131 APOPTOSIS AND DEVELOPMENT OF IN VITRO-PRODUCED BOVINE EMBRYOS EXPOSED TO SUPRAPHYSIOLOGICAL CONCENTRATIONS OF INSULIN-LIKE GROWTH FACTOR-1 (IGF-1)

2009 ◽  
Vol 21 (1) ◽  
pp. 165
Author(s):  
M. A. Velazquez ◽  
H. Niemann
Keyword(s):  
Laser Scanning ◽  
Apoptotic Cell ◽  
Polycystic Ovary ◽  
Statistical Significance ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Control Group ◽  
Fisher Exact Test ◽  
Bovine Embryos

It has been hypothesized that high non-physiological IGF-1 levels are partially responsible for the recurrent pregnancy loss observed in women with the polycystic ovary syndrome (Eng GS et al. 2007 Diabetes 56, 2228–2234). The aim of this study was to determine the effect of supraphysiological concentrations of IGF-1 on blastocyst production and the occurrence of apoptosis in bovine embryos, which are a good model for human embryo development (Baumann CG et al. 2007 Mol. Reprod. Dev. 74, 1345–1353). COC obtained by slicing from abattoir ovaries were matured (TCM-199, Sigma) for 24 h and fertilized (Fert-TALP) for 18 h (Day 0) in vitro. Two different IGF-1 (Recombinant human IGF-1, R&D Systems GmbH, Wiesbaden, Germany) concentrations (supraphysiological = 1000 ng mL–1 and physiological = 100 ng mL–1) were added to the culture media (Synthetic oviduct fluid/BSA) and compared with a control group (no IGF-1 supplementation). On Day 8, blastocyst rates (22 replicates) were recorded and DNA degradation was detected in blastocyst nuclei using a cell death detection kit (Roche Diagnostics GmbH, Mannheim, Germany) based on the terminal deoxinucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) principle. Embryos (n = 27 [control], n = 29 [both IGF-1 groups]) from 4 replicates were examined by confocal laser scanning microscopy. Data were analyzed by ANOVA and the Fisher exact test using the SigmaStat 2.0 software package (Jandel Scientific, San Rafael, CA). Cleavage was numerically improved by both, 1000 (59.1 ± 1.8) and 100 (58.2 ± 2.8) ng IGF-1 over controls (53.5 ± 2.2), but the differences did not reach statistical significance (P = 0.22). The proportion of hatched blastocysts was enhanced by 100 (5.8 ± 1.0, P = 0.03) and 1000 (5.1 ± 0.7, P = 0.03) ng IGF-1 compared to controls (2.8 ± 0.6). Total blastocyst rate was increased by 100 ng IGF-1 (34.4 ± 1.9, P = 0.02) over controls (28.3 ± 1.7), but not by 1000 ng IGF-1 (29.1 ± 1.6 P = 0.75). The 100 ng IGF-1 group (38.5 ± 3.7) had fewer degenerated embryos (P = 0.01) compared to 1000 ng IGF-1 (49.7 ± 3.3). The proportion of embryos displaying at least one apoptotic cell was greater in the 1000 ng IGF-1 group over controls (96% v. 77% P = 0.04). The number of blastomeres with TUNEL-positive nuclei per embryo was higher in the supraphysiological group (5.5 ± 0.6, P < 0.001) compared with the control (2.3 ± 0.4) and the physiological group (2.5 ± 0.3). There were no significant differences between the control and the 100 ng IGF-1 group in this regard (P = 0.49). In conclusion, supraphysiological concentrations of IGF-1 do not increase blastocyst production but increase levels of apoptosis in bovine embryos produced in vitro. M. A. V. is in the PhD program of the University of Veterinary medicine, Hannover, Germany, and is supported by the German Academic Exchange Service (DAAD)

scholarly journals Susceptibility of Mature Staphylococcus Biofilms to Chinese Herbal Decoction Sanhuang Jiedu: An In Vitro Study

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Shaoe Zhang ◽  
Xiao Wang ◽  
Xiaotao Shi ◽  
Honglue Tan ◽  
Himanshu Garg
Keyword(s):  
Laser Scanning ◽  
Multidrug Resistant ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Control Group ◽  
Antibiofilm Activity ◽  
Dependent Manner ◽  
Chinese Herbal ◽  
Titanium Surfaces

Background. External socking and washing with the Chinese herbal Sanhuang Jiedu decoction (SHJD) can effectively control local limb infections with bone and implant exposure. However, the antibiofilm activities of this decoction in vitro have not yet been investigated. Therefore, the aim of this study was to examine the effects and characteristics of SHJD on the mature biofilms of multidrug-resistant staphylococci on a titanium surface. Methods. Biofilm-forming methicillin-resistant Staphylococcus epidermidis ATCC 35984 and S. aureus ATCC 43330, and non-biofilm-forming S. epidermidis ATCC 12228 were selected as the experimental strains. The mature biofilms were prepared on titanium surfaces. The five experimental groups were based on dilution concentrations (DC) of SHJD: the control group (biofilm incubated with 0.85% NaCl solution), the SHJD (DC:1/8) group (initial SHJD solution was diluted 1/8), the SHJD (DC:1/4) group, the SHJD (DC:1/2) group, and the SHJD (DC:1/1) group (initial SHJD solution). The effects of SHJD on the mature biofilms were observed with the bacterial spread plate method, crystal violet (CV) staining, scanning electron microscopy, and confocal laser scanning microscopy. Results. After culture in tryptic soy broth for 72 h, ATCC 43300 and ATCC 35984 produced mature biofilms and ATCC 12228 did not. The optical density value of ATCC 12228 was 0.11 ± 0.02 , significantly lower than that of ATCC 35984 ( 0.42 ± 0.05 ) or ATCC 43300 ( 0.41 ± 0.03 ) ( P < 0.05 ). The mature biofilms of ATCC 43300 and ATCC 35984 clearly disintegrated when incubated for 12–24 h with SHJD (DC:1/1) or SHJD (DC:1/2), showing only scattered bacterial adhesion. In the SHJD (DC:1/4) group, although many residual bacterial colonies still clustered together, presenting a biofilm structure, it was very looser than that in the SHJD (DC:1/8) group in which the biofilm was similar to that in the control group. For ATCC 12228, only colony adhesion was observed, and the number of colonies decreased as the concentration of SHJD or the culture period increased. The quantitative results for the bacterial spread plate and CV staining showed significant differences between the SHJD groups ( P < 0.05 ). Conclusion. SHJD has antibiofilm activity against multidrug-resistant Staphylococcus strains. It weakens or disrupts already-formed mature biofilms on titanium surfaces in a concentration- and incubation time-dependent manner.

scholarly journals Analysis of Shear Bond Strength and Morphology of Er:YAG Laser-Recycled Ceramic Orthodontic Brackets

2016 ◽  
Vol 2016 ◽  
pp. 1-6 ◽  
Author(s):  
Ruo-qiao Han ◽  
Kai Yang ◽  
Ling-fei Ji ◽  
Chen Ling
Keyword(s):  
Bond Strength ◽  
Shear Bond Strength ◽  
Laser Scanning ◽  
Er:Yag Laser ◽  
Statistical Significance ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Control Group ◽  
Test Machine ◽  
Ceramic Brackets

Objective. The aim of this study was to compare the recycling of deboned ceramic brackets via an Er:YAG laser or via the traditional chairside processing methods of flaming and sandblasting; shear bond strength and morphological changes were evaluated in recycled brackets versus new brackets.Materials and Methods. 3M Clarity Self-Ligating Ceramic Brackets with a microcrystalline base were divided into groups subjected to flaming, sandblasting, or exposure to an Er:YAG laser. New ceramic brackets served as a control group. Shear bond strengths were determined with an Electroforce test machine and tested for statistical significance through analysis of variance. Morphological examinations of the recycled ceramic bracket bases were conducted with scanning electron microscopy and confocal laser scanning microscopy. Residue on the bracket base was analyzed with Raman spectroscopy.Results. Faded, dark adhesive was left on recycled bracket bases processed via flaming. Adhesive was thoroughly removed by both sandblasting and exposure to an Er:YAG laser. Compared with new brackets, shear bond strength was lower after sandblasting (p<0.05), but not after exposure to an Er:YAG laser. The Er:YAG laser caused no damage to the bracket.Conclusion. Er:YAG lasers effectively remove adhesive from the bases of ceramic brackets without damaging them; thus, this method may be preferred over other recycling methods.

scholarly journals Effects of cigarette smoking on the growth of Streptococcus mutans biofilms: an in vitro study

2021 ◽  
Author(s):  
Ye Han
Keyword(s):  
Treatment Group ◽  
Cigarette Smoking ◽  
Streptococcus Mutans ◽  
Laser Scanning ◽  
Water Soluble ◽  
Laser Scanning Microscopy ◽  
Extracellular Polysaccharides ◽  
Confocal Laser ◽  
Control Group

Abstract This study aimed to investigate the differences in growth and virulence (EPSs and acidogenicity) of Streptococcus mutans biofilms (S. mutans) according to the different times of cigarette smoking (CS) treatment. S. mutans biofilms (74-hour-old) were formed on saliva-coated hydroxyapatite disks. The biofilms were treated with CS at different times per day (one time, three times, and six times/day). The control group did not receive CS treatment. Acidogenicity, dry weight, colony-forming units, water-soluble/insoluble extracellular polysaccharides, and intracellular polysaccharides were analyzed and confocal laser scanning microscopy images were obtained of the 74-h-old biofilms. The 74-h-old biofilms on sHA discs in the 6 times/day CS treatment group showed the lowest biofilm accumulation and extracellular polysaccharide amount compared with the control group and other CS treatment groups. In the CLSM study, the biofilms in the six times/day CS treatment group also showed the lowest bacterial count (live and dead cells) and EPS biovolume. CS has an obvious inhibition on the growth of S. mutans biofilms, the degree of inhibition is proportional to the number of CS treatments.

scholarly journals Curcumin induces apoptosis in trophoblast model cell line

2018 ◽  
Vol 27 (2) ◽  
Author(s):  
Tatit Nurseta ◽  
Yahya Irwanto ◽  
I W.A. Wiyasa ◽  
Rahajeng Rahajeng ◽  
Imelda Imelda ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Nuclear Translocation ◽  
Hydatidiform Mole ◽  
In Vitro Study ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Control Group ◽  
Trophoblast Cells ◽  
Enos Expression

Background: Several studies have reported that curcumin exerts chemopreventive effects in various type of cancers, through several mechanisms, however, the effect of curcumin on carcinogenesis in patients with hydatidiform mole has not yet been investigated. This study was conducted to evaluate the effect of curcumin on apoptosis, proliferation, and nuclear translocation of endothelial nitricoxide synthase in trophoblast cells induced by estradiol in complete hydatidiform mole (CHM).Methods: In this in vitro study, trophoblast cells were divided into six groups, the control group (trophoblast cells were exposed to 100 pg/mL of 17-β estradiol) and the treatment group (trophoblast cells were exposed to 100 pg/mL of 17-β estradiol in the presence of curcumin with doses: 50, 100, 200, 400, and 800 µM). At the end of study, the cell proliferation was analyzed using MTT assay and apoptosis with TUNEL test in each group thropoblast cell. eNOS translocation was assayed using confocal laser scanning microscopy at the various dose of curcumin.Results: Curcumin at the doses of 200, 400, and 800 µM significantly decreased the proliferation and increased the apoptotic index in curcumin-treated group compared to those in the control group (p<0.05). All doses of curcumin treatment significantly decreased the nuclear eNOS expression compared to that in the control group. The three highest doses of curcumin increased cytoplasmic eNOS expression compared to that in control group.Conclusion: Curcumin inhibits the proliferation and modulates the apoptosis of trophoblast cells induced by estradiol in CHM involvement.

Vacuoles Induced in Mammalian Cells by Helicobacter Cytotoxin are Acidic Organelles

1990 ◽  
Vol 48 (3) ◽  
pp. 168-169
Author(s):  
M. H. Chestnut ◽  
C. E. Catrenich
Keyword(s):  
Acridine Orange ◽  
Mammalian Cells ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Ulcer Disease ◽  
Gastroduodenal Disease ◽  
H Pylori

Helicobacter pylori is a non-invasive, Gram-negative spiral bacterium first identified in 1983, and subsequently implicated in the pathogenesis of gastroduodenal disease including gastritis and peptic ulcer disease. Cytotoxic activity, manifested by intracytoplasmic vacuolation of mammalian cells in vitro, was identified in 55% of H. pylori strains examined. The vacuoles increase in number and size during extended incubation, resulting in vacuolar and cellular degeneration after 24 h to 48 h. Vacuolation of gastric epithelial cells is also observed in vivo during infection by H. pylori. A high molecular weight, heat labile protein is believed to be responsible for vacuolation and to significantly contribute to the development of gastroduodenal disease in humans. The mechanism by which the cytotoxin exerts its effect is unknown, as is the intracellular origin of the vacuolar membrane and contents. Acridine orange is a membrane-permeant weak base that initially accumulates in low-pH compartments. We have used acridine orange accumulation in conjunction with confocal laser scanning microscopy of toxin-treated cells to begin probing the nature and origin of these vacuoles.

scholarly journals Effect of dissolution rate and subsequent ion release on cytocompatibility properties of borophosphate glasses

2019 ◽  
Vol 5 (1) ◽  
pp. 85-97
Author(s):  
Nusrat Sharmin ◽  
Mohammad S. Hasan ◽  
Md. Towhidul Islam ◽  
Chengheng Pang ◽  
Fu Gu ◽  
...  
Keyword(s):  
Dissolution Rate ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Ion Release ◽  
Scanning Calorimetry ◽  
Cell Culture Study ◽  
Borophosphate Glasses ◽  
Mg63 Cells

AbstractPresent work explores the relationship between the composition, dissolution rate, ion release and cytocompatibility of a series of borophosphate glasses. While, the base glass was selected to be 40mol%P2O5-16mol%CaO-24mol%MgO-20mol%Na2O, three B2O3 modified glass compositions were formulated by replacing Na2O with 1, 5 and 10 mol% B2O3. Ion release study was conducted using inductively coupled plasma atomic emission spectroscopy (ICP-AES). The thermal scans of the glasses as determined by differential scanning calorimetry (DSC) revealed an increment in the thermal properties with increasing B2O3 content in the glasses. On the other hand, the dissolution rate of the glasses decreased with increasing B2O3 content. To identify the effect of boron ion release on the cytocompatibility properties of the glasses, MG63 cells were cultured on the surface of the glass discs. The in vitro cell culture study suggested that glasses with 5 mol% B2O3 (P40B5) showed better cell proliferation and metabolic activity as compares to the glasses with 10 mol% (P40B10) or with no B2O3 (P40B0). The confocal laser scanning microscopy (CLSM) images of live/dead stained MG63 cells attached to the surface of the glasses also revealed that the number of dead cells attached to P40B5 glasses were significantly lower than both P40B0 and P40B10 glasses.

scholarly journals Cephalosporin nitric oxide-donor prodrug DEA-C3D disperses biofilms formed by clinical cystic fibrosis isolates of Pseudomonas aeruginosa

2019 ◽  
Vol 75 (1) ◽  
pp. 117-125 ◽  
Author(s):  
Odel Soren ◽  
Ardeshir Rineh ◽  
Diogo G Silva ◽  
Yuming Cai ◽  
Robert P Howlin ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Clinical Isolates ◽  
Nitric Oxide Donor ◽  
Confocal Laser ◽  
Broth Microdilution Method

Abstract Objectives The cephalosporin nitric oxide (NO)-donor prodrug DEA-C3D (‘DiEthylAmin-Cephalosporin-3′-Diazeniumdiolate’) has been shown to initiate the dispersal of biofilms formed by the Pseudomonas aeruginosa laboratory strain PAO1. In this study, we investigated whether DEA-C3D disperses biofilms formed by clinical cystic fibrosis (CF) isolates of P. aeruginosa and its effect in combination with two antipseudomonal antibiotics, tobramycin and colistin, in vitro. Methods β-Lactamase-triggered release of NO from DEA-C3D was confirmed using a gas-phase chemiluminescence detector. MICs for P. aeruginosa clinical isolates were determined using the broth microdilution method. A crystal violet staining technique and confocal laser scanning microscopy were used to evaluate the effects of DEA-C3D on P. aeruginosa biofilms alone and in combination with tobramycin and colistin. Results DEA-C3D was confirmed to selectively release NO in response to contact with bacterial β-lactamase. Despite lacking direct, cephalosporin/β-lactam-based antibacterial activity, DEA-C3D was able to disperse biofilms formed by three P. aeruginosa clinical isolates. Confocal microscopy revealed that DEA-C3D in combination with tobramycin produces similar reductions in biofilm to DEA-C3D alone, whereas the combination with colistin causes near complete eradication of P. aeruginosa biofilms in vitro. Conclusions DEA-C3D is effective in dispersing biofilms formed by multiple clinical isolates of P. aeruginosa and could hold promise as a new adjunctive therapy to patients with CF.

scholarly journals Use of electromagnetic stimulation on an Enterococcus faecalis biofilm on root canal treated teeth in vitro

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Beatriz H. D. Panariello ◽  
Justin K. Kindler ◽  
Kenneth J. Spolnik ◽  
Ygal Ehrlich ◽  
George J. Eckert ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Root Canal ◽  
Laser Scanning ◽  
Confocal Imaging ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Microscopy Imaging ◽  
Electromagnetic Stimulation ◽  
Root Canal Disinfection

AbstractRoot canal disinfection is of utmost importance in the success of the treatment, thus, a novel method for achieving root canal disinfection by electromagnetic waves, creating a synergistic reaction via electric and thermal energy, was created. To study electromagnetic stimulation (EMS) for the disinfection of root canal in vitro, single rooted teeth were instrumented with a 45.05 Wave One Gold reciprocating file. Specimens were sterilized and inoculated with Enterococcus faecalis ATCC 29,212, which grew for 15 days to form an established biofilm. Samples were treated with 6% sodium hypochlorite (NaOCl), 1.5% NaOCl 1.5% NaOCl with EMS, 0.9% saline with EMS or 0.9% saline. After treatments, the colony forming units (CFU) was determined. Data was analyzed by Wilcoxon Rank Sums Test (α = 0.05). One sample per group was scored and split for confocal laser scanning microscopy imaging. There was a significant effect with the use of NaOCl with or without EMS versus 0.9% saline with or without EMS (p = 0.012 and 0.003, respectively). CFUs were lower when using 0.9% saline with EMS versus 0.9% saline alone (p = 0.002). Confocal imaging confirmed CFU findings. EMS with saline has an antibiofilm effect against E. faecalis and can potentially be applied for endodontic disinfection.

scholarly journals Cocktail of carbohydrases from Aspergillus niger: an economical and eco-friendly option for biofilm clearance from biopolymer surfaces

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Arashdeep Kaur ◽  
Sanjeev Kumar Soni ◽  
Shania Vij ◽  
Praveen Rishi
Keyword(s):  
Aspergillus Niger ◽  
Laser Scanning ◽  
Statistical Modelling ◽  
Laser Scanning Microscopy ◽  
Enzyme Treatment ◽  
Confocal Laser ◽  
Abiotic Surfaces ◽  
Enzyme Cocktail ◽  
Natural Variant

AbstractBiofilm formation on both biotic and abiotic surfaces accounts for a major factor in spread of antimicrobial resistance. Due to their ubiquitous nature, biofilms are of great concern for environment as well as human health. In the present study, an integrated process for the co-production of a cocktail of carbohydrases from a natural variant of Aspergillus niger was designed. The enzyme cocktail was found to have a noteworthy potential to eradicate/disperse the biofilms of selected pathogens. For application of enzymes as an antibiofilm agent, the enzyme productivities were enhanced by statistical modelling using response surface methodology (RSM). The antibiofilm potential of the enzyme cocktail was studied in terms of (i) in vitro cell dispersal assay (ii) release of reducing sugars from the biofilm polysaccharides (iii) the effect of enzyme treatment on biofilm cells and architecture by confocal laser scanning microscopy (CLSM). Potential of the enzyme cocktail to disrupt/disperse the biofilm of selected pathogens from biopolymer surfaces was also assessed by field emission scanning electron microscopy (FESEM) analysis. Further, their usage in conjunction with antibiotics was assessed and it was inferred from the results that the use of enzyme cocktail augmented the efficacy of the antibiotics. The study thus provides promising insights into the prospect of using multiple carbohydrases for management of heterogeneous biofilms formed in natural and clinical settings.

scholarly journals pH and Reduction Dual-Responsive Bi-Drugs Conjugated Dextran Assemblies for Combination Chemotherapy and In Vitro Evaluation

Polymers ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 1515
Author(s):  
Xiukun Xue ◽  
Yanjuan Wu ◽  
Xiao Xu ◽  
Ben Xu ◽  
Zhaowei Chen ◽  
...  
Keyword(s):  
Combination Chemotherapy ◽  
Laser Scanning ◽  
Rational Design ◽  
A549 Cells ◽  
Chemotherapeutic Agents ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Tumor Resistance ◽  
Oxidized Dextran

Polymeric prodrugs, synthesized by conjugating chemotherapeutic agents to functional polymers, have been extensively investigated and employed for safer and more efficacious cancer therapy. By rational design, a pH and reduction dual-sensitive dextran-di-drugs conjugate (oDex-g-Pt+DOX) was synthesized by the covalent conjugation of Pt (IV) prodrug and doxorubicin (DOX) to an oxidized dextran (oDex). Pt (IV) prodrug and DOX were linked by the versatile efficient esterification reactions and Schiff base reaction, respectively. oDex-g-Pt+DOX could self-assemble into nanoparticles with an average diameter at around 180 nm. The acidic and reductive (GSH) environment induced degradation and drug release behavior of the resulting nanoparticles (oDex-g-Pt+DOX NPs) were systematically investigated by optical experiment, DLS analysis, TEM measurement, and in vitro drugs release experiment. Effective cellular uptake of the oDex-g-Pt+DOX NPs was identified by the human cervical carcinoma HeLa cells via confocal laser scanning microscopy. Furthermore, oDex-g-Pt+DOX NPs displayed a comparable antiproliferative activity than the simple combination of free cisplatin and DOX (Cis+DOX) as the extension of time. More importantly, oDex-g-Pt+DOX NPs exhibited remarkable reversal ability of tumor resistance compared to the cisplatin in cisplatin-resistant lung carcinoma A549 cells. Take advantage of the acidic and reductive microenvironment of tumors, this smart polymer-dual-drugs conjugate could serve as a promising and effective nanomedicine for combination chemotherapy.

Sign in / Sign up

Export Citation Format

Share Document