A2G Is a New System of α-2-Globulin Allotypes in Pig Blood Serum

Combinations of four -2-globulin allotypes were studied for their distribution in pigs of nine different breeds and hybrid groups. Based on this analysis, a new, previously unpublished polyallelic genetic system designated A2G was postulated. The complex alleles of this system control -2-globulin allotypes and are suggested to be encoded by genes of two subloci. One of these subloci is virtually monomorphic, whereas the other has at least four allelic variants.     

Serum TGF-β1 as a Biomarker for Type 2 Diabetic Nephropathy: A Meta-Analysis of Randomized Controlled Trials

Background

Abnormal expression of serum TGF-β1 was found in patients with diabetic nephropathy. However, the association of TGF-β1 with the risk of diabetic nephropathy remains unknown. The present study was undertaken to investigate whether such an association exists.

Methods

We searched the Chinese VIP, Wangfang, China National Knowledge Infrastructure, PubMed, Embase, and Google Scholar databases for relevant studies and extracted all eligible data. Stata12 software was used for statistical analysis.

Results

Nine reports met our criteria and were used for data extraction. There were 264 patients and 227 healthy controls from qualified reports in this meta-analysis. The results suggested that serum TGF-β1 levels were significantly up-regulated in patients with diabetic nephropathy; the instrumental variable was 3.94 (95% confidence interval 3.20–4.68, p<0.01).

Conclusions

Meta-analysis suggested that elevated serum TGF-β level in patients with diabetes is associated with a high risk of nephropathy. Further studies are required to validate these observations.     

Evaluation of Serum and Tissue Levels of α-Tocopherol

This study was designed to test the effect of supplementation of several antioxidants, including α-tocopherol, on the clinical reduction of premalignant oral lesions. Samples of oral mucosa and serum were taken from baseline to 9 months of supplementation from patients with premalignant oral lesions and analyzed for α-tocopherol by HPLC. Statistical increases in both serum and tissue α-tocopherol were found after supplementation. There was no statistical relationship between α-tocopherol and β-carotene levels.     

The Binding of Benzopyranes to Human Serum Albumin. A Structure–Affinity Study

The binding of several benzopyranes to serum albumin was studied by equilibrium dialysis at pH7.4 in a 67mM sodium phosphate buffer at 37°C. The equilibrium data were analyzed using a computer program for curve fitting. The binding isotherm for warfarin, 4-hydroxycoumarin, 4-chromanol, coumarin, 3-acetylcoumarin, and benzoic acid can be described by two stoichiometric dissociation constants. Elimination of the 4-hydroxyl group in the coumarin chemical structures decreases the binding affinity of the compounds on the primary binding site of serum albumin, with 4-chromanol the smallest ligand which binds to seroalbumin with high affinity. Thus, the affinity of 4-benzopyranol and the 4-hydroxybenzopyranones greater than that of benzopyranones. On the other hand, elimination of the 2-oxo group in the benzopyranone chemical structures decreases affinity for the secondary binding site.     

Glycoproteogenomics: A Frequent Gene Polymorphism Affects the Glycosylation Pattern of the Human Serum Fetuin/α-2-HS-Glycoprotein

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Highlights

• •Frequent genetic polymorphism affects the glycosylation pattern of fetuin.
• •Personalized in-depth proteoform profiling of fetuin purified from 20 donors.
• •Classification of serum donors into three different genotypes.
• •Septic patients show increased level of fucosylation at N-glycolation site N176.

     

On the existence of a possible A2A-D2-β-Arrestin2 complex: A2A agonist modulation of D2 agonist-induced β-arrestin2 recruitment

Given that coactivation of adenosine A(2A) (A(2A)R) and dopamine D(2) (D(2)R) receptors results in the coaggregation, cointernalization, and codesensitization of the A(2A)R and D(2)R and the role of scaffolding protein β-arrestin2 in the desensitization, internalization, and signaling of G-protein-coupled receptors, in this study we explored the ability of the A(2A)R agonist CGS21680 in A(2A)R-D(2)R-coexpressing cells to modulate the D(2)R agonist-induced recruitment of β-arrestin2 to the D(2)R by means of proximity-based bioluminescence resonance energy transfer (BRET(2)) and co-trafficking analysis. We found evidence that CGS21680 can increase the maximal BRET(2) signal between β-arrestin2(RLuc) and D(2L)R(GFP2) upon D(2)R activation, by increasing the potency of the D(2)R agonist to exert this action. In addition, this change was associated with an increased formation of cytoplasmic clusters containing β-arrestin2(GFP2) and D(2L)R(YFP) as seen from the co-trafficking analysis. Furthermore, the A(2A)R agonist advanced the time for the increase in Akt phosphorylation obtained with the D(2)R agonist. Finally, using a novel bioinformatics approach to predict the protein-protein interface, we have also found that amino acid pro-triplets TNY, LLS, RAF, and VSR may be crucial for the -induced β-arrestin2 recruitment by A(2A)R-D(2)R heteromers. Taken together, the results indicate that the antagonistic A(2A)R-D(2)R allosteric receptor-receptor interaction in A(2A)R-D(2)R heteromers favors β-arrestin2 recruitment to the D(2L)R protomer with subsequent cointernalization associated with a reduced time onset of Akt phosphorylation followed by a rapid dephosphorylation. Thus, β-arrestin2 action becomes more rapid and short-lasting and, in this way, mimics G-protein-mediated signaling.     

Sirtuin1: A Promising Serum Protein Marker for Early Detection of Alzheimer’s Disease

Sirtuin (SIRT) pathway has a crucial role in Alzheimer’s disease (AD). The present study evaluated the alterations in serum sirtuin1 (SIRT1) concentration in healthy individuals (young and old) and patients with AD and mild cognitive impairment (MCI). Blood samples were collected from 40 AD and 9 MCI patients as cases and 22 young healthy adults and 22 healthy elderly individuals as controls. Serum SIRT1 was estimated by Surface Plasmon Resonance (SPR), Western Blot and Enzyme Linked Immunosorbent Assay (ELISA). A significant (p<0.0001) decline in SIRT1 concentration was observed in patients with AD (2.27±0.46 ng/µl) and MCI (3.64±0.15 ng/µl) compared to healthy elderly individuals (4.82±0.4 ng/µl). The serum SIRT1 concentration in healthy elderly was also significantly lower (p<0.0001) compared to young healthy controls (8.16±0.87 ng/µl). This study, first of its kind, has demonstrated, decline in serum concentration of SIRT1 in healthy individuals as they age. In patients with AD and MCI the decline was even more pronounced, which provides an opportunity to develop this protein as a predictive marker of AD in early stages with suitable cut off values.     

Serum Antibodies against CD28– A New Potential Marker of Dismal Prognosis in Melanoma Patients

Background

Autoantibodies against CD28 have been found in patients with autoimmune and atopic diseases. These antibodies may act as superagonists and activate T cells but may also be antagonistic or induce immunosuppressive effects by activating regulatory T cells. Autoimmunity in melanoma patients has been discussed controversially.

Objective

We investigated 230 melanoma patients for the occurrence of CD28 antibodies and the effect of the latter on overall and progress-free survival.

Methods

We constructed an ELISA assay to measure CD28 serum antibodies. 230 patients with melanoma and a control-group of 625 patients consistent of 212 patients with virus hepatitis b or c, 149 patients with allergies, 78 patients with psoriasis, 46 patients with plasmocytoma and 140 healthy blood donors were investigated for the occurrence of CD28 antibodies.

Results

CD28 abs occur at a higher percentage in patients with melanoma and in patients with viral hepatitis than in other groups investigated (p<0.001). Occurrence of CD28 abs is significantly higher in patients receiving interferons independent from the underlying disease (p<0.001). In vitro CD28 serum antibodies have an inhibitory effect on the CD28 receptor as they lead to reduced stimulation of Jurkat cells. Presence of CD28 was correlated with a higher risk of dying from melanoma (p = 0.043), but not with a significantly shortened overall survival or progression-free survival.

Conclusion

Interferon therapy appears to induce the production of CD28 abs. In light of reports that these CD28 abs induce immunosuppressive Tregs and – as our data show – that they are inhibitors of CD28 receptor mediated stimulation, the continuation of therapies with interferons in melanoma patients developing CD28 antibodies should be critically reconsidered, since our data indicate a worse outcome of patients with CD28 abs.     

Detection of RASSF1A and RAR? Hypermethylation in Serum DNA from Breast Cancer Patients

Breast cancer is fast emerging as the leading cancer amongst females, especially in young females in metropolitan cities in India. The epigenetic alterations involved in the onset and progression of breast cancer may serve as biomarkers for early detection and prognosis of the disease. Furthermore, using body fluids such as serum offers a non-invasive method to procure multiple samples for such analyses. In this study, we examined methylation status of two normally unmethylated but biologically significant cancer genes, RAS association domain family protein 1A (RASSF1A) and Retionic acid receptor ? (RAR?) by Methylation Specific PCR (MSP) in invasive ductal carcinomas of the breast and paired serum DNA. RASSF1A was found to be methylated in 17 of 20 (85%) breast tumors; while sera from 15 of 20 (75%) of the patients showed concordant methylated RASSF1A, with a sensitivity of 88%. RAR? was methylated in 2/20 (10%) breast tumors. A gene unmethylated in the tumor DNA was always found to be unmethylated in the matched serum DNA for both RASSF1A and RAR? genes; hence specificity was 100%. Immunohistochemical analysis of RAR? protein in 15 breast carcinoma patients harboring unmethylated RAR? in tumors and serum DNA showed the expression of RAR? protein in tumors and paired normal breast tissues, confirming the MSP findings, suggesting that RAR? promoter is functional in these cases. This study underscores the potential utility of DNA methylation based screening of serum, a readily accessible body fluid, as a surrogate marker for early detection of breast cancer.      

Serum Bile Acids in Repaired Tetralogy of Fallot: A Marker for Liver and Heart?

Background and Aims

Patients with repaired tetralogy of Fallot may develop chronic right ventricular dysfunction and hepatic congestion over time. We hypothesized that bile acid metabolism is altered in repaired tetralogy of Fallot patients and therefore sought to correlate right ventricular indices with serum bile acid levels.

Methods

Indexed right ventricular end diastolic volume, as assessed by cardiac magnetic-resonance imaging, was classified as <100ml/m² (Group 1, n = 5), 100–150ml/m² (Group 2, n = 18), and >150ml/m² (Group 3, n = 6) in 29 patients with repaired tetralogy of Fallot. Pulmonary regurgitation fraction and right ventricular ejection fraction were calculated. The serum bile acid profile, including 15 species, in these patients was determined by liquid chromatography coupled with mass spectrometry.

Results

Serum bile acid levels increased from Group 1 to Group 3 (2.5 ± 0.7; 4.1 ± 2.5; 6.0 ± 2.8 μmol/l, respectively) with significantly increased bile acid values in Group 3 compared to Group 1 (p≤0.05). In Group 3, but not in Group 1 and 2, a significant increase in glycine-conjugated bile acids was observed. Pulmonary regurgitation fraction increased (12 ± 1; 28 ± 16; 43 ± 3%, Groups 1–3, respectively) and right ventricular ejection fraction decreased (48.4 ± 6.4; 48.5 ± 6.5; 42.1 ± 5.3%, Groups 1–3, respectively) with rising indexed right ventricular end diastolic volume.

Conclusions

These preliminary results suggest that serum bile acid levels are positively correlated with indexed right ventricular end-diastolic volume in patients with repaired tetralogy of Fallot; however, this needs to be confirmed in a larger patient cohort.     

Development of a Highly Sensitive Immuno-PCR Assay for the Measurement of α-Galactosidase A Protein Levels in Serum and Plasma

Fabry disease is an X-linked genetic disorder caused by defects in the α-galactosidase A (GLA) gene, and heterogeneous mutations lead to quantitative and/or qualitative defects in GLA protein in male patients with Fabry disease. Random X-chromosomal inactivation modifies the clinical and biochemical features of female patients with Fabry disease. Functional polymorphisms have been frequently reported in recent times, and these increase the difficulty of understanding the pathogenetic basis of the disease. To date, GLA protein level has been measured using an enzyme-linked immunosorbent assay (ELISA). However, ELISA is not highly sensitive due to the high background noise. In this paper, we introduce a novel application of the immuno-polymerase chain reaction (PCR) method (termed Multiple Simultaneous Tag [MUSTag]) for measurement of the GLA protein level in blood samples. We compared the sensitivities of the MUSTag method with plates or magnetic beads with those of ELISA for recombinant human GLA and found that the apparent maximal sensitivity was higher for the former than for the latter. We then measured the GLA concentrations in serum and plasma from male patients with classic Fabry disease (Male Fabry), females with Fabry disease (Female Fabry), male subjects harboring the functional polymorphism p.E66Q (E66Q), and control (Control) subjects. Our results revealed that compared to the MUSTag plate and ELISA, the MUSTag beads assay afforded a clearer estimation of the GLA protein levels in the serum and plasma with minimal or no background noise, although all the methods could differentiate between the Male Fabry, E66Q, and Control groups. The Female Fabry group showed characteristic heterogeneity, which was consistent with the X-linked inheritance. This novel method is expected to be useful for the sensitive determination of GLA level in blood and elucidation of the pathogenetic basis of Fabry disease.     

Serum selenium predicts levels of F2-isoprostanes and prostaglandin F2α in a 27 year follow-up study of Swedish men

Low concentrations of selenium (Se) predict mortality and cardiovascular diseases in some populations. The effect of Se on in vivo indicators of oxidative stress and inflammation, two important features of atherosclerosis, in human populations is largely unexplored. This study investigated the longitudinal association between serum selenium (s–Se) and a golden standard indicator of oxidative stress in vivo (8-iso-prostaglandin F_2α, a major F₂-isoprostane), an indicator of cyclooxygenase (COX)-mediated inflammation (prostaglandin F_2α), high sensitive C-reactive protein (hsCRP), interleukin-6 (IL-6) and serum amyloid A protein (SAA) in a follow-up study of 27 years. The s–Se was measured in 615 Swedish men at 50 years of age in a health investigation. The status of oxidative stress and inflammation was evaluated in a re-investigation 27 years later by quantification of urinary 8-iso-PGF_2α and 15-keto-dihydro- PGF_2α (a major metabolite of PGF_2α) and serum hsCRP, SAA and IL-6. Men in the highest quartile of s–Se at age 50 had decreased levels of 8-iso-PGF_2α compared to all lower quartiles and decreased levels of PGF_2α compared to all lower quartiles at follow-up. These associations were independent of BMI, diabetes, hyperlipidemia, hypertension, smoking, α-tocopherol and β-carotene at baseline. The s–Se was not associated with hsCRP, SAA or IL-6 at follow-up. In conclusion, high concentrations of s–Se predict reduced levels of oxidative stress and subclinical COX-mediated (but not cytokine-mediated) inflammation in a male population. The associations between Se, oxidative stress and inflammation, respectively, might be related to the proposed cardiovascular protective property of Se.     

Reassessing the Detection of B-Virus–Specific Serum Antibodies

B virus, a natural pathogen of macaques, can cause a fatal zoonotic disease in humans. Serologic screening of macaques by titration ELISA (tELISA, screening test) and by Western blot analysis (WBA, confirmatory test) is one of the principle measures to prevent human infection. Here we slightly modified these 2 tests and reevaluated their correlation. We developed a high-throughput tELISA and used it to screen 278 sera simultaneously against the homologous BV antigen and the heterologous antigens of Papiine herpesvirus 2 and Human herpesvirus 1. More sera (35.6%) were positive by the BV-ELISA than by the HVP2-ELISA (21.6%) or HSV1-ELISA (19.8%). The superiority of the homologous tELISA over the heterologous tELISA was prominent in low-titer sera. WBA confirmed only 21% of the tELISA-positive sera with low or intermediate antibody titers. These sera might have contained antibodies to conformational epitopes that could not be detected by WBA, in which denatured antigens are used, but that could be detected by tELISA, which detects both linear and conformational epitopes. WBA confirmed 82% of the tELISA high-titer sera. However, WBA defined the remaining 18% of sera, which were negative by tELISA, as nonnegative. This finding can be attributed to the difficulties encountered with the subjective interpretation of results by WBA. Together, the current results indicate the inadequacy of WBA as a confirmatory assay for sera with low antibody titers.Abbreviations: BV, B virus (Macacine herpesvirus 1); CLIA, Clinical Laboratory Improvement Amendments; EU, ELISA units; HSV, Human herpesvirus; HVL, langur herpesvirus; HVM, Herpesvirus managabey; HVP, Papiine herpesvirus; SA8, simian agent 8; tELISA, titration ELISA; UN, uninfected; WBA, Western blot analysisB virus (BV; Macacine herpesvirus 1), which is endemic in all species of macaques (natural hosts), is a member of the genus Simplexvirus, subfamily Alphaherpesvirinae and family Herpesviridae. Alphaherpesviruses are characterized by the ability to establish a neurotropic, generally asymptomatic, infection in their natural hosts. Macaques spread BV within a group by contact with macaques that are shedding virus during an acute or intermittently reactivated infection. BV is closely related to 2 well-characterized human alphaherpesviruses, Human herpesvirus (HSV) types 1 and 2, to simian agent 8 (SA8; Cercopithecine herpesvirus 2), which is endemic in African green monkeys (Cercopithecus aethiops), and to 2 recently identified simian alphaherpesviruses, HVP2 (Papiine herpesvirus 2) in baboons (Papio spp.) and langur herpesvirus (HVL), which is endemic in langur monkeys (Presbytis spp.)^5,6,7 and which has not been officially classified or named.⁹ Recently, sera from a group of sooty mangabey monkeys (Cercocebus atys) were found to crossreact serologically with other simian simplexviruses.^6,8,11 It was assumed that crossreactive antibodies were induced by a putative alphaherpesvirus that is endemic in mangabeys. This virus was provisionally named as Herpesvirus managabey (HVM) and is pending taxonomic evaluation.Each of the simplex viruses has remarkable host specificity in nature. However, cross-species infections with BV have been reported. BV is the only nonhuman primate alphaherpesvirus that infects humans. When it does so, BV causes an often-fatal zoonotic disease in 80% of untreated humans.^{7,10,21,23,32,33}BV is transmitted through bites, scratches, or contact with infected oral or genital body fluids. In addition, the virus can be transmitted via fomites and from human to human through contact with contaminated wounds. Virus replication occurs in epithelial or fibroblast cells at the epidermal or dermal site of virus entry; however, BV also enters the peripheral nervous system via axons without replicating locally in surrounding epithelial cells, as has been reported for other simplex viruses.^20,23 Once BV enters peripheral nerves, life-long latency is established in the dorsal root of spinal ganglia or cranial ganglia of infected hosts. BV undergoes periodic reactivation in macaques as well as in humans that survive this zoonosis. In both macaques and humans, BV can be reactivated in the ganglia, generally resulting in anterograde travel of the virus and replication at the original site of infection.^10,33 This event results in virus shedding from infected cells, an event that can be detected by PCR or virus isolation if samples are collected during this event. However, because virus shedding is unpredictable and sporadic, identification of BV infection by means of PCR or virus isolation is rare. A more practical approach to identifying infected macaques or humans is the use of serologic methods for identifying antibodies specific for BV, although the shortcomings of this approach are appreciated when screening sera from subjects that are in the midst of a primary infection but have not yet produced detectable levels of antibodies or from BV-infected subjects that lack detectable antibody because of waning levels or anergy.Because of the high lethality of BV to humans and life-long infection in survivors that lack effective strategies to clear this virus, BV antigen is produced under BSL4 conditions according to federal guidelines and under strict biosecurity regulations.³ Many laboratories in the United States, Europe, and Asia cannot produce BV antigens because of these restrictions and therefore use alternative crossreacting (heterologous) herpesvirus antigens such as HVP2 and HSV1 for the detection of antibodies to BV.^{10,19,26,29,34} Our previous studies¹⁶ indicated that using heterologous viruses in serologic assays for detecting BV antibodies contributes to increased false-negative results.Serologic diagnosis of BV infection in macaques at the National B Virus Resource Laboratory has been based on 2 principal tests that meet standards proscribed by the Clinical Laboratory Improvement Amendments:⁴ the titration ELISA (tELISA) and Western blot analysis (WBA).^15,31 tELISA detects antibody in sera from most BV-infected macaques by using the complex mixture of BV antigens that is present in lysates from infected cells and adsorbed onto polystyrene microtiter plates. These infected-cell lysates are prepared by using nondenaturing detergents. Quality-control assessment of each antigen lot is performed, including determinations of protein concentration, antigen mass, immunoreactivity with macaque serum pools, and reactivity in WBA assays.WBA was chosen as the confirmatory test for the BV-screening tELISA; in addition, WBA serves as the primary screening assay for detecting antibodies to BV, HSV1, and HSV2 in human sera collected after exposure in the workplace.^27,31 The relationship between ELISA and WBA is complex, and therefore the agreement between the tests may be lower than expected given that WBA detects only conformationally independent epitopes, whereas ELISA identifies both linear and conformational epitopes (unless the epitopes are cryptic).²⁸ In addition, the different antibodies detected by each test might be induced at distinctly different time points after infection, so that what one assay detects, the other may not.^2,8,9 Despite the difficulties encountered and due to the lack of a better alternative, both assays are required to maximize the likelihood of correctly identifying whether BV antibodies are present in a sample.Because the identification of BV-infected macaques in any colony, especially SPF colonies, is of great medical and economic significance, assays should be designed to identify low levels of antibody as soon as possible after infection.^12,31 For BV, as for other alphaherpesviruses, the presence of serum antibodies likely indicates the presence of virus that is either in an active lytic state (and actively replicating) or in an inactive, latent state. It is therefore important to have both a sensitive assay, to prevent missing low-responders, and a highly specific assay, to avoid obtaining false-negative results that can lead to costly veterinary and medical decisions.In our continuing efforts to improve the accuracy of the tELISA for BV, we recently modified and automated it.¹⁸ This process included validation experiments that enabled us to reexamine some features of the BV serologic diagnosis.Here we describe additional modifications of the tELISA and its adaptation to an automated, high-throughput 384-well format. This format enabled us to assess the sensitivities of tELISA using homologous compared with heterologous antigens in the same run. In addition, we evaluated the correlation between tELISA and WBA and discuss the effectiveness of WBA as a confirmatory test for tELISA. All assays were developed and performed inhouse in accordance with CLIA regulations,⁴ because each test either was used or could be used for identification of BV antibodies in human sera as well as macaque sera. Although CLIA does not regulate veterinary assays, we feel strongly that the tests described are superior because of the strict quality control that must be maintained for testing human samples.     

Rapid Immunochromatographic Detection of Serum Anti-α-Galactosidase A Antibodies in Fabry Patients after Enzyme Replacement Therapy

We developed an immunochromatography-based assay for detecting antibodies against recombinant α-galactosidase A proteins in serum. The evaluation of 29 serum samples from Fabry patients, who had received enzyme replacement therapy with agalsidase alpha and/or agalsidase beta, was performed by means of this assay method, and the results clearly revealed that the patients exhibited the same level of antibodies against both agalsidase alpha and agalsidase beta, regardless of the species of recombinant α-galactosidase A used for enzyme replacement therapy. A conventional enzyme-linked immunosorbent assay supported the results. Considering these, enzyme replacement therapy with agalsidase alpha or agalsidase beta would generate antibodies against the common epitopes in both agalsidase alpha and agalsidase beta. Most of the patients who showed immunopositive reaction exhibited classic Fabry phenotype and harbored gene mutations affecting biosynthesis of α-galactosidase A. As immunochromatography is a handy and simple assay system which can be available at bedside, this assay method would be extremely useful for quick evaluation or first screening of serum antibodies against agalsidase alpha or agalsidase beta in Fabry disease with enzyme replacement therapy.     

A novel synthesis of 2-deoxy-α-glycosides

The key step of the synthesis involves the reaction of glycals [3,4,6-tri-O-acetyl-d-glucal (1), the new glycal derivative 4-O-acetyl-1,5-anhydro-2,6-dideoxy-3-C-methyl-3-O-methyl-l-ribo-hex-l-enitol (2), and 3-acetamido-4,6,-di-O-acetyl-1,5-anhydro-2,3-dideoxy-d-arabino-hex-l-enitol (3)] with 1.5 molar equivalents of several alcohols in the presence ofN-bromosuccinimide in acetonitrile to give mainly the corresponding 2-bromo-2-deoxy-α-glycopyranosides (4-21). The glycopyranosides (4-8 and16-21) from1 and3 have the α-d-manno configuration and those (10-15) from2 have the α-l-altro configuration. The yields are high from1, virtually quantitative from2, and moderate from3. Debromination of the 2-bromo-2-deoxy compounds with tributylstannane and a radical initiator gives the corresponding 2-deoxy-α-glycopyranosides (22-38) in quantitative yields. In particular, the branched-chain glycal2 reacts with alcohols to give exclusively the corresponding α-glycopyranosides (27-32) of cladinose in strikingly high overall yields. The stereoselectivity and regiospecificity of the bromination reaction are described. 1,3-Dibromo-5,5-dimethylhydantoin andN-bromoacetamide are also found to be useful for the reaction.     

The Down-Regulation of Neuroligin-2 and the Correlative Clinical Significance of Serum GABA Over-Expression in Hirschsprung’s Disease

The goal of this study was to investigate the expression level of neuroligin-2 in different colon tissue segments of children with Hirschsprung’s disease (HSCR) and the correlative clinical significance of serum Gamma-Aminobutyric Acid (serum GABA) in HSCR. Neuroligin-2 was assessed by Immunohistochemistry staining method on routine paraffin section from different colon tissue segments of HSCR (ganglionic colonic segment, transitional colonic segment and aganglionic colonic segment). Western-blot analysis and real-time fluorescence quantitative PCR(qRT-PCR) were applied to compare and evaluate the expression levels of neuroligin-2 from three segments of HSCR, and we used Enzyme-linked Immunosorbent Assay (ELISA) method to detect and compare the serum GABA between HSCR and non-HSCR. Immunohistochemistry staining demonstrated that intensive neuroligin-2 staining was detected in the ganglion cells in the ganglionic colonic and transitional colonic segments from the HSCR children; however, neuroligin-2 staining was down-regulated significantly in the aganglionic colonic segments. The expression levels of neuroligin-2 mRNA and protein in the aganglionic colonic segment were decreased compared to the ganglionic colonic segment and transitional colonic segment (P < 0.05). And the level of serum GABA was significantly higher in HSCR than that in non-HSCR. The expression of neuroligin-2 varies from different segments of HSCR. The down-regulation of neuroligin-2 in aganglionic colonic segments may be correlated with the excessive intestine contraction and further result in HSCR. The over-expression of serum GABA may be considered as a new diagnostic method of HSCR.     

Serum amyloid A and inflammasome activation: A link to breast cancer progression?

Breast cancer is the most frequently diagnosed cancer in women globally. Although there have been many significant advances made in the diagnosis and treatment of breast cancer, numerous unresolved challenges remain, which include prevention, early diagnosis, metastasis and recurrence. The role of inflammation in cancer development is well established and is believed to be one of the leading hallmarks of cancer progression. Recently, the role of the inflammasome, a cytosolic multiprotein complex, has received attention in different cancers. By contributing to the activation of inflammatory cytokines the inflammasome intensifies the inflammatory cascade. The inflammasome can be activated through several pathways, which include the binding of pattern associated molecular patterns (PAMPs) and damage associated molecular patterns (DAMPs) to toll-like receptors (TLRs). Serum amyloid A (SAA), a non-specific acute-phase protein, can function as an endogenous DAMP by binding to pattern recognition receptors like TLRs on both breast cancer cells and cancer associated fibroblasts (CAFs). SAA can thus stimulate the production of IL-1β, thereby creating a favourable inflammatory environment to support tumour growth. The aim of this review is to highlight the possible role of SAA as an endogenous DAMP in the tumour microenvironment (TME) thereby promoting breast cancer growth through the activation of the NLRP3 inflammasome.