Changes in morphology and biochemical indices in browning callus derived from <Emphasis Type="Italic">Jatropha curcas</Emphasis> hypocotyls

Callus browning is a typical feature of callus cultures derived from the hypocotyl of Jatropha curcas. Brown callus results in decreased regenerative ability, poor growth and even death. In this study, we investigated the effect of browning on callus morphology and biochemical indices. Light microscopy and scanning electron microscopy showed striking differences in callus morphology. During browning, chlorophylls and carotenoids concentrations decreased steadily. Polyphenol oxidase (PPO) and peroxidase (POD) enzymatic activities patterns were similar during callus culture with a higher activity level at week 3 compared to week 2 or later weeks. Grey relation degree analysis indicated that PPO played a more important role than POD in enzymatic callus browning. Polyacrylamide gel electrophoresis results showed differences between browning and non-browning callus. Gas chromatography–mass spectrometry results showed that saturated and unsaturated fatty acid quantities differed significantly but there was little difference in fatty acid composition between non-browning and browning callus. Differences in 17, 18.4 and 25 kDa protein concentrations were also observed in browning and non-browning callus using sodium dodecyl sulfate–polyacrylamide gel electrophoresis.     

昆虫细胞免疫反应中的吞噬、集结和包囊作用

细胞免疫是昆虫天生免疫系统中很重要的部分, 包括了由血细胞介导的一系列吞噬、 集结和包囊等作用。本文讨论了近年来在昆虫细胞免疫方面的研究进展, 包括参与昆虫细胞免疫的血细胞类型, 识别外来异物的受体因子, 影响免疫活性的一些酶和化学物质等。另外还就吞噬模式, 以及集结和包囊过程中粘附态细胞的形成等加以讨论。     

Targeted inhibition of the EGFR pathways enhances Zn‐BC‐AM PDT‐induced apoptosis in well‐differentiated nasopharyngeal carcinoma cells

Epidermal growth factor receptor (EGFR), a receptor often expressed in nasopharyngeal carcinoma (NPC) cells, is one of the recently identified molecular targets in cancer treatment. In the present study, the effects of combined treatment of Zn‐BC‐AM PDT with an EGFR inhibitor AG1478 were investigated. Well‐differentiated NPC HK‐1 cells were subjected to PDT with 1 µM of Zn‐BC‐AM and were irradiated at a light dose of 1 J/cm² in the presence or absence of EGFR inhibitor AG1478. Specific protein kinase inhibitors of downstream EGFR targets were also used in the investigation. EGFR, Akt, and ERK were found constitutively activated in HK‐1 cells and the activities could be inhibited by the EGFR inhibitor AG1478. A sub‐lethal concentration of AG1478 was found to further enhance the irreversible cell damage induced by Zn‐BC‐AM PDT in HK‐1 cells. Pre‐incubation of the cells with specific inhibitors of EGFR (AG1478), PI3k/Akt (LY294002), or MEK/ERK (PD98059) before light irradiation were found to enhance Zn‐BC‐AM PDT‐induced formation of apoptotic cells. The efficacy of Zn‐BC‐AM PDT can be increased through the inhibition of EGFR/PI3K/Akt and EGFR/MEK/ERK signaling pathways in NPC cells. Combination therapy with Zn‐BC‐AM PDT and EGFR inhibitors may further be developed for the treatment of advanced NPC. J. Cell. Biochem. 108: 1356–1363, 2009. © 2009 Wiley‐Liss, Inc.     

XC1028 from Xanthomonas campestris adopts a PilZ domain‐like structure without a c‐di‐GMP switch

The crystal structure of XC1028 from Xanthomonas campestris has been determined to a resolution of 2.15 Å using the multiple anomalous dispersion approach. It bears significant sequence identity and similarity values of 64.10% and 70.09%, respectively, with PA2960, a protein indispensable for type IV pilus‐mediated twitching motility, after which the PilZ motif was first named. However, both XC1028 and PA2960 lack detectable c‐di‐GMP binding capability. Although XC1028 adopts a structure comprising a five‐stranded β‐barrel core similar to other canonical PilZ domains with robust c‐di‐GMP binding ability, considerable differences are observed in the N‐terminal motif; XC1028 assumes a compact five‐stranded β‐barrel without an extra long N‐terminal motif, whereas other canonical PilZ domains contain a long N‐terminal sequence embedded with an essential “c‐di‐GMP switch” motif. In addition, a β‐strand (β1) in the N‐terminal motif, running in exactly opposite polarity to that of XC1028, is found inserted into the parallel β3/β1′ strands, forming a completely antiparallel β4↓β3↑β1↓β1′↑ sheet in the canonical PilZ domains. Such dramatic structural differences at the N‐terminus may account for the diminished c‐di‐GMP binding capability of XC1028, and suggest that interactions with additional proteins are necessary to bind c‐di‐GMP for type IV fimbriae assembly. Proteins 2009. © 2008 Wiley‐Liss, Inc.     

Obesity and genetics regulate microRNAs in islets, liver, and adipose of diabetic mice

Type 2 diabetes results from severe insulin resistance coupled with a failure of β cells to compensate by secreting sufficient insulin. Multiple genetic loci are involved in the development of diabetes, although the effect of each gene on diabetes susceptibility is thought to be small. MicroRNAs (miRNAs) are noncoding 19–22-nucleotide RNA molecules that potentially regulate the expression of thousands of genes. To understand the relationship between miRNA regulation and obesity-induced diabetes, we quantitatively profiled approximately 220 miRNAs in pancreatic islets, adipose tissue, and liver from diabetes-resistant (B6) and diabetes-susceptible (BTBR) mice. More than half of the miRNAs profiled were expressed in all three tissues, with many miRNAs in each tissue showing significant changes in response to genetic obesity. Furthermore, several miRNAs in each tissue were differentially responsive to obesity in B6 versus BTBR mice, suggesting that they may be involved in the pathogenesis of diabetes. In liver there were approximately 40 miRNAs that were downregulated in response to obesity in B6 but not BTBR mice, indicating that genetic differences between the mouse strains play a critical role in miRNA regulation. In order to elucidate the genetic architecture of hepatic miRNA expression, we measured the expression of miRNAs in genetically obese F2 mice. Approximately 10% of the miRNAs measured showed significant linkage (miR-eQTLs), identifying loci that control miRNA abundance. Understanding the influence that obesity and genetics exert on the regulation of miRNA expression will reveal the role miRNAs play in the context of obesity-induced type 2 diabetes.     

Suppressive effects of swainsonine on C6 glioma cell in vitro and in vivo

Swainsonine, an extract from Astragalus membranaceus, is known for its anti-cancer effects and could prevent metastases. In order to investigate the effects and mechanisms of swainsonine in C6 glioma cells, we carry out correlated experiments in vitro and in vivo. After treatment with swainsonine, the effective dose and IC₅₀ value of swainsonine in the C6 glioma cell were examined using the MTT assay. Cell cycle distribution and apoptotic rates were analyzed using FCM and [Ca²⁺]_i was measured by LSCM. Expressions of p16 and p53 protein were evaluated by immunocytochemical methods. Simultaneously, glioma-bearing rats were administered swainsonine at doses of 2, 4 and 8 mg/kg body wt. The inhibition rate was calculated and pathological sections were observed. The results indicated that the growth of C6 glioma cells is inhibited by swainsonine in vitro, with an IC₅₀ value within 24 h of 0.05 μg/ml. Increases in swainsonine correlate with S phase percentages of 11.3%, 11.6% and 12.4%, respectively. Moreover, the expression of apoptosis inhibiting p53 and p16 protein decreases gradually. Tumor weight in vivo decreased clearly and HE dyeing of tumor tissue showed gray, its texture was soft, with necrosis and hemorrhagic concentrated inward. Swainsonine could inhibit the proliferation of C6 glioma cells in vitro and the growth of C6 glioma in vivo. The mechanisms of swainsonine-induced apoptosis may relate with the expression of apoptosis-related genes and overloading-[Ca²⁺]_i-induced endoplasmic reticulum stress.     

Clinical observation of Danhong Injection (herbal TCM product from Radix Salviae miltiorrhizae and Flos Carthami tinctorii) in the treatment of traumatic intracranial hematoma

Danhong Injection (DHI), a Chinese Materia Medica standardized product extracted from Radix Salviae Miltiorrhizae and Flos Carthami tinctorii, has the actions of promoting blood circulation and resolving stasis to promote regeneration. The clinical therapeutic effects of DHI on traumatic intracranial hematoma (TICH) were observed. Eighty patients with TICH were randomly assigned to trial group and a control group (40 patients per group), and all were administered with routine medication. Additionally, DIH was administered intravenously to patients in the trial group. Pre and post-treatment GCS was observed in the two groups, along with GOS after therapy. The intracranial hematoma absorption, hemorheological changes, and changes in coagulation indexes pre- and post-treatment were evaluated. The results indicated that GCS and GOS after therapy for the trial group were superior to those for the control group (p<0.05). There was a significant post-treatment difference in the intracranial hematoma absorption between the two groups (p<0.01). Each hemorheological index in the trial group improved significantly as compared with that of the control group (p<0.05 or p<0.01). The plasma levels of fibrinogen and D-dimer in the trial group were significantly decreased after therapy (p<0.01). These results suggest that DHI is conducive to the recovery of patients with TICH.     

Development of a high analytical performance-xanthine biosensor based on layered double hydroxides modified-electrode and investigation of the inhibitory effect by allopurinol

The determination of xanthine has considerable importance in clinical and food quality control. Therefore, in this present work, we developed a novel xanthine biosensor based on immobilization of xanthine oxidase (XnOx) by attractive materials layered double hydroxides (LDHs). Amperometric detection of xanthine was evaluated by holding the modified electrode at 0.55V (versus saturated calomel electrode (SCE)). Due to the special properties of LDHs, such as chemical inertia, mechanical and thermal stability, anionic exchange ability, high porosity and swelling properties, XnOx/LDHs-modified electrode exhibited a developed analytical performance. The biosensor provided a linear response to xanthine over a concentration range of 1 x 10(-6)M to 2 x 10(-4)M with a sensitivity of 220 mAM(-1)cm(-2) and a detection limit of 1x10(-7)M based on S/N=3. In addition, the immobilized XnOx layers have been characterized using atomic force microscopy under both air atmosphere and liquid environment, which exhibited the interesting swelling phenomenon of LDHs. The investigation of inhibition of XnOx by allopurinol was carried out using this XnOx/LDHs-modified electrode. The experimental results indicated that inhibitory effect could be achieved by allopurinol with a quasi-reversible competitive type.     

Xanthine oxidase/laponite nanoparticles immobilized on glassy carbon electrode: Direct electron transfer and multielectrocatalysis

In this work, colloidal laponite nanoparticles were further expanded into the design of the third-generation biosensor. Direct electrochemistry of the complex molybdoenzyme xanthine oxidase (XnOx) immobilized on glassy carbon electrode (GCE) by laponite nanoparticles was investigated for the first time. XnOx/laponite thin film modified electrode showed only one pair of well defined and reversible cyclic voltammetric peaks attributed to XnOx–FAD cofactor at about −0.370 V vs. SCE (pH 5). The formal potential of XnOx–FAD/FADH₂ couple varied linearly with the increase of pH in the range of 4.0–8.0 with a slope of −54.3 mV pH⁻¹, which indicated that two-proton transfer was accompanied with two-electron transfer in the electrochemical reaction. More interestingly, the immobilized XnOx retained its biological activity well and displayed an excellent electrocatalytic performance to both the oxidation of xanthine and the reduction of nitrate. The electrocatalytic response showed a linear dependence on the xanthine concentration ranging from 3.9 × 10⁻⁸ to 2.1 × 10⁻⁵ M with a detection limit of 1.0 × 10⁻⁸ M based on S/N = 3.     

Characterization and flocculating properties of a novel bioflocculant produced by <Emphasis Type="Italic">Bacillus circulans</Emphasis>

We studied a novel bioflocculant, PX, that is produced from Bacillus Bacillus circulans X3, and has excellent flocculating activity with regard to its characterization and flocculating properties. The bioflocculant was purified from supernatant by ethanol precipitation, dialysis and gel permeation chromatography (GPC). The major component of PX was an acid polysaccharide including uronic (19.8%), pyruvic (6.5%) and acetic acids (0.7%). It consisted of galactose, mannose, xylitol, rhamnose and galacturonic acid in an approximate molar ration of 5:4.1:3:2:1.2. The molecular weight of PX was about 4.85 × 10⁴ Da as determined by GPC. The infrared spectrum of the bioflocculant indicated the presence of carboxyl, hydroxyl, amino and methoxyl groups. Studies of the flocculating properties revealed that it was stable at 60–100°C and pH 4–10. Moreover, it could flocculate a kaolin suspension over a wide range of pH and temperature in the presence of CaCl₂.