Parkinson's disease(PD) is characterized by alphasynucleinopathy that affects all levels of the braingut axis including the central, autonomic, and enteric nervous systems. Recently, it has been recognized that th...Parkinson's disease(PD) is characterized by alphasynucleinopathy that affects all levels of the braingut axis including the central, autonomic, and enteric nervous systems. Recently, it has been recognized that the brain-gut axis interactions are significantly modulated by the gut microbiota via immunological,neuroendocrine, and direct neural mechanisms. Dysregulation of the brain-gut-microbiota axis in PD may be associated with gastrointestinal manifestations frequently preceding motor symptoms, as well as with the pathogenesis of PD itself, supporting the hypothesis that the pathological process is spread from the gut to the brain. Excessive stimulation of the innate immune system resulting from gut dysbiosis and/or small intestinal bacterial overgrowth and increased intestinal permeability may induce systemic inflammation, while activation of enteric neurons and enteric glial cells may contribute to the initiation of alpha-synuclein misfolding.Additionally, the adaptive immune system may be disturbed by bacterial proteins cross-reacting with human antigens. A better understanding of the brain-gutmicrobiota axis interactions should bring a new insight in the pathophysiology of PD and permit an earlier diagnosis with a focus on peripheral biomarkers within the enteric nervous system. Novel therapeutic options aimed at modifying the gut microbiota composition and enhancing the intestinal epithelial barrier integrity in PD patients could influence the initial step of the following cascade of neurodegeneration in PD.展开更多
Helicobacter pylori (H. pylori) infection underlies gastric ulcer disease, gastric cancer and duodenal ulcer disease. The disease expression reflects the pattern and extent of gastritis/gastric atrophy (i.e., duodenal...Helicobacter pylori (H. pylori) infection underlies gastric ulcer disease, gastric cancer and duodenal ulcer disease. The disease expression reflects the pattern and extent of gastritis/gastric atrophy (i.e., duodenal ulcer with non-atrophic and gastric ulcer and gastric cancer with atrophic gastritis). Gastric and duodenal ulcers and gastric cancer have been known for thousands of years. Ulcers are generally non-fatal and until the 20<sup>th</sup> century were difficult to diagnose. However, the presence and pattern of gastritis in past civilizations can be deduced based on the diseases present. It has been suggested that gastric ulcer and duodenal ulcer both arose or became more frequent in Europe in the 19<sup>th</sup> century. Here, we show that gastric cancer and gastric ulcer were present throughout the 17<sup>th</sup> to 19<sup>th</sup> centuries consistent with atrophic gastritis being the predominant pattern, as it proved to be when it could be examined directly in the late 19<sup>th</sup> century. The environment before the 20<sup>th</sup> century favored acquisition of H. pylori infection and atrophic gastritis (e.g., poor sanitation and standards of living, seasonal diets poor in fresh fruits and vegetables, especially in winter, vitamin deficiencies, and frequent febrile infections in childhood). The latter part of the 19<sup>th</sup> century saw improvements in standards of living, sanitation, and diets with a corresponding decrease in rate of development of atrophic gastritis allowing duodenal ulcers to become more prominent. In the early 20<sup>th</sup> century physician’s believed they could diagnose ulcers clinically and that the diagnosis required hospitalization for “surgical disease” or for “Sippy” diets. We show that while H. pylori remained common and virulent in Europe and the United States, environmental changes resulted in changes of the pattern of gastritis producing a change in the manifestations of H. pylori infections and subsequently to a rapi展开更多
“Lingzhi”is a mushroom that has been renowned in China for more than 2,000 years because of its claimed medicinal properties plus its symbolic fortune.“Lingzhi”has high economic value mostly as a dietary supplemen...“Lingzhi”is a mushroom that has been renowned in China for more than 2,000 years because of its claimed medicinal properties plus its symbolic fortune.“Lingzhi”has high economic value mostly as a dietary supplement in the modern market especially in East Asia,and its medicinal functions have become a hot study topic.For over a century,the highly prized medicinal fungus,known as“Lingzhi”in East Asia,has been assigned to Ganoderma lucidum,a species originally described from Europe.Molecular studies in recent years have revealed that the commercially cultivated‘G.lucidum’(“Lingzhi”)in East Asia is a different species from the true G.lucidum.The present study aims to clarify the species identity of“Lingzhi”based on morphological studies and analysis of rDNA nuc-ITS sequences,and additional gene fragments of mt-SSU,RPB1,RPB2,and TEF1-αof“Lingzhi”were provided.All Ganoderma species that mostly resemble“Lingzhi”in phylogeny and/or morphology were included for analysis.We propose a new species G.lingzhi for“Lingzhi”,which has an East Asia distribution.The most striking characteristics which differentiate G.lingzhi from G.lucidum are the presence of melanoid bands in the context,a yellow pore surface and thick dissepiments(80–120μm)at maturity.G.curtisii is most closely related to G.lingzhi in phylogeny and is from North America.Ganoderma flexipes,G.multipileum,G.sichuanense,G.tropicum and‘G.tsugae’,are also closely related with G.lingzhi and are reported from China.These species are compared and discussed.‘Ganoderma tsuage’reported from China is determined as conspecific with G.lucidum,hence the distribution of G.lucidum extends from Europe to northeastern China.展开更多
Background Previous studies have indicated that thrombi n (TM) may play a major role in brain edema after intracerebral hemorrhages (ICHs). However, the mechanism of TM-induced brain edema is poorly understood. In th...Background Previous studies have indicated that thrombi n (TM) may play a major role in brain edema after intracerebral hemorrhages (ICHs). However, the mechanism of TM-induced brain edema is poorly understood. In this study, we explored the effect of TM on the permeability of the blood brain barrier (BBB) and investigated its possible mechanism, aiming at providing a potential target for brain edema therapy after ICHs.Methods TM or TM + cathepsin G (CATG) was stereotaxically injected into the right caudate nucleus of Sprague-Dawley rats in vivo. BBB permeability was measured by Evans-Blue extravasation. Brain water content was determined by the dry-wet weight method. Brain microvascular endothelial cells were then cultured in vitro. After TM or TM+CATG was added to the endothelial cell medium, changes in the morphology of cells were dynamically observed by phase-contrast light microscopy, and the expression of matrix metalloproteinase-2 (MMP-2) protein was measured by immunohistochemical method.Results BBB permeability increased at 6 hours after a TM injection into the ipsilateral caudate nucleus (P<0.05), peaked between 24 hours (P<0.01) and 48 hours (P<0.05) after the injection, and then declined. Brain water content changed in parallel with the changes in BBB permeability. However, at all time points, BBB permeability and brain water content after a TM+CATG injection were not significantly different from the respective parameters in the control group (P>0.05). TM induced endothelial cell contraction in vitro in a time-dependent manner and enhanced the expression of MMP-2 protein. After incubation with TM+CATG, cell morphology and MMP-2 expression did not change significantly as compared to the control group (P>0.05).Conclusions Increased BBB permeability may be one of the mechanisms behind TM-induced cerebral edema. TM induces endothelial cell contraction and promotes MMP-2 expression by activating protease activated receptor-1 (PAR-1), possibly leading to the opening of the BBB.展开更多
β-Thalassemia is a global health issue, caused by mutations in the HBB gene. Among these mutations, HBB -28 （A〉G） mutations is one of the three most common mutations in China and Southeast Asia patients with β-th...β-Thalassemia is a global health issue, caused by mutations in the HBB gene. Among these mutations, HBB -28 （A〉G） mutations is one of the three most common mutations in China and Southeast Asia patients with β-thalassemia. Correcting this mutation in human embryos may prevent the disease being passed onto future generations and cure anemia. Here we report the first study using base editor （BE） system to correct disease mutant in human embryos. Firstly, we produced a 293T cell line with an exogenous HBB -28 （A〉G） mutant fragment for gRNAs and targeting efficiency evaluation. Then we collected primary skin fibroblast cells from a β-thalassemia patient with HBB -28 （A〉G） homozygous mutation. Data showed that base editor could precisely correct HBB -28 （A〉G） mutation in the patient＇s primary cells. To model homozygous mutation disease embryos, we consb＇ucted nuclear transfer embryos by fusing the lymphocyte or skin fibroblast cells with enucleated in vitro matured （IVM） oocytes.Notably, the gene correction efficiency was over 23.0% in these embryos by base editor. Although these embryos were still mosaic, the percentage of repaired blastomeres was over 20.0%. In addition, we found that base editor variants, with narrowed deamination window, could promote G-to-A conversion at HBB -28 site precisely in human embryos. Collectively, this study demonstrated the feasibility of curing genetic disease in human somatic cells and embryos by base editor system.展开更多
AIM: To investigate the relationship between gastric dysmotility,gastrointestinal hormone abnormalities, and neuroendocrine cells in gastrointestinal mucosa in patients with functional dyspepsia (FD).METHODS: Gastric ...AIM: To investigate the relationship between gastric dysmotility,gastrointestinal hormone abnormalities, and neuroendocrine cells in gastrointestinal mucosa in patients with functional dyspepsia (FD).METHODS: Gastric emptying was assessed with solid radiopaque markers in 54 FD patients, and the patients were divided into two groups according to the results, one with delayed gastric emptying and the other with normal gastric emptying. Seventeen healthy volunteers acted as normal controls. Fasting and postprandial plasma levels and gastroduodenal mucosal levels of gastrointestinal hormones gastrin, somatostatin (SS) and neurotensin (NT)were measured by radioimmunoassay in all the subjects.G cells (gastrin-producing cells) and D cells (SS-producing cells) in gastric antral mucosa were immunostained with rabbit anti-gastrin polyclonal antibody and rabbit anti-SS polyclonal antibody, respectively, and analyzed quantitatively by computerized image analysis.RESULTS: The postprandial plasma gastrin levels, the fasting and postprandial plasma levels and the gastric and duodenal mucosal levels of NT were significantly higher in the FD patients with delayed gastric emptying than in those with normal gastric emptying and normal controls. The number and gray value of G and D cells and the G cell/D cell number ratio did not differ significantly between normal controls and the FD patients with or without delayed gastric emptying.CONCLUSION: Our findings suggest that the abnormalities of gastrin and NT may play a role in the pathophysiology of gastric dysmotility in FD patients, and the abnormality of postprandial plasma gastrin levels in FD patients with delayed gastric emptying is not related to the changes both in the number and gray value of G cells and in the G cell/D cell number ratio in gastric antral mucosa.展开更多
AIM: To screen and investigate the effective g RNAs against hepatitis B virus(HBV) of genotypes A-D.METHODS: A total of 15 g RNAs against HBV of genotypes A-D were designed. Eleven combinations of two above g RNAs(dua...AIM: To screen and investigate the effective g RNAs against hepatitis B virus(HBV) of genotypes A-D.METHODS: A total of 15 g RNAs against HBV of genotypes A-D were designed. Eleven combinations of two above g RNAs(dual-g RNAs) covering the regulatory region of HBV were chosen. The efficiency of each g RNA and 11 dual-g RNAs on the suppression of HBV(genotypes A-D) replication was examined by the measurement of HBV surface antigen(HBs Ag) or e antigen(HBe Ag) in the culture supernatant. The destruction of HBV-expressing vector was examined in Hu H7 cells co-transfected with dual-g RNAs and HBVexpressing vector using polymerase chain reaction(PCR) and sequencing method, and the destruction of ccc DNAwas examined in Hep AD38 cells using KCl precipitation, plasmid-safe ATP-dependent DNase(PSAD) digestion, rolling circle amplification and quantitative PCR combined method. The cytotoxicity of these g RNAs was assessed by a mitochondrial tetrazolium assay.RESULTS: All of g RNAs could significantly reduce HBs Ag or HBe Ag production in the culture supernatant, which was dependent on the region in which g RNA against. All of dual g RNAs could efficiently suppress HBs Ag and/or HBe Ag production for HBV of genotypes A-D, and the efficacy of dual g RNAs in suppressing HBs Ag and/or HBe Ag production was significantly increased when compared to the single g RNA used alone. Furthermore, by PCR direct sequencing we confirmed that these dual g RNAs could specifically destroy HBV expressing template by removing the fragment between the cleavage sites of the two used g RNAs. Most importantly, g RNA-5 and g RNA-12 combination not only could efficiently suppressing HBs Ag and/or HBe Ag production, but also destroy the ccc DNA reservoirs in Hep AD38 cells.CONCLUSION: These results suggested that CRISPR/Cas9 system could efficiently destroy HBV expressing templates(genotypes A-D) without apparent cytotoxicity. It may be a potential approach for eradication of persistent HBV ccc DNA in chronic HBV infection patients.展开更多
进入新世纪以来,可持续发展在世界范围内成为一个更加严峻的话题,为了更有效地控制和利用我们的环境资源,许多国家纷纷制定了相关的建筑环境评估体系。本文介绍了当今四个具影响力的环境评估体系,包括英国的 BREEAM体系,美国的 L E E D...进入新世纪以来,可持续发展在世界范围内成为一个更加严峻的话题,为了更有效地控制和利用我们的环境资源,许多国家纷纷制定了相关的建筑环境评估体系。本文介绍了当今四个具影响力的环境评估体系,包括英国的 BREEAM体系,美国的 L E E DTM 系统,澳大利亚的NABERS以及中国的 GBCAS。通过对比阐述了四个体系不同的发展历程、内容、权重及特别之处。展开更多
Heterotrimeric G proteins are known to function as messengers in numerous signal transduction pathways.The nullmutation of RGA(rice heterotrimeric G protein α subunit),which encodes the α subunit of heterotrimeric G...Heterotrimeric G proteins are known to function as messengers in numerous signal transduction pathways.The nullmutation of RGA(rice heterotrimeric G protein α subunit),which encodes the α subunit of heterotrimeric G proteinin rice,causes severe dwarfism and reduced responsiveness to gibberellic acid in rice.However,less is known aboutheterotrimeric G protein in brassinosteroid(BR)signaling,one of the well-understood phytohormone pathways.In thepresent study,we used root elongation inhibition assay,lamina inclination assay and coleoptile elongation analysis todemonstrated reduced sensitivity of dl mutant plants(caused by the null mutation of RGA)to 24-epibrassinolide(24-epiBL),which belongs to brassinosteroids and plays a wide variety of roles in plant growth and development.Moreover,RGA transcript level was decreased in 24-epiBL-treated seedlings in a dose-dependent manner.Our results show thatRGA is involved in rice brassinosteroid response,which may be beneficial to elucidate the molecular mechanisms of Gprotein signaling and provide a novel perspective to understand BR signaling in higher plants.展开更多
文摘Parkinson's disease(PD) is characterized by alphasynucleinopathy that affects all levels of the braingut axis including the central, autonomic, and enteric nervous systems. Recently, it has been recognized that the brain-gut axis interactions are significantly modulated by the gut microbiota via immunological,neuroendocrine, and direct neural mechanisms. Dysregulation of the brain-gut-microbiota axis in PD may be associated with gastrointestinal manifestations frequently preceding motor symptoms, as well as with the pathogenesis of PD itself, supporting the hypothesis that the pathological process is spread from the gut to the brain. Excessive stimulation of the innate immune system resulting from gut dysbiosis and/or small intestinal bacterial overgrowth and increased intestinal permeability may induce systemic inflammation, while activation of enteric neurons and enteric glial cells may contribute to the initiation of alpha-synuclein misfolding.Additionally, the adaptive immune system may be disturbed by bacterial proteins cross-reacting with human antigens. A better understanding of the brain-gutmicrobiota axis interactions should bring a new insight in the pathophysiology of PD and permit an earlier diagnosis with a focus on peripheral biomarkers within the enteric nervous system. Novel therapeutic options aimed at modifying the gut microbiota composition and enhancing the intestinal epithelial barrier integrity in PD patients could influence the initial step of the following cascade of neurodegeneration in PD.
基金Supported by In part by the Office of Research and Development Medical Research Service Department of Veterans Affairs,Public Health Service grants No.DK062813 and No.DK56338 which funds the Texas Medical Center Digestive Diseases Center
文摘Helicobacter pylori (H. pylori) infection underlies gastric ulcer disease, gastric cancer and duodenal ulcer disease. The disease expression reflects the pattern and extent of gastritis/gastric atrophy (i.e., duodenal ulcer with non-atrophic and gastric ulcer and gastric cancer with atrophic gastritis). Gastric and duodenal ulcers and gastric cancer have been known for thousands of years. Ulcers are generally non-fatal and until the 20<sup>th</sup> century were difficult to diagnose. However, the presence and pattern of gastritis in past civilizations can be deduced based on the diseases present. It has been suggested that gastric ulcer and duodenal ulcer both arose or became more frequent in Europe in the 19<sup>th</sup> century. Here, we show that gastric cancer and gastric ulcer were present throughout the 17<sup>th</sup> to 19<sup>th</sup> centuries consistent with atrophic gastritis being the predominant pattern, as it proved to be when it could be examined directly in the late 19<sup>th</sup> century. The environment before the 20<sup>th</sup> century favored acquisition of H. pylori infection and atrophic gastritis (e.g., poor sanitation and standards of living, seasonal diets poor in fresh fruits and vegetables, especially in winter, vitamin deficiencies, and frequent febrile infections in childhood). The latter part of the 19<sup>th</sup> century saw improvements in standards of living, sanitation, and diets with a corresponding decrease in rate of development of atrophic gastritis allowing duodenal ulcers to become more prominent. In the early 20<sup>th</sup> century physician’s believed they could diagnose ulcers clinically and that the diagnosis required hospitalization for “surgical disease” or for “Sippy” diets. We show that while H. pylori remained common and virulent in Europe and the United States, environmental changes resulted in changes of the pattern of gastritis producing a change in the manifestations of H. pylori infections and subsequently to a rapi
基金financed by the National Natural Science Foundation of China(Project Nos.30910103907,31070022).
文摘“Lingzhi”is a mushroom that has been renowned in China for more than 2,000 years because of its claimed medicinal properties plus its symbolic fortune.“Lingzhi”has high economic value mostly as a dietary supplement in the modern market especially in East Asia,and its medicinal functions have become a hot study topic.For over a century,the highly prized medicinal fungus,known as“Lingzhi”in East Asia,has been assigned to Ganoderma lucidum,a species originally described from Europe.Molecular studies in recent years have revealed that the commercially cultivated‘G.lucidum’(“Lingzhi”)in East Asia is a different species from the true G.lucidum.The present study aims to clarify the species identity of“Lingzhi”based on morphological studies and analysis of rDNA nuc-ITS sequences,and additional gene fragments of mt-SSU,RPB1,RPB2,and TEF1-αof“Lingzhi”were provided.All Ganoderma species that mostly resemble“Lingzhi”in phylogeny and/or morphology were included for analysis.We propose a new species G.lingzhi for“Lingzhi”,which has an East Asia distribution.The most striking characteristics which differentiate G.lingzhi from G.lucidum are the presence of melanoid bands in the context,a yellow pore surface and thick dissepiments(80–120μm)at maturity.G.curtisii is most closely related to G.lingzhi in phylogeny and is from North America.Ganoderma flexipes,G.multipileum,G.sichuanense,G.tropicum and‘G.tsugae’,are also closely related with G.lingzhi and are reported from China.These species are compared and discussed.‘Ganoderma tsuage’reported from China is determined as conspecific with G.lucidum,hence the distribution of G.lucidum extends from Europe to northeastern China.
文摘Background Previous studies have indicated that thrombi n (TM) may play a major role in brain edema after intracerebral hemorrhages (ICHs). However, the mechanism of TM-induced brain edema is poorly understood. In this study, we explored the effect of TM on the permeability of the blood brain barrier (BBB) and investigated its possible mechanism, aiming at providing a potential target for brain edema therapy after ICHs.Methods TM or TM + cathepsin G (CATG) was stereotaxically injected into the right caudate nucleus of Sprague-Dawley rats in vivo. BBB permeability was measured by Evans-Blue extravasation. Brain water content was determined by the dry-wet weight method. Brain microvascular endothelial cells were then cultured in vitro. After TM or TM+CATG was added to the endothelial cell medium, changes in the morphology of cells were dynamically observed by phase-contrast light microscopy, and the expression of matrix metalloproteinase-2 (MMP-2) protein was measured by immunohistochemical method.Results BBB permeability increased at 6 hours after a TM injection into the ipsilateral caudate nucleus (P<0.05), peaked between 24 hours (P<0.01) and 48 hours (P<0.05) after the injection, and then declined. Brain water content changed in parallel with the changes in BBB permeability. However, at all time points, BBB permeability and brain water content after a TM+CATG injection were not significantly different from the respective parameters in the control group (P>0.05). TM induced endothelial cell contraction in vitro in a time-dependent manner and enhanced the expression of MMP-2 protein. After incubation with TM+CATG, cell morphology and MMP-2 expression did not change significantly as compared to the control group (P>0.05).Conclusions Increased BBB permeability may be one of the mechanisms behind TM-induced cerebral edema. TM induces endothelial cell contraction and promotes MMP-2 expression by activating protease activated receptor-1 (PAR-1), possibly leading to the opening of the BBB.
基金We are grateful to Dr. Qi Zhou for helpful suggestions. This work was supported by National Key R＆D Program of China （2017YFC1001901 and 2017YFC1001600）, the Science and Technology Planning Project of Guangdong Province （2015B020228002）, the Guangzhou Science and Technology Project （201707010085） and the National Natural Science Foundation of China （Grant No. 81771579）.
文摘β-Thalassemia is a global health issue, caused by mutations in the HBB gene. Among these mutations, HBB -28 （A〉G） mutations is one of the three most common mutations in China and Southeast Asia patients with β-thalassemia. Correcting this mutation in human embryos may prevent the disease being passed onto future generations and cure anemia. Here we report the first study using base editor （BE） system to correct disease mutant in human embryos. Firstly, we produced a 293T cell line with an exogenous HBB -28 （A〉G） mutant fragment for gRNAs and targeting efficiency evaluation. Then we collected primary skin fibroblast cells from a β-thalassemia patient with HBB -28 （A〉G） homozygous mutation. Data showed that base editor could precisely correct HBB -28 （A〉G） mutation in the patient＇s primary cells. To model homozygous mutation disease embryos, we consb＇ucted nuclear transfer embryos by fusing the lymphocyte or skin fibroblast cells with enucleated in vitro matured （IVM） oocytes.Notably, the gene correction efficiency was over 23.0% in these embryos by base editor. Although these embryos were still mosaic, the percentage of repaired blastomeres was over 20.0%. In addition, we found that base editor variants, with narrowed deamination window, could promote G-to-A conversion at HBB -28 site precisely in human embryos. Collectively, this study demonstrated the feasibility of curing genetic disease in human somatic cells and embryos by base editor system.
文摘AIM: To investigate the relationship between gastric dysmotility,gastrointestinal hormone abnormalities, and neuroendocrine cells in gastrointestinal mucosa in patients with functional dyspepsia (FD).METHODS: Gastric emptying was assessed with solid radiopaque markers in 54 FD patients, and the patients were divided into two groups according to the results, one with delayed gastric emptying and the other with normal gastric emptying. Seventeen healthy volunteers acted as normal controls. Fasting and postprandial plasma levels and gastroduodenal mucosal levels of gastrointestinal hormones gastrin, somatostatin (SS) and neurotensin (NT)were measured by radioimmunoassay in all the subjects.G cells (gastrin-producing cells) and D cells (SS-producing cells) in gastric antral mucosa were immunostained with rabbit anti-gastrin polyclonal antibody and rabbit anti-SS polyclonal antibody, respectively, and analyzed quantitatively by computerized image analysis.RESULTS: The postprandial plasma gastrin levels, the fasting and postprandial plasma levels and the gastric and duodenal mucosal levels of NT were significantly higher in the FD patients with delayed gastric emptying than in those with normal gastric emptying and normal controls. The number and gray value of G and D cells and the G cell/D cell number ratio did not differ significantly between normal controls and the FD patients with or without delayed gastric emptying.CONCLUSION: Our findings suggest that the abnormalities of gastrin and NT may play a role in the pathophysiology of gastric dysmotility in FD patients, and the abnormality of postprandial plasma gastrin levels in FD patients with delayed gastric emptying is not related to the changes both in the number and gray value of G cells and in the G cell/D cell number ratio in gastric antral mucosa.
基金Supported by Natural Science Foundation of China,No.81471938the National S and T Major Project for Infectious Diseases,No.2013ZX10002-002 and No.2012ZX10002-005111 Project,No.B07001
文摘AIM: To screen and investigate the effective g RNAs against hepatitis B virus(HBV) of genotypes A-D.METHODS: A total of 15 g RNAs against HBV of genotypes A-D were designed. Eleven combinations of two above g RNAs(dual-g RNAs) covering the regulatory region of HBV were chosen. The efficiency of each g RNA and 11 dual-g RNAs on the suppression of HBV(genotypes A-D) replication was examined by the measurement of HBV surface antigen(HBs Ag) or e antigen(HBe Ag) in the culture supernatant. The destruction of HBV-expressing vector was examined in Hu H7 cells co-transfected with dual-g RNAs and HBVexpressing vector using polymerase chain reaction(PCR) and sequencing method, and the destruction of ccc DNAwas examined in Hep AD38 cells using KCl precipitation, plasmid-safe ATP-dependent DNase(PSAD) digestion, rolling circle amplification and quantitative PCR combined method. The cytotoxicity of these g RNAs was assessed by a mitochondrial tetrazolium assay.RESULTS: All of g RNAs could significantly reduce HBs Ag or HBe Ag production in the culture supernatant, which was dependent on the region in which g RNA against. All of dual g RNAs could efficiently suppress HBs Ag and/or HBe Ag production for HBV of genotypes A-D, and the efficacy of dual g RNAs in suppressing HBs Ag and/or HBe Ag production was significantly increased when compared to the single g RNA used alone. Furthermore, by PCR direct sequencing we confirmed that these dual g RNAs could specifically destroy HBV expressing template by removing the fragment between the cleavage sites of the two used g RNAs. Most importantly, g RNA-5 and g RNA-12 combination not only could efficiently suppressing HBs Ag and/or HBe Ag production, but also destroy the ccc DNA reservoirs in Hep AD38 cells.CONCLUSION: These results suggested that CRISPR/Cas9 system could efficiently destroy HBV expressing templates(genotypes A-D) without apparent cytotoxicity. It may be a potential approach for eradication of persistent HBV ccc DNA in chronic HBV infection patients.
文摘进入新世纪以来,可持续发展在世界范围内成为一个更加严峻的话题,为了更有效地控制和利用我们的环境资源,许多国家纷纷制定了相关的建筑环境评估体系。本文介绍了当今四个具影响力的环境评估体系,包括英国的 BREEAM体系,美国的 L E E DTM 系统,澳大利亚的NABERS以及中国的 GBCAS。通过对比阐述了四个体系不同的发展历程、内容、权重及特别之处。
基金This project was supported by the Major State Basic Research Program of China （2005CB 120806）, National Natural Science Foundation of China for Distinguished Young Scholars （30525026） and the State Transgenic Plant Project （JY04-A-01）
文摘Heterotrimeric G proteins are known to function as messengers in numerous signal transduction pathways.The nullmutation of RGA(rice heterotrimeric G protein α subunit),which encodes the α subunit of heterotrimeric G proteinin rice,causes severe dwarfism and reduced responsiveness to gibberellic acid in rice.However,less is known aboutheterotrimeric G protein in brassinosteroid(BR)signaling,one of the well-understood phytohormone pathways.In thepresent study,we used root elongation inhibition assay,lamina inclination assay and coleoptile elongation analysis todemonstrated reduced sensitivity of dl mutant plants(caused by the null mutation of RGA)to 24-epibrassinolide(24-epiBL),which belongs to brassinosteroids and plays a wide variety of roles in plant growth and development.Moreover,RGA transcript level was decreased in 24-epiBL-treated seedlings in a dose-dependent manner.Our results show thatRGA is involved in rice brassinosteroid response,which may be beneficial to elucidate the molecular mechanisms of Gprotein signaling and provide a novel perspective to understand BR signaling in higher plants.
