Medulloblastoma is the most common malignant pediatric brain tumor. Some are thought to originate from cerebellar granule neuron progenitors （CGNPs） that fail to undergo normal cell cycle exit and differentiation. T...Medulloblastoma is the most common malignant pediatric brain tumor. Some are thought to originate from cerebellar granule neuron progenitors （CGNPs） that fail to undergo normal cell cycle exit and differentiation. The contribution of microRNAs to the initiation and progression of medulloblastoma remains poorly understood. Increased expression of the miR-183-96-182 cluster of microRNAs has been noted in several aggressive sub- groups. We identified that expression of miR-183-96-182 was higher in medulloblastomas with Pten gene loss in the background of the activated sonic hedgehog （Shh） signaling pathway. Ectopic miR-183-96-182 expression in CGNPs synergized with exogenous Shh to increase proliferation and its role depended on hedgehog signaling ac- tivation. Our findings suggest a new microRNA cluster, the miR-183-96-182, functionally collaborates with the Shh signaling pathway in the development of medulloblastomas in mice.展开更多
目的探讨食管鳞状细胞癌成纤维细胞hedgehog通路是否应答SHH蛋白。方法分离、原代培养食管鳞状细胞癌成纤维细胞,用Western blot方法检测培养的细胞是否为成纤维细胞,SHH蛋白与细胞孵育后提取细胞总RNA,荧光定量PCR方法检测细胞中hedge...目的探讨食管鳞状细胞癌成纤维细胞hedgehog通路是否应答SHH蛋白。方法分离、原代培养食管鳞状细胞癌成纤维细胞,用Western blot方法检测培养的细胞是否为成纤维细胞,SHH蛋白与细胞孵育后提取细胞总RNA,荧光定量PCR方法检测细胞中hedgehog通路是否应答。结果 Western blot结果显示培养细胞表达间质细胞标记Vimentin和Fibronectin,不表达上皮细胞标记E-cadherin。三株细胞中有两株应答SHH蛋白,一株不应答。结论食管鳞状细胞癌中,成纤维细胞对SHH配体是否应答存在着不同的情况,因此,检测间质中Hh通路的活性可为临床上选择对Hh抑制剂应答的肿瘤提供重要的生物标记。展开更多
Odontogenic keratocysts(OKCs) are common cystic lesions of odontogenic epithelial origin that can occur sporadically or in association with naevoid basal cell carcinoma syndrome(NBCCS). OKCs are locally aggressive, ca...Odontogenic keratocysts(OKCs) are common cystic lesions of odontogenic epithelial origin that can occur sporadically or in association with naevoid basal cell carcinoma syndrome(NBCCS). OKCs are locally aggressive, cause marked destruction of the jaw bones and have a propensity to recur. PTCH1 mutations(at ～80%) are frequently detected in the epithelia of both NBCCS-related and sporadic OKCs, suggesting that PTCH1 inactivation might constitutively activate sonic hedgehog(SHH) signalling and play a major role in disease pathogenesis. Thus, small molecule inhibitors of SHH signalling might represent a new treatment strategy for OKCs. However, studies on the molecular mechanisms associated with OKCs have been hampered by limited epithelial cell yields during OKC explant culture. Here, we constructed an isogenic PTCH1^(R135 X/+) cellular model of PTCH1 inactivation by introducing a heterozygous mutation, namely, c.403 C>T(p.R135 X), which has been identified in OKC patients, into a human embryonic stem cell line using the clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated 9(Cas9) system. This was followed by the induction of epithelial differentiation. Using this in vitro isogenic cellular model, we verified that the PTCH1 R135 X/+heterozygous mutation causes ligand-independent activation of SHH signalling due to PTCH1 haploinsufficiency. This activation was found to be downregulated in a dose-dependent manner by the SHH pathway inhibitor GDC-0449. In addition, through inhibition of activated SHH signalling, the enhanced proliferation observed in these induced cells was suppressed, suggesting that GDC-0449 might represent an effective inhibitor of the SHH pathway for use during OKC treatment.展开更多
文摘Medulloblastoma is the most common malignant pediatric brain tumor. Some are thought to originate from cerebellar granule neuron progenitors （CGNPs） that fail to undergo normal cell cycle exit and differentiation. The contribution of microRNAs to the initiation and progression of medulloblastoma remains poorly understood. Increased expression of the miR-183-96-182 cluster of microRNAs has been noted in several aggressive sub- groups. We identified that expression of miR-183-96-182 was higher in medulloblastomas with Pten gene loss in the background of the activated sonic hedgehog （Shh） signaling pathway. Ectopic miR-183-96-182 expression in CGNPs synergized with exogenous Shh to increase proliferation and its role depended on hedgehog signaling ac- tivation. Our findings suggest a new microRNA cluster, the miR-183-96-182, functionally collaborates with the Shh signaling pathway in the development of medulloblastomas in mice.
文摘目的探讨食管鳞状细胞癌成纤维细胞hedgehog通路是否应答SHH蛋白。方法分离、原代培养食管鳞状细胞癌成纤维细胞,用Western blot方法检测培养的细胞是否为成纤维细胞,SHH蛋白与细胞孵育后提取细胞总RNA,荧光定量PCR方法检测细胞中hedgehog通路是否应答。结果 Western blot结果显示培养细胞表达间质细胞标记Vimentin和Fibronectin,不表达上皮细胞标记E-cadherin。三株细胞中有两株应答SHH蛋白,一株不应答。结论食管鳞状细胞癌中,成纤维细胞对SHH配体是否应答存在着不同的情况,因此,检测间质中Hh通路的活性可为临床上选择对Hh抑制剂应答的肿瘤提供重要的生物标记。
基金supported by the National Natural Science Foundation of China (81671006, 81300894)Beijing Natural Science Foundation (7172238)
文摘Odontogenic keratocysts(OKCs) are common cystic lesions of odontogenic epithelial origin that can occur sporadically or in association with naevoid basal cell carcinoma syndrome(NBCCS). OKCs are locally aggressive, cause marked destruction of the jaw bones and have a propensity to recur. PTCH1 mutations(at ～80%) are frequently detected in the epithelia of both NBCCS-related and sporadic OKCs, suggesting that PTCH1 inactivation might constitutively activate sonic hedgehog(SHH) signalling and play a major role in disease pathogenesis. Thus, small molecule inhibitors of SHH signalling might represent a new treatment strategy for OKCs. However, studies on the molecular mechanisms associated with OKCs have been hampered by limited epithelial cell yields during OKC explant culture. Here, we constructed an isogenic PTCH1^(R135 X/+) cellular model of PTCH1 inactivation by introducing a heterozygous mutation, namely, c.403 C>T(p.R135 X), which has been identified in OKC patients, into a human embryonic stem cell line using the clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated 9(Cas9) system. This was followed by the induction of epithelial differentiation. Using this in vitro isogenic cellular model, we verified that the PTCH1 R135 X/+heterozygous mutation causes ligand-independent activation of SHH signalling due to PTCH1 haploinsufficiency. This activation was found to be downregulated in a dose-dependent manner by the SHH pathway inhibitor GDC-0449. In addition, through inhibition of activated SHH signalling, the enhanced proliferation observed in these induced cells was suppressed, suggesting that GDC-0449 might represent an effective inhibitor of the SHH pathway for use during OKC treatment.
