosis (E), 35 women with benign gynecological disorders but without endometriosis (NE, non-endometriosis) as a comparative group, and a control group consisting of 18 healthy subjects (C). The serum concentrations of IL-1ß, IL-6, hs-CRP, YKL-40, PRL and CA 125 were significantly higher in the E group (median values: 0.41 pg/mL, 2.42 pg/mL, 2.33 mg/L, 79.30 ng/mL, 21.88 ng/mL and 68.00 U/mL, respectively) than in the control group (median values: 0.21 pg/mL, 0.98 pg/mL, 0.52 mg/L, 49.77 ng/mL, 12.08 ng/mL and 12.20 U/mL respectively), with the significance of p = 0.011, p < 0.001, p = 0.028, p = 0.005, p < 0.001 and p < 0.001, respectively. The IgG concentrations were significantly lower in the endometriosis group (median value: 1061.21 mg/dL) as compared to healthy women (median value: 1210.50 mg/dL; p = 0.025). Significant differences in concentrations of IL-6 (p = 0.040), hs-CRP (p = 0.007) and CA 125 (p < 0.001) were observed in stage III vs. stage IV of endometriosis. Significantly higher concentrations of IL-6 (p = 0.010), hs-CRP (p = 0.037) and PRL (p < 0.001) were observed in the NE group vs. the control group. Only CA 125 concentrations were significantly higher in endometriosis patients as compared to the non-endometriosis group (p < 0.001). The proposed panel of inflammatory markers, especially IL-6, PRL and CA 125, may become a useful tool to identify women with advanced endometriosis who could qualify for treatment.

Sirtuins-The New Important Players in Women's Gynecological Health.

The participation of sirtuins in the regulation of oxidative stress and inflammation lies at the basis of their possible modes of action and is related to their expression in various cell structures; their location in the mitochondria and blood plasma has been indicated as of primary importance. Despite many existing studies, research on sirtuins continues to present an opportunity to discover new functions and dependencies, especially when it comes to women's gynecological health. Sirtuins have a significant role in both the formation and the course of many gynecological diseases. Their role is particularly important and well documented in the course of the development of cancer within the female reproductive organs; however, disturbances observed in the ovary and oocyte as well as in follicular fluid are also widely investigated. Additionally, sirtuins take part in some gynecological disturbances as regulative factors in pathways associated with insulin resistance, glucose and lipids metabolism disorders. In this review, we would like to summarize the existing knowledge about sirtuins in the manner outlined above.

Sirtuins as Important Factors in Pathological States and the Role of Their Molecular Activity Modulators.

Sirtuins (SIRTs), enzymes from the family of NAD+-dependent histone deacetylases, play an important role in the functioning of the body at the cellular level and participate in many biochemical processes. The multi-directionality of SIRTs encourages scientists to undertake research aimed at understanding the mechanisms of their action and the influence that SIRTs have on the organism. At the same time, new substances are constantly being sought that can modulate the action of SIRTs. Extensive research on the expression of SIRTs in various pathological conditions suggests that regulation of their activity may have positive results in supporting the treatment of certain metabolic, neurodegenerative or cancer diseases or this connected with oxidative stress. Due to such a wide spectrum of activity, SIRTs may also be a prognostic markers of selected pathological conditions and prove helpful in assessing their progression, especially by modulating their activity. The article presents and discusses the activating or inhibiting impact of individual SIRTs modulators. The review also gathered selected currently available information on the expression of SIRTs in individual disease cases as well as the biological role that SIRTs play in the human organism, also in connection with oxidative stress condition, taking into account the progress of knowledge about SIRTs over the years, with particular reference to the latest research results.

Preliminary Study on Selected Markers of Oxidative Stress, Inflammation and Angiogenesis in Patients with Bladder Cancer.

In recent years, bladder cancer (BC) has been reported as one of the most commonly occurring cancers among older people, and its detection is still difficult. Therefore, there is a need to search for additional useful markers of disease. Some studies indicate the important roles of inflammation and oxidative stress (OS) in bladder tumour pathogenesis. The aim of this study was to examine the levels of selected markers of OS, inflammation and angiogenesis in blood plasma/serum samples derived from patients with BC, and a healthy control group. Moreover the degrees of change and strength of correlation between values of the analysed markers and tumour stage or grade were estimated. Concentrations of: malondialdehyde (MDA) and advanced oxidation protein products (AOPP), and total antioxidant status (TAS) divided into slow (TAS-s) and fast (TAS-f) antioxidants (spectrophotometric measurement), angiogenin (ANG) (immunoenzymatic method) and C-reactive protein (CRP) (immunoturbidimetric method) were determined in both the studied groups. The majority of values of the examined parameters were significantly higher among patients, while subfractions of TAS were significantly lower in comparison to the control group. Moreover, different values and different strengths of correlation between the examined parameters and cancer stage or grade were noticed. The most significant changes for CRP were observed in T2 and for MDA in G3, while the lowest TAS-f activity was revealed in G1 patients. Increased values of OS parameters, angiogenesis and inflammation markers, in combination with reduced TAS subfractions activity in BC are important in its pathogenesis and will be helpful in estimation of patients' condition.

Long-term changes of salivary exoglycosidases and their applicability as chronic alcohol-drinking and dependence markers.

OBJECTIVES: Investigation of long-term dynamic changes of salivary activity/output of exoglycosidases, deglycosylation processes and their applicability as alcohol markers. METHODS: Exoglycosidase (α-fucosidase (FUC), ß-galactosidase (GAL), ß-glucuronidase (GLU), ß-hexosaminidase (HEX, HEX A and HEX B isoenzymes) and α-mannosidase (MAN)) activities were measured in the saliva of healthy social drinking controls (C), alcohol-dependent non-smokers (ANS) and alcohol-dependent smokers (AS) at the 1st, 15th, 30th and 50th day of abstinence after chronic alcohol drinking. RESULTS: The activity of exoglycosidases was 2-3-fold (MAN), 2-6 fold (FUC), 8-25-fold (HEX A) and 19-40-fold (GLU) higher in the ANS and AS groups than in controls, and had good/excellent sensitivity, specificity and accuracy. The higher outputs of exoglycosidases were in the AS and ANS groups than in controls at the 1st day (GLU, HEX A) and at the 50th day (GLU, FUC, MAN) of abstinence. We found numerous correlations between alcohol-drinking days with GLU and HEX A, alcohol amounts with HEX A and duration of alcohol dependence with FUC and MAN activity/output. CONCLUSIONS: Salivary exoglycosidases/deglycosylation processes were still very high up to 50 days after the end of alcohol consumption. We found markers of chronic alcohol consumption (HEX A), alcohol dependence (FUC and MAN) and chronic alcohol consumption and dependence (GLU).

The content of immunomodulatory glycoepitopes in seminal plasma glycoproteins of fertile and infertile men.

According to a concept of fetoembryonic defence, protein-carbohydrate interaction may be involved in the regulation of maternal immunity that prevents rejection of allograft spermatozoa, embryo and fetus. In the present study we focussed on the evaluation of the expression of glycoepitopes that may be of crucial importance in this process: LewisY (LeY) and LewisX (LeX) as well as terminal sialylation. Polyacrylamide gel electrophoresis with sodium dodecyl sulphate was used to separate seminal plasma samples of fertile (n=10) and infertile (n=103) men; these were then probed with lectins specific to fucose (Lotus tetragonolobus agglutinin and Ulex europaeus agglutinin) and sialic acid (Sambucus nigra agglutinin and Maackia amurensis agglutinin). Differential expression of α2,3-bound sialic acid was found in six out of seven analysed bands, whereas differences in the other analysed glycoepitopes were found in fewer numbers of bands. Mass spectrometry analysis focussed on the identification of proteins carrying glycans with immunomodulatory epitopes, including fibronectin, lactoferrin, clusterin, zinc-α2-glycoprotein, prostate acid phosphatase and prostate-specific antigen; these should be submitted to further detailed analysis.

Comparison of haptoglobin and alpha1-acid glycoprotein glycosylation in the sera of small cell and non-small cell lung cancer patients.

INTRODUCTION: Cancer-related carbohydrate epitopes, which are regarded as potential diagnostic and prognostic biomarkers, are carried on the main acute phase proteins. It is not clear, however, if the glycosylation profile is similar in different glycoproteins, or it is protein specific to some extent. The aim of the study was to compare fucosylation, α2,3 sialylation and expression of sialyl-Lewisx epitopes (sLe(x)) in the serum as a whole, AGP and haptoglobin of small cell (SCLC) and non-small cell lung cancer (NSCLC) patients with respect to healthy subjects as well as the cancer stage and its histological type. MATERIAL AND METHODS: Thirty-three NSCLC, 13 SCLC patients and 20 healthy volunteers were included in the study. Carbohydrate epitopes were detected by means of their reactivity with specific lectins and monoclonal anti-sLe(x) antibodies in direct or dual-ligand ELISA tests. RESULTS: Significantly increased fucosylation was found in total serum in both cancer groups and in NSCLC haptoglobin. No difference was observed in SCLC haptoglobin or α1-acid glycoprotein in both cancer groups. Also α2,3 sialylation was elevated in total serum, but not in α1-acid glycoprotein. This type of sialylation was undetectable in haptoglobin by means of MAA reactivity, in both healthy and cancer subjects. Complete sLe(x) antigens were overexpressed in total NSCLC serum and SCLC AGP, and their level was considerably lowered in cancer haptoglobin. DISCUSSION: Typical acute phase proteins, haptoglobin and AGP, exhibit different glycosylation profiles in lung cancer. Alterations observed in haptoglobin reflected the disease process better than those in AGP. Comparison of haptoglobin and AGP glycosylation to that observed in total serum suggests that some efficient carriers of disease-altered glycoproteins still remain unidentified.

Seminal plasma glycoproteins in male infertility and prostate diseases: is there a chance for glyco-biomarkers?

CONTEXT: Disturbed protein-carbohydrate interactions may underlie the molecular mechanism of some diseases of the male reproductive tract, including infertility and prostate diseases. OBJECTIVE: To summarize the current knowledge on the glycosylation patterns of glycodelin-S, fibronectin, prostate-specific antigen, and α(1)-acid glycoprotein. RESULTS: Some rare glycoepitopes have been found in seminal plasma glycoproteins: high-mannose and polylactosamine-type glycans, and N-glycans containing N-acetyl-galactosamine. The glycosylation profiles occur altered in pathological conditions. CONCLUSION: Further detailed studies may lead up to indicate the biomarkers useful in the management of male reproductive tract disorders.

Terminal monosaccharide screening of synovial immunoglobulins G and A for the early detection of rheumatoid arthritis.

The expressions of some terminal glycotopes of synovial immunoglobulins G, A, and M were analysed in relation to rheumatoid arthritis (RA) progression defined according to early and advanced radiological changes in patients' hands. The relative amounts of terminal monosaccharides were determined by lectin-immunoblotting of immunoglobulin preparations using appropriate lectins able to recognize alpha2,6-linked (Sambucus nigra agglutinin) and alpha2,3-linked (Maackia amurensis agglutinin) sialic acid, galactose (Ricinus communis agglutinin I), N-acetylglucosamine (Griffonia simplicifolia agglutinin II) as well as alpha1,6-linked (Aleuria aurantia lectin), alpha1,3-linked (Lotus tetragonolobus agglutinin), and alpha1,2-linked (Ulex europaeus agglutinin) fucose. The results indicate differences between early and advanced RA stages in the terminal sugar exposition of synovial IgG and IgA, but not IgM. The galactose-deficient glycotope with exposed N-acetylglucosamine of the synovial 33.1-kDa IgG fragment appeared exclusively in the early stage of RA. In contrast, this glycotope of intact synovial IgG and IgA was present in both groups, although with higher proportions in advanced RA. The proportions of the sialyl and fucosyl determinants of intact synovial A and G immunoglobulins were clearly lower in the early RA group than in the advanced. The analysis of terminal oligosaccharide exposition in IgG, IgG fragments, and IgA present in the synovial fluid of RA patients might be applicable as a stage-specific marker in the diagnosis and therapy of RA patients.