β-羟基丁酸改善β淀粉样蛋白1-42诱导小鼠海马神经元HT22细胞的能量障碍

背景:阿尔茨海默病患者存在严重的脑能量障碍,近年来基于酮体干预的脑能量拯救策略在阿尔茨海默病的治疗中越来越受到重视。目的:探讨β-羟基丁酸能否改善β淀粉样蛋白1-42(β-amyloid protein 1-42,Aβ_(1-42))诱导的小鼠海马神经元HT22细胞能量障碍。方法:将HT22细胞分为4组,分别为对照组、β-羟基丁酸组、Aβ_(1-42)组、Aβ_(1-42)+β-羟基丁酸组。使用相应试剂盒检测HT22细胞的存活率、ATP水平、α-酮戊二酸脱氢酶活性、Na^(+)K^(+)-ATP酶活性、线粒体膜电位及活性氧水平。结果与结论:与对照组相比,Aβ_(1-42)组HT22细胞的存活率、ATP水平、α-酮戊二酸脱氢酶活性、Na^(+)K^(+)-ATP酶活性、线粒体膜电位均显著降低(P<0.05),活性氧水平显著升高(P<0.05)。与Aβ_(1-42)组相比,Aβ_(1-42)+β-羟基丁酸组HT22细胞的存活率、ATP水平、α-酮戊二酸脱氢酶活性、Na^(+)K^(+)-ATP酶活性、线粒体膜电位均显著升高(P<0.05),活性氧水平显著降低(P<0.05)。结果表明:β-羟基丁酸提高了线粒体生物能量功能和细胞存活率,最终改善了Aβ_(1-42)诱导的HT22细胞能量障碍。

Autophagy occurs within an hour of adenosine triphosphate treatment after nerve cell damage:the neuroprotective effects of adenosine triphosphate against apoptosis

After hypoxia, ischemia, or inflammatory injuries to the central nervous system, the damaged cells release a large amount of adenosine triphosphate, which may cause secondary neuronal death. Autophagy is a form of cell death that also has neuroprotective effects. Cell Counting Kit assay, monodansylcadaverine staining, flow cytometry, western blotting, and real-time PCR were used to determine the effects of exogenous adenosine triphosphate treatment at different concentrations （2, 4, 6, 8, 10 mmol/L） over time （1, 2, 3, and 6 hours） on the apoptosis and autophagy of SH-SY5Y cells. High concentrations of extracellular adenosine triphosphate induced autophagy and apoptosis of SH-SYSY cells. The enhanced autophagy first appeared, and peaked at 1 hour after treatment with adenosine triphosphate. Cell apoptosis peaked at 3 hours, and persisted through 6 hours. With prolonged exposure to the adenosine triphosphate treatment, the fraction of apoptotic cells increased. These data suggest that the SH-SY5Y neural cells initiated autophagy against apoptosis within an hour of adenosine triphosphate treatment to protect themselves against injury.

Inhibitory effects of extracellular adenosine triphosphate on growth of esophageal carcinoma cells

AIM： To study the growth inhibitory effects of ATP on TE-13 human squamous esophageal carcinoma cellsin vitro.METHODS： NTT assay was used to determine the inhibition of proliferation of ATP or adenosine （ADO） on TE-13 cell line. The morphological changes of TE-13 cells induced by ATP or ADO were observed under fluorescence light microscope by acridine orange （AO）/ethidium bromide （EB） double stained cells. The intemudeosomal fragmentation of genomic DNA was detected by agarose gel electrophoresis. The apoptotic rate and cell cycle after treatment with ATP or ADO were determined by flow cytometry.RESULTS： ATP and ADO produced inhibitory effects on TE-13 cells at the concentration between 0.01 and 1.0 mmol/L. The ICs0 of TE-13 cells exposed to ATP or ADO for 48 and 72 h was 0.71 or 1.05, and 0.21 or 0.19 mmol/L, respectively. The distribution of cell cycle phase and proliferation index （PI） value of TE-13 cells changed, when being exposed to ATP or ADO at the concentrations of 0.01, 0.1, and 1 mmol/L for 48 h. ATP and ADO inhibited the cell proliferation by changing the distribution of cell cycle phase via either G0/G1 phase （ATP or ADO, 1 mmol/L） or S phase （ATP, 0.1 mmol/L） arrest. Under light microscope, the tumor cells exposed to 0.3 mmol/L ATP or ADO displayed morphological changes of apoptosis. A ladder-like pattern of DNA fragmentation was obtained from TE-13 cells treated with 0.1-1 mmol/L ATP or ADO in agarose gel electrophoresis. ATP and ADO induced apoptosis of TE-13 cells in a dose-dependent manner at the concentration between 0.03 and 1 mmol/L. The maximum apoptotic rate of TE-13 cells exposed to ATP or ADO for 48 h was 16.63% or 16.9%, respectively.CONCLUSION： ATP and ADO inhibit cell proliferation, arrest cell cycle, and induce apoptosis of TE-13 cell line.

Adenosine triphosphate promotes locomotor recovery after spinal cord injury by activating mammalian target of rapamycin pathway in rats

The mammalian target of rapamycin （mTOR） pathway plays an important role in neuronal growth, proliferation and differentiation. To better understand the role of mTOR pathway involved in the induction of spinal cord injury, rat models of spinal cord injury were established by modified Allen＇s stall method and interfered for 7 days by intraperitoneal administration of mTOR activator adenosine triphosphate and mTOR kinase inhibitor rapamycin. At 1-4 weeks after spinal cord injury induction, the Basso, Beattie and Bresnahan locomotor rating scale was used to evaluate rat locomotor function, and immunohistochemical staining and western blot analysis were used to detect the expression of nestin （neural stem cell marker）, neuronal nuclei （neuronal marker）, neuron specific enolase, neurofilament protein 200 （axonal marker）, glial fibrillary acidic protein （astrocyte marker）, Akt, mTOR and signal transduction and activator of transcription 3 （STAT3）. Results showed that adenosine triphosphate-mediated Akt/mTOR/STAT3 pathway increased endogenous neural stem cells, induced neurogenesis and axonal growth, inhibited excessive astrogliosis and improved the locomotor function of rats with spinal cord injury.

Protective effect of exogenous adenosine triphosphate on hypothermically preserved rat liver

AIM:To clarify the protective effect of exogenous adenosine triphosphate(ATP)on hypothermically preserved rat livers. METHODS:Establishment of continuous hypothermic machine perfusion model,detection of nucleotides in hepatocytes with HPLC,measurement of activities of LDH and AST in the perfusate,observation of histopathological changes in different experiment groups,and autoradiography were carried out to reveal the underlying mechanism of the protective effect of ATP. RESULTS:The intracellular levels of ATP and EC decreased rapidly after hypothermic preservation in control group,while a higher ATP and EC level,and a slower decreasing rate were observed when ATP-MgCl_2 was added to the perfusate (P<0.01).As compared with the control group,the activities of LDH and AST in the ATP-MgCl_2 group were lower(P<0.05). Furthermore,more severe hepatocyte damage and neutrophil infiltration were observed in the control group.Radioactive [α-^(32)P]ATP entered the hypothermically preserved rat hepatocytes. CONCLUSION:Exogenous ATP has a protective effect on rat livers during hypothermical preservation.However,Mg^(2+) is indispensable,addition of ATP alone produces no protective effect.The underlying mechanism may be that exogenous ATP enters the hypothermically preserved rat liver cells.

Pre-treatment role of inosine triphosphate pyrophosphatase polymorphism for predicting anemia in Egyptian hepatitis C virus patients

AIM: To investigate and clarify, for the first time, the role of inosine triphosphate pyrophosphatase (ITPA ) polymorphism in Egyptian chronic hepatitis C virus (HCV) patients.METHODS:The human genomic DNA of all patients was extracted from peripheral blood cells in order to determine the single nucleotide polymorphism (SNP) of ITPA (rs1127354). SNP genotyping was performed by real time polymerase chain reaction (PCR, ABI TaqMan allelic discrimination kit) for 102 treatment-naive Egyptian patients with chronic HCV. All patients had no evidence of cardiovascular or renal diseases. They received a combination treatment of pegylated interferon α (PEG-IFNα) as a weekly subcutaneous dose plus an oral weight-adjusted dose of ribavirin (RBV). The majority received PEG-IFNα2a (70.6%) while 29.4% received PEG-IFNα2b. The planned duration of treatment was 24-48 wk according to the viral kinetics throughout the course of treatment. Pre-treatment liver biopsy was done for each patient for evaluation of fibrosis stage and liver disease activity. The basal viral load level was detected quantitatively by real time PCR while viral load throughout the treatment course was performed qualitatively by COBAS TaqMan assay. RESULTS: Ninety-three patients (91.2%) had ITPA SNP CC genotype and 9 (8.8%) had non-CC genotype (CA and AA). The percentage of hemoglobin (Hb) decline was higher for CC patients than for non-CC patients, particularly at weeks 4 and 8 (P=0.047 and 0.034, respectively). During the first 12 wk of treatment, CC patients had significantly more Hb decline > 3 g/dL than non-CC patients: 64.5% vs 22.2% at weeks 8 and 12, respectively, (P=0.024 and 0.038). Reduction of the amount of the planned RBV dose was significantly higher for CC patients than non-CC patients during the first 12 wk (18% ± 12.1% vs 8.5% ± 10.2%, P=0.021). The percentage of CC patients with RBV dose reduction was significantly greater than that of non-CC patients (77.4% vs 44.4%, P=0.044). Multivariate analysis identified only the percentage of RBV dose as a predictor for Hb decline. Platelet decline was significantly higher in non-CC patients than CC patients at weeks 12, 24 and 48 (P=0.018, 0.009 and 0.026, respectively). CONCLUSION: Rs1127354 ITPA polymorphism plays a decisive role in protecting against treatment-induced anemia and the need for RBV dose reduction in Egyptian HCV patients.

Efficient production of cyclic adenosine monophosphate from adenosine triphosphate by the N-terminal half of adenylate cyclase from Escherichia coli

In this study,we aimed at developing an efficient biocatalytic process for bio-production of cyclic adenosine monophosphate(c AMP)from adenosine triphosphate(ATP).First,adenylate cyclase from Escherichia coli MG1655(EAC)and Bordetella Pertussis(BAC)were expressed in E.coli BL21(DE3)and comparatively analyzed for their activities.As a result,EAC from E.coli MG1655 exhibited a higher activity.However,amount of EAC were obtained in an insoluble form.Therefore,we expressed the first 446 amino acids of EAC(EAC446)to avoid the inclusion body.The effects of induction temperature,incubation time,and incubation p H were further evaluated to improve the expression of EAC446.Subsequently,the reaction process for the production of c AMP with ATP as a starting material was investigated.As none of c AMP was detected in the whole-cell based biocatalytic process,the reaction catalyzed by the crude enzyme was determined for c AMP production.What's more,the reaction temperature,reaction p H,metal ion additives and substrate concentration was optimized,and the maximum c AMP production of 18.45 g·L^-1was achieved with a yield of 95.4%after bioconversion of 6 h.

Role of Deoxyribonucleoside Triphosphates(dNTPs)in Cell Transformation

Deoxyribenucleoside triphosphate (dNTP) pools were measured in normal BALB/c3T3 cells, transformation-treated cells and transformed cells with reverse-phase HPLC. The fluctuation of dNTP pools was similar after transformation treatment with alkylating mutagen glycidyl methacrylate(GMA) or Nmethyl- N'- nitro N- nitrosoguanidine (MNNG ). However,the gap between deoxyguanosine triphosphate + deoxyadenosine triphosphate (dGTP + dATP) pools and deoxythymidine triphosphate + deoxycytidine triphosphate (dTTP + dCTP) pools was greatly intensified. The measurements also indicated that the dNTP pools in transformed cells were quite different from those in normal cells. The results suggested that dNTP pools may play an important role in cell transformation

Symptomatic improvement in an acute, non-traumatic spine pain model with a combination of uridine triphosphate, cytidine monophosphate, and hydroxocobalamin

Rationale: In a previously published trial on spinal acute non-traumatic pain, peripheral neuro- regenerative combination of UTP, CMP and hydroxocobalamin presented unexpected analgesicproperties. Objective: To corroborate analgesiceffects of UTP, CMP and hydroxocobalamin combination in a self-paired evolutionary model. Methods: Mean VAS scores from pretreatment, V2 (5th treatment day) and V3 (10th treatment day) were plotted and statistically analyzed (ANOVA) for differences. PFQ scores from pretreatment, V2, and V3 were analyzed using the chisquare test. Results: The difference between V3 and pretreatment mean VAS scores was statistically significant (p < 0.0001). The improvement in PFQ scores throughout the study was found to be statistically significant (p < 0.0001). Conclusion: The combination of UTP, CMP and hydroxocobalamin seems to have analgesic properties in mediumterm use. The complex peripheral neu-roregenerative pharmacodynamics of this combination provides a plausible basis for this finding. Further randomized studies are needed to explore this combination for the indication of neuropathic pain due to spinal structure involvement.

Polyphosphatase PPN1 of Saccharomyces cerevisiae Is a Deoxyadenosine Triphosphate Phosphohydrolase

The Saccharomyces cerevisiae polyphosphatase PPN1 (uniprot/Q04119) degrades inorganic polyphosphates both by cleaving Pi from the chain end and by fragmenting long-chain polymers into shorter ones. In this study, we have found a new activity of this protein: it releases phosphate from dATP. The dATP phosphohydrolase activity of pure PPN1 was ~7-fold lower compared to the exopolyphosphatase activity. This activity was strongly stimulated by Co2+ ions, as well as by ammonium ions, and inhibited by heparin and pyrophosphate similar to the exopolyphosphatase activity of PPN1. The Km value for dATP was 0.88 ± 0.14 mM. The dATP phosphohydrolase activity in the cells of PPN1-overexpressing yeast strain was several-fold higher than that in the parent strain. The other exopolyphosphatase of S. cerevisiae, PPX1, did not split Pi from dATP.

Adenosine triphosphate induced cell death: Mechanisms and implications in cancer biology and therapy

Adenosine triphosphate(ATP)induced cell death(AICD)is a critical cellular process that has garnered substantial scientific interest for its profound relevance to cancer biology and to therapeutic interventions.This comprehensive review unveils the intricate web of AICD mechanisms and their intricate connections with cancer biology.This review offers a comprehensive framework for comprehending the multifaceted role of AICD in the context of cancer.This is achieved by elucidating the dynamic interplay between systemic and cellular ATP homeostasis,deciphering the intricate mechanisms governing AICD,elucidating its intricate involvement in cancer signaling pathways,and scrutinizing validated key genes.Moreover,the exploration of AICD as a potential avenue for cancer treatment underscores its essential role in shaping the future landscape of cancer therapeutics.

Changes in P2Y purinoreceptor-mediated intracellular calcium signal pathways results in inositol-1, 4, 5-triphosphate-sensitive calcium stores in rat small trigeminal ganglion neurons

BACKGROUND： Most of the currently available information on purinergic receptors （P2Rs） involved in pain transmission is based on results obtained in dorsal root ganglion or the spinal cord. However, the mechanism of P2Rs in trigeminal neuralgia remains unclear. OBJECTIVE： To investigate changes in the P2R-mediated calcium signaling pathway in nociceptive trigemJnal ganglion neurons. DESIGN, TIME AND SETTING： In vitro experiments were conducted at the Patch-Clamp Laboratory of Comprehensive Experiment Center of Anhui Medical University, China from September 2008 to June 2009. MATERIALS： Thapsigargin, caffeine, suramin, and adenosine 5＇-triphosphate were purchased from Sigma, USA. METHODS： Using Fura-2-based microfluorimetry, intracellular calcium concentration （[Ca^2＋]i） was measured in freshly isolated adult rat small trigeminal ganglion neurons before and after drug application. MAIN OUTCOME MEASURES： Fluorescent intensities were expressed as the ratio F340/F380 to observe [Ca^2＋]i changes. RESULTS： In normal extracellular solution and Ca^2＋-free solution, application of thapsigargin （1 μmol/L）, a sarcoplasmic reticulum Ca^2＋ pump adenosine 5＇-triphosphate inhibitor, as well as caffeine （20 mmol/L）, a ryanodine receptor agonist, triggered [Ca^2＋]i increase in small trigeminal ganglion neurons. A similar response was induced by application of adenosine 5＇-triphosphate （100 μmol/L）. In Ca^2＋-free conditions, adenosine 5＇-triphosphate-induced [Ca^2＋]i transients in small trigeminal ganglion neurons were inhibited in cells pre-treated with thapsigargin （P 〈 0.01）, but not by caffeine （P 〉 0.05）. In normal, extracellular solution, adenosine 5＇-triphosphate-induced [Ca^2＋]i transients in small trigeminal ganglion neurons were partly inhibited in cells pre-treated with thapsigargin （P 〈 0.05）. CONCLUSION： Inositol-1,4, 5-triphosphate （IP3）- and ryanodine-sensitive Ca^2＋ stores exist in rat nociceptive trigeminal ganglion neurons. Two pathways are involved in the purinoreceptor-mediated [Ca^2＋]i rise observed in nociceptive trigeminal ganglion neurons. One pathway involves the metabotropic P2Y receptors, which are associated with the IP3 sensitive Ca^2＋store, and the second pathway is coupled to ionotropic P2X receptors that induce the Ca^2＋ influx.

Nicotinamide Adenine Dinucleotide and Adenosine Triphosphate Oscillations Caused by Gradual Entry of Substrates within Mitochondria

Nicotinamide adenine dinucleotide (NAD) oscillation was observed when the isolated mitochondria were immersed in a pyruvate solution. In addition, when an adenosine diphosphate (ADP) was added to the mitochondrial suspension containing pyruvate, adenosine triphosphate (ATP) oscillation was observed as well as NADH oscillation. At this time, the pH within mitochondria also oscillated. It was found that the oscillatory reaction of NADH caused by the membrane permeation of pyruvate continues, causing the oscillation of NADH and H+ in the subsequent reactions. The pH oscillation led to the ATP oscillation. It is considered that the oscillatory reaction caused by the gradual entry of pyruvate into mitochondria was thought to be carried over to both the citric acid cycle and the respiratory chain, ultimately leading to the ATP oscillation in oxidative phosphorylation. Similarly, it was found that membrane permeation of malate causes the gradual occurrence of NADH, at which point NADH oscillates, followed by an oscillatory reaction of the respiratory chain, and finally ATP oscillation. It was found that the oscillations of NADH and ATP occur without going through the citric acid cycle. Oscillations of NADH and other intermediates in both the citric acid cycle and respiratory chain were also confirmed by experiments using semipermeable membranes. These results support our hypothesis that the gradual entry of the substrate by membrane permeation triggers an oscillatory reaction of the enzyme, which is also carried over to subsequent reactions.

Electroacupuncture improves neuropathic pain Adenosine, adenosine 5'-triphosphate disodium and their receptors perhaps change simultaneously

Applying a stimulating current to acupoints through acupuncture needles–known as electroacupuncture–has the potential to produce analgesic effects in human subjects and experimental animals. When acupuncture was applied in a rat model, adenosine 5-triphosphate disodium in the extracellular space was broken down into adenosine, which in turn inhibited pain transmission by means of an adenosine A1 receptor-dependent process. Direct injection of an adenosine A1 receptor agonist enhanced the analgesic effect of acupuncture. The analgesic effect of acupuncture appears to be mediated by activation of A1 receptors located on ascending nerves. In neuropathic pain, there is upregulation of P2X purinoceptor 3 （P2X3） receptor expression in dorsal root ganglion neurons. Conversely, the onset of mechanical hyperalgesia was diminished and established hyperalgesia was significantly reversed when P2X3 receptor expression was downregulated. The pathways upon which electroacupuncture appear to act are interwoven with pain pathways, and electroacupuncture stimuli converge with impulses originating from painful areas. Electroacupuncture may act via purinergic A1 and P2X3 receptors simultaneously to induce an analgesic effect on neuropathic pain.

肾移植患者ABCA1及GPIHBP1水平与术后血脂的相关性

目的分析尿毒症肾移植患者三磷酸腺苷结合盒亚家族A成员1(ABCA1)及糖基磷脂酰肌醇锚定高密度脂蛋白结合蛋白1(GPIHBP1)水平与术后血脂的相关性。方法回顾性分析2015年3月至2023年1月郑州大学第一附属医院肾移植科收治的97例尿毒症肾移植患者的临床资料,按照术后血脂情况分为血脂正常组(n=34)和血脂紊乱组(n=63),比较两组患者术后1个月、3个月、6个月、12个月的ABCA1、GPIHBP1、血脂(高密度脂蛋白胆固醇、低密度脂蛋白、甘油三酯及总胆固醇)水平和他克莫司血药浓度,采用Pearson相关分析法分析术后ABCA1及GPIHBP1与血脂水平及他克莫司血药浓度的相关性,并比较两组患者随访12个月内并发症的发生情况。结果血脂紊乱组患者术后1个月、3个月、6个月、12个月的ABCA1、GPIHBP1、总胆固醇、甘油三酯、低密度脂蛋白胆固醇及他克莫司血药浓度明显高于血脂正常组,差异均有统计学意义(P<0.05);血脂紊乱组患者术后1个月、3个月、6个月及12个月的高密度脂蛋白胆固醇为(1.05±0.14)mmol/L、(1.14±0.18)mmol/L、(1.18±0.16)mmol/L、(1.23±0.17)mmol/L,均低于同时间段血脂正常组的(1.17±0.21)mmol/L、(1.37±0.28)mmol/L、(1.42±0.24)mmol/L、(1.50±0.32)mmol/L,差异均有统计学意义(P<0.05)。尿毒症肾移植患者的ABCA1与总胆固醇、他克莫司血药浓度呈正相关(P<0.05),与高密度脂蛋白胆固醇呈负相关(P<0.05),而GPIHBP1与总胆固醇、甘油三酯、他克莫司血药浓度均呈正相关(P<0.05)。血脂紊乱组患者的并发症总发生率为49.21%,明显高于血脂正常组的23.53%,差异有统计学意义(P<0.05)。结论尿毒症肾移植患者ABCA1及GPIHBP1基因水平与他克莫司血药浓度及术后血脂水平有一定相关性,检测ABCA1及GPIHBP1基因水平能够为尿毒症肾移植患者术后管理提供一定依据。

建立检测猕猴三磷酸腺苷结合盒转运蛋白G2的mRNA相对表达水平的RT-qPCR方法

目的本研究旨在建立一种实时荧光定量PCR方法,用于检测猕猴三磷酸腺苷结合盒转运蛋白G2(adenosine triphosphate-binding cassette transporter protein G2,ABCG2)mRNA的基因转录水平。方法使用NCBI上GenBank数据库猕猴(Macaca mulatta)的ABCG2核苷酸序列号NM_001032919.1及内参GAPDH核苷酸序列号NM_001195426.1,借助Primer premier 5.0软件设计PCR引物。提取猕猴新鲜肾组织的总RNA,并反转录合成cDNA。接着,利用PCR引物进行实时荧光定量PCR扩增,并根据反应体系中荧光的变化情况定量分析ABCG2的mRNA相对表达水平。结果PCR产物测序结果显示,扩增的ABCG2和GAPDH核苷酸序列与NCBI上猕猴的序列同源性分别为90.91%和91.14%。ABCG2和GAPDH的扩增效率均达到80%~120%,实时荧光定量PCR标准曲线的熔解曲线为单峰,R2接近1。结论本研究建立的检测猕猴ABCG2 mRNA实时荧光定量检测方法,为研究高尿酸血症的发病机制以及新药开发奠定基础。

氧化石墨烯-DNA纳米探针用于三磷酸腺苷的检测与药物递送

癌细胞内三磷酸腺苷(ATP)浓度异常与肿瘤发生发展过程密切相关,因此,快速、准确地检测细胞内外ATP水平具有重要意义。盐酸阿霉素(DOX)是一种广泛使用的抗癌药物,能嵌入DNA碱基对,并通过抑制DNA复制和转录诱导细胞凋亡。氧化石墨烯(Graphene oxide, GO)由于具有毒性低、比表面积大和易功能化,可以有效、稳定地负载DNA纳米探针进入细胞等优点而被广泛应用。然而,复杂环境中的生物分子容易通过物理吸附竞争性结合到GO表面,导致假阳性信号。基于此,提出了一种新型的GO-DNA纳米探针,并将其应用于ATP的检测与抗癌药物DOX靶向递送。以ATP的核酸适配体与其互补链杂交形成双链DNA(dsDNA),并通过G-C碱基对负载DOX,利用互补链延伸的poly A序列吸附到GO表面构建了GO-dsDNA-DOX纳米探针,能极大程度地降低复杂环境中物理吸附引起的干扰,减少假阳性信号产生。ATP与适配体特异性结合会导致DOX释放,根据其荧光“off-on”实现ATP的定量分析,DOX荧光强度与ATP含量在0.08~8.0 mmol/L范围内呈现良好的线性关系,线性方程为IF=3.0897c+129.08,检测限(3σ/k)为0.059 mmol/L,且方法具有良好的选择性和抗干扰能力。细胞毒性及荧光成像结果表明,负载DOX的纳米探针在人乳腺癌细胞(MCF-7)内有明显的药物释放,显著诱导细胞凋亡。该研究建立了一种免修饰、简单和快速的ATP含量检测分析方法,并利用癌细胞内高浓度ATP实现靶向药物递送,降低了对正常细胞的毒副作用,为癌症的治疗提供了新思路。

ABCA3作为非小细胞肺癌的预后生物标志物及其与免疫细胞浸润的相关性

目的 本研究旨在基于生物信息学分析探讨ABCA3在非小细胞肺癌——肺腺癌(lung adenocarcinoma,LUAD)中的临床意义和生物学功能。方法 基于癌症基因组图谱(the Cancer Genome Atlas,TCGA),提取LUAD中ABCA3表达和相应的临床信息。使用R4.2.2软件进行统计分析。采用Wilcoxon符号秩和检验比较TCGA LUAD队列中肿瘤组织与对照组之间ABCA3的表达,以及临床病理参数与ABCA3表达水平之间的关系。根据ABCA3表达的中位数,将TCGA LUAD患者分为高表达组和低表达组。使用survival包和survminer包分析ABCA3与LUAD患者总生存(overall survival,OS)期和无进展生存(progression free survival,PFS)期的关系,并进行单因素和多因素Cox回归分析确定与LUAD预后相关的危险因素。使用rms包构建预测LUAD患者1年、2年和3年OS率的列线图模型。使用DESeq2包对高表达组和低表达组患者进行差异表达基因(differential expressed genes,DEGs)分析,并使用cluster Profil er包对DEGs进行基因本体(Gene Ontology,GO)数据库和基因组京都百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)富集分析。利用CIBERSORT算法预测LUAD肿瘤样本和正常样本中22种浸润性免疫细胞的丰度,并分析ABCA3表达与免疫浸润细胞和免疫检查点之间的相关性。结果 ABCA3基因在LUAD肿瘤组织中的表达均低于配对和非配对样本正常组织(P<0.05)。ABCA3高表达LUAD患者的OS (P=0.031)和PFS (P=0.044)明显高于低表达患者。T2分期(P=0.004)和T3分期(P=0.044)患者ABCA3的表达显著低于T1分期患者。多因素Cox回归分析显示,T分期、N分期和ABCA3表达是影响LUAD预后的独立影响因素(P<0.05)。基于ABCA3表达、T分期和N分期构建的列线图预测模型的C-index为0.86,提示模型与实际结果较为一致。校准曲线中的偏差修正线接近理想曲线,提示列线图模型的预测结果与观测结果较吻合。基因集富集分析显示ABCA3高表达组主要与花生四烯酸代谢、补体和凝血级联、肠道免疫网络免疫球蛋白A产生等有关。ABCA3的表达与NRP1、TRFRSF14、TNFSF15、CD40LG、HHLA2、CD28呈明显正相关,而与IDO1、CD70、TNFSF4、CD276和VTCN1呈明显负相关(P<0.05)。12种免疫浸润细胞在高表达组和低表达组存在明显差异(P<0.05)。此外,14种免疫浸润细胞与ABCA3表达呈现明显相关性(P<0.05),包括巨噬细胞、CD4+T细胞和树突细胞。结论 ABCA3的表达与LUAD患者预后和免疫细胞浸润有关,可能是预测非小细胞肺癌预后的有价值生物标志物。

二参真武汤对心肾阳虚型慢性心力衰竭大鼠心肌能量代谢的影响

目的:探讨二参真武汤对心肾阳虚型慢性心力衰竭(CHF)大鼠心肌能量代谢的影响。方法:60只大鼠随机选取8只设为空白组,剩余52只通过甲状腺切除后给予0.02%阿霉素腹腔注射建立CHF模型,将建模大鼠随机分为模型组、阳性对照组、二参真武汤低剂量组、二参真武汤高剂量组,每组8只,连续灌胃28 d。给药结束后,ELISA法测空白组、模型组血清三碘甲状腺原氨酸(T_(3))、甲状腺素(T_(4))、氨基末端B型利钠肽(NT-proBNP)和各组心肌组织腺嘌呤核苷三磷酸(ATP)、腺嘌呤核苷二磷酸(ADP)、B型脑钠肽(BNP)、肌酸激酶同工酶(CK-MB)、Na^(+)-K^(+)-ATP酶、Ca^(2+)-ATP酶。彩色多普勒心脏超声检测心功能;HE染色法观察心脏病理形态;天狼星染色法观察心肌胶原纤维沉积形态;电镜观察心肌线粒体超微结构。结果:造模后大鼠血清NT-proBNP上升,T_(3)、T_(4)下降(均P<0.01)。与空白组相比,模型组大鼠左心室射血分数(LVEF)、左心室短轴缩短率(LVFS)下降,左室收缩末期内径(LVIDs)、左心室收缩末期容积(LVESV)上升(均P<0.01)。ATP、ADP、Na^(+)-K^(+)-ATP酶、Ca^(2+)-ATP酶下降,BNP、CK-MB上升(均P<0.01)。心肌细胞肿胀,边缘模糊,胶原纤维沉积增多,线粒体数量减少、嵴模糊。与模型组相比,阳性对照组及二参真武汤低剂量组、二参真武汤高剂量组LVEF、LVFS上升、LVIDs、LVESV下降,ATP、ADP、Na^(+)-K^(+)-ATP酶、Ca^(2+)-ATP酶上升,BNP、CK-MB下降(均P<0.01);肿胀心肌细胞减少,胶原纤维沉积面积缩小,心肌线粒体损伤减轻。结论:二参真武汤可有效改善CHF大鼠心肌损伤,对心脏有保护作用,其机制可能是通过调节ATP、ADP改善心肌能量代谢。

亚慢性PM_(2.5)和O_(3)共同暴露对大鼠鼻黏膜ATP总量及ATP酶活性的影响

目的:探讨细颗粒物(fine particle matter,PM_(2.5))和臭氧(ozone,O_(3))亚慢性共同暴露对大鼠鼻黏膜组织腺嘌呤核苷三磷酸(adenosine triphosphate,ATP)总量及ATP酶活性的影响。方法:采用随机数字表法将20只雄性Sprague Dawley(SD)大鼠均分为对照组和暴露组,每组各10只,分别饲养于常规清洁级环境和本团队既往所搭建的大气污染物暴露系统中,连续暴露208 d。暴露期间,监测暴露系统内PM_(2.5)和O_(3)浓度,采用自测和站点数据相结合的方法对暴露系统内PM_(2.5)和O_(3)进行综合评估。在暴露第208天处死大鼠取心、肝、脾、肾、睾丸等主要器官和鼻黏膜组织,称量各脏器重量并计算脏器系数。利用所收集鼻黏膜组织,使用生物发光法测定ATP总量,使用分光光度法检测Na^(+)-K^(+)-ATP酶和Ca^(2+)-ATP酶活性。使用两独立样本t检验比较各指标组间差异。结果:自第3周开始至暴露结束,暴露组大鼠体质量高于对照组(P<0.05),两组间脏器系数差异无统计学意义。暴露组日均PM_(2.5)浓度为(30.68±19.23)μg/m3,O_(3)最大8 h浓度(O_(3)-8 h)为(82.45±35.81)μg/m3。暴露组大鼠鼻黏膜组织ATP化学发光值(792.4±274.1)IU/L低于对照组(1126.8±218.1)IU/L,鼻黏膜组织Na^(+)-K^(+)-ATP酶活性(1.53±0.85)U/mg低于对照组(4.31±1.60)U/mg(P<0.05)。对照组和暴露组鼻黏膜组织的蛋白含量分别为(302.14±52.51)mg/L和(234.58±53.49)mg/L,Ca^(2+)-ATP酶活性分别为(0.81±0.27)U/mg和(0.99±0.73)U/mg,组间差异均无统计学意义。结论:亚慢性PM_(2.5)和O_(3)共同暴露可能影响大鼠鼻黏膜组织供氧能力。