High Molecular Weight Glycoprotein Research Articles

Mucin-Inspired Polymeric Fibers for Herpes Simplex Virus Type 1 Inhibition.

Mucus lines the epithelial cells at the biological interface and is the first line of defense against multiple viral infections. Mucins, the gel-forming components of mucus, are high molecular weight glycoproteins and crucial for preventing infections by binding pathogens. Consequently, mimicking mucins is a promising strategy for new synthetic virus inhibitors. In this work, synthetic mucin-inspired polymers (MIPs) as potential inhibitors of herpes simplex virus 1 (HSV-1) are investigated. By using a telechelic reversible addition-fragmentation chain-transfer (RAFT) polymerization technique, a new dendronized polysulfatep(G1AAm-OSO3)PDSwith an amide-backbone similar to the native mucin glycoproteins is synthesized.p(G1AAm-OSO3)PDSshows mucin-like elongated fiber structure, as revealed in cryo-electron microscopy (cryo-EM) imaging, and its HSV-1 inhibition activity together with its previously reported methacrylate analoguep(G1MA-OSO3)PDS is tested. Both of the sulfated MIPs show strong HSV-1 inhibition in plaque reduction assays with IC50 values in lower nanomolar range (<3× 10-9 m) and demonstrate a high cell compatibility (CC50>1.0mgmL-1) with lower anticoagulant activity than heparin. In addition, the prophylactic and therapeutic activity of both MIPs is assessed in pre- and post-infection inhibition assays and clearly visualize their high potential for application using fluorescent microscopy imaging of infected cells.

Pregnancy Associated Plasma Protein-A (PAPP-A) as a Marker to Distinguish Normotensive with Early 2nd Trimester and Late 3rd Trimester Onset of Preeclampsia

Introduction: Preeclampsia is a hypertensive condition that occurs after 20 weeks of gestation accompanied by target organ damage. Complications of preeclampsia can cause intrauterine fetal growth retardation, and placental hypoperfusion, even in the most serious situations, namely termination of pregnancy and death of the fetus and/or mother. Pregnancy-associated plasma protein-A (PAPP-A) is a high molecular weight glycoprotein that is produced in the placenta and secreted into the maternal bloodstream. However, based on several studies that have been conducted, there is uncertainty in the results of assessing PAPP-A levels obtained in pregnant women in the second and third trimesters. Aim: Proving differences in PAPP-A levels in the second and third trimesters in the incidence of early-onset preeclampsia and normotensive pregnancy. Methods: An analytic observational study with a cross-sectional approach was carried out in the delivery room of RSUP Dr. Kariadi Semarang, Halmahera Health Center, Ngesrep Health Center, Bulu Health Center, and private midwife practice in Semarang City. The subjects of the study were six 2nd-trimester preeclampsia patients, fourteen 3rd-trimester preeclampsia patients, and twenty normotensive pregnancy patients who met the inclusion and exclusion criteria. Data were analyzed using Mann Whitney with a significance of p&lt;0.05 Results: There was a significant difference in PAPP-A levels (p&lt;0.001) between the preeclampsia and normotensive pregnancy groups, whereas PAPP-A levels were higher in the preeclampsia group. There were significant differences in PAPP-A levels (p&lt;0.001) between the 2nd-trimester preeclampsia, 3rd-trimester preeclampsia, and normotensive pregnancies, where the highest PAPP-A levels were found in the 2nd-trimester preeclampsia group. Conclusion: There was a significant difference in PAPP-A levels between the second and third trimesters of early-onset preeclampsia compared to normotensive pregnancies, where PAPP-A levels were higher in the second and third trimesters of early-onset preeclampsia. Elevated PAPP-A levels in the second and third trimesters are associated with an increased risk of early-onset preeclampsia.

Fecal-adherent mucus is a non-invasive source of primary human MUC2 for structural and functional characterization in health and disease

The O-glycoprotein Mucin-2 (MUC2) forms the protective colon mucus layer. While animal models have demonstrated the importance of Muc2, few studies have explored human MUC2 in similar depth. Recent studies have revealed that secreted MUC2 is bound to human feces. We hypothesized human fecal MUC2 (HF-MUC2) was accessible for purification and downstream structural and functional characterization. We tested this via histologic and quantitative imaging on human fecal sections; extraction from feces for proteomic and O-glycomic characterization; and functional studies via growth and metabolic assays in vitro. Quantitative imaging of solid fecal sections showed a continuous mucus layer of varying thickness along human fecal sections with barrier functions intact. Lectin profiling showed HF-MUC2 bound several lectins but was weak to absent for Ulex europaeus 1 (α1,2 fucose-binding) and Sambucus nigra agglutinin (α2,6 sialic acid-binding), and did not have obvious b1/b2 barrier layers. HF-MUC2 separated by electrophoresis showed high molecular weight glycoprotein bands (∼1-2 MDa). Proteomics and Western analysis confirmed the enrichment of MUC2 and potential MUC2-associated proteins in HF-MUC2 extracts. MUC2 O-glycomics revealed diverse fucosylation, moderate sialylation, and little sulfation vs. porcine colonic MUC2 and murine fecal Muc2. O-glycans were functional and supported the growth of Bacteroides thetaiotaomicron (B. theta) and short-chain fatty acid (SCFA) production in vitro. MUC2 could be similarly analyzed from inflammatory bowel disease stools, which displayed an altered glycomic profile and differential growth and SCFA production by B. theta vs. healthy samples. These studies describe a new non-invasive platform for human MUC2 characterization in health and disease.

Mucins: an overview of functions and biological activity

This review aims to provide novel evidence on the function of mucins in defense of epithelia and to spot mucin changes in the epithelial surface.&#x0D; High molecular weight glycoproteins known as mucins are distinguished by their substantial O-glycosylation. The cell surface-associated mucins are divided into two categories as surface and gel-forming mucins. These are among the significant mucins expressed by the surface epithelia. Recent developments in functional assays have evaluated their functions in preserving corneal, conjunctival, respiratory, and digestive epithelia. The presentation includes changes in mucin and mucin O-glycan production in epithelial surface illnesses, including infection, non-autoimmune dry eye, autoimmune dry eye, and allergy.&#x0D; Mucins are high molecular weight glycoproteins characterized by their extensive O-glycosylation. Recent advances using functional assays have allowed the examination of their roles in protecting epithelial tissues. Alterations in mucin and mucin O-glycan biosynthesis in epithelial surface disorders, including allergy, non-autoimmune dry eye, cancers, and infection, are presented.

Mucin1 utterance in oral squamous cell carcinoma: A cancer maker and target for nanotheranostics

Mucins are cell bound high molecular weight glycoproteins which are secreted by epithelial cells. Total 21 mucin variants are identified till date. Mucin1 (MUC1) is a transmembrane glycoprotein, which when reacts with beta-catenin, can able to enter the nucleus to activate T-cell factor/leukocyte enhancing factor 1 transcription factors and gene expression, after which it may inhibit cell-cell and cell-stroma interactions and function as a signal transducer, leading to tumor progression.To compare and correlate the expression and positive intensity of MUC1 in oral squamous cell carcinoma, oral epithelial dysplasia and normal oral mucosa using Immunohistochemistry. This study included a total of 45 cases in which the study groups are oral squamous cell carcinoma (n=15), oral epithelial dysplasia (n=15) and control of normal oral mucosa (n=15), which are analysed for the expression of anti MUC1 rabbit monoclonal antibody using immunohistochemical technique.The mucin1 positive cells in the study groups were as follows, 53.3% cases in OSCC, 13.3% cases in OED and none showed positivity in normal oral mucosa. The results obtained were statistically analysed using Kruskal-Wallis test and there was a statistically significant difference in score between the different tissue groups, Kruskal – Wallis H score = 13.034, p = 0.001.There is progressive increase in the MUC1 expression from oral epithelial dysplasia to OSCC. This utterance might be due to suppression of inhibitory proteins for MUC1 immunoexpression in mature atypical squamous cells as well as proposed to act both as an anti-adhesive and adhesive molecule.

Krebs von den Lungen-6 (KL-6) in cerebrospinal fluid from neurosarcoidosis patients.

Krebs von den Lungen-6 (KL-6) is a high molecular weight (MW) glycoprotein mainly secreted by type II pneumocytes because of lung damage or during regeneration. Neurosarcoidosis (NS), where sarcoid granulomas involve the nervous system, occurs in 5-20% of patients with sarcoidosis. No data is currently available on KL-6 in serum or CSF of NS patients. The present study compared KL-6 concentrations in serum and CSF of NS patients versus others with neurodegenerative (ND) or chronic inflammatory demyelinating (DM) diseases. Nine NS patients (mean age 46.2 years, range 16-61 years, M/F 5/4), nine patients with a chronic neurodegenerative disease (mean age 53.1 years, range 37-65 years, M/F 5/4) and nine patients with a chronic demyelinating disease (mean age 46.3 years, range 18-65 years, M/F 5/4) were retrospectively enrolled. Measurable CSF concentrations of KL-6 were detected in 7/9 NS patients but in no ND or DM patients. No significant differences in CSF concentrations of ACE were observed between the three groups (p=0.0819). In NS patients, CSF concentrations of KL-6 were directly correlated with CSF albumin index (r=0.98; p<0.0001), albumin (r=0.979, p=0.0001), IgG (r=0.928, p=0.0009) and total protein concentrations (r=0.945, p=0.0004). KL-6 is a high MW protein, under physiological conditions it is unlikely to cross the blood-brain barrier. We found KL-6 in CSF from NS and not from ND and DM patients. The finding sustains the specificity of changes in KL-6 in this granulomatous disease, suggesting it as a candidate biomarker for recognition of NS.

Mucin expression in liver cancer

Increasing incidence, difficulties in early diagnosis, and a high mortality rate in liver cancer (LC) determine the relevance of studying the mechanisms of its development. The aim of the work is to evaluate the expression of high molecular weight glycoproteins MUC-1, MUC-13 in liver cancer. The object of study is LC tissue samples of 65 patients from the archives and 34 blood serum samples from patients with morphologically confirmed LC. The age of subjects was 26– 97 years. The level of antibodies to MUC-1 and MUC-13 was studied by ELISA. The reference value ranges of MUC-1 (0.250 ± 0.10 ng/ml) and MUC-13 (0.321 ± 0.13 ng/ml) in the blood serum of healthy individuals were established. The concentration of antibodies to MUC-1 and MUC-13 in the blood serum in RP was significantly higher than that in practically healthy individuals. The concentration of MUC-1 and MUC-13 in the LC tumor tissue was higher than that in the blood serum of apparently healthy individuals and LC patients. With a confirmed LC diagnosis, the level of antibodies to MUC-1 in the blood serum, which exceeds 0.373 ng/ml, and the level of antibodies to MUC-13, which is more than 0.939 ng/ml, may indicate a high risk of a tumor process.

Group A streptococci induce stronger M protein-fibronectin interaction when specific human antibodies are bound.

Group A streptococcus (GAS) is a highly adapted, human-specific pathogen that is known to manipulate the immune system through various mechanisms. GAS' M protein constitutes a primary target of the immune system due to its spatial configuration and dominance on the bacterial surface. Antibody responses targeting the M protein have been shown to favor the conserved C region. Such antibodies (Abs) circumvent antigenic escape and efficiently bind to various M types. The ability of GAS to bind to fibronectin (Fn), a high molecular weight glycoprotein of the extracellular matrix, has long been known to be essential for the pathogen's evolutionary success and fitness. However, some strains lack the ability to efficiently bind Fn. Instead, they have been found to additionally bind Fn via the A-B domains of their M proteins. Here, we show that human Abs can induce increased Fn-binding affinity in M proteins, likely by enhancing the weak A-B domain binding. We found that this enhanced Fn binding leads to a reduction in Ab-mediated phagocytosis, indicating that this constitutes a GAS immune escape mechanism. We could show that the Fc domain of Abs is necessary to trigger this phenomenon and that Ab flexibility may also play a key role. We, moreover, saw that our Abs could enhance Fn binding in 3 out of 5 emm type strains tested, belonging to different clades, making it likely that this is a more generalizable phenomenon. Together our results suggest a novel synergistic interplay of GAS and host proteins which ultimately benefits the bacterium.

Capillary gel electrophoresis of very high molecular weight glycoproteins. Commercial and tailor-made gels for analysis of human monomeric and secretory immunoglobulin A

Capillary gel electrophoresis (CGE) has been widely used for analysis of proteins according to their size. However, to our knowledge, this technique has not been optimized to immunoglobulin A (IgA) analysis, a protein of current and emerging high interest in several fields. IgA is the first barrier of human body against pathogens. This protein in human milk and colostrum is essential for immune protection of newborns and treatment of milk for storage in Human Milk Banks may alter IgA. The emerging use of IgA as therapeutic treatment also encourages the development of analysis methods for this class of immunoglobulins.IgA is far more heterogeneously glycosylated and complex than the well-studied IgG molecules. IgA in serum is mainly monomeric (mIgA) with about 160 kDa, while in secretions such as saliva, milk, colostrum, etc, secretory immunoglobulin A (sIgA) is the predominant form. This is a dimer where both monomers are linked by the J-chain and the secretory component accounting all together for a MW higher than 400 kDa including the glycans. This size is far from the 225 kDa MW for which commercial CGE kits are intended.The general rules governing CGE behavior of analytes cannot be directly applied to every protein. Addressing studies directed specifically to target proteins is specially needed for the large size and highly complex target analytes of this study. In this work the effect of several factors on CGE analysis of human serum and colostrum IgA is studied. The feasibility of performing analysis of both IgA classes using a commercial CGE kit is shown. In addition, this work introduces another novelty by preparing tailor-made reproducible gel buffers and to characterize them in terms of dynamic viscosity, conductivity, and electroosmotic flow mobility in bare fused silica capillaries. The possibility of analyzing mIgA and sIgA in less than 10 min using these tailor-made gels is demonstrated. Inter-day variation (RSD) for the main peak of sIgA is 0.25% for migration time (tm) and 0.27% for percentage corrected peak area (Acorr).

Enhancing the technofunctionality of γ-aminobutyric acid enriched germinated wheat by modification of arabinoxylan, gluten proteins and liquid lamella of dough

To enhance the technofunctionality of germinated wheat enriched with γ-aminobutyric acid, xylanase (Xyn) and glucose oxidase (Gox) were incorporated with emphasis on modifying the key components. Combination of Xyn and Gox enhanced steamed bread quality with optimum loaf volume and textural property. Continuous and dense gluten network was facilitated and improved viscoelasticity of dough. Water solubility of arabinoxylan (AX) enhanced with Xyn and the molecular weight was more homogeneous distributed throughout bread making process with Xyn and Gox. Polymerization behavior of α-/γ-gliadin and glutenin was suppressed in steamed bread, while incorporation of AX to insoluble proteins was enhanced by enzymes. In addition, the promoted formation of high molecular weight glycoprotein in the liquid lamella of dough enhanced the thermal stability of foams and contribute to superior quality of steamed bread. Results demonstrated that germinated wheat could be exploited as a functional ingredient with desirable technofunctionality by modification of the components.

Genome-wide bidirectional CRISPR screens identify mucins as host factors modulating SARS-CoV-2 infection

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a range of symptoms in infected individuals, from mild respiratory illness to acute respiratory distress syndrome. A systematic understanding of host factors influencing viral infection is critical to elucidate SARS-CoV-2–host interactions and the progression of Coronavirus disease 2019 (COVID-19). Here, we conducted genome-wide CRISPR knockout and activation screens in human lung epithelial cells with endogenous expression of the SARS-CoV-2 entry factors ACE2 and TMPRSS2. We uncovered proviral and antiviral factors across highly interconnected host pathways, including clathrin transport, inflammatory signaling, cell-cycle regulation, and transcriptional and epigenetic regulation. We further identified mucins, a family of high molecular weight glycoproteins, as a prominent viral restriction network that inhibits SARS-CoV-2 infection in vitro and in murine models. These mucins also inhibit infection of diverse respiratory viruses. This functional landscape of SARS-CoV-2 host factors provides a physiologically relevant starting point for new host-directed therapeutics and highlights airway mucins as a host defense mechanism.

The subcommissural organ maintains features of neuroepithelial cells in the adult mouse.

The subcommissural organ (SCO) is a part of the circumventricular organs located in the dorsocaudal region of the third ventricle at the entrance of the aqueduct of Sylvius. The SCO comprises epithelial cells and produces high molecular weight glycoproteins, which are secreted into the third ventricle and become part of Reissner's fibre in the cerebrospinal fluid. Abnormal development of the SCO has been linked with congenital hydrocephalus, a condition characterized by excessive accumulation of cerebrospinal fluid in the brain. In the present study, we characterized the SCO cells in the adult mouse brain to gain insights into the possible role of this brain region. Immunohistochemical analyses revealed that expression of Pax6, a transcription factor essential for SCO differentiation during embryogenesis, is maintained in the SCO at postnatal stages from P0 to P84. SCO cells in the adult brain expressed known neural stem/progenitor cell (NSPC) markers, Sox2 and vimentin. The adult SCO cells also expressed proliferating marker PCNA, although expression of another proliferation marker Ki67, indicating a G2/M phase, was not detected. The SCO cells did not incorporate BrdU, a marker for DNA synthesis in the S phase. Therefore, the SCO cells have a potential for proliferation but are quiescent for cell division in the adult. The SCO cells also expressed GFAP, a marker for astrocytes or NSPCs, but not NeuN (for neurons). A few cells positive for Iba1 (microglia), Olig2 (for oligodendrocytes) and PDGFRα (oligodendrocyte progenitors) existed within or on the periphery of the SCO. These findings revealed that the SCO cells have a unique feature as secretory yet immature neuroepithelial cells in the adult mouse brain.

Technical report: In-gel sample preparation prior to proteomic analysis of bovine faeces increases protein identifications by removal of high molecular weight glycoproteins

Bovine faecal composition is complex and a knowledge gap exists in the understanding of the bovine faecal proteome. In the present study, in-gel sample preparation (IGSP) of faecal samples prior to proteomics showed an increase in the number of proteins identified in faecal samples compared to those processed by filter-aided sample preparation (FASP). The optimised sample preparation method removed high molecular weight glycoproteins as part of the clean-up process of the faecal samples, and in combination with in-gel digestion before liquid chromatography with tandem mass spectrometry (LC-MS/MS). The use of IGSP led to enhanced protein identification with increases in the number of peptides identified and in the percent coverage of proteins in the bovine faecal samples. SignificanceCharacterization of faecal proteins has the potential to increase our understanding of host responses to changes such as diet, disease and drug-treatment. In-gel sample preparation prior to proteomics can be used to remove high molecular weight glycoproteins and reduce protein/peptide loss in FASP. This method of sample preparation will have application not only in the investigation of bovine faecal extracts but also in studies where large molecules such as glycoproteins or oligosaccharides could have detrimental influences on sample preparation involving ultrafiltration.

Historical aspects of muscle research in the Dubowitz Neuromuscular Centre: the Hammersmith days

Historical aspects of muscle research in the Dubowitz Neuromuscular Centre: the Hammersmith days

Immunotoxin Targeting Cell-Bound MUC1 on Myelomonocytic and Monocytic Leukemic Cells

Cell surface molecules aberrantly or overexpressed by myeloid leukemic cells represent potential disease-specific therapeutic targets for antibodies. MUC1, a polymorphic type I high molecular weight glycoprotein represents such a target. MUC1 is cell surface molecule with extracellular, transmembrane, and cytoplasmic domains; the cytoplasmic tail participates in intracellular signaling. Cleavage of MUC1 yields two unequal chains: a large extracellular alpha subunit bound in strong non-covalent interaction to a smaller beta subunit containing the transmembrane and cytoplasmic domains. Essentially all anti-MUC1 antibodies reported to date target the highly immunogenic MUC1 alpha chain. Because the MUC1 alpha chain binds the cell only intermittently in an ‘on-and-off’ manner, agents directed against the alpha chain will not effectively target MUC1+ cells. In contrast, the MUC1 SEA domain represents a stable structure fixed to the cell surface at all times. We therefore generated mAbs that specifically recognize the cell-bound MUC1 SEA domain. One of them, a partially humanized murine mAb termed DMB-5F3 was used to examine the expression of MUC1 on leukemic cells by flow cytometry. DMB-5F3 was found to strongly bind not all AML, but rather a specific set of AML subtypes: Chronic Myelomonocytic Leukemia (CMML); Acute Myelomonocytic and Monocytic Leukemia (AML-M4 and AML-M5), and Juvenile Myelomonocytic Leukemia (JMML). Other AML subtypes did not express MUC1. To test the cytotoxic ability of anti-MUC1 antibodies against MUC-expressing AML, we generated a humanized, dimeric single-chain DMB-5F3 scFv fragments fused to recombinant gelonin, a type I ribosome-inactivating toxin which itself is incapable of cell binding or internalization into mammalian cells to which a 6xHis-tag was added. The anti-MUC1 scFv-toxin cytometrically identified using an anti-6xHis-tag was found to bind MUC1 positive cell line. We examined the cytotoxic effects of anti MUC1 scFV-toxin using the MUC-1 positive AML cell lines MV411 and MOLM14 (monocytic leukemic subtype). MUC1 scFV-toxin was found to be highly cytotoxic on MUC1+ AML cell lines, resulting in direct cell death. Our data strongly support the use of an anti-MUC 1 antibody-drug conjugate to selectively target and inhibit the growth of MUC1-expressing AML. DisclosuresNo relevant conflicts of interest to declare.

A pan-cancer analysis of MUC family genes as potential biomarkers for immune checkpoint therapy.

2598 Background: Mucin (MUC) is a family of high-molecular weight glycoproteins and increased mucin production occurs in many malignant tumors. Recent studies have found relationship between mutations of some MUC family genes and efficacy of immunotherapy. Here, we explored the associations of MUC family genes (MUC2, MUC3A, MUC4, MUC5B, MUC6, MUC12, MUC16, MUC17, MUC19) mutation with ICI response based on multidimensional data from multiple solid tumors. Methods: 15 solid tumor types of TCGA genomic data for 6138 patients was used to evaluate tumor mutational burden (TMB) differences between MUC family genes mutation group and wildtype group. TMB was calculated as the total count of nonsynonymous mutations in coding sequence. Neoantigens of 3039 samples across 11 solid tumor types were obtained from The Cancer Immunome Atlas. A pan-cancer immunotherapy cohort (Broad/Dana-Farber, Nat Genet 2018, N = 249) was used to explore the relationship between mutations of MUC family genes and its efficacy of immunotherapy. Results: The most common mutated MUC genes (frequency &gt; 5%) were MUC16 (25.3%), MUC17 (10.8%), MUC5B (10.5%), MUC4 (8.6%), and MUC2 (5.1%). The data between MUC mutation group and wild type group showed a significant difference (P &lt; 0.01) in TMB. Median TMB across fifteen tumors in MUC mutation group and wild type group is 7.04 mutations per Mb and 2.07 mutations per Mb, separately. The TNB between two group is also showed a significant difference (P &lt; 0.01). Median TNB across nine types tumor in MUC mutation group and wild type group is 121.5 neoantigens and 34.0 neoantigens, separately. A multivariable analysis across the pan-cancer cohort using Cox proportional-hazards regression demonstrated that KMT2C mutation was associated with better OS (hazard ratio, 0.66; 95%CI, 0.45-0.99; P = 0.042), adjusting for sex and cancer type. Conclusions: The results indicated that MUC family genes mutation was associated with a higher TMB and TNB. Analysis of immunotherapy cohort data showed MUC family was associated with better OS. These findings indicate that these genes mutation may serve as a predictive biomarker for ICI in solid tumor patients.

RvD2 stimulates conjunctival goblet cell secretion by activating multiple Ca 2+ ‐dependent intracellular signaling pathways

Background Resolvin (Rv)D2 is biosynthesized from docosahexaenoic acid (DHA) and was identified from murine self-resolving exudates during the resolution phase of self-limited acute inflammation in vivo. RvD2 was identified in non-diseased human tears, which indicates the potential physiological role of RvD2 on the ocular surface. We determined the action of RvD2 on conjunctival goblet cells and the Ca2+- dependent signaling pathways involved. Method Goblet cells were cultured from male rat conjunctiva. Signaling pathways were studied by measuring intracellular [Ca2+] ([Ca2+]i) using fura 2/AM. Mucin secretion was determined using an enzyme-linked lectin assay (ELLA). Results Administration of RvD2 at 10-10M-10-7M to conjunctival goblet cells increased both the [Ca2+]i and the secretion of high molecular weight glycoprotein including the secretory mucin MUC5AC in a dose-dependent manner. The depletion of extracellular calcium significantly blocked the RvD2-induced [Ca2+]i increase. Pre-treatment with phospholipase (PL) C inhibitor U73122 (10-6M), significantly blocked the [Ca2+]i increase and mucin secretion induced by RvD2 10-8M, U73343 as the negative control failed to lower the RvD2-induced [Ca2+]i increase and mucin secretion. Treatment with 2APB 10-6M, the inositol trisphosphate (IP)3 receptor inhibitor, significantly reduced the RvD2-induced [Ca2+]i increase and the mucin secretion. Pre-treatment with 1-butanol 0.3%, the inhibitor of PLD, significantly blocked the [Ca2+]i increase induced by RvD2 10-8M, but not mucin secretion. The PLA2 inhibitor aristolochic acid 10-5M did not block the RvD2-induced [Ca2+]i increase, but successfully downregulated the RvD2-induced mucin secretion. Conclusion RvD2 induces an increase in [Ca2+]i level and mucin secretion in conjunctival goblet cells. The [Ca2+]i increase is dependent upon activation of PLC and PLD, but not PLA2 pathways. The mucin secretion is dependent on PLA2 and PLC, but not PLD pathways. RvD2 may act as a regulator of conjunctival goblet cells physiologically.

Correlations between colorectal cancer biomarkers

This short review describes the main biomarkers used in research and clinical practice for colon cancer. There are several categories of biomarkers, depending on the substrate they represent or the method of harvesting. We will discuss about genetic biomarkers like microsatellite instability, KRAS, BRAF etc. Blood-based biomarkers available to monitor patients with colorectal cancer are represented by CEA and carbohydrate antigen 19-9 (CA19-9). CEA, a high molecular weight glycoprotein, is found in embryonic tissue and colorectal malignancies. We also present the stool markers. These markers are useful for screening patients at risk to develop colon cancer. The markers are also used to monitor the progression of the disease and the results of systemic therapy.

Immunocytochemistry of In vitro produced mucin from HT29- MTX cell line

Mucus represents the first line of defense against pathogens and damage in the human intestine. the majorcomponents of mucus are mucins which are high molecular weight glycoproteins providing potential bindingsites for both pathogenic and commensal microorganisms. To study host-enteric pathogen interactions,human intestinal cell models are widely used. Epithelial cell lines derived from human colon carcinomasproviding useful experimental alternatives as getting normal intestinal cells is not easily applicable.In the current study, we described a condition enhances HT29-MTX cells to produce continuous mucuslayer. Immunocytochemistry was used to visualize Muc 2, the most secreted mucin from HT29-MTX cellsusing anti Muc2 antibody.The results showed that incubation of HT-29 MTX cells with shaking and providing cells with fresh mediumonce to twice daily for three weeks can be the best way to mimic the natural behavior of the intestine whichenhanced the seeded cells to produce the mucin in vitro.

Characterization of the glycans involved in sea urchin Paracentrotus lividus reversible adhesion

Sea urchins have hundreds of specialized adhesive organs, the tube feet, which play a key role in locomotion, substrate attachment and food capture. Tube feet are composed by two functional units: a proximal cylindrical stem that is mobile and flexible, attached to a distal flattened disc that produces adhesive secretions. Oral tube feet discs possess a specialized duo-glandular epidermis that produces adhesive and de-adhesive secretions, enabling strong but reversible adhesion to the substrate. Due to the growing interest in biomimetic adhesives, several studies have been carried out to characterize sea urchin adhesives, and up to date, it has been shown that it is composed by proteins and glycans. The protein fraction has been the subject of several studies, that pin-pointed several adhesion-related candidates. Contrastingly, little is known about the glycans that compose sea urchin adhesives. This study aims at contributing to this topic by focusing on the characterization of the glycosidic fraction of the adhesive secreted by the sea urchin Paracentrotus lividus (Lamarck, 1816), using a battery of 22 lectins, applied to 3 complementary techniques. Our results show that five lectins label exclusively the disc adhesive epidermis and simultaneously the secreted adhesive, being, therefore, most likely relevant for sea urchin adhesion. In addition, it was possible to determine that the glycosidic fraction of the adhesive is composed by a high molecular weight glycoprotein containing N-acetylglucosamine oligomers.