BACKGROUND Guanine nucleotide-binding protein,alpha stimulating(GNAS)mutations are characteristic of intraductal papillary mucinous neoplasms(IPMNs).Pancreatic ductal adenocarcinomas(PDACs)harboring GNAS mutations ori...BACKGROUND Guanine nucleotide-binding protein,alpha stimulating(GNAS)mutations are characteristic of intraductal papillary mucinous neoplasms(IPMNs).Pancreatic ductal adenocarcinomas(PDACs)harboring GNAS mutations originate in IPMNs.GNAS is a complex imprinted locus that produces five transcripts regulated by differential methylated regions,NESP55,GNASAS,GNASXL,GNAS1A,and GNAS.AIM To evaluate if methylation changes in the differential methylated regions of GNAS locus contributed to malignant progression of pancreatic cysts.METHODS GNAS locus methylation was analyzed in archival pancreatic cyst fluid(PCF)obtained by endoscopic ultrasound with fine-needle aspiration by methylation specific–multiplex ligation dependent probe amplification.Results were normalized and analyzed using Coffalyser.Net software.RESULTS Fifty-two PCF samples obtained by endoscopic ultrasound with fine-needle aspiration and previously characterized for KRAS and GNAS mutations were studied.The final diagnoses were surgical(11)and clinicopathological(41),including 30 benign cysts,14 pre-malignant cyst,and eight malignant cysts.Methylation changes at NESP55,GNASAS,GNAS1A,and especially GNASXL were more frequent in malignant cysts,and NESP55 and GNASAS were useful for diagnosis.A combined variable defined as“GNAS locus methylation changes”was significantly associated with malignancy(6/8 malignant cysts and only 2/20 benign cysts)and improved classification.Hypermethylation in both maternally(NESP55)and paternally(GNASXL)derived promoters was found in 3/3 PDACs.CONCLUSION This is the first study to identify methylation changes in the GNAS locus,improving the diagnosis of malignant pancreatic cysts and suggesting a role in progression to PDAC.展开更多
Green macroalgae,e.g.,Ulva lactuca,are valuable bioactive sources of nutrients;but algae recalcitrant cell walls,composed of a complex cross-linked matrix of polysaccharides,can compromise their utilization as feedstu...Green macroalgae,e.g.,Ulva lactuca,are valuable bioactive sources of nutrients;but algae recalcitrant cell walls,composed of a complex cross-linked matrix of polysaccharides,can compromise their utilization as feedstuffs for monogastric animals.This study aimed to evaluate the ability of pre-selected Carbohydrate-Active enZymes(CAZymes)and sulfatases to degrade U.lactuca cell walls and release nutritive compounds.A databank of 199 recombinant CAZymes and sulfatases was tested in vitro for their action towards U.lactuca cell wall polysaccharides.The enzymes were incubated with the macroalga,either alone or in combination,to release reducing sugars and decrease fluorescence intensity of Calcofluor White stained cell walls.The individual action of a polysaccharide lyase family 25(PL25),an ulvan lyase,was shown to be the most efficient in cell wall disruption.The ulvan lyase treatment,in triplicate measures,promoted the release of 4.54 g/L(P<0.001)reducing sugars,a mono-and oligosaccharides release of 11.4 and 11.2 mmol/100 g of dried alga(P<0.01),respectively,and a decrease of 41.7%(P<0.001)in cell wall fluorescence,in comparison to control.The ability of ulvan lyase treatment to promote the release of nutritional compounds from alga biomass was also evaluated.A release of some monounsaturated fatty acids was observed,particularly the health beneficial 18:1c9(P<0.001).However,no significant release of total fatty acids(P>0.05),proteins(P?0.861)or pigments(P>0.05)was found.These results highlight the capacity of a single recombinant ulvan lyase(PL25 family)to incompletely disrupt U.lactuca cell walls.This enzyme could enhance the bioaccessibility of U.lactuca bioactive products with promising utilization in the feed industry.展开更多
基金Supported by a Research Grant from Sociedade Portuguesa de Endoscopia Digestiva in 2018.
文摘BACKGROUND Guanine nucleotide-binding protein,alpha stimulating(GNAS)mutations are characteristic of intraductal papillary mucinous neoplasms(IPMNs).Pancreatic ductal adenocarcinomas(PDACs)harboring GNAS mutations originate in IPMNs.GNAS is a complex imprinted locus that produces five transcripts regulated by differential methylated regions,NESP55,GNASAS,GNASXL,GNAS1A,and GNAS.AIM To evaluate if methylation changes in the differential methylated regions of GNAS locus contributed to malignant progression of pancreatic cysts.METHODS GNAS locus methylation was analyzed in archival pancreatic cyst fluid(PCF)obtained by endoscopic ultrasound with fine-needle aspiration by methylation specific–multiplex ligation dependent probe amplification.Results were normalized and analyzed using Coffalyser.Net software.RESULTS Fifty-two PCF samples obtained by endoscopic ultrasound with fine-needle aspiration and previously characterized for KRAS and GNAS mutations were studied.The final diagnoses were surgical(11)and clinicopathological(41),including 30 benign cysts,14 pre-malignant cyst,and eight malignant cysts.Methylation changes at NESP55,GNASAS,GNAS1A,and especially GNASXL were more frequent in malignant cysts,and NESP55 and GNASAS were useful for diagnosis.A combined variable defined as“GNAS locus methylation changes”was significantly associated with malignancy(6/8 malignant cysts and only 2/20 benign cysts)and improved classification.Hypermethylation in both maternally(NESP55)and paternally(GNASXL)derived promoters was found in 3/3 PDACs.CONCLUSION This is the first study to identify methylation changes in the GNAS locus,improving the diagnosis of malignant pancreatic cysts and suggesting a role in progression to PDAC.
基金Fundaçao para a Ciencia e Tecnologia(FCT,Lisbon,Portugal)through grant PTDC/CAL-ZOO/30238/2017 associated post-doc contract to MMC,CIISA(Project UIDB/00276/2020)a PhD fellowship to DFC(SFRH/BD/126198/2016).
文摘Green macroalgae,e.g.,Ulva lactuca,are valuable bioactive sources of nutrients;but algae recalcitrant cell walls,composed of a complex cross-linked matrix of polysaccharides,can compromise their utilization as feedstuffs for monogastric animals.This study aimed to evaluate the ability of pre-selected Carbohydrate-Active enZymes(CAZymes)and sulfatases to degrade U.lactuca cell walls and release nutritive compounds.A databank of 199 recombinant CAZymes and sulfatases was tested in vitro for their action towards U.lactuca cell wall polysaccharides.The enzymes were incubated with the macroalga,either alone or in combination,to release reducing sugars and decrease fluorescence intensity of Calcofluor White stained cell walls.The individual action of a polysaccharide lyase family 25(PL25),an ulvan lyase,was shown to be the most efficient in cell wall disruption.The ulvan lyase treatment,in triplicate measures,promoted the release of 4.54 g/L(P<0.001)reducing sugars,a mono-and oligosaccharides release of 11.4 and 11.2 mmol/100 g of dried alga(P<0.01),respectively,and a decrease of 41.7%(P<0.001)in cell wall fluorescence,in comparison to control.The ability of ulvan lyase treatment to promote the release of nutritional compounds from alga biomass was also evaluated.A release of some monounsaturated fatty acids was observed,particularly the health beneficial 18:1c9(P<0.001).However,no significant release of total fatty acids(P>0.05),proteins(P?0.861)or pigments(P>0.05)was found.These results highlight the capacity of a single recombinant ulvan lyase(PL25 family)to incompletely disrupt U.lactuca cell walls.This enzyme could enhance the bioaccessibility of U.lactuca bioactive products with promising utilization in the feed industry.
