A novel aged mouse model of recurrent intracerebral hemorrhage in the bilateral striatum

The current animal models of stroke primarily model a single intracerebral hemorrhage(ICH)attack,and there is a lack of a reliable model of recurrent ICH.In this study,we established 16-month-old C57 B L/6 male mouse models of ICH by injecting collagenaseⅦ-S into the left striatum.Twenty-one days later,we injected collagenaseⅦ-S into the right striatum to simulate recurrent ICH.Our results showed that mice subjected to bilateral striatal hemorrhage had poorer neurological function at the early stage of hemorrhage,delayed recovery in locomotor function,motor coordination,and movement speed,and more obvious emotional and cognitive dysfunction than mice subjected to unilate ral striatal hemorrhage.These findings indicate that mouse models of bilateral striatal hemorrhage can well simulate clinically common recurrent ICH.These models should be used as a novel tool for investigating the pathogenesis and treatment targets of recurrent ICH.

基于增广Lyapunov函数的干扰攻击不确定网络系统的有限时间控制

研究了具有结构不确定性和干扰攻击的网络系统的有限时间控制问题。通过构造一个新的增广Lyapunov函数,研究了系统有限时间稳定的充分条件。通过引入线性矩阵不等式方法,将充分条件归结为一个依赖于网络诱导时延和外部干扰界的可行性问题。然后,提出了状态反馈控制器的设计策略,使网络系统更适合工程实际。数值算例说明了该方法的有效性。

Polymorphisms of some cytokines and chronic hepatitis B and C virus infection

AIM： To study the relationship between the polymorphisms in some cytokines and the outcome of hepatitis B virus （HBV） and hepatitis C virus （HCV） infection. METHODS： Samples were obtained from 203 patients infected with HBV and/or HCV while donating plasma in 1987, and 74 controls were obtained from a rural area of North China. Antibodies to HBV or HCV antigens were detected by enzyme-linked imrnunoassay. The presence of viral particles in the serum was determined by nested reverse-transcriptase polymerase chain reaction （PCR）. Hepatocellular injury, as revealed by alanine aminotransferase （ALT） and aspartate aminotransferase level, was detected by a Beckman LX-20 analyzer. DNA was extracted from blood cells. Then, the single nucleotide polymorphisms of IL-2-330, IFN-γ＋874, IL-10-1082/-592 and IL-4-589 were investigated by restriction fragment length polymorphism-PCR or sequence specific primer-PCR.RESULTS： Persistent infection with HBV, HCV, and HBV/HCV coinfection was associated with IL-2-330 TT genotype and T allele, IFN-γ＋874 AA genotype, and IL-10-1082 AA genotype. The clinical outcome of HBV and/or HCV infection was associated with IL-2-330 TT genotype and T allele, IFN-γ＋874 AA genotype, and IL-10-1082 AA genotype. IL-2-330 GG genotype frequency showed a negative correlation with clinical progression, IL-10-1082 AA genotype frequency showed a positive correlation and IL-10-1082 AG genotype frequency showed a negative correlation with clinical progression. HCV RNA positive expression was associated with IL-10-1082 AA genotype and the A allele frequency. Abnormal serum ALT level was associated with IL-10-592 AC genotype frequency and IL-4-589 CC genotype, CT genotype, and the C allele. CONCLUSION： These results suggest that polymorphisms in some cytokine genes influence persistent HBV and HCV infection, clinical outcome, HCV replication, and liver damage.

Effect of artesunate supplementation on bacterial translocation and dysbiosis of gut microbiota in rats with liver cirrhosis

AIM: To evaluate the effect of artesunate(AS) supplementation on bacterial translocation(BT) and gut microbiota in a rat model of liver cirrhosis. METHODS: Fifty-four male Sprague-Dawley rats were randomly divided into a normal control group(N), a liver cirrhosis group(M) and a liver cirrhosis group intervened with AS(MA). Each group was sampled at 4, 6 and 8 wk. Liver cirrhosis was induced by injection of carbon tetrachloride(CCl4), intragastric administration of 10% ethanol, and feeding a high fat diet. Rats in the MA group were intragastrically administered with AS(25 mg/kg body weight, once daily). Injuries of the liver and intestinal mucosa were assessed by hematoxylineosin or Masson's trichrome staining. Liver index was calculated as a ratio of the organ weight(g) to body weight(g). The gut microbiota was examined by automated ribosomal intergenic-spacer analysis of fecal DNA. BT was assessed by standard microbiological techniques in the blood, mesenteric lymph nodes(MLNs), liver, spleen, and kidney. RESULTS: Compared to group N, the body weight was reduced significantly in groups M and MA due to the development of liver cirrhosis over the period of 8 wk. The body weight was higher in group MA than in group M. The liver indices were significantly elevated at 4, 6 and 8 wk in groups M and MA compared to group N. AS supplementation partially decreased the liver indices in group MA. Marked histopathologic changes in the liver and small intestinal mucosa in group M were observed, which were alleviated in group MA. Levels of pro-inflammatory interleukin-6 and tumor necrosis factor-α were significantly elevated at 8 wk in ileal homogenates in group M compared to group N, which were decreased after AS supplementation in group MA. The dysbiosis of gut microbiota indicated by the mean diversity(Shannon index) and mean similarity(Sorenson index) was severe as the liver cirrhosis developed, and AS supplementation had an apparent intervention effect on the dysbiosis of gut microbiota at 4 wk. The occurrence of BT was increased in the liver of group M compared to that of group N. AS supplementation reduced BT in group MA at 8 wk. BT also occurred in the MLNs, spleen, and kidney, which was reduced by AS supplementation. BT was not detected in the blood in any group.CONCLUSION: Dysbiosis of gut microbiota, injury of intestinal mucosal barrier and BT occurred as liver cirrhosis progressed, which might enhance inflammation and aggravate liver injury. AS may have other nonantimalarial effects that modulate gut microbiota,inhibit BT and alleviate inflammation, resulting in a reduction in CCl4, alcohol and high fat-caused damages to the liver and intestine.

PBX3 promotes migration and invasion of colorectal cancer cells via activation of MAPK/ERK signaling pathway

AIM: To investigate the role of pre-B-cell leukemia homeobox (PBX)3 in migration and invasion of colorectal cancer (CRC) cells.

Intermittent hypoxia is involved in gut microbial dysbiosis in type 2 diabetes mellitus and obstructive sleep apnea-hypopnea syndrome

BACKGROUND Obstructive sleep apnea(OSA)-hypopnea syndrome(OSAHS)has been recognized as a comorbidity of type 2 diabetes mellitus(T2DM);more than half of T2DM patients suffer from OSAHS.Intermittent hypoxia(IH)plays an important role in metabolic diseases,such as obesity and OSAHS,through various mechanisms,including altering the gut microecological composition and function.Therefore,it is important to study the role of gut microbiota in T2DM patients with OSAHS,which has a high incidence and is prone to several complications.AIM To assess whether IH is involved in altering the fecal microbiome in T2DM patients with OSAHS.METHODS Seventy-eight participants were enrolled from Henan Province People’s Hospital and divided into healthy control(HC,n=26),T2DM(n=25),and T2DM+OSA(n=27)groups based on their conditions.The fecal bacterial DNA of the research participants was extracted and subjected to 16S ribosomal RNA sequencing.The clinical indices,such as insulin resistance index,homocysteine(HCY)concentration,and the concentrations of inflammatory factors in the peripheral blood,were assessed and recorded.RESULTS Group T2DM+OSA had the highest apnea-hypopnea index(AHI)(2.3 vs 3.7 vs 13.7),oxygen desaturation index(0.65 vs 2.2 vs 9.1),HCY concentration(9.6μmol/L vs 10.3μmol/L vs 13.81μmol/L)and C-reactive protein(CRP)concentrations(0.3 mg/L vs 1.43 mg/L vs 2.11 mg/L),and lowest mean oxygen saturation(97.05%vs 96.6%vs 94.7%)among the three groups.Twelve and fifteen key differences in amplicon sequence variants were identified when comparing group T2DM+OSA with groups T2DM and HC,respectively.We found progressively decreased levels of Faecalibacterium,Eubacterium,and Lachnospiraceae,and an increase in the level of Actinomyces,which strongly correlated with the HCY,CRP,fasting plasma glucose,and hemoglobin A1c concentrations,AHI,mean oxygen saturation,and insulin resistance index in group T2DM+OSA(P<0.05).CONCLUSION For T2DM patients with OSAHS,IH may be involved in selective alterations of the gut microbiota,which may affect the pathophysiological development of T2DM and DM-related complications.

Site-directed mutagenesis identified the key active site residues of alcohol acyltransferase PpAAT1 responsible for aroma biosynthesis in peach fruits

The aroma of peach fruit is predominantly determined by the accumulation of γ-decalactone and ester compounds.A previous study showed that the biosynthesis of these aroma compounds in peach fruit is catalyzed by PpAAT1,an alcohol acyltransferase.In this work,we investigated the key active site residues responsible for γ-decalactone and ester biosynthesis.A total of 14 candidate amino acid residues possibly involved in internal esterification and 9 candidate amino acid residues possibly involved in esterification of PpAAT1 were assessed via site-directed mutagenesis.Analyses of the in vitro enzyme activities of PpAAT1 and its site-directed mutant proteins(PpAAT1-SMs)with different amino acid residue mutations as well as the contents of γ-decalactone in transgenic tobacco leaves and peach fruits transiently expressing PpAAT1 and PpAAT1-SMs revealed that site-directed mutation of H165 in the conserved HxxxD motif led to lost enzymatic activity of PpAAT1 in both internal esterification and its reactions,whereas mutation of the key amino acid residue D376 led to the total loss ofγ-decalactone biosynthesis activity of PpAAT1.Mutations of 9 and 7 other amino acid residues also dramatically affected the enzymatic activity of PpAAT1 in the internal esterification and esterification reactions,respectively.Our findings provide a biochemical foundation for the mechanical biosynthesis ofγ-decalactone and ester compounds catalyzed by PpAAT1 in peach fruits,which could be used to guide the molecular breeding of new peach species with more favorable aromas for consumers.

Efficacy of different antibiotics in treatment of children with respiratory mycoplasma infection

BACKGROUND Respiratory infections in children are common pediatric diseases caused by pathogens that invade the respiratory system.Children are considerably susceptible to Mycoplasma pneumoniae infection.There has been widespread clinical attention on treatment strategies for this disease.AIM To analyze the clinical efficacy of different antibiotics in treating pediatric respiratory mycoplasma infections.METHODS We included 106 children with a confirmed diagnosis of respiratory mycoplasma infection who were admitted to our hospital from April 2017 to July 2019 and grouped them using a random number table.Among them,53 children each received clarithromycin or erythromycin.The clinical efficacy of both drugs was evaluated and compared.We performed the multiplex polymerase chain reaction(MP-PCR)test and determined the MP-PCR negative rate in children after the end of the treatment course.We compared the incidence of toxic and side effects,including nausea,diarrhea,and abdominal pain;further,we recorded the length of hospitalization,antipyretic time,and drug costs.Additionally,we evaluated and compared the compliance of the children during treatment.RESULTS The erythromycin group showed a significantly higher total effective rate of clinical treatment than the clarithromycin group.MP-PCR test results showed that the clarithromycin group had a significantly higher MP-PCR negative rate than the erythromycin group.Moreover,children in the clarithromycin group had shorter fever time,shorter hospital stays,and lower drug costs than those in the erythromycin group.The clarithromycin group had a significantly higher overall drug adherence rate than the erythromycin group.The incidence of toxic and side effects was significantly lower in the clarithromycin group than in the erythromycin group(P<0.05).CONCLUSION Our findings indicate that clarithromycin has various advantages over erythromycin,including higher application safety,stronger mycoplasma clearance,and higher medication compliance in children;therefore,it can be actively promoted.

High Mixing Entropy Enhanced Energy States in Metallic Glasses

Owing to the nonequilibrium nature,the energy state of metallic glasses(MGs)can vary a lot and has a critical influence on the physical properties.Exploring new methods to modulate the energy state of glasses and studying its relationship with properties have attracted great interests.Herein,we systematically investigate the energy state,mixing entropy and physical properties of Zr–Ti–Cu–Ni–Be multicomponent high entropy MGs by experiments and simulations.We find that the energy state increases along with the increase of mixing entropy.The yield strength and thermal stability of MGs are also enhanced by high mixing entropy.These results may open a new door on regulation of energy states and thus physical properties of MGs.

Reply to“Comment on‘High Mixing Entropy Enhanced Energy States in Metallic Glasses'”

We thank Dr.Ristic and Dr.Babic for their interests in our work and for reminding us about their works on metallic glasses.Metallic glasses are a family of materials with disordered atomic packing structure,which makes it quite challenging to design new advanced metallic glasses and makes it charming to understand the relative physical mechanisms.

Value of increased fetal cardio thoracic ratio in predicting adverse pregnancy outcome in 11-13^(+6)weeks

Objective:To explore the value of increased fetal cardio thoracic ratio(CTR)in predicting adverse fetal pregnancy outcome during 11-13+6 weeks.Methods:The ultrasonographic features,chromosome or gene detection and the pregnancy outcome of 86 fetuses with increased CTR at 11-13+6 weeks’gestation were retrospectively analyzed.Fetuses were divided into non-structural malformation group(46 cases)and structural malformation group(40 cases)according to results of fetal ultrasound examination.The differences of CTR and nuchal translucency thickness(NT)between the two groups were compared by t test.The area under the curve(AUC)was calculated by constructing the receiver operating curve(ROC)to determine the best diagnostic threshold,sensitivity and specificity of CTR and NT in predicting chromosomal abnormalities.Results:All the 86 fetuses had serious problems such as chromosome or gene abnormalities,or structural malformations.The CTR and NT of the structural malformation group were significantly higher than those of the non-structural malformation group,and the difference was statistically significant(P<0.05).Through villous biopsy of 60 fetuses,47 cases(78.3%,47/60)with haemoglobin Bart’s disease,10 cases(16.7%,10/60)with chromosomal abnormalities were detected,and none abnormalities were detected in other 3 cases.According to the ROC curve,the AUC for CTR and NT to predict chromosomal abnormalities are 0.691 and 0.954,respectively,and the diagnostic cut-off values are 0.57 and 3.6 mm,respectively.The sensitivity is 60.0%and 100%,the specificity is about 79.6%and 85.7%,respectively.Follow-up showed that 5 cases of fetal were death intrauterine,and the remaining 81 cases were induced to labor.The specificity of CTR increase in predicting adverse pregnancy outcome was 100%.Conclusion:The increase of CTR in 11-13+6 weeks of gestation indicates that the fetus may have serious problems and poor clinical prognosis.Attention should be paid to the accurate evaluation of CTR in early pregnancy so as to provide reference for the prediction of fetal pregnancy outcome.

Surface vitrification of carbon by laser treatment: Insights of glass formation from molecular dynamics

Microstructural modification of carbon materials,such as carbon fibers(Cf)and pyrolytically deposited carbon,is important for engineering applications.However,the regulation of these materials is not an effortless task.To understand the impacts of thermal spikes from pulsed laser processing on the structural adaptation of amorphous carbon(a-C),we performed melt quenching by molecular dynamics(MD)simulations.Our results confirm that the vitrification behavior of carbon can be tuned by adjusting the cooling rate(R),which is controlled by the thermal spikes of laser processing.Moreover,we set up a two-step way to locate the critical cooling rate(R_(c))of monoatomic carbon,which is refined by the sharp change in the environmental similarity parameter.Using this novel technique,we demonstrate that the ordering degree and the local atomic motif can be largely modified by going across a bar of 100 K/ps,which is extracted as the critical cooling rate to ensure the complete amorphization of carbon.This approach provides a criterion for both experimentally processing and theoretically simulating a-C structures.Therefore,this work provides guidelines on how to tune the amorphous carbon structures of engineering materials and provides an outlook for the wonderland of amorphous carbon materials.

Friction and wear properties of PM cam materials with different boron contents

In this study,effects of B addition on the sintering densification,microstructure,hardness,friction and wear properties of sintered Fe-2.4C-4Cr-1Mo-0.5P-0.7Si-2.5Cu(in wt%)were investigated.In spite of the decreased sintered density,the addition of B changes the phase composition of the materials and their ratio.Moreover,hardness of either the matrix or the liquid solidification structure dramatically increases.These changes in micro structure result in higher friction coefficient and lower wear loss.It is observed that the addition of0.1 wt%B offers the optimum friction and wear properties with a running-in period of only 30 s and wear volume loss of 0.006 mm^(3) under the testing conditions.Such friction and wear properties are superior to those of the other two widely used cam materials,cast iron and 45 steel.

基于熵和剪切带开发新型非晶合金

非晶合金是非晶材料家族中的新成员,由于缺乏原子排列长程有序特征,具有独特的性能和广泛应用前景.然而,非晶合金的形成、成分设计和形变问题在该领域长期未得到解决,制约着新型非晶合金的高效开发.传统上,多组元非晶合金的成分设计主要遵循相图研究范式,以温度调控为出发点,确定低液相线的成分为落脚点,尤其强调深共晶成分.然而,对于缺乏相图的多组元体系则力所不及,亟需新的理论突破.本文综述了非晶转变中的热力学与动力学基本规律,重点探讨了热力学与动力学性质之间的关联性,尤其是梳理了材料熔化熵与熔点粘度、熔点等多个非晶形成经典参量之间的内在关联,提出并论证了熔化熵和振动熵为非晶形成的关键热力学参量,可有效指导高性能非晶合金的设计和表征.结合实际应用中对非晶材料的工程要求,讨论了化学成分调控对非晶合金剪切带的萌生密度和扩展稳定性的影响,并总结了在单调加载和疲劳加载条件下剪切带行为对非晶合金断裂过程的作用.最后展望了振动熵和构型熵之间争议问题,提出未来工作中需要发展具有物理意义玻璃形成能力指标和可靠数据库的需求,混合加载模式下的韧性以及疲劳和韧性之间的权衡等问题.本文为进一步建立高性能非晶材料成分设计路径提供了参考与指导.

Electrocatalytic characterization of Ni-Fe-TiO_(2) overlayers for hydrogen evolution reaction in alkaline solution

The Ni-Fe-TiO_(2) overlayers on mild steel strips were prepared by electrochemical deposition.The layers were characterized morphologically by confocal laser scanning microscopy and scanning electron microscopy(SEM) coupled with energy-dispersive spectroscopy(EDS)analysis.The layers exhibit a quasi-three-dimensional(3D)morphology in which the crystalline,TiO_(2),is embedded.Electrocatalytic activity of the Ni-Fe-TiO_(2) layers for the hydrogen evolution reaction(HER) was assessed by using pseudo-steady-state polarization curves and electrochemical impedance spectroscopy(EIS) in alkaline solution.The results were compared with the properties of Ni-Fe electrodes and used for determining the mechanism and kinetics of HER.In comparison with Ni-Fe electrodes,the synthesized Ni-Fe-TiO_(2) electrodes present higher catalytic activity for HER due to the increase in the real surface area and high intrinsic elec trocatalytic activity of titanium dioxide.The present study provides valuable insight for exploring practical applications of Ni-based alloys as hydrogen evolution electrodes.

Engineering a New Chloroplastic Photorespiratory Bypass to In crease Photosynthetic Efficiency and Productivity in Rice

Over the past few years, three photorespiratory bypasses have been introduced into plants, two of which led to observable in creases in photos yn thesis and biomass yield. However, most of the experiments were carried out using Arabidopsis under controlled environmental conditions, and the increases were only observed under low-light and short-day conditions. In this study, we designed a new photorespiratory bypass (called GOC bypass), characterized by no reducing equivalents being produced during a complete oxidation of glycolate into CO2 catalyzed by three rice-self-originating enzymes, i.e., glycolate oxidase, oxalate oxidase, and catalase. We successfully established this bypass in rice chloroplasts using a multi-gene assembly and transformation system. Transgenic rice plants carrying GOC bypass (GOC plants) showed significant increases in photosynthesis efficiency, biomass yield, and nitrogen content, as well as several other CO2-enriched phe no types under both greenhouse and field conditions .Grain yield of GOC plants varied depending on seeding season and was increased significantly in the spring. We further demonstrated that GOC plants had significant advantages under high-light conditions and that the improvements in GOC plants resulted primarily from a photosynthetic CO2-concentrating effect rather than from improved energy balance. Taken together, our results reveal that engineering a newly designed chloroplastic photorespiratory bypass could increase photosynthetic efficiency and yield of rice plants grown in field conditions, particularly under high light.

A Synthetic Photorespiratory Shortcut Enhances Photosynthesis to Boost Biomass and Grain Yield in Rice

Several photorespiratory bypasses have been introduced into plants and shown to improve photosynthesis by increasing chloroplastic C02 concentrations or optimizing energy balance.We recently reported that an engineered GOC bypass could increase photosynthesis and productivity in rice.However,the grain yield of GOC plants was unstable,fluctuating in different cultivation seasons because of varying seed setting rates.In this study,we designed a synthetic photorespiratory shortcut(the GCGT bypass)consisting of genes en-coding Oryza sativa glycolate oxidase and Escherichia coli catalase,glyoxylate carboligase,and tartronic semialdehyde reductase.The GCGT bypass was guided by an optimized chloroplast transit peptide that targeted rice chloroplasts and redirected 75% of carbon from glycolate metabolism to the Calvin cycle,identical to the native photorespiration pathway.GCGT transgenic plants exhibited significantly increased biomass production and grain yield,which were mainly attributed to enhanced photosynthesis due to increased chloroplastic C02 concentrations.Despite the increases in biomass production and grain yield,GCGT transgenic plants showed a reduced seed setting rate,a phenotype previously reported for the GOC plants.Integrative transcriptomic,physiological,and biochemical assays revealed that photosynthetic car-bohydrates were not transported to grains in an efficient manner,thereby reducing the seed setting rate.Taken together,our results demonstrate that the GCGT photorespiratory shortcut confers higher yield by promoting photosynthesis in rice,mainly through increasing chloroplastic C02 concentrations.

Laves Phase in 22Cr-27Ni-2Ti-Al Austenitic Valve Steel

The Thermo Calc software was employed to calculate the equilibrium phase of 22Cr-27Ni-2Ti-Al austenitic valve steel, and the Laves phase in this steel was investigated after solid solution and aging by means of scanning electron microscopy （SEM）, transmission electron microscopy （TEM） and X- ray diffraction （XRD）. Results show that the microstructure of the experimental steel consists of γ, γ′, Laves and NbC. I.aves phases mainly precipitate on grain boundaries, with granular and fibrous morphologies. The size of the granular Laves phase is about 50-600 nm, and the fihrous Laves phase is composed of short strip Laves phases which are 300-600 nm in length and 50 150 nm in width. These two morphologies of Laves phase coarsen with increasing aging temperature, and this steel possesses the highest strength and hardness at aging temperature of 760 ℃.

Hot Deformation Behavior of F6NM Stainless Steel

The hot deformation behavior of F6NM stainless steel was investigated by hot compression test in a Gleeble-1500D thermal-mechanical simulator. The flow strain-stress curves were obtained and the corresponding metallographic observation of this steel under different deformation conditions was also carried out. This steel exhibi- ted dynamic recrystallization （DRX） in the temperature range of 1 273- 1473 K and the strain rate range of 0.01- 0.1 s^-1. The activation energy for hot deformation was determined to be 457.91 kJ/mol, and the hot deformation equations were also established. The flow instability zone was determined and could be divided into two regions. The first one was located in the temperature range of 1 173- 1 348 K and the strain rate range of 0. 056-10 s^-1 , while the second one is in the temperature range of 1398-1448 K and the strain rate range of 1.25-10 s^-1. In the end, the optimum conditions for hot working were provided.

Microstructure and graphitization behavior of diamond/SiC composites fabricated by vacuum vapor reactive infiltration

To inhibit the graphitization of diamond under high temperature and low pressure, diamond/SiC composites were firstly fabricated by a rapid gaseous Si vacuum reactive infiltration process. The microstructure and graphitization behavior of diamond in the composites under various infiltration temperatures and holding time were investigated. The thermal conductivity of the resul- tant materials was discussed. The results show that the diamond-to-graphite transition is effectively inhibited at temperature of as high as 1600 ℃ under vacuum, and the substantial graphitization starts at 1700 ℃. The microstructure of those ungraphitized samples is uniform and fully densified. The inhibition mechanisms of graphitization include the isolation of the catalysts from diamond by a series of protective layers, high pressure stress applied on diamond by the reaction-bonded SiC, and the moderate gas-solid reaction. For the graphitized samples, the boundary between diamond and SiC is coarse and loose. The graphitization mechanism is considered to be an initial detachment of the bilayers from the diamond surfaces, and subsequently flattening to form graphite. The ungraphitized samples present higher thermal conductivity of about 410 W.m-1.K-1 due to the fine interfacial structure. For the graphitized samples, the thermal conductivity decreases significantly to 285 W.m-1.K-1 as a result of high interfacial thermal resistance.