Immunohistochemical study of the canonical and non-canonical Wnt signaling pathway in colorectal carcinoma and non-neoplastic mucosa / Estudo imunoistoquímico da via de sinalização canônica e não canônica da proteína Wnt em carcinoma colorretal e em mucosa não neoplásica

Colorectal cancer is linked to several signaling pathways such as Wnt pathway. Our objective is to detect and verify the integrity of protein members of Wnt signaling pathway in colorectal carcinoma and non-neoplastic colorectal tissue. Sixty-four patients with colorectal carcinoma provided samples of colorectal neoplasia and non-neoplastic tissues, which were prepared in tissue microarray blocks and subjected to immunohistochemical analysis. The primary antibodies used were Wnt-1, Wnt-2, Wnt-5a Frizzled-1, Frizzled-5 and axin. Immunoexpression of Wnt-2 protein was significantly lower in colorectal tumor tissue and axin protein immunoexpression was significantly higher in tumor tissue. There was no significant difference in the expression of Wnt-1, Wnt-5a, Frizzled-1 and Frizzled-5 proteins in both tissues. The higher expression of Wnt-2 protein in non-neoplastic colorectal tis- sue suggests the participation during the hyperproliferative stage of colorectal mucosa. The increased axin protein immunoexpression in colorectal tumor suggests a decrease in the formation of the beta-catenin destructor complex. (AU)

O câncer colorretal está ligado a várias vias de sinalização, como a via Wnt. Nosso objetivo é detectar e verificar a integridade das proteínas da via de sinalização Wnt no carcinoma colorretal e no tecido colorretal não neoplásico. Sessenta e quatro pacientes com carcinoma colorretal forneceram amostras de neoplasia e tecidos não neoplásicos, que foram colocadas em blocos de tissue microarray e submetidas à análise imuno-histoquímica. Os anticorpos primários utilizados foram Wnt-1, Wnt-2, Wnt-5a Frizzled-1, Frizzled-5 e axina. A imunoexpressão da proteína Wnt-2 foi significativamente menor no tecido tumoral e a imunoexpressão da proteína axina foi significativamente superior no tecido do tumor. Não houve diferença significativa na expressão de Wnt-1, Wnt-5a, frizzled-1 e nas proteínas Frizzled 1 e 5 em ambos os tecidos. A maior expressão de Wnt-2 da proteína no tecido colorretal não neoplásico sugere a participação desta proteína durante o estágio de hiperproliferação da mucosa colorretal. O aumento da imunoexpressão da proteína axina no tumor colorretal sugere uma diminuição na formação do complexo de destruição da proteína beta-catenina. (AU)

The relationship between frizzled 6 gene polymorphisms and neural tube defects in children of northern Han Chinese population / 中华神经科杂志

Objective To study the association of single nucleotide polymorphisms (SNPs) of the frizzled 6(FZD6) gene with neural tube defects(NTDs) in a northern Han Chinese population.Methods Three nonsynonymous SNPs in the FZD6 gene (rs827528,rs3808553,rs12549394) were examined.The SNPs were genotyped by polymerase chain reaction (PCR) and sequencing in 135 NTD patients and matched normal controls.The allele,genotype and haplotype frequencies were calculated and analyzed to examine the association between FZD6 SNPs and NTDs.Results Both T allele and TT genotype frequencies of the rs3808553 polymorphism in the NTDs group were significantly higher than those in the controls,and children with T allele and TT genotype were associated with increased risk of NTDs (OR =1.575,95％ CI 1.112-2.230,P =0.010 and OR =2.811,95％ CI 1.325-5.967,P =0.023 respectively).There were no significant differences among different genotypes or alleles in both rs827528 and rs12549394.Haplotypes AG-C and A-T-C were found associated with NTDs in the case-control study (OR =0.560,95％ CI 0.378-0.830,P=0.004 and OR=1.670,95％CI 1.126-2.475,P =0.011 respectively).Conclusions The rs3808553 polymorphism of FZD6 is obviously associated with NTDs in children of northern Han Chinese population.The TT genotype may increase the risk for NTDs.The rs827528 and rs12549394 polymorphisms of FZD6 may have no association with NTDs.

Non-association Between Polymorphisms of the Frizzled Receptor Genes and Bone Mineral Density in Postmenopausal Korean Women

We investigated the association between single nucleotide polymorphisms (SNPs) in the frizzled (FZD) genes in the Wnt signal pathway and circulating osteoprotegerin (OPG), soluble receptor activator of NF-kappaB ligand (sRANKL) levels, bone turnover markers, and bone mineral density (BMD) in postmenopausal women. The SNPs in the FZD1, FZD5, FZD6, FZD7, and FZD9 genes were analyzed by direct sequencing in 371 postmenopausal Korean women. Levels of serum OPG, sRANKL, osteocalcin, C-telopeptide of type I collagen, calcium, parathyroid hormone and calcitonin, and BMD at the lumbar spine and femoral neck were measured. The SNPs in the FZD1, FZD5, FZD7, and FZD9 genes, and in exon 2 of the FZD6 gene were not observed. No significant differences in the adjusted BMD of lumbar spine and femoral neck and serum levels of OPG, sRANKL, and bone markers were noted among the single or haplotype genotypes of the L345M and E664A SNPs in the FZD6 gene and the distributions of these single or haplotype genotypes were not different according to the bone mass status. In conclusion, the polymorphisms of the FZD genes are not associated with BMD of the lumbar spine and femoral neck, bone turnover markers, or circulating OPG-sRANKL in Korean women.

Non-association Between Polymorphisms of the Frizzled Receptor Genes and Bone Mineral Density in Postmenopausal Korean Women

We investigated the association between single nucleotide polymorphisms (SNPs) in the frizzled (FZD) genes in the Wnt signal pathway and circulating osteoprotegerin (OPG), soluble receptor activator of NF-kappaB ligand (sRANKL) levels, bone turnover markers, and bone mineral density (BMD) in postmenopausal women. The SNPs in the FZD1, FZD5, FZD6, FZD7, and FZD9 genes were analyzed by direct sequencing in 371 postmenopausal Korean women. Levels of serum OPG, sRANKL, osteocalcin, C-telopeptide of type I collagen, calcium, parathyroid hormone and calcitonin, and BMD at the lumbar spine and femoral neck were measured. The SNPs in the FZD1, FZD5, FZD7, and FZD9 genes, and in exon 2 of the FZD6 gene were not observed. No significant differences in the adjusted BMD of lumbar spine and femoral neck and serum levels of OPG, sRANKL, and bone markers were noted among the single or haplotype genotypes of the L345M and E664A SNPs in the FZD6 gene and the distributions of these single or haplotype genotypes were not different according to the bone mass status. In conclusion, the polymorphisms of the FZD genes are not associated with BMD of the lumbar spine and femoral neck, bone turnover markers, or circulating OPG-sRANKL in Korean women.

Negative feedback regulation of Wnt signaling by Gbetagamma-mediated reduction of Dishevelled

Wnt signaling is known to be important for diverse embryonic and post-natal cellular events and be regulated by the proteins Dishevelled and Axin. Although Dishevelled is activated by Wnt and involved in signal transduction, it is not clear how Dishevelled-mediated signaling is turned off. We report that guanine nucleotide binding protein beta 2 (Gnb2; Gbeta2) bound to Axin and Gbeta2 inhibited Wnt mediated reporter activity. The inhibition involved reduction of the level of Dishevelled, and the Gbeta2gamma2 mediated reduction of Dishevelled was countered by increased expression of Axin. Consistent with these effects in HEK293T cells, injection of Gbeta2gamma2 into Xenopus embryos inhibited the formation of secondary axes induced either by XWnt8 or Dishevelled, but not by beta-catenin. The DEP domain of Dishevelled is necessary for both interaction with Gbeta2gamma2 and subsequent degradation of Dishevelled via the lysosomal pathway. Signaling induced by Gbeta2gamma2 is required because a mutant of Gbeta2, Gbeta2 (W332A) with lower signaling activity, had reduced ability to downregulate the level of Dishevelled. Activation of Wnt signaling by either of two methods, increased Frizzled signaling or transient transfection of Wnt, also led to increased degradation of Dishevelled and the induced Dishevelled loss is dependent on Gbeta1 and Gbeta2. Other studies with agents that interfere with PLC action and calcium signaling suggested that loss of Dishevelled is mediated through the following pathway: Wnt/Frizzled-->Gbetagamma-->PLC-->Ca+2/PKC signaling. Together the evidence suggests a novel negative feedback mechanism in which Gbeta2gamma2 inhibits Wnt signaling by degradation of Dishevelled.