[Advances in the study of the role of adenosine 2A receptor in liver injury].

Adenosine, as an endogenous purine nucleoside, is widely distributed in various tissues and organs of the body. It binds to adenosine receptors to regulate a variety of important biological processes. Adenosine 2A receptors have a close relationship with the occurrence and development of various clinical diseases. This article reviews the research progress of adenosine 2A receptors in non-alcoholic fatty liver disease, acute immune hepatitis, liver ischemia-reperfusion injury, liver fibrosis, etc., in order to provide new research strategies for the prevention and treatment of these diseases.

Sulfated polysaccharides and immune response: promoter or inhibitor?

Sulfated polysaccharides, which frequently connect to core protein, are expressed not only on cell surface but also throughout the extracellular matrix. Besides providing structural integrity of cells, sulfated polysaccharides interact with a variety of sulfated polysaccharides-binding proteins, such as growth factors, cytokines, chemokines and proteases. Sulfated polysaccharides play two-edged roles, inhibitor and promoter, in immune response. Some sulfated polysaccharides act as the immunosuppressor by blocking inflammatory signal transduction induced by proinflammatory cytokines, suppressing the activation of complement and inhibiting the process that leukocytes adhere to and pass through endothelium. On the contrary, the interaction between immune cells and sulfated polysaccharides produced by bacteria, endothelial cells and immune cells initiate the occurrence of immune response. It promotes the processes of recognizing and arresting antigen, migrating transendothelium, moving into and out of immune organ and enhancing the proliferation of lymphocyte. The structure of sulfated polysaccharides, such as molecular weight and sulfated sites heterogeneity, especially the degree of disaccharide sulfation, position of the sulfate moiety and organization of sulfated domains, may play critical role in their controversial effects. As a consequence, the interaction between sulfated polysaccharides and sulfated polysaccharide-binding proteins may be changed by modifying the structure of sulfated polysaccharides chains. The administration of drug targeting sulfated polysaccharide-protein interaction may be useful in treating inflammatory related diseases.

The POU homeodomain protein Oct-1 binds Cis-regulatory element essential for the human GnRH upstream promoter activity in JEG-3 cells.

In previous studies, we have localized four specific nuclear protein-binding elements in the human GnRH upstream promoter. To test whether these four elements are reproductive tissue specific, we placed the four elements upstream to a thymidine kinase (TK) promoter/luciferase reporter gene, and transfected the constructs into human placental choriocarcinoma (JEG-3) cells. The 272-bp fragment (-994 to -723) containing the four elements can drive heterologous TK promoter expression in JEG-3 cells about 15 times more than that of basal TK promoter activity. Deletion of element 4 (E4, -987/-968) significantly decreased (4-fold) the luciferase activity. Further deletion of the other elements (E3 individual, -960/-940 or E3 and E2 in combination, -919/-896) only slightly decreased the luciferase activity. In contrast, deletion of element 1 (E1, -876/-851) caused a 2-fold loss of luciferase activity and elimination of E2 and E3 only lost less than 2-fold of the luciferase activity. Study performed with 5' end deletion of this region confirmed these observations. Furthermore, E4 DNA-protein complex can be supershifted by Oct-1 antibody, indicating that Oct-1 binds to E4. These results clearly demonstrated that all four elements are required to confer tissue-specific expression of the hGnRH gene in JEG-3 cells. However, the E4 is the most important for the tissue-specific expression of the hGnRH gene in JEG-3 cells. Oct-1 factor binds with E4 element and may be involved in the mediation of the human GnRH upstream promoter activity.

A novel perfusion system for animal cell cultures by two step sequential sedimentation.

A novel perfusion system was developed for high density culture of animal cells. The system consists of an airlift bioreactor, a setting tank and a flat settler. Both the settling tank and flat settler have two connecting pipes for transporting the cells from and back to the reactor, respectively. Thus, the cell flow in the settlers can be controlled in uni-direction, avoiding the countercurrent flow of the cells. During perfusion cultures, the cells firstly settled in the settling tank, then, unsettled cells in the tank were transferred to the flat settler for re-settling. With the application of the system to hybridoma cell cultures, it was found that the maximum viable cell density, monoclonal antibody concentration and average productivity were 1.31 x 107 cells ml-1, 400 mg l-1 and 461 mg l-1 d-1, respectively, which were much higher than those of a batch culture. Both theoretical analysis and experimental results showed a much higher separation efficiency in such a two-step sedimentation system than that in a conventional one-step sedimentation system. In addition, the volumetric ratio of the sedimentation devices to the culture volume in our developed system is much lower, which may be potentially useful on an industrial-scale.

Effects of morphine on rheological properties of rat red blood cells.

To evaluate the effect of morphine on red blood cells, in vivo and in vitro rat models of morphine dependence were established. Rheological properties of rat red cells were measured by ektacytometry; the biophysical changes in the membrane of rat red cells were measured by the Fourier-Transformed Infrared technique (FT-IR) and the fluorescence depolarization method. The results show that the membrane fluidity of red cells was greatly reduced by morphine and the secondary structure of membrane proteins was changed. This suggests that morphine affects the rat red cell membrane directly, rather than through opioids-receptors.

A study of effects of WGA and ConA on RBC membrane receptors using a new ektacytometric method.

With a new ektacytometry, we studied the relation between the microstructure of red blood cell (RBC) membrane and the rheological properties of RBCs in a shear flow field of low viscosity. The main contributions of this paper are as follows: 1. The hemorheological meanings of the orientation index (DI)or and the small deformation index (DI)d were explored. (DI)or is an overall rheological index depending on the deformability and morphology of RBCs. The better the physiological shape of RBCs is maintained, the greater the (DI)or is. (DI)d can be used to describe the lipid fluidity of RBC membrane. Such an explanation for the meaning of (DI)d has been forcefully supported by our experiments using electron spin resonance (ESR) and fluorescence polarization. 2. The influence of wheat germ agglutinin (WGA) of different concentrations on the lipid fluidity of membrane is different from that of concanavalin A (ConA). The lipid fluidity of membrane changes with WGA concentration treating RBCs and there is a maximum value for the membrane fluidity at a specific concentration of WGA. However, the deformability of membrane described by the integrate deformation index (IDI) monotonically decreased with the increase in WGA concentration treating RBCs. 3. It is concluded that the increase in the lipid fluidity of red cell membrane is not necessarily associated with the improvement of RBC deformability.

An animal model to study erythrocyte senescence with a narrow time window of erythrocyte production.

Using the method of inducing spherocytic anemia in the rabbit with antibody serum, we have developed an animal model in which red blood cells (RBCs) can nearly grow synchronously. With this model, we determined that the surface charge density on the RBC membrane decreased with cell aging. The change was not linear, being much more profound in the latter half of RBC life span. There was a positive correlation between the mean RBC density and its "age" (r = 0.847, p < 0.01). However, the density distribution of the RBCs at the same "age" showed a broad range, and the density values for RBC groups with different ages showed considerable overlap. This indicates that the density gradient technique can be used to separate RBC population into fractions with different mean ages, but has a low resolving power for obtaining individual RBCs of a given "age".

An animal model to study erythrocyte senescence with a narrow time window of erythrocyte production: alterations in osmotic fragility and deformability of erythrocytes during their life span.

Using the model in which the entire RBC population was nearly synchronously produced following the induction of spherocytic anemia in the rabbit with antibody serum, we determined the changes of RBC osmotic fragility and deformability with aging. The results showed that the osmotic fragility increased with the RBC aging process in a nonlinear manner, being much more profound in the later part of the RBC life span. The RBC deformation index (DI) was measured by an ektacytometry. It is found that the DI decreased with RBC aging in a nonlinear fashion, with increasingly greater changes in the later part of the RBC life span. The alterations of RBC mechanical properties with aging may be attributable to a number of factors, including changes of RBC size and shape, and the viscoelasticity of the cytoplasm and membrane.

Changes of biophysical behavior of k562 cells for p16 gene transfer.

p16 gene was transferred into human erythroleukemia cell line K562 that had been subjected to p16 deletion. The changes of biophysical behavior of K562 cells after gene transfer were studied with the micropipette technique. The micropipette data were analyzed with a standard solid viscoelastic model. In comparison to untransfected control K562 cells and the cells transfected with an empty vector, the p16-transfected K562 cells showed an increase in the elastic element K(1), which is inversely proportional to the maximum deformation over a long period of time, whereas the viscous element mu and the other elastic element K(2) showed no significant difference. The results indicate that K562 cells became more rigid after p16 transfer. The p16-transfected K562 cells also had a higher surface charge density. This study contributes to our knowledge about the suppression effect of p16 gene on tumor cell migration and about its use in gene therapy.

[Tissue culture induced translocation conferring powdery mildew resistance between wheat and Dasypyrum villosum and its marker-assisted selection].

Glutamate oxaloacetate transaminase (GOT) electrophoretic analyses were performed in 175 regenerants arising from immature embryos of crosses between wheat (Triticum aestivum L.) and 6D/6V substitution stocks. The GOT-V2 coding specific enzyme band was absent in two regenerants, designated 98R149 and 98R159 respectively, originated from cross of Yi 4095 and 6D/6V substitution stock c.v. RW15. Pm21 gene linked SCARs (Sequence Characterized Amplified Regions) analysis indicated that 6VS chromosome arms existed in 98R149 and 98R159. Fluorescence in situ hybridization with total genomic DNA extracted from Dasypyrum villosum (L.) as a probe confirmed the occurrence of translocation between 6V chromosome and a wheat one in the two regenerants mentioned above. 98R149 and 98R159 were immune to powdery mildew (Erysiphe graminisDC. f. sp. tritici) inoculation with mix races collected from Hebei Province. The results of the present paper added another feasible example of useful translocations via tissue culture.

[On the relation between substance P and physostigmine on cardiovascular control in rabbit brain].

Using an electromagnetic flowmeter technique, the cardiac output as measured by blood flow in the aortic arch was measured during intracerebroventricular injection (icv.) of Substance P (SP) and physostigmine in 47 anesthetized rabbits. Carotid artery blood pressure and heart rate were recorded. After icv of SP (20 micrograms/20 microliters) or physostigmine (60 micrograms/20 microliters), both cardiac output and mean artery pressure were increased. Pretreatment with icv. of atropine (150 micrograms/20 microliters) did not alter the effect of SP, but the effect of physostigmine was blocked by pretreatment with SP blocker (25 micrograms/20 microliters). These findings suggest that cerebral SP is involved in cholinergic mechanisms on the central control of blood pressure.

A study of RBC aggregation-sedimentation phenomenon with conductivity method.

In this paper, we present a conductivity method for measuring the RBC (red blood cell) aggregation-sedimentation phenomenon and the theoretical analysis of this phenomenon. A theory describing the RBC aggregation-sedimentation phenomenon with the conductivity method is proposed and the formula of the l-t function which depicts the RBC aggregation-sedimentation process is given. We put forward the aggregation index and sedimentation factor describing the RBC aggregation-sedimentation phenomenon for the first time.

Public awareness of stuttering in Shanghai, China.

This study reports the results of an investigation of knowledge and public awareness of stuttering among laypersons in Shanghai, China. A total number of 1968 respondents answered a questionnaire including questions about prevalence, onset, gender distribution and occurrence in different cultures, cause, treatment, intelligence and heredity of stuttering. Stuttering appears to be a disorder that most people in the Shanghai area are familiar with, and for several aspects a considerable number of respondents held views that are in line with current knowledge about stuttering. For some aspects though knowledge is limited, indicating a need for disseminating scientific information on stuttering. The results of the present study are compared with a similar Belgium study.

A nanogold-quenched fluorescence duplex probe for homogeneous DNA detection based on strand displacement.

A nanogold-quenched fluorescence duplex probe has been developed for lighting up homogenous hybridization assays. This novel probe is constructed from two strands of different lengths, and labeled by nanogold and a fluorophore at the long-strand 5'-end and the short-strand 3'-end, respectively. The two tags are in close contact, resulting in complete quenching of the probe fluorescence. If perfectly complemented to the nanogold-labeled strand, a long target oligonucleotide would displace the short fluorophore-labeled strand, and as a result, restore the fluorescence. By using nanogold in the probe, an extremely high quenching efficiency (99.1%) and removal of free fluorophore-labeled strand is achieved. The signal-to-noise ratio and the detection limit (50 pmol L(-1)) of homogenous assays are therefore improved significantly, in comparison with similar probes using organic acceptors. Moreover, the probe has a great inhibition effect on hybridization to a mismatched oligonucleotide. This effect provides the assay with a high specificity, and particularly the assay has great potential in applications for discriminating variations in sequences. The assay sensitivity could be markedly enhanced by using fluorescent materials in the signal strand that are brighter and not quenched by nucleobases.

[A clinical study on the effect of kaisailu on alleviating neonatal jaundice].

The effects of kaisailu defecation on alleviating neonatal jaundice were studied. 203 case of newborns fed with total mother breast milk were divided into two groups: experimental group (100 cases) received routine treatment and kaisailu (10 ml per rectum) control group (103 cases) only received routine treatment. The findings were as follows: excretory time of meconium in experimental group was more ahead than that in control group; the excretory meconium volume of the first time was obviously higher in experimental group; time of meconium changed yellow was distinctly shorter and the bilirubin index (measured by transcortical bilirubin method) lowered more rapidly in experimental group than that in control group. The results suggested that kaisailu defecation can alleviate neonatal jaundice.

Application of statistically-based experimental designs for the optimization of eicosapentaenoic acid production by the diatom Nitzschia laevis.

Statistically based experimental designs were applied to the optimization of medium components and environmental factors for eicosapentaenoic acid (EPA) production by the diatom Nitzschia laevis in heterotrophic conditions. First, the Plackett-Burman design was used to evaluate the effects of variables including medium components and environmental factors on cell growth and EPA production. Among these variables, NaCl, CaCl(2), PI metal solution, pH, and temperature were identified to have the significant effects (with confidence level > 90 %). Subsequently, the concentrations of NaCl, CaCl(2), PI metal solution as well as the values of pH and temperature were optimized using central composite design. The cell growth and EPA production were found to respectively correlate to NaCl, CaCl(2), pH, and temperature that could be represented by second-order polynomial models. The optimal values of the four parameters were determined by response surface and numerical analyses as 8 g/L NaCl, 0.10 g/L CaCl(2), pH 8.5 and 19.8 degrees C for cell dry weight (DW), and 14 g/L NaCl, 0.10 g/L CaCl(2), pH 8.5 and 18 degrees C for EPA production, respectively. The subsequent verification experiments confirmed the validity of the models. This optimization strategy led to a DW of 9 g/L, an EPA yield of 280 mg/L and an EPA productivity of 28 mg/L/d, respectively, which were considerably higher than those obtained in the previous studies.

A perfusion-cell bleeding culture strategy for enhancing the productivity of eicosapentaenoic acid by Nitzschia laevis.

A perfusion-cell bleeding culture strategy was developed for enhancing the productivity of eicosapentaenoic acid (EPA) by the diatom Nitzschia laevis. As the strategy combined the concepts of continuous culture and perfusion culture, it allowed continuously and simultaneously harvesting the algal cells and removing inhibitory compounds during the cultivation. Compared with a single operation of continuous culture, the perfusion-cell bleeding culture greatly enhanced the steady-state biomass concentration, biomass productivity, EPA yield, EPA productivity and glucose utilization efficiency. The perfusion-cell bleeding culture also allowed higher biomass productivity and EPA productivity than the single perfusion culture did. At a bleeding rate of 0.67 day(-1) and a perfusion rate of 0.6 day(-1), the EPA productivity achieved 175 mg l(-1) day(-1), which is the highest ever reported in microalgal cultures.