Effect of acetyl L-carnitine on human retinal pigment epithelium-19 cells in hypoxic conditions

AIM:To investigate the effect of acetyl-L-carnitine(ALCAR)on cell viability,morphological integrity,and vascular endothelial growth factor(VEGF)expression in human retinal pigment epithelium(ARPE-19)cells using a hypoxic model.METHODS:In the first set of experiments,the optimal CoCl_(2) dose was determined by exposing ARPE-19 cell cultures to different concentrations.To evaluate the effect of ALCAR on cell viability,five groups of ARPE-19 cell culture were established that included a control group,a sham group(200μM CoCl_(2)),and groups that received 1,10 and 100 mM doses of ALCAR combined with 200μM CoCl_(2),respectively.The cell viability was measured by MTT assay.The morphological characteristics of cells were observed by an inverted phase contrast microscope.The levels of VEGF and HIF-1α secretion by ARPE-19 cells were detected by enzyme linked immunosorbent assay(ELISA)assay.RESULTS:ARPE-19 cells were exposed to different doses of CoCl_(2) in order to create a hypoxia model.Nevertheless,when exposed to a concentration of 200μM CoCl_(2),a notable decrease in viability to 83% was noted.ALCAR was found to increase the cell viability at 1 mM and 10 mM concentrations,while the highest concentration(100 mM)did not have an added effect.The cell viability was found to be significantly higher in the groups treated with a concentration of 1 mM and 10 mM ALCAR compared to the Sham group(P=0.041,P=0.019,respectively).The cell viability and morphology remained unaffected by the greatest dose of ALCAR(100 mM).The administration of 10 mM ALCAR demonstrated a statistically significant reduction in the levels of VEGF and HIF-1α compared with the Sham group(P=0.013,P=0.033,respectively).CONCLUSION:The findings from the current study indicate that ALCAR could represent a viable therapeutic option with the potential to open up novel treatment pathways for retinal diseases,particular relevance for age-related macular degeneration(AMD).However,to fully elucidate ALCAR’s application potential in retinal diseases,additional investigation is necessary to clearly define the exact mechanisms involved.

Unveiling osteoprotective potential of biologically active naringenin in rats with dexamethasone-induced osteoporosis

Objective To investigate the protective effects of naringenin(NRG)against dexamethasone(DEX)-induced osteoporosis(OP)in rats.Methods Molecular docking of NRG was done with AutoDock Vina 1.2.0 software.Forty-eight female Wistar rats were randomly divided into six groups(n=8 each):normal control(NC),DEX(7 mg/kg,i.m.),NRG-low(NRG-L;25 mg/kg,i.g.),NRG-medium(NRG-M;50 mg/kg,i.g.),NRG-high(NRG-H;100 mg/kg,i.g.),and alendronate(ALN;0.25 mg/d,i.g.)groups.OP was induced by administering DEX once a week for five weeks in all groups except NC group.Begining in the third week after the initial DEX administration,the rats in NRG-L,NRG-M,NRG-H,and ALN groups received the corresponding treatments daily for three weeks,while NC and DEX groups received no additional treatment.Serum samples were collected at the end of the experiment for biochemical,bone turnover,antioxidant,lipid profile,and inflammatory cytokine analyses.Femur bones underwent physical parameter testing and histopathological examination.Results The molecular docking results illustrated that NRG docked with calcitonin(CT),lowdensity lipoprotein(LDL),bone morphogenetic protein(BMP),vascular endothelial growth factor(VEGF)receptor,forkhead transcription factors,and osteoprogenitor cells showed good binding energy.In rats administered with 25,50,and 100 mg/kg NRG,there was a significant enhancement in serum biochemical indices,characterized by a reduction in tartrate-resistant acid phosphatase(TRAP),parathyroid hormone(PTH),and an elevation in osteocalcin(OC)and CT levels(P<0.05,P<0.01,and P<0.001,respectively).Despite no significant changes in thickness,weight,and length(P>0.05),there was a marked increase in bone mineral density(BMD)(P<0.01,P<0.001,and P<0.001,respectively).Antioxidant enzyme markers showed significant upregulation,with higher glutathione,superoxide dismutase,and catalase,and a concurrent decrease in malondialdehyde(MDA)(P<0.05,P<0.01,and P<0.001,respectively).The lipid profile also improved significantly,with lower cholesterol(CH),triglycerides(TG),and low-density lipoprotein(LDL)levels,and an increase in high-density lipoprotein(HDL)level(P<0.05,P<0.01,and P<0.001,respectively).Inflammatory cytokine levels were reduced,as evidenced by decreases in tumor necrosis factor(TNF),interleukin(IL)-6,and IL-1β(P<0.05,P<0.01,and P<0.001,respectively).Furthermore,histological alterations revealed obvious improvements,and the body weight of rats treated with NRG showed an increase compared with DEX group.Conclusion These findings imply that NRG exhibited a protective effect against DEX-induced OP in rats as it promotes the bone formation process by increasing the number of bone turnover markers including OC and CT,and restoring of antioxidant status,lipid metabolism,and inflammatory markers.

促红素联合多糖铁复合物及左卡尼汀治疗维持性血液透析患者肾性贫血的临床价值

目的 分析研究促红素联合多糖铁复合物及左卡尼汀治疗维持性血液透析患者肾性贫血的临床价值。方法 选取2015年3月至2016年7月之间本院收治的94例维持性血液透析肾性贫血患者作为研究对象，以上患者均知晓并自愿参与本次研究，将其按随机数字表法分为观察组与对照组，每组47例，再予以不同的治疗方式。对照组患者使用促红素皮下注射治疗，观察组患者使用促红素皮下注射联合口服多糖铁复合物及左卡尼汀静脉注射治疗。两组患者治疗前后Hb、Hct、ALB、SF等指标的改善情况、治疗后临床症状的改善情况、不良反应发生率、治疗前后的SAS评分、SDS评分比较。结果 治疗前，两组患者的Hb、Hct、ALB、SF指标、SAS评分、SDS评分差异均无统计学意义（均P＞0.05）；治疗后，观察组患者的Hb、Hct、ALB、SF指标、SAS评分、SDS评分均显著优于对照组，差异有统计学意义（P＜0.05）。治疗后，观察组患者的临床症状改善情况较为显著，不良反应发生率仅为4.26%；对照组患者的临床症状改善情况不甚明显，不良反应发生率高达19.15%；组间比较，差异有统计学意义（P＜0.05）。结论 给予维持性血液透析肾性贫血患者使用促红素皮下注射联合口服多糖铁复合物及左卡尼汀静脉注射进行治疗，可显著改善患者的临床症状，降低不良反应的发生率，消除患者的不良情绪，恢复患者的机体平衡，值得临床推广应用。

市售千金子饮片质量分析

目的通过对收集的市售千金子饮片进行含量测定,为制定千金子质量标准提供依据。方法以脂肪油、浸出物、秦皮乙素和七叶树苷为指标,对市售的6个样品进行含量测定。结果各地千金子样品脂肪油组分、醚溶性成分差别不大,水溶性成分、醇溶性成分、秦皮乙素和七叶树苷差异较大。结论各地市售样品在内在质量上存在较大差异。

血管活性因子与慢性乙型肝炎关系的研究

目的 探讨慢性乙型肝炎患者血浆内皮素 (ET)、一氧化氮 (NO)、心钠素 (ANP)和降钙素基因相关肽 (CGRP)水平及相互关系。方法 测定 60例慢性乙肝、60例乙肝后肝硬化患者ET、NO、ANP和CGRP及肝功能水平 ,并选择 30例健康者作为对照。结果 慢性乙肝、肝硬化患者ET、NO明显升高 ,CGRP明显降低 ,ANP无变化 ,肝硬化患者治疗后ET明显下降 ,CGRP明显上升 ,慢性乙肝、肝硬化患者治疗后NO明显下降 ,但仍高于对照。结论 ET、NO、ANP和CGRP相互作用 ,与慢性乙型肝炎的发生、发展过程有关。

Possible involvement of integrin signaling pathway in the process of recovery from restraint stress in rats

Objective To search novel genes or pathways involved in the recovery process after restraint stress in rats. Methods We compared the hypothalamus transcriptional profiles of two different recovery patterns （fast recovery vs slow recovery） from restraint stress in rats using oligonucleotide microarray, the recovery pattern was determined by the decrement of plasma adrenocorticotropic-hormone （ACTH） and corticosterone levels during one hour recovery period after stress. A real-time quantitative RT-PCR was applied to validate the differential expressed genes. Results Analysis of the microarray data showed that most of genes were not differentially expressed between fast recovery group and slow recovery group. Among the differentially expressed genes we found that talin, together with serine/threonine protein phosphatase PPl-beta catalytic subunit （PP-1B） and integrin α-6 precursor （VLA-6） genes, were at least 1.5 fold upregulated in the fast recovery group, while junctional adhesion molecule 1 （F11r） was 1.5 fold down-regulated in the fast recovery group. Conclusion The results implied that integrin signaling pathway may be involved in the recovery from restraint stress in rats. The present study provided a global overview of hypothalamus transcriptional profiles during the process of recovery from the restraint stress in rats. The integrin signaling pathway seems to be involved in the recovery process, which deserves further study to clarify the integrin-mediated recovery mechanism after restraint stress.

Changes of serum adrenocorticotropic hormone and cortisol levels during sleep seizures

Objective Measuring the serum concentrations of adrenocorticotropic hormone （ACTH） and cortisol in epileptic seizures during sleep to investigate their link to the EEG changes. Methods Pre-surgical evaluation was performed by videoEEG monitoring using 24 channel recording. Thirty six epilepsy patients could be attributed to two groups： 28 patients had spontaneous seizures, and the other 8 patients whose seizures were induced by bemegride. Another 11 persons with confirmed psychogenic non-epileptic seizures （PNES） served as control group. Blood samples were obtained at five points： wake （08：00 a.m.）, sleep （00：00 a.m.）, and shortly before, during and after an epileptic seizure. The serum ACTH and cortisol were measured and analyzed by chemiluminescent immunoassay. Results The levels of ACTH and cortisol in serum underwent significant changes： declining below the average sleep-level shortly before seizures, increasing during seizures, and far above the average wake-level after seizures （P 〈 0.001）. Such changes did not occur in the control group （P 〉 0.05）. The ACTH and cortisol levels had no significant difference between spontaneous group and bemegride-induced group （P 〉 0.05）. Conclusion The serum concentrations of ACTH and cortisol during sleep seizures are linked with pre-ictal and ictal EEG changes in epilepsy patients.

Therapeutic effect of microencapsulated porcine retinal pigmented epithelial cells transplantation on rat model of Parkinson's disease

Object To investigate the therapeutic effect of microencapsulated porcine retinal pigmented epithelial cells （RPE-M） transplantation on rat model of Parkinson＇s disease （PD）. Methods Primary porcine RPE cells were harvested by enzyme digestion and expanded in culture medium. Determine the levels ofdopamine （DA） and homovanillic acid （HVA） by high performance liquid chromatography electrochemical （HPLC） assay, and the levels of brain-derived neurotrophic factor （BDNF） and glial-derived neurotrophic factor （GDNF） were detected by ELISA. Alginate-polylysine-alginate （APA） microencapsulated cells were produced by using a high voltage electrostatic system. PD rat model was established by unilateral injection of 6-hydroxydopamine （6-OHDA） into the medial forebrain bundle （MFB）. After that, the RPE-M was transplanted into the corpus striatum of PD rat, and then the rotation test scores were recorded and biochemical changes of the corpus striatum were tested. Results The levels of DA, HVA, BDNF and GDNF secreted by RPE were stable in the RPE culture supernatant and were not changed by the microencapsulation. Eighty-three percent rats developed PD by unilateral lesion of 6-OHDA in the MFB. The RPE-M transplantation had therapeutic effect on 33% PD rats. Conclusion Porcine RPE cells grow actively in vitro and could secrete DA, HVA, BDNF, and GDNF constantly, which does not be affected by the passage culture and the APA miroencapsulation. RPE-M transplantation of may be a curative therapy for PD.

Preparation and Physicochemical Characterization of Quercetin-HP-β-CD Inclusion Complexes

Aim To prepare and characterize the QURC-HP-β-CD inclusion complexes and investigate the thermodynamic parameters of the process. Methods QURC-HP-β-CD inclusion complexes were prepared by the grinding method. The equilibrium inclusion constants and thermodynamic parameters were determinated by phase solubility analysis. Dissolution tests were performed to study the dissolution rate of inclusion complexes. The formation of inclusion complexes was confirmed by differential scanning calorimetry （ DSC）, infrared spectroscopy （IR） , powder X-ray diffractometry （PXRD） and scanning electron microscopy （SEM）. Results The aqueous solubility of quercetin was greatly increased （ about 37 folds） by inclusion technique, and the initial dissolution rate was markedly improved （10 folds） in the first 5 min. The results of DSC and SEM photographs showed that quercetin crystal disappeared in inclusion complexes, which indicated the formation of new phase. FT-IR spectra showed that the carbonyl quercetin crystal grinding method. absorption band of quercetin was shifted. PXRD showed that the diffraction peak of disappeared. Conclusion QURC-HP-β-CD inclusion complexes are produced by the The solubility of quercetin is improved by the inclusion technique.

Effect of Quercetin on the Proliferation and Mitochondrial Transmembrane Potential of CBRH-7919 Cells

[Objective] To investigate the effect of quercetin on the proliferation and mitochondrial transmembrane potential of CBRH-7919 cells. [Method] The CBRH-7919 cells of hepatocarcinoma were cultured in vitro. After treated with different concentrations of quercetin, the OD405 nm of CBRH-7919 cells was detected by using the acid phosphatase assy （APA）; morphologic changes of the cells were observed under inverted microscope; the mitochondrial transmembrane potential （△ψm） intensity changes of CBRH-7919 cells were analyzed by flow cytometry after stained with Rhodamine 123. [Result] Quercetin inhibited the proliferation of CBRH-7919 cells significantly, and the growth inhibitory effect presented time- and dose-dependent relationship. Typical decrease of cell density was observed by optical microscopy on the quercetin-treated cells. With the effect of 10 μg/ml quercetin on CBRH-7919 cells for 12, 24 and 48 h, the percentage of Rhodamine 123 stained hypofluorescence cells increased, while the mitochondrial transmembrane potential（△ψm） intensity of CBRH-7919 cells decreased. [Conclusion] Quercetin could inhibit the proliferation of CBRH-7919 cells in vitro, causing the decrease in mitochondrial transmembrane potential.

Effect of Quercetin on CYP1A2, CYP2E1, CYP3A2 Activities and its Inhibitory Mechanism Studies in Rat Liver Microsomes

Aim To assess the potential effect of quercetin （QU）, an natural plant estrogen, on CYP1A2, CYP2E1, and CYP3A2 activities in rat liver microsomes; and to identify the magnitude of inhibitory effect and the probable inhibitory mechanism of QU. Methods QU and specific substrate were concurrently incubated, with HPLC detection of the substrate metabolites for data analysis. The magnitude of inhibitory effect of QU on CYP3A2 was compared with those of ketoconazole （Ket） and erythromycin （Ery）. The mechanism of its inhibitory effect on CYP3A2 and CYP2E1 was derived from Lineweaver-Burk plots. Results HPLC methods were in good linear relationship with r〉0.999 1. Relative standard deviations for intra-day and inter-day were〈8.4%. Recovery of each analyte in the concentrations studied was between 91.1% and 107.6 %. QU （up to 8 μmol·L^-1） showed potent induction to CYP1A2 （338.1% of the negative control）while inhibited CYP2E1 （49.2% of the negative control） and CYP3A2 （60.3% of the negative control） activity. The magnitude of inhibitory effect for QU on CYP3A2 was between those for Ket and Ery （Ket〉QU〉Ery）. QU exhibited competitive inhibition of CYP3A2 dextromethorphan N-demethylation reaction and expressed noncompetitive inhibition of CYP2E1 chlorzoxazone-6-hydroxylation reaction. Conclusion HPLC assay has been validated with precision and accuracy. QU is an effective inhibitor of several CYP isoforms. It may cause relevant drug-drug interactions with CYP3A substrates. As a plant flavonoid, QU has potential not only in molecular advantage but also in CYP450 module capability for further application in cancer chemotherapy.

The Chemical Constituents from the Bulbs of Ornithogalum caudatum

A new natural product (1) together with 26 know compounds were isolated from the Bulbs of Ornithogalum caudatum. Their structures were established on the basis of spectral analyses as n-butyl pyroglutamate (1), nonadecyl alcohol(2), eicosanol(3), behenic acid(4), b-sitosterol(5), stigmasterol(6), glycerol 1-monocerotate(7), pyrocatechol(8), p-ethoxybenzoic acid(9), p-coumarinic acid(10), protocatechuric acid(11), ursolic acid(12), betulinic acid(13), fumaric acid(14), succinic acid(15), uracil(16), xanthine(17), quercetin(18), kaempferol (19), isorham-netin(20), adenosine(21), daucosterol(22), stigmasterol 3-O-b-D-glucopyranoside(23), quercetin 3-O-b-D-glucopyra-noside(24), kaempferol 3-O-b-D-glucopyranoside(25), rutin(26), and kaempferol 3-O-b-rutinoside(27). All of them, except compound 5, were isolated from this plant for the first time.

A New Antifungal Flavonol Glycoside from Hypericum perforatum

In addition to six known flavonoids quercitrin, hyperoside, avicularin, rutin, quercetin and kaemferol, a new flavonol glycoside named 6″_O_acetyl quercetin 3_O_β_ D _alloside (1) was isolated from the aerial parts of Hypericum perforatum L. The structures were determined on the basis of spectroscopic methods (UV, IR, FAB_MS, 1H_NMR and 13 C NMR). Antifungal assay of all compounds showed that metabolite 1, quercitrin and quercetin were inhibitory to the growth of phytopathogenic fungus Helminthosporium sativum Pamel King et Bakke with minimum inhibitory concentrations (MICs) of 25, 50 and 50 μg/mL, respectively. Moreover, glycoside 1 and quercitrin were also shown to be able to inhibit the growth of Fusarium graminearum Schw. with MIC of 100 μg/mL. The MICs of ketoconazole used as control against the test fungi were 0.5 μg/mL in our assay.

Influence of Replacing Bacterial Pheophytin with Pheophytin on the Picosecond Fluorescence from Reaction Centers of Purple Bacteria

Fluorescence spectra of native purple bacterial reaction center (RC) and bacterial pheophytin (Bphe) replaced RCs were obtained from 600 nm to 900 nm with a selective excitation at 597 nm. With the help of measuring the fluorescence from bacterial chlorophyll, bacterial pheophytin and plant pheophytin, the corresponding components in the RCs are classified for fluorescence emission. Results showed that pheophytin substitution influences the composition of fluorescence spectra. Therefore, four, three and two components were obtained from fluorescence spectra of native RC, Bphe B_replaced RC and Bphe A,B _replaced RC, respectively. Fluorescence components are well correlated to the binding of plant pheophytin. The decay of excited state of primary electron donor P in different RCs was also studied by measuring the fluorescence decay at 686.4, 674.1 and 681.1 nm, respectively. The decaying kinetics changed in different RCs, indicating that pheophytin replacement influenced the energy transduction and primary photochemical reaction in purple bacterial reaction centers.

Effects of Ca^(2+)concentrations on accumulations of mineral elements in the components of Pteroceltis tatarinowii

The bark of Pteroce/tis tatarinowii is a raw material for manufacturing XuanPaper. The effects of Ca^(2+) concentrations on the accumulation of mineral elements in the bark,leaf and root of Pteroceltis tatarinowii were studied under controlled conditions. The types ofHoagland nutrient solution with three Ca^(2+) concentrations levels (200, 400 and 600 μg·g^(-1))and a control (without Ca^(2+)) were designed to culture Pteroceltis tatarinowii. After 6 months,contents of seven mineral elements including Ca, K, Mg, Mn, Zn, Cu and Na in the root, leaf and barkwere analyzed. The results indicated that Ca accumulations content in the root, leaf and bark hadpositively relation with Ca^(2+) concentrations (200, 400, 600 μg · g^(-1)), and the order of theCa content in the three components was root>leaf>bark. Ca content in the root treated with 600 μg·g^(-1) Ca^(2+) concentrations was 5.5 times as high as that of the control, and about 1.4 times ashigh as that of the root treated in 200 and 400 μg/g Ca^(2+) concentrations respectively. On thecontrary, K and Mg contents in the root, leaf and bark were negatively related to Ca^(2+)concentrations, especially in the bark, and their accumulation trend followed the order ofleaf>root>bark. K content in the bark treated with 600 μg ·g^(-1) Ca^(2+) concentrations was 39.3%of that of the control, and was 79.0% and 91.8% of that of the bark treated with 200 μg ·g^(-1)and 400 μg ·g^(-1) Ca^(2+) concentrations respectively; Mg content in the bark treated with 600μg ·g^(-1) Ca^(2+) concentrations was 23.4% of that of the control, and was 27.1% and 35.4% ofthat of the bark treated with 200 and 400 μg·g^(-1) Ca^(2+) concentrations respectively. Comparedwith the control, the general tendency of Mn, Zn and Cu content decreased with increasing of Ca^(2+)concentrations and their contents were in the order: root>leaf>bark. Based on the results of thisstudy, the experiment has been useful for providing academic bases in improving the bark quality ofPteroceltis tatarinowii on non-limestone soil.

不同部位注射重组人促红素的减痛效果评价

目的探讨不同部位注射重组人促红素的减痛效果。方法按数字随机法随机抽取2016年5月份41名患者，选择上臂三角肌下缘注射为对照组（三角肌组），肘关节伸外侧无痛感区域皮下注射为观察组（肘关节组），同一工作人员执行注射重组人促红素，在同一患者每日注射不同的部位，即肘关节伸外侧区域与上臂三角肌下缘皮下注射交替进行，每次注射完毕都让患者在脸谱表情＋评分的疼痛评估图上指出其感觉程度，并分别记录同一患者不同方法注射时患者的疼痛评分、患者满意度及护士操作方便程度并进行分析比较。结果肘关节组的患者疼痛评分、患者满意度、护士操作方便程度均优于三角肌组，差异均有统计学意义（均P〈0．05）。结论经肘关节伸侧无痛感区域注射刺激性较强的重组人促红素可降低患者的疼痛程度、提高患者的满意度、方便护士操作。

Morphological and behavioral consequences of recurrent seizures in neo-natal rats are associated with glucocorticoid levels

Objective It is well documented that epilepsy can increase neurogenesis in certain brain regions and cause behavioral alternations in patients and different epileptic animal models. A series of experimental studies have demonstrated that neurogenesis is regulated by various factors including glucocorticoid （CORT）, which can reduce neurogenesis. Most of studies in animal have been focused on adulthood stage, while the effect of recurrent seizures to immature brain in neonatal period has not been well established. This study was designed to investigate how the recurrent seizures occurred in the neonatal period affected the immature brain and how CORT regulated neurogenesis in immature animals. Methods Neonatal rats were subjected to 3 pilocarpine-induced seizures from postnatal day 1 to day 7. Then neurogenesis at different postnatal ages （i.e. P8, P12, P22, P50） was observed. Behavioral performance was tested when the rats were mature （P40）, and plasma CORT levels following recurrent seizures were simultaneously monitored. Results Rats with neonatal seizures had a significant reduction in the number of Bromodeoxyuridine （BrdU） labeled cells in the dentate gyrus compared with the control groups when the animals were euthanized on P8 or P12 （P 〈 0.05）; whereas there was no difference between the two groups on P22. Until P50, rats with neonatal seizures had increased number of BrdU-labeled cells compared with the control group （P 〈 0.05）. In Morris water maze task, pilocarpine-treated rats were significantly slower than the control rats at the first and second day, and there were no differences at other days. In probe trial, there was no significant difference in time spent in the goal quadrant between the two groups. Endocrine studies showed a correla- tion between the number of BrdU positive cells and the CORT level. Sustained increase in circulating CORT levels was observed following neonatal seizures on P8 and P12. Conclusion Neonatal recurrent seizures can biphasely modulate neurogenesis over different time windows with a down-regulation at early time and up-regulation afterwards, cause persistent deficits in cognitive functions of adults, and increase the circulating CORT levels. CORT levels are related with the morphological and behavioral consequences of recurrent seizures.