维生素C联合替莫唑胺对胶质瘤细胞的毒性作用及其机制

目的:探讨维生素C(VC)联合替莫唑胺(TMZ)对胶质瘤细胞活力的毒性作用及其机制。方法:在体外条件下培养人胶质瘤细胞BMG-1和SHG44细胞,设对照组(不施加VC与TMZ)、TMZ组(0.2 mmol/L)、VC(0.5mmol/L)+TMZ(0.2 mmol/L)组,TMZ(0.2 mmol/L TMZ)+U0126(10μmol/L)组,每组实验重复3次。采用MTT实验检测细胞生存率;流式细胞术和Annexin V-FITC/PI染色检测细胞凋亡情况; ROS检测试剂盒检测活性氧簇(ROS)水平,Western blot检测与凋亡、自噬及ERK通路相关蛋白的表达。结果:与对照组比较,TMZ组胶质瘤细胞的存活率显著下降(P<0.05)。与TMZ组比较,VC+TMZ组胶质细胞瘤细胞的存活率显著下降(P<0.01),VC+TMZ组中细胞凋亡率显著升高,且Bax、Cleaved caspase-3及Cleaved PARP蛋白表达显著增加,Bcl-2表达显著降低,而ROS水平及细胞自噬率显著降低,LC3-Ⅱ/LC3-1表达显著降低,p62表达显著增加(P均<0.05)...     

Trypanosoma cruzi: effect of benznidazole therapy combined with the iron chelator desferrioxamine in infected mice

Iron chelators have been employed in various studies aimed at evaluating the relationship between the iron status of the host and the development of infection. In the present study, the effects of benznidazole (BZ) therapy in combination with the iron chelator desferrioxamine (DFO) on the development of infection in mice inoculated with Trypanosoma cruzi Y strain have been investigated. Infected mice treated with DFO presented lower levels of parasitemia compared with infected untreated animals. Therapy with BZ for 21 days, with or without DFO, led to decreased parasitemia and reduced mortality, but BZ in combination with DFO treatment for 35 days (BZ/DFO-35) gave 0% mortality. All infected groups presented lower levels of iron in the liver, but serum iron concentrations were greater in DFO-35 and BZ/DFO-35, whereas hemoglobin levels were higher in BZ/DFO-35 and lower in DFO-35 compared with other treated groups. The percentage cure, determined from negative hemoculture and PCR results in animals that had survived for 60 days post-infection, was 18% for BZ and BZ/DFO-35, 42% for BZ combined with DFO for 21 days, and 67% for DFO-35. The results demonstrate that modification in iron stores increases BZ efficacy.     

缓释型促性腺激素释放激素与生长激素治疗儿童中枢性性早熟的疗效观察

目的用生长激素与促性腺激素释放激素类似物(Gonadotropin-releasing hormone analogues,GHA)联合治疗中枢性性早熟女性患儿对其最终成人身高的影响.方法生长激素(GH)与促性腺激素释放激素类似物(GHA)联合治疗4例中枢性性早熟女性患儿半年,对比治疗前后患儿的第二性征,骨龄发育,性激素及最终成人身高的变化.结果第二性征的发育停止,骨龄发育被控制,实际生活年龄与骨年龄的比值提高(平均0.79→0.84);血LH对促性腺激素释放激素的反应及血浆雌激素水平平均已降至青春期前,分别为(平均25.79±10.60mlu/ml→1.13±0.21mlu/ml)及(平均64.87±27.51pg/ml→3.03±1.87pg/ml);预测最终成人身高增加(平均149.60±4.31cm→156.75±3.84cm)差异具有显著性(P＜0.05).结论生长激素与GHA联合治疗中枢性性早熟患儿,不仅能抑制第二性征发育,而且能有效改善最终成人身高,无任何毒付作用.     

The effects of chronic estradiol treatment on working memory deficits induced by combined infusion of beta-amyloid (1-42) and ibotenic acid

Estrogen limits in vitro neuron death induced by application of beta-amyloid, the cytotoxic peptide linked to Alzheimer's disease. However, the ability of estrogen to protect neurons and preserve cognitive function in vivo following exposure to beta-amyloid has not been demonstrated. Our objective was to evaluate the potential of estrogen to reduce spatial working memory deficits in female rats induced by administration of a neurotoxic form of beta-amyloid in combination with the excitotoxin, ibotenic acid. The interaction of beta-amyloid with excitotoxic factors may underlie cognitive deficits associated with Alzheimer's disease. Therefore, to create an experimental model typical of early Alzheimer's disease a low dose of ibotenic acid was administered with beta-amyloid into the dorsal hippocampus. Ovariectomized rats were implanted subcutaneously with Silastic capsules that produce physiological levels of 17beta-estradiol 10 days before bilateral intrahippocampal injections of aggregated beta-amyloid (1-42) and ibotenic acid. Capsules remained in situ throughout behavioral testing. When tested 3-10 weeks after neurotoxin treatment, females without estrogen capsules exhibited delay-dependent impairments in working memory performance on a water maze and a radial arm maze. Females treated with estrogen and combined neurotoxins displayed working memory performance comparable to unlesioned females on both tasks. Neurotoxin treatment increased immunoreactivity for glial fibrillary acidic protein but this measure was unaffected by estradiol treatment indicating that estrogen did not limit glial proliferation. Results indicate that estrogen prevented deficits in spatial working memory induced by neurotoxin treatments intended to mimic the pathology of early Alzheimer's disease.     

N-acetyl-GLP-1: a DPP IV-resistant analogue of glucagon-like peptide-1 (GLP-1) with improved effects on pancreatic beta-cell-associated gene expression

Glucagon-like peptide-1(7-36)amide (GLP-1) is a key insulinotropic hormone with the reported potential to differentiate non-insulin secreting cells into insulin-secreting cells. The short biological half-life of GLP-1 after cleavage by dipeptidylpeptidase IV (DPP IV) to GLP-1(9-36)amide is a major therapeutic drawback. Several GLP-1 analogues have been developed with improved stability and insulinotropic action. In this study, the N-terminally modified GLP-1 analogue, N-acetyl-GLP-1, was shown to be completely resistant to DPP IV, unlike native GLP-1, which was rapidly degraded. Furthermore, culture of pancreatic ductal ARIP cells for 72 h with N-acetyl-GLP-1 indicated a greater ability to induce pancreatic beta-cell-associated gene expression, including insulin and glucokinase. Further investigation of the effects of stable GLP-1 analogues on beta-cell differentiation is required to assess their potential in diabetic therapy.     

Anti EGFR scFv::FTH1/FTH1纳米粒子的制备及其在哮喘治疗中的应用

哮喘作为常见的呼吸性疾病之一,严重危害人类的健康,造成了严重的医疗负担。最新研究表明表皮生长因子受体 (Epidermal growth factor receptor,EGFR) 参与哮喘的发生发展。为构建靶向EGFR用于哮喘治疗的纳米粒子,通过基因工程方法将抗EGFR的单链抗体 (Single chain antibody fragment,scFv) 修饰在铁蛋白重链亚基 (Human ferritin H-chain,FTH1) N端,采用混合复性的方式成功构建anti EGFR scFv::FTH1/FTH1纳米粒子。通过透射电子显微镜对纳米粒子的结构进行分析,结果表明,纳米粒子可以自组装成中空笼状结构,粒径约为12 nm。采用SDS-PAGE对纯化后的纳米粒子进行半定量分析发现,亚基物质的量之比为FTH1︰anti EGFR scFv::FTH1=7︰3。在哮喘小鼠动物模型上发现Anti EGFR scFv::FTH1/FTH1纳米粒子能有效抑制哮喘小鼠肺组织中杯状细胞增生及粘液分泌,对胶原沉积纤维化也有一定抑制效果。这些研究为铁蛋白应用于哮喘的治疗提供了良好的基础。     

Involvement of purinergic P2Y1R in antidepressant-like effects of electroacupuncture treatment on social isolation stress mice

Depression is a common neuropsychiatric disorder with high incidence and disability. Electroacupuncture (EA) is effective in the treatment of depression. However, the underlying mechanisms are not fully understood. Social isolation stress during post-weaning period can impair purinergic signaling in the brain of rodents and has emerged as a major risk factor for depression. The purpose of this study was to investigate the involvement of P2Y1 receptor (P2Y1R) in the antidepressant-like effects of EA. In this study, C57BL/6 mice were randomly assigned to group-housed (GH) or social isolated (SI) groups at post-natal day 21. After 6 weeks of social isolation, EA was performed on acupoints “Bai-hui” (GV20) and “Yin-tang” (GV29), or non-acupoints for 4 weeks. The SI mice received either intracerebroventricular injection of a selective P2Y1R agonist, MRS2365 (1 nmol); or a selective P2Y1R antagonist, MRS2179 (2 μmol), before and after EA. We found that SI mice exhibited depression-like behaviors accompanied with anxiety-like behaviors. The expressions of P2Y1R were well co-localized with GFAP-positive astrocytes and increased in the prefrontal cortex and hippocampus of SI mice. After treated with MRS2179, the depression-like behaviors of SI mice were attenuated, but not with MRS2365. Meanwhile, we found that EA could attenuate social isolation caused depression- and anxiety-like behaviors, and inhibited the up-regulation of P2Y1R in the prefrontal cortex and hippocampus of SI mice. Notably, the positive effects of EA on depression-like behaviors of SI mice could be reversed by MRS2365, while MRS2365 had no effect on the anxiolytic-like effects of EA. Therefore, we provide new evidence that EA could ameliorate depression- and anxiety-like behaviors in social isolation stress mice, and P2Y1R was involved in the antidepressant-like effects of EA.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11302-021-09827-1.     

长脉冲1064nm激光联合酮康唑软膏封包病甲治疗甲真菌病的临床疗效观察

目的观察长脉冲1064 nm激光联合酮康唑软膏封包病甲治疗甲真菌病的临床疗效。方法 将73例甲真菌病门诊患者随机分为两组:联合组(37例,病甲75个)采用长脉冲1064 nm激光联合酮康唑软膏封包病甲治疗;对照组(36例,病甲87个)单纯使用激光治疗。每周1次,连续治疗8周,治疗6个月后随访评估。比较两种方法治疗甲真菌病的临床有效率,观察1064 nm激光联合酮康唑软膏封包病甲治疗甲真菌病患者的临床疗效。结果 联合组治疗甲真菌病的总的临床有效率(74.67%)高于对照组(63.22%),3种临床类型在联合组的临床有效率均高于对照组,尤其近端甲下型(PSO)的临床有效率明显提高,但差异无统计学意义;当9≤SCIO≤15时,联合组治疗甲真菌病的临床有效率高于对照组,差异有统计学意义。结论 对于9≤SCIO≤15的甲真菌病患者,长脉冲1064 nm激光联合酮康唑软膏封包病甲治疗甲真菌病的临床疗效显著高于单纯激光治疗。     

AMP-activated protein kinase kinase: detection with recombinant AMPK alpha1 subunit

The AMP-activated protein kinase (AMPK) is a heterotrimeric serine/threonine protein kinase important for the responses to metabolic stress. It consists of a catalytic alpha subunit and two non-catalytic subunits, beta and gamma, and is regulated both by the allosteric action of AMP and by phosphorylation of the alpha and beta subunits catalyzed by AMPKK(s) and autophosphorylation. The Thr172 site on the alpha subunit has been previously characterized as an activating phosphorylation site. Using bacterially expressed AMPK alpha1 subunit proteins, we have explored the role of Thr172-directed AMPKKs in alpha subunit regulation. Recombinant alpha1 subunit proteins, representing the N-terminus, have been expressed as maltose binding protein (MBP) 6x His fusion proteins and purified to homogeneity by Ni(2+) chromatography. Both wild-type alpha1(1-312) and alpha1(1-312)T172D are inactive when expressed in bacteria, but the former can be fully phosphorylated (1 mol/mol) on Thr172 and activated by a surrogate AMPKK, CaMKKbeta. The corresponding AMPKalpha1(1-392), an alpha construct containing its autoinhibitory sequence, can be similarly phosphorylated, but it remains inactive. In an insulinoma cell line, either low glucose or 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) treatment leads to activation and T172 phosphorylation of endogenous AMPK. Under the same conditions of cell incubation, we have identified an AMPKK activity that both phosphorylates and activates the recombinant alpha1(1-312), but this Thr172-directed AMPKK activity is unaltered by low glucose or AICAR, indicating that it is constitutively active.     

Comparison of independent and combined chronic metabolic effects of GIP and CB1 receptor blockade in high-fat fed mice

GIP receptor antagonism with (Pro³)GIP protects against obesity, insulin resistance, glucose intolerance and associated disturbances in mice fed high-fat diet. Furthermore, cannabinoid CB₁ receptor antagonism with AM251 reduces appetite and body weight gain in mice. The present study has examined and compared the effects of chronic daily administrations of (Pro³)GIP (25 nmol/kg body weight), AM251 (6 mg/kg body weight) and a combination of both drugs in high-fat fed mice. Daily i.p. injection of (Pro³)GIP, AM251 or combined drug administration over 22 days significantly (P < 0.05 to <0.01) decreased body weight compared with saline-treated controls. This was associated with a significant (P < 0.05 to <0.01) reduction of food intake in mice treated with AM251. Plasma glucose levels and glucose tolerance were significantly (P < 0.05) lowered by 22 days (Pro³)GIP, AM251 or combined drug treatment. These changes were accompanied by a significant (P < 0.05) improvement of insulin sensitivity in all treatment groups. In contrast, AM251 lacked effects on glucose tolerance, metabolic response to feeding and insulin sensitivity in high-fat mice when administered acutely. These data indicate that chemical blockade of GIP- or CB₁-receptor signaling using (Pro³)GIP or AM251, respectively provides an effective means of countering obesity and related abnormalities induced by consumption of high-fat energy-rich diet. AM251 lacks acute effects on glucose homeostasis and there was no evidence of a synergistic effect of combined treatment with (Pro³)GIP.     

人工红树林湿地系统净化污水研究进展

咸淡水交接处这种特殊的生境造就了红树植物“抗污”和“降污”的功能,但能否将自然红树林湿地作为污水处理场所目前仍有争议。按照构建的人工红树林湿地的类型,分为三大类：模拟潮汐湿地系统、人工红树林污水处理系统以及红树林种植-养殖生态耦合系统。第一类系统模拟了自然海滩上的潮涨潮落的情形,第二类系统与一般人工湿地相类似,而第三类系统是指在养殖塘种植红树植物以去除有机物和营养盐。不同的类型反映了该领域研究工作的不断深入,展现了红树林湿地净化污水从理论探索向实际应用不断靠近的过程。应用人工红树林湿地净化污水具有较大的可行性,前景可观。     

Synthesis and BZR affinity of pyrazolo[1,5-a]pyrimidine derivatives. Part 1: study of the structural features for BZR recognition

Examination of the pharmacophoric points of the pyrazolo[1,5-a]pyrimidine derivatives, ligands for BZR, previously published led us to the design of a novel class of 3,6-diaryl-4,7-dihydro-pyrazolo[1,5-a]pyrimidin-7-ones and to determine the groups involved in the BZR recognition.     

Plant Sar1 isoforms with near-identical protein sequences exhibit different localisations and effects on secretion

In plants, differentiation of subdomains of the endoplasmic reticulum (ER) dedicated to protein export, the ER export sites (ERES), is influenced by the type of export-competent membrane cargo to be delivered to the Golgi. This raises a fundamental biological question: is the formation of transport intermediates at the ER for trafficking to the Golgi always regulated in the same manner? To test this, we followed the distribution and activity of two plant Sar1 isoforms. Sar1 is the small GTPase that regulates assembly of COPII (coat protein complex II) on carriers that transport secretory cargo from ER to Golgi. We show that, in contrast to a tobacco Sar1 isoform, the two Arabidopsis Sar1 GTPases were localised at ERES, independently of co-expression of Golgi-destined membrane cargo in tobacco cells. Although both isoforms labelled ERES, one was found to partition with the membrane fraction to a greater extent. The different distribution of fluorescent fusions of the two isoforms was influenced by the nature of an amino acid residue at the C-terminus of the protein, suggesting that the requirements for membrane association of the two GTPases are not equal. Furthermore, functional analyses based on the secretion of the bulk flow marker α-amylase indicated that over-expression of GTP-restricted mutants of the two isoforms caused different levels of ER export inhibition. These novel results indicate a functional heterogeneity among plant Sar1 isoforms.     

An antagonistic 5-HT receptor system in the auricles of the systemic heart complex of Sepia officinalis L. (Cephalopoda) shows 5-HT1 and 5-HT4 subtype properties

In pharmacological bioassays on isolated ring-shaped auricle preparations of Sepia officinalis, the action of the specific 5-hydroxytryptamine (5-HT) agonists 8-OH-DPAT (5-HT1a), CP-93129 (5-HT1b), TFMPP (5-HT1b) and RS-67333 (5-HT4) on these autonomously contractile compartments was studied. 8-OH-DPAT and CP-93129 induced mainly positive effects on frequency and tone on the isotonically suspended auricles. The positive effect of 8-OH-DPAT on frequency was blocked by the specific 5-HT1a antagonist NAN-190. The 5-HT1b agonist TFMPP caused similar effects on tone and a positive impact on the auricular amplitude. The highly specific 5-HT4 agonist RS-67333 induced an effect opposite to the action of 5-HT1 agonists inducing mainly negative effects on frequency, amplitude and tone, causing a diastolic standstill at a concentration of 10(-6) M. These negative effects were blocked by the adenylyl cyclase inhibitor SQ-22,536 in the absence of a diastolic standstill. The opposing action of 5-HT1 and 5-HT4 agonists on auricular contractile activity suggests that an antagonistic 5-HT-receptor system exists within the auricular myocardial cells of S. officinalis, probably consisting of 5-HT1- and 5-HT4-like subtypes. The results also suggest that adenylyl cyclase acts as the intracellular target enzyme of both signal transduction mechanisms.     

New metal complexes with 5-(1H-imidazol-4-ylmethyl)aminoisophthalic acid: Syntheses, structures, electrochemistry and electrocatalysis

Three new coordination polymers {[Cu(HL)(H₂O)]·H₂O}_n (1), [Ag(H₂L)]_n (2), and {[Co(HL)(phen)(H₂O)]·8H₂O}_n (3) [H₃L = 5-(1H-imidazol-4-ylmethyl)aminoisophthalic acid, phen = 1,10-phenanthroline] have been synthesized under hydrothermal conditions. The results of X-ray diffraction analysis revealed that complex 1 displays (3, 3)-connected 2D network with (4, 8²) topology, while complexes 2 and 3 have infinite 1D chain structure, in which one of the two carboxylic groups of H₂L⁻/HL²⁻ is uncoordinated. The 2D layers of 1 or the 1D chains of 2 and 3 are further linked together by hydrogen bonds and π-π interactions to form 3D supramolecular frameworks. Moreover, the electrochemical properties of complexes 1 and 2 have been studied by modified glassy carbon electrodes of 1 (Cu-GCE) and 2 (Ag-GCE), and the results indicate that the Cu-GCE and Ag-GCE show one-electron redox peaks. In addition, both Cu-GCE and Ag-GCE have good electrocatalytic activities toward the reduction of H₂O₂ in phosphate buffer (pH 5.5) solution.     

High spatio-temporal resolution measurement of A1R and A2AR interactions combined with Iem-spFRET and E-FRET methods

A₁R-A_2AR heterodimers regulate striatal glutamatergic neurotransmission. However, few researches about kinetics have been reported. Here, we combined Iem-spFRET and E-FRET to investigate the kinetics of A₁R and A_2AR interaction. Iem-spFRET obtains the energy transfer efficiency of the whole cell. E-FRET gets energy transfer efficiency with high spatial resolution, whereas, it was prone to biases because background was easily selected due to manual operation. To study the interaction with high spatio-temporal resolution, Iem-spFRET was used to correct the deviation of E-FRET. In this paper, A₁R and A_2AR interaction was monitored, and the changes of FRET efficiency of the whole or/and partial cell membrane were described. The results showed that activation of A₁R or A_2AR leads to rapid aggregation, inhibition of A₁R or A_2AR leads to slow segregation, and the interaction is reversible. These results demonstrated that combination of Iem-spFRET and E-FRET could measure A₁R and A_2AR interaction with high spatio-temporal resolution.     

Binding profiles of cholesterol ester transfer protein with current inhibitors: a look at mechanism and drawback

Although the pharmacological inhibition of cholesterol ester transport protein (CETP) has been proposed as a method of preventing and treating cardiovascular disease (CVD), the adverse effects of current inhibitors have cast doubt on the interaction mechanisms of inhibitors and CETP. In response, a molecular dynamics simulation was used to investigate their interaction and shed light on the lipid exchange mechanism of CETP. Results showed that torcetrapib, anacetrapib, and evacetrapib can induce the incremental rigidity of CETP, yet decrease the stability of Helix X and the hydrophobic tunnel of CETP, with passable binding abilities (ΔG_bind, ?61.08, ?64.23, and ?61.57 kcal mol^?1). During their binding processes, Van der Waals components (ΔE_vdw + ΔG_SA) play a dominant role, and the inhibitory effects closely correlated with residues Cys13, Val198, Gln199, Ser230, His232, and Phe263, which could reduce the flexibility of N- and C- termini and Helix X, as well as the stability of hydrophobic tunnel, into which the three inhibitors could enter and promote the formation of intramolecular H-bonds such as Thr138–Asn192 and Arg37–Glu186. Additionally, the three inhibitors could restrain the formation of an opening at the CETP N-terminal, which given the other findings suggests the tunneling mechanism of CETP transfer. The paper closes with an explanation of conceivable causes of the insufficient efficacy of the inhibitors, and puts forward the rationality in targeting the CETP distal end for CVD therapies.     

Detection of tetanus-induced effects in linearly lined-up micropatterned neuronal networks: application of a multi-electrode array chip combined with agarose microstructures

K-7174, a GATA-specific inhibitor, is a putative anti-inflammatory agent that attenuates effects of inflammatory cytokines in certain cell types. However, molecular mechanisms involved have not been elucidated. We found that, in glomerular podocytes, induction of monocyte chemoattractant protein 1 (MCP-1) and inducible nitric oxide synthase (iNOS) by TNF-alpha was abrogated by K-7174. It was correlated with unexpected induction of unfolded protein response (UPR) evidenced by: (1) induction of endogenous indicators 78 kDa glucose-regulated protein and CCAAT/enhancer-binding protein-homologous protein, and (2) suppression of an exogenous indicator, endoplasmic reticulum stress-repressive alkaline phosphatase. In podocytes, induction of UPR by either tunicamycin, thapsigargin, A23187 or AB5 subtilase cytotoxin completely reproduced the suppressive effect of K-7174. Furthermore, K-7174-elicited UPR abrogated induction of MCP-1 and iNOS not only by TNF-alpha but also by medium conditioned by activated macrophages. These results suggested a novel, UPR-dependent mechanism underlying the anti-inflammatory potential of K-7174.     

Preventive effect of several drugs againstPseudomonas aeruginosa infection and the toxicity of combined tumor necrosis factor with lipopolysaccharide: Relationship between lethality and the arachidonic cascade

The participation of tumor necrosis factor (TNF) and lipopolysaccharide (LPS) inPseudomonas aeruginosa (Pa) infection was examined. The lethal challenge of Pa or TNF and LPS injection could be prevented by pretreatment with anti-TNF antibody, polymyxin B, ONO 1078, or Shosaiko-to. The combined effects of TNF and LPS may be deeply related to the lethality of Pa infection. The activities of leukotriene(LT) C₄/D₄/E₄ or platelet activating factor (PAF) were also related to the lethality of Pa infection, probably due to the subsequently produced TNF which acts in combination with LPS. Activating the host defence mechanism with biological response modifiers like Chinese medicines was effective against Pa infection. One mechanism could involve an activity as an LT inhibitor or PAF antagonist. Following the administration of TNF and/or LPS, the serum levels of arachidonic cascade products underwent various changes. With a combination of TNF and LPS, there was a synergistic increment of prostaglandins, thromboxane, and LT. Following pretreatment with Shosaiko-to, suppression of LTs was dominant even with the combination of TNF and LPS, which might be related to the lethality of the infection or combined TNF with LPS.