梨幼果中脱落酸和吲哚-3-乙酸的简易气相色谱分析

采用溴甲基五氟苯(PFB-Br)对其衍生、酯化成各具有5个氟的两种衍生物,直接进样测定梨幼果中的脱落酸和吲哚-3-乙酸的含量,能减少前处理的复杂步骤,脱落酸和吲哚-3-乙酸回收率分别为95.8%、84.5%,具有较高的可靠性。     

Antagonistic effect of indole-3-acetic acid on kinetin-stimulated amaranthin accumulation in the cotyledons of Amaranthus mangostanus seedlings

Light triggered the initiation of amaranthin biosynthesis in cotyledons of Amaranthus mangostanus L. seedlings. Cytokinin induced amaranthin synthesis in the dark and increased the accumulation of amaranthin under light irradiation. No studies have explored whether indole-3-acetic acid (IAA) can affect kinetin-induced amaranthin accumulation in seedlings of A. mangostanus L. In this study, we found that IAA inhibited both the kinetin- and light-induced synthesis of amaranthin. In the dark, 10.0 mg l^?1 IAA caused a 68% reduction in amaranthin production after induction by 5.0 mg l^?1 kinetin. In the presence of light, 10.0 mg l^?1 IAA resulted in a 50% decrease in amaranthin synthesis following induction by 5.0 mg l^?1 kinetin. In addition, IAA could reverse kinetin-induced amaranthin accumulation under red, blue, or far-red light conditions. Our results suggest that IAA had an antagonistic effect on the light-induced or cytokinin-stimulated accumulation of amaranthin in the cotyledons of A. mangostanus L. seedlings.     

The roles of cytokinins and abscisic acid in the pericarp of litchi (Litchi chinensis Sonn.) in determining fruit size

Summary

Variations in the concentrations of cytokinins (CTKs) and abscisic acid (ABA) were studied in the pericarp of litchi (Litchi chinensis Sonn.) fruit using the large-fruited cv. ‘Erdanli’ (55.3 g per fruit) and the small-fruited cv. ‘Huaizhi’ (20.9 g per fruit), as well as large fruit (32.4 g) from early blooms and small (20.8 g) fruit from late blooms on the same inflorescences of cv. ‘Feizixiao’ from the same commercial orchard in Guangdong, China in 2000. ‘Erdanli’ had higher concentrations of CTKs than cv. ‘Huaizhi’ on three out of six sampling dates during fruit development, and lower concentrations of abscisic acid (ABA) from 40 d after anthesis. In cv. ‘Feizixiao’, fruit from early blooms had higher concentration of CTKs than fruit from late blooms at all sampling dates, and lower concentrations of ABA on three out of five sampling dates. Therefore, the former had a higher CTK:ABA ratio. These data suggest that a high CTK:ABA ratio favours fruit growth in litchi.     

油菜素内酯和脱落酸调控葡萄果实花色苷合成的研究

【目的】为研究2,4-表油菜素内酯(2,4-epibrassinolide,EBR)和脱落酸(abscisic acid,ABA)处理对葡萄花色苷合成与可溶性固形物含量、果皮苯丙氨酸解氨酶(PAL)和类黄酮糖基转移酶(UFGT)活性以及果实内源ABA含量的影响,探索油菜素内酯调控葡萄果实成熟及花色苷合成的机理。【方法】以酿酒葡萄‘赤霞珠’(Cabernet Sauvignon)和‘烟73’(Yan 73)为试材,在葡萄转色前分别用0.1、0.4、0.8 mg.L-1EBR,1mg.L-1Brz(brassinazole,BR生物合成抑制剂)和200 mg.L-1ABA,均匀喷施于葡萄果实,在葡萄成熟过程中测定葡萄果皮花色苷含量及PAL和UFGT酶活性,同时测定果实ABA和可溶性固形物含量。【结果】在果实着色初期,‘赤霞珠’和‘烟73’葡萄果皮PAL和UFGT活性及果实内源ABA含量均逐渐升高,当果实接近成熟花色苷含量基本稳定时ABA含量开始降低。与对照相比,0.4mg.L-1EBR和200 mg.L-1ABA处理显著增加了果实内源ABA含量,提高了果皮PAL和UFGT活性,促进了果皮花色苷的合成和果实可溶性固形物的积累。0.1 mg.L-1和0.8 mg.L-1EBR处理总体增加了果实内源ABA含量,促进了花色苷的合成和可溶性固形物的积累,并提高了UFGT和PAL酶活性,但比0.4 mg.L-1EBR处理提高的幅度小,且差异显著。1 mg.L-1Brz处理使果实ABA的合成推迟,果实可溶性固形物含量、果皮PAL和UFGT活性以及花色苷含量均低于对照,但差异不显著。【结论】外源EBR和ABA处理促进了葡萄成熟和花色苷合成,在不同浓度EBR处理中,以0.4 mg.L-1处理效果较好;内源ABA可能参与了EBR对葡萄成熟和花色苷合成的调控。     

干旱胁迫下水杨酸、脱落酸和海藻提取物对苹果幼苗抗旱性及养分吸收的影响

以盆栽苹果砧木M9T337矮化幼苗为试材,研究了水杨酸(SA)、脱落酸(ABA)和海藻提取物(SE)对重度干旱胁迫下苹果幼苗生长、光合色素、抗氧化酶活性、光合特性、渗透调节物质以及养分吸收的影响。结果表明:重度干旱胁迫显著抑制了苹果幼苗的营养生长,降低了叶片抗氧化酶活性、渗透调节物质含量、光合作用及植株养分含量。喷施SA、ABA和SE均促进了干旱胁迫下幼苗的生长,提高了叶片光合色素含量以及净光合速率(Pn)、气孔导度(Gs)、胞间CO2浓度(Ci)和蒸腾速率(Tr),同时也提高了苹果叶片中超氧化物歧化酶(SOD)、过氧化物酶(POD)和过氧化氢酶(CAT)活性以及可溶性蛋白、脯氨酸等渗透调节物质的含量,降低了丙二醛(MDA)含量,有效缓解了干旱胁迫对植株造成的伤害。与SA和ABA相比,喷施SE在促进苹果幼苗营养生长、提高叶绿素a含量和叶片SOD和CAT活性、增加可溶性蛋白和脯氨酸含量、提高植株养分含量方面效果更显著。     

The proteome research of human lens epithelial cells by two-dimensional gel electrophoresis and mass spectrometry

AIM: To establish the techniques in the proteome research of human lens epithelial cells,including the techniques of two-dimensional electrophoresis and mass spectrometry.METHODS: Total protein of cultured human lens epithelial cells was extracted with two kinds of different methods.The proteins were separated using immobilized pH gradients 2-DE and visualized by silver staining.The digitized images obtained by GS-800 scaner were then analyzed with PDQuest software in order to establish the differential expression profiles.The differential expressed protein spots were cut from the gels using proteomework spot cutter and subjected to in-gel digestion with trypsin.The digested peptide separation was conducted by a Finnigan LCQ MS coupled with a Surveyor HPLC system. RESULTS: A high resolution and reproducible 2-ED image was successfully obtained.The maps of 2-DE showed that lens proteins were in the section of pH 4-7 which the relative molecular weight was 17-72 kD.Relative molecular weight of more abundant proteins was localized at 19-50 kD,as well as the isoelectric points were found to lie between PI 5-7.Two of these proteins were identified by mass spectrometry and database queries.CONCLUSION: A stable protein maps of human lens epithelial cells is constructed.The technique will be used in human lens research to characterize physiological processes and diseases.     

Effects of ethylene,abscisic acid and auxin on fruit abscission in water dropwort (Oenanthe stolonifera DC.)

Fruit abscission in water dropwort (Oenanthe stolonifera DC.) was investigated by measuring fruit removal force (FRF) affected by fruit ages and plant hormones. FRF decreased rapidly 30 days after anthesis (DAA). Ethylene evolution increased rapidly at 35 DAA and ABA concentration increased gradually after 20 DAA. ACC and ethephon decreased the FRF of fruit explants and the promoting effect of ACC was delayed by silver thiosulphate and cycloheximide, but not by norbornadiene and actinomycin D. ABA enhanced fruit abscission without increasing ethylene evolution. However, the observation that CoCl₂ delayed abscission induced by ABA indicates possible involvement of ethylene in the ABA effect. Treatment with NAA, fenoprop and IAA had no effect on 30 DAA fruit explants, but IAA delayed abscission of 25 DAA explants. This suggests that the effect of auxins may differ with fruit age when used as control agents.     

高效液相色谱-串联质谱法测定葡萄和土壤中噻苯隆残留消解动态

【目的】评价噻苯隆在葡萄上的使用安全性。【方法】采用高效液相色谱-串联质谱法建立快速准确的噻苯隆在葡萄及土壤中的残留分析方法,进行2 a两地的消解动态及最终残留试验。【结果】建立的噻苯隆在葡萄及土壤中的残留分析方法表明:噻苯隆在0.01~1.00 mg·L-1内线性关系良好,灵敏度、准确度均符合检测要求。采用该方法研究噻苯隆在葡萄中的消解动态,对葡萄及果园土壤的最终残留进行了测定,结果表明,噻苯隆在葡萄样品中的消解曲线均符合一级动力学方程,半衰期为0.8~5.7 d,最终残留试验表明,在到达安全间隔期14 d时噻苯隆在葡萄中最高残留量为0.020 0mg·kg-1,远低于国家限量标准。【结论】噻苯隆在葡萄中属于易降解农药,降解较快。     

Influence of high concentrations of cytokinins on the production of somatic embryos by germinating seeds of tomato,aubergine and pepper

Summary

Somatic embryos of tomato, aubergine and pepper were initiated from intact seedlings when seeds were cultured on medium containing 6-benzylaminopurine (BAP) or thidiazuron (TDZ). The percentage of explants producing somatic embryos was highest for aubergine on media containing low concentrations of BAP, i.e. 0–10 mg l^–1, for tomato at 15–20 mg l^–1 and for pepper at 40–80 mg l^–1. Aubergine and tomato produced fewer somatic embryos per responsive seedling when cultured on medium containing TDZ, and pepper did not produce any somatic embryos on media containing 0–20 mg l^–1 1 TDZ. Morphogenesis of the seedlings producing somatic embryos was similar for all the genotypes, i.e. the seedlings were dwarf, only the cotyledons expanded, development of the apical meristem and the root were suppressed and a ring-like crown of nodular tissue developed at the base of the hypocotyl from which somatic embryos were initiated. Co-cultivation of tomato and aubergine seeds with seeds of pepper in media containing 0, 5, 10, 15 and 20 mg l^–1 BAP inhibited somatic embryogenesis in tomato and aubergine instead of assisting somatic embryogenesis in pepper. This is discussed in relation to the recent findings for the induction of somatic embyrogenesis in peanut (Arachis hypogea L.) and the role of BAP and TDZ.     

Mechanism of acylation stimulating protein resistance induced by fatty acid in 3T3-L1 adipocytes and preadipocytes

AIM: To evaluate the potential acylation stimulating protein (ASP) resistance in both adipocytes and preadipocytes under the conditions by which insulin resistance is produced by the stimulation of free fatty acids (FFA), and to explore the mechanism of ASP resistance on post-receptor level. METHODS: 3T3-L1 preadipocytes were induced to differentiate. Then the cells were treated with oleate or palmitate at concentration of 0 mmol/L (FFA-free DMEM/F12), 0.125 mmol/L, 0.5 mmol/L or 1.0 mmol/L overnight. Glucose transport was assessed by ［3H］ 2-deoxyglucose uptake to evaluate insulin resistance and ASP resistance. Both non-FFA treated and FFA treated 3T3-L1 cells were cultured with ASP at concentration of 5.0 μmol/L for 4 h, then the cell proteins were extracted, and the expressions of guanine nucleotide binding protein beta (Gβ), guanine nucleotide-binding protein alpha-q/11(Gαq/11), phosphorylated-protein kinase Cα (p-PKCα) and phosphorylated-protein kinase Cζ (p-PKCζ) were measured by Western blotting. RESULTS: Both adipocytes and preadipocytes were responsive to ASP. ASP stimulation increased glucose transport by 198% in adipocytes and by 287% in preadipocytes (P<0.01 vs PBS). FFA at concentration of 0.125 mmol/L did not change ASP-stimulated glucose transport significantly, but high dose of oleate or palmitate effectively reduced the ASP response with a significant reduction by 47% (P<0.05 for oleate) and 34% (P<0.05 for palmitate) at 1 mmol/L FFA in adipocytes. Similarly in preadipocytes, glucose uptake rates were decreased by 43% (P<0.05 for oleate) and 62% (P<0.01 for palmitate) at 1 mmol/L FFA. Effects were comparable to those obtained with insulin. After overnight incubation with oleate or palmitate in adipocytes and preadipocytes, Gβ, Gαq/11, p-PKCα and p-PKCζ were downregulated both in the absence of ASP treatment and in the presence of ASP treatment in adipocytes. At concentration of 1.0 mmol/L, oleate inhibited the expressions of ASP-induced Gβ, Gαq/11, p-PKCα and p-PKCζ in adipocytes by 47％, 44%, 39% (P<0.05, P<0.01) and 20% (P>0.05), respectively. Palmitate also effectively blocked the expressions of ASP (at concentration of 1.0 mmol/L)-induced Gβ, Gαq/11, p-PKCα and p-PKCζ by 50%, 43%, 44% and 43% (P<0.05, P<0.01) in adipocytes. In preadipocytes, oleate only inhibited ASP-induced p-PKCα and p-PKCζ significantly by 39% and 19%, respectively (P<0.05). However, overnight exposure of 3T3-L1 preadipocytes to 1 mmol/L palmitate leaded to 45%, 50%, 52% and 21% (P<0.05, P<0.01) inhibition of ASP-induced expressions of Gβ, Gαq/11, p-PKCα and p-PKCζ, respectively. CONCLUSION: Oleate and palmitate inhibit ASP-mediated stimulation of glucose transport both in adipocytes and preadipocytes. The study provides direct evidence of ASP resistance under the condition of insulin resistance induced by FFA in a cellular model. The mechanism of action involves both changes in expression of C5L2 as well as signaling parameters. Fatty acid-induced ASP resistance may contribute to the physiological abnormalities associated with insulin resistance and obesity phenotype.     

Molecular cloning of three malic acid related genes MdPEPC, MdVHA-A, MdcyME and their expression analysis in apple fruits

Malic acid (MA) in apple fruit is the predominant organic acid associated with taste, flavour and juice quality. In this study, three full-length cDNAs of MdPEPC, MdVHA-A and MdcyME were cloned from apple fruit. They encoded cytosolic phosphoenolpyruvate carboxylase (MdPEPC, EC 4.1.1.31), subunit A of vacuolar H⁺-ATPase (MdVHA, EC 3.6.1.3) and cytosolic NADP-dependent malic enzyme (MdcyME, EC 1.1.1.40), respectively, for MA synthesis, transportation and degradation. Real-time quantitative PCR discovered that the expression levels of three genes varied with development stages, and that their expression patterns differed between low acid (LA) and high acid (HA) genotypes. In addition, enzyme activity assay showed that PEPC and VHA contributed positively to MA accumulation during fruit development both in LA and HA, while cyME did negatively.     

Models for predicting the mass of apricot fruits by geometrical attributes (cv. Shams,Nakhjavan, and Jahangiri)

Fruits with the similar weight and uniform shape are in high demand in terms of marketing value. Therefore, an awareness of grading fruit based on weight is crucial. A part of this research was aimed to present some physical properties of three Iranian apricot cultivars (Shams, Nakhjavan, and Jahangiri). In addition, apricot mass was predicted by different physical characteristics with linear and nonlinear models as three different classifications: (1), single or multiple variable regressions of apricot dimensional characteristics. (2), single or multiple variable regressions of apricot projected areas and (3), estimating apricot mass based on its volume. All properties considered in the current study were found to be statistically significant at the 1% probability level. The highest and the lowest dimensional characteristics were found for Jahangiri and Nakhjavan cultivars, respectively. Sphericity values had significant difference among the tested cultivars. The latter property values were 0.971, 0.917, and 0.973 for Shams, Nakhjavan, and Jahangiri cultivars, respectively. Based on the results, the surface area of the Jahangiri cultivar was found to be 6458.35 mm², followed by the Shams and Nakhjavan cultivars, which had a mean of 6115.36, and 4395.25 mm², respectively. In this paper, the coefficients of static friction of the cultivars on three different surfaces are also reported. The results showed that mass modeling of apricot based on minor diameter and three projected areas are the most appropriate models in the first and the second classifications, respectively. In third classification, the best model was obtained on the basis of the actual volume as M = 0.997V_m + 0301, R² = 0.98, R.S.E. = 1.711 with R² = 0.98. It was concluded that the suitable grading system of apricot mass is based on minor diameter as nonlinear relation: M = 0.0019c^2.693, R² = 0.96, R.S.E. = 2.2.     

Rapid metabolic profiling of <Emphasis Type="Italic">Nicotiana tabacum</Emphasis> defence responses against <Emphasis Type="Italic">Phytophthora nicotianae</Emphasis> using direct infrared laser desorption ionization mass spectrometry and principal component analysis

Background  

Successful defence of tobacco plants against attack from the oomycete Phytophthora nicotianae includes a type of local programmed cell death called the hypersensitive response. Complex and not completely understood signaling processes are required to mediate the development of this defence in the infected tissue. Here, we demonstrate that different families of metabolites can be monitored in small pieces of infected, mechanically-stressed, and healthy tobacco leaves using direct infrared laser desorption ionization orthogonal time-of-flight mass spectrometry. The defence response was monitored for 1 - 9 hours post infection.     

Changes of IL-4, IL-10, IL-12 levels in rat serum and lung during acute respiratory distress syndrome induced by oleic acid

AIM:To observe the dynamic changes of interleukin-4(IL-4), IL-10, IL-12 in rat serum and lung tissues during acute respiratory distress syndrome (ARDS).METHODS:The ARDS model of rats was induced by intravenous injection of oleic acid. The levels of IL-4, IL-10, IL-12 in serum and the supernatant of lung tissues were measured by enzyme linked immunosorbent assay (ELISA).RESULTS:The Levels of serum and lung IL-10, IL-12 in ARDS rats were increased in 4 h, 8 h, 16 h group compared with control group. The levels in IL-10 in serum in 16 h group and IL-10 in lung tissues of 8 h group were lower than that in 4 h group. The Levels of IL-4 in serum in 4 h, 8 h group were higher than that in control group, while IL-4 in 16 h group was lower than that in 8 h group. IL-4 of lung tissues in 4 h, 8 h, 16 h group were increased significantly, but in 16 h group were lower than that in 8 h group. The biggest changes of pulmonary coefficient and histopathology were observed at 4 h after injection of oleic acid.CONCLUSIONS:IL-4, IL-10 and IL-12 might play important roles in inflammatory reaction induced by oleic acid. The pro-and anti-inflammatory cytokines produced successively during ARDS.The relationship between unbalanced cytokines and lung injury in ARDS needs to be further studied.     

Promotion of seed germination and subsequent seedling growth of loquat (Eriobotrya japonica,Lindl) by moist-chilling and GA3 applications

The germination of loquat seeds faces certain problems. The present research was designed to study the promotion of the germination of loquat seeds by moist-chilling and GA₃ applications. The results showed that loquat seeds display an endogenous dormancy that can be released by moist-chilling treatment for a certain period. In this respect, the best treatment was moist-chilling for 3 weeks at 5 ± 1 °C or 1 week of moist-chilling followed by soaking in 250 ppm GA₃ solution for 20 h. These treatments significantly increased germination percentage (88 and 85%, respectively) and decreased time to 50% germination (T50) (31.5 and 40.7 days, respectively) compared to control (51% and 56 days, respectively). Also, the characteristics of the obtained seedlings were much better than the control seedlings. In addition, the 3-week moist-chilled seeds contained the highest soluble protein concentration and were characterized by the synthesis of new protein band of 161.7 kDa that was absent in all other treatments. This treatment lead to the absence of five polypeptides bands (222.5, 201.5, 109.5, 71.1 and 49.3 kDa), which were synthesized in GA₃ treatment, and the presence of a higher number of polypeptide bands compared with those of other moist-chilling periods and the control treatments. However, increasing the moist-chilling period over 3 weeks significantly decreased both germination percentage and T50. The combination between GA3 and moist-chilling treatments produced differential effects on seed germination, soluble protein and the number of protein bands depending on the length of the moist-chilling period. Although GA3 application on un-chilled seeds resulted in more synthesis of protein bands than other tested treatments, it did not improve the germination process. The concentration of soluble inorganic phosphorus of the tested seeds was negatively (r = −0.57^*) and the concentration of soluble organic phosphorus positively (r = +0.49^*) correlated with the germination percentage. It was concluded that treatment of moist-chilling for 3 weeks or 1 week moist-chilling followed by 250 ppm GA₃ is recommended for promoting the germination process of loquat seeds and improving growth characteristics of the subsequent seedlings.     

Regulation by reactive oxygen species of matrix metalloproteinase-1, 3 and tissue inhibitor of matrix metalloproteinase-1 in smooth muscle cells

AIM: To understand whether reactive oxygen species promote the rupture of atherosclerotic plaques by regulating the balance of matrix metalloproteinase-1, 3 (MMP-1, 3) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) in smooth muscle cells. METHODS: Aortic smooth muscle cells from 4-6months-healthy abortive fetuses were incubated for 24 hours with xanthine (100 μmol/L) and xanthine oxidase (5 U/L) in vitro . MMP-1, 3 and TIMP-1 in the concentrated culture media were measured by Western blotting ( n =3 independent experiments). RESULTS: Incubation with xanthine/xanthine oxdiase decreased the amount of MMP-1 in the aortic smooth muscle cells (21.2%±5.5% of the control group), and pro-MMP-1 was activated completely. Reactive oxygen species (ROS) also activated pro-MMP-3, and increased the production of MMP-3 in the aortic smooth muscle cells. On the other hand, ROS inhibited the production of TIMP-1 in the aortic smooth muscle cells. CONCLUSION: It is complicated that ROS regulates the balance of MMPs and TIMPs. ROS may contribute to matrix degradation and the rupture in the atherosclerotic plaques.     

miR-556-3p regulates viability,migration and invasion of endometrial cancer cells by targeting SASH1

AIM To investigate whether microRNA-556-3p （miR-556-3p） regulates the viability, migration and invasion of endometrial cancer cells by targeting SASH1 gene. METHODS The expression of miR-556-3p, and the mRNA and protein levels of SASH1 in endometrial cancer tissues were detected by RT-qPCR and Western blot. Anti-miR-556-3p or pcDNA-SASH1 was transfected into endometrial cancer Ishikawa cells. The cell viability was detected by MTT assay, the migration and invasion abilities of the cells were detected by Transwell chamber method, and the protein expression levels of cyclin D1, p21, matrix metalloproteinase-2 （MMP-2） and MMP-9 were detected by Western blot. StarBase prediction and dual-luciferase reporter experiments were used to analyze the targeting relationship between miR-556-3p and SASH1. Anti-miR-556-3p and si-SASH1 were co-transfected into the Ishikawa cells, and their effects on cell viability, migration and invasion were examined by the methods described above. RESULTS Compared with adjacent tissues, the expression of miR-556-3p in endometrial cancer tissues was increased significantly, and the expression of SASH1 at mRNA and protein levels was decreased significantly （P<0.05）. Inhibition of miR-556-3p expression or induction of SASH1 over-expression obviously reduced the viability of Ishikawa cells, the number of migratory cells, the number of invasive cells and the protein levels of cyclin D1, MMP-2 and MMP-9, and dramatically increased the protein level of p21 （P<0.05）. miR-556-3p targeted SASH1 and negatively regulated its expression. Knock-down of SASH1 expression reversed the inhibitory effect of miR-556-3p expression inhibition on the viability, migration and invasion of Ishikawa cells. CONCLUSION Inhibition of miR-556-3p expression suppresses the viability, migration and invasion of endometrial cancer cells. The mechanism is related to the regulation of its target gene SASH1.     

miR-140-3p inhibits viability,invasion and migration of non-small-cell lung cancer A549 cells by targeting PD-L1

AIM: To explore the target relationship between microRNA-140-3p (miR-140-3p) and programmed cell death ligand 1 (PD-L1) and their effect on the viability, migration and invasion of non-small-cell lung cancer A549 cells.METHODS: RT-qPCR was used to detect the miR-140-3p expression in HLF-1, A549 and H1299 cells, and then the A549 cells with the most significant difference were selected as the subsequent research object. TargetScan software and dual-luciferase reporter assay were performed to predict and confirm the target relationship between miR-140-3p and PD-L1. RT-qPCR and Western blot were used to determine the effects of miR-140-3p mimic and inhibitor on PD-L1 expression level. MTT assay was used to detect the viability of A549 cells. Transwell assay was performed to detect the migration and invasion abilities of the A549 cells.RESULTS: miR-140-3p was significantly down-regulated in the A549 cells and H1299 cells (P<0.05). Transfection with miR-140-3p mimic decreased the expression of PD-L1 and inhibited the viability, migration and invasion of the A549 cells. Transfection with pcDNA3.0-PD-L1 reversed the inhibitory effect of miR-140-3p on the viability, migration and invasion of the A549 cells.CONCLUSION: miR-140-3p inhibits the viability, migration and invasion of A549 cells by targeting PD-L1.     

Fluid-phase endocytosis in Citrus juice cells is independent from vacuolar pH and inhibited by chlorpromazine,an inhibitor of PI-3 kinases and clathrin-mediated endocytosis

Summary

Compared to their more acidic relatives, sweet lime (Citrus limettioides Tan.) is distinguished by having a high vacuolar pH and a low sugar content. To determine whether variations in vacuolar pH are linked to different sugar uptake systems, resulting in contrasting sugar levels, we investigated developmental changes in V- (tonoplast-bound) and P-(plasmalemma-bound) ATPase activities, and the capacity of energised plasmalemma and tonoplast vesicles to take-up sucrose. Both plasmalemma- and tonoplast-bound ATPase activities, increased during fruit development. Artificially energised plasmalemma vesicles accumulated sucrose against a concentration gradient, whereas tonoplast vesicles did not show a similar uptake capacity, even in the presence of ATP. When juice cells were incubated with sucrose for 18 h with the endocytic marker, dextran-Texas Red (d-TR), an intense red fluorescence was observed in the vacuole. Sucrose-inducible uptake of d-TR was suppressed by the clathrin- and PI-3 kinase-mediated endocytosis inhibitor, chlorpromazine, but not by the caveolin-mediated endocytosis inhibitor, filipin. However, 3-dimensional Z-stack confocal images demonstrated the formation and presence of endocytic vesicles 0.5 – 2 µm in diameter, which were considerably larger than vesicles formed by clathrin-assisted scaffolding. The data indicated that: (i) most sucrose uptake into the vacuole is not mediated by a tonoplast-bound carrier; (ii) uptake of external solutes into the vacuole of C. limettioides juice cells is independent of vacuolar pH; and (iii) sucrose uptake occurs largely by a non-selective endocytic mechanism composed of vesicles, consistent with pinocytosis.     

Effect of HIPK2 gene on viability,apoptosis and JAK2/STAT3 signaling pathway in NRK-52E renal tubular epithelial cells induced by hypoxia and reoxygenation

AIM: To investigate the effect of homeodomain-interacting protein kinase 2 (HIPK2) on the viabi-lity, apoptosis and JAK2/STAT3 signaling pathway in NRK-52E renal tubular epithelial cells induced by hypoxia and reoxygenation (H/R). METHODS: HIPK2 small interfering RNA (siRNA) was transfected into NRK-52E cells by Lipofectamine^TM 2000, and normal control group (control group) and negative control group (HIPK2-NC group) were set up. After H/R, the cell viability was measured by CCK-8 assay, the apoptotic rate and Ca²⁺ fluorescence intensity were analyzed by flow cytometry, and the protein levels of Ki67, cleaved caspase-3, caspase-12, Bcl-2, Bax, p-JAK2 and p-STAT3 were determined by Western blot. RESULTS: Compared with control group, the protein expression of HIPK2 in the NRK-52E cells was significantly decreased after transfection with HIPK2 siRNA (P<0.05). Compared with control group, the cell viability and the protein expression of Ki67 and Bcl-2 in H/R group were also significantly decreased, and the apoptotic rate, the Ca²⁺ fluorescence intensity and the protein levels of cleaved caspase-3, caspase-12, Bax, p-JAK2 and p-STAT3 were significantly increased (P<0.05). Compared with H/R group, the cell viability and the protein expression of Ki67 and Bcl-2 in HIPK2-siRNA+H/R group were significantly increased, while the apoptotic rate, the Ca²⁺ fluorescence intensity and the protein levels of cleaved caspase-3, caspase-12, Bax, p-JAK2 and p-STAT3 were significantly decreased (P<0.05). CONCLUSION: Inhibition of HIPK2 gene expression promotes H/R-induced growth of NRK-52E renal tubular epithelial cells, and reduces the apoptosis. The mechanism is related to down-regulating the JAK2/STAT3 signaling pathway.