Drug activity against Onchocerca gutturosa males in vitro: a model for chemotherapeutic research on onchocerciasis

1987 ◽  
Vol 61 (4) ◽  
pp. 271-281 ◽  
Author(s):  
Simon Townson ◽  
C. Connelly ◽  
A. Dobinson ◽  
R. Muller
Keyword(s):  
Culture System ◽  
Serum Proteins ◽  
Good Condition ◽  
Calf Serum ◽  
New Drugs ◽  
Foetal Calf Serum ◽  
Partial Recovery ◽  
Feeder Cells ◽  
Compound Identification

ABSTRACTAn in vitro system for chemotherapeutic research using adult male Onchocerca gutturosa has been developed as a model for O. volvulus. Using a culture system consisting of medium MEM+10% heat inactivated foetal calf serum (IFCS)+LLCMK2 (monkey kidney) feeder cells in an atmosphere of 5% CO2 in air, we examined the effects of a range of antiparasitic drugs on worm motility. Ivermectin, levamisole, furapyrimidone, Mel W, chloroquine, metrifonate, flubendazole, amoscanate and the Ciba-Geigy compounds CGP 6140, CGP 20′376 and CGI 17658 either immobilized or significantly reduced motility levels at a concentration of 5x10−5M or less within a 7-day period. Worms were affected at very low concentrations by ivermectin (effective conc. to reduce motility levels to 50% of controls, 3.14x10−8M), levamisole (7.95x10−8M), CGP 6140 (8.87x10−9M) and CGP 20′376 (2.78x10−8M). Difficulties were experienced in accurately repeating the immotile endpoint for levamisole due to an inconsistent partial recovery of motility. Over a 7-day period diethylcarbamazine had little effect on motility levels, while suramin caused a slight increase in activity compared to controls at some timepoints. Subsequent experiments demonstrated some differences in drug efficacy depending on the presence or absence of serum and feeder cells in the culture system probably because of drug avidly binding to serum proteins. However, serum and cells were found to be essential ingredients of the culture system to maintain worms in good condition, indicating that new drugs should be evaluated both in the presence and absence of serum and cells. Comparisons were made between the responses of O. gutturosa and Brugia pahangi to certain drugs and these species were found to significantly differ in their sensitivities to ivermectin and a novel compound (Wellcome), indicating that Onchocerca parasites should be used wherever possible for compound identification and development intended for the treatment of onchocerciasis. The in vitro system described here, using male O. gutturosa, provides a basis for further research and a practical alternative to O. volvulus.

Maturation of porcine oocytes for 33 or 44 hours in vitro in the presence or absence of serum

1998 ◽  
Vol 1998 ◽  
pp. 180-180
Author(s):  
K. Rust ◽  
M.E. Staines ◽  
GJ. McCallum ◽  
N.S. Prathalingam ◽  
S.A. Edwards ◽  
...  
Keyword(s):  
Disease Transmission ◽  
Calf Serum ◽  
International Markets ◽  
Foetal Calf Serum ◽  
Maturation Time ◽  
Nuclear Maturation ◽  
Defined Media ◽  
Porcine Embryo ◽  
Disease Risks

Porcine embryo production in vitrois providing the impetus for the development of cryopreservation strategies aimed at welfare-friendly domestic and international marketing and movement of stock in a manner that minimises risks of disease transmission. In the context of disease risks, defined media, which avoid the use of serum and other biohazardous products, are likely to become essential in the production of embryos for international markets. In preparing for this situation, the present comparative study investigated in vitronuclear maturation of porcine oocytes in the presence of either foetal calf serum (FCS) or polyvinyl alcohol (PVA). In addition, the effect of restricting the maturation time to 33 rather than 44 hours was examined.

166 EFFECT OF INDIVIDUAL CULTURE SYSTEM ON IN VITRO DEVELOPMENT OF IN VITRO-MATURED - IN VITRO-FERTILIZED BOVINE EMBRYOS

2012 ◽  
Vol 24 (1) ◽  
pp. 195
Author(s):  
R. Nishii ◽  
K. Imai ◽  
H. Koyama ◽  
O. Dochi
Keyword(s):  
Culture System ◽  
Calf Serum ◽  
Control Culture ◽  
Total Cell ◽  
Bovine Embryo ◽  
Bovine Embryos ◽  
Culture Systems ◽  
Individual Culture ◽  
Single Culture

An individual in vitro culture system for bovine embryo needs to be developed for the study of embryo developmental competence. The objective of this study was to examine the effect of individual culture systems on the development of in vitro-matured–in vitro-fertilized bovine embryos. Two individual culture systems were compared. Cumulus–oocyte complexes (COC) were collected by aspiration of ovarian follicles (diameter, 2 to 5 mm) obtained from a local abattoir. The COC were matured in TCM-199 supplemented with 5% calf serum and 0.02 AU mL–1 of FSH. Groups of 20 COC were incubated in 100-μL droplets of IVM media at 38.5°C under an atmosphere of 5% CO2 for 20 h. After 18 h of gamete co-culture (3 × 106 sperm mL–1), the presumptive zygotes were cultured in CR1aa medium supplemented with 5% calf serum at 38.5°C under an atmosphere of 5% CO2, 5% O2 and 90% N2 for 9 days (fertilization = Day 0). The presumptive zygotes were randomly assigned to 1 of the following 3 treatments: single culture (1 zygote was cultured in a 5-μL droplet), well-of-the-well (WOW; Sugimura et al. 2010 Biol. Reprod. 83, 970–978) culture (25 zygotes were cultured individually in each 125-μL droplet) and control culture (25 zygotes were cultured in a 125-μL droplet). Embryo development was evaluated for cleavage and blastocyst rates, on Day 2 and Day 7 to 9 after IVF, respectively. The rates of development up to the blastocyst stage and total cell number in the blastocysts, determined by an air-drying method, were investigated. The cleavage and blastocyst rates were analysed by the chi-square test and the total cell numbers were analysed by ANOVA. The cleavage rates were significantly higher in the control and WOW groups than in the single-culture group (P < 0.01) and the blastocyst rates were significantly lower in the single-culture group than in the control culture group (P < 0.05; Table 1). The total cell numbers (mean ± s.d.) of blastocysts did not significantly differ between the single culture (154.6 ± 21.8), control culture (155.2 ± 22.5) and WOW culture (159.8 ± 27.0) groups. These results indicate that although the blastocyst rate was lower in the single-culture system than in the WOW or group culture system, in vitro-matured–in vitro-fertilized bovine embryos can be cultured using the single-culture system. Moreover, the quality of blastocysts developed by the single-culture system is the same as that of blastocysts developed using the other 2 culture systems. Table 1.Effect of different culture methods for bovine embryo development

Oesophagostomum radiatum: inhibition of in vitro development by newborn calf serum

1990 ◽  
Vol 64 (1) ◽  
pp. 9-14
Author(s):  
I. J. East ◽  
C. J. Fitzgerald
Keyword(s):  
Inhibitory Action ◽  
Natural Infection ◽  
Calf Serum ◽  
Foetal Calf Serum ◽  
Newborn Calf Serum ◽  
In Vitro Development ◽  
Specific Antibodies ◽  
Serum Inhibition ◽  
Vitro Development

ABSTRACTOesophagostomum radiatum developed to fourth stage larvae after 14 days in in vitro culture. However, development was totally inhibited if the standard 50% foetal calf serum in the medium was replaced by newborn calf serum. Inhibition did not occur with serum from cattle immune to O. radiatum through natural infection or experimental vaccination irrespective of the titre of specific antibodies to O. radiatum in each serum. The inhibitory action of NCS could be abolished by heat treatment at 56°C for 1 h but not by dialysis or repeated freeze-thawing. The inhibition was not consistent with observed differences in the activity of 19 enzymes in the various sera or the absence of various thiol-containing stimulants of worm development.

Echinococcus granulosus: observations on strobilar development in in vitro monophasic culture

1995 ◽  
Vol 69 (2) ◽  
pp. 173-175 ◽  
Author(s):  
F. Ponce Gordo ◽  
C. Cuesta Bandera
Keyword(s):  
Liquid Medium ◽  
Echinococcus Granulosus ◽  
Calf Serum ◽  
Foetal Calf Serum ◽  
Stages Of Development ◽  
Protein Substrate ◽  
Cultivation Technique ◽  
Biphasic Medium

AbstractThe in vitro cultivation technique of Echinococcus granulosus protoscoleces usually states the necessity of a biphasic medium with a solid protein substrate for strobilar development to take place; otherwise, in a monophasic medium, protoscoleces follow a vesicular development. However, in some monocphasic cultures, the development of several strobilate individuals (in different quantities and stages of development, depending on the culture) were observed. The only known diference form cultures made previously and snice, where the development was vesicular, was the batch of foetal calf serum used in the constitution of the liquid medium, and this is presumed to be the cause of this unexpected strobilar development.

The Factor Affecting the Spreading of Chondrocytes upon Inorganic Substrate

1973 ◽  
Vol 13 (1) ◽  
pp. 193-204
Author(s):  
M. TAKEICHI
Keyword(s):  
Conditioned Medium ◽  
Calf Serum ◽  
Fresh Medium ◽  
Foetal Calf Serum ◽  
Chick Embryos ◽  
Embryonic Cells ◽  
Mass Cultures ◽  
Rounded Form ◽  
Inorganic Substrate

The effect of conditioned medium (CM) prepared from mass-cultures of chick embryonic cells was studied on the spreading behaviour of chondrocyte derived from sterna of 16-day-old chick embryos. Freshly dissociated chondrocytes exhibited a quite rounded form, and this shape did not change when they were cultured with fresh medium (Eagle's MEM + 6% foetal calf serum) in vitro for several days. Non-dialysable material(s) in CM added into the fresh medium stimulated the formation of pseudopods of chondrocytes, without primarily affecting the synthesis of chondroitin sulphates. Such an activity of CM was not lost after boiling, but it was lost following treatment with proteases. The chondrocytes covered with newly deposited acidmucopoly-saccharides were insensitive to the effect of CM, but they became sensitive to form pseudopods after treatment of the cells with chondroitinase. These results suggest that CM contains a macromolecular material(s) to enhance the motility or adhesiveness of chondrocytes.

Cell-associated proteases affect tumour cell migration in vitro

1979 ◽  
Vol 36 (1) ◽  
pp. 241-252
Author(s):  
J. Varani ◽  
W. Orr ◽  
P.A. Ward
Keyword(s):  
Cell Migration ◽  
Human Serum ◽  
Protease Inhibitors ◽  
Trypsin Inhibitor ◽  
Calf Serum ◽  
Lima Bean ◽  
Caproic Acid ◽  
Foetal Calf Serum ◽  
Migratory Activity

The in vitro migratory activity of mouse fibrosarcoma cells in medium containing either foetal calf serum or normal human serum was studied. These 2 sera were studied because foetal calf serum contains high levels of protease inhibitor activity while human serum contains much less. The cells migrated actively in medium with foetal calf serum but migration was greatly inhibited in human serum-containing medium. When protease inhibitors such as soybean trypsin inhibitor, lima bean trypsin inhibitor and bovine pancreas trypsin inhibitor were added to human serum-containing medium cell migration was supported almost as effectively as in medium with foetal calf serum. Addition of epsilon-amino-n-caproic acid to human serum or depletion of the plasminogen from human serum did not enable it to support enhanced migration. epsilon-amino-n-caproic acid actually inhibited migration. A variant cell population with elevated levels of caseinolytic activity and elevated levels of activity against the substrate n-acetyl-DL-phenylalanine-beta-naphthyl ester (a substrate specific for chymotrypsin-like enzymes) was isolated from the parent cells. When the variant cells were compared to the parent cells regarding migratory activity in foetal calf serum or human serum-containing medium, the variant cells showed much less activity. Only a few, widely scattered variant cells migrated in the human serum-containing medium. These data suggest that a cell-associated factor interferes with the migration of the cells in medium with human serum. This factor apparently is neutralized in medium sontaining human serum to which protease inhibitors with antitrypsin activity have been added.

Initial phases of the rat testis differentiation in vitro

Development ◽  
1984 ◽  
Vol 83 (1) ◽  
pp. 15-31
Author(s):  
Roxane Agelopoulou ◽  
Solange Magre ◽  
Evangélie Patsavoudi ◽  
Alfred Jost
Keyword(s):  
Amniotic Membrane ◽  
Sertoli Cells ◽  
Calf Serum ◽  
Basal Medium ◽  
Foetal Calf Serum ◽  
Clear Cells ◽  
Testis Differentiation ◽  
Sex Chromatin ◽  
Testicular Differentiation

Rat gonadal primordia with their supporting mesonephroi were explanted in vitro at the undifferentiated stage (12 days 16h after fertilization), at the outset of testicular differentiation (13 days 9 h) or when already containing seminiferous cords. The younger foetuses were sexed with the sex chromatin test in the amniotic membrane. The basal medium was CMRL 1066 and the culture period, 1 to 4 days. Testicular differentiation resulted from the appearance of large clear cells, the primordial Sertoli cells, and from their aggregation into seminiferous cords. Addition of 15 % foetal calf serum to the medium prevented the differentiation of seminiferous cords, but large clear cells appeared. In testes from 14- or 15-day-old foetuses, the seminiferous cords disintegrated under the influence of serum. The serum did not prevent the differentiation of Sertoli cells, but impaired organogenesis or maintenance of the early seminiferous cords. The results support previous histological observations on the initial stages of testicular differentiation.

Response of preimplantation rat blastocysts in vitro to extracellular uterine luminal components, serum and hormones

1977 ◽  
Vol 25 (1) ◽  
pp. 265-277
Author(s):  
M.A. Surani
Keyword(s):  
Calf Serum ◽  
Giant Cells ◽  
Biologically Active ◽  
Foetal Calf Serum ◽  
Active Components ◽  
Rat Serum ◽  
Pregnant Females ◽  
Trophoblast Giant Cells

The influence of extracellular environmental factors on preimplantation rat blastocysts was tested by determining the number of embryos which escaped from their zonae pellucidae, followed by attachment and outgrowth of trophoblast giant cells, after 72 h in culture Uterine luminal ocmponents from individual females, or hormones, were included in Dulbecco's medium which contained 4 mg/ml bovine serum albumin. In about 20% of cases, uterine fluids were embryotonic. However, uterine fluids from day-5 pregnant females, the day of implantation in the rat, were more potent in these tests than uterine fluids obtained from ovariectomized females treated with progesterone alone. The potency of a mixture of the 2 fluids was also high. Uterine fluids obtained at 14 h after an injection of oestradiol and progesterone to the ovariectomized females, were also effective in these tests. Rat serum and foetal calf serum were effective too, but steroids or insulin alone in the medium had no detectable influence on embryos. Serum or uterine luminal proteins appear to be essential for maintaining the viability of the blastocysts and for inducing the responses observed here. In the uterine fluids, some proteins released into the lumen after treatment of females with oestradiol and progesterone appear to be the biologically active components. Differences in the responses of blastocysts in vitro are compared with those in vivo.

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