Green fluorescent protein (GFP) fused to the F_actin binding domain of mouse talin labels the actin cytoskeleton in the immature pollen of stable transformed rice (Oryza sativa L.) plants. Actin microfilaments could b...Green fluorescent protein (GFP) fused to the F_actin binding domain of mouse talin labels the actin cytoskeleton in the immature pollen of stable transformed rice (Oryza sativa L.) plants. Actin microfilaments could be visualized only in the late_developmental stage of the immature pollen. During this developmental stage, microfilaments, initially composed of very short fibrils, develop into a very complex and novel network that sometimes totally and sometimes partially encloses the vegetative nucleus and the spherical shaped generative cell in the central cytoplasm of the immature pollen. The behavior of the actin microfilamentous structure throughout the late_developmental stage of the immature pollen is extremely dynamic, and the likelihood of this structure in generating forces for vegetative nucleus and generative cell movement in the immature pollen has been discussed. No actin filaments were visualized in the spherical generative cells.展开更多
Selecting oocytes that are most likely to develop is crucial for in vitro fertilization and animal cloning. Brilliant cresyl blue (BCB) staining has been used for oocyte selection in large animals, but its wider uti...Selecting oocytes that are most likely to develop is crucial for in vitro fertilization and animal cloning. Brilliant cresyl blue (BCB) staining has been used for oocyte selection in large animals, but its wider utility needs further evaluation. Mouse oocytes were divided into those stained (BCB+) and those unstained (BCB-) according to their ooplasm BCB coloration. Chromatin configurations, cumulus cell apoptosis, cytoplasmic maturity and developmental competence were compared between the BCB+ and BCB- oocytes. The effects of oocyte diameter, sexual maturity and gonadotropin stimulation on the competence of BCB+ oocytes were also analyzed. In the large- and medium-size groups, BCB+ oocytes were larger and showed more surrounded nucleoli (SN) chromatin configurations and higher frequencies of early atresia, and they also gained better cytoplasmic maturity (determined as the intracellular GSH level and pattern of mitochondrial distribution) and higher developmental potential after in vitro maturation (IVM) than the BCB-oocytes. Adult mice produced more BCB+ oocytes with higher competence than the prepubertal mice when not primed with PMSG. PMSG priming increased both proportion and developmental potency of BCB+ oocytes. The BCB+ oocytes in the large-size group showed more SN chromatin configurations, better cytoplasmic maturity and higher developmental potential than their counterparts in the medium-size group. It is concluded that BCB staining can be used as an efficient method for oocyte selection, but that the competence of the BCB+ oocytes may vary with oocyte diameter, animal sexual maturity and gonadotropin stimulation. Taken together, the series of criteria described here would allow for better choices in selecting oocytes for better development.展开更多
Objective To investigate the mechanism of immune hyporesponsiveness induced by donor-antigen- unloaded recipient-derived immature dendritic cell (imDC) of liver grafts in rats. Methods Forty Sprague-Dawley rats (d...Objective To investigate the mechanism of immune hyporesponsiveness induced by donor-antigen- unloaded recipient-derived immature dendritic cell (imDC) of liver grafts in rats. Methods Forty Sprague-Dawley rats (donor) and forty male Wistar rats (recipient) were randomly divided into 4 groups: control, cyclosporine A (CsA), mature DC (mDC), and imDC groups respectively, with 10 donor rats and 10 recipient rats in each group. Recipient rats in CsA group were treated with 10 mg-kg-~'d-I CsA starting day 2 after the transplantation. Recipients in the mDC or imDC groups were given Wistar rat derived mDCs (1 × 10^6/rat) or imDCs (1 × 10^6/rat) via dorsal vein of the penis respectively 1 day before the transplantation. In each group, 5 recipients were kept for determination of survival time and the other 5 rats were executed at day 10 after transplantation. Blood samples were collected for the measurement of serum alanine aminotransferase (ALT), total bilirubin (TBIL), interleukin 2 (IL-2), interferon gamma (IFN-γ), IL-4, and IL-10 levels. Liver tissue was harvested for HE staining and acute rejection evaluation. Expression levels of Fas-L/Fas in the grafts were detected by immunohistochemical staining; andWestern blot was used to detect the expression level of Scurfm. Results The survival time of CsA and imDC groups was significantly longer than that of control and mDC groups (all P〈0.05). The levels of serum ALT and TBIL in the control group (2072.20±217.93 IU/L and 147.42±22.02pmol/L) and mDC group (2117.00±285.13 IU/L and 141.58±20.82 pmol/L) were significantly higher than those in the CsA group (59.68±13.48 IU/L and 15.40±2.13 pmol/L) or imDC group (50.80±9.63 IU/L and 14.44±3.49 pmol/L) (all P〈0.05). In the CsA and imDC groups, the levels of IL-2 (22.52±3.75 pg/mL and 22.12±3.90 pg/mL) and IFN-γ (309.20±25.19 pg/mL and 321.00±21.64 pg/mL) were significantly lower, but the levels of IL-4 (297.60±25.07 pg/mL and 277.00±22.47 pg/mL) and IL-10 (1226.00±140.49 pg/mL and 1423.00±106.39 pg/mL) were higher than those of the control (IL-2:147.78±12.80 pg/mL, IFN-γ: 1758.60±106.22 pg/mL, IL-4:17.40±4.77pg/mL, IL-10: 81.00+ 9.47 pg/mL) and mDC groups (IL-2:142.34±9.29 pg/mL, IFN-7:1835.00±82.63 pg/mL, IL-4: 15.60+ 3.96 μg/mL, IL-10: 68.80± 11.23 pg/mL) (all P〈0.01). The expression level of Scurfin protein on CD4+CD25 + T cells of the imDC group (1.34±0.29) was significantly higher than that in the control (0.72±0.13), CsA (0.37±0.11), and mDC groups (0.78±0.17) (all P〈0.05).展开更多
Whole-genome bisulfite sequencing(WGBS)allows single-base resolution and genome-wide profiling of DNA methylation in plants and animals.This technology provides a powerful tool to identify genes that are potentially c...Whole-genome bisulfite sequencing(WGBS)allows single-base resolution and genome-wide profiling of DNA methylation in plants and animals.This technology provides a powerful tool to identify genes that are potentially controlled by dynamic changes of DNA methylation and demethylation.However,naturally occurring epimutants are rare and genes under epigenetic regulation as well as their biological relevances are often difficult to define.In tomato,fruit development and ripening are a complex process that involves epigenetic control.We have taken the advantage of the tomato epimutant Colourless non-ripening(Cnr)and performed comparative mining of the WGBS datasets for the Cnr and Sl CMT3-silenced Cnr fruits.We compared DNA methylation profiles for the promoter sequences of approximately 5,000 bp immediately upstream of the coding region of a list of20 genes.Differentially methylated regions were found for some of these genes.Virus-induced gene silencing(VIGS)of differentially methylated gene Sl DET1 or Sl PDS resulted in unusual brown pigmentation in Cnr fruits.These results suggest that comparative WGBS coupled with VIGS can be used to identify genes that may contribute to the colourless unripe phenotype of fruit in the Cnr epimutant.展开更多
hb(hunchback) is a contributing factor in anteroposterior axial patterning of insects.Although the hb function in Locusta migratoria manilensis has been investigated,its expression pattern remains unknown.Here,the mou...hb(hunchback) is a contributing factor in anteroposterior axial patterning of insects.Although the hb function in Locusta migratoria manilensis has been investigated,its expression pattern remains unknown.Here,the mouse polyclonal antibody was produced against Hb fusion protein,and then its expression pattern during oogenesis and embryogenesis of L.migratoria manilensis was examined by immunohistochemical staining.Hb protein was detected in the oocyte nucleus which was positioned centrally within the developing oocyte.The oocyte nucleus gradually moved to the posterior end of the egg along with the oocyte maturing.In freshly laid eggs,Hb formed gradient at the posterior end of the egg,and then hb was expressed as a band in the middle of the blastodisc.As the blastodisc differentiated into the head and trunk,the expression region became wide,which would develop into spatial gnathal and thoracic segments.With abdominal segmentation,the expression domain in the gnathal and thoracic region became faint and eventually faded out,while the Hb expression domain appeared at the posterior growth zone in a discontinuous expression manner.The hb expression pattern of L.migratoria manilensis is greatly similar to that of other locusts,such as Schistocerca americana and another L.migratoria.Compared with other insects,hb expression is conserved in the gnathal and thoracic domains,while divergent in oogenesis and abdomen.展开更多
文摘Green fluorescent protein (GFP) fused to the F_actin binding domain of mouse talin labels the actin cytoskeleton in the immature pollen of stable transformed rice (Oryza sativa L.) plants. Actin microfilaments could be visualized only in the late_developmental stage of the immature pollen. During this developmental stage, microfilaments, initially composed of very short fibrils, develop into a very complex and novel network that sometimes totally and sometimes partially encloses the vegetative nucleus and the spherical shaped generative cell in the central cytoplasm of the immature pollen. The behavior of the actin microfilamentous structure throughout the late_developmental stage of the immature pollen is extremely dynamic, and the likelihood of this structure in generating forces for vegetative nucleus and generative cell movement in the immature pollen has been discussed. No actin filaments were visualized in the spherical generative cells.
基金Acknowledgments This study was supported by grants from the China National Natural Science Foundation (Nos. 30430530 and 30571337) and from the Momentous Research Project of the China Ministry of Science and Technology (No. 2006CB944003).
文摘Selecting oocytes that are most likely to develop is crucial for in vitro fertilization and animal cloning. Brilliant cresyl blue (BCB) staining has been used for oocyte selection in large animals, but its wider utility needs further evaluation. Mouse oocytes were divided into those stained (BCB+) and those unstained (BCB-) according to their ooplasm BCB coloration. Chromatin configurations, cumulus cell apoptosis, cytoplasmic maturity and developmental competence were compared between the BCB+ and BCB- oocytes. The effects of oocyte diameter, sexual maturity and gonadotropin stimulation on the competence of BCB+ oocytes were also analyzed. In the large- and medium-size groups, BCB+ oocytes were larger and showed more surrounded nucleoli (SN) chromatin configurations and higher frequencies of early atresia, and they also gained better cytoplasmic maturity (determined as the intracellular GSH level and pattern of mitochondrial distribution) and higher developmental potential after in vitro maturation (IVM) than the BCB-oocytes. Adult mice produced more BCB+ oocytes with higher competence than the prepubertal mice when not primed with PMSG. PMSG priming increased both proportion and developmental potency of BCB+ oocytes. The BCB+ oocytes in the large-size group showed more SN chromatin configurations, better cytoplasmic maturity and higher developmental potential than their counterparts in the medium-size group. It is concluded that BCB staining can be used as an efficient method for oocyte selection, but that the competence of the BCB+ oocytes may vary with oocyte diameter, animal sexual maturity and gonadotropin stimulation. Taken together, the series of criteria described here would allow for better choices in selecting oocytes for better development.
基金Supported by Science and Technology Planning Project of Yunnan Province,China (2007CA007)
文摘Objective To investigate the mechanism of immune hyporesponsiveness induced by donor-antigen- unloaded recipient-derived immature dendritic cell (imDC) of liver grafts in rats. Methods Forty Sprague-Dawley rats (donor) and forty male Wistar rats (recipient) were randomly divided into 4 groups: control, cyclosporine A (CsA), mature DC (mDC), and imDC groups respectively, with 10 donor rats and 10 recipient rats in each group. Recipient rats in CsA group were treated with 10 mg-kg-~'d-I CsA starting day 2 after the transplantation. Recipients in the mDC or imDC groups were given Wistar rat derived mDCs (1 × 10^6/rat) or imDCs (1 × 10^6/rat) via dorsal vein of the penis respectively 1 day before the transplantation. In each group, 5 recipients were kept for determination of survival time and the other 5 rats were executed at day 10 after transplantation. Blood samples were collected for the measurement of serum alanine aminotransferase (ALT), total bilirubin (TBIL), interleukin 2 (IL-2), interferon gamma (IFN-γ), IL-4, and IL-10 levels. Liver tissue was harvested for HE staining and acute rejection evaluation. Expression levels of Fas-L/Fas in the grafts were detected by immunohistochemical staining; andWestern blot was used to detect the expression level of Scurfm. Results The survival time of CsA and imDC groups was significantly longer than that of control and mDC groups (all P〈0.05). The levels of serum ALT and TBIL in the control group (2072.20±217.93 IU/L and 147.42±22.02pmol/L) and mDC group (2117.00±285.13 IU/L and 141.58±20.82 pmol/L) were significantly higher than those in the CsA group (59.68±13.48 IU/L and 15.40±2.13 pmol/L) or imDC group (50.80±9.63 IU/L and 14.44±3.49 pmol/L) (all P〈0.05). In the CsA and imDC groups, the levels of IL-2 (22.52±3.75 pg/mL and 22.12±3.90 pg/mL) and IFN-γ (309.20±25.19 pg/mL and 321.00±21.64 pg/mL) were significantly lower, but the levels of IL-4 (297.60±25.07 pg/mL and 277.00±22.47 pg/mL) and IL-10 (1226.00±140.49 pg/mL and 1423.00±106.39 pg/mL) were higher than those of the control (IL-2:147.78±12.80 pg/mL, IFN-γ: 1758.60±106.22 pg/mL, IL-4:17.40±4.77pg/mL, IL-10: 81.00+ 9.47 pg/mL) and mDC groups (IL-2:142.34±9.29 pg/mL, IFN-7:1835.00±82.63 pg/mL, IL-4: 15.60+ 3.96 μg/mL, IL-10: 68.80± 11.23 pg/mL) (all P〈0.01). The expression level of Scurfin protein on CD4+CD25 + T cells of the imDC group (1.34±0.29) was significantly higher than that in the control (0.72±0.13), CsA (0.37±0.11), and mDC groups (0.78±0.17) (all P〈0.05).
基金supported by Ministry of Agriculture of the People’s Republic of Chinathe National Transgenic Program of China (2016ZX08009001-004 to Yiguo Hong)+4 种基金National Natural Science Foundation of China (31370180 to Yiguo Hong, 31601765 to Weiwei Chen)Hangzhou Normal University Pandeng Program (201108 to Yiguo Hong)the Hangzhou City Government Innovative Program for Science Excellence (20131028 to Yiguo Hong)Zhejiang Provincial Natural Science Foundation (LY14C010005 to Nongnong Shi)the UK Biotechnology and Biological Sciences Research Council (BBS/E/H/00YH0271 to Yiguo Hong)
文摘Whole-genome bisulfite sequencing(WGBS)allows single-base resolution and genome-wide profiling of DNA methylation in plants and animals.This technology provides a powerful tool to identify genes that are potentially controlled by dynamic changes of DNA methylation and demethylation.However,naturally occurring epimutants are rare and genes under epigenetic regulation as well as their biological relevances are often difficult to define.In tomato,fruit development and ripening are a complex process that involves epigenetic control.We have taken the advantage of the tomato epimutant Colourless non-ripening(Cnr)and performed comparative mining of the WGBS datasets for the Cnr and Sl CMT3-silenced Cnr fruits.We compared DNA methylation profiles for the promoter sequences of approximately 5,000 bp immediately upstream of the coding region of a list of20 genes.Differentially methylated regions were found for some of these genes.Virus-induced gene silencing(VIGS)of differentially methylated gene Sl DET1 or Sl PDS resulted in unusual brown pigmentation in Cnr fruits.These results suggest that comparative WGBS coupled with VIGS can be used to identify genes that may contribute to the colourless unripe phenotype of fruit in the Cnr epimutant.
基金supported by the National Natural Science Foundation of China (Grant No. 30700435)Program of Natural Science Foundation of Chongqing (Grant No. CSTC2009BB1387)
文摘hb(hunchback) is a contributing factor in anteroposterior axial patterning of insects.Although the hb function in Locusta migratoria manilensis has been investigated,its expression pattern remains unknown.Here,the mouse polyclonal antibody was produced against Hb fusion protein,and then its expression pattern during oogenesis and embryogenesis of L.migratoria manilensis was examined by immunohistochemical staining.Hb protein was detected in the oocyte nucleus which was positioned centrally within the developing oocyte.The oocyte nucleus gradually moved to the posterior end of the egg along with the oocyte maturing.In freshly laid eggs,Hb formed gradient at the posterior end of the egg,and then hb was expressed as a band in the middle of the blastodisc.As the blastodisc differentiated into the head and trunk,the expression region became wide,which would develop into spatial gnathal and thoracic segments.With abdominal segmentation,the expression domain in the gnathal and thoracic region became faint and eventually faded out,while the Hb expression domain appeared at the posterior growth zone in a discontinuous expression manner.The hb expression pattern of L.migratoria manilensis is greatly similar to that of other locusts,such as Schistocerca americana and another L.migratoria.Compared with other insects,hb expression is conserved in the gnathal and thoracic domains,while divergent in oogenesis and abdomen.