OBJECTIVETo study the effect of extracellular DNA(eDNA) on the formation of Streptococcus mutans(Sm) biofilms during different growth periods in sucrose environment.

METHODSSm biofilms were established on smooth glass surfaces under the environment of 1% sucrose and cultured in the condition of 37 ℃, 5% O2, 85% N2 and 10% CO2. Samples were randomly divided into four groups based on fourculture time(6,12, 24 and 48 h), respectively. Each group was further divided into two subgroups: control group(without deoxyribonuclease Ⅰ[DNaseⅠ] treatment) and test group(with DNaseⅠtreatment). DNaseⅠ was added 1 h advance in the treatment group to a final concentration of 100 U/ml. Each sample was stained with mixed SYTO-9/PI fluorescent dye. Confocal laser scanning microscopy was used for biofilm observation and scanning. The total biomass, the thickness and the volume of red fluorescence of each biofilm sample were measured following three-dimensional reconstruction using the softwear of Imaris 7.2.3.

RESULTSUnder the environment of 1% sucrose, the Sm bacterial adhesion and distribution density increased over time, the quantity of eDNA and membrane-damaged bacteria which were indicated by red fluorescence also increased within 24 h but dropped later. The biofilm biomasses of Sm biofilm in 6, 12, 24 and 48 h DNaseⅠ treatment group reduced significantly(P<0.05) compared to those in the corresponding control groups by 81.3%, 85.0%, 90.1% and 12.4%, respectively. The biofilm thicknesses in each DNase Ⅰ treatment group (except 6 h group) also reduced significantly(P<0.05) compared to those in the corresponding control group by 34.4%, 45.6% and 23.6%, respectively. The quantities of eDNA and membrane-damaged bacteria reduced in each treatment group except 48 h group compared to that in the corresponding control group.

CONCLUSIONSUnder the environment of 1% sucrose, eDNA plays an important role in promoting the formation of Sm biofilm.

Bacterial Adhesion ; drug effects ; Biofilms ; growth & development ; DNA ; physiology ; Deoxyribonuclease I ; pharmacology ; Microscopy, Confocal ; Streptococcus mutans ; physiology ; Sucrose ; Sweetening Agents ; Temperature

The objective of the study was to analyse Streptococcus mutans biofilms grown under different dietary conditions by using multifaceted methodological approaches to gain deeper insight into the cariogenic impact of carbohydrates. S. mutans biofilms were generated during a period of 24 h in the following media: Schaedler broth as a control medium containing endogenous glucose, Schaedler broth with an additional 5% sucrose, and Schaedler broth supplemented with 1% xylitol. The confocal laser scanning microscopy (CLSM)-based analyses of the microbial vitality, respiratory activity (5-cyano-2,3-ditolyl tetrazolium chloride, CTC) and production of extracellular polysaccharides (EPS) were performed separately in the inner, middle and outer biofilm layers. In addition to the microbiological sample testing, the glucose/sucrose consumption of the biofilm bacteria was quantified, and the expression of glucosyltransferases and other biofilm-associated genes was investigated. Xylitol exposure did not inhibit the viability of S. mutans biofilms, as monitored by the following experimental parameters: culture growth, vitality, CTC activity and EPS production. However, xylitol exposure caused a difference in gene expression compared to the control. GtfC was upregulated only in the presence of xylitol. Under xylitol exposure, gtfB was upregulated by a factor of 6, while under sucrose exposure, it was upregulated by a factor of three. Compared with glucose and xylitol, sucrose increased cell vitality in all biofilm layers. In all nutrient media, the intrinsic glucose was almost completely consumed by the cells of the S. mutans biofilm within 24 h. After 24 h of biofilm formation, the multiparametric measurements showed that xylitol in the presence of glucose caused predominantly genotypic differences but did not induce metabolic differences compared to the control. Thus, the availability of dietary carbohydrates in either a pure or combined form seems to affect the cariogenic potential of S. mutans biofilms.
Bacterial Load ; drug effects ; Bacteriological Techniques ; Biofilms ; drug effects ; Cariogenic Agents ; metabolism ; pharmacology ; Culture Media ; Dental Enamel ; microbiology ; Fluorescent Dyes ; Gene Expression Regulation, Bacterial ; drug effects ; Gene Expression Regulation, Enzymologic ; drug effects ; Genotype ; Glucose ; metabolism ; Glucosyltransferases ; metabolism ; Humans ; Microbial Viability ; drug effects ; Microscopy, Confocal ; Polysaccharides, Bacterial ; biosynthesis ; Streptococcus mutans ; drug effects ; enzymology ; metabolism ; Sucrose ; metabolism ; pharmacology ; Sweetening Agents ; metabolism ; pharmacology ; Tetrazolium Salts ; Time Factors ; Up-Regulation ; Xylitol ; metabolism ; pharmacology

Streptococcus mutans is a common Gram-positive bacterium and plays a significant role in dental caries. Tobacco and/or nicotine have documented effects on S. mutans growth and colonization. Sortase A is used by many Gram-positive bacteria, including S. mutans, to facilitate the insertion of certain cell surface proteins, containing an LPXTGX motif such as antigen I/II. This study examined the effect of nicotine on the function of sortase A to control the physiology and growth of S. mutans using wild-type S. mutans NG8, and its isogenic sortase-defective and -complemented strains. Briefly, the strains were treated with increasing amounts of nicotine in planktonic growth, biofilm metabolism, and sucrose-induced and saliva-induced antigen I/II-dependent biofilm formation assays. The strains exhibited no significant differences with different concentrations of nicotine in planktonic growth assays. However, they had significantly increased (P≤0.05) biofilm metabolic activity (2- to 3-fold increase) as the concentration of nicotine increased. Furthermore, the sortase-defective strain was more sensitive metabolically to nicotine than the wild-type or sortase-complemented strains. All strains had significantly increased sucrose-induced biofilm formation (2- to 3-fold increase) as a result of increasing concentrations of nicotine. However, the sortase-defective strain was not able to make as much sucrose- and saliva-induced biofilm as the wild-type NG8 did with increasing nicotine concentrations. These results indicated that nicotine increased metabolic activity and sucrose-induced biofilm formation. The saliva-induced biofilm formation assay and qPCR data suggested that antigen I/II was upregulated with nicotine but biofilm was not able to be formed as much as wild-type NG8 without functional sortase A.
Amino Acid Motifs ; Aminoacyltransferases ; drug effects ; genetics ; Antigens, Bacterial ; drug effects ; Bacterial Adhesion ; drug effects ; Bacterial Proteins ; drug effects ; genetics ; Biofilms ; drug effects ; Cysteine Endopeptidases ; drug effects ; genetics ; Dose-Response Relationship, Drug ; Humans ; Mutation ; genetics ; Nicotine ; administration & dosage ; pharmacology ; Peptidoglycan ; drug effects ; genetics ; Saliva ; physiology ; Streptococcus mutans ; drug effects ; enzymology ; growth & development ; Sucrose ; pharmacology

OBJECTIVETo observe the effect of hesperidin on behavior and hypothalamic-pituitary-adrenal (HPA) axis of ratmodel of chronic stress-induced depression.

METHODChronic unpredictable mild stress (CUMS) was used to establish the rat depression model. Sixty male SD rats were divided randomly into six groups: the normal group, the model group, the hesperidin (40, 80, 160 mg x kg(-1)) group and the positive fluoxetine (10 mg x kg(-1)) group. They were orally administered with drugs for three weeks. The sucrose preference test and the forced swimming test (FST) were assayed to detect animal behavior. The levels of corticosterone (CORT) in serum, mRNA of corticotropin release factor (CRF) in hypothalamus as well as protein expression of glucocorticoid receptor (GR) in paraventricular nucleus (PVN) were determined to clarify the anti-depression effect and mechanism of hesperidin.

RESULTCompared with the model group, rats in the hesperidin (40, 80, 160 mg x kg(-1)) treatment group showed significant increase in the sucrose consumption and decrease in the immobility time in FST to varying degrees. Meanwhile, the excessively high serum CORT and adrenal index of CUMS rats were reversed by treatment with hesperidin. In addition, hesperidin inhibited CRF mRNA expression in hypothalamus and up-regulated GR protein expression in PVN among CUMS rats.

CONCLUSIONHesperidin could effectively improve the behavior of CUMS rats and show the anti-depression effect. Its mechanisms may be related to the function of regulating HPA axis.

Administration, Oral ; Animals ; Behavior, Animal ; drug effects ; Corticosterone ; blood ; Corticotropin-Releasing Hormone ; genetics ; metabolism ; Depression ; drug therapy ; etiology ; Fluoxetine ; administration & dosage ; Gene Expression Regulation ; drug effects ; Hesperidin ; administration & dosage ; pharmacology ; Hypothalamo-Hypophyseal System ; drug effects ; physiopathology ; Hypothalamus ; metabolism ; Male ; Models, Animal ; Pituitary-Adrenal System ; drug effects ; physiopathology ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, Glucocorticoid ; metabolism ; Stress, Psychological ; complications ; drug therapy ; Sucrose ; metabolism ; Swimming ; Up-Regulation

OBJECTIVETo study the effect of sucrose and plant growth substances of different concentrations on the induction of test-tube tuberous roots of Rehmannia glutinosa, in order to establish an efficient system for the induction of test-tube tuberous roots from leaves of R. glutinosa.

METHODLeaves from test-tube seedlings of 85-5 R. glutinosa were used as explants. After rooting induction, they were transferred to medium with orthogonal design for inducing test-tube tuberous roots of R. glutinosa.

RESULT AND CONCLUSIONNAA played a significant role in induction of test-tube tuberous roots of R. glutinosa, followed by sucrose and 6-BA. With leaves from test-tube seedlings as the explants, the optimal medium for inducing test-tube tuberous roots of R. glutinosa was MS + BA 3.0 mg x L(-1) + NAA 0.1 mg x L(-1) + sucrose 7%. The study provides an efficient induction system for studies on artificial seeds and secondary metabolism with test-tube tuberous roots of R. glutinosa.

Benzyl Compounds ; Dose-Response Relationship, Drug ; Kinetin ; pharmacology ; Naphthaleneacetic Acids ; pharmacology ; Plant Growth Regulators ; pharmacology ; Plant Leaves ; drug effects ; growth & development ; Plant Roots ; drug effects ; growth & development ; Purines ; Rehmannia ; drug effects ; growth & development ; Seedlings ; drug effects ; growth & development ; Sucrose ; pharmacology ; Tissue Culture Techniques ; instrumentation ; methods

OBJECTIVETo cultivate adventitious roots of Hypericum perforatum in bioreactors, in order to seek for suitable conditions for adventitious growth.

METHODThe effect of IBA concentration, sugar type and concentration, inoculum volume and air volume of adventitious roots on the cultivation of adventitious roots of H. perforatum was observed in a 5 L air-lift bioreactor.

RESULTAdventitious roots of H. perforatum were cultivated in a MS culture dish. With the increase of IBA concentration, the propagation coefficient of adventitious roots of H. perforatum was on the rise. The IBA concentration ranging between 1.25-1.75 mg x L(-1) was suitable for the growth of adventitious roots. Adventitious roots grew best with sucrose in MS medium, with the propagation coefficient up to 22.15. When sucrose concentration was 30 g x L(-1), fresh weight, dry weight and propagation coefficient reached the maximum value. An adventitious root reactor with an inoculum volume of 20 g was favorable for the growth of adventitious roots. The air volume of reactors of 0.075 vvm (air volume/culture volume per minute) was favorable for the growth of adventitious roots, with the significant increase in the propagation coefficient of adventitious roots. In the amplification experiment, we found that the cultivation conditions of adventitious roots in a 5 L bioreactor was completely applicable to that in 10 and 20 L bioreactors, and adventitious roots grew well in a large bioreactor.

CONCLUSIONIBA concentration, sugar type and concentration, inoculum volume and air volume had a significant effect on the growth of adventitious roots.

Air ; Biomass ; Bioreactors ; Carbohydrates ; pharmacology ; Dose-Response Relationship, Drug ; Hypericum ; drug effects ; growth & development ; Indoles ; pharmacology ; Plant Roots ; drug effects ; growth & development ; Sucrose ; pharmacology ; Tissue Culture Techniques ; instrumentation ; methods

The purpose of our study was to evaluate the effect of photodynamic therapy (PDT), using erythrosine as a photosensitizing agent and a dental halogen curing unit as a light source, on Streptococcus mutans in a biofilm phase. The S. mutans biofilms were formed in a 24-well cell culture cluster. Test groups consisted of biofilms divided into four groups: group 1: no photosensitizer or light irradiation treatment (control group); group 2: photosensitizer treatment alone; group 3: light irradiation alone; group 4: photosensitizer treatment and light irradiation. After treatments, the numbers of colony-forming unit (CFU) were counted and samples were examined by confocal laser scanning fluorescence microscopy (CLSM). Only group 4 (combined treatment) resulted in significant increases in cell death, with rates of 75% and 55% after 8 h of incubation, and 74% and 42% at 12 h, for biofilms formed in brain-heart infusion (BHI) broth supplemented with 0% or 0.1% sucrose, respectively. Therefore, PDT of S. mutans biofilms using a combination of erythrosine and a dental halogen curing unit, both widely used in dental clinics, resulted in a significant increase in cell death. The PDT effects are decreased in biofilms that form in the presence of sucrose.
Bacterial Load ; drug effects ; Bacteriological Techniques ; Biofilms ; drug effects ; Curing Lights, Dental ; classification ; Erythrosine ; therapeutic use ; Humans ; Microbial Viability ; drug effects ; Microscopy, Confocal ; Photochemotherapy ; methods ; Photosensitizing Agents ; therapeutic use ; Sonication ; Streptococcus mutans ; drug effects ; Sucrose ; pharmacology ; Time Factors

This study is to observe anti-lipotoxic effect of sesamin on renovascular hypertensive rats fed with a high-fat, high-sucrose diet. Thirty-four complex model rats were induced by two-kidney, one-clip method and on high-fat and refined-carbohydrate diet for thirteen weeks. From the fifth week, intragastric administration of sesamin (120, 60 and 30 mg x kg(-1) x d(-1)) lasted for eight weeks. Blood pressure (BP), blood fat (BF), blood glucose (BG), free fatty acids (FFA), insulin (Ins), tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 were determined. Pathological changes of pancreas, perirenal fat and liver were semiquantitatively analyzed. In sesamin (120 and 60 mg x kg(-1) x d(-1)) group, it was found that there were decrease of levels of BP, BF, BG, TNF-alpha, IL-6 and FFA, improvement of insulin resistance and glucose tolerance, alleviation of body weight, humid weight of fat, liver and pancreas and their organ index, and reduction of islet cell hyperplasia and amount of lipid droplet vacuoles in lipocyte and hepatocyte. It is implied that sesamin had anti-lipotoxic effect and its mechanism may be closely associated with the amelioration of insulin resistance via reducing lipidoses in hepatocyte and inflammatory adipokines such as TNF-alpha and IL-6.
Adipocytes ; drug effects ; Animals ; Anticholesteremic Agents ; administration & dosage ; pharmacology ; Antihypertensive Agents ; administration & dosage ; pharmacology ; Blood Glucose ; metabolism ; Blood Pressure ; drug effects ; Body Weight ; drug effects ; Cholesterol ; blood ; Diet, High-Fat ; Dioxoles ; administration & dosage ; pharmacology ; Dose-Response Relationship, Drug ; Fatty Acids, Nonesterified ; blood ; Glucose Tolerance Test ; Hypertension, Renovascular ; blood ; pathology ; Insulin ; blood ; Insulin Resistance ; Interleukin-6 ; blood ; Islets of Langerhans ; pathology ; Lignans ; administration & dosage ; pharmacology ; Liver ; pathology ; Male ; Pancreas ; pathology ; Rats ; Rats, Sprague-Dawley ; Sucrose ; Triglycerides ; blood ; Tumor Necrosis Factor-alpha ; blood

OBJECTIVETo explore the feasibility of ultra-rapid freezing of human spermatozoa in the cryogenic vial with different concentrations of sucrose solution.

METHODSWe divided 40 normal semen samples prepared with the routine swim-up technique into 6 aliquots, 1 as the control and the other 5 cryopreserved with sucrose solution at the concentrations of 0.15, 0.20, 0.25 and 0.30 mol/L, respectively. After thawing, we determined and compared the motility, progressive motility and plasma membrane integrity of the sperm among the 6 groups.

RESULTSThe motility, progressive motility and plasma membrane integrity of the sperm were significantly lower after thawing than before cryopreservation ([96.2 +/- 1.8]%, [93.8 +/- 2.8]% and [99.0 +/- 0.8 ]%) (P<0.05). Post-thawing sperm motility was (55.5 +/- 6.3)% in the 0.20 mol/L sucrose group, significantly higher than in the 0.15, 0.25 and 0.30 mol/L groups ([45.9 +/- 6.6]%, [50.4 +/- 9.4]% and [45.5 +/- 11.2]%) (P<0.05), and it was (53.6 +/- 5.0)% in the conventional freezing group, with no statistically significant difference from the 0.20 and 0.25 mol/L sucrose cryopreservation groups (P> 0.05), but remarkably higher than in the 0.15 and 0.30 mol/L groups (P<0.05). Post-thawing progressive sperm motility exhibited no statistically significant differences between the 0.20 mol/L sucrose and conventional freezing groups ([44.4 +/- 7.4]% vs [42.3 +/- 8.1]%, P>0.05), but markedly higher in both than in the 0.15, 0.25 and 0.30 mol/L sucrose groups ([37.1 +/- 8.3 ]%, [33.1 +/- 9.2]% and [22.0 +/- 9.1]%) (P<0.05). Post-thawing plasma membrane integrity was significantly higher in the 0.20 mol/L sucrose cryopreservation group ( [70.1 +/- 6.9]%) than in either the conventional freezing group ([63.1 +/- 6.8]%) or the 0.15, 0.25 and 0.30 mol/L sucrose groups ([57.7 +/- 8.3]%, [63.5 +/- 10.7]% and [57.8 +/- 12.9]%) (P<0.05).

CONCLUSIONAs a simple, safe and effective method, ultra-rapid freezing with sucrose solution at the final concentration of 0.20 mol/L can be used for the cryopreservation of human spermatozoa.

Cell Membrane ; drug effects ; Cryopreservation ; methods ; Humans ; Male ; Semen Preservation ; methods ; Sperm Motility ; drug effects ; Sucrose ; administration & dosage ; pharmacology

Taxus suspension cell culture has the potential to provide a sustainable source of anticancer drug paclitaxel (Taxol) and other taxoids. In the cell culture of Taxus chinensis, Taxuyunnanine C (Tc) is the primary taxoid. To design a rational strategy for redirecting the precursor fluxes from other taxoids into paclitaxel production, we employed Real-time Quantitative PCR (RQ-PCR) to understand the dynamic profiling of key biosynthetic pathway genes of palcitaxel and taxoids during the culture process. Six genes (TASY, TDAT, T5alphaH, TalphaH, T10betaH and T14betaH) were quantified under the process condition of double elicitation by 2,3-dihydroxylpropanyl jasmonate (DHPJA) (100 micromol/L on day 7 and day 12), and sucrose feeding (20 g/L) on day 7. This process treatment led to a high accumulation of Tc at (554.46 +/- 21.28) mg/L 8 days after the first elicitation. Then 9 days after the second elicitation, Tc production was as high as (997.72 +/- 1.51) mg/L. The early pathway genes TASY and TDAT were significantly up-regulated by 182-fold and 98-fold, respectively for the first DHPJA elicitation and by 208-fold and 131-fold, respectively for the second elicitation. The induction occurred after each elicitation lasted for about 24 h before their abundances decreased. Things are somewhat different in the case of the other four genes T5alphaH, TalphaH, T10betaH and T14betaH. For gene TalphaH, it was highly up-regulated by 3061-fold for the first DHPJA elicitation and by 1016-fold for the second elicitation. For the other three genes T5alphaH, T10betaH, T14betaH, they were up-regulated by 13-fold, 38-fold and 20-fold, respectively for the first DHPJA elicitation and by 7-fold, 16-fold and 6-fold, respectively for the second elicitation. The RQ-PCR results showed that there is tight correlation between gene expression and Tc accumulation. Gene expression was in accordance with Tc yield. Elicitation could improve expression of six genes. While along with culture course, high expression of the genes weakened. Elicitation for the second time would promote high expression of the genes again.
Cell Culture Techniques ; Culture Media ; Cyclopentanes ; pharmacology ; Gene Expression Regulation, Plant ; Oxylipins ; pharmacology ; Plant Growth Regulators ; pharmacology ; Sucrose ; pharmacology ; Taxoids ; metabolism ; Taxus ; genetics ; metabolism ; Transcriptome