Microvascular damage, neuroinflammation and extracellular matrix remodeling in Col18a1 knockout mice as a model for early cerebral small vessel disease.

Collagen type XVIII (COL18) is an abundant heparan sulfate proteoglycan in vascular basement membranes. Here, we asked (i) if the loss of COL18 would result in blood-brain barrier (BBB) breakdown, pathological alterations of small arteries and capillaries and neuroinflammation as found in cerebral small vessel disease (CSVD) and (ii) if such changes may be associated with remodeling of synapses and neural extracellular matrix (ECM). We found that 5-month-old Col18a1-/- mice had elevated BBB permeability for mouse IgG in the deep gray matter, and intravascular erythrocyte accumulations were observed brain-wide in capillaries and arterioles. BBB permeability increased with age and affected cortical regions and the hippocampus in 12-month-old Col18a1-/- mice. None of the Col18a1-/- mice displayed hallmarks of advanced CSVD, such as hemorrhages, and did not show perivascular space enlargement. Col18a1 deficiency-induced BBB leakage was accompanied by activation of microglia and astrocytes, a loss of aggrecan in the ECM of perineuronal nets associated with fast-spiking inhibitory interneurons and accumulation of the perisynaptic ECM proteoglycan brevican and the microglial complement protein C1q at excitatory synapses. As the pathway underlying these regulations, we found increased signaling through the TGF-ß1/Smad3/TIMP-3 cascade. We verified the pivotal role of COL18 for small vessel wall structure in CSVD by demonstrating the protein's involvement in vascular remodeling in autopsy brains from patients with cerebral hypertensive arteriopathy. Our study highlights an association between the alterations of perivascular ECM, extracellular proteolysis, and perineuronal/perisynaptic ECM, as a possible substrate of synaptic and cognitive alterations in CSVD.

COL18A1 as the Candidate Gene for the Prognostic Marker of Breast Cancer According to the Analysis of the DNA Copy Number Variation by Array CGH / 한국유방암학회지

PURPOSE: We tried to select and validate the candidate gene for the prognostic marker of breast cancer by comparing the analysis of copy number variation (CNV) between normal breast tissues and breast cancer tissues by performing array comparative genomic hybridization (CGH). METHODS: Array CGH was performed with using the fresh frozen tissues of 77 breast cancer patients. We selected the clones with more than a 20% frequency of gain or loss, and the clones with gain or loss in more than 2 consecutive clones. We finally selected the clones that were statistically significant on the survival analysis. We searched for the candidate gene that belonged to the candidate clones and we selected the final candidate gene that is assumed to be most related to the carcinogenesis of breast cancer by searching for information of the individual gene. We performed RT-PCR to validate the RNA expression of the final candidate gene with using the breast tissues of another 20 breast cancer patients. RESULTS: Eleven (10 in the gain group and 1 in the loss group) clones were finally selected as candidate clones. The significant CNVs with gain were found in the regions of 1q23.1, 1q41, 1q44, 5p15.33, 8q21.3, 15q26.3, 17q12 and 21q22.3 and the significant CNV with loss was found in 14q32.33. COL18A1 (21q22.3) was selected as the final candidate gene and the RT-PCR results revealed that the expression of COL18A1 was up-regulated in the cancer tissues of 18 of the other 20 (90%) breast cancer patients. CONCLUSION: We selected COL18A1 (21q22.3) as the candidate gene for the prognostic marker of breast cancer by comparing the analysis of CNVs from the array CGH. The RNA of COL18A1 was over-expressed in breast cancer tissue, as determined by RT-PCR.

COL18A1 as the Candidate Gene for the Prognostic Marker of Breast Cancer According to the Analysis of the DNA Copy Number Variation by Array CGH / 한국유방암학회지

PURPOSE: We tried to select and validate the candidate gene for the prognostic marker of breast cancer by comparing the analysis of copy number variation (CNV) between normal breast tissues and breast cancer tissues by performing array comparative genomic hybridization (CGH). METHODS: Array CGH was performed with using the fresh frozen tissues of 77 breast cancer patients. We selected the clones with more than a 20% frequency of gain or loss, and the clones with gain or loss in more than 2 consecutive clones. We finally selected the clones that were statistically significant on the survival analysis. We searched for the candidate gene that belonged to the candidate clones and we selected the final candidate gene that is assumed to be most related to the carcinogenesis of breast cancer by searching for information of the individual gene. We performed RT-PCR to validate the RNA expression of the final candidate gene with using the breast tissues of another 20 breast cancer patients. RESULTS: Eleven (10 in the gain group and 1 in the loss group) clones were finally selected as candidate clones. The significant CNVs with gain were found in the regions of 1q23.1, 1q41, 1q44, 5p15.33, 8q21.3, 15q26.3, 17q12 and 21q22.3 and the significant CNV with loss was found in 14q32.33. COL18A1 (21q22.3) was selected as the final candidate gene and the RT-PCR results revealed that the expression of COL18A1 was up-regulated in the cancer tissues of 18 of the other 20 (90%) breast cancer patients. CONCLUSION: We selected COL18A1 (21q22.3) as the candidate gene for the prognostic marker of breast cancer by comparing the analysis of CNVs from the array CGH. The RNA of COL18A1 was over-expressed in breast cancer tissue, as determined by RT-PCR.

Collagen XVIII/endostatin expression in experimental endotoxemic acute renal failure

Acute renal failure (ARF) is a frequent complication of Gram-negative sepsis, with a high risk of mortality. Lipopolysaccharide (LPS)-induced ARF is associated with hemodynamic changes that are strongly influenced by the overproduction of nitric oxide (NO) through the cytokine-mediated up-regulation of inducible NO synthase. LPS-induced reductions in systemic vascular resistance paradoxically culminate in renal vasoconstriction. Collagen XVIII is an important component of the extracellular matrix expressed in basement membranes. Its degradation by matrix metalloproteases, cathepsins and elastases results in the formation of endostatin, claimed to have antiangiogenic activity and to be a prominent vasorelaxing agent. We evaluated the expression of endostatin/collagen XVIII in an endotoxemic ARF model. ARF was induced in C57BL/6 mice by intraperitoneal injection of LPS (10 mg/kg) followed by sacrifice 4 and 12 h later. Kidney tissue was the source of RNA and protein and the subject of histological analysis. As early as 4 h after LPS administration, blood urea, creatinine and NO levels were significantly increased compared to control. Endostatin/collagen XVIII mRNA levels were 0.71 times lower than sham-inoculated mice 4 h after LPS inoculation, returning to normal levels 12 h after LPS inoculation. Immunohistological examination revealed that acute injury caused by LPS leads to an increase of endostatin basement membrane staining in association with the decrease of CD31 endothelial basement membrane staining. These results indicate that in the early phase of endotoxemic ARF the endostatin levels were not regulated by gene expression, but by the metabolism of collagen XVIII.

COL18A1 is highly expressed during human adipocyte differentiation and the SNP c.1136C > T in its "frizzled" motif is associated with obesity in diabetes type 2 patients

Collagen XVIII can generate two fragments, NC11-728 containing a frizzled motif which possibly acts in Wnt signaling and Endostatin, which is cleaved from the NC1 and is a potent inhibitor of angiogenesis. Collagen XVIII and Wnt signaling have recently been associated with adipogenic differentiation and obesity in some animal models, but not in humans. In the present report, we have shown that COL18A1 expression increases during human adipogenic differentiation. We also tested if polymorphisms in the Frizzled (c.1136C>T; Thr379Met) and Endostatin (c.4349G>A; Asp1437Asn) regions contribute towards susceptibility to obesity in patients with type 2 diabetes (113 obese, BMI =30; 232 non-obese, BMI < 30) of European ancestry. No evidence of association was observed between the allele c.4349G>A and obesity, but we observed a significantly higher frequency of homozygotes c.1136TT in obese (19.5 percent) than in non-obese individuals (10.9 percent) [P = 0.02; OR = 2.0 (95 percentCI: 1.07-3.73)], suggesting that the allele c.1136T is associated to obesity in a recessive model. This genotype, after controlling for cholesterol, LDL cholesterol, and triglycerides, was independently associated with obesity (P = 0.048), and increases the chance of obesity in 2.8 times. Therefore, our data suggest the involvement of collagen XVIII in human adipogenesis and susceptibility to obesity.

Colágeno XVIII pode gerar dois fragmentos, um correspondendo à região NC11-728 contendo o motivo ''frizzled'', o qual possivelmente atua na sinalização Wnt, e outro correspondendo a Endostatina, que é clivada a partir da região NC1 e é uma potente inibidora de angiogênese. Colágeno XVIII e a via de sinalização Wnt foram recentemente associados à diferenciação adipogênica e obesidade em alguns modelosanimais, porém ainda não em humanos. No presente trabalho, mostramos que os níveis de expressão gênica do COL18A1 aumentam durante o processo de diferenciação adipogênica em humanos. Também testamos se polimorfismos localizados no motivo ''Frizzled'' (c.1136C > T; Thr379Met) e na região da Endostatina (c.4349G > A; Asp1437Asn) contribuem na predisposição a obesidade em pacientes com diabetes tipo 2. (113 obesos, BMI > 30; 232 não-obesos, BMI < 30) de ancestralidade Européia. Nenhuma evidência de associação entre o alelo c.4349G > A e obesidade foi observada, contudo, observamos uma freqüência significativamente maior de homozigotos c.1136TT em obesos (19.5 por cento) do que em não-obesos (10.9 por cento)[P = 0.02; OR = 2.0 (95 por centoCI: 1.07-3.73)], sugerindo que o alelo c.1136T está associado com obesidade conforme ummodelo recessivo. Este genótipo manteve-se associado à obesidade (P = 0.048) mesmo após o controle das variáveis colesterol, LDL e triglicérides, e confere um risco 2.8 vezes maior de obesidade. Portanto, nossos dados sugerem o envolvimento do colágeno XVIII em adipogênese humana e predisposição a obesidade.

Mutations in collagen 18A1 (COL18A1) and their relevance to the human phenotype

Colágeno XVIII, uma proteoglicana, é um componente das membranas basais (MBs). Existem três isoformas distintas que diferem apenas na região N-terminal, mas que apresentam um padrão específico de expressão nos diferentes tecidos e durante o desenvolvimento. A clivagem da região C-terminal produz endostatina, um inibidor de angiogênese. Na sua região N-terminal, há um motivo "frizzled'', o qual parece estar envolvido com a sinalização de Wnt. Mutações no gene COL18A1 causam a síndrome de Knobloch (SK), uma condição de herança autossômica recessiva caracterizada por degeneração vítreo - retiniana, degeneração de mácula e encefalocele occipital. Esta revisão discute o efeito tanto de alelos raros como polimórficos no fenótipo, mostrando que deficiência de uma das isoformas de colágeno XVIII é suficiente para causar SK e que alelos nulos causando deficiência de todas as isoformas de colágeno XVIII estão associadas a alterações oculares mais graves. Esta revisão, além de ilustrar a importância funcional do colágeno XVIII no desenvolvimento do olho e na manutenção de sua estrutura, também mostra que esta proteína tem um papel funcional importante em outros tecidos e órgão, como no sistema nervoso central e rim.