Identifying the factors that facilitate or hinder association between galectins and transmembrane mucins is not only critical to understanding the organization of the epithelial glycocalyx, but also may be exploited for potential therapeutic development. Synthetic glycopolymers that emulate natural mucins have been developed during the past few years to study how the structure of mucin glycans and their spatial arrangements along the mucin’s polypeptide backbone affect the interactions with carbohydrate- binding proteins [39,40]. Glycopolymers functionalized with lipid tails have been introduced into membranes of live cells such as ldlD CHO, a cell type lacking endogenous mucins . Here, we show that glycopolymers decorated with pendant cellobiose- and lactose-glycans incorporate into cultures of stratified human corneal epithelial cells (Figure 4), known to contain apical islands of undifferentiated and differentiated cells, the latter featuring glycosylated transmembrane mucins [41,42]. Increasing the amount of cellobiose on the cell surface via glycopolymer insertion enhanced rose bengal uptake (Figure 5), suggesting that interfer- ence with surface recognition of endogenous lactosyl residues impairs barrier function at the ocular surface. Unexpectedly, insertion of lactose-containing glycopolymers, which have the capacity to bind galectin-3, did not enhance barrier function in our three-dimensional culture system; in fact, the regions of rose bengal uptake detected were similar to those of control cultures. A possible explanation is that lactose-containing glycopolymers incorporate into the glycocalyx but fail to compete for galectin-3 binding in the presence of endogenous glycosylated mucins— natural ligands for galectin-3 on apical surfaces [10,30]. Alterna- tively, lactose-containing glycopolymers may incorporate into undifferentiated apical cells with poorly glycosylated mucins, but in insufficient quantities to efficiently induce lattice formation. As restoring barrier function is essential to the treatment of ocular surface disease, further research is required to elucidate the underlying causes that may impair the gain of glycocalyx barrier function when synthetic glycopolymers are used.
The samples were dehydrated using graded ethanol (S27074102, Sinopharm Chemical Reagent Co., Ltd., China), ﬁxed with dimethylbenzene (10023418, Sino- pharm Chemical Reagent Co., Ltd.), and embedded in parafﬁn (SA633001, Sinopharm Chemical Reagent Co., Ltd.), followed by sectioning into slices (Polysine Adhe- sion Slides; SLI-2002, Fuzhou Maixin Biotech. Co., Ltd., China). Subsequently, the sections were dewaxed with xylene. Then the samples were dehydrated using graded ethanol, and washed with PBS. Antigen retrieval was conducted using a pressure cooker: parafﬁn sections were put into the pressure cooker and pressurization was increased slowly. After the pressure cooker emitted steam for 5–6 min, the parafﬁn sections were removed from the heat source and put into the cold water. Then, 1 drop of peroxidase (Beijing Zhongshan Golden Bridge Biotech. Co., Ltd., China) was added to each slice to block the reaction. After, the section was incubated at room tempera- ture for 10 min, washed with PBS, and normal non-immune serum was added and slides were incubated for 10 min. Subsequently, the antibodies were added (mouse anti- human ezrin (ab4069, 1:100), galectin-3 monoclonal anti- body (ab2785, 1:100), PBS as blank control) (All purchased from Santa Cruz Company, USA) and incubated overnight and washed with PBS. Then, biotin-labeled secondary antibody was added (69314360, Sinopharm Chemical Reagent Co., Ltd.) and incubated for 10 min, followed by washing with PBS. Additionally, streptavidin enzyme was added and samples were incubated for 10 min. Again, samples were washed with PBS, and added with DAB solution (SP-9000-D, Beijing Zhongshan Golden Bridge Biotech. Co., Ltd.). After washing with PBS, the samples were hematoxylin stained (CTS-1097, Fuzhou Maixin Bio- tech. Co.), colored in blue and dehydrated with gradient ethanol. Xylene was used for transparency and neutral gum (DAB-0033, Fuzhou Maixin Biotech. Co.) was used for sealing.
Cytokeratin-19 (CK-19) expression in thyroid nodules is in general intense and diffuse in papillary carcinoma and heterogeneous labeling in carcinoma and in follicu- lar adenoma, with nil or low expression in other benign lesions [30,82]. Galectins, especially galectin-3, are sug- gested to play a role in the pathogenesis of well- differentiated thyroid carcinoma, particularly in papil- lary carcinoma and, therefore, it is one of the mar- kers most commonly used to assist in distinguishing thyroid lesions. Hector Battifora mesothelial-1 (HBME-1) has been demonstrated to be important as a thyroid marker of follicular origin, with greater affinity to ma- lignant lesions when compared to benign lesions. Because of that, they are the three most used immuno- markers in pathology practice and each of them had different rates of false-negatives and false-positive results and some authors advocate that a panel of the three markers might be more helpful than the use of a single immunomarker, improving the specificity, posi- tive and negative predictive value and thus diagnostic accuracy .
Schistosoma mansoni s ynthesizes GalNAcb1-4(Fuca1-3)GlcNAc(Lac-DiNAc) structures (N-acetylgalactosamine b1-4 N- acetylglucosamine), which are known to interact with galectin-3 and elicit an intense humoral response. Antigens derived from the eggs and adult worms are continuously drained to MLNs and induce a polyclonal B cell activation. In the present work, we observed that chronically-infected galectin-3 -/- mice exhibited a significant reduced amount of macrophages and B lymphocytes followed by drastic histological changes in B lymphocyte and plasma cell niches in the MLNs. The lack of galectin-3 favored an increase in the lymphoid follicle number, but made follicular cells more susceptible to apoptotic stimuli. There were an excessive quantity of apoptotic bodies, higher number of annexin V + /PI - cells, and reduced clearance
Figure 2. Galectin-3 enhances gastric cancer cell migration/invasion by increasing PAR-1 expression. A, mRNA and protein levels of galectin-3 and PAR-1 detected by RT-PCR and western blot analysis in SNU-638 cells, infected with lenti-virus containing LacZ or galectin-3, and then transfected with PAR-1 siRNA or scRNA for a negative control. b-actin was used as a loading control. B and C, Migration (B) and invasion (C) assays were conducted of SNU-638 cells infected with lenti-virus containing LacZ or galectin-3 cells, and then transfected with PAR-1 siRNA or scRNA for a negative control. Results was shown as histogram (* p,0.001 vs. Cont group). D, Schematic model of PAR-1 promoter with AP-1 binding site, Primers used for ChIP assay were prepared to detect AP-1 binding site (2463,2474) from 2666 to 2307. E, Galectin-3 interacts with c-Jun and fra-1 (as, an AP-1 complex ) in MKN-28 cells. Immunoprecipitation was performed as described in ‘‘Material and Methods’’, and then galectin-3, c-Jun and fra-1 detected by western blot analysis. Whole cell lysates (WCLs) were used as a positive control. F, Analysis of chromatin immunoprecipitation assay using antibodies to galectin-3, c-Jun and Fra-1 in MKN-28 cells transfected with scRNA and galectin-3 siRNA. PCR primer for the PAR-1 gene promoter was used to detect promoter fragment in immunoprecipitates. Input lane, total genomic DNA used as control for the PCR reaction. G, Luciferase activity of AP-1 in lacZ and galectin-3 over-expressing cells. Luciferase assay was performed using AP-1 expression luciferase vector transfection to lacZ and galectin-3 over-expressing cells (* p,0.001 vs. Cont). b-galactoside was used as negative control.
-invading and cells of invasive fronts in CMMT were notable exceptions to this impaired expres- sion of galectin-3-binding sites. A report of Holíková et al., (2002), demonstrated that a2, 6- but not a2, 3-linked N-acetyl-D-neuraminic acid is implicated in masking galectin-3 binding sites in squamous epithelial carcinomas. Hence, in parallel to the use of the Gal3/AP probe, before and after neuraminidase treatment, we assessed the reactivity profile of two plant lectins which recognize distinct types of sialylation. The binding pattern of a 2, 6-Neu5Ac-recognizing plant lectin (i.e. SNA) was inversely associated to Gal3/AP ability to bind to the CMMT tissues. In contrast to a2, 6-, expression of a2, 3- linked sialic acid, assessed with MAL, was not associated to galectin-3 binding site negative areas. Several reports have shown that while only occasionally binding to a2, 6-sialylated glycans, galectin-3 binds well to a2, 3-sialylated glycans (Hirabayashi et al., 2002; de Melo et al ., 2007). Thus, the findings of the present study suggest that, although probably not exclusively, a2, 6 sialylation can be an on/off switch for galectin-3 binding in situ.
O KRAS é a isoforma RAS mais frequentemente mutada em vários cancros, incluindo no cancro colorretal (CCR; 30-50% dos casos), que é uma importante causa de morte em todo o mundo. As mutações mais frequentes neste oncogene, mutações pontuais nos codões 12, 13 e 61, que maioritariamente resultam em substituições G12V, G12D, ou G13D, têm um impacto importante nas decisões terapêuticas. As plataformas de sinalização KRAS na membrana são estabilizadas pela proteína galectina-3 (Gal-3). Gal-3, a única galectina tipo “quimera” dentro da família das galectinas, tem inúmeros ligandos intra e extracelulares, desempenhando papéis centrais em várias funções celulares. Alterações no perfil de expressão da Gal-3 estão correlacionadas com uma variedade de tumores, incluindo CCR, estando envolvidas no crescimento tumoral, transformação, apoptose, angiogénese, adesão, invasão e metastização. A transformação mediada pela Gal-3 é parcialmente atribuída à sua interação específica com KRAS e consequente ativação das suas vias de sinalização. Os resultados fenotípicos dessa interação não estão bem estabelecidos, constituindo assim um importante tema de investigação com possíveis implicações terapêuticas. Por sua vez, p16 é um conhecido supressor tumoral que desempenha vários papéis adicionais, inclusivamente na regulação da angiogénese, apoptose, anoikis, imortalização e senescência. A p16 parece estar relacionada com KRAS e Gal-3, exercendo a sua função de supressão tumoral por regulação negativa de ambas as proteínas para atingir reversão de resistência a anoikis.
Results of immunohistochemical scoring are sum- marized by tumor type in Table 1, representative illustrations being shown in Figures 1 to 3. Reactivity was generally cytoplasmic and sometimes nuclear, espe- cially in the tumors showing strong galectin-3 expression. In summary, strong (3+) immunoreactivity was noted in most of the specimens (61%), followed by 2+ (22%), and 1+ (13%). Only 4% of the cases were immunonegative, including 2 hemangiopericytomas of the dura, and 1 each metastatic carcinoma of the prostate, choroid plexus carcinoma, grade III ependymoma, glioblastoma, leio- myosarcoma, and Langerhans cell histiocytosis. Mimics of SCO, including granular cell tumor and pituicytoma were strongly immunoreactive (100% and 75% of cases, respectively). Other tumors demonstrating 3+ immuno- reactivity included melanoma (78%), nerve sheath tumors (75%), metastatic nonsmall cell carcinoma (74%), and meningiomas (58%) (Table 1). Sarcomas showed variable TABLE 1. (continued)
For both wound healing and the formation of a fibrotic lesion, circulating monocytes enter the tissue and differentiate into fibroblast-like cells called fibrocytes and pro-fibrotic M2a macrophages, which together with fibroblasts form scar tissue. Monocytes can also differ- entiate into classically activated M1 macrophages and alternatively activated M2 macro- phages. The proteases thrombin, which is activated during blood clotting, and tryptase, which is released by activated mast cells, potentiate fibroblast proliferation and fibrocyte dif- ferentiation, but their effect on macrophages is unknown. Here we report that thrombin, tryp- tase, and the protease trypsin bias human macrophage differentiation towards a pro-fibrotic M2a phenotype expressing high levels of galectin-3 from unpolarized monocytes, or from M1 and M2 macrophages, and that these effects appear to operate through protease-acti- vated receptors. These results suggest that proteases can initiate scar tissue formation by affecting fibroblasts, fibrocytes, and macrophages.
Non-alcoholic steatohepatitis (NASH) and resultant liver fibrosis is a major health problem without effective therapy. Some data suggest that galectin-3 null mice are resistant to the development of NASH with fibrosis. We examined the ability of two complex carbohydrate drugs that bind galectin-3, GM-CT-01 and GR-MD-02, to treat NASH with fibrosis in a murine model. GR-MD-02 treatment resulted in marked improvement in liver histology with significant reduction in NASH activity and collagen deposition. Treatments seemed also to improve both glomerulopathy and interstitial fibrosis observed in kidneys. The improvement in liver histology was evident when animals were treated early in disease or after establishment of liver fibrosis. In all measures, GM-CT-01 had an intermediate effect between vehicle and GR-MD-02. Galectin-3 protein expression was increased in NASH with highest expression in macrophages surrounding lipid laden hepatocytes, and reduced following treatment with GR-MD-02, while the number of macrophages was unchanged. Treatment with GR-MD-02 also reduced the expression of pathological indicators including iNOS, an important TH1 inflammatory mediator, CD36, a scavenger receptor for lipoproteins on macrophages, and a-smooth muscle actin, a marker for activated stellate cells which are the primary collagen producing cells in liver fibrosis. We conclude that treatment with these galectin-3 targeting drugs improved histopathological findings of NASH and markedly reduced fibrosis in a murine model of NASH. While the mechanisms require further investigation, the treatment effect is associated with a reduction of galectin-3 expressed by activated macrophages which was associated with regression of NASH, including hepatocellular fat accumulation, hepatocyte ballooning, intra-portal and intra-lobular inflammatory infiltrate, and deposition of collagen. Similar effects were found with GM-CT-01, but with approximately four-fold lower potency than GR-MD-02. The results, in combination with previous experiments in toxin-induced fibrosis, suggest that these galectin-targeting drugs may have potential in human NASH with fibrosis.
To occur the transition from normal epithelium to squamous cell carcinoma is a necessary a for multiple stages process, such as smoking and alcohol abuse and a possible association with HPV infection. Several types of molecular markers have been studied in cancer of larynx, including proteins associated with apoptosis (bcl-2 and PARP-1) and proteins involved in many biological process such as galectin-3. In this study, analyses of qualitative and quantitative immunohistochemistry was performed for bcl-2, PARP-1 and galectin-3 in 65 patients diagnosed with laryngeal squamous cell carcinoma divided into in situ laryngeal carcinomas(LSCC S ), laryngeal squamols cells carcinomas without metastases (LSCC WT ) and with metastasis (LSCC W ) and cervical lymph nodes (CL). HPV detection and typing was performed by PCR and the HPV types evaluated were HPV 6, 11, 16, 18, 31 and 33. In quantitative of galectin-3 there was observed a significant increase of expression in invasive laryngeal squamous cell carcinoma (LSCC WT and LSCC W ) compared with in situ laryngeal carcinomas (LSCC S ), may indicating that this protein could be a good marker for progression of laryngeal carcinoma. For PARP-1 and bcl-2 protein there was no difference in the levels of expression in all groups studied. In qualitative analysis PARP-1 showed a homogenous immunolabeling in both high and low among the groups. In relation to Galectin-3, it was observed a predominance of cases with high expression, unlike the protein bcl-2 where the expression prevalence was low in all cases of laryngeal carcinoma and their metastatic lymph nodes. Of the 65 patients, 55 (84.6%) were positive for beta-globin and 7 (12.7%) of 55 patients were positive for HPV. Because of a low incidence of HPV in the cases studied, it was not possible correlate the proteins bcl-2, PARP-1 and Galectin-3 with the presence of HPV.
Their importance depends on the context of use, because the purpose for which is used has an influence in its required sensitivity, specificity and analytical details. Bogoslovsky et al  identified qui te a few contexts of use: “identify patients who may require acute neuroimaging (CT or MRI); select patients at risk for secondary brain injury processes (e.g., increase of intrac- erebral pressure, hemorrhage growth, expansion of cerebral edema, ischemia or neuroinflamma- tion); aid in counseling with symptoms provided at discharge; identify patients who are at risk for developing postconcussive syndrome, posttraumatic epilepsy or chronic traumatic encephalopa- thy; predict outcome with respect to poor or good recovery; and inform decisions regarding when to return to work or to play.” Additionally, the presence of copeptin, galectin-3, MMP-9 and occludin allowed discrimination between injury groups and healthy controls .
proteins with high sequence homologies but opposite effects on cell survival (Figure 1). As clearly shown in the Figure, Bcl-2 and Bcl-X L are negative regulators of apoptosis, whereas Bax, Bad, and Bak contribute to a positive signal in the regulation of cell death (77). Transfection of galectin-3 cDNA into leukemia T cells conferred resistance to ap- optosis induced by Fas ligation and stauro- sporine (76). Of particular interest, galectin- 3 showed a significant sequence similarity to Bcl-2, mainly concentrated in the functional BH1 (NWGR) domain. Although this para- digm between galectins-1 and -3 seems to be attractive, it should not be conceived as a general principle. Future studies should chal- lenge this paradigm in the context of other physiological systems. Consistently, expres- sion of galectins-1 and -3 was found to be differentially regulated during mammalian gestation (43,44). Iglesias et al. (44) reported the coexistence of mitogenic galectin-3 and apoptotic galectin-1 (43) in ovine placenta. Interestingly, galectin-3 expression was found to be decreased in term ovine pla- centa, in comparison with the middle of the gestation period. In contrast, no significant decrease was observed in galectin-1 expres- sion. In this context, one might speculate that differential expression of both ß-galac- toside-binding lectins in ovine placenta could be associated with selective requirements at different developmental stages of gestation. Furthermore, the participation of other members of the galectin family in the control of programmed cell death should also be considered. In this sense, galectin-9, a novel
Gastric cancer remains the fourth most commonly diagnosed cancer and is the second leading cause of cancer related deaths worldwide . Galectins are a group of proteins that bind b- galactosides through evolutionarily conserved sequence elements of the carbohydrate recognition domain (CRD)[2,3]. The galectin family contributes to neoplastic transformation, tumor cell survival, tissue invasion and metastasis of several types of cancer, including gastric cancer . Previous studies showed that the expression level of galectin-3 was significantly higher in the gastric carcinomas compared to adjacent normal tissue. Galectin-3 and galectin-4 may be useful tumor markers for gastric cancers with respect to tumor progression and potentiality of lymph node metastasis especially in certain histological types of gastric cancer [5,6]. Galectin-9 (Gal-9) is a new member of the galectin protein family . In recent years, an important role has emerged for the
model. Even in this case the scalar potential involving dou- blets of the residual gauge symmetry does not coincide with the potential of the MSSM. In the present work, we have considered that the supersymmetry is broken at the same time as the 3-3-1 gauge symmetry. Hence, we have to con- sider the complete 3-3-1 scalar potential. It means that in the Duong and Ma supersymmetric model there are no doubly charged charginos and exotic charged squarks. ~b! In Ref. @34# it was assumed that some of the VEVs have zero value, unlikely we have considered all ~but s 2 8 0 and s 1 0 ) of them different from zero. Hence we are able to obtain realistic quark and charged lepton masses, as can be seen from Eqs. ~20!, ~21!, and ~22!. In Ref. @34# some of these masses have to be generated by radiative corrections @40#. From ~a! and ~b! we see that the supersymmetric 3-3-1 model considered in this work has different phenomenological features from the supersymmetric 3-3-1 model of Duong and Ma.
In spite of significant advances in elucidating the role of galectin-1 (Gal-1) within models of autoimmune conditions, focusing on T- and B- cells [14-16], the effects of this protein toward the cells of innate immunity have not been stud- ied in such detail. We began the present study by validating this effect using a classical model of acute inflammation induced by zymosan in mice [17, 18]. Furthermore, the pharmacologi- cal treatment with human recombinant (hr) Gal- 1 was investigated in innate immune cell re- cruitment, particularly polymorphonuclear leu- kocytes (PMNs) and mononuclear phagocytic cells, and in the expression of the adhesion molecules ơ2-integrin (CD11b) and L-selectin (CD62L) during the process of PMN transmigration in the mouse mesentery.
Os espectros de excitação dos sólidos foram obtidos à tempe- ratura ambiente e são apresentados nas Figuras 4 e 5, independente da concentração dos íons ativadores e sensibilizadores, observou-se considerável predominância da BTC em menores comprimento de onda entre 275 à 375 nm, em relação às bandas relativas as transi- ções f-f do íon Eu 3+ . Os espectros apresentam perfis semelhantes