The surface morphology and fine structure of CHO (Chinese hamster ovary) cells following enucleation.

Chinese hamster ovary cells grown in monolayer culture and exposed to cytochalasin B were enucleated by centrifugation. Thereafter, the karyoplasts (the nucleated parts obtained from the bottoms of the centrifuge tubes) and the cytoplasts (the enucleated cytoplasmic parts attached to the coverslips) were allowed to recover and subsequently were examined by scanning and transmission electron microscopy. Microscopy of thin sections revealed that the karyoplasts, limited by an intact plasma membrane, contain an intact nucleus surrounded by a layer of cytoplasm that includes ribosomes, mitochondria, and fragments of the endoplasmic reticulum, but no centrioles or microtubules. The cytoplasts, similarly examined, appear to contain all cytoplasmic organelles and systems, including centrioles and microtubules. The karyoplasts, when replated in fresh medium adhere to the substrate but remain essentially spherical and are incapable of motility.

They disintegrate in about 72 hr. The cytoplasts, under identical conditions, recover a shape similar to that of the whole Chinese hamster ovary cell and display some motility. They generally survive not more than 48 hr. It appears that this enucleation procedure consistently separates the nucleus and limited cytoplasm from the centrosphere and microtubule-containing cytoplasts and Centrifuge tube , furthermore, that the formdetermining and motility mechanisms reside in the cytoplast and function without nuclear participation for the short period of viability.

Essential role of IL-10/STAT3 in chronic stress-induced immune suppression.

• Stress can either enhance or suppress immune functions depending on a variety of factors such as duration of stressful condition. Chronic stress has been demonstrated to exert a significant suppressive effect on immune function.
• However, the mechanisms responsible for this phenomenon remain to be elucidated. Here, male C57BL/6 mice were placed in a 50-ml conical centrifuge tube with multiple punctures to establish a chronic restraint stress model.
• Serum IL-10 levels, IL-10 production by the splenocytes, and activation of STAT3 in the mouse spleen were assessed. We demonstrate that IL-10/STAT3 axis was remarkably activated following chronic stress. Moreover, TLR4 and p38 MAPK play a pivotal role in the activation of IL-10/STAT3 signaling cascade.
• Interestingly, blocking antibody against IL-10 receptor and inhibition of STAT3 by STAT3 inhibitor S3I-201 attenuates stress-induced lymphocyte apoptosis. Inhibition of IL-10/STAT3 dramatically inhibits stress-induced reduction in IL-12 production. Furthermore, disequilibrium of Th1/Th2 cytokine balance caused by chronic stress was also rescued by blocking IL-10/STAT3 axis.
• These results yield insight into a new mechanism by which chronic stress regulates immune functions. IL-10/STAT3 pathway provides a novel relevant target for the manipulation of chronic stress-induced immune suppression.
• Free radical scavenging activities of water-soluble extracts from some natural sources, health foods, and antioxidant substances were measured using the JES-FR30 JEOL spectrometer.
• The objective was to develop a standardized method whereby comparison could be made between the radical scavenging activities of complex mixtures. Scavenging of hydroxyl radical was determined using DMPO.
• Activity was calibrated using a standard material, L-ascorbic acid 2-[3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-2H -1- benzopyran-6yl-hydrogen phosphate] potassium salt (EPC-K1), an analog of vitamin C and vitamin E which is water soluble and stable at room temperature. The order of greatest hydroxyl radical scavenging activity was green tea extract, pine bark extract (Pycnogenol), Ginkgo Biloba extract (EGb 761), a flavonoid blend of several fruit and vegetable extracts (GNLD), and Bio-Normalizer (Sun-O Corp).
• Activity was determined after treatment of samples with ascorbic acid oxidase. This treatment revealed the presence of ascorbate in some natural extracts and commercial preparations. The pine bark extract was the most heat resistant and had ascorbate-like activity in the preparations.
• Scavenging of superoxide anion was determined using the spin trap, 5,5-dimethyl-1-pyrroline-N-oxide (DMPO), and analyzed by comparison with a standard curve made with superoxide dismutase. Comparison of the water solubilized components of natural source antioxidants showed that filtrates fractionated using centrifuge type Millipore filter tubes (M.W. < 100,000; M.W. < 10,000) also had almost the same SOD-like activity. Samples were also treated with ascorbate oxidase or by heating (100 degrees C for 10 min). The order of activity, from greatest to least, was Ginkgo biloba extract EGb 761, pycnogenol, beta-catechin, tea and BioNormalizer.

Field observations of the Asian longhorned beetle (Anoplophora glabripennis) mating behavior in China suggested that a female-produced contact pheromone was almost certainly involved in sex recognition.

Gas chromatography-mass spectrometry (GC-MS) analysis of A. glabripennis adults’ whole body cuticular extracts indicates that a series of long-chain hydrocarbons comprise the cuticular waxes of both sexes. Although for the most part the GC profiles are similar for the two sexes, five monounsaturated compounds were consistently more abundant in samples from females than in those from males. These compounds were identified as (Z)-9-tricosene, (Z)-9-pentacosene, (Z)-7-pentacosene, (Z)-9-heptacosene, and (Z)-7-heptacosene in the approximate ratio of 1:2:2:8:1, respectively. Antennal and palpi contact to a polypropylene micro-centrifuge tube coated with a synthetic mixture of the five compounds stimulated copulatory behavior in males.

A modification that entails the use of buffering during extraction was made to further improve results for certain problematic pesticides (e.g., folpet, dichlofluanid, chlorothalonil, and pymetrozine) in a simple, fast, and inexpensive method for the determination of pesticides in produce. The method, known as the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method for pesticide residues in foods, now involves the extraction of the sample with acetonitrile (MeCN) containing 1% acetic acid (HAc) and simultaneous liquid-liquid partitioning formed by adding anhydrous MgSO4 plus sodium acetate (NaAc).

The extraction method is carried out by shaking a centrifuge tube which contains 1 mL of 1% HAc in MeCN plus 0.4 g anhydrous MgSO4 and 0.1 g anhydrous NaAc per g sample. The tube is then centrifuged, and a portion of the extract is transferred to a tube containing 50 mg primary secondary amine sorbent plus 150 mg anhydrous MgSO4/mL of extract. After a mixing and centrifugation step, the extract is transferred to autosampler vials for concurrent analysis by gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/tandem mass spectrometry. Independent of the original sample pH, the use of buffering during the extraction yields pH <4 in the MeCN extract and >5 in the water phase, which increases recoveries of both acid- and base-sensitive pesticides.

The method was evaluated for 32 diverse pesticides in different matrixes, and typical percent recoveries were 95 +/- 10, even for some problematic pesticides. Optional solvent exchange to toluene prior to GC/MS analysis was also evaluated, showing equally good results with the benefit of lower detection limits, but at the cost of more time, material, labor, and expense.