136 CLOSED SYSTEM OF GEL-LOADING-TIP VITRIFICATION FOR IN VITRO-PRODUCED BOVINE BLASTOCYSTS BEFORE AND AFTER BIOPSY

2007 ◽  
Vol 19 (1) ◽  
pp. 185
Author(s):  
K. Tominaga ◽  
F. Iwaki ◽  
E. Yamaguchi ◽  
Y.-M. Dou ◽  
I. Kijihana ◽  
...  
Keyword(s):  
Closed System ◽  
Open Systems ◽  
Sucrose Solution ◽  
Survival Rates ◽  
Japanese Black Cattle ◽  
Significant Difference ◽  
Before And After ◽  
Survival Assay ◽  
In Vitro Survival

Ultra-rapid cooling by direct plunging of gel-loading-tip (GL-tip) into liquid nitrogen (LN2) contributed to the high post-warm survival rates of in-vitro-produced (IVP) bovine embryos (Tominaga and Hamada 2001 J. Reprod. Dev. 47, 267–273). Since GL-tip vitrification and other ultra-rapid vitrification methods have potential risk of microbiological contamination in the LN2, the current study was undertaken to develop a closed system of GL-tip vitrification for IVP bovine blastocysts before and after biopsy. Day 7 blastocysts were produced in vitro (Tominaga et al. 2000 Theriogenology 53, 1669–1680), and biopsied for sex determination (Tominaga and Hamada 2004 Theriogenology 61, 1181–1191). The blastocysts were exposed first to 10% ethylene glycol (EG) +10% DMSO in TCM-199/20% FCS for 2 min at 37°C, and then placed in vitrification medium with 20% EG + 20% DMSO + 0.6 M sucrose in TCM 199/20% FCS for 30 s at 37°C. Three to 4 blastocysts in 0.6 µL of the vitrification medium were aspirated into a GL-tip (NK-tip; Nippon Medical & Chemical Instruments Co., Ltd., Osaka, Japan). Immediately before being plunged into LN2, the tip of the GL-tip was sealed by dipping into 100% glycerol at 4°C, and the opposite open end of the GL-tip was sealed by inserting a holding stick under LN2 vapor. After warming in 0.25 M sucrose solution at 37°C, the glycerol-seal was removed from the tip by gentle shaking. The blastocysts diluted in a two-step manner were cultured up to 72 h for survival assay. Within the 72 h of culture, intact embryos that initiated hatching and biopsied embryos that re-expanded were considered as surviving. Two more biopsied blastocysts derived from ovum pickup, IVF, and IVC were vitrified–warmed in the closed system, and then transferred to 2 recipients. The results indicate no significant difference in the post-thaw survival rates of either intact or biopsied blastocysts in both closed and open systems (Table 1). One recipient became pregnant and delivered a female calf. Gestation period (287 days) and birth weight (22 kg) were within the normal range of the Tajima strain of Japanese Black cattle. We reached the conclusion that the closed system of GL-tip vitrification was simple and highly effective for IVP bovine blastocysts before and after biopsy. Table 1. In vitro survival of intact and biopsied bovine blastocysts after GL-tip vitrification in closed and open systems

scholarly journals A novel in vitro survival assay of small intestinal stem cells after exposure to ionizing radiation

2014 ◽  
Vol 55 (2) ◽  
pp. 381-390 ◽  
Author(s):  
Motohiro Yamauchi ◽  
Kensuke Otsuka ◽  
Hisayoshi Kondo ◽  
Nobuyuki Hamada ◽  
Masanori Tomita ◽  
...  
Keyword(s):  
Stem Cells ◽  
Ionizing Radiation ◽  
Intestinal Stem Cells ◽  
Survival Assay ◽  
Small Intestinal ◽  
In Vitro Survival

scholarly journals Methods to assess the viability of cryopreserved Jatropha curcas L. seed germplasm

2016 ◽  
Vol 18 (2) ◽  
pp. 391-398 ◽  
Author(s):  
A.N. SALOMÃO ◽  
I.R.I. SANTOS ◽  
S.C.B.R. JOSÉ ◽  
J.P. DA SILVA ◽  
B.G. LAVIOLA
Keyword(s):  
Jatropha Curcas ◽  
Germination Percentage ◽  
Experimental Sample ◽  
Embryonic Axes ◽  
Medicinal Uses ◽  
Jatropha Curcas L ◽  
Significant Difference ◽  
Before And After ◽  
Potential Applications

ABSTRACT Jatropha curcas L. is a plant species with many potential applications, especially medicinal uses (hypoglycemic, anti-inflammatory, haemostatic, healing, anti-tumor). The objective of this study was to test germination in moist paper rolls for whole seeds and in vitro for excised embryonic axes, in an attempt to identify the best method to assess the quality of J. curcas seed germplasm, cryopreserved with different water contents. The experimental sample with a 6.2% moisture content (MC) was divided in subsamples which were hydrated and dehydrated for 0 (control), 4, 8, 11 and 24h. The initial germination percentages were 63% for whole seeds and 81% for excised embryonic axes. After exposure to liquid nitrogen (LN), germination percentages were 48% (whole seeds) and 57% (excised embryonic axes). There was no significant difference between germination percentages in embryonic excised from seeds subjected or not subjected to freezing, with different MC. In contrast, there was a reduction of the whole seed germination percentage when exposed to LN (contrast = 0.17, standard error = 0.04, t = 4.09, p = 0.001) and not for the hydration and dehydration treatments. The methodology based on in vitro cultures of the embryonic axis isolated from seeds stored in LN with distinct MC values was more efficient than the standard germination test to evaluate the viability of J. curcas seeds before and after LN storage.

In vitro efficacy of ganciclovir against feline herpesvirus type 1 and assessment of ocular tolerability in healthy cats

2020 ◽  
pp. 1098612X2094436
Author(s):  
Andrew C Lewin ◽  
Chin-Chi Liu ◽  
Christopher Alling ◽  
Pilar Camacho-Luna ◽  
Bruna Miessler ◽  
...  
Keyword(s):  
Blood Chemistry ◽  
Ocular Disease ◽  
Herpesvirus Type ◽  
Significant Difference ◽  
Feline Herpesvirus ◽  
Before And After ◽  
Clinically Significant ◽  
Complete Blood Counts ◽  
Ocular Tolerability

Objectives Feline herpesvirus-1 (FHV-1) is a prevalent cause of ocular disease in cats and limited topical options for treatment currently exist. The first objective of this study was to confirm the efficacy of ganciclovir against FHV-1 in vitro. The second objective was to assess the safety and ocular tolerability of topically applied ganciclovir eye gel (GEG) in healthy cats. Methods FHV-1 was used to infect tissue culture wells covered in maximally confluent Crandall–Rees feline kidney cells prior to the addition of three molarities of ganciclovir (8.9 µM, 17.8 µM and 89 µM) before being incubated for 48 h. Ganciclovir efficacy in vitro was then assessed using standard plaque reduction assay. Commercially available GEG (0.15%) was applied q8h to one randomly chosen eye of four healthy cats for 7 days. Commercially available lubricating eye gel (LEG) was applied to the opposite eye q8h. Complete blood counts (CBC), blood chemistry panels (CHEM) and urinalysis (UA) were performed on all cats before and after the study period. Ocular lesions were assessed daily using a standardized scheme. Results Ganciclovir led to a significant reduction in FHV-1 plaque number, area and diameter at all tested molarities in vitro. The highest molarity assessed (89 µM) caused a 100% reduction in viral plaque number. There was no significant difference in lesion scores between eyes receiving GEG and LEG. Animals remained healthy throughout the study period with CBC, CHEM and UA showing no clinically significant alterations. Conclusions and relevance Based on the in vitro results, ganciclovir appears to be effective against FHV-1 in vitro. When applied q8h as a commercial 0.15% gel to a small group of cats with normal eyes, this medication was well tolerated. Taken together, these data suggest this medication warrants further investigation in cats with ocular disease caused by FHV-1.

Endothelium-dependent arterial wall tone elasticity modulated by blood viscosity

2002 ◽  
Vol 282 (2) ◽  
pp. H389-H394 ◽  
Author(s):  
Edmundo I. Cabrera Fischer ◽  
Ricardo L. Armentano ◽  
Franco M. Pessana ◽  
Sebastián Graf ◽  
Luis Romero ◽  
...  
Keyword(s):  
Blood Viscosity ◽  
Muscle Activation ◽  
Arterial Wall ◽  
Mock Circulation ◽  
Significant Difference ◽  
Before And After ◽  
Strain Relationship ◽  
Instantaneous Pressure

The role of blood viscosity on arterial wall elasticity before and after deendothelization (DE) was studied. Seven ovine brachiocephalic arteries were studied in vitro under physiological pulsatile flow conditions achieved by a mock circulation loop. Instantaneous pressure and diameter signals were assessed in each arterial segment. Incremental elastic modulus ( E inc) was calculated using the slope of the pure elastic stress-strain relationship. There was no significant difference between E inc values before and after DE (3.11 vs. 3.16 107 dyn/cm2) at a blood viscosity of 2.00 mPa · s. Increases in blood viscosity (2.50, 3.00, 3.50, and 4.00 mPa · s) always resulted in decreases of E inc before DE; inversely, increases in blood viscosity resulted in increases of E inc after DE. These values of E inc, for identical levels of blood viscosity, were always significantly lower ( P< 0.05) before DE than those obtained after DE. Arterial wall elasticity assessed through E inc was strongly influenced by blood viscosity, probably due to presence or absence of endothelium relaxing factors or to direct shear smooth muscle activation when endothelial cells are removed.

scholarly journals Therapeutic effectiveness of frozen platelet concentrates for transfusion

Blood ◽  
1981 ◽  
Vol 57 (2) ◽  
pp. 243-249 ◽  
Author(s):  
HM Lazarus ◽  
EA Kaniecki-Green ◽  
SE Warm ◽  
M Aikawa ◽  
RH Herzig
Keyword(s):  
Vapor Phase ◽  
Platelet Concentrate ◽  
In Vitro Tests ◽  
Platelet Concentrates ◽  
Emergency Situations ◽  
Ultrastructural Studies ◽  
Significant Difference ◽  
Before And After ◽  
The Difference

Abstract Six patients received platelet concentrate transfusions from their HLA- identical siblings. Platelet concentrates were administered either fresh, or after being frozen in 10% dimethylsulfoxide, at a slow controlled rate (1 degree C/min) or rapidly (approximately 8 degrees C/min) in the vapor-phase of a liquid nitrogen refrigerator. The median freeze-thaw loss was 13.5%. The mean 1-hr and 20-hr corrected increments in platelet count were calculated for fresh platelet concentrates transfused before and after transfusion with controlled- rate frozen and vapor-phase frozen platelet concentrates. There was no significant difference among the first and second transfusion of fresh platelet concentrates, nor was the difference observed between fresh and controlled-rate frozen platelet concentrates significant. The difference between fresh and vapor-phase frozen platelet concentrates, and between controlled-rate frozen and vapor-phase frozen platelet concentrates were highly significant (p < 0.01). In vitro tests of aggregation using ristocetin and platelet ultrastructural studies paralleled the transfusion experience. Our results indicate that HLA- identical platelet concentrates can be successfully frozen and thawed for transfusion if a slow, controlled rate of freezing is employed. The use of HLA-identical frozen platelet concentrates may be important in emergency situations for the refractory patient and potentially for the establishment of a platelet concentrate bank.

Effect of Single Dose In Vivo Propranolol Therapy on In Vitro Adhesion of Human SS RBC.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1234-1234 ◽  
Author(s):  
Laura M. De Castro ◽  
Jude C. Jonassaint ◽  
Jennifer G. Johnson ◽  
Milena Batchvarova ◽  
Marilyn J. Telen
Keyword(s):  
Safety Data ◽  
Study Drug ◽  
Flow Chamber ◽  
In Vivo Studies ◽  
Dependent Manner ◽  
Number Of Patients ◽  
Significant Difference ◽  
Before And After

Abstract Sickle red blood cells (SS RBC) are abnormally adhesive to both endothelial cells (ECs) and components of the extracellular matrix (ECM). Epinephrine (epi) has been shown to elevate cAMP in SS RBC and increase adhesion of SS RBC to ECs in a protein kinase A-dependent manner. In vitro and in vivo studies performed in our lab have led to the hypothesis that adrenergic stimuli such as epi may initiate or exacerbate vaso-occlusion and thus contribute to the association of vaso-occlusive events with physiologic stress. We are conducting a prospective, dose-escalation pilot clinical study to investigate whether in vivo administration of one dose of propranolol either down-regulates baseline SS RBC adhesion in vitro or prevents its upregulation by epi. In addition, this study will provide additional safety data regarding the use of propranolol in normotensive patients with sickle cell disease (SCD). Figure Figure To date, we have completed the first two dose cohorts. 11 subjects (9 SS and 1 Sβ° thalassemia; 7 females, 3 males) have participated. No severe adverse events were noted. Cohorts 1 and 2 had mean pre-propranolol blood pressure (BP) of 116 (5.9 SD)/ 60.4 (3.98 SD) and 106.8 (4.68 SD)/ 58 (3.9 SD), respectively; this difference was not statistically significant. Minimal and asymptomatic changes in BP were noted in both cohorts after drug administration, with biphasic systolic and diastolic BP nadirs at 45 and 240 minutes. No clinically significant changes in heart rate were observed. Adhesion studies were performed using a graduated height flow chamber on the day of RBC collection. RBC adhesion to ECs was studied before and after epi stimulation and was measured at sheer stresses ranging from 1 to 3 dyne/cm2. Baseline adhesion measurements were validated by comparing percent (%) adhesion assayed at 2 different times within 7 days—at screening and before propranolol dose on the study drug day. We observed no significant difference in adhesion at the 2 different time points without propranolol. Comparison of % adhesion of epi-stimulated RBC to ECs before and 1 hour after propranolol showed that propranolol given in vivo significantly inhibited both non-stimulated and epi-stimulated SS RBC adhesion (p=0.04 and p=0.001, respectively). Lastly, comparison of SS RBC adhesion at both drug doses confirmed the drug-related inhibition of adhesion (p&lt;0.004). We conclude that propranolol administered in vivo decreases SS RBC baseline adhesion to ECs and substantially abrogates epi-stimulated adhesion to ECs, as measured in vitro. Although we have thus far studied only a small number of patients and low propranolol doses, we expect to confirm these results with the 3rd cohort, in which a higher dose of propranolol will be used. If our findings continue to show that propranolol can decrease both SS RBC baseline and epi-stimulated adhesion to ECs, study of propranolol on a larger scale would be warranted in order to ascertain its safety and efficacy as an anti-adhesive therapy in SCD.

scholarly journals The difference of the salivary volume before and after drinking the rosella tea (Hibiscus sabdariffa)

2010 ◽  
Vol 22 (3) ◽  
Author(s):  
Mutiara Indah Permata Sari Islami ◽  
Edeh Roletta Haroen ◽  
Sri Tjahajawati
Keyword(s):  
In Vitro Study ◽  
T Test ◽  
Hibiscus Sabdariffa ◽  
Test Analysis ◽  
Community Benefits ◽  
Significant Difference ◽  
Before And After ◽  
Quasi Experimental ◽  
The Difference

Introduction: Roselle plants (Hibiscus sabdariffa) is one of the herbs that began to be used by the community. Benefits of this plant is quite a lot for health. The portion taken for consumption is the red flower petals. oselle tea is one of the sour beverages which can affect the salivary gland secretion. The purpose of this study is to analyzed the difference of salivary volume before and after drinking roselle tea. Methods: This study has been conducted to 40 students of Faculty of Dentistry, Padjadjaran University, ranging from 18-23 years of age with good general condition. This study is quasi-experimental in vitro study using the paired test analysis with α = 0,05 of the data collected from salivary volume. Results: The result of study indicates that the average of salivary volume before drinking roselle tea is 1,90 milliliter. After drinking roselle tea, the average of salivary volume is 4,54 milliliter. The result of paired test analysis shows that t-test is 16,172 and t-table is 2,022. The value of t-test > t-table. Result of statistic shown there is significant difference of salivary volume before and after drinking roselle tea. Conclusion: There is a difference of salivary volume before and after drinking roselle tea.

scholarly journals Influence of scarification method on seed germination of the terrestrial orchid Anacamptis laxiflora (Lam.)

2021 ◽  
Vol 5 (1) ◽  
pp. 15-23
Author(s):  
Gwenaëlle Deconninck ◽  
Argyrios Gerakis
Keyword(s):  
Infection Rate ◽  
Sucrose Solution ◽  
Germination Rate ◽  
Terrestrial Orchid ◽  
Significant Difference ◽  
Terrestrial Orchids ◽  
Sexual Propagation ◽  
Centrifugation Method ◽  
Bacteria And Fungi

AbstractA critical step during in vitro sexual propagation of terrestrial orchids is the treatment of the microscopic seeds with a disinfecting solution that kills bacteria and fungi attached to the seeds. This treatment is necessary to prevent infection of the culture vessels. At the same time, the treatment serves to scarify the seeds, a process that disrupts seed dormancy and initiates germination. The literature is inconclusive with respect to the proper combination of disinfecting solution strength and treatment duration. Both factors should be adapted to each species to guarantee minimal infection rate without damaging the embryo. This research aims to compare three disinfection/scarification methods for seeds of Anacamptis laxiflora (Lam.): (i) soaking in 0.5% NaClO, (ii) soaking in 0.5% NaClO, then centrifugation, and (iii) presoaking the seeds in sucrose solution, then soaking in 0.5% NaClO. The seeds were soaked in the disinfecting solution for 5 to 85 min. Following scarification, the seeds were sown in modified Malmgren nutrient medium. Infected and germinated vessels were counted at 41 and 189 d after sowing. We found that the longer the chemical treatment, the lower the infection rate, and the higher the germination rate. There was no significant difference in germination rate between the NaClO and the NaClO-plus-centrifugation method; in fact, the slight savings in disinfection time effected by centrifugation were more than offset by the added complexity of the method. Moreover, we found that centrifugation significantly delays germination. The sucrose presoak-plus-NaClO method was superior to plain NaClO, as the sucrose stimulates the germination of microbial spores on the surface of the seeds, making them easier to kill. Perhaps seeds with thicker testa as well as whole immature capsules could benefit even more from the pretreatment in sucrose solution.

scholarly journals Anticariogenic potential of white cheese, xylitol chewing gum, and black tea

2018 ◽  
Vol 12 (02) ◽  
pp. 199-203 ◽  
Author(s):  
Pinar Gul ◽  
Nilgun Akgul ◽  
Nilgun Seven
Keyword(s):  
Ph Value ◽  
Sucrose Solution ◽  
Black Tea ◽  
Critical Value ◽  
White Cheese ◽  
Plaque Ph ◽  
Significant Difference ◽  
Before And After ◽  
Tukey Honestly Significant Difference ◽  
Post Hoc

ABSTRACT Objective: This study aimed to examine the effects of these foods on plaque pH and the potential development of tooth decay. Materials and Methods: Plaque pH was measured using the sampling method before and after 1, 5, 10, 20, 30, 45, and 60 min following consumption of these foods individually and after rinsing with a 10% sucrose solution. Statistical analysis was performed using one-way ANOVA and Tukey honestly significant difference post hoc tests (α = 0.05). Results: Although there were statistically significant differences in all test groups except the BT (P = 0.620) and sucrose + XCG (P = 0.550) groups in time, none of the participants chosen for this study were having a plaque pH value anywhere close to the critical value (pH = 5.5). Conclusion: WC, BT, and XCG are advisable as anticariogenic foods because pH values are not below critical value.

Supplementation of culture medium with knockout serum replacement improves the survival of bovine secondary follicles when compared with other protein sources during in vitro culture

Zygote ◽  
2019 ◽  
Vol 28 (1) ◽  
pp. 32-36
Author(s):  
D.S. Gomes ◽  
L.B. Aragão ◽  
M.F. Lima Neto ◽  
P.A.A. Barroso ◽  
L.R.F.M. Paulino ◽  
...  
Keyword(s):  
Bovine Serum ◽  
Culture Medium ◽  
Survival Rates ◽  
Basal Membrane ◽  
Histological Evaluation ◽  
Serum Replacement ◽  
Before And After ◽  
Chi Squared ◽  
Knockout Serum Replacement

SummaryThe present study evaluated the effect of knockout serum replacement (KSR), fetal bovine serum (FBS) and bovine serum albumin (BSA) on the viability and growth of bovine secondary follicles cultured in vitro for 12 days. To this end, secondary follicles were isolated (185–202 μm) and cultured in vitro in TCM-199+ medium supplemented with KSR (5% and 10%), FBS (5% and 10%) or BSA (3 mg/ml) at 38.5°C with 5% CO2 in air. Follicular diameters were evaluated on days 0, 4, 8 and 12. After 12 days of culture, follicular survival analysis was performing by using calcein-AM and ethidium homodimer. Before and after culture, follicles were fixed in paraformaldehyde for histological evaluation. Follicular diameter at different days of culture were compared using the Kruskal–Wallis test, while the percentages of viable follicles were analyzed by chi-squared test (P < 0.05). Results showed that follicles cultured in the presence of KSR at both concentrations presented higher follicular survival rates than those cultured in control medium alone or supplemented with FBS or BSA. Conversely, the presence of KSR, BSA or FBS did not increase follicular diameter after 12 days of culture. Histology analysis showed that, among the tested treatments, follicles cultured in the presence of KSR had preserved rounded oocytes, juxtaposed granulosa cells and intact basal membrane. In conclusion, supplementation of culture medium with KSR increases the follicular survival of bovine secondary follicles cultured in vitro.

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